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EuropeanNeuropsychopharmacology

The Journal of the European College of Neuropsychopharmacology

VOLUME 23 (2013) Supplement 1

VOLUME 23 (2013)

ELSEVIER B.V. and ECNP

AMSTERDAM – BOSTON – LONDON – NEW YORK – OXFORD – PARIS – PHILADELPHIA – SAN DIEGO – ST LOUIS

Abstracts of the

Nice, France

2013 ECNP Workshop on Neuropsychopharmacologyfor Young Scientists in Europe7-10 March 2013

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iii

Contents

MOLECULAR NEUROPSYCHOPHARMACOLOGY

LecturesS.01.01 Induced pluripotent stem cells and in vitro models of neurodevelopmental disorders

J. Price*, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S1S.01.02 Novel strategies for probing 3D structures of GPCRs

F. Marshall*, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S2

PostersP.1.001 3-Nitropropionic acid induces autophagy by forming mitochondrial permeability transition pore, not by activating

mitochondrial fissionM.E. Solesio*, S. Saez-Atienzar, J. Jordan, M.F. Galindo, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S3

P.1.002 Pharmacological characterisation of positive allosteric modulators acting on the metabotropic glutamate receptor 2K.A. Bennett*, A. Weaver, F. Marshall, C.J. Langmead, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . S4

P.1.003 Inflammatory response in the brain of rats exposed to chronic mild stressC. Zecchillo*, F. Macchi, R. Molteni, G. Racagni, M. Papp, M.A. Riva, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . S5

P.1.004 RNAi-mediated serotonin transporter suppression rapidly increases serotonergic neurotransmission and hippocampalneurogenesisA. Ferres-Coy*, F. Artigas, A. Bortolozzi, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S6

P.1.005 Role of cannabinoid CB1 receptors in modulation of dopamine output in the prefrontal cortex associated with food restrictionin ratsV. Licheri*, G. Talani, L. Dazzi, F. Biggio, C. Utzeri, V. Lallai, S. Lutzu, G. Biggio, E. Sanna, Italy . . . . . . . . . . . . . . S7

P.1.006 Oxytocin controls CRF gene expression through TORC3: implications for stress-related disordersB. Jurek*, D.A. Slattery, Y. Liu, I.D. Neumann, G. Aguilera, E.H. van den Burg, Germany . . . . . . . . . . . . . . . . . . . S7

P.1.007 A proteomic and functional analysis reveals that 5-HT6 receptors modulate neuronal differentiation by recruitment of Cdk5F. Duhr*, M. Seveno, C. Mannoury la Cour, D. Dupuis, M.J. Millan, J. Bockaert, P. Marin, S. Chaumont-Dubel, France . . S8

P.1.008 V1B/CRF1 receptor heterodimerisation as a key mechanism of vasopressin and corticotropin-releasing factor synergismJ. Mion*, V. Boulay, M.J. Millan, G. Guillon, M. Corbani, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S9

P.1.009 Cell synchronisation as a tool to optimise expression of metabotropic glutamate receptors in inducible mammalian expressionsystemB. Chruscicka*, P. Branski, G. Burnat, A. Pilc, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S10

P.1.010 The identification of protein tyrosine phosphatase, non-receptor type 1 in hippocampal modulation of food anticipatorybehaviourE. Kostrzewa*, L.A.W. Verhagen, C. Gelegen, H.A. van Lith, D.A. Collier, M. Mitsogiannis, E. de Vries, M. van Gestel,R.A. Adan, M.J.H. Kas, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S11

P.1.011 Identification of a significant role for the ventral hippocampus in neuropeptide S-elicited anxiolysisI.A. Ionescu*, J. Dine, J. Stepan, Y.C. Yen, L. Herrmann, F. Holsboer, C.T. Wotjak, R. Landgraf, M. Eder, U. Schmidt, Germany S12

P.1.012 Stress-induced vulnerability of presynaptic glutamatergic terminals and effect of desipramineN. Nava*, M. Popoli, L. Musazzi, G. Wegener, J.R. Nyengaard, Denmark . . . . . . . . . . . . . . . . . . . . . . . . . . . . S13

P.1.013 Chronic (−)cannabidiol produces antidepressant-like effects in bulbectomised mice, acting on 5-HT1A and CB1 receptorfunctionalityR. Linge*, A. Pazos, A. Diaz, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S13

P.1.014 Intracerebroventricular administration of interleukin-1b elevates brain kynurenic acid and disrupts prepulse inhibition inC57BL/6 miceM. Larsson*, L. Schwieler, G. Engberg, S.B. Powell, S. Erhardt, Sweden . . . . . . . . . . . . . . . . . . . . . . . . . . . . S14

P.1.015 Time-dependent adaptations at the level of dopamine D2 receptor in stress-resilient ratsD. Zurawek*, A. Faron-Gorecka, M. Kusmider, M. Kolasa, P. Gruca, M. Papp, M. Dziedzicka-Wasylewska, Poland . . . . . S15

P.1.016 Modulation of dopamine D2L and D3 receptor signalling and cell surface-cytosol trafficking by dysbindin in CHO cellsN. Schmieg*, C. Rocchi, R. Maggio, M.J. Millan, C. Mannoury la Cour, France . . . . . . . . . . . . . . . . . . . . . . . . S16

P.1.017 Histaminergic regulation of presumed serotonin neurons in the dorsal raphe nucleusK. Panetta*, D. Belelli, J. Lambert, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S16

P.1.018 Dissecting the neural mechanisms underlying impaired threat detection in the Ahi1 knockout mouseA. Lotan*, T. Lifschytz, A. Slonimsky, S. Abedat, Y. Fellig, H. Cohen, O. Lory, G. Goelman, B. Lerer, Israel . . . . . . . . . S17

P.1.019 Role of the 5-HT2A receptor in the mechanism of action of antidepressant drugs: a translational human–mouse studyG. Quesseveur*, A.C. Petit, F. Gressier, R. Colle, D.J. David, A.M. Gardier, C. Verstuyft, E. Corruble, B.P. Guiard, France . S18

P.1.020 The role of CREB/BDNF/TrkB signalling in the zinc deficiency model of depressionU. Doboszewska*, B. Szewczyk, M. Sowa-Kucma, K. Mlyniec, G. Nowak, Poland . . . . . . . . . . . . . . . . . . . . . . . S19

P.1.021 Chronic administration of haloperidol in rats and its effect on microglial cell density and whole brain weight and volumeP.S. Bloomfield*, O.D. Howes, V. de Paola, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S20

P.1.022 Role of trace amine-associated receptor 1 (TAAR1) in the modulation of the dopaminergic system and cortico-striatal signallingS. Espinoza*, I. Sukhanov, G. Lignani, L. Medrihan, S. Maggi, G. Giannotti, F. Fumagalli, F. Benfenati, V. Tucci, R. Gainetdinov,Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S21

P.1.023 Survival role of embryonal proteins in Alzheimer’s disease linked dementia via regulation of oxidative stress and level ofcatecholaminesK. Yenkoyan*, M. Aghajanov, Armenia, Republic of . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S22

P.1.024 Endocannabinoid-mediated plasticity at inhibitory synapses on dopamine cells as a marker of vulnerability to addictionC. Sagheddu*, M. Melis, M. Pistis, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S22

iv Contents

BEHAVIOURAL PHARMACOLOGY

LecturesS.02.01 Genetic models of psychiatric disorders: phenotyping mutant mice

J. Waddington*, Ireland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S25S.02.02 Developmental models of schizophrenia and their behavioural characterisation

K.C.F. Fone*, D.J.G. Watson, A. McIntosh, M.V. King, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S25

PostersP.2.001 Citalopram given to stressed and control pregnant rats causes sex-dependent changes in behaviour and CRH mRNA expression

in their offspringI. Zohar*, M. Weinstock, Israel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S27

P.2.002 Behavioural, molecular and glutamatergic changes in a developmental model of schizophrenia, and reversal by a 5-HT6receptor antagonistM.V. King*, O. Negm, P. Tighe, S. Knapp, P. Wigmore, K.C.F. Fone, United Kingdom . . . . . . . . . . . . . . . . . . . . . S27

P.2.003 Sub-anaesthetic ketamine modulates intrinsic blood oxygen level-dependent (BOLD) connectivity between the hippocampusand the prefrontal cortex in the ratN. Gass*, A. Sartorius, A.J. Schwarz, E. Schenker, C. Risterucci, M. Spedding, L. Zheng, A. Meyer-Lindenberg, W. Weber-Fahr,Germany . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S28

P.2.004 The dopamine b-hydroxylase inhibitor nepicastat suppresses different chocolate-motivated behaviours in ratsA. Zaru*, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S29

P.2.005 The trans-isomer of resveratrol acutely improves motivation in ratJ. Samardzic*, L. Gojkovic-Bukarica, D.I. Obradovic, Serbia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S30

P.2.006 Effects of serotonin (5-HT)1B receptor ligands on amphetamine-seeking behaviour in ratsJ. Miszkiel*, E. Przegalinski, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S31

P.2.007 Tryptophan depletion impairs emotion recognition in healthy womenM. Defrancesco*, W. Parson, J. Marksteiner, E.A. Deisenhammer, J. Kemmler, H. Niederstatter, H. Hinterhuber, Austria . . S32

P.2.008 5XFAD mouse model of Alzheimer’s disease: a dissociation between brain pathology and behavioural phenotypeK. Sonn*, K. Jaako, R.K. Jain, A. Zharkovsky, Estonia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S33

P.2.009 Effect of early life experiences on brain structure and function: neurogenesis and decision makingM. Loi*, S. Koricka, L. de Visser, M.J.H. Kas, P.J. Lucassen, M. Joels, The Netherlands . . . . . . . . . . . . . . . . . . . . S33

P.2.010 Effects of L-DOPA and sarizotan treatments in a parkinsonian rat model of depressionN. Schintu*, X. Zhang, A.A. Mathe, P. Svenningsson, Sweden . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S34

P.2.011 GluA1 and its Postsynaptic density protein 95 (PSD-95)/Discs large/Zonula occludens-1 (PDZ)-interaction: a role inexperience-dependent behavioural plasticity in the forced swim testF. Freudenberg*, V. Marx, V. Mack, L.E. Layer, M. Klugmann, P.H. Seeburg, R. Sprengel, T. Celikel, Germany . . . . . . . S35

P.2.012 Chronic modulation of 5-HT4 and 5-HT6 serotonergic receptors: a new hope in the treatment of cognitive deficits?A. Quiedeville*, T. Freret, V. Bouet, M. Boulouard, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S36

P.2.013 Modelling serotonergic neuromodulation of behavioural performance in spatial working memoryM. Cano-Colino*, R. Almeida, A. Compte, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S37

P.2.014 Serotonin receptor 2B knockout mice present schizophrenic-like behaviourP.M. Pitychoutis*, L. Maroteaux, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S37

P.2.015 Low performing rats model the inattentive subtype of adult ADHD in the 5-choice continuous performance task (5C-CPT)A. Tomlinson*, K.M. Marshall, J.C. Neill, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S38

P.2.016 Exposure to an alternative reward does not reduce cocaine-seeking behaviourC. Nicolas*, C. Lafay-Chabassier, M. Solinas, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S39

P.2.017 The hallucinogen 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) disrupts cortical function: reversal by antipsychotic drugsM.S. Riga*, F. Artigas, P. Celada, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S40

P.2.018 Characterisation of the effects of partial agonist of a4b2* nACh receptor cytisine in the two-choice serial reaction time taskG. Makshakov*, O. Dravolina, A. Bespalov, E. Kayukova, M. Dorofeikova, E. Zvartau, Russia . . . . . . . . . . . . . . . . S41

P.2.019 The mismatch hypothesis: a new way of linking early experiences and adult environment to vulnerability to stressS. Santarelli*, K.V. Wagner, J. Hartmann, X.D. Wang, M.V. Schmidt, Germany . . . . . . . . . . . . . . . . . . . . . . . . . S42

P.2.020 Disruption of 5-HT7 receptors accelerates age-related episodic-like memory declineG. Beaudet*, M. Brehin, T. Freret, G. Nee, V. Delaunay, M. Boulouard, E. Paizanis, France . . . . . . . . . . . . . . . . . . S42

P.2.021 Role of adenosine (A)2A receptor in nicotine addiction − pharmacological and genetic aspectsJ. Czyzyk*, E. Nowak, M. Bader, K. Fuxe, M. Filip, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S43

P.2.022 Oxytocin in the central nucleus of the amygdala mediates social buffering of fearE. Rickenbacher*, M. Moita, Portugal . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S44

P.2.023 The antagonist of 5-HT7 receptors, SB-269970, and amisulpride both reverse ketamine-induced cognitive inflexibility in ratsD. Rafa*, A. Nikiforuk, P. Popik, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S45

P.2.024 The effects of positive allosteric modulators of a7 nicotinic receptors on rats’ performance in the odour span testA. Potasiewicz*, A. Nikiforuk, P. Popik, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S46

P.2.025 Ventral medial prefrontal cortex inactivation reduces context-induced reinstatement of nicotine seekingR.F. Struik*, J. Peters, S. Jonkman-Tielemans, T.J. De Vries, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . S46

P.2.026 Pro-inflammatory cytokines induce anhedonia in mice and increase monoamine transporter activity in the nucleus accumbensF. van Heesch*, J. Prins, K.G.C. Westphal, G.A.H. Korte-Bouws, B. Olivier, A.D. Kraneveld, S.M. Korte, The Netherlands . S47

P.2.027 a7 nicotinic acetylcholine receptor (a7 nAChR) agonists or antagonists as potential cognition enhancers?N.P. Van Goethem*, J. Prickaerts, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S48

P.2.028 Enhanced vulnerability to cocaine reinforcing effects in female H/Rouen mice selectively bred for depressive-like behaviourV. Rappeneau*, A.L. Morel, P.H. Luppi, J.M. Vaugeois, M. El Yacoubi, A. Berod, France . . . . . . . . . . . . . . . . . . . S49

P.2.029 Serotonin regulates hippocampal synaptic plasticity and object memory in miceS.P. Fernandez*, A. Gruart i Masso, P. Gaspar, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S50

P.2.030 Are nutritional variables associated with cognition in stimulant-dependence?C.F. Whitelock*, K.D. Ersche, United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S50

Contents v

P.2.031 The CBA/J mouse as a genetic model of visceral hypersensitivity with co-morbid anxiety and depression: role of glutamatetransportR.D. Moloney*, T.G. Dinan, J.F. Cryan, Ireland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S51

P.2.032 Nanoparticles as disease-modifying mediators for brain therapy: focus on Huntington’s diseaseB.M.D.C. Godinho*, J.R. Ogier, R. Darcy, C.M. O’Driscoll, J.F. Cryan, Ireland . . . . . . . . . . . . . . . . . . . . . . . . S52

P.2.033 Does serotonin depletion augment or counteract the aggression-provoking effect of testosterone in mice?E. Studer*, J. Naslund, L. Westberg, E. Eriksson, Sweden . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S53

P.2.034 Swim-stress exposure effects on nociceptive behaviour and the endocannabinoid system in two rat strains differing in stressresponsivityE.M. Jennings*, B. Okine, W.M. Olango, M. Roche, D.P. Finn, Ireland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S54

P.2.035 ‘Anxious’ rats exhibit an enhanced anxiogenic response to acute SSRI treatment and indices of heightened serotonergictransmissionJ. Naslund*, E. Studer, R. Petterson, H. Nissbrandt, E. Eriksson, Sweden . . . . . . . . . . . . . . . . . . . . . . . . . . . . S54

EPIGENETICS: TOWARDS NEW DRUG TARGETS

LecturesS.03.01 The epigenetics of brain disorders and their inheritance − focus on cognition

I. Mansuy*, Switzerland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S57

PostersP.3.001 Valproic acid treatment prevents the development of deficit in sensorimotor gating in adult prenatally

methylazoxymethanol-treated ratsJ. Latusz*, E. Bator, P. Mordalska, K. Wedzony, M. Mackowiak, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S57

P.3.002 The effect of valproic acid on changes in methylation pattern of histone H3 induced by prenatal MAM administration in mPFCE. Bator*, J. Latusz, P. Mordalska, K. Wedzony, M. Mackowiak, Poland . . . . . . . . . . . . . . . . . . . . . . . . . . . . S58

P.3.003 Chronic early-life stress programs hippocampal neurogenesis and cognitive function: a role for epigenetics?E.F.G. Naninck*, L. Hoeijmakers, M. Engel, P.J. Lucassen, A. Korosi, The Netherlands . . . . . . . . . . . . . . . . . . . . S59

P.3.004 Possible interplay between BDNF and dynorphin in bipolar disorder: role of epigenetic mechanismsA. Di Francesco*, C. D’Addario, B. Dell’Osso, M.C. Palazzo, B. Benatti, D. Galimberti, B. Arosio, A.C. Altamura,M. Maccarrone, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S60

P.3.005 Early environment affects activity based anorexia genetic susceptibility and epigenetic programming of neurodevelopmentalgenes in miceE. Pjetri*, E.V.S. Hessel, H. Oppelaar, P.N.E. de Graan, B. Olivier, M.J.H. Kas, The Netherlands . . . . . . . . . . . . . . . S61

P.3.006 Epigenetic modifications of bdnf gene at hippocampal level induced by chronic ethanol intake in C57BL/6J miceE. Stragier*, R. Massart, M. Hamon, L. Lanfumey, France . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S61

P.3.007 Stress-induced miRNA changes in depression: peripheral biomarker or pathophysiology?S. Kalman*, K. Garbett, A. Vereczkei, R.C. Shelton, K. Mirnics, Hungary . . . . . . . . . . . . . . . . . . . . . . . . . . . . S62

P.3.008 S-Adenosyl-methionine impairs forced swimming-induced behavioural immobility by inhibiting gene expression indentate gyrus neuronsE.A. Saunderson*, A.F. Trollope, M. Gutierrez-Mecinas, A.A. Shaikh, H. Spiers, J. Mill, J.M.H.M. Reul, United Kingdom . . S63

P.3.009 Differential effects of prenatal stress on serotonin transporter deficient mice: the role of epigenetic programmingK. Schraut*, S.B. Jakob, G. Kenis, A.G. Schmitt, S. Kneitz, C.J. Scholz, G. Ortega, H. Steinbusch, D.L.A. van den Hove,K.P. Lesch, Germany . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S64

P.3.010 Time-dependent effects of antidepressant treatments on miRNome expression profile in hippocampus of ratsM. Seguini*, D. Tardito, A. Mallei, G. Racagni, M. Popoli, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S65

CLINICAL NEUROPSYCHOPHARMACOLOGY

LecturesS.04.01 Disrupted cerebral connectivity and brain disorders: linking circuits to genes

A. Meyer-Lindenberg*, H. Tost, E. Bilek, Germany . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S67

PostersP.4.001 Clinical implications of polarity index of drugs in maintenance treatment of bipolar disorder: a naturalistic study

D. Popovic*, E. Vieta, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S68P.4.002 Genetic differences in drug-metabolising enzymes: can they be used to predict antidepressant treatment response?

K. Hodgson*, K.E. Tansey, R. Uher, O.S.P. Daivs, K.J. Aitchison, P. McGuffin, United Kingdom . . . . . . . . . . . . . . . S69P.4.003 Publication pressure and burn out among Dutch medical professors: a nationwide survey

J. Tijdink*, Y.M. Smulders, A.C.M. Vergouwen, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S70P.4.004 Early onset of lithium prophylaxis as possible good prognostic factor for staging bipolar disorder

S. Brioschi*, L. Franchini, C. Colombo, E. Smeraldi, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S71P.4.005 Effects of cigarette smoking on schizophrenia treatment with olanzapine

K. Zoric*, N. Zivkovic, G. Djokic, Serbia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S71P.4.006 Reward processing in unaffected siblings of schizophrenia patients: a functional magnetic resonance imaging study

M. de Leeuw*, M. Vink, R.S. Kahn, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S72P.4.007 Associations of antiepileptic drugs with anxiety and depressive symptoms in paediatric epilepsy: a pilot study

D. Stevanovic*, J. Jancic, Serbia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S73P.4.008 Lithium-regulated genes in lymphoblastoid cells from bipolar-affective patients

S. Kittel-Schneider*, M. Hilscher, S. Schreck, C.J. Scholz, A. Reif, Germany . . . . . . . . . . . . . . . . . . . . . . . . . . S74P.4.009 Age at onset and cognitive impairment in schizophrenia: an ecological cross-sectional study with stabilised patients

A. Caldiroli*, M. Buoli, E. Caletti, R.A. Paoli, S. Zago, A.C. Altamura, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . S74P.4.010 Association of personality features with lithium prophylactic response

D. Dembinska-Krajewska*, S. Kliwicki, M. Chlopocka-Wozniak, J. Rybakowski, Poland . . . . . . . . . . . . . . . . . . . . S75

vi Contents

P.4.011 Clinical features and drug-induced side effects in early versus late antidepressant respondersC. Fabbri*, A. Marsano, M. Balestri, A. Serretti, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S76

P.4.012 Effects of second generation antipsychotics on cognitive domains measured with the Matrics Consensus Cognitive Batteryin early psychosesI. Montalvo*, M. Creus, A. Gutierrez-Zotes, L. Ortega, R. Monseny, T. Feliu, J. Franch, E. Vilella, J. Labad, Spain . . . . . S77

P.4.013 Modulation of brain structure by catechol O-methyltransferase Val158Met polymorphism in chronic cannabis usersA. Batalla*, C. Soriano-Mas, M. Torrens, J.A. Crippa, S. Bhattacharyya, L. Blanco-Hinojo, B.J. Harrison, M. Farre, J. Pujol,R. Martın-Santos, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S78

P.4.014 Fear conditioning and context learning in relation to treatment outcome: a study in patients with panic disorder and social phobiaP. Duits*, J.M.P. Baas, I.M. Engelhard, M.A. van den Hout, D.C. Cath, The Netherlands . . . . . . . . . . . . . . . . . . . S79

P.4.015 Long term therapy with methylphenidate induces modest effects on growth in ADHD childrenC. Balia*, A. Anedda, F. Granitzio, S. Carucci, A. Zuddas, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S80

P.4.016 Regional differences of SERT occupancy in major depression: an in vivo PET study using [11C]DASBP. Baldinger*, G.S. Kranz, M. Savli, W. Wadsak, D. Haeusler, A. Hahn, M. Mitterhauser, C. Philippe, S. Kasper,R. Lanzenberger, Austria . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S80

P.4.017 Post training noradrenaline reuptake inhibition modulates fear memory consolidation and abolishes long term fear responsesin humansJ. Almeida*, J. Tulen, F. van der Veen, S. Kushner, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S81

P.4.018 General error monitoring system dysfunction in obsessive compulsive disorder patients: an event-related potential studyL. Carmi*, U. Alyagon, A. Zangen, R. Dar, J. Zohar, Israel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S82

P.4.019 Insight and recovery in schizophrenic patients: an observational studyD. Cannavo*, C. Concerto, E. Battaglia, O. Bianchini, E. Aguglia, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . S83

P.4.020 Allopurinol for mania − a randomised trial of allopurinol vs. placebo as add-on treatment in manic bipolar patientsS. Burshtein*, M. Weiser, A.A. Gershon, G. Marian, N. Vlad, I.G. Grecu, E. Tocari, A. Tiugan, M. Hotineanu, J.M. Davis, Israel S84

P.4.021 Genome-wide association analysis and pathway analysis in predicting antidepressant treatment responseN. Antypa*, A. Drago, A. Serretti, Italy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S85

P.4.022 Neural correlates of generalised anxiety disorder without comorbid depression: preliminary data from a functional MRI studyK. Hilbert*, U. Lueken, K. Beesdo-Baum, Germany . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S86

P.4.023 Habenular nuclei in different phases of major depressive disorder: a magnetic resonance imaging volumetric studyJ. De Diego-Adelino*, M. Carceller, M. Serra-Blasco, Y. Vives-Gilabert, B. Gomez-Anson, D. Puigdemont, E. Alvarez,V. Perez, M.J. Portella, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S86

P.4.024 Using positron emission tomography to investigate microglial activation in alcohol dependence: preliminary findingsN.J. Kalk*, Q. Guo, R. Cherian, D. Erritzoe, A. Waldman, K. Dar, R.N. Gunn, D.J. Nutt, E.A. Rabiner, A.R. Lingford-Hughes,United Kingdom . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S87

P.4.025 Effects of deep brain stimulation of prelimbic and infralimbic areas of the prefrontal cortex of the ratL. Jimenez-Sanchez*, L. Perez-Caballero, A. Castane, X. Lopez-Gil, L. Campa, M. Galofre, E. Berrocoso, A. Adell, Spain . S88

P.4.027 Six-month follow-up study of repetitive transcranial magnetic stimulation in the treatment of resistant major depressive disorderC. Concerto*, D. Cannavo, F. Magnano SanLio, R. Ricceri, G. Lanza, E. Aguglia, Italy . . . . . . . . . . . . . . . . . . . . S89

P.4.028 Neural correlates of stress-related hypothalamic–pituitary–adrenal axis activity in neuroticismA. Boehringer*, F. Lederbogen, H. Tost, L. Haddad, A. Meyer-Lindenberg, Germany . . . . . . . . . . . . . . . . . . . . . S90

P.4.029 Antioxidant status and fatty acids in adolescents with Asperger syndrome and first episodes of early-onset psychosisM.G. Moron-Nozaleda*, M. Alvarez-Blazquez, C.M. Dıaz-Caneja, E. Dulin, C. Moreno, M.C. Guisasola, C. Arango,M. Parellada, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S91

P.4.030 Bright light dose correlates with change in striatal serotonin transporter binding in healthy ScandinaviansB. Mc Mahon*, A.S. Andersen, L. Feng, K.K. Holst, M.K. Madsen, S. Lehel, M.M. Herth, P. Iversen, L. Hasholt, G.M. Knudsen,Denmark . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S92

P.4.031 Effects of mental distress on cognitive functioning in patients admitted for cardiac rehabilitation after acute coronary eventsJ. Burkauskas*, J. Brozaitiene, R. Bunevicius, Lithuania . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S92

P.4.032 Neuroinflammation in temporal cortex in schizophrenia patientsT.F. van der Doef*, M. Yaqub, M.G. Bossong, R. Boellaard, A.D. Windhorst, A.A. Lammertsma, R.S. Kahn, B.N.M. vanBerckel, The Netherlands . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S93

P.4.033 Cerebrospinal fluid biomarkers of brain injury in bipolar disorderJ. Jakobsson*, E. Palsson, C.J. Ekman, C. Sellgren, A.G.M. Johansson, H. Zetterberg, K. Blennow, M. Landen, Sweden . . S94

P.4.034 Glucose abnormalities in newly diagnosed, medication-naıve patients with bipolar disorder, mania, and psychosisC. Garcia-Rizo*, B. Kirkpatrick, E. Fernandez-Egea, C. Oliveira, I. Grande, J. Undurraga, E. Vieta, M. Bernardo, Spain . . S95

P.4.035 Clinical improvement and plasmatic concentrations of fluoxetine in major depressive disorder (MDD), obsessive–compulsivedisorder (OCD) and generalised anxiety disorder (GAD)A. Blazquez*, S. Mas, M.T. Plana, A. Lafuente, I. Mendez, L. Lazaro, Spain . . . . . . . . . . . . . . . . . . . . . . . . . . S96

P.4.036 Corticotropin-releasing hormone and serotonin − the neuropsychopharmacology of fear learning deficit from rodents to humansI. Heitland*, L. Groenink, E.Y. Bijlsma, R.S. Oosting, J.L. Kenemans, J.M.P. Baas, The Netherlands . . . . . . . . . . . . . S97

P.4.037 HMGB1 signalling alters T-cell functioning in response to redox status in depressed patients: effect on glucocorticoid receptorfunctionJ. Rybka*, A. Cattaneo, K. Kedziora-Kornatowska, D. Kupczyk, J. Kedziora, Poland . . . . . . . . . . . . . . . . . . . . . . S98

P.4.038 Effect of neuregulin-1 gene functional variant and environmental factors on alcohol use disorderM. Vaht*, E. Kiive, K. Laas, J. Parik, T. Veidebaum, J. Harro, Estonia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S98

Author Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S101Keyword Index . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . S105

S1

Molecular neuropsychopharmacology

Lectures

S.01.01 Induced pluripotent stem cells andin vitro models of neurodevelopmentaldisorders

J. Price1 °. 1King’s College London, Institute ofPsychiatry, London, United Kingdom

There is increasing interest in developing cellular modelsof psychiatric disorders [1]. On the one hand, the idea of acellular model of a complex disorder such as schizophreniaor autism would seem absurd. These are profoundly humanconditions, which surely cannot be reduced to events ina tissue culture dish. Moreover, one is very aware of thecheckered history of animal models in psychiatry. Cancellular models hope for more success, particularly as thepivotal features of these disorders are likely to involvefeatures only emergent at the system level?The prospects for cellular models rest on a series of

technical and scientific advances of recent years. First, thegenetics of autism, schizophrenia, and bipolar disorderhas produced latterly some strong candidates, either fromgenome wide association studies (GWAS), or from copynumber variations (CNVs). This means that we can lookfor the causative features of disease among the phenotypesassociated with specific genetic variants. Second, therehave been a number of pivotal advances in stem cellbiology. Several sources of human stem cells are nowavailable to researchers including somatic and fetal stemcells, such as neural stem cell populations, and pluripotentcells, such as embryonic stem (ES) cells and more recentlyinduced pluripotent (iPS) cells. These cells allow twosignificant advances. First, neural development can berecapitulated in culture. Not completely of course, but stemcells in vitro can enact in vitro many of the seminal featuresof neural development, such as neurogenesis, gliogenesis,axon outgrowth, and synapse formation. This includesmany of the features that are putatively disorganised inneurodevelopmental disorders. Second, by combining ourunderstanding of the genetics with our current ability tomanipulate genomes, we can generate human cells in vitrothat carry precisely the genetic variants that are foundassociated with disease. So, we can assay precisely howthese variants impact development, and thereby entertainprecise hypotheses about the developmental bases of

psychiatric disorders. For example, our laboratory has usedneural progenitor cells taken from the human hippocampusto study the effects of stress factors, such as corticosterone,and antidepressant drugs on hippocampal neurogenesis [2].We have been able reproduce many features of depressionobserved in both patients and animal models, but thetractability of the culture system has allowed us to studythe molecular basis of disease aetiology that would havebeen impossible in whole organism approachesiPSC technology represents the pinnacle of this

approach. This allows researchers to take somatic cellsfrom a patient − typically from skin, hair, or blood −and ‘reprogram’ them to give pluripotent cells, essentiallyindistinguishable in their potential from the pluripotentcells that otherwise exist only at the earliest stages ofembryonic development. These iPSCs can be used toderive neural tissue (or indeed any tissue type) in amanner that largely mimics normal development, andcan be used to study the deviation from normality thataccompanies neurodevelopmental disorders. For example,we have grown hair keratinocytes from autistic patientscarrying microdeletions of SHANK3, a structural proteinof the synapse. We are now actively engaged in studyingsynaptogenesis in neurons derived from these iPSCs andcomparing this to what we see in control cells.Among the phenotypes that can be studied in these

cellular models are cellular phenotypes (the differentiationof neurons, the growth of axons, etc) but also molecularphenotypes, such as gene expression and epigeneticvariation. For example, we have discovered recently thatneural stem cells in vitro show random monoallelic geneexpression [3]. This is a phenomenon whereby some cellsexpress just a single allele at a particular genetic locus.It appears to be random, because some clones of neuralstem cells express one allele, other sister clones fromthe same donor express the alternate allele, while a thirdset of clones are biallelic expressing both alleles. Thechoice seems to be made randomly at some point duringdevelopment, then maintained robustly right through thedifferentiation of the stem cells into neurons and glia.We estimate that probably 5 to 10% of expressed geneshave this property. This is of interest in the aetiologyof psychiatric disorders for two reasons. First, if itoccurs during normal development (which has yet to bedemonstrated), then it would mean that the brain would bea mosaic of clones of cells, each with a unique combination

S2 Molecular neuropsychopharmacology

of mono-allelically expressed genes. Thus each clonewould be subtly different. It is easy to imagine how thiscould drive familiar features such as discordance betweentwins. Second, there is an over representation among thesemono-allelic loci of genes associated with both autism andschizophrenia. We do not yet know how to interpret thisfinding, but it suggests a particular involvement of theseloci in neural development, and a sensitivity of these locito gene dosage.These multiple avenues mean that increasingly disease

aetiology can be approached in tissue culture systems. Howfar this endeavour can go remains to be seen, and the iPStechnology in particular has much promise, but limiteddelivery so far. Surely, much disease pathophysiology willbe beyond the scope of these unsophisticated cellularmodels. Nonetheless, much of the proximal impact ofgenetic variants is likely to appear at the cellular level,and these approaches should aid our understanding of theseearly steps. Finally, the potential of cellular systems as drugdiscovery assays and for target identification and validationhas not escaped the attention of the pharmaceuticalindustry. This could be an important development, giventhe paucity of new therapies in the field of psychiatry.

Reference(s)

[1] Bray, N.J., Kapur, S., and Price, J. 2012. Investigatingschizophrenia in a “dish”: possibilities, potential andlimitations. World Psychiatry 11, 153–155.

[2] Anacker, C., Zunszain, P.A., Cattaneo, A., Car-valho, L.A., Garabedian, M.J., Thuret, S., Price, J.,and Pariante, C.M. 2011. Antidepressants increasehuman hippocampal neurogenesis by activating theglucocorticoid receptor. Mol Psychiatry 16, 738–750.

[3] Jeffries, A.R., Perfect, L.W., Ledderose, J., Schalkwyk,L.C., Bray, N.J., Mill, J., and Price, J. 2012. StochasticChoice of Allelic Expression in Human Neural StemCells. Stem Cells 30, 1938–1947.

Disclosure statement: I act as consultant to ReNeuronLtd., a UK biotech company developing stem cells fortherapeutic and drug discovery applications.

S.01.02 Novel strategies for probing3D structures of GPCRs

F. Marshall1 °. 1Heptares Therapeutics, CSO,Hertfordshire, United Kingdom

G protein coupled receptors are a major class ofcell surface proteins responding to hormones andneurotransmitters. Around 40% of drugs on the market −particularly in the CNS area − mediate their activitythrough GPCRs including drugs such as olanzapine for

schizophrenia, opioids for pain and dopamine agonists forParkinson’s disease, however there have been few recentsuccesses with only 10 new GPCRs being drugged inthe last decade. New GPCRs of interest including thereceptors for neuropeptides and lipids have proved moredifficult to find modulators with good in vivo activity,including selectivity, safety and pharmacokinetics. Highthroughput screening is frequently used to discover startingpoints for GPCR drug discovery, however oftentimes theleads derived from this approach have a high molecularweight and lipophilicity. Such compounds often fail duringclinical development due to poor pharmacokinetics and offtarget toxicity.The use of modern biophysical and structure based

techniques in drug discovery have been very effectivelyapplied to soluble targets such as enzymes. Structuralbiology of membrane proteins including GPCRs has beenhampered by low levels of expression, heterogeneityof the protein, flexibility, the existence of differentconformational states, instability when removed from themembrane and lack of domains for making contacts incrystal lattices. A range of protein engineering strategieshave now been developed which address many of theseissues and have resulted in the structures of over 15 GPCRsbeing solved in the last few years [1].Heptares Therapeutics uses the technique of confor-

mational thermostabilisation to generate GPCRs with anincreased stability in a chosen conformational state [2].These stabilised receptors are known as StaRs. StaRs havebeen produced for over 30 different GPCRs in differentsub families including family A, B and C. An increase inthermostability is measured by solubilising the protein indetergent and heating in the presence of a ligand of definedpharmacology. The melting point or Tm is defined as thetemperature at which 50% of the protein is unfolded. Theaddition of a small number of point mutations results ina pronounced increase in the Tm of as much as 30ºC.As well as having an increased thermostability StaRs canbe purified in harsh short chain detergents that are moreuseful for crystallisation.Fluorescence size exclusion chromatography (FSEC) is

a technique whereby GFP is fused to the C-terminus ofthe receptor. The solubilised protein is analysed by FSECand the shape of the peak eluting from the column isindicative of the quality of the protein. A sharp single peakshows a monodisperse protein suitable for crystallisation.Native GPCRs generally show a broad profile with severalsubpeaks. This is improved by the addition of ligand,thermostabilising mutations as well as modifications tothe protein such as truncations. Prior to crystallisation theprotein construct is optimised by making truncations andremoving sites for post-translational modification. GPCRsare usually expressed in the baculovirus insect cell system

Molecular neuropsychopharmacology S3

and purified on an IMAC Nickel affinity column usinga His-tag at the C terminus. Other expression systemsthat have been used for structural studies with GPCRsinclude yeast, E. coli and mammalian cells. Crystallisationscreens are set up in a wide range of different detergentconditions. Two approaches are routinely used: vapourdiffusion in short chain detergents and lipidic cubicphase (LCP) crystallisation. The lipidic cubic phase isa lipid matrix which produces a more native, membrane-like environment. Extensive screening and optimisationof crystallisation conditions is then required to obtainthe best diffracting crystals. Crystallisation in LCP isfrequently facilitated by making fusion proteins with theGPCRwhich increase crystallisation contacts. For exampleT4 lysozyme may be fused into the third intracellularloop of the receptor. Antibodies may also be included ascrystallisation chaperones. The resulting GPCR crystalsare small and must be taken to specialised microfocussedX-ray beams at the synchrotron. Selection of the ligand forco-crystallisation studies is also important. In the absenceof thermostabilising mutations a high affinity stabilisingligand is required to obtain crystals. The presence ofthermostabilising mutations reduces this requirement andenables multiple co-crystal structures of weaker ligands tobe obtained. This is important when using co-structuresduring the lead optimisation stage of drug discovery.Once the structure is solved a model of the receptor

structure is obtained and this is used for virtual screeningand structure based drug discovery. X-ray structures arehighly enabling for GPCR drug discovery as they allowdrug candidates to be designed which fit efficiently into theligand binding pocket [3]. Compounds can be optimisedfor affinity, kinetics and selectivity using structure basedapproaches. Heptares have applied this approach to arange of CNS discovery projects including an adenosineA2A antagonist for the treatment of Parkinson’s disease,an orexin receptor antagonist for the treatment of sleepdisorders and a highly selective muscarinic M1 agonistfor the treatment of Alzheimer’s disease.

Reference(s)[1] Katritch V, Cherezov V, Stevens RC. (2012). Diversity

and modularity of G protein-coupled receptorstructures. Trends Pharmacol Sci. 33(1):17−27.

[2] Tate CG. (2012). A crystal clear solution fordetermining G-protein-coupled receptor structures.Trends Biochem Sci. 37(9):343−52.

[3] Congreve M, Langmead C, Marshall FH. (2011).The use of GPCR structures in drug design. AdvPharmacol. 62:1−36.

Disclosure statement: I am an employee of HeptaresTherapeutics a company working on GPCR structure baseddrug discovery

Posters

P.1.001 3-Nitropropionic acid induces autophagyby forming mitochondrial permeabilitytransition pore, not by activatingmitochondrial fission

M.E. Solesio1 °, S. Saez-Atienzar2, J. Jordan2,M.F. Galindo1. 1Complejo Hospitalario UniversitarioAlbacete, Unidad de NeuropsicofarmacologıaTraslacional, Albacete, Spain; 2Universidad deCastilla-La Mancha Facultad de Medicina, Grupode Neurofarmacologıa, Albacete, Spain

Purpose of study: Huntington’s disease is a neurodegen-erative process associated with mitochondrial alterations.Inhibitors of the electron-transport channel complex II,such as 3-nitropropionic acid (3NP), are used to studythe molecular and cellular pathways involved in thisdisease [1]. We studied the effect of 3NP on mitochondrialmorphology and dynamics, as well as its involvement inmacrophagy and apoptosis activation.

Methods used: Pharmacological and biochemicalmethods were used to characterise the effects of3NP on autophagy and mitochondrial dynamics andapoptosis, in a dopaminergic cell culture model (SH-SY5Y). 24 h before the transfections, SH-SY5Y cellswere plated. The day after, there were transfected withGFP-LC3, GFP-Drp1 or GFP-Bax to ascertain theirrole and intracellular localisation after 3NP treatment,using confocal microscopy. Mitochondrial morphologywas studied by transfecting our cell cultures with thechimeric protein pDs-Red2Mito and then visualising themusing confocal microscopy. In some experiments, dataobtained by confocal microscopy were corroborated byWB analysis.

Summary of results: Untreated SH-SY5Y cells pre-sented long, tubular and filamentous net of mitochondria.After 3NP (5mM) treatment, mitochondria became shorterand rounder (filamentous mitochondria: 74±5 in controlvs 16±2 in 3NP-treated cells). 3NP induces formationof mitochondrial permeability transition pore, both in cellcultures and in isolated liver mitochondria, and this processwas inhibited by cyclosporine A. After 3 h of treatment,3NP induces autophagy activation (4±2 dotted cells incontrol vs 17±1.2 in 3NP-treated cells). Participation ofthe mitochondrial fission pathway was excluded because3NP did not induce translocation of the dynamin-relatedprotein 1 (Drp1) to the mitochondria (8±2 cells withDrp1 translocated in control vs 11±5 in 3NP treated-cells). The Drp1 inhibitor Mdivi-1 did not affect theobserved changes in mitochondrial morphology, (47±2cells with fragmented mitochondria in control vs 52±3 in


3NP treated cells). After 12 h of 3NP addition, we observedBax mitochondrial translocation, (26±5 Bax traslocationunder control conditions vs 41±2 after 3NP treatment).Finally, scavengers of reactive oxygen species failed toprevent mitochondrial alterations, while cyclosporine A,but not Mdivi-1, prevented the generation of ROS. Datashown are mean±SEM, unless otherwise stated. Statisticalsignificance of differences between groups was determinedby ANOVA followed by a Newman–Keuls post hocanalysis. The level of statistical significance was set atP< 0.05.

Conclusions: There was a direct correlation betweenformation of mitochondrial permeability transition poreand autophagy induced by 3NP treatment. Activationof autophagy preceded the apoptotic process and wasmediated, at least partly, by formation of reactive oxygenspecies and mitochondrial permeability transition pore.3NP induces mitochondrial swelling at short time, thusDrp1 is not able to migrate to the mitochondria. Moreover,cell apoptosis is activated through the intrinsic pathway.

Reference(s)

[1] Beal, M.F., Brouillet, E., Jenkins, B.G., Ferrante, R.J.,Kowall, N.W., Miller, J.M., Storey, E., Srivastava, R.,Rosen, B.R., Hyman, B.T., 1993. Neurochemical andhistological characterisation of striatial excitotoxiclesions produced by mitochondrial toxin 3-nitro-propionic acid. J Neurosci 13, 4181–4192.

Disclosure statement: Work supported by SAF2008–05143-C03−1 from Ministerio de Ciencia e Innovacionand PI2007/55 Consejerıa de Sanidad from Junta deComunidades de Castilla-La Mancha (to JJ) and by‘Incorporacion de grupos emergentes’ FIS CARLOSIII (EMER07/023) and FIS-FEDER (PI080693) andPI11/00736 (to MFG). MES is a FIS-FEDER grant fellow.

P.1.002 Pharmacological characterisation ofpositive allosteric modulators acting onthe metabotropic glutamate receptor 2

K.A. Bennett1 °, A. Weaver1, F. Marshall1,C.J. Langmead1. 1Heptares Therapeutics, Department ofpharmacology, Hertfordshire, United Kingdom

There are eight subtypes of metabotropic glutamate(mGlu) receptors which bind the neurotransmitterglutamate. The mGlu2 and mGlu3 receptors are expressedpre-synaptically in the cortex, thalamus, striatum, amyg-dala and hippocampus. Hyperactivity in glutamatergictransmission in these regions is associated with anxietydisorders and psychosis and may be ameliorated bypharmacological activation of mGlu2. The selectivity and

tolerance issues faced by orthosteric agonists of mGlureceptors are circumvented by allosteric modulators, whichmodulate the affinity (a) and/or efficacy (b) of orthostericligands. Here we show how reported positive allostericmodulators (PAMs) of mGlu2 (BINA, LY487379 and JNJ-40068782) affect the affinity and efficacy of glutamate.Binding studies were established using the orthosteric

antagonist [3H]LY341495 in HEK293 cell membranestransiently expressing mGlu2. A three-way competitionbinding assay was used to study the interaction between[3H]LY341495, glutamate and the PAMs (BINA, 2mM−2nM; LY487379 and JNJ-40068782, 30mM−10 nM)allowing estimation of affinity co-operativity betweenPAM and glutamate (a′). Data was analysed globally usingthe simple allosteric ternary complex model [1,2]. Aninducible mGlu2 expressing Jump-In HEK cell line (LifeTechnologies) was used to measure Gai/o activity usinga commercially available cAMP kit (HTRF; CisBio). Re-ceptor expression was induced with 1mg/ml doxycycline(16 h) and, prior to challenge, 3U/ml glutamate pyruvatetransaminase and 5mM sodium pyruvate was appliedto eliminate endogenous glutamate. When necessary,cells were pre-treated with PAM (15min) before theaddition of glutamate. Data was analysed globally usingthe operational model of allosterism [3] to elucidatenet affinity/efficacy co-operativity parameters (ab) andefficacy co-operativity factors (b).[3H]LY341495 bound with high affinity (Kd ±S.E.M =

0.61±0.03 nM) in mGlu2 expressing cell membranes andwas completely displaced by glutamate (pKi 5.69±0.13)and the orthosteric agonists LY367385 and LY354740 (pKivalues 4.73±0.08 and 7.68±0.15, respectively). Althoughthey did not modulate equilibrium [3H]LY341495 binding(a = 1), BINA, LY487379 and JNJ-40068782 all increasedglutamate affinity (a′; Table 1). Functional analysisrevealed, in this system, all three PAMs behaved aspartial allosteric agonists and were able to increase thepotency of glutamate (glutamate pEC50 4.26±0.09). Thecombined co-operativity values (ab; Table 1) obtainedfrom the functional data were consistent with positiveallosteric modulation (ab> 1). Derivation of b (i.e abilityof PAM to modulate efficacy; Table 1) revealed thatBINA and LY487379 were weak positive modulators ofglutamate efficacy whilst JNJ-40068782 behaved as a‘neutral’ modulator (i.e JNJ-40068782 co-binding did notalter the efficacy of glutamate).Activation of mGlu2/3 receptors negatively feeds back

to reduces glutamate release at glutamatergic synapses.PAMs of mGlu2 have the ability to increase the responseto endogenous glutamate − reducing glutamate release.The results from this study highlight subtle differencesin the ability of mGlu2 PAMs BINA, LY487379 andJNJ-40068782 to modulate the affinity and efficacy of


glutamate. LY487379 equally modulated glutamate affinityand efficacy, BINA modulated affinity to a greater extentthan efficacy whilst JNJ-40068782 acted solely to increaseaffinity of glutamate.

Table 1. Properties of mGlu2 PAMs

Compound Co-operativity inbinding(a′; mean±S.E.M.n = 3)

Net co-operativityeffect(ab; mean±S.E.M.n = 3)

Co-operativity infunction(b; mean±S.E.M.n = 3)

BINA 4.81±0.10 23.44±1.48 2.96±0.37LY487379 2.25±0.25 5.50±1.39 2.11±0.41JNJ-40068782 6.65±0.44 8.95±1.27 1.16±0.13

Reference(s)

[1] Lazareno S, Birdsall NJ., 1995. Detection, quanti-tation, and verification of allosteric interactions ofagents with labeled and unlabeled ligands at G protein-coupled receptors: interactions of strychnine andacetylcholine at muscarinic receptors. Mol Pharmacol.Aug;48(2):362−78.

[2] Bradley SJ, Langmead CJ, Watson JM, Challiss RA.,2011. Quantitative analysis reveals multiple mecha-nisms of allosteric modulation of the mGlu5 receptorin rat astroglia. Mol Pharmacol May;79(5):874−85.Feb 14.

[3] Leach K, Sexton PM, Christopoulos A., 2007.Allosteric GPCR modulators: taking advantage ofpermissive receptor pharmacology. Trends PharmacolSci. Aug;28(8):382−9.

P.1.003 Inflammatory response in the brain ofrats exposed to chronic mild stress

C. Zecchillo1 °, F. Macchi1, R. Molteni1, G. Racagni1,M. Papp2, M.A. Riva1. 1University of Milan, ScienzeFarmacologiche e Biomolecolari, Milan, Italy; 2Instituteof Pharmacology, Polish Academy of Sciences, Krakow,Poland

Major depressive disorder (MDD) is a common disorderthat represents a leading cause of disability in the world. Itis thought to originate from the interaction between suscep-tibility genes and environmental events, such as stress, towhich an individual can be exposed in different momentsof life [1]. One of the major problems of depression isthe relevant percentage of patients who do not show anadequate response to antidepressant therapy, as well as thehigh rate of relapse. Growing evidence suggests that theactivation of the inflammatory/immune system contributesto the pathogenesis of depression. In particular, depression

shows comorbidity with cancer, arthritis rheumatoid, car-diovascular and neurodegenerative diseases, characterisedby inflammatory alterations [2]. In addition, elevatedblood levels of the pro-inflammatory cytokines includinginterleukin (IL)-1b, IL-6 and tumour necrosis factor(TNF)-a are commonly found in depressed patients [3]. Arole for inflammation in depression is also supported by thefindings that cytokine administration induces depressivesymptoms, as occurs in the 30% of hepatitis C patients whoare treated with the immune activator interferon-alpha.On these bases, it is important to characterise the

changes of immune/inflammatory response in animalmodels of depression in order to establish their relationshipwith the depressive phenotype as well as the involvementin antidepressant response.In order to do this we investigated the inflammatory

response of rats exposed to a chronic mild stress (CMS)paradigm, which represents a well-established animalmodel of depression, for 8 weeks. Moreover, a groupof animals (sham or CMS) were chronically treated withthe antidepressant imipramine (10mg/kg/day starting fromweek 2), in order to evaluate the ability of the antidepres-sant treatment to interfere with inflammatory alterations.As expected, chronic mild stress caused a gradual

decrease in the consumption of 1% sucrose solution overthe 8-week period. When investigating the molecularchanges of inflammatory markers, we found that thegene expression of two cytokines, IL-1b and IL-6, wassignificantly increased in the hippocampus of stressedanimals (+49%, p< 0.05; +49%, p< 0.01). Also themRNA levels for CD11b, a marker of microglia activation,were increased after CMS (+83%, p< 0.01). Moreover,we found that chronic imipramine treatment was ableto normalise the depressive phenotype caused by CMSparadigm, although it did not alter the changes of theinflammatory response (IL-1b (+47%), IL-6 (+61%), andCD11b (+119%)).In summary, these data provide support for a link

between inflammation and depression, suggesting thata depressive state may be associated with significantalterations of the inflammatory response in selectedbrain regions. Interestingly, the failure of antidepressanttreatment to normalise the inflammatory changes, whilecorrecting the ‘anhedonic’ phenotype, suggests that clas-sical pharmacological intervention may not work on thefull spectrum of changes that characterise the depressivephenotype. This concept, when translated to humans, maybe relevant for the presence of residual symptoms that areassociated with enhanced the risk of relapse.

Reference(s)

[1] Caspi, A., Sugden, K., Moffitt, TE., Taylor, A., Craig,IW., Harrington, H., McClay, J., Mill, J., Martin, J.,


Braithwaite, A., Poulton, R., 2003. Influence of lifestress on depression: moderation by a polymorphismin the 5-HTT gene. Science. 301(5631):386−9.

[2] Anisman, H., Merali, Z., Hayley, S. 2008. Neurotrans-mitter, peptide and cytokine processes in relation todepressive disorder: comorbidity between depressionand neurodegenerative disorders. Prog Neurobiol.85(1):1−74.

[3] Connor, TJ., Leonard, BE. 1998. Depression, stressand immunological activation: the role of cytokines indepressive disorders. Life Sci. 1998;62(7):583–606.

P.1.004 RNAi-mediated serotonin transportersuppression rapidly increasesserotonergic neurotransmission andhippocampal neurogenesis

A. Ferres-Coy1 °, F. Artigas1, A. Bortolozzi1. 1IIBB-CSIC(IDIBAPS), Neurochemistry and Neuropharmacology,Barcelona, Spain

Despite extensive research, the neurobiology of majordepressive disorder (MDD) remains poorly understooddue to lack of biomarkers, relatively low rates ofheritability, and heterogeneity of precipitating factors,including stress. Serotonin (5-HT) participates in theaetiology and treatment of MDD. In fact, selectiveserotonin reuptake inhibitors (SSRIs) are the most widelyprescribed antidepressant drugs. However, SSRIs takeweeks to months to produce a therapeutic responseand are only moderately effective, leaving more thanone-third of depressed individuals resistant to drugtreatments [1]. The serotonin transporter (SERT) is akey regulator of serotonergic neurotransmission, as itcontrols the active fraction of 5-HT. RNA interference(RNAi) plays a critical role in regulating gene expressionand normal cell development. It also provides newalternatives to pharmacological treatments to modulatebrain neurotransmission through the use of exogenoussmall interference RNA (siRNA). Previously, we showedthe feasibility to silence the expression of 5-HT1A-autoreceptor in 5-HT neurons [2,3]. Here we used amultifaceted approach to assess the ability of RNAi toreduce SERT expression in dorsal raphe of adult mice. Weexamined downstream changes on brain variables linked toantidepressant efficacy and compared SERT-siRNA effectswith those of a standard SSRI (fluoxetine) treatment.Local SERT-siRNA infusion for 4-days (0.7 nmol/dose)suppressed SERT expression in DR (40%). This wasaccompanied by a selective and widespread reduction ofSERT-binding sites and protein levels throughout the brain.Moreover, a 4-day regimen with intra-DR SERT-siRNA

modified brain variables considered to be key markersof antidepressant action, such as: (a) reduced expressionand sensitivity of 5-HT1A-autoreceptors; (b) augmentedextracellular 5-HT in DR-projecting areas such as striatumand hippocampus; (c) increased hippocampal neurogenesis(BdrU-labelled cells to 144±8% of vehicle-treated mice)and (d) increased plasticity-associated gene expression(BDNF, VEGF and Arc) in hippocampus. In contrast,a 4-day regimen with fluoxetine (20mg/kg/day) didnot alter any of these variables and only started tomodify them after 15-day treatments. Furthermore, theintranasal administration of conjugated SERT-siRNA for7-days, but not fluoxetine, reversed the behaviouraldysfunction induced by three weeks of corticosteronetreatment (reduced immobility time in tail suspension test,reduced latency to feed in response to stress in noveltysuppressed feeding test and increased the preference andintake of sucrose solution compared with water). Thesefindings highlight the critical role of SERT in the controlof serotonergic function, including serotonin-mediatedneural plasticity. They also support the use of siRNAtargeting serotonergic genes (SERT, 5-HT1A-autoreceptor)as a new generation of antidepressant therapies with apotential greater efficacy faster onset of action than currenttreatments.

Reference(s)

[1] Wong, M.L., Licinio, J., 2001. Research and treatmentapproaches to depression. Nature Rev Neurosci 2,343–351.

[2] Ferres-Coy, A., Santana, N., Castane, A., Cortes, R.,Carmona, M.C., Toth, M., Montefeltro, A., Artigas, F.,Bortolozzi, A., 2013. Acute 5-HT1A autoreceptorknockdown increases antidepressant responses andserotonin release in stressful conditions. Psychophar-macology (Berl) 225, 61−76.

[3] Bortolozzi, A., Castane, A., Semakova, J., San-tana, N., Alvarado, G., Cortes, R., Ferres-Coy, A.,Fernandez, G., Carmona, M.C., Toth, M., Perales, J.C.,Montefeltro, A., Artigas, F., 2012. Selective siRNA-mediated suppression of 5-HT1A autoreceptors evokesstrong anti-depressant-like effects. Mol Psychiatry 17,612–623.

Disclosure statement: Supported by Spanish Ministry ofScience and Innovation − CDTI’s CENIT program (DEN-DRIA consortium), Project No. CEN-20101023CDTI;Instituto de Salud Carlos III PI10/00290–FEDER andCentro de Investigacion Biomedica en Red de SaludMental (CIBERSAM, P91C).


P.1.005 Role of cannabinoid CB1 receptorsin modulation of dopamine output inthe prefrontal cortex associated withfood restriction in rats

V. Licheri1 °, G. Talani2, L. Dazzi1, F. Biggio1, C. Utzeri1,V. Lallai1, S. Lutzu1, G. Biggio1,2, E. Sanna1,2.1University of Cagliari, Life and Environmental Sciences,Section of Neuroscience, Monserrato (CA), Italy; 2Instituteof Neuroscience National Research Council, Life andEnvironmental Sciences, Monserrato (CA), Italy

An extensive body of literature has documented theimportance of endocannabinoid system in the regulationof appetite and feeding behaviour in mammals. Thecannabinoid CB1 receptor is abundant in the prefrontalcortex (PFC), which plays key roles in working memoryand reward. Principal dopaminergic afferents arisingfrom the ventral tegmental area (VTA) are crucial forPFC function and dopamine type 2 (D2) as well asCB1 receptors are co-expressed on g-aminobutyric acid(GABA)-containing terminals in the PFC [2].To elucidate the role of CB1 receptors in the regulation

of dopamine release in the PFC associated with feedingbehaviour in rats, we exposed Sprague-Dawley rats to arestricted food regimen, with availability of food limitedto a 2-h period daily for 3 weeks. Control rats were givenad libitum access to food. Microdialysis, Western-blot aswell as patch clamp experiments were performed in orderto test our hypothesis.Food-restricted (FR) rats showed a marked increase

in the extracellular dopamine concentration in the PFC80min before food presentation, with the concentrationpeaking during food consumption and returning to baselineafter food removal. These changes were attenuated by theCB1 antagonist SR141716A but were unaffected by theagonist WIN 55212-2. Patch clamp experiments performedin principal medial PFC (mPFC) neurons revealed thatduring the anticipatory phase before food presentationthere is a significant decrease of the inhibitory effect ofWIN 55212-2 on GABAergic spontaneous inhibitory post-synaptic currents (sIPSC) frequency in FR rats comparedwith control animals (one-way ANOVA: p = 0.017 vscontrols). Moreover, endocannabinoid-dependent DSI ofGABAergic inhibition was abolished in FR rats duringthis phase (one-way ANOVA: p = 0.001 vs controls). Thebasal sIPSC frequency was also reduced in mPFC neuronsof FR rats compared with control animals, suggestive ofan altered control of presynaptic GABA release (one-wayANOVA: p = 0.0001 vs controls). A complex innervationsbetween mPFC and VTA has been previously described byother authors. Briefly, evidence suggests that endocannabi-

noids may modulate dopamine release in the PFC indi-rectly by activating CB1 receptors located on presynapticGABAergic terminals, thereby inhibiting GABA releaseand resulting in disinhibition of glutamatergic neuronsthat project to the VTA. An increase in glutamate releasein the VTA would stimulate dopaminergic neurons thatproject back to the mPFC and thereby increase the releaseof dopamine in this region [3]. Current-clamp recordingsrevealed an increased excitability in both mPFC and VTAneurons of FR animals, relative to control animals, aneffect that correlates well with the decrease of GABArelease observed in mPFC inhibitory synapses (t-test:p< 0.05 vs controls). Finally, CB1 expression in the PFCwas reduced in FR rats before food presentation comparedwith controls accordingly with other previews reports [1].Together, these data support a role for the endocannabi-

noid system in regulation of dopamine release in the PFC,and they suggest that the feeding-associated increase indopamine output in the PFC of FR rats might be due todown-regulation of CB1 in this brain region that in turnregulates the function of the whole meso-cortical neuronalcircuitry.

Reference(s)[1] Bello, N.T., Coughlin, J.W., Redgrave, G.W., Laden-

heim, E.E., Moran, T.H., Guarda, A.S, 2012. Dietaryconditions and highly palatable food access alter ratcannabinoid receptor expression and binding density.Physiol Behav 105: 720–726.

[2] Chiu, C.Q., Puente, N., Grandes, P., Castillo, P.E.,2010. Dopaminergic modulation of endocannabinoid-mediated plasticity at GABAergic synapses in theprefrontal cortex. J Neurosci 30: 7236–7248.

[3] Egerton, A., Allison, C., Brett, R.R., Pratt, J.A., 2006.Cannabinoids and prefrontal cortical function: insightsfrom preclinical studies. Neurosci Biobehav Rev 30:680–695.

P.1.006 Oxytocin controls CRF gene expressionthrough TORC3: implications forstress-related disorders

B. Jurek1 °, D.A. Slattery1, Y. Liu2, I.D. Neumann1,G. Aguilera2, E.H. van den Burg1. 1Universityof Regensburg, Dept of Neurobiology and AnimalPhysiology, Regensburg, Germany; 2Eunice KennedyShriver National Institute of Child Health and HumanDevelopment, Section on Endocrine Physiology, BethesdaMD, USA

Increased fear and stress responses following a threateningstimulus are considered normal emotional and physiolog-ical reactions. However, they become pathological when


exaggerated or prolonged, and can then lead to severepsychological and somatic disorders including depressionand anxiety. The neuropeptide corticotropin-releasingfactor (CRF), which is produced in the hypothalamus, isa key regulator of both fear/anxiety and stress responses/reactions. It controls the hypothalamo–pituitary–adrenal(HPA)-axis to stimulate the release of the stress hormonecortisol or corticosterone via ACTH. Its expression inturn is regulated by the transcription factor CREBand its cofactor TORC (transducer of regulated CREBactivity, a.k.a. CRTC) that, upon an appropriate stimulus,translocates from the cytoplasm to the nucleus to form acomplex with CREB and activate gene transcription bybinding to a promoter region [1].We have shown that another neuropeptide, namely

oxytocin, is anxiolytic within the hypothalamus, andhas anti-stress properties. The underlying molecularmechanisms of these effects are unknown, yet areimportant for the quest for pharmaca that may dampen(exaggerated) stress responses. As CRF cells expressthe oxytocin receptor, oxytocin might inhibit HPA-axisactivity directly through altering CRF gene expressionor CRF release. Therefore, in the present study, wehypothesised that oxytocin inhibits CRF gene transcriptionthrough prevention of CREB phosphorylation and/orTORC translocation and assessed this using a combinationof in vivo and in vitro assays.Icv administration of oxytocin in male rats delayed

and prolonged restraint stress-induced CRF transcription,as measured by hnRNA levels using quantitative real-time PCR. CRF hnRNA levels were significantly lowerin oxytocin-infused rats when compared with vehicle-infused rats 10min after the onset of the stressor(p< 0.05, n = 4). In contrast, while CRF hnRNA levelsreturned to basal values in the vehicle-treated group after30min, they remained elevated in the OXT-treated group(p< 0.05, n = 8). In line with these in vivo findings,Western blot analyses using primary hypothalamic neuronsshowed that the oxytocin receptor agonist TGOT loweredthe maximum CRF hnRNA response to forskolin by20% (p< 0.05, n = 7) after 30min of stimulation, butprolonged forskolin-induced CRF expression (up to 90minof stimulation; p< 0.05, n = 7). In a hypothalamic cellline (H32), TGOT did not influence forskolin-inducedCREB phosphorylation nor TORC2 nuclear levels atany timepoint tested, but delayed the forskolin-inducedincrease of nuclear TORC3 concentrations from 10 to60min of incubation (10min: p< 0.05; 60min: p< 0.001;n = 4−8). TGOT/oxytocin without forskolin/stress showedno effect on any parameter tested.These results indicate that oxytocin controls CRF gene

expression by modulating nuclear TORC3 levels, thusdecelerating CRF synthesis during the initial phase of

an acute stress response. Indeed, this was mirrored via adelay in peak plasma ACTH levels from 10min in thevehicle-treated group to 15min in the oxytocin-treatedgroup (interaction factor time × treatment; F(3,28) = 3.372,p< 0.05). Taken together, our findings indicate thatoxytocin acts on the CRF system to modulate the acuteHPA-axis response to a stressor. Therefore, drugs, such asoxytocin, that target TORC3 activity may represent novelcandidates for the treatment of stress-related disorders.

Reference(s)

[1] Liu, Y., Knobloch, S., Grinevich, V., Aguil-era, G., 2011. Stress induces parallel changes incorticotrophin-releasing hormone (CRH) transcriptionand nuclear translocation of transducer of regulatedcAMP response element-binding activity 2 in hypotha-lamic CRH neurones. J Neuroendocrinol, 216–223.

P.1.007 A proteomic and functional analysisreveals that 5-HT6 receptorsmodulate neuronal differentiationby recruitment of Cdk5

F. Duhr1 °, M. Seveno2, C. Mannoury la Cour3,D. Dupuis3, M.J. Millan3, J. Bockaert1, P. Marin1,S. Chaumont-Dubel1. 1Institut de GenomiqueFonctionnelle (IGF), Neurobiologie, Montpellier, France;2Institut de Genomique Fonctionnelle (IGF), Plateformede Proteomique Fonctionnelle, Montpellier, France;3Institut de Recherche Servier, Psychopharmacologie,Croissy sur Seine, France

Purpose of the Study: The serotonin 5-HT6 receptor isexpressed in CNS regions involved in the pathogenesisof disorders like Alzheimer’s disease and schizophrenia.5-HT6 receptors are a promising target for treatmentof the accompanying cognitive deficits, since theirblockade consistently enhances mnemonic performance inrodents [1]. Paradoxically, still little is known about 5-HT6receptor-associated signalling pathways, an issue we haveaddressed by a proteomic approach. Previously, we showedphysical association of the 5-HT6 receptor with severalmembers of the mTOR pathway and demonstrated thatmTOR recruitment by prefrontal 5-HT6 receptors perturbscognition in schizophrenia [2]. Here we show that 5-HT6receptors interact with Cyclin-dependent kinase 5 (Cdk5),a protein which controls actin cytoskeleton dynamics andmodulates neurodevelopmental processes such as neuronmigration, neurite outgrowth, synaptogenesis and dendriticspine morphogenesis.

Methods and Results: The present study explored therole of Cdk5, under the control of 5-HT6 receptors, in


the differentiation of NG108−15 neuroblastoma cells andrat striatal neurons in primary culture, assessed by neuriteoutgrowth. Expression of voltage-gated Ca2+ channels wasalso analysed in NG108−15 cells using Fura2 imaging. Theunpaired Student’s t-test and one-way ANOVA followed byDunnett’s test were performed (using the Prism software)for two-sample and multiple comparisons, respectively.Expressing 5-HT6 receptors in NG108−15 neuroblastomatriggered neurite outgrowth and induced expression offunctional voltage-gated Ca2+ channels. These effectswere not further enhanced by an agonist (WAY181,187,1mM) but were prevented by SB258,585 (10mM),a selective 5-HT6 antagonist. Thus, upon SB258,585treatment, cells showed a decrease in neurite length of48.4% compared to untreated cells (22.5±1.5mm vs.43.5±1mm, n = 4, p< 0.0001). Expression of a dominant-negative (DN) Cdk5 or treating cells with roscovitine(pharmacological inhibitor of Cdk5) likewise inhibitedNG108−15 cells differentiation induced by 5-HT6 receptorexpression (control: 69.8±3.6mm; DN Cdk5: 42.8±3mm,p< 0.0001; roscovitine: 28.1±1.5mm, n = 3, p< 0.0001).SB258,585 also impaired the association of 5-HT6receptor with Cdk5 in NG108−15 cells, as determined byco-immunoprecipitation, suggesting that this interactionwas necessary for induction of differentiation. Treatingstriatal neurons with either SB258,585 or roscovitineimmediately after seeding decreased neurite length, as de-termined 24 hrs later (untreated: 29.9±0.7mm; SB258,585:23.7±0.6mm, p< 0.0001; roscovitine: 20.2±1.0mm,n = 4, p< 0.0001). Conversely, exposure of neurons toWAY181,187 did not significantly affect neurite outgrowth(p = 0.43). Further supporting a role of endogenouslyexpressed receptors in differentiation of striatal neurons,silencing 5-HT6 receptor expression in cultured neuronssignificantly reduced neurite length (control siRNA:29.6±0.8mm, 5-HT6 receptor siRNA: 17.2±0.9mm, n = 3,p< 0.0001).

Conclusions: Complementing work indicating that5-HT6 receptors modulate neuronal migration [3], thepresent data show that 5-HT6 receptors developmentallypromote neuronal differentiation and reveal a criticalrole for Cdk5 in this process. These novel insightsinto molecular substrates underlying neurodevelopmentaleffects of 5-HT6 receptors are of potential importanceto the pathophysiology and treatment of early-onsetCNS conditions like autism-spectrum disorder andschizophrenia.

Reference(s)

[1] Mitchell, E.S., and Neumaier, J.F. 2005 5-HT6receptors: a novel target for cognitive enhancement.Pharmacol Ther 108, 320–333.

[2] Meffre, J., Chaumont-Dubel, S., Mannoury laCour, C., Loiseau, F., Watson, D.J., Dekeyne, A.,Seveno, M., Rivet, J.M., Gaven, F., Deleris, P.,Herve, D., Fone, K.C., Bockaert, J., Millan, M.J.,and Marin, P. 2012 5-HT(6) receptor recruitment ofmTOR as a mechanism for perturbed cognition inschizophrenia. EMBO Mol Med 4, 1043–1056.

[3] Riccio, O., Potter, G., Walzer, C., Vallet, P., Szabo, G.,Vutskits, L., Kiss, J.Z., and Dayer, A.G. 2009 Excessof serotonin affects embryonic interneuron migrationthrough activation of the serotonin receptor 6. MolPsychiatry 14, 280–290.

P.1.008 V1B/CRF1 receptor heterodimerisationas a key mechanism of vasopressin andcorticotropin-releasing factor synergism

J. Mion1 °, V. Boulay1, M.J. Millan2, G. Guillon1,M. Corbani1. 1Institut de Genomique FonctionnelleCNRS UMR 5203 − INSERM U.661 Universites deMontpellier I et II, Endocrinologie, Montpellier, France;2Institut de Recherches SERVIER, Unite de Recherche etDecouverte, Croissy-sur-Seine, France

Purpose of the study: Vasopressin (AVP) andCorticotropin-Releasing Factor (CRF) are involved in thestress response, mainly by regulating ACTH secretionfrom the pituitary and by increasing catecholamineand corticosteroid secretion from the adrenal medulla.In these structures, AVP and CRF act in synergismvia V1B and CRF1 receptors, respectively. Recently,we suggested that synergism operates via both secondmessenger crosstalk and putative mechanism involvingreceptors heterodimerisation [1]. To further validate thislast original mechanism, we monitored the influence ofreceptor heterodimerisation selectivity and of receptorheterodimerisation disruption on functional synergism.

Methods: We co-expressed in HEK293 cells eitherCRF1 or CRF2 receptors on one hand, and either V1Bor V1A receptors on the other hand. For each pair oftransfected receptors, the degree of physical interactionwas evaluated by BRET experiments. AVP receptors weretagged with the Renilla Luciferase and CRF receptors weretagged with Yellow Fluorescent Protein at their C-terminaldomain. In contrast to a previous study by Young et al. [2],tagging the receptors at their C terminus preserved theirfunctionality and was a key element for the success of thisapproach. For each pair of transfected receptors, functionalsynergism was measured by respective second messengeraccumulation: Inositol Phosphates (IPs) for V1B and V1Aand cAMP for CRF1 and CRF2). We also generated amutant of CRF1 receptor unable to heterodimerise with


V1B receptor and looked for its ability to induce functionalsynergism.

Results: Only for V1B/CRF1 receptors, there wasa clear association between receptor heterodimerisation(BRETmax reach from x1 = Fluorescence/Luminescence =0.005 to x2 = 0.02) and crosstalk-independent potentiationin both IPs (27% under 10 nM AVP and 10 nM CRF)and cAMP (29% under 1 nM CRF and 10 nM AVP)signalling pathways. By contrast, for co-transfection ofV1B and CRF2 receptors, no molecular association couldbe observed (no saturation curve) and hormone-stimulatedsecond messenger potentiation only reflected signallingcrosstalk. Evidence for the existence of V1A/CRF1heterodimers was found (x1 = 0.015; x2 = 0.03) but theyconformationally differed to V1B/CRF1 heterodimers,and potentiation of signalling independent of secondmessenger crosstalk could only be observed for formationof IPs (13% under 10 nM AVP and 10 nM CRF).To further correlate heterodimerisation and crosstalk-

independent potentiation, we generated a chimericalCRF1-CRF2 receptor. BRET experiments show that thismutant has lost capacity to heterodimerise with V1Breceptors. Moreover, despite it still activated cAMPpathway in a dose-dependent manner, this CRF1 mutantwas no longer able to potentiate vasopressin-stimulatedIPs accumulation via a mechanism independent of secondmessengers crosstalk.

Conclusion: These results suggest that AVP-CRFsynergism at least partially reflects the existence ofV1B/CRF1 heterodimers, and that their formation modifiessignalling at these sites. Studies underway are in linewith the existence of molecular physical association ofV1B/CRF1 heterodimers in native tissues. Further, thepresent work is consistent with a role for V1B/CRF1heterodimers controlling ACTH secretion − and theactivity of corticolimbic structures controlling mood − inthe pathogenesis of stress-related psychiatric conditionslike depression, anxiety disorders and schizophrenia.

Reference(s)

[1] Murat, B., Devost, D., Andres, M., Mion, J., Boulay, V.,Corbani, M., Zingg, H.H., Guillon, G., 2012 V1band CRHR1 receptor heterodimerization mediatessynergistic biological actions of vasopressin and CRH.Mol Endocrinology 26(3), 502−20.

[2] Young, S.F., Griffante, C., Aguilera, G., 2007 Dimer-ization between vasopressin V1b and corticotropinreleasing hormone type 1 receptors. Cell MolNeurobiol 27, 439–461.

Disclosure statement: Work supported by the ANR,SERVIER (France) and the FRM.

P.1.009 Cell synchronisation as a tool tooptimise expression of metabotropicglutamate receptors in induciblemammalian expression system

B. Chruscicka1 °, P. Branski1, G. Burnat1, A. Pilc1.1Polish Academy of Sciences, Neurobiology, Krakow,Poland

Background: The metabotropic glutamate receptors(mGluRs) are class C G protein-coupled receptors thatplay an important neuromodulatory role throughout thebrain, as such they are involved in a number of psychiatricand neurological disorders like anxiety, depression andschizophrenia [1]. Expression of metabotropic glutamatereceptors in heterologous mammalian cells is a methodfor their functional characterisation, and tool to study theagonist, antagonist and allosteric effects which are veryattractive targets for therapeutic intervention. However, theconstitutive expression of mGluRs seems to have a toxiceffect on the HEK293 cells. Thus, an inducible expressionsystem (in which the mGluRs expression is activated inthe presence of tetracycline) is used [2]. Unfortunately anexpression of receptors was confirmed in approximately30% of cells after tetracycline induction.

Aim: Our goal was to increase the number of HEK293cells with an induced expression of mGlu2 receptor usingsynchronisation of cell cycle.

Methods: cDNA fragment encoding mGlu2 receptorwas cloned into a pcDNA5/FRT/TO vector under thecontrol of a tetracycline regulated promoter. HEK293 cellscontaining inducible mammalian expression system (Flp-In T-REx host cells line from Invitrogen) were treatedwith GeneJuice transfection reagent, and then selected byhygromycin. Synchrony of stably transfected cells wasperformed by means of three different synchronisationprotocols. To obtain G2/M phase block, cells weresupplemented with nocodazole for 18 h. To arrest cellsat the beginning of S phase a double thymidine blockwas used (16 h and 18 h). G0/G1 population of cells wasobtained by serum starvation for 48 h [3]. The efficacyof cell synchronisation was verified by flow cytometricanalysis of DNA content after propidium iodide-staining.For evaluation a surface expression of mGlu2 receptor flowcytometer immunofluorescence staining was performed.

Results: We received 45.2±0.9% cells at the beginningof S phase after double thymidine block, 60.6±0.9% cellsin M phase after nocodazole treatment and 77.9±8.0%cells in G0/G1 phase after serum starvation. Untreatedcells are characterised by 52.7±1.8% cells in G0/G1,31.3±0.9% in S and 15.0±1.0% in G2/M phase ofcycle. Cell synchronisation seems to have statistically


significant effect on mGlu2 receptor expression level[One-way ANOVA, F(3.41) = 22.8, p< 0.0001]. MoreoverDunnett’s Multiple Comparison Test analysis shows thatlevel of mGlu2 receptor surface expression in cells afterdouble thymidine block was significantly higher comparedto receptor expression level in asynchronously growingcells (p< 0.01). In contrast, the expression level of mGlu2receptor was significantly decreased in M phase blockedcells (p< 0.01) and cells after serum starvation (p< 0.01).

Conclusions: The results indicate that modification ofthe inducible expression system by cell synchronisationseems to have significant effect on surface mGluR2expression level.

Reference(s)

[1] Palucha, A., Pilc, A., 2007. Metabotropic glutamatereceptor ligands as possible anxiolytic and antidepres-sant drugs. Pharmacology & Therapeutics 115, 116–147.

[2] Van Craenenbroeck, K., Vanhoenacker, P., Ley-sen, J.E., Haegeman, G., 2001. Evaluation of thetetracycline- and ecdysone-inducible systems for ex-pression of neurotransmitter receptors in mammaliancells. European Journal of Neuroscience 14, 968–976.

[3] Yu, J.N., Ma, S.F., Miao, D.Q., Tan, X.W., Liu, X.Y.,Lu, J.H., Tan, J.H., 2006. Effects of cell cycle status onthe efficiency of liposome-mediated gene transfectionin mouse fetal fibroblasts. Journal of Reproductionand Development 52, 373–382.

P.1.010 The identification of protein tyrosinephosphatase, non-receptor type 1in hippocampal modulation of foodanticipatory behaviour

E. Kostrzewa1 °, L.A.W. Verhagen1, C. Gelegen1,H.A. van Lith2, D.A. Collier3, M. Mitsogiannis1, E. deVries1, M. van Gestel1, R.A. Adan1, M.J.H. Kas1.1Rudolf Magnus Institute of Neuroscience, Neuroscienceand Pharmacology, Utrecht, The Netherlands; 2UtrechtUniversity, Faculty of Veterinary Medicine, Utrecht, TheNetherlands; 3King’s College, Institute of Psychiatry,London, United Kingdom

Background: Activity based anorexia (ABA) is a rodentmodel of hyperactivity evoked by food restriction.The paradoxical hyperactivity during food restriction isreminiscent of that observed in patients suffering fromeating disorders. One of the parameters of this model,food anticipatory activity (FAA) enables us to studyneurobiological mechanisms that synchronise the timing

of food availability and changes in arousal in order to beprepared for upcoming relevant environmental events [1].

Methods: To identify chromosomes contributing toFAA, 21 mouse strains from a chromosome substitutionpanel (host strain expressing FAA (C57BL/6J) and adonor strain lacking FAA (A/J)) were tested using theABA model (n = 8−10 mice per strain). Subsequently, wegenerated an F2 population from chromosome substitutionstrains 2 that lacks FAA. F2 mice (n = 125) weregenotyped and phenotyped to fine map a locus onmouse chromosome 2 associated with FAA. In addition,genome-wide micro-array gene expression analysis wasperformed to identify differentially expressed genes in thehippocampus of F2 mice selected for their high or lowFAA-levels. To functionally validate the candidate gene wegenerated shRNA viral vector (pAAV-shbase containingGFP cassette, titer 7.8×1012 genomic copies/ml) andinjected C57BL/6J mice in the dentate gyrus (DG) threeweeks prior to the ABA model. We verified virus spreadusing ISH for GFP and assessed the potential neuronaldamage by performing IHC for NeuN and ISH formiRNA124. Level of candidate gene knockdown wasquantified by radioactive ISH. Animals’ body weightchanges, food intake and running wheel activity (RWA)during ABA were recorded.

Results: We genetically mapped a Quantitative TraitLocus on mouse chromosome 2 associated with FAA.The combined genetic fine mapping and genome-widegene expression data revealed Ptpn1 (protein tyrosinephosphatase, non-receptor type 1) as a candidate genefor FAA. To validate this candidate gene, we performedstereotactic injections of a virus containing anti-Ptpn1shRNA. The virus infected neurons in DG and partly in CAfields without detectable neuronal damage. The expressionof Ptpn1 in DG was significantly reduced in the anti-Ptpn1 group in comparison to the control virus injectedgroup (Student t-test: t(8) = 2.50, p< 0.05). There were nodifferences in body weight, food intake nor baseline RWA(Student t-test: t(8) = −0.493, p = 0.635) between anti-Ptpn1 and control virus treated mice. Similarly, total RWAduring the restriction phase did not differ significantlybetween the groups (Student t-test: t(8) = 0.126, p = 0.903respectively). Normalised FAA, (total FAA as a percentageof total RWA), was found to be significantly decreasedin the Ptpn1 knockdown group compared to the controlgroup (Student t-test: t(8) = 2.459, p< 0.05). A positiveassociation was found between the level of Ptpn1 mRNAexpression in the DG and the magnitude of FAA(r = 0.642, p = 0.045).

Conclusions: This data confirm that FAA levels areassociated with Ptpn1 gene expression in the DG. As Ptpn1is a key regulator of insulin and leptin receptor sensitivity,these findings provide a possible mechanism through


which metabolic hormones mediate the timing of arousalnecessary to be prepared for upcoming environmentalevents.

Reference(s)

[1] Kas, M.J., Adan, R.A., 2011. Animal models of eatingdisorder traits. Curr Top Behav Neurosci 6:209−27.

P.1.011 Identification of a significant rolefor the ventral hippocampus inneuropeptide S-elicited anxiolysis

I.A. Ionescu1 °, J. Dine2, J. Stepan2, Y.C. Yen3,L. Herrmann1, F. Holsboer4, C.T. Wotjak3, R. Landgraf5,M. Eder2, U. Schmidt1. 1Max-Planck-Institut furPsychiatrie, RG Molecular Psychotraumatology,Munchen, Germany; 2Max-Planck-Institut fur Psychiatrie,RG Neuronal Networks Dynamics, Munchen, Germany;3Max-Planck-Institut fur Psychiatrie, RG NeuronalPlasticity, Munchen, Germany; 4Max-Planck-Institut furPsychiatrie, MPI of Psychiatry, Munchen, Germany;5Max-Planck-Institut fur Psychiatrie, RG BehavioralNeuroendocrinology, Munchen, Germany

Purpose of the study: Past research increasingly pointstowards a dual role of the hippocampus, where the dorsalhippocampus is involved in memory-related tasks and theventral hippocampus (VH) modulates emotional processes,including fear and anxiety. Whereas pharmacologicaltargeting of the VH has been associated with anxiogeniceffects, reports on VH-mediated anxiolysis are still scarce.Here, we describe for the first time a role for the VH in theanxiolytic effects of neuropeptide S (NPS), a neuropeptidewith a high potential as a novel therapy for anxietydisorders such as posttraumatic stress disorder (PTSD).

Methods: NPS uptake in the hippocampus was investi-gated by intranasal administration of fluorophore-labelledNPS. Effects of NPS treatment on hippocampal expressionof candidate proteins involved in the glutamatergic systemand in synaptic plasticity were examined by real-time PCRand immunoblotting in C57BL6/N mice as well as intwo mouse models of anxiety disorders, namely the highanxiety behaviour (HAB) mice and a mouse model ofPTSD. To elucidate the effects of NPS treatment on VHphysiology, field potential recordings were conducted inVH slices. The impact of NPS treatment on neuronalactivity flow in the VH was measured using voltage-sensitive dye imaging (VSDI). Finally, the role of the VHin behavioural effects of NPS was examined by intra-VHinjection of NPS or vehicle and monitoring of locomotionin the open field and of anxiety-like behaviour in thelight-dark test and in the elevated plus maze (EPM).

Results: Our experiments showed uptake of fluores-cently labelled NPS in the CA1, CA2 and CA3 regionsand in the dentate gyrus (DG) of the murine hippocampus,indicating hippocampal expression of the functional NPSreceptor (NPSR). On a molecular level, intranasal NPStreatment modulates hippocampal expression of the GluR1subunit of AMPA receptors, of the glutamate transporterGlt-1 and of synapsin isoforms differentially in C57BL6/Nmice and in HAB mice [1]. Additionally, NPS increaseshippocampal synapsin expression in the mouse model ofPTSD, where synapsin expression had previously beenfound decreased [2]. On a physiological level, intranasaladministration of NPS led to decreased paired-pulsefacilitation and long-term potentiation (LTP) at CA3-CA1synapses in C57BL6/N mice as well as in HAB mice.Moreover, in the VSDI assay, bath application of NPS toVH slices resulted in weakened stimulus-evoked activityflow from DG to CA1. Finally, intra-VH injection of NPSsignificantly decreased anxiety-like behaviour on the EPM.

Conclusions: Here, we identified the VH as a novel tar-get region of NPS that had not been previously associatedwith NPS-mediated anxiolysis. In the hippocampus, NPStreatment may contribute to modulation of hippocampalsynaptic plasticity by regulating expression of proteinsinvolved in the glutamatergic circuit and in synapticplasticity. Furthermore, we prove that targeting the VH isalso sufficient to mediate NPS-elicited anxiolysis in vivo,possibly by modulating amygdala activity via bidirectionalconnections to and from the ventral CA1 region. Inconclusion, our data point towards an important role ofthe hippocampus in mediating anxiolytic effects of NPS.

Reference(s)

[1] Ionescu, I.A., Dine, J., Yen, Y.-C., Buell, D.R.,Herrmann, L., Holsboer, F., Eder, M., Landgraf, R.,Schmidt, U., 2012. Intranasally administered Neu-ropeptide S (NPS) exerts anxiolytic effects followinginternalization into NPS receptor-expressing neurons.Neuropsychopharmacology 37, 1323−37.

[2] Herrmann, L., Ionescu, I.A., Henes, K., Golub, Y.,Wang, N.X.R., Buell, D.R., Holsboer, F., Wotjak, C.T.,Schmidt, U., 2012. Long-lasting hippocampal synapticprotein loss in a mouse model of posttraumatic stressdisorder. PLoS One 7, e42603.


P.1.012 Stress-induced vulnerability ofpresynaptic glutamatergic terminals andeffect of desipramine

N. Nava1 °, M. Popoli2, L. Musazzi2, G. Wegener1,J.R. Nyengaard3. 1Center for Psychiatric Research,Aarhus University, Aarhus, Denmark; 2Laboratoryof Neuropsychopharmacology and FunctionalNeurogenomics, University of Milano, Milano, Italy;3Stereology & Electron Microscopy Laboratory, AarhusUniversity, Aarhus, Denmark

Background: Growing evidence has highlighted a deepimpairment in stress-related disorders of limbic structuressuch as amygdala, hippocampus and prefrontal cortex,where glutamatergic transmission is considered to beprevalent [1,2].A number of studies have shown the cumulative

effects of stress and its major mediators, glucocorticoids,on brain volume and dendritic remodelling, in bothhumans and rodents. Nevertheless, very little is knownon the ultrastructural changes exerted by behaviouralstress on glutamatergic synapses responsible for neuronalcommunication. Excitatory synapses provide synapticcommunication by neurotransmitters which are storedwithin the presynaptic terminal in morphologically distinctpools of vesicles, namely the readily-releasable pool ofvesicles (RRP), docked to the active zone and readyfor release, and the reserve pool of vesicles. When aneurotransmitter is released, exchange of information takesplace through interaction of glutamate with receptorssitting on the post-synaptic density.Alterations of such synaptic ultrastructure might result

in impairment of glutamatergic release and transmission.Aim: The main goal of the present study was

therefore to shade light on the consequences of acutebehavioural stress and chronic antidepressant treatmenton the ultrastuctural features of asymmetric terminals,commonly regarded as excitatory, within the medialprefrontal cortex (mPFC).

Methods: Rats were treated chronically with eitherdesipramine (DMI) or vehicle (2 weeks). At the endof the treatment, animals were subjected to acute Foot-Shock (FS) stress and soon after they were deeplyanaesthetised and perfused. mPFC subareas (prelimbicarea, dorsal anterior cingulate and medial precentral area)were identified, based on their noticeable cytoarchitecturalcharacteristics, and overall volume quantified with theCavalieri estimator [3]. Through serial section electronmicroscopy, asymmetric synapses were identified and thenumber of vesicles in the readily-releasable pool (RRP)and the reserve-pool estimated. Extension of the post-

synaptic density as well as of the active zone area wasmeasured; presynaptic terminal volume was also assessed.

Results: Acute behavioural stress induced a strongincrease in the number of vesicles docked to thepresynaptic membrane and ready for release, which wasprevented by chronic treatment with DMI. Post-synapticdensity area was strongly reduced after acute FS-stress;previous treatment with DMI only partially preventedthis change. Moreover, following acute stress, the volumeof presynaptic glutamatergic terminals was reduced. Noeffect was observed on the volume of medial prefrontalcortex.

Conclusions: In the present study we showed that acutestress was able to significantly change the ultrastructure ofglutamatergic terminals, by inducing a large increase in thenumber of the RRP vesicles, as well as decreasing the areaof the post-synaptic density and the volume of the wholepresynaptic terminal. Chronic treatment with DMI seemedto only partially prevent such stress-induced changes.Identifying the effects of stress on excitatory transmissionmay provide further knowledge for developing drugsdirectly targeting the glutamatergic system.

Reference(s)

[1] Musazzi, L., Milanese, M., Farisello, P., Zappettini, S.,Tardito D., Barbiero, V.S., Bonifacino T., Mallei A.,Baldelli P., Racagni G., Raitieri M., Benfenati F.,Bonanno G., Popoli M., 2010. Acute stress increasesdepolarization-evoked glutamate release in the ratprefrontal/frontal cortex: the dampening action ofantidepressants. PloS One 5(1):e8566.

[2] Ansell E.B., Rando K., Tuit K., Guarnaccia J., SinhaR., 2012. Cumulative adversity and smaller graymatter volume in medial prefrontal, anterior cingulate,and insula regions. Biol Psychiatry 72:57−64.

[3] Van Eden C.G., Uylings H.B. 1985. Cytoarchitectonicdevelopment of the prefrontal cortex in the rat. J CompNeurol 241:253−67.

P.1.013 Chronic (−)cannabidiol producesantidepressant-like effects inbulbectomised mice, acting on5-HT1A and CB1 receptor functionality

R. Linge1 °, A. Pazos1, A. Diaz1. 1University Cantabria/CIBERSAM, Instituto de Biomedicina y Biotecnologia deCantabria, Santander, Spain

Depression is one of the most prevalent psychiatricdisorders in our society. The classic antidepressant drugs,such as the selective serotonin reuptake inhibitors (SSRIs),act raising the monoamine levels in different regions. In


spite of the fact that >90% these antidepressants enhancemonoaminergic transmission by blocking neurotransmitterreuptake, the fast neurotransmitter increase in presynapticareas activates autoreceptors such as the 5-HT1A,inhibiting neuronal firing. Thus, the therapeutic effect isonly obtained after some weeks of sustained treatment.As the classic monoaminergic hypothesis fails to explain

all the features of the pathology, other hypotheses havebeen proposed involving other neurotransmitter receptorsas the cannabinoid receptors.In this sense, the major non-psychoactive constituent

of marihuana, (−)cannabidiol (CBD), has been recentlyproposed as a potential therapy for mood disorders [1].The acute administration of this drug has been provento have an anxiolytic effect. However, the consequencesof chronic treatment, particularly in an animal model ofdepression, remain unknown.In this study we have tested the effect of chronic

treatment with CBD in the bilateral olfactory bulbectomy(OBX) mice model of comorbid depression and anxiety,both in behavioural paradigms and in the functionality ofCB1 and 5-HT1A receptors in different brain areas.

Materials and Methods: Mice were bulbectomisedor sham operated and after a recovery period of onemonth, they received CBD or vehicle resulting into fourexperimental groups. CBD 50mg/kg/day was administeredintraperitoneally for 3 days followed by CBD 10mg/kg/dayfor 11 days. Aversive open field and sucrose intake testswere conducted for behavioural characterisation at the endof the treatment. The animals were sacrificed and the brainswere removed, frozen and cut into 14mm slices. G-proteincoupling to CB1 and 5-HT1A receptors was analysed by[35S]GTPgS binding in brain areas relevant for depressivepathology, using WIN55,212−2 and 8-OH-DPAT agonists,respectively.

Results: CBD treatment prompted a decrease inperipheral hyperactivity of OBX mice in the open field,but no modification was reported in the total locomotoractivity of sham counterparts. As for the central activity,no significant changes were noticed, either in OBX orin sham animals. In the sucrose intake test, OBX miceexhibited a decrease in sucrose consumption, resemblingan anhedonic state that was reversed by two weeksCBD treatment. Regarding receptor functionality, 8-OH-DPAT-stimulated [35S]GTPgS autoradiography revealedthat 5-HT1A functionality was decreased in OBX limbicareas, such as amygdala, hippocampus and entorhinalcortex, and it was reverted by chronic administrationof CBD. On the other hand, WIN55,212−2-stimulated[35S]GTPgS autoradiography showed an increased CB1functionality in hypothalamus of the OBX group comparedto their sham counterparts, and CBD restored the normalactivity.

Conclusions: Taken together, these results indicate thatCBD improves the depression-like features of OBX mice,acting like classic antidepressant drugs. The modulationof 5-HT1A and CB1 receptors functionality in limbic areascould underlie these effects.

Reference(s)

[1] El-Alfy AT, Ivey K, Robinson K, Ahmed S,Radwan M, Slade D, Khan I, ElSohly M,Ross S, 2010. Antidepressant-like effect of delta9-tetrahydrocannabinol and other cannabinoids isolatedfrom Cannabis sativa L. Pharmacol Biochem Behav.95(4):434−42.

P.1.014 Intracerebroventricular administration ofinterleukin-1b elevates brain kynurenicacid and disrupts prepulse inhibition inC57BL/6 mice

M. Larsson1 °, L. Schwieler1, G. Engberg1, S.B. Powell2,S. Erhardt1. 1Karolinska Institute, Physiology andPharmacology, Stockholm, Sweden; 2University ofCalifornia, Department of Psychiatry, San Diego, USA

Purpose: Patients with schizophrenia and bipolar disorderdisplay elevated central levels of the pro-inflammatorycytokine interleukin-1b (IL-1b) [1] and of the endogenousNMDA receptor antagonist kynurenic acid (KYNA) [2].Both KYNA and IL-1b are predominantly elevatedin those bipolar patients that have had a psychoticepisode. Moreover, in similarity with the psychotomimeticcompounds ketamine and PCP, pharmacologically elevatedlevels of KYNA in rodents have been shown to disruptprepulse inhibition (PPI) [3]. The aim of the present studywas to investigate if IL-1b influences the synthesis of brainKYNA in mice and if administration of IL-1b affects PPI.

Methods: Adult C57BL/6 mice were injected intra-cerebroventricularly (i.c.v.) with 0.5, 1, 5, or 10 ng of IL-1bin PBS under isoflurane anaesthesia. Control animalsrecieved vehicle only. Six hours post-injection the animalswere swiftly sedated with isoflurane and then euthanisedby cervical dislocation and their brains were harvested.KYNA was then quantified by means of reversed phasehigh performance liquid chromatography. Another cohortof mice received i.c.v. injections of 0.5, 5, or 50 ng IL-1band were tested for PPI deficits at several time pointspost-injection.

Results: Administration of 0.5 ng IL-1b, but not 1, 5, or10 ng, significantly elevated brain KYNA levels comparedto vehicle (9.49±1.88 nM vs. 3.40±0.51 nM, one-wayANOVA with Bonferroni’s post hoc test, F(4,30) = 4.737,


p< 0.05). I.c.v. administration of IL-1b significantlydisrupted PPI at the lowest dose only (0.5 ng).

Conclusion: Present results support the hypothesis thatIL-1b and KYNA are important players in the patho-physiology of psychotic diseases, such as schizophreniaand bipolar disorder. Notably, only administration of thelowest dose IL-1b had a PPI disruptive effect, indicatingthat this effect may be mediated by the increased brainKYNA concentrations observed at this dose. Presentdata are also in line with recent in-vitro data from ourlaboratory showing that IL-1b, by induction of tryptophan2,3-dioxygenase, increase KYNA production in humancortical astrocytes.

Reference(s)

[1] Soderlund, J., Schroder, J., Nordin, C., Samuels-son, M., Walther-Jallow, L., Karlsson, H., Erhardt, S.,Engberg, G., 2009. Activation of brain interleukin-1beta in schizophrenia. Mol Psychiatry 14, 1069−71.

[2] Erhardt, S., Blennow, K., Nordin, C., Skogh, E.,Lindstrom, LH., Engberg, G., 2001. Kynurenic acidlevels are elevated in the cerebrospinal fluid of patientswith schizophrenia. Neurosci. Lett. 313, 96−98.

[3] Erhardt, S., Schwieler, L., Emanuelsson, C., Geyer,MA., 2004. Endogenous kynurenic acid disruptsprepulse inhibition. Biological Psychiatry. 56, 255–260.

P.1.015 Time-dependent adaptations at thelevel of dopamine D2 receptor instress-resilient rats

D. Zurawek1 °, A. Faron-Gorecka1, M. Kusmider1,M. Kolasa1, P. Gruca2, M. Papp2, M. Dziedzicka-Wasylewska1. 1Polish Academy of Sciences, Departmentof Biochemical Pharmacology, Krakow, Poland; 2PolishAcademy of Sciences, Department of BehaviouralPharmacology, Krakow, Poland

Recent research has focussed on the mechanisms thatunderlie the susceptibility to stress. Long-lasting mildstress, in contrast to single short-term traumatic event,may be a more important factor leading to depression.Few studies have examined the molecular differencescharacteristic for stress-resilient individuals [1] therefore,factors that contribute to stress-resilience may be worth ofinvestigation. Stress exerts strong influence on mesolimbicdopamine system and elicits specific responses dependingon duration and type of aversive stimuli [2]. Therefore,the main purpose of this study was to examine stress-induced changes in mesoaccumbal dopamine D2 mRNAexpression and D2 receptor protein density in two groups

of animals with different behavioural responses to CMSparadigm.We used Chronic Mild Stress [3], a well-established

behavioural animal model of depression, in which malerats were subjected to unpredictable, variable stressconditions for 2 and 5 consecutive weeks. After 2 weeks ofthe CMS we identified three different groups of animals:1. control, 2. stress-reactive which significantly decreaseddrinking of a palatable sucrose solution (F2,29 = 14.74,p< 0.005 vs. control) and 3. stress-resilient animalswith unaltered drinking profile (F2,29 = 14.74, p> 0.05 vs.control). Similar groups were identified after 5 weeksof the CMS: 1. control, 2. stress-reactive (F2,29 = 10.79,p< 0.05 vs. control) and 3. stress-resilient (F2,29 = 10.79,p> 0.05 vs. control). [3H]Domperidone was used asa ligand to label dopamine D2 receptors in nucleusaccumbens (Nacc) shell and core, and ventral tegmentalarea (VTA). In situ hybridisation method was used toexamine dopamine D2 receptor mRNA changes in alltested brain regions. Data were analysed using appropriateANOVA analysis of variances followed by Bonferroni’spost hoc test.

Table 1. The effect of 2 and 5 weeks of CMS on specific [3H]domperidone binding in various regions of the ratbrain

2 weeks CMS 5 weeks CMS

Control Stress-reactive Stress-resilient Control Stress-reactive Stress-resilient

Nacc shell 45.69±3.937 34.14±4.236 19.95±2.829*** 36.44±4.690 19.88±2.827** 23.45±2.802*Nacc core 46.70±4.649 32.49±3.979 21.44±3.097*** 45.59±7.104 23.42±3.506* 32.08±4.228Medial VTA 46.45±4.732 43.55±7.477 30.72±4.740 34.99±4.522 17.49±1.653* 35.02±5.901Lateral VTA 44.18±4.474 40.69±6.720 20.59±3.019** 24.35±2.361 13.10±1.822* 28.11±5.271Data represent the mean±SEM; n = 10 (fmol/mg tissue).*p< 0.05, **p< 0.01, ***p< 0.001 vs. control.

CMS affected the mesolimbic dopamine circuit instress-resilient animals after 2 weeks, and in stress-reactivegroup of rats after 5 weeks, as observed by a decrease in thelevel of dopamine D2 receptor protein without alterationsin D2 mRNA expression. The increase in D2 mRNAexpression returned the dopamine D2 receptor densityto control levels in stress-resilient rats after 5 weeks ofCMS, while that effect was not observed in stress-reactiveanimals.

Conclusions: Despite earlier blunting, stress-resilientanimals, but not stress-reactive animals, effectivelyadapted to the extended stress and coped with it byincreasing dopamine D2 receptor mRNA expression andprotein density in mesoaccumbens system after 5 weeks ofstress. Stress-reactive rats did not show such adaptations orit may be delayed, leading to anhedonia, a core symptomof depression.

Reference(s)

[1] Blugeot A, Rivat C, Bouvier E, Molet J, Mouchard A,Zeau B, Bernard C, Benoliel JJ, Becker C (2011)Vulnerability to depression: from brain neuroplas-


ticity to identification of biomarkers. J Neurosci31(36):12889−99.

[2] Cabib S, Giardino L, Calza L, Zanni M, Mele A,Puglisi-Allegra S (1998) Stress promotes majorchanges in dopamine receptor densities within themesoaccumbens and nigrostriatal systems. Neuro-science 84(1):193–200.

[3] Willner P, Towell A, Sampson D, Sophokleous S,Muscat R (1987) Reduction of sucrose preference bychronic unpredictable mild stress, and its restorationby a tricyclic antidepressant. Psychopharmacology(Berl) 93(3):358−64.

P.1.016 Modulation of dopamine D2L and D3

receptor signalling and cell surface-cytosol trafficking by dysbindin inCHO cells

N. Schmieg1 °, C. Rocchi2, R. Maggio2, M.J. Millan1,C. Mannoury la Cour1. 1Institut de Recherche Servier,Unite Recherche et Decouverte Neuroscience, Croissy-sur-Seine, France; 2University of L’Aquila, Biotechnologicaland Applied Clinical Sciences Department, L’Aquila,Italy

Purpose of the studies: Schizophrenia is a severeneuropsychiatric disorder with many risk factors, bothenvironmental and genetic. As regards genetic factors, dys-tobrevin binding protein 1 (dysbindin) is a susceptibilitygene and its levels are reduced in schizophrenia [1]. Fur-ther, genetically reducing dysbindin expression enhancedcell surface expression of D2 receptors in frontal cortex [2].In view of these observations suggesting altered control ofdopaminergic transmission in schizophrenia by dysbindin,we investigated the influence of dysbindin overexpressionon signalling and cell surface localisation of human (h)dopamine D2L versus D3 receptors stably expressed inCHO cells.

Methods and Results: CHO cells stably expressing D2Lor D3 receptors (Bmax (in fmol/mg protein) 8283±438 and9766±347, respectively) were transiently transfected withdysbindin (3mg) with no significant effect on Bmax values.Employing the non-membrane penetrant radioligand, [3H]-methoxysulpride, it was shown that in dysbindin-treatedcompared to control cells (100%) there was a reductionin the density of hD2L and hD3 receptors to 61±5 and59±4%, respectively, without any change in radioligandaffinity (KD): 0.22±0.05 and 0.23±0.05, for hD2L andhD3 receptors, respectively. In line with these data, usingAlphascreen Surefire™ assays, dysbindin overexpressionwas found to markedly decrease the inhibitory effect ofDA stimulation on forskolin-activated adenylyl cyclase

activity. As compared to control values (defined as 100%),values in the presence of dysbindin were 62±7% and41±5% for hD2L and hD3 receptors, respectively −as compared to 98±6% for D1 receptors, which werenot affected in a control study. As shown by WesternImmunoblotting, in the presence of dysbindin, and ascompared to control (100%) values, the maximal effectof DA on Akt (Ser473 and Thr308), GSK-3b (Ser9) andERK1/2 phosphorylation at hD2L receptors was 42±3,58±5 and 48±3% respectively and, at hD3 receptors, itwas 41±4, 53±3 and 22±4%, respectively. Despite thesealterations in maximal effect there was, by contrast, littleor no change in the potency of DA for all measures ofhD2L and hD3 receptor activation. Finally, employing theinhibitor of clathrin/caveolar-mediated receptor internal-isation, methyl-beta-cyclodextrin (MbCD), we acquiredevidence that this mechanism may be involved in theinfluence of dysbindin upon signalling and membrane-cytosol shuttling at both hD2L and hD3 receptors.

Conclusions: These data show that overexpression ofdysbindin markedly reduces the efficacy (but not potency)of DA-mediated signalling at hD2L and hD3 receptorsas concerns both inhibition of adenylyl cyclase andrecruitment of Akt/GSK-3b and ERK1/2 pathways. Theseobservations were parallelled by decreases in cell surfaceexpression of hD2L and hD3 receptors and may, at leastpartially, reflect a decrease of receptor density involvingclathrin/caveolar-mediated internalisation. These datasupport studies of dysbindin silencing in suggestingthat abnormalities in dysbindin control of D3 and D2Lreceptor signalling and localisation may be related to thepathogenesis and symptoms of psychosis.

Reference(s)[1] Ghiani C.A. and Dell’Angelica E.C., 2011. Dysbindin-

containing complexes and their proposed functions inthe brain: From zero to (too) many in a decade. ASNneuro 3. pii: e00058.

[2] Ji, Y., Yang, F., Papaleo, F., Wang, H.X., Gao, W.J.,Weinberger, D.R., Lu, B., 2009. Role of dysbindinin dopamine receptor trafficking and cortical GABAfunction. Proc Natl Acad. Sci. 106, 19593–19598.

P.1.017 Histaminergic regulation of presumedserotonin neurons in the dorsal raphenucleus

K. Panetta1 °, D. Belelli1, J. Lambert1. 1University ofDundee, Division of Neuroscience, Dundee Scotland,United Kingdom

Introduction: Sleep is essential for our cognitive andphysical wellbeing, yet 25% of the UK population


suffer from sleep disorders which can have majorconsequences on their daily lives. Neural control of thesleep-wake cycle results from the complex interaction ofneurotransmitters systems, which arise from anatomicallyand chemically-defined brain structures. Serotonergic andhistaminergic neurons of the dorsal raphe nucleus (DRN)and tuberomammillary nucleus (TMN) respectively areproposed to be wake-promoting neurons, although howthey act in concert to influence behavioural state isnot well understood [1]. H1 receptor antagonists andcertain selective serotonin (5-HT) receptor antagonistshave been shown promote sleep in humans [2,3]highlighting the possible importance of these compoundsas sleep therapeutics. The purpose of this study was toutilise electrophysiological techniques to better understandhow 5-HT neurons within the DRN are regulatedby histamine.

Methods and Results: Firstly extracellular single-unitrecordings were performed from putative 5-HT neuronswithin mouse brain slices containing the DRN. Thebath application of 10mM histamine caused an increasein the firing frequency of all cells tested. Secondlywhole-cell voltage-clamp recordings (VH = −60mV) wereperformed from putative 5-HT neurons within mousebrain slices containing the DRN. Under these recordingconditions, the bath application of 10mM or local pressuredelivery of 300mM, (5−10ms, 10psi) histamine causedan inward current associated with increased membranenoise, indicative of the opening of associated ion channels.To identify the receptors underlying the histamine-evoked current, the effects of selective ligands of thehistamine receptor subtypes (H1, H2 and H3 receptors(Rs) known to be present in the DRN were investigated.The bath application of the H1R antagonist, histabudifen(20−60mM) completely abolished the histamine-inducedinward current. In contrast, both the H2R antagonist,ranitidine (10mM), and the H3R antagonist thioperamide(1mM) had no effect on the histamine-induced current.Interestingly, the bath application of either of two struc-turally distinct H1 inverse agonists, mepyramine (300nM)and dimethendene (100nM) also inhibited the histamine-induced inward current, but additionally caused an outwardshift in the holding current thus, suggesting the presence ofan endogenous tonic histaminergic conductance. However,the competitive antagonist histabudifen while blocking thehistamine-evoked response did not produce an outwardcurrent, thus suggesting H1Rs to be constitutively (i.e.spontaneously) active rather than being gated by ambientlevels of histamine.

Conclusions: Collectively, these results demonstratethat histamine is able to modulate presumed 5-HT neuronsand moreover that a histamine tone is present in the DRN.Importantly, the findings reported here provide the first

example of a spontaneous H1 receptor activity in the DRN.Future studies will assess the physiological relevance ofthis constitutive activity in vivo to explore whether H1inverse agonists may be used as therapeutic agents to treatsleep disorders.

Reference(s)

[1] Saper, C.B., Scammell, T.E., Lu, J., (2005). Hypothala-mic regulation of sleep and circadian rhythms. Nature437, 1257–1263.

[2] Landolt, H.P., Meier, V., Burgess, H.J., et al.(1999). Serotonin-2 receptors and human sleep: effectof a selective antagonist on EEG power spectra.Neuropsychopharmacology 21, 455–466.

[3] Morairty, S.R., Hedley, L., Flores, J., Martin, R.,Kilduff, T.S., (2008). Selective 5HT2A and 5HT6receptor antagonists promote sleep in rats. Sleep 31,34−44.

P.1.018 Dissecting the neural mechanismsunderlying impaired threat detection inthe Ahi1 knockout mouse

A. Lotan1 °, T. Lifschytz1, A. Slonimsky1, S. Abedat2,Y. Fellig3, H. Cohen4, O. Lory5, G. Goelman5,B. Lerer1. 1Biological Psychiatry Laboratory, Hadassah-Hebrew University Medical Center, Jerusalem, Israel;2Cardiovascular Research Center, Hadassah-HebrewUniversity Medical Center, Jerusalem, Israel; 3Deptof Pathology, Hadassah-Hebrew University MedicalCenter, Jerusalem, Israel; 4Anxiety and Stress ResearchUnit, Faculty of Health Sciences Ben-Gurion Universityof the Negev, Beer-Sheva, Israel; 5MRI Laboratory,Hadassah-Hebrew University Medical Center, Jerusalem,Israel

Purpose: The Abelson helper integration site (AHI1) geneplays a pivotal role in brain development. Studies by ourgroup, replicated by several others, have demonstratedassociation of single nucleotide polymorphisms in AHI1with susceptibility to schizophrenia and autism [1,2].To elucidate the mechanism whereby alteration inAHI1 expression may be implicated in the pathogenesisof endophenotypes related to severe neuropsychiatricdisorders, we studied Ahi1 heterozygous knockout mice(Ahi1+/−). We employed behavioural paradigms thatmodel different aspects of schizophrenia, pharmacologicalchallenges, histological studies and resting state functionalmagnetic resonance imaging (rsfMRI).

Methods: We evaluated the behavioural effects ofreduced Ahi1 expression by comparing performance ofAhi1+/− and control wildtype mice (Ahi1+/+) on tests


that model positive (prepulse inhibition, MK-801 inducedhyperlocomotion), negative (sociability) and cognitive(Morris water maze) aspects of schizophrenia, as wellas tests measuring anxiety such as elevated plus maze(EPM), light dark box (LDB) and open field (OF) tests.Serum cortisol and core body temperature were measuredfollowing behavioural and pharmacological experiments.We sought structural abnormalities in the brains of Ahi1+/−

mice by light microscopy and functional disconnectivityby rsfMRI.

Results:Western blots showed lower Ahi1 protein levelsin Ahi1+/− mice, although the difference was significantonly in newborns. We did not detect abnormalities onbehavioural measures reflecting positive, negative andcognitive domains of schizophrenia. On the other, weobserved consistently lower levels of anxiety in Ahi1+/−

mice. Compared to Ahi1+/+, Ahi1+/− mice spent moretime in the OF arena centre (p< 0.01), the EPM openarms (p< 0.01) and the LDB lit zone (p< 0.001). Serumcortisol levels and core body temperature, two highlyconsistent endophenotypes relevant to the mammalianstress response, were significantly lower in Ahi1+/−

than Ahi1+/+ mice after exposure to environmentalstress in the EPM (p = 0.02 and p = 0.03, respectively).However, following a direct anxiogenic challenge withcaffeine 80mg/kg i.p., cortisol levels were similaracross genotypes. Detailed examination of brain structureusing H&E staining revealed normal morphology andcytoarchitecture in Ahi1+/− mice. Surface areas of boththe cerebellum, which is grossly distorted in Ahi1-null mice, and the amygdala, which is implicated inthreat detection, were not reduced. However, rsfMRIdata indicated that a dense network of symmetricalcorticolimbic connections and highly significant functionalconnections between the cerebellum and amygdala, bothevident in Ahi1+/+ mice, were markedly diminished inAhi1+/− mice (p< 0.01).

Conclusions: Our results indicate that decreasedAhi1 expression is associated with a defect in threatdetection and/or in generation of an adequate stressresponse. Although we did not observe structuralbrain abnormalities in Ahi1+/− mice, rsfMRI suggestedcorticolimbic disconnectivity. The present findings implythat neurodevelopmental abnormalities associated withreduced expression of Ahi1 during the fetal and neonatalperiod may have significant long-term behavioural andfunctional consequences in the adult. Such a mechanismis consistent with currently postulated neurodevelopmentaltheories of schizophrenia and autism. The present dataprovide a translational evidence for the role of AHI1, agene replicably associated with severe neuropsychiatricdisorders, in the pathogenesis of endophenotypes relatedto threat detection and anxiety.

Reference(s)

[1] Lerer, B., Segman, R.H., Hamdan, A., Kanyas, K.O.,Kohn, Y., Korner, M., Lanktree, M., Kaadan, M.,Turetsky, N., Yakir, A., Kerem, B., Macciardi, F.,2003. Genome scan of Arab Israeli families mapsa schizophrenia susceptibility gene to chromosome6q23 and supports a locus at chromosome 10q24.Mol Psychiatry 8, 488−98.

[2] Amann-Zalcenstein, D., Avidan, N., Kanyas, K.,Ebstein, R.P., Kohn, Y., Hamdan, A., Ben-Asher, E.,Karni, O., Mujaheed, M., Segman, R.H., Maier, W.,Macciardi, F., Beckmann, J.S., Lancet, D., Lerer, B.,2006. AHI1, a pivotal neurodevelopmental gene,and C6orf217 are associated with susceptibility toschizophrenia. Eur J Hum Genet 14, 1111−9.

P.1.019 Role of the 5-HT2A receptor in themechanism of action of antidepressantdrugs: a translational human–mousestudy

G. Quesseveur1 °, A.C. Petit2, F. Gressier2, R. Colle2,D.J. David1, A.M. Gardier1, C. Verstuyft3, E. Corruble2,B.P. Guiard1. 1University of Paris Sud 11, EA 3544Laboratoire de Neuropharmacologie, Chatenay MalabryCedex, France; 2University of Paris Sud 11, INSERMU 669 Department of Psychiatry Bicetre UniversityHospital, Le Kremlin Bicetre, France; 3Hopital Bicetre,Service de Genetique Moleculaire Pharmacogenetique etHormonologie, Le Kremlin Bicetre, France

Selective serotonin reuptake inhibitors (SSRIs) arecommonly prescribed for the treatment of majordepression. However, 50% of depressive patients donot respond adequately to these medications. Althoughevidence incriminates the overactivation of the 5-HT1Aautoreceptor in this poor response [1], other serotonergicreceptors could be recruited to modulate the therapeuticactivity of SSRIs [2]. In agreement with this hypothesis,growing arguments suggest that variants at gene encodingthe 5-HT2A receptor are associated with antidepressantresponses [3] but the results of pharmacogenetic studiesin human are still matter of debate.The purpose of this translational study was to

determine the effects of 5-HT2A receptor inactivation onthe electrophysiological, neurochemical and behaviouralactivity of SSRIs in mice. On the other hand, thiswork evaluated the impact of two putatively functionalsingle nucleotide polymorphisms of the 5-HT2A receptorgene (rs6313 and rs6314) [3] on SSRIs responses indepressed patients from the French cohort METADAP.In particular, we studied the percentage of responders and


the improvement of their depression scores from variousrelevant scales.In wild-type 5-HT+/+1A mice, the acute administration

of the SSRIs escitalopram or fluoxetine decreased thefiring rate of dorsal raphe (DR) 5-HT neurons, whilethe administration of the selective 5-HT1A receptorantagonist WAY100635 reversed this effect. Remarkably,the electrophysiological response induced by both SSRIspersisted in 5-HT+/+1A mice pretreated with WAY100635 orin 5-HT−/−1A mice thereby demonstrating the involvementof another serotonergic receptor type in the inhibitoryactivity. The observation that the 5-HT2A receptor an-tagonist MDL100907 also reversed escitalopram-induceddecrease in DR 5-HT neuronal activity indicates that thesimultaneous blockade of 5-HT1A and 5-HT2A receptors isrequired to prevent the acute inhibitory effects of SSRIsupon the serotonergic system. It also suggests that theactivity of SSRIs might be enhanced in 5-HT−/−2A miceafter chronic treatment. However, the genetic inactivationof the 5-HT2A receptor significantly attenuated the abilityof repeated administration of escitalopram or fluoxetine toincrease the firing rate of DR 5-HT neurons and reducedtheir antidepressant-like effects in the tail suspension testor the novelty suppressed feeding paradigm. Finally, theenhancement of adult hippocampal neurogenesis inducedby prolonged administration of SSRIs was blunted in5-HT−/−2A mice.In depressed patients, rs6313 and rs6314 genetic

variants were not associated with SSRIs response. Weextended this observation to the fact that separatelyanalysed neither escitalopram nor fluoxetine responseswere altered for the rs6313. In marked contrast, atrend toward a lower percentage of improvement ofdepression score was detected after escitalopram treatmentin homozygous individuals for the C allele of the rs6314(p = 0.06).Altogether, these preclinical and clinical data indicate

that a functional variant of the 5-HT2A receptor genemay be associated with a poor SSRIs response resulting,at least in part, from an impairment of serotonergicneurotransmission. Although this study has to becompleted by determining the consequence of the Callele on the function/expression of the 5-HT2A receptorin the human brain, these results could be of particularimportance to select appropriate antidepressant treatmentaccording to the patient’s genotype.

Reference(s)

[1] Gardier AM, Malagie I, Trillat AC, Jacquot C, ArtigasF. Role of 5-HT1A autoreceptors in the mechanism ofaction of serotoninergic antidepressant drugs: recentfindings from in vivo microdialysis studies. FundamClin Pharmacol. 1996;10(1):16−27.

[2] Quesseveur G, Nguyen HT, Gardier AM, Guiard BP.5-HT2 ligands in the treatment of anxiety anddepression. Expert Opin Investig Drugs. 2012Nov;21(11):1701−25.

[3] Serretti A, Drago A, De Ronchi D. HTR2A genevariants and psychiatric disorders: a review of currentliterature and selection of SNPs for future studies.Curr Med Chem. 2007;14(19):2053−69. Review.

P.1.020 The role of CREB/BDNF/TrkB signallingin the zinc deficiency model ofdepression

U. Doboszewska1 °, B. Szewczyk1, M. Sowa-Kucma1,K. Mlyniec2, G. Nowak1. 1Polish Academy of SciencesInstitute of Pharmacology, Department of Neurobiology,Krakow, Poland; 2Jagiellonian University MedicalCollege, Department of Biochemical Toxicology, Krakow,Poland

Background: Induced zinc deficiency in adult ratssignificantly decreases the number of progenitor cellsand immature neurons in the dentate gyrus of the rathippocampus. Zinc deficiency, which occurs during bothlactation and adulthood, increases hippocampal apoptosis.This suggests that the trace element zinc may be involvedin the regulation of hippocampal neurogenesis during theearly stages of life through to adulthood [1]. Neurotrophinsare an important class of signalling molecules essential forthe development of the central nervous system (CNS).Brain-derived neurotrophic factor (BDNF) is the bestcharacterised of these neurotrophins in terms of its rolein synaptic plasticity as well as its potential role inthe pathology and treatment of a variety of psychiatricdisorders. BDNF gene expression is increased by thetranscription factor cyclic AMP response-element bindingprotein (CREB). Mature BDNF signals via the high-affinity tropomyosine-related kinase B receptor (TrkB).BDNF signalling via TrkB receptors divides into threepathways, all of which converge on the transcription factorCREB, which in turn up-regulates gene expression [2].

Aim: To examine whether zinc deprivation induceschanges in the protein levels of the phosphorylated formsof CREB (pCREB), BDNF and TrkB in the prefrontalcortex and hippocampus of rats.

Methods:Male Sprague-Dawley rats (5-week-old) werefed a 50mgZn/kg (control group) or a 3mgZn/kg (zinc-deficient group) diet for four weeks. The protein levels ofpCREB, BDNF and TrkB were measured using westernblotting.

Results: Four weeks of dietary zinc deprivation resultedin a significant decrease in the protein levels of pCREB,


BDNF and TrkB in the hippocampus but not in theprefrontal cortex. The level of pCREB protein wasdecreased by 91% whereas the level of BDNF protein wasdecreased by 73% in the hippocampus of the zinc-deficientrats when compared with that of the control animals. Atthe same time point a decrease (by 40%) in TrkB proteinwas observed in the hippocampus of the zinc-deficientgroup compared to the control group.

Conclusions: The previous studies indicated thatdietary-induced zinc deficiency leads to the developmentof depressive like behaviour (e.g. increase in theimmobility time in the forced swim test) and thatexperimentally induced zinc deficiency might be a modelof depression. The present data suggests that pCREB,BDNF and TrkB expression is modulated by zincdeficiency and that the decreased expression of theseproteins after dietary zinc deprivation is linked to thedepressive like behaviour in the zinc deficiency paradigm.It seems that the hippocampus, the region of the brain thatplays a critical role in neurogenesis, is more susceptibleto the biochemical changes caused by zinc deficiencythan the prefrontal cortex. This is consistent with dataobtained during experiments where acute and chronicstress paradigms were found to decrease the expressionof BDNF in the rodent hippocampus [1].

Reference(s)

[1] Levenson, C.W., Morris, D., 2011. Zinc and neuro-genesis: making new neurons from development toadulthood. Adc Nutr 2, 96–100.

[2] Autry, A.E., Monteggia, L.M., 2012. Brain-derivedneurotrophic factor and neuropsychiatric disorders.Pharmacol Rev 64, 238–258.

P.1.021 Chronic administration of haloperidolin rats and its effect on microglial celldensity and whole brain weight andvolume

P.S. Bloomfield1 °, O.D. Howes1, V. de Paola2. 1MRCClinical Sciences Centre, Psychiatric Imaging, London,United Kingdom; 2MRC Clinical Sciences Centre,Neuroplasticity and Disease, London, United Kingdom

Introduction: Patients with schizophrenia are routinelyprescribed antipsychotic medication. Magnetic resonanceimaging (MRI) in newly medicated patients has demon-strated how, over a 5 year period, there is an averagewhole brain structural deficit of 8%, with regional lossesof up to 20%, when compared to baseline [1]. Anotherstudy investigating schizophrenic patients demonstratean increase in binding with the [C11]PK11195 positron

emission tomography (PET) radiotracer, which acts asa marker of neuroinflammation [2]. While these arechanges since the start of medication, it is not possibleto differentiate between changes arising from medicationand those occurring with the progression of the disease.Here we dose naıve rats with haloperidol (Hal) to test thehypotheses that anitpsychotic drugs induce brain volumeloss and microglial activation.

Methods: We used subcutaneous drug pellets to slowlyrelease Hal over a two-week period in randomised mixedcages of control (n = 9) and medicated (n = 9) male SpragueDawley rats. The drug pellets released a 0.05mg/kg/daydose, control animals were implanted with a placebo pellet.The dose used is a comparatively low dose in terms ofpreclinical literature, with a similarly low D2 receptoroccupancy when compared with clinical data (calculatedfrom Samha et al., 2008 [3]).Following perfusion, brains were dissected for post

mortem analysis. The weights of brains were measuredand whole brain volumes were calculated using waterdisplacement. The brain tissue was then processed forimmunohistochemistry, where changes in microglial cellnumber were determined to assess neuroinflammatoryassociated changes.

Results: Here we show that there is a significantreduction in both brain weight and volume in Hal treatedanimals (2.57 g (±0.088 SEM), 1583.3mm3 (±105.4SEM)) when compared with placebo controls (2.78 g(±0.111 SEM), 2166.7mm3 (±153.7 SEM)) (p< 0.05,independent samples t-test). Immunohistochemical quan-tification of colocalised Iba-1 and DAPI stains was used todetermine the number of microglial cells in the following800mm2 regions of interest (ROIs); the prefrontal cortex,hippocampus and ventral striatum. There was a consistentincrease in microglial number across the 3 regions indrug treated animals (see table), which trended towardsignificance (p = 0.07).

Mean Iba-1 positive cells counts in cortical ROIs

PFC(±SEM)

Hippocampus(±SEM)

Striatum(±SEM)

Placebo 59.0 (±11.1) 25.3 (±5.1) 69.2 (±16.1)Haloperidol 62.0 (±12.4) 51.3 (±13.2) 85.3 (±19.4)

Conclusions: The results we have seen here showhow a low dose of a typical antipsychotic medicationis able to produce significant changes in the brains ofnaıve rats. Further investigation is required to determinethe full effects of these changes and the mechanisminvolved. An experiment investigating the effects of ahigher dose regime is in progress and will be usedto investigate inflammatory changes further alongside


cortical morphology. While evaluating the results of thispreliminary investigation, the impact of function requiresconsideration, as the changes we see here may not bedetrimental to cognition. We propose further investigationinto the impact of such cortical changes on function.

Reference(s)

[1] Thompson, P.M., et al., Mapping adolescent brainchange reveals dynamic wave of accelerated gray mat-ter loss in very early-onset schizophrenia. Proceedingsof the National Academy of Sciences, 2001. 98(20):p. 11650–11655.

[2] Doorduin, J., et al., Neuroinflammation in schizo-phrenia-related psychosis: A PET Study. Journal ofNuclear Medicine, 2009. 50(11): p. 1801–1807.

[3] Samaha, A.-N.L., et al., Less Is More: Antipsychoticdrug effects are greater with transient rather thancontinuous delivery. Biological Psychiatry, 2008.64(2): p. 145–152.

P.1.022 Role of trace amine-associatedreceptor 1 (TAAR1) in the modulationof the dopaminergic system andcortico-striatal signalling

S. Espinoza1 °, I. Sukhanov1, G. Lignani1, L. Medrihan1,S. Maggi1, G. Giannotti2, F. Fumagalli2, F. Benfenati1,V. Tucci1, R. Gainetdinov1. 1Italian Institute ofTechnology, NBT, Genova, Italy; 2University of Milan,Pharmacology, Milan, Italy

Purpose of the study: Mammalian Trace AmineAssociated Receptor 1 (TAAR1) is a G protein-coupledreceptor (GPCR) that is mainly expressed in limbic regionsand monoaminergic nuclei, such as ventral tegmentalarea, dorsal raphe and nucleus coeruleus [1]. Thereis evidence indicating that TAAR1 is involved in themodulation of dopaminergic system [2]. In mice lackingTAAR1 (TAAR1-KO mice), amphetamine induces morepronounced locomotor stimulation and dopamine release.Moreover, it has been reported that D2 receptor function isaltered in TAAR1-KO mice. Dopamine system is involvedin many physiological functions and has been implicatedin various pathological states such as schizophrenia andParkinson’s disease. Previously, we showed that TAAR1and D2 could heterodimerise and this interaction couldinfluence TAAR1 signalling in vitro and D2-inducedbehaviour in vivo [3]. In this study, our purpose was todescribe how TAAR1 could influence dopamine system invivo, in particular we evaluated TAAR1 role in D2-relatedsignalling in the striatum.

Methods used: Binding: the total number of D1 and D2receptors was evaluated by binding experiments. Westernblot: total form and phosphorylated form of variousproteins was investigated with the analysis of differentbrain regions extracts. BRET: b-arrestin2 recruitment toD2L receptor was monitored by using a BRET approach.Electrophysiology: NMDA and AMPA functionality wasmeasured by patch clamp recordings in slices. Behaviour:deficits of cortico-striatal circuitry were assessed using atiming task, namely, fix interval and peak interval.

Results: With binding experiments, we found that D2but not D1 number was decreased in striatum of TAAR1-KO mice. Moreover, we analysed D2 receptor-relatedsignalling, both G protein-dependent as well as G protein-independent pathway. In striatum of TAAR1-KO mice, asignificant reduction in AKT and GSK-3b phosphorylationwas found as well as a reduction of b-catenin, with nodifference in phosphorylation of DARPP32, ERK andCREB, demonstrating that the AKT/GSK3 pathway wasactivated. In order to understand the mechanism of thisactivation, we also studied b-arrestin2 recruitment to D2Rusing BRET and its role in vivo in striatum of TAAR1-KO mice. While in vitro it seems that TAAR1 does notmodulate b-arrestin2 recruitment to D2R, in vivo we foundan alteration of the complex AKT/PP2A. Since TAAR1modulates dopaminergic neurons located in VTA we madeinvestigations in the prefrontal cortex, particularly NMDAreceptor functionality. By western blot we noted thatNMDA receptor was less phosphorylated at the serine 896suggesting a reduction in NMDA functionality. By usingpatch clamp recordings we found that NMDA receptorlocated in pyramidal neurons in layer V were less active.Finally, we performed a timing task, namely a fix intervaland peak interval. While it seems that the timing it is notaltered in TAAR1-KO mice, it is evident a deficit in thegeneral performance, suggesting a deficit in cognition andan alteration in cortico-striatal circuits.

Conclusions: These data indicates that TAAR1 is ableto modulate dopamine system, in particular D2 receptorfunctions in striatum, and also prefrontal cortex NMDAfunctions, neurons excitability and some behavioursrelated to cortico-striatal pathways.

Reference(s)

[1] Lindemann L and Hoener MC (2005) A renaissance intrace amines inspired by a novel GPCR family. TrendsPharmacol Sci 26(5):274–281.

[2] Sotnikova TD, Caron MG, and Gainetdinov RR, Traceamine-associated receptors as emerging therapeutictargets. Mol Pharmacol 76 (2009) 229−35.

[3] Espinoza S, Salahpour A, Masri B, Sotnikova TD,Messa M, Barak LS, Caron MG, and Gainetdinov RR,


Functional interaction between trace amine-associatedreceptor 1 and dopamine D2 receptor. Mol Pharmacol80 (2011) 416−25.

P.1.023 Survival role of embryonal proteins inAlzheimer’s disease linked dementia viaregulation of oxidative stress and levelof catecholamines

K. Yenkoyan1 °, M. Aghajanov1. 1Yerevan StateMedical University, Department of Biochemistry, Yerevan,Armenia, Republic of

Purpose of the study: Main event in the pathogenesisof such a neurodegenerative disorder as Alzheimer’sdisease (AD) is generation and deposition of amyloidbeta (Ab). Oxidative stress and generation of free radicalspecies have implications in the formation of Ab andits subsequent neurotoxicity. Degeneration of aminergicbrainstem nuclei such as the locus coeruleus which isvery vulnerable to the free radicals and the selectiveexhaustion of catecholamines near the locus coeruleusof brainstem can appear as a potential pathogenesisof AD. That’s why our study was focussed to findsome parallels between oxidative stress and changesin catecholamines concentration experimental model ofAD, and find new biological agents which will beable to regulate them. As a bioregulator we chosethe complex of proteoglycans of embryonal genesis(PEG). PEG contains the pool of proteoglycans ofembryonal genesis which are associated with alpha-fetoprotein, chorionic gonadotrophin, beta1-glycoprotein,carcinoembryonic antigen, and the carbohydrate antigensCa-19−9 and Ca-125.

Methods used: The experimental model of AD wasmade in rats by intracerebroventricular injection ofaggregated Ab (fragment 25−35). The animals weredivided into five groups: the control group consistedof vehicle-treated animals; the 1st experimental group(PEG-control) was subcutaneously injected with PEG(0.5mg/100 g) only; the 2nd experimental group was i.c.v.injected with aggregated Ab; the 3rd experimental groupwas subcutaneously injected with PEG (0.5mg/100 g)7 days before Ab injection (group PEG-1); the 4th

experimental group was subcutaneously administeredPEG (0.5mg/100 g) 7 days before Ab injection andon the 31st day after it (group PEG-2). Oxidativestress and antioxidant capacity were measured by useof chemoluminescence method (spontaneous, induced byUV and Fe2+ ions) in cerebral cortex, hippocampusand brainstem. Adrenergic structures of brainstem werestudied by glyoxylic acid condensation using luminescence

microscope and accompanied by the HPLC study ofnorepinephrine (NE) and metabolites of adrenergics.

Summary of results and Conclusion: After injectionof Ab received data testify the increase in the level ofchemoluminescence (both spontaneous and inducible) inthe cerebral cortex, hippocampus and brainstem. On theother hand the increase in intensity of luminescence ofcatecholamine granules near the locus coeruleus was alsodetected. At the same time HPLC results show highelevation in concentration of NE in the brainstem. InPEG treated animals all types of chemoluminescence inmentioned brain structures decreased and the antioxidantcapacity increased without any significant differencebetween PEG-1 and PEG-2 groups. Almost no changesof adrenergic structures were shown compared with thecontrol rats in all PEG treated groups. At the same timein PEG-1 group concentration of NE was less than incontrol, whereas in PEG-2 group it was about controllevel. It should be certainly pointed that the resultsof biochemical determination of NE and metabolitesof monoamines in brainstem were in unison with theluminescent study. Summarising, it seems that regulationof oxidative stress and disturbance in adrenergic structurescan be of importance for neuronal rescue in Alzheimer’spathology and regulation of these both processes can leadto neuroprotection.

P.1.024 Endocannabinoid-mediated plasticity atinhibitory synapses on dopamine cellsas a marker of vulnerability to addiction

C. Sagheddu1 °, M. Melis1, M. Pistis1. 1University ofCagliari, Biomedical Sciences, Cagliari, Italy

Addiction is a psychiatric disorder, whose aetiologyinvolves interaction of inherited predispositions andenvironmental factors. Both clinical and preclinicalfindings indicate that there are important genetic variationsin vulnerability to drug addiction, and that such differencesmay be mediated by the same biological mechanisms.Addictive drugs share the properties of being self-administered by laboratory animals, and of activatingthe brain reward circuitry, which stems from the ventraltegmental area (VTA) where dopamine (DA) cells arelocated. Endocannabinoids serve as retrograde signallingmolecules at many synapses in the brain, including theVTA, and regulate reward seeking by modulating DAsignalling. Among synaptic inputs that regulate DA neuronimpulse activity, those arising from the newly identifiedrostromedial tegmental nucleus (RMTg) play a majorrole [1,2].


In this study we took advantage of significantdifferences between pairs of lines of rats selectively bredfor their voluntary alcohol preference or aversion, thatis the Sardinian alcohol-preferring (sP) or nonpreferring(sNP) rat line. We investigated their electrophysiologicalproperties and synaptic plasticity both in vivo and in vitro.Extracellular single unit recordings in anaesthetised ratsrevealed a difference in baseline firing activity of DAneurons between sP and sNP rats consistent with ourprevious study [3]. More particularly, sP rats showed anincreased spontaneous neuronal activity. Remarkably, thisincrease was parallelled by a reduced strength of RMTginputs. This enhanced impulse activity of DA cells in sPrats negatively correlated with the duration of inhibitionelicited by electrical stimulation of the RMTg.Since DA neurons can escape afferent inputs by releas-

ing endocannabinoids, we examined one form of short-term synaptic plasticity mediated by endocannabinoids atinhibitory synapses in the brain that is depolarisation-induced suppression of inhibition (DSI). VTA DA neuronsmake no exception to this rule and express this formof plasticity. We have found that DSI is differentlyexpressed by two discrete sets of inhibitory synapsesarising from rostral and caudal afferents onto VTA DAneurons. This phenomenon is selectively mediated by theendocannabinoid 2-arachidonoylglycerol (2-AG), whichactivates presynaptic type 1-cannabinoid (CB1) receptors.However, the two discrete DSI do not seem to dependupon differences in CB1 number and/or function, since thedose-response relationship curves showed no differences.On the other hand, we found that the difference in DSIcan be ascribed to the rate 2-AG is degraded. Thus, 2-AGby differently depressing inhibitory synapses arising fromeither rostral or caudal afferents might indirectly alter DAneuron functional state, and enhance the responsivenessof the reward pathway to phasic DA.Given that sP rats are vulnerable phenotype, and that

they possess this endocannabinoid-mediated form of shortterm plasticity, our results suggest that differences inthe equipment of the endocannabinoid system machinerymight control specific sources of vulnerability.

Reference(s)

[1] Lecca S, Melis M, Luchicchi A, Muntoni AL,Pistis M., 2012. Inhibitory inputs from rostromedialtegmental neurons regulate spontaneous activityof midbrain dopamine cells and their responsesto drugs of abuse. Neuropsychopharmacology.Apr;37(5):1164−76.

[2] Lecca S, Melis M, Luchicchi A, Ennas MG,Castelli MP, Muntoni AL, Pistis M., 2011. Effectsof drugs of abuse on putative rostromedial tegmental

neurons, inhibitory afferents to midbrain dopaminecells. Neuropsychopharmacology 36(3):589–602.

[3] Melis M, Pillolla G, Perra S, Colombo G, Muntoni AL,Pistis M., 2009. Electrophysiological properties ofdopamine neurons in the ventral tegmental area ofSardinian alcohol-preferring rats. Psychopharmacol-ogy 201(4):471−81.

S25

Behavioural pharmacology

Lectures

S.02.01 Genetic models of psychiatric disorders:phenotyping mutant mice

J. Waddington1 °. 1Royal College of Surgeons in Ireland,Molecular and Cellular Therapeutics, Dublin, Ireland

This presentation reviews the application of mutant mousephenotypes to the study of neuropsychiatric disorders ingeneral and psychotic illness in particular, with schizo-phrenia as the primary exemplar. It considers these issuesat the levels of behavioural, psychopharmacological andcellular phenotypes of putative import for understandingdisease processes and drug development. Mutant modelsappear to be heuristic at two main levels; firstly, byindicating the functional roles of neuronal componentsthought to be of relevance to the putative pathobiology ofpsychotic illness, they help resolve overt behavioural andunderlying cellular processes regulated by those neuronalcomponents; secondly, by indicating the functional roles ofgenes associated with risk for psychotic illness, they helpresolve overt behavioural and underlying cellular processesregulated by those risk genes. This presentation focussesinitially on models of dopaminergic and glutamatergicdysfunction. Then, it considers advances in the geneticsof schizophrenia and mutant models relating to replicablerisk genes. There is continuing need to address notonly numerous technical challenges but also to developmore ‘real world’ paradigms that reflect the milieuof gene × environment and gene × gene interactionsthat characterise psychotic illness and its response toantipsychotic drugs. Thus, the presentation extends thisdiscussion by exemplifying two new, variant approachesin mutant mice that may serve as prototypes for advancingunderstanding of disease processes and drug development.An enduring challenge is the translational relationshipbetween mouse phenotype, clinical psychopathology and,to the extent known, disease pathobiology.

S.02.02 Developmental models of schizophreniaand their behavioural characterisation

K.C.F. Fone1 °, D.J.G. Watson1, A. McIntosh1, M.V. King1.1The University of Nottingham, School of BiomedicalSciences, Queen’s Medical Centre, Nottingham, UnitedKingdom

Schizophrenia is a chronic debilitating neuropsychiatricdisorder which affects approximately 1% of the population.Symptoms fall into three categories; positive (such asauditory and visual hallucinations and thought disorder),negative (e.g. emotional blunting, social withdrawal andanhedonia) and cognitive dysfunction (including impairedexecutive function, working memory and attention). Thecause of schizophrenia remains unclear but the risk ofdeveloping this disorder is influenced by both genetic andenvironmental factors. Reliable, predictive animal modelsof complex psychiatric disorders, such as schizophrenia,are therefore essential both to increase our understandingof its neurobiological basis and to develop drugs withimproved efficacy. Animal models offer a more rapidplatform to monitor disease progression than in man,the ability to perform invasive monitoring of anatomicaland molecular changes that underlie the disease, and totest novel medicine impossible in patients. Numerousanimal models of schizophrenia have been producedeither by drug-administration, neuronal lesioning, geneticmanipulation or exposure to early-life adversity [1]. Theseinclude: (i) repeated administration of amphetamine orthe NMDA receptor antagonist, phencyclidine, to producechronic changes in dopaminergic and/or glutamatergicfunction, (ii) gestational exposure to the antimitotic agent,methylazoxymethanol, to alter neurogenesis, (iii) neonatalibotenic acid lesion of the ventral hippocampus and(iv) alteration of the early-life environment, whose long-lasting impact on anatomical and behavioural deficits in theresultant rats will be compared. Most rodent models havebehavioural phenotype changes which resemble ‘positive-like’ symptoms of schizophrenia, probably reflectingaltered mesolimbic dopamine function, but fewer modelsalso show altered social interaction and learning andmemory impairment analogous to the negative andcognitive symptoms, respectively. As the negative andcognitive impairments in schizophrenia are resistant totreatment with current antipsychotics, even after remission

S26 Behavioural pharmacology

of the psychosis, it is clearly vital that animal modelsshould replicate these changes. All useful animal modelsshould have the appropriate triad of face (symptomhomology), construct (replicate the theoretical neurobi-ological rationale and pathology) and predictive (show theexpected pharmacological response to treatment by knownantipsychotics and potential new compounds) validity tothe clinical disorder being modelled. For schizophrenia asuitable constellation of behavioural and neurochemicalabnormalities would include postpubertal onset, lossof hippocampal and cortical connectivity and function,limbic dopamine dysregulation, cortical glutamatergichypofunction, vulnerability to stress, abnormal response toreward, social withdrawal and cognitive impairment. Thesefeatures will be discussed using the neurodevelopmentalrat model of schizophrenia produced by rearing rats insocial isolation from weaning [2]. A perplexing problemis how to evaluate core symptoms of mood disorders likefeelings and thoughts which are uniquely human traits. Afurther problem is that there is no current ‘gold standard’medication available to treat symptoms which can beused as a definitive positive control in preclinical studies.Furthermore, many of the current antipsychotics may havea small therapeutic window of effect before sedation andother non-specific motor suppressant actions confoundinterpretation in tasks designed to assess negative andcognitive function. Within a colony rats display a definedsocial structure and develop a hierarchy which playsa critical impact on their development. Thus socialdeprivation of rat pups from the age of weaning (byplacing them in separate cages from littermates) altersbrain development and causes behavioural deficits atadulthood which are unaltered by social re-integration inlater life. Post-weaning social isolation of rats induceslocomotor hyperactivity, enhanced responses to novelty(neophobia), sensorimotor gating deficits and learning andmemory deficits. Collectively, these behavioural changeshave been termed the “isolation syndrome” and severalof these resemble core symptoms of schizophrenia.The hyperactivity may relate to changes in mesolimbicdopamine activity which reflect changes in positivesymptoms seen in schizophrenia and are accompaniedby an increased proportion of striatal DHigh2 receptorswhich may contribute to dopamine supersensitivitydemonstration the utility of animal models to determineunderlying molecular mechnaisms of the behaviouralimpact observed. Furthermore when isolation is combinedwith neonatal treatment with phencyclidine in the form ofa ‘dual adverse hit’ resultant offspring show reduced socialinteraction with an age- and weight-matched conspecificwhich may serve as a very useful index of the negativesymptoms of schizophrenia that has proved very difficult tomodel in rodents. Several of the neurobiological changes

in the brain of isolation-reared rats resemble some ofthose seen in patients with schizophrenia, suggesting themodel has good construct validity. For example, isolationreared rats show a selective reduction in prefrontal cortexvolume measured by MRI, accompanied by decreaseddendritic spine density and altered spine morphologyand a reduction in parvalbumin positive GABAergicchandelier cartridges on interneurones in the prefrontalcortex. Cytoskeletal alterations and a selective loss ofparvalbumin and calbindin-containing interneurones havealso been identified in the hippocampus of isolation rearedrats, similar to changes seen in schizophrenic patients.This presentation will describe the behavioural andneurochemical abnormalities and findings from treatmentwith current antipsychotics (such as risperidone) or noveldrugs acting on dopaminergic [3] and non-dopaminergictargets, such as 5-HT6 receptor antagonists, mGluR2/3agonists, memantine and nicotinic agonists, to reversethese changes and illustrate the predictive validity of theparadigm.

Reference(s)

[1] Jones, C.A., Watson, D.J.G. & Fone K.C.F. 2011.Animal models of schizophrenia. Brit J Pharmacol164, 1162–1194.

[2] Fone, K.C.F. & Porkess, M.V. 2008. Behaviouraland neurochemical effects of post-weaning socialisolation in rodents − relevance to developmentalneuropsychiatric disorders. Neurosci & Biobehav Rev32, 1087–1102.

[3] Watson, D.J.G., Loiseau, F., Ingaleseni, M., Mil-lan, M.J. & Fone, K.C.F. 2012. Selective blockadeof dopamine D3 receptors enhance while D2 receptorantagonism impairs social novelty discrimination andnovel object recognition in rats: a key role for theprefrontal cortex. Neuropsychopharmacol 37, 770–786.

Disclosure statement: The work associated with thispresentation has been funded by several Pharmaceuti-cal companies including GSK, Shire Pharmaceuticals,F.Hoffmann-La Roche, Institut de Recherches Servier andLaboratorios Dr Esteve.

Behavioural pharmacology S27

Posters

P.2.001 Citalopram given to stressed and controlpregnant rats causes sex-dependentchanges in behaviour and CRH mRNAexpression in their offspring

I. Zohar1 °, M. Weinstock1. 1Hebrew University ofJerusalem, Pharmacology, Jersualem, Israel

Rationale: Exposure to stressful life events duringpregnancy increases the likelihood of anxiety anddepression in the young and adult offspring of humans andexperimental animals. An increasing number of pregnantwomen exposed to stress and/or suffering from depressionare being treated with antidepressant drugs that areselective serotonin reuptake inhibitors (SSRIs) but it isnot clear whether any benefits of treatment outweigh thepotential adverse effects on the offspring.

Objectives: To compare the effect of maternal treatmentwith citalopram, a frequently prescribed SSRI, onbehaviour in prenatally-stressed (PS) and control ratsof both sexes. Since we had previously found that theincreased anxiety induced by prenatal stress was associatedwith sex-dependent alterations in the gene expression ofmembers of the corticotropin-releasing hormone (CRH)family in the amygdala and the paraventricular nucleusin the hypothalamus (PVN) [1] we also determined howthese were affected by maternal citalopram treatment.

Methods: Citalopram (10mg/kg/day) was administeredin the drinking fluid to pregnant individually housedrats from day 7 of gestation until after their pups wereweaned when aged 21 days. Half of the pregnant rats weresubjected to once-daily varied stress from day 14−21 ofgestation. Each stressor was shown to elevate maternalplasma COR more than threefold and there was no signof adaptation to them. Offspring behaviour and geneexpression of the CRH family, CRHR1, CRHR2 and CRHbinding protein (CRH-BP) was evaluated in the amygdalaand the PVN in adulthood.

Results: Stressed females were more anxious thancontrols three weeks after parturition and citalopramtreatment prevented the increased anxiety. Adult PSmales and females were also more anxious than controlsin the elevated plus maze and showed depressive-likebehaviour in the forced swim test. Maternal citalopramtreatment reduced anxiety in PS females but not in malesand did not affect depressive-like behaviour in eithersex. Moreover, citalopram treatment of control mothersinduced depressive-like behaviour in the offspring of bothsexes. In both the amygdala and the PVN, PS females hada higher expression of CRH mRNA and lower expression

of CRHR2 mRNA than controls and males had a lowerexpression of CRH-BP and of CRHR2 which could explaintheir increased anxiety. Citalopram treatment had littleeffect on CRH signalling in PS males in both brain areasbut reduced the expression of CRH, its binding proteinand receptors in PS females in the amygdala and increasedCRHBP in the PVN.

Conclusion: Although citalopram can prevent anxietyin stressed rat mothers it does not normalise behaviour oftheir offspring and even induces anxiety in those of controlrats. It is possible that blockade of the 5HT transporter bycitalopram at a critical time during development could leadto reduced activation of postsynaptic 5HT1A receptorswhich changes CRH signalling in the amygdala and thePVN and induces depressive-like behaviour in adulthood.

Reference(s)

[1] Zohar I, Weinstock M., 2011. Differential effect ofprenatal stress on the expression of corticotrophin-releasing hormone and its receptors in the hy-pothalamus and amygdala in male and female rats.J Neuroendocrinol 23(4):320–328.

P.2.002 Behavioural, molecular andglutamatergic changes in adevelopmental model of schizophrenia,and reversal by a 5-HT6 receptorantagonist

M.V. King1 °, O. Negm2, P. Tighe2, S. Knapp1,P. Wigmore1, K.C.F. Fone1. 1The University ofNottingham, School of Biomedical Sciences, Nottingham,United Kingdom; 2The University of Nottingham,Molecular Medical Sciences, Nottingham, UnitedKingdom

Post-weaning social isolation in the rat, a neurodevelop-mental model of schizophrenia, impairs memory in severaltasks which map to distinct cognitive domains deficientin this disorder, including hippocampal-dependent novelobject discrimination (NOD) and conditioned emotionalresponses (CER). These impairments are accompanied bystructural alterations within the hippocampus, includingreduced cell proliferation and survival, dendritic lengthand spine density. Reduced parvalbumin and calbindin-containing GABA interneurones and decreased expressionof Vesicular Glutamate Transporter 1 (VGLUT1) indicatedisruption of inhibitory and excitatory neurotransmissionand also resemble changes seen in schizophrenia [1]. Incontrast 5-HT6 receptor antagonists reverse drug-induceddeficits in NOD and CER, and elevate hippocampal cellproliferation and glutamate efflux in group-housed rats [2],


so may potentially reverse isolation-induced cognitiveimpairments. The aims of this study were to utiliseprotein microarray, immunohistochemical and glutamatemicrosensor techniques to further characterise isolation-induced hippocampal changes, and examine the ability ofa 5-HT6 receptor antagonist, SB-399885, to reverse theneurochemical and behavioural alterations in this model.Male Lister hooded rats (University of Nottingham or

Charles River UK) were weaned on post-natal day 21−24and housed individually or in groups (3−4). Rats receivedminimal handling until assessment (starting 5−6 weekslater) of locomotor activity, NOD, and contextual andcue-mediated CER. Rats received i.p. vehicle (1% Tween80, 1ml/kg) or 10mg/kg SB-399885 on six occasions(n = 11/housing-treatment combination) either 30min priorto testing or immediately after CER acquisition topreclude potential nociceptive/affective confounds. Ratswere killed 24 h after the final injection and hippocampicollected for quantification of VGLUT1−3 expressionusing Western blots, cell proliferation using Ki67immunohistochemistry and expression of intracellularsignalling molecules using protein microarray. Glutamatesignalling in hippocampal slices from separate groupsof drug-free rats (n = 7/housing condition) was measuredusing enzyme-coated microsensors.Isolation-rearing prevented discrimination of the novel

object during NOD (P> 0.05 versus familiar, whereasgroup-housed P< 0.001), such that the discriminationratio (DR) was significantly reduced from 0.68±0.02 to0.56±0.03 (P< 0.01). Isolation-rearing also reduced cue-mediated freezing during CER retention from 245±12s to162±24s (P< 0.01), without affecting contextual freezing.SB-399885 restored NOD (P< 0.001 versus familiar)such that the DR increased to 0.71±0.03 (P< 0.01),and partially reversed the CER deficit (freezing duration193±17s) and the increase in VGLUT2 and decrease inKi67 expression (P> 0.05 versus isolate and group vehiclecontrols in each case). Isolation also elevated Rac/CDC42(Rho GTPases which regulate microtubule stabilisationand dendrite morphogenesis, and have been implicatedin schizophrenia), while SB-399885 decreased TAK1 andpSTAT3 (members of the SAPK/JNK signalling cascade,whose stimulation via Rac/CDC42 reduces pSTAT3) inisolates only. Isolation tended to lower basal extracellularglutamate in hippocampal slices, from 4.32±1.40mM to1.99±0.41mM (P= 0.0708), but the combination of 3mMSB-399885 and 120mM KCl stimulated glutamate releaseirrespective of housing.The 5-HT6 receptor antagonist reversed isolation-

induced cognitive impairments and partially reverseddeficits in hippocampal cell proliferation in this neu-rodevelopmental model of schizophrenia. These findingsfurther support the use of 5-HT6 antagonists to treat

cognitive dysfunction, and the value of isolation-rearedrats to investigate the underlying neurobiology ofschizophrenia and evaluate novel treatments for thecognitive symptoms.

Reference(s)

[1] Jones, C.A., Watson, D.J., Fone, K.C., 2011. Animalmodels of schizophrenia. Br J Pharmacol 164:1162–1194.

[2] King, M.V., Marsden, C.A., Fone, K.C., 2008. Arole for the 5-HT1A, 5-HT4 and 5-HT6 receptors inlearning and memory. Trends Pharmacol Sci 29:482–492.

P.2.003 Sub-anaesthetic ketamine modulatesintrinsic blood oxygen level-dependent(BOLD) connectivity between thehippocampus and the prefrontal cortexin the rat

N. Gass1 °, A. Sartorius1, A.J. Schwarz2, E. Schenker3,C. Risterucci4, M. Spedding3, L. Zheng1, A. Meyer-Lindenberg5, W. Weber-Fahr1. 1Central Institute ofMental Health Medical Faculty Mannheim University ofHeidelberg, Neuroimaging, Mannheim, Germany; 2EliLilly and Company, Translational Medicine, Indianapolis,USA; 3Institut de Recherches Servier, Neuroscience DrugDiscovery Unit, Croissy s/Seine, France; 4F. Hoffmann-LaRoche, CNS Biomarker Pharmaceuticals Division, Basel,Switzerland; 5Central Institute of Mental Health MedicalFaculty Mannheim University of Heidelberg, Departmentof Psychiatry and Psychotherapy, Mannheim, Germany

Purpose: Resting state fMRI (rsfMRI) is well-establishedin humans and has been shown to be sensitive topharmacological modulation. More recently, consistentintrinsic connectivity networks have been demonstratedin the rat, but drug effects on rsfMRI are only beginningto be characterised in the rodent. The utility of rsfMRIas a translational biomarker depends on (1) its sensitivityto pharmacological modulation in preclinical species and(2) the degree of convergence with effects using the samecompound in humans. Ketamine, a potent N-methyl-D-aspartate (NMDA) receptor antagonist, is of substantialcurrent interest both (a) as a pharmacological modelof glutamatergic dysfunction in psychiatric disease [1],and (b) as a rapidly acting antidepressant, effective intreatment-resistant depressive patients [2]. The aim ofthis work was to systematically characterise the effectsof ketamine on rsfMRI in the rat.

Methods: Male Sprague-Dawley rats (368–447 g) in4 parallel groups (N= 10/group) received either vehicle


(saline) or one of three sub-anaesthetic doses ofS-ketamine (5, 10 and 25mg/kg; s.c.). Rats were scannedunder 0.14mg/kg/h medetomidine infusion. Three 8.5-min rsfMRI datasets were acquired from each rat at9.4T scanner: pre-injection and 15 and 30min post-injection. Blood samples were taken from each rat afterscanning to determine exposure to ketamine. Seed-basedconnectivity mapping was used to test the hypothesisthat ketamine modulates functional connectivity within thehippocampal-prefrontal system. An ROI–ROI correlationanalysis, based on a parcellation of the brain into 44 atlas-derived regions, was also performed to profile the ketamineeffects throughout the brain.

Results: Pharmacokinetic/pharmacodynamic (PK/PD)image analysis revealed an increased functional con-nectivity between the hippocampus (except its ventralpart) and regions in the prefrontal cortex that positivelycorrelated with ketamine plasma levels. The effectswere strongest 30min post-injection. ROI–ROI correlationanalysis revealed dose-dependent increases in connectivitywithin prefrontal cortex (PFC) structures and between thePFC and cortical structures, in particular the temporal andparietal association cortices, 15min post-injection.

Conclusions: The observed increases in functionalconnectivity reveal possible neural mechanisms underlyingestablished behavioural effects of ketamine, includingincreased wakefulness and locomotor activity, and areconsistent with ketamine-induced increases in corticalEEG gamma band coherence. This pattern might resultfrom increased glutamate levels which could then bindother type of glutamate receptors (e.g. AMPA receptors)and also induce synaptogenesis similar to long-termpotentiation. Additionally, PFC increased connectivitycould also reflect a psychotomimetic aspect observedafter ketamine intake in humans. This study providesfurther evidence that rsfMRI is a sensitive probe ofcentral pharmacological effects in preclinical species, andcharacterizes the effects of ketamine − a tool compoundof considerable current interest in psychiatry research −on rsfMRI in the rat. These results provide an importantcomparator to (a) other preclinical modalities and(b) analogous rsfMRI experiments emerging in humans.

Reference(s)[1] Large, C., 2007. Do NMDA receptor antagonist

models of schizophrenia predict the clinical efficacy ofantipsychotic drugs? J Psychopharmacol 21(3): 283.

[2] Zarate, C.A. Jr., Singh, J.B., Carlson, P.J.,Brutsche, N.E., Ameli, R., Luckenbaugh, D.A.,Charney, D.S., Manji, H.K., 2006. A randomized trialof an N-methyl-D-aspartate antagonist in treatment-resistant major depression. Arch Gen Psychiatry63(8):856−64.

P.2.004 The dopamine b-hydroxylase inhibitornepicastat suppresses differentchocolate-motivated behaviours in rats

A. Zaru1 °. 1Neuroscience Institute − National ResearchCouncil of Italy, Department of Medicine, Monserrato(CA), Italy

Background and Aims: Chocolate is the most craved foodand its excessive consumption may lead to overweight andaddiction-like behaviours, including stress-induced relapseto chocolate-seeking and -taking. Animal models indicatethat the reinforcing and motivational effects of abusivedrugs and palatable foods are mediated by similar neuronalcircuitries and molecular mechanisms [1]; accordingly,compounds affecting the reinforcing and motivationalproperties of abusive drugs may also affect the reinforcingand motivational properties of palatable foods. Recent datademonstrated that the selective dopamine b-hydroxylaseinhibitor, nepicastat, suppressed the reinstatement ofcocaine-seeking behaviour in rats [2]. The present studywas therefore designed to investigate the effect ofnepicastat on different chocolate-motivated behaviours inrats.

Methods: Male Wistar rats were trained to self-administer a chocolate-flavoured beverage (CFB; 5%powdered Nesquik® in water) under an operant (lever-responding) procedure in daily 30-min sessions [3]. Aftertraining, rats were allocated to 3 separate experiments,assessing the effect of nepicastat (0, 25, 50, and 100mg/kg;i.p.) on: (a) reinforcing properties of CFB [to thisend, rats were exposed to a fixed ratio 10 scheduleof reinforcement, in which the response requirement(RR; i.e., the ‘cost’ − in terms of number of lever-responses − of each CFB presentation) was kept fixedthroughout the session] (Experiment 1); (b) motivationalproperties of CFB [rats were exposed to a progressive ratioschedule of reinforcement, in which RR was progressivelyincreased after the delivery of each reinforcer; the lowestratio not completed (breakpoint) was taken as measureof the motivational properties of CFB] (Experiment 2);(c) reinstatement of CFB-seeking [after extinction of CFBself-administration, non-reinforced lever-responding forCFB was triggered by the non-contingent presentation(‘priming’) of CFB] (Experiment 3). Experiment 4assessed the effect of nepicastat on spontaneous locomotoractivity.

Results:All rats easily acquired and steadily maintainedCFB self-administration, with baseline values of ~1200lever-responses and ~50 ml/kg CFB consumed per session.In Experiment 1, treatment with 25, 50, and 100mg/kgnepicastat reduced by ~15%, 35%, and 45%, respectively,the number of lever-responses for CFB [F(3,33) = 14.85,


P< 0.0001]. In Experiment 2, treatment with 25, 50, and100mg/kg nepicastat reduced by ~15%, 35%, and 65%,respectively, the breakpoint for CFB [F(3,24) = 10.60,P< 0.0005]. In Experiment 3, limited and non-contingentpresentation of CFB robustly reinstated lever-responding;treatment with 25, 50, and 100mg/kg nepicastat reducedby ~50%, 60%, and 95%, respectively, the number of lever-responses (CFB-seeking) [F(3,28) = 10.45, P< 0.0001]. InExperiment 4, no dose of nepicastat affected locomotoractivity [F(3,31) = 1.57, P> 0.05].

Conclusions: Treatment with non-sedative doses ofnepicastat reduced the reinforcing and motivationalproperties of CFB and totally suppressed the reinstatementof CFB-seeking in rats. The suppressing effect ofnepicastat on reinstatement of CFB-seeking replicatedthe drug effect on reinstatement of cocaine-seeking [2],suggesting that common neuronal circuitries underlierelapse-like behaviours for different rewards. Based onthe mechanism of action of nepicastat, these results implya role for brain noradrenaline and dopamine in CFB-seeking and -taking behaviours. These results suggestthat nepicastat is potentially effective in the treatment of‘chocoholism’ and addictive-like, food-related behaviours.

Reference(s)[1] Volkow, N.D., Wise, R.A., 2005. How can drug

addiction help us understand obesity? Nat Neurosci8, 555–560.

[2] Schroeder, J.P., Cooper, D.A., Schank, J.R.,Lyle, M.A., Gaval-Cruz, M., Ogbonmwan, Y.E.,Pozdeyev, N., Freeman, K.G., Iuvone, P.M., Ed-wards, G.L., Holmes, P.V., Weinshenker, D., 2010.Disulfiram attenuates drug-primed reinstatementof cocaine seeking via inhibition of dopamineb-hydroxylase. Neuropsychopharmacology 35, 2440–2449.

[3] Maccioni, P., Pes, D., Carai, M.A.M., Gessa, G.L.,Colombo, G., 2008. Suppression by the cannabinoidCB1 receptor antagonist, rimonabant, of the rein-forcing and motivational properties of a chocolate-flavoured beverage in rats. Behav Pharmacol 19, 197–209.

P.2.005 The trans-isomer of resveratrol acutelyimproves motivation in rat

J. Samardzic1 °, L. Gojkovic-Bukarica1, D.I. Obradovic1.1Medical Faculty University of Belgrade, PharmacologyClinical Pharmacology and Toxicology, Belgrade, Serbia

Purpose of the study: Resveratrol (3,5,4′-trihydroxy-trans-stilbene), a natural non-flavonoid polyphenol antiox-idant, exists as two geometric isomers: cis- and trans-.

It has been shown that more stable trans-resveratroldemonstrates a variety of pharmacological activitiesincluding antioxidant, anti-inflammatory, neuroprotectiveproperties and amelioration of learning and memoryimpairment [1−3]. However, its behavioural profile stillremains controversial. The goal of the present study wasto examine the influence of trans-resveratrol and compareits dose-response effects on memory and depression-likebehaviour.

Methods used: We independently studied the effectsof trans-resveratrol (5−20mg/kg) on retention versusacquisition of active avoidance (AA) and depression-like behaviour in the forced swim test (FST). AAtest was performed in automated two-way shuttle boxesand programming recording units (Campden Instruments,Sileby, UK). In the first part of the study, the activeavoidance task was elaborated by 100-trial 2-day sessions,and in the second part the influence of trans-resveratrolon the acquisition rate was checked in a procedurelasting five consecutive days, with 50 trials per day. FSTwas performed in a glass cylinder, 45 cm high, 20 cmdiameter filled with water up to a height of 30 cm, with atemperature of 21−23ºC. Male Wistar rats were exposed totwo swimming sessions (an initial 15-min pretest session,followed 24 h later by a 5-min test session). A ratwas considered immobile whenever it floated passivelyin the water and only made movements necessary tokeep its head above the water line. We also tested thelocomotion to exclude the excitatory or inhibitory effects.The measurement of locomotor activity was performedin a clear Plexiglas box (40×25×35 cm), for 30min,without any habituation period. For all the experiments,the behaviour of the animals was recorded by a digitalcamera, and the data were analysed by one-way ANOVA,followed by Dunnett’s t test.

Summary of results: Treatment with trans-resveratrolsignificantly affected retrieval of avoidance responses onthe second day of shuttle box testing (F(4,45) = 2.88,p< 0.05). Dunnett’s test indicated that the trans-resveratrolavoidance-facilitatory dose was 20mg/kg. However, itdid not induce significant differences in acquisitionrate during 5 days training. In FST, during the testsession, the average immobility time of animals, inseconds, for the vehicle and resveratrol (5, 10, 20mg/kg)was 145.3, 147.3, 94.7 and 66.0, respectively. ANOVAindicated statistically significant effects of resveratrol(F(3,20) = 50.95, p< 0.001). Dunnett’s analysis showedthat resveratrol significantly decreased immobility time atthe doses of 10 and 20mg/kg, exerted acute antidepressant-like effects. ANOVA did not show a significant effect oftreatment on the total immobility time of the animalsduring 30min of monitoring of spontaneous locomotoractivity (F(3,20) = 1.68, p> 0.05).


Conclusions: Our results experimentally supportthe findings that under certain circumstances, trans-resveratrol, produces acute memory-enhancing andantidepressant-like effects. Furthermore, these effects werenot confounded by locomotor influences. This study willbe extended in the further characterising the behaviouralprofile of resveratrol, as well as clarifying the probablecentral mechanism of its action.

Reference(s)

[1] Kumar, A., Naidu, P.S., Seghal, N., Padi, S.S.,2007. Neuroprotective effects of resveratrol againstintracerebroventricular colchicine-induced cognitiveimpairment and oxidative stress in rats. Pharmacology79, 17−26.

[2] Ranney, A., Petro, M.S., 2009. Resveratrol protectsspatial learning in middle-aged C57BL/6 mice fromeffects of ethanol. Behav Pharmacol 20, 330–336.

[3] Xu, Y., Wang, Z., You, W., Zhang, X., Li, S., Barish,P.A., Vernon, M.M., Du, X., Li, G., Pan, J., Ogle, W.O.,2010. Antidepressant-like effect of trans-resveratrol:Involvement of serotonin and noradrenaline system.Eur Neuropsychopharmacol 20, 405–413.

P.2.006 Effects of serotonin (5-HT)1B receptorligands on amphetamine-seekingbehaviour in rats

J. Miszkiel1 °, E. Przegalinski1. 1Polish Academy ofSciences, Department of Pharmacology, Krakow, Poland

Background: Amphetamine and cocaine addiction is aserious medical and social problem; however, despite manyyears of research, there is still no efficient therapy [1].Both drugs belong to the psychostimulants and amongother behavioural effects they show rewarding properties.These effects − evoked by enhanced activation of the braindopamine system, especially by elevated extracellulardopamine level in the nucleus accumbens − determine theiraddictive properties [2]. Interestingly, it was establishedthat manipulation of the serotonin (5-HT)1B receptor canmodify the behavioural effects of these psychostimulantsincluding their reinforcing activity [3]. However, the roleof the 5-HT1B receptor in amphetamine self-administrationand extinction/reinstatement model is still unclear. In orderto substantiate a role of those receptors in incentivemotivation for amphetamine we used the extinction/reinstatement model to examine the effects of the5-HT1B receptor ligand on reinforcement of extinguishedamphetamine-seeking behaviour.

Methods: Male Wistar rats (250–300 g) were trainedto self-administer amphetamine in a standard operant

chambers, during 2-hour daily sessions, 6 days per week(maintenance). The FR schedule was gradually elevatedfrom FR1 to FR5 once animals showed the stableresponding. Every each 5 completed active lever pressesresulted in a 5-s infusion of amphetamine and a 5-spresentation of a stimulus complex. Rats were maintainedon the 0.06mg/kg/infusion amphetamine dose until theymet the criteria. When rats met the self-administrationcriteria, the extinction procedure was begun on thefollowing day. During the extinction animals receivedintravenous saline infusions (instead of amphetamine)without presentation of the conditioned stimulus (lightand sound). Once they reached the extinction criteria ratswere divided into separate groups to run reinstatementexperiment. Reinstatement was triggered by amphetamineinjection (i.p.) or the presentation of the stimulus complexpaired with threshold dose of amphetamine (i.p.). Separategroups of rats were pretreated with either SB 214461(a 5-HT1B receptor antagonist; 2.5−7.5mg/kg; i.p.), CP94253 (a 5-HT1B receptor agonist; 0.03−5mg/kg; i.p.)or both before the appropriate test sessions. Data wereanalysed by t-Student test or by one- or two-way analysisof variance followed by Dunnett or Newman–Keuls tests.

Results: The 5-HT1B receptor antagonists SB 216641(5–7.5mg/kg) attenuated the amphetamine (1.5mg/kg)-[F(3,24) = 9.351, p< 0.001] and amphetamine-associatedcue combined with the threshold dose of amphetamine-induced [F(3,25) = 4.808, p< 0.01] reinstatement ofamphetamine-seeking behaviour. The 5-HT1B recep-tor agonist CP 94253 (1.25−5mg/kg) also inhibitedamphetamine-seeking behaviour induced by amphetamine(1.5mg/kg) [F(5,38) = 7.963, p< 0.0001] but not by acue combined with threshold dose of amphetamine-induced [F(4,31) = 0.507] reinstatement. The inhibitoryeffect of CP 94253 on amphetamine-seeking behaviourremained unaffected [F(1,21) = 0.16] by this 5-HT1Breceptor antagonist.

Conclusion: Our results indicate that tonic activationof 5-HT1B receptors is involved in amphetamine-and amphetamine associated cue combined with thethreshold dose of amphetamine-induced reinstatement ofamphetamine-seeking behaviour and that the inhibitoryeffects of 5-HT1B receptor antagonists on these phenomenaare directly related to motivational aspects of amphetamineabuse. The inhibitory effect of CP 94253 on amphetamine-seeking behaviour seems to be unrelated to the 5-HT1Breceptor activation and may result from a general reductionof motivation.

Reference(s)

[1] Jupp, B., Lawrence, A.J. 2010. New horizons fortherapeutics in drug and alcohol abuse. PharmacolTher. 125, 138–168.


[2] Pontieri, F.E., Tanda, G., Di Chiara, G. 1995.Intravenous cocaine, morphine, and amphetaminepreferentially increase extracellular dopamine in the‘shell’ as compared with the ‘core’ of the rat nucleusaccumbens. Proc Natl Acad Sci U S A. 19, 12304–12308.

[3] Miszkiel, J., Filip, M., Przegalinski, E. 2011. Roleof serotonin (5-HT)1B receptors in psychostimulantaddiction. Pharmacol Rep. 63, 1310–1315.

P.2.007 Tryptophan depletion impairs emotionrecognition in healthy women

M. Defrancesco1 °, W. Parson2, J. Marksteiner3,E.A. Deisenhammer1, J. Kemmler1, H. Niederstatter2,H. Hinterhuber1. 1Medical University Innsbruck,Department of General Psychiatry, Innsbruck, Austria;2Medical University Innsbruck, Institute of LegalMedicine, Innsbruck, Austria; 3LKH Hall, Department ofPsychiatry, Innsbruck, Austria

Purpose of the study: The brain serotonin systemmodulates neural circuits that regulate emotion and moodin humans [1]. Tryptophan depletion (TD) is an establishedmethod to decrease cerebral serotonin levels [2]. The sero-tonin transporter promoter polymorphism (5-HTTLPR)is suggested to influence emotion recognition [3]. Thisstudy examined the effect of TD on the recognition ofemotional faces in healthy women carrying a s-allele ofthe 5-HTTLPR polymorphism.

Methods: A prospective clinical study was conductedfor 38 healthy women carrying an s-allele of the5-HTTLPR polymorphism. Participants completed com-puterised tests (Penn Emotion Recognition Test, and PennEmotion Acuity Task) and a visual analogue scale (VAS)before, five and seven hours after the administration ofacute TD. Blood samples were taken at these three timepoints to analyse biological markers (plasma serotonin,total plasma tryptophan (Trp), 5-hydroxyindolacetic acid(5-HIAA), long neutral amino acids (LNAAs), Trp /LNAAratio). Comparability of the two genetic subtypes withrespect to socio-demographic variables was checked bymeans of the adequate statistical tests (t-test, Mann–Whitney U-test, chi-square test), depending on thetype of variable. Repeated-measures analysis of variance(ANOVA) was performed to analyse the biochemicalparameters in the course of time (baseline, 5 hours and7 hours after TD). Genotype (5-HTTLPR) was included inthe model as a between-subjects factor allowing statisticaltesting for both a main effect of genotype and time-by-genotype interaction.

Results: Thirty-eight female participants with a meanage of 23.18±1.96 (mean±SD), mean years of educationof 16.65±1.69 and a mean BMI of 21.14±2.31 completedthe study. All participants had euthymic depressionscores in Beck Depression Inventory at baseline testing0.97±1.97. After TD, mood (VAS) decreased significantlywithin 5 hours (p< 0.001) and then increased againafter 7 hours (p = 0.020) compared to 5 hours afterTD. Further, total plasma Trp, 5-HIAA, and Trp/LNAAratio were significantly decreased by TD over time.Concerning genotype, repeated measures ANOVA showeda significant time-by-genotype interaction regarding thetime course of Trp plasma levels (p = 0.038). Homozygouscarriers of the 5-HTTLPR s-allele (−10.6mg/ml ±2.5)showed a significantly stronger reduction of plasma Trpthan heterozygous individuals (−8.2mg/ml ±3.0) betweenbaseline and 5 hours after TD (p = 0.026). Comparedto baseline, subjects recognised angry faces significantlybetter 7 hours after TD (p< 0.001). TD increased thespecificity for the emotions sad and happy but decreasedthe sensitivity for the emotion sad over the course of thetest day.

Conclusions: In summary, we found that TD impairsthe sensitivity for negative emotions in faces inhealthy females carrying the s-allele of the 5-HTTLPRpolymorphism. In addition, our data suggest that changesin plasma levels of tryptophan, 5-HIAA and the Trp/LNAAratio are directly associated with the mood lowering effectof TD in genetically vulnerable healthy females. Theseresults substantiate the putative role of the serotonergicsystem in the processing of emotional faces.

Reference(s)

[1] Heninger G.R., Charney D.S., and Sternberg D.E.1984. Serotonergic function in depression. Prolactinresponse to intravenous tryptophan in depressedpatients and healthy subjects. Archives of GeneralPsychiatry 41: 398–402.

[2] Marsh A.A., Finger E.C., Buzas B., Soliman N.,Richell R.A., Vythilingham M., Pine D.S., GoldmanD., and Blair R.J.R. 2006. Impaired recognition offear facial expressions in 5-HTTLPR S-polymorphismcarriers following tryptophan depletion. Psychophar-macology 189: 387–394.

[3] Roiser J.P., Muller U., Clark L., and Sahakian B.J.2007. The effects of acute tryptophan depletion andserotonin transporter polymorphism on emotionalprocessing in memory and attention. InternationalJournal of Neuropsychopharmacology 10: 449–461.


P.2.008 5XFAD mouse model of Alzheimer’sdisease: a dissociation between brainpathology and behavioural phenotype

K. Sonn1 °, K. Jaako1, R.K. Jain1, A. Zharkovsky1.1University of Tartu, Dept. of Pharmacology, Tartu,Estonia

Alzheimer’s disease (AD) is a neurodegenerative dis-ease clinically characterised by progressive cognitiveimpairment and pathologically by extracellular b-amyloid(Ab)-containing senile plaques, intraneuronal aggregatesof hyperphosphorylated t-protein and neurofilaments aswell as synaptic loss. The amyloid cascade hypothesispostulates that deposition of toxic Ab species is theprimary cause of AD. 5XFAD mice is a transgenic modelof AD known for rapid brain accumulation of Ab [1].Our aim was to characterise behaviour and brain

findings of 5XFAD mice dynamically in order to gaininsight into pathophysiology of AD and to identify possiblenew drug targets. Male and female 5XFAD mice and wild-type (WT) littermates of 2 and 6 months of age were used;n = 5−15.Morris water maze test was performed on 6 consecutive

days. Training lasted 4 days, platform was made visibleon day 5 and removed on day 6. Two months old 5XFADmice displayed impaired learning (no decrease in timeto find hidden platform during training days), but notmemory (% time spent in quadrant of the pool whereplatform had been) compared to WT mice. Swimmingvelocity and vision did not differ between groups at2 months. At 6 months of age, there was no difference inlearning, memory or vision between 5XFAD and wild-typemice. However, swimming velocity of 5XFAD mice wassignificantly lower at 6 months compared to WT mice.Elevated plus maze task was performed on 3 consecutive

days. 5XFAD mice demonstrated a decrease in % of openarm entries on days 2 and 3 at 2, but not 6 months ofage. At 2 but not 6 months, 5XFAD mice showed a trendtowards increased % of open arm entries compared toWT mice.At 2 months, 5XFAD mice had a significant increase

in microglial activation (demonstrated immunohistochem-ically by a greater Iba-1 positive area fraction inhippocampus and amygdala) compared to WT mice.The difference increased further by 6 months. Diameterof Congo-red positive Ab plaques in hippocampus andamygdala significantly increased in 5XFAD mice between2 and 6 months of age.The number of polysialylated neural cell adhesion

molecule (PSA-NCAM)-positive cells in the hippocampuswas significantly higher in 2, but not in 6 monthsold 5XFAD mice. By 6 months, the number of PSA-

NCAM-positive cells decreased significantly in 5XFADand WT mice with no difference between the groups.Immunoblotting showed significantly higher levels ofPSA-NCAM in 6 months old and a trend for increasein 2 months old 5XFAD mice.Levels of synaptic proteins synaptophysin and PSD-

95 and neurofilament H as well as phosphorylatedneurofilament H were measured by Western blot inhippocampus and cortex of 5XFAD and WT mice at 2 and6 months. No difference in the levels of either synapticor axonal markers was found in either age group betweenexperimental groups.Our data suggest existence of compensatory mecha-

nisms in 5XFAD mice such as enhanced neurogenesis orneurite outgrowth which need identification. Additionally,our findings raise the question of suitability of 5XFADmice as an AD model.

Reference(s)

[1] Oakley H, Cole SL, Logan S et al., 2006. Intra-neuronal beta-amyloid aggregates, neurodegeneration,and neuron loss in transgenic mice with fivefamilial Alzheimer’s disease mutations: potentialfactors in amyloid plaque formation. The Journal ofNeuroscience 26, 10129–10140.

P.2.009 Effect of early life experiences on brainstructure and function: neurogenesisand decision making

M. Loi1 °, S. Koricka1, L. de Visser1, M.J.H. Kas1,P.J. Lucassen2, M. Joels1. 1Rudolf Magnus Institute ofNeuroscience, Neuroscience and Pharmacology, Utrecht,The Netherlands; 2Swammerdam Institute of Life Sciences,Centre for Neuroscience, Amsterdam, The Netherlands

Exposure to stress during early postnatal life can alteradult brain structure and function and can e.g. enhancestress responsiveness later in life and increase vulnerabilityfor psychiatric disorders. Previous studies suggest that4 days of treatment with mifepristone prevents a reductionin hippocampal neurogenesis induced by chronic stressthrough a blockage of the glucocorticoid receptors [1].To investigate effects of early life stress on neurogenesis

and decision making, we used the maternal deprivation(MD) paradigm in rats that leads to increased levels ofcirculating corticosterone (CORT). First we wish to studythe effects of MD on neurogenesis at postnatal day (PND)29. Second, we shall assess the effects of MD on decision-making processes during young-adulthood. Finally, theeffects of glucocorticoid receptor blockade mifepristone(RU38486) during early adolescence will be examinedboth on structural and behavioural alterations inducedby MD.


Wistar rats were exposed to 24 h MD at PND3. OnPND21, pups were group housed. At PND 26−28 halfof the maternal deprived and control groups receivedeither mifepristone or vehicle (VEH) treatment throughoral gavage administration. At PND29, females wereperfused. Proliferation and neurogenesis were quantifiedusing immunohistochemistry for Ki-67 and doublecortin,respectively. To assess funcional effects of MD onexecutive functioning, 12 week old male rats performedthe Rodent Iowa Gambling Task (r-IGT) that measures theability to choose a long-term advantageous option withimmediate moderate rewards, over a long-term disadvan-tageous option with immediate high rewards. After the lasttrial, brains were processed for c-fos immunochemistry.Good performing rats (no MD, and MD treated with

mifepristone groups), showed a higher fraction of visitsinto the advantageous arm: NoMD+VEH (mean±SEM:0.6±0.2 n = 8); NoMD+RU38486 (mean±SEM: 0.5±0.1n = 11); MD+RU38486 (mean±SEM: 0.5±0.1 n = 12)compared to saline-treated maternal deprived animals(mean±SEM: 0.2±0.1 n = 12). Good performing rats,during the training, improved in choosing the long-termadvantageous arm while MD+VEH rats did not improve.Post-hoc comparison revealed significant difference in per-formance between NOMD+VEH vs MD+VEH (p = 0.015)as well as between MD+MIF compared to MD+VEH(p = 0.025). Good performing rats showed a lower fractionof visits into the empty arms (mean±SEM: 0.3±0.08) thanpoor performing rats (mean±SEM: 0.5±0.08), a measureof non-reward related exploration.Our data show MD impairs adult decision-making

performance, as deprived rats did not learn to choosefor the long-term advantageous option. Treatment withRU38486 during early adolescence rescued the negativeeffects of MD and improved task performance. As r-IGTperformance relies on the integrity of the prefrontal cortex,we hypothesize the poor performance in the r-IGT may becaused by MD effects on prefrontal cortex development.

Reference(s)

[1] Oomen CA, Mayer JL, de Kloet ER, Joels M,Lucassen PJ., 2007. Brief treatment with the gluco-corticoid receptor antagonist mifepristone normalizesthe reduction in neurogenesis after chronic stress. EurJ Neurosci 26: 3395–3401.

Disclosure statement: Supported by Corcept Inc. PJL issupported by the Dutch Brain Foundation.

P.2.010 Effects of L-DOPA and sarizotantreatments in a parkinsonian rat modelof depression

N. Schintu1 °, X. Zhang1, A.A. Mathe1, P. Svenningsson1.1Karolinska Institute, Clinical Neuroscience, Stockholm,Sweden

Parkinson’s disease (PD) is the second most common neu-rodegenerative disorder. The diagnostic symptoms of PDare based on the motor dysfunctions, bradykinesia, tremor,rigidity and postural imbalance. The standard drug for PDis L-DOPA, but its therapeutic efficacy is gradually lostand dyskinesias appear after 5−10 years of treatment. Onepossible explanation for the development of dyskinesia isthat, when most dopaminergic neurons are degenerated,L-DOPA is converted to dopamine in serotoninergic neu-rons and thus released in a non-controlled manner [1]. Inthis context, serotonin 5-HT1A agonists, such as sarizotan,have been found to have antidyskinetic properties. Besidemotor symptoms, PDs patients have non-motor symptomsof which depression is the most frequent. Because of theincreased awareness of the negative impact of depressionfor the quality of life of PD patients, there is a growinginterest in developing optimised treatments that alleviateboth motor and non-motor symptoms of PD.In this study, unilateral 6-OHDA lesioning was

performed in the Flinders Sensitive Line (FSL), a rat strainassociated with distinct behavioural and neurochemicalfeatures of major depression, and their littermates, theFlinders Resistant Line (FRL). Two weeks later, ratswere injected with apomorphine (1mg/kg) and theircontrolateral rotations were measured over 30 minutes todetermine the success of nigrostriatal denervation. The ef-fectiveness of the lesion was further confirmed postmortemby dopamine transporter autoradiography in the striatum.Four weeks after surgery, rats were administered withsaline, L-DOPA/benserazide (10/7.5mg/kg), sarizotan(2.5mg/kg) alone or in combination with L-DOPA oncedaily for 23 days. Once per week, rotational behaviourand abnormal involuntary movements (AIMs) [2] weremeasured. 30 minutes after the last injection, ratswere sacrificed and brains rapidly removed for in situhybridisation study of immediate early genes includingc-fos in the striatum.The results showed that the first administration

of L-DOPA and L-DOPA/sarizotan induced similarcontralateral turning behaviour in both FRL and FSLrats. After chronic treatment with L-DOPA, a significantincrease in the number of rotations was observed inthe FRL group. In contrast, FRL rats receiving chronicco-administration of sarizotan/L-DOPA did not developbehavioural sensitisation. In the FSL group, chronic


treatment with neither L-DOPA nor L-DOPA/sarizotaninduced any significant behavioural sensitisation. Chronictreatment with L-DOPA and L-DOPA/sarizotan inducedAIMs in both lines of rats. However, FRL rats treatedwith L-DOPA spent more time in AIMs compared toFSL rats. Moreover, co-administration with sarizotanreduced significantly L-DOPA-induced AIMs in FRL, butnot in FSL rats. In situ hybridisation analysis showedthat treatment with L-DOPA and L-DOPA/sarizotansignificantly increased c-fos mRNA levels in the lesionedside as compared to the nonlesioned side of salinetreated FRL and FSL rats. Interestingly, c-fos mRNAlevel in FSL group after chronic L-DOPA treatmentwas significantly lower than in FRL group receivingthe same treatment. Moreover, co-administration withsarizotan reduced significantly L-DOPA-induced c-fosin FRL, but not in FSL, rats. The results suggestthat parkinsonian animals with distinct depressive-likephenotypes react differently to PD treatments in termsof dyskinetic behaviour and striatal neuronal-like activity.

Reference(s)[1] Carta M, Carlsson T, Kirik D, Bjorklund A. Dopamine

released from 5-HT terminals is the cause of L-DOPA-induced dyskinesia in parkinsonian rats. Brain. 2007Jul;130(Pt 7):1819−33. Epub 2007 Apr 23.

[2] Pinna A, Tronci E, Schintu N, Simola N, Volpini R,Pontis S, Cristalli G, Morelli M. A new ethy-ladenine antagonist of adenosine A(2A) receptors:behavioural and biochemical characterization as anantiparkinsonian drug. Neuropharmacology. 2010Mar;58(3):613−23.

P.2.011 GluA1 and its Postsynaptic densityprotein 95 (PSD-95)/Discs large/Zonulaoccludens-1 (PDZ)-interaction: a rolein experience-dependent behaviouralplasticity in the forced swim test

F. Freudenberg1 °, V. Marx2, V. Mack3, L.E. Layer3,M. Klugmann4, P.H. Seeburg3, R. Sprengel3, T. Celikel2.1University of Wurzburg, Department of PsychiatryPsychosomatics and Psychotherapy, Wurzburg, Germany;2Radboud University Nijmegen, Donders Institute forBrain Cognition and Behavior Centre for Neuroscience,Nijmegen, The Netherlands; 3Max Planck Institutefor Medical Research, Department of MolecularNeurobiology, Heidelberg, Germany; 4University ofNew South Wales, Department of Physiology, Sidney,Australia

Although current pharmacological treatment for depres-sion mainly targets the monoaminergic system, recent

studies indicate that glutamatergic neurotransmission ismore principally involved in the neuropathology ofdepression [1]. Importantly, pharmacological blockadeof NMDA receptors (e.g. with ketamine) has relativelylong-lasting antidepressant effects with rapid onset. Theantidepressant properties of ketamine require activationand synaptic incorporation of GluA1-containing AMPAreceptors [2]. Moreover, hippocampal samples fromclinically depressed patients display reduced mRNA levelsfor GluA1. These findings argue that GluA1-dependentsynaptic plasticity might be critically involved in thedevelopment of depression [1,2].Here we tested this hypothesis by exposing wild

type (WT) and GluA1 transgenic mice to two sessionsof forced swimming, 24-hours apart (FST1 and FST2respectively), and assessed experience-dependent expres-sion of behavioural despair by quantification of latency toimmobility and cumulative immobility (Table 1).

Table 1. Latency to immobility and cumulative immobility during the first (FST1)and second day (FST2) of testing for all genotypes.

Genotype Latency to immobility (s) Cumulative immobility (%)

FST1 FST2 FST1 FST2

WT (N= 10) 96.1±9.5 35.7±8.5** 46.3±5.3 60.5±2.6**GluA1−/− (N = 7) 284.0±66.8 244.7±75.4 25.9±3.2 31.4±6.4TG8.1 (N= 9) 221.3±61.3 210.8±59.0 26.7±1.1 26.2±2.0A1.1 (N= 7) 327.3±41.7 102±16.2*** 27.7±2.4 35.5±3.6*WT-Cre (N= 7) 201.7±37.2 65.1±9.9*** 35.6±2.3 47.4±2.2**DdHpc (N= 6) 120.5±15.3 132.2±39.6 40.0±1.8 39.3±3.0DvHpc (N= 13) 142.9±10.3 148.5±36.0 38.9±1.7 36.9±1.8Statistical significance after post-hoc Tukey’s t-test (FST1 vs FST2): *p<0.05,**p< 0.01, ***p< 0.001.

In WT mice latency was significantly reduced andimmobility was significantly increased during FST2(compared to FST1), suggesting successful experience-dependent induction of behavioural despair. Interestingly,mice globally lacking GluA1-containing AMPA receptors(GluA1−/− mice) were impaired in the experience-dependent expression of behavioural despair in the FST, assuggested by comparable latency and immobility on bothdays of testing. By transgenically expressing a GFP-taggedGluA1-subunit (GFPGluA1) in the GluA1−/− background(A1.1 mice) we were able to rescue this deficit. However,when the GFPGluA1-subunit lacked the final C-terminalamino acid (leucine) (TG8.1 mice), responsible for PDZ-interaction of GluA1 [3] the deficit seen in GluA1−/− miceremained (i.e. unchanged latency and immobility), arguingthat GluA1–PDZ interaction is critical for expression oflearned behavioural despair.To investigate the neural circuits involved, we selectively

removed GluA1 in hippocampus of mice with ‘floxed’GluA1 alleles by stereotaxic delivery of a Cre-expressing recombinant adeno-associated virus (rAAV).While control mice (WT mice injected with the Cre-


expressing rAAV; WT-Cre) showed normal reduction inlatency and increased immobility, mice preferentiallylacking GluA1 in dorsal (DdHpc) or ventral (DvHpc)hippocampus did not display any changes in latency andimmobility across sessions and were indistinguishablefrom GluA1−/− mice, suggesting that hippocampal GluA1-containing AMPA receptors are critical for the experience-dependent expression of behavioural despair in the FST.Taken together, this study provides evidence that

hippocampal GluA1-containing AMPA receptors and theirinteraction with PDZ-domain proteins are critical for theinduction of learned behavioural despair, thereby linkingmechanisms of hippocampal GluA1-dependent synapticplasticity with the pathophysiology of depression.

Reference(s)

[1] Skolnick, P., Popik, P., Trullas, R., 2009. Glutamate-based antidepressants: 20 years on. Trends inpharmacological sciences 30, 563–569.

[2] Duman R.S., Voleti B., 2012. Signaling pathwaysunderlying the pathophysiology and treatment ofdepression: novel mechanisms for rapid-acting agents.Trends in neurosciences 35, 47−56.

[3] Cai C., Coleman S.K., Niemi K., Keinanen K., 2002.Selective binding of synapse-associated protein 97 toGluR-A alpha-amino-5-hydroxy-3-methyl-4-isoxazolepropionate receptor subunit is determined by a novelsequence motif. The Journal of biological chemistry277:31484–31490.

P.2.012 Chronic modulation of 5-HT4 and 5-HT6

serotonergic receptors: a new hope inthe treatment of cognitive deficits?

A. Quiedeville1 °, T. Freret1, V. Bouet1, M. Boulouard1.1Universite de Caen Basse-Normandie, Groupe Memoireet Plasticite comportementale − EA4259, Caen, France

Drugs designed for patients suffering from neurodegener-ative diseases like Alzheimer’s act on a single moleculartarget, and have most of the time only symptomaticeffects. As hitting a single target seems more and moreinadequate, Multiple Medication Therapies (MMT) aregaining interest. With the aim to use a low dose of eachcomponent and therefore to diminish possible adverseeffects, MMT could represent a real improvement in thetreatment of neurodegenerative diseases.Serotonin is a central neurotransmitter known to

be involved in a great variety of physiological andcognitive functions, including learning and memoryprocesses. Among the fourteen subtypes of serotoninreceptors, the 5-HT4 and 5-HT6 receptors (5-HT4R

and 5-HT6R) are interesting therapeutic targets forthe treatment of memory disorders and their co-modulation could represent an innovative MMT strategy.Neurochemical and behavioural studies have shown thatactivation of 5-HT4R or blockade of 5-HT6R bothimprove memory performances. Accordingly, a therapeuticapproach combining a simultaneous modulation of thesetwo receptors could be an interesting and innovativestrategy in the treatment of memory disorders.In order to find the best therapeutic strategy, we first

selected sub-efficient doses of a 5-HT4R partial agonist(RS 67333) and of a 5-HT6R antagonist (SB-271046),administered chronically (14 days; i.p.) to adult maleNMRI mice. We chose to assess their effects in theobject recognition test, given that it assesses episodicmemory, which is the first memory affected by normalor pathological aging and Alzheimer’s disease. The testwas performed in a Y-maze [1], with an inter-sessionsinterval of 72 h, a duration that does not allow significantrecognition in normal mice and therefore is appropriate toobserve promnesiant effects. This test sets up a criterionof 20s of exploration for both objects, thus the explorationtime of the novel object will be compared to the chancelevel of 10s with a Student t test.Results showed that chronic activation of 5-HT4R by

RS 67333 improved episodic-like memory performances,at the dose of 5mg/kg [time spent exploring thenovel object, mean±SEM, Student t test versus 10 s,11.17±0.37, p = 0.004], but not at 1mg/kg [11.00±0.67,p = 0.073]. Chronic blockade of 5-HT6R by SB-271046improved memory performances at very low doses suchas 0.001mg/kg [11.42±0.34, p = 0.0007], but not below.We demonstrated that chronic activation of 5-HT4R

or blockade of 5-HT6R, both at very low doses,improved episodic-like memory performances. Besides,a preliminary study assessing the effects of a chronicco-modulation of these two receptors with sub-efficientdoses of RS 67333 and SB-271046 showed the highpotential synergistic effect of this strategy. These resultsbring new hopes in finding an effective strategy torestore memory performances both in physiological andpathological situations, along with potential suppressionof adverse effects.

Reference(s)

[1] Leger, M., Quiedeville, A., Paizanis, E., Natkunara-jah, S., Freret, T., Boulouard, M., Schumann-Bard, P.,2012. Environmental enrichment enhances episodic-like memory in association with a modified neuronalactivation profile in adult mice. PLoS ONE 7(10):e48043. doi:10.1371/journal.pone.0048043.


P.2.013 Modelling serotonergic neuromodulationof behavioural performance in spatialworking memory

M. Cano-Colino1 °, R. Almeida2, A. Compte1.1IDIBAPS, Theoretical Neurobiology, Barcelona, Spain;2Karolinska Institute, Department of Neuroscience,Stockholm, Sweden

The executive functions of prefrontal cortex (PFC) areregulated by ascending monoaminergic systems. In par-ticular, the dorsal PFC is associated with spatial workingmemory (SWM), linked to persistent neuronal activity,and it receives anatomical projections from the brainstemmonoamine centres. Catecholamine regulation of SWMhas been studied at mechanistic and computational levels.However, the role of serotonin (5-HT) in SWM isunclear. Thus, while 5-HT is clearly associated withorbitofrontal cortex functions, its association with SWMis controversial, with positive and negative reports. Anindirect association between PFC 5-HT and SWM issuggested by the pro-cognitive effects of antipsychoticdrugs targeting 5-HT receptors. Several neurocognitiveproblems are core deficits of schizophrenia and SWMdeficits mark a genetic predisposition for the disease.These weak associations between 5-HT and SWM are

in contrast with neurobiological observations predictinga stronger link. Indeed, 5-HT receptors are denselyexpressed in various PFC subfields, and serotonergicprojections from the raphe nuclei target specificallyPFC and modulate neuronal activity. Moreover, 5-HT2Areceptor activation/blockade in dorsal PFC reducespersistent neuronal activity during SWM tasks.Here, we sought to develop a computational model of

serotonergic modulation of the PFC network in order toaddress the possible reasons for this discrepancy. We useda computational network model of spiking neurons [1]which is consistent with PFC neurophysiological data frombehaving monkeys in SWM. This model falls within thegeneral class of attractor models proposed to accountfor behavioural deficits of schizophrenia, and providesa rich set of behavioural output predictions that can beexperimentally tested. We incorporated 5-HT receptormechanisms in the model following electrophysiologicalevidence, and we tested the effects of 5-HT receptorligands on network simulation performance during SWM.We found that 5-HT modulated network’s SWM

performance non-monotonously via 5-HT2A and especiallyvia 5-HT1A receptors, following an inverted-U shape.This may partly explain the weak behavioural effectsof serotonergic agents in previous SWM studies.Our simulations showed that errors committed at low

and high 5-HT concentrations are due to differentnetwork dynamics instabilities, suggesting that these twoconditions can be distinguished experimentally based ontheir dependency with delay length, on the responseconfidence declared in error trials and on PFC activationcontrasts in neuroimaging studies. We derived specificpredictions regarding the expected behavioural effects ofserotonin agents in two classic working-memory tasks: anoculomotor delayed-response task and a delayed match-to-sample task. In brief, the probability of memorystorage in the oculomotor task and the rate of missesin the match-to-sample task will be most sensitive toserotonin modulation, depending on delay duration forincreases but not for decreases in baseline 5-HT. Our studyunderscores the relevance of identifying different errortypes in SWM tasks in order to reveal the associationbetween neuromodulatory systems and SWM. Testingthese model-derived predictions in electrophysiologicaland psychophysical studies of SWM will help advanceour understanding of the neural basis of SWM and itsneuromodulation by 5-HT receptors.

Reference(s)

[1] Compte A., Brunel N., Goldman-Rakic P.S., WangX.J., 2000. Synaptic mechanisms and networkdynamics underlying spatial working memory in acortical network model. Cereb Cortex 10:910–923.

P.2.014 Serotonin receptor 2B knockout micepresent schizophrenic-like behaviour

P.M. Pitychoutis1 °, L. Maroteaux1. 1INSERM UMR-S839 Institut du Fer a Moulin, Universite Pierre et MarieCurie F75005, Paris, France

Purpose: Impulsivity, very broadly defined as actionwithout foresight, novelty-seeking and hyperlocomotion,shares common playground with numerous mentaldisorders, among them chronic substance abuse, attentiondeficit hyperactivity disorder, and schizophrenia [1]. In arecent study conducted in a Finnish cohort, we reportedthat a population-specific serotonin 2B receptor (5-HT2BR)stop-codon (i.e. HTR2B Q20*) predisposes to severeimpulsivity [2]. Moreover, the genetic ablation of the5-HT2BR gene in mice (5-HT−/−2B ) yielded a hyperloco-motor and novelty-seeker phenotype, characterised byenhanced impulsivity, as assessed in the delay discountingtask [2,3]. In the same cohort, psychosis was numericallymore prevalent in HTR2B Q*20 carriers, with early-onset schizophrenia being observed in this group [2].However, the extent to which 5-HT2BR is implicated in theneurobiology of schizophrenia has never been investigated


per se. Therefore, in the present study we investigated theeffects of the genetic ablation of the 5-HT2B receptor,across a battery of schizophrenia-relevant behaviouralparadigms and demonstrate herein that loss of functionof 5-HT2BR confers a wide spectrum of schizophrenic-like behavioural and psychopharmacological phenotypesin mice.

Methods: Adult male 5-HT−/−2B mice and their respectivewild-type (WT) control mice were used in the presentstudy. All mice were tested in the prepulse inhibitionof acoustic startle (PPI) and the three-compartmentalsociability and preference for social novelty tests. Putativeattentional and cognitive deficits were assessed in thenovel object recognition (NOR), the contextual and cuedfear conditioning and the latent inhibition (LI) paradigms.Locomotor response to the systemic administration ofthe psychostimulants dizocilpine (i.e. MK-801) andamphetamine was also assessed. Dopamine (DA) andglutamate content, as well as the mRNA expression ofdopaminergic and glutamatergic receptors was assessed inthe dorsal striatum (dSTR) and the nucleus accumbens(NAC) of 5-HT−/−2B and WT mice.

Results: Domains related to the positive, negativeand cognitive symptom-clusters of schizophrenia appearto be affected in 5-HT−/−2B mice. In particular, geneticablation of the 5-HT2BR induced deficits in sensorimotorgating (i.e. impaired PPI performance) and selectiveattention (i.e. lack of LI establishment). Moreover,5-HT−/−2B mice exhibited increased locomotor responseto the psychostimulant properties of dizocilpine andamphetamine, as well as deficits in social interaction, NORand fear memory that closely resemble the behaviouralalterations observed in schizophrenic patients. At theneurochemical level, ablation of the 5-HT2BR induceda region-selective decrease of DA concentrations in thedSTR and glutamate content in the NAC accompanied byrespective alterations in dopaminergic and glutamatergicreceptors’ expression.

Conclusions: Herein we report that 5-HT2BR deficiencymay induce a wide spectrum of schizophrenic-likebehavioural phenotypes accompanied by robust region-distinctive dopaminergic and glutamatergic neurochem-ical alterations. The latter implies an important rolefor 5-HT2BR in the regulation of dopaminergic andglutamatergic activity. This genetic mouse model holdsheuristic value in elucidating the relevant neurodevelop-mental, epigenetic, and physiological mechanisms thatmay be sensitive to 5-HT2BR genetic variations. Giventhat many currently marketed atypical antipsychotic drugspresent high affinity for the 5-HT2BR our findings bearbroader significance for the use of these drugs in psychoticdisorders.

Reference(s)

[1] Humby, T., Wilkinson, L.S., 2011. Assaying dissocia-ble elements of behavioural inhibition and impulsivity:translational utility of animal models. Curr OpinPharmacol 11, 534–539.

[2] Bevilacqua, L., Doly, S., Kaprio, J., Yuan, Q.,Tikkanen, R., Paunio, T., Zhou, Z., Wedenoja, J.,Maroteaux, L., Diaz, S., Belmer, A., Hodgkin-son, C.A., Dell’osso, L., Suvisaari, J., Coccaro, E.,Rose, R.J., Peltonen, L., Virkkunen, M., Goldman, D.,2011. A population-specific HTR2B stop codonpredisposes to severe impulsivity. Nature 468, 1061–1066.

[3] Doly, S., Valjent, E., Setola, V., Callebert, J., Herve, D.,Launay, J.M., Maroteaux, L., 2008. Serotonin 5-HT2Breceptors are required for 3,4-methylenedioxymeth-amphetamine-induced hyperlocomotion and 5-HTrelease in vivo and in vitro. J Neurosci 28, 2933–2940.

P.2.015 Low performing rats model theinattentive subtype of adult ADHD in the5-choice continuous performance task(5C-CPT)

A. Tomlinson1 °, K.M. Marshall2, J.C. Neill1. 1Universityof Bradford, School of Pharmacy, Bradford West Yorkshire,United Kingdom; 2University of Manchester, School ofPharmacy, Manchester, United Kingdom

The 5-choice continuous performance task (5C-CPT) isan enhanced version of the 5-choice serial reaction timetask (5-CSRTT). The 5C-CPT assesses vigilance in away that is similar to the human CPT, in comparisonto the 5-CSRTT that assesses sustained attention.The 5C-CPT allows for the measurement of responseinhibition in terms of false-alarm responding to non-signalstimuli [1,2]. Disturbances in attention and inhibitorycontrol play a central role in the symptomatology ofADHD. Measuring response inhibition is important whenvalidating animal models of ADHD because increasedfalse-alarm responding is often mediated by impairmentsin performance of the CPT in humans. Selection ofrats within a normal ‘population’ that display reducedsustained attention in 5-CSRTT training sessions mayprovide a more translational model of ADHD [3]. Thepresent study extends these findings by selecting rats thatdisplay reduced sustained attention and vigilance in boththe 5-CSRTT and the 5C-CPT.The aim of the current study was to investigate

the effects of psychostimulant and non-stimulant drugs


on attention, impulsivity and performance in adult ratsseparated into high and low-performers in the 5C-CPT.The effects of acute methylphenidate (MPH) 0.5, 1.0,

and 2.0mg/kg i.p, and atomoxetine (ATMX) 0.2, 1.0,2.0mg/kg i.p were assessed in the 5C-CPT in femaleLister-hooded rats (n = 40). Animals were trained for 60sessions and then divided into two groups (high and lowperformers) based on set criteria. Animals were challengedon test days by increasing the inter-trial interval (ITI) from5s to 10s.ATMX significantly reduced premature responding

in low performers at 1.0 and 2.0mg/kg (p< 0.05).ATMX (2.0mg/kg) significantly increased % accuracy andreduced false alarm rate (p< 0.05) in poor performers.ATMX (1.0mg/kg, 2.0mg/kg) significantly increasedsensitivity index in low performers (p< 0.05; p< 0.01),and significantly decreased responsivity index (p< 0.05)MPH at 2.0mg/kg significantly improved accuracy(p< 0.05), and reduced premature responding (p< 0.05)in low performers and increased premature responding inhigh performers (p< 0.05). MPH significantly increasedcorrect rejections in high performers at 2.0mg/kg(p< 0.05), and decreased correct rejections at all doses inlow performers (p< 0.01, p< 0.05, p< 0.05 respectively).MPH significantly increased sensitivity index in lowperformers at 0.5 and 1.0mg/kg (p< 0.01).In summary, ATMX (noradrenaline reuptake inhibitor,

non-stimulant) reduced impulsivity in low performersin both the 5-CSRTT (premature responses) and in the5C-CPT (false alarm rate). ATMX enhanced sustainedattention in the 5CSRTT and vigilance in 5C-CPT in lowperformers. Methylphenidate (dopamine/noradrenaline re-uptake inhibitor, psychostimulant) enhanced sustainedattention in the 5-CSRTT, vigilance in the 5C-CPT (%correct rejections) and marginally reduced impulsivity inlow performers in the 5C-CPT. This would suggest thatlow performers are more sensitive to the effects of bothstimulant and non-stimulant drugs. These data provide thevalidation of a rat model for the inattentive subtype ofadult ADHD. The model utilises the 5C-CPT to select forthose with deficits in sustained attention and vigilance,which can then be enhanced by ADHD medication.

Reference(s)

[1] Young, J.W., Light, G.A., Marston, H.M., Sharp, R.& Geyer, M.A. 2009. The 5-choice continuousperformance test: evidence for a translational test ofvigalence for mice PLoS One, 4, e4227.

[2] Young, J.W., Powell, S.B., Scott, C.N., Zhou, X. &Geyer, M.A. 2011. The effect of reduced dopamineD4 receptor expression in the 5-choice continuousperformance task: Separting response inhibition from

premature responding Behaviour and Brain Reseach,222, 183–192.

[3] Puumala, T., Ruotsalainen, S., Jakala, P., Rickkinen, P.& Sirvio, J. 1996. Behavioural and pharmacologicalstudies on the validation of a new animal model forattention deficit hyperactivity disorder. NeurobiologyLearning and Memory 66, 198–211.

P.2.016 Exposure to an alternative reward doesnot reduce cocaine-seeking behaviour

C. Nicolas1 °, C. Lafay-Chabassier2, M. Solinas1.1Laboratoire de Neurosciences Experimentales etCliniques-LNEC, INSERM U-1084 Universite dePoitiers, Poitiers Cedex, France; 2Laboratoire deNeurosciences Experimentales et Cliniques-LNECINSERM U-1084 Universite de Poitiers, Departmentof Clinical Pharmacology, Poitiers University Hospital,Poitiers Cedex, France

Exposure to positive environmental conditions, suchas environmental enrichment, decreases vulnerabilityto drugs and reduces the risks of relapse to drug-seeking behaviour [1]. Environmental enrichment iscomprised of several elements such as social interactions,novelty and sensory and physical activity, which havebeen described to be rewarding in rodents. Becauseaccess to alternative rewards decreases addiction-relatedbehaviours [2], it could be speculated that environmentalenrichment produces its positive effects on addiction byacting as an alternative reward [3]. On the other hand,whereas in most studies alternative rewards are presentedtogether with drugs, environmental enrichment is providedchronically as a living environment that is temporallyand physically distinct to drug-related conditions. Herewe investigated whereas chronic access to a naturalreinforcer such as sucrose, during period of withdrawalunder conditions similar to those used for environmentalenrichment, could reduce drug craving.For this, we let rats to self-administer cocaine during 10

experimental sessions (0.15mg/injection, 6 hours per day)whereas yoked-saline rats received only saline injections.At the end of the self-administration training period, ratswere subjected to a withdrawal period of 30 days duringwhich they were separated into three groups: a) the firstgroup had a constant access to one bottle of water and onebottle containing sucrose (10% w/v, sucrose continuousSC); b) the second group had access to water and sucroseintermittently (10% w/v, sucrose intermittent, SI) andc) the third group was a control group and had accessto two bottles of water (water, WAT). At the end of the


withdrawal period, rats were tested for cocaine seekingbehaviour during a single session that lasts 6 hours.We found that, during abstinence, SC and SI rats avidly

consumed sucrose: they drank 3 times more than WATrats and they completely ignored water bottles. Sucroseconsumption and preference were constant over the entireperiod of withdrawal and did not differ between cocaineand yoked-saline rats. After the withdrawal period of30 days, rats that were exposed to cocaine show highlevels of drug-seeking behaviour but no difference wasfound among SC, SI and WAT rats.Altogether, these results show that exposure to a natural

reward such as sucrose during withdrawal period does notaffect cocaine seeking behaviour. These results suggestthat exposure to an alternative reward that is temporallyand physically distinct from drug-related environments isnot sufficient to reduce the risks of relapse. In addition,they suggest that elements of enrichment other thanalternative reinforcement play a more important rolein reducing drug seeking or, at least, that alternativereinforcement must be combined with other elements ofenrichment to be effective in decreasing drug seeking.

Reference(s)

[1] Chauvet, C., Lardeux, V., Goldberg, SR., Jaber, M.,Solinas, M., 2009. Environmental enrichment reducescocaine seeking and reinstatement induced by cues andstress but not by cocaine. Neuropsychopharmacology34(13):2767−78.

[2] Ahmed, S.H., 2005. Imbalance between drug andnon-drug reward availability: A major risk factor foraddiction. Review, Eur J Pharmacol 5;526(1−3):9−20.

[3] Solinas, M., Thiriet, N., Chauvet, C., Jaber, M., 2010.Prevention and treatment of drug addiction by environ-mental enrichment. Prog Neurobiol 92(14):572−92.

P.2.017 The hallucinogen 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT)disrupts cortical function: reversal byantipsychotic drugs

M.S. Riga1 °, F. Artigas1, P. Celada1. 1IIBB-CSIC(IDIBAPS) CIBERSAM, Neuropharmacology andNeurochemistry, Barcelona, Spain

5-MeO-DMT, a natural hallucinogen component ofAyahuasca (Amazonian beverage), is a non-selectiveserotonin 5-HT1A/5-HT2A receptor agonist. Its potentialinterest in schizophrenia lies in its ability to mimicpsychotic symptoms such as hallucinations. We previouslyreported that other hallucinogens (the non-competitiveNMDA-R antagonist phencyclidine and the 5-HT2A/2C

agonist DOI) markedly disrupt cortical synchrony inthe low frequency range (<4Hz) in rodent prefrontalcortex (PFC), an effect reversed by antipsychoticdrugs [1−3].The aims of the present study are: (1) to examine

whether 5-MeO-DMT disrupts cortical synchrony in PFC,(2) to examine the ability of antipsychotic drugs to reverseits effects, and (3) to identify other brain areas sensitiveto the action of the hallucinogen. We used electrophys-iological techniques − single unit extracellular recordingof mPFC pyramidal neurons, local field potential (LFP)and epidural electrocorticogram (ECoG) recordings −and blood oxygen level-dependent (BOLD) functionalmagnetic resonance imaging (fMRI) in anaesthetisedanimals. Drugs were administrated intravenously (i.v.) inrat and subcutaneously (s.c.) in mice. Statistical analyseswere conducted using Student’s t-test, one- or two-wayANOVA followed by multiple comparison test. Statisticalsignificance was set at p< 0.05.5-MeO-DMT (0.1mg/kg i.v.), in combination with

clorgyline (MAO-A inhibitor, to prevent peripheraldegradation of 5-MeO-DMT) altered pyramidal discharge(Student’s t-test; p< 0.001; n = 42) and concurrentlyreduced low frequency cortical oscillations (LFCO;<4Hz) to 64±2% of basal values (one-way ANOVA;p< 0.000001; n = 58) in rat PFC. Likewise, 5-MeO-DMT (1mg/kg s.c.) differentially reduced LFCO inthe PFC of wild-type (WT) mice (to ~50% of basalvalues) and 5-HT2A knockout mice (KO2A) (transiently to~75% of basal values) (two-way ANOVA; p< 0.03). The5-MeO-DMT-induced reduction in LFCOwas significantlyreversed by M100907 (5-HT2A receptor antagonist) andWAY-100635 (5-HT1A receptor antagonist) in the rat.Likewise, the effect produced by 5-MeO-DMT in KO2Amice was fully prevented by WAY-100635. Overall, thesedata indicate that 5-MeO-DMT reduces cortical synchronyvia activation of 5-HT2A, and to a lesser extent, 5-HT1Areceptors. The antipsychotic drugs haloperidol, clozapineand risperidone and the mGluR2/3 agonist LY379268reversed 5-MeO-DMT effects on LFCO.Moreover, fMRI studies showed significant alterations

in BOLD response in several cortical areas (PFC, primaryvisual (V1), somatosensory (S1) and auditory (Au1) cor-tices). Further electrophysiological experiments indicatedthat 5-MeO-DMT reduced LFCO in V1, S1 and Au1 areasin rats and WT mice. Interestingly, 5-MeO-DMT alteredLFCO in V1 of KO2A mice.Together with previous findings [1−3], the present

results indicate that reductions in LFCO are a commonneurophysiological signature of hallucinogens. The rever-sal of these effects by antipsychotic drugs with differentmechanisms of action suggests a clear association withtheir therapeutic activity, regardless of their initial target.


This supports the usefulness of the LFCO model in PFCto examine the neurobiological basis of hallucinationsand in target identification during antipsychotic drugdevelopment.Moreover the present results point to the prefrontal

and sensorial cortical areas as sites of action of thishallucinogen and suggest the involvement of 5-HT1Areceptors in the action of indoleamine hallucinogens, inaddition to their well-known action on 5-HT2A receptors.

Reference(s)

[1] Kargieman L, Santana N, Mengod G, Celada P,Artigas F (2007). Antipsychotic drugs reverse thedisruption in prefrontal cortex function produced byNMDA receptor blockade with phencyclidine. ProcNatl Acad Sci U S A 104: 14843–14848.

[2] Celada P, Puig MV, Diaz-Mataix L, Artigas F(2008). The hallucinogen DOI reduces low-frequencyoscillations in rat prefrontal cortex: reversal byantipsychotic drugs. Biol Psychiatry 64: 392–400.

[3] Kargieman L, Riga MS, Artigas F, Celada P(2012). Clozapine reverses phencyclidine-induceddesynchronization of prefrontal cortex through a5-HT(1A) receptor-dependent mechanism. Neuropsy-chopharmacology 37: 723–733.

P.2.018 Characterisation of the effects ofpartial agonist of a4b2* nACh receptorcytisine in the two-choice serialreaction time task

G. Makshakov1 °, O. Dravolina1, A. Bespalov1,E. Kayukova1, M. Dorofeikova1, E. Zvartau1.1Pavlov State Medical University, Valdman Instituteof Pharmacology, St. Petersburg, Russia

It has been suggested that nicotine has attention-enhancingproperties and attenuates response inhibition. Nicotine wasfound to improve target detection and to decrease reactiontime, but also to impair the ability to withhold prematureresponses (i.e. to increase impulsivity). These effects ofnicotine are thought to be mediated by a4b2*, a4b4*and a3b2* nicotinic acetylcholine receptor (nAChR)subtypes [1]. The purpose of this study was to investigateeffects of cytisine, a partial agonist of a4b2* nAChR, onthe performance in the two-choice serial reaction time task(2-CSRTT).The study was conducted in male Wistar rats. At the

beginning of the training, all animals (n = 10) were placedon food restriction maintaining 85% of their free-feedingweight. Experiment sessions lasted for 30min or untilcompletion of 100 trials, whichever occurred first. Rats

were trained to detect a brief visual stimulus (three lightbulbs) presented randomly in one of two locations and topress the lever beneath highlighted bulbs during the stimu-lus presentation (1 s) or the limited hold period (2 s). Sucha response (‘correct’) was followed by a food pellet presen-tation. ‘Incorrect’ responses or failure to respond withinthe limited hold period (‘omissions’) were recorded andpunished by the extinction of the house light for 5 s (‘time-out’ period). Just after a ‘correct’ response and collectionof the reward or an ‘incorrect’ response/an omission andtime-out, the hole on the opposite wall was illuminated anda nose-poke was required to initiate the next trial. Leverresponses before trial initiation (‘pre-initiation’ responses)were recorded but had no programmed consequence. Aftertrial initiation, animals had to avoid lever pressing for5 s before visual stimulus presentation, otherwise theywere punished by time-out and premature responses wererecorded. Criterion of 70% correct responding and lessthan 10% omissions during two consecutive sessionsrequired to proceed the drug treatment phase. Variousdoses of cytisine (0, 0.3, 1.0 and 3.0mg/kg, s.c.) andnicotine (0, 0.075, 0.15, 0.3 and 0.6mg/kg, s.c.) wereadministered in a pseudo-random order.Both compounds had no reliable effects on percent

of ‘correct’ responses and number of premature,perseverative, and time-out responses, although nicotinetended to increase premature responses. Latencies to makea ‘correct’ response or to collect a food reward were alsounchanged from baseline. However, administration of thehighest dose of cytisine (3.0mg/kg) attenuated the numberof completed trials due to significantly increased numberof ‘pre-initiation’ responses. Further analysis found thatonly number of ‘pre-initiation’ responses (on both levers)performed after a ‘correct’ response became persistentlyelevated following cytisine administration.In summary, the results of the present study demonstrate

that cytisine induced performance disruption indicative ofincreased ‘pre-initiation’, but not premature responding.Such deficits might reflect an aspect of responsedisinhibition, other than loss of impulse control, thatrepresents an additional measure of compulsivity relatedto cognitive inflexibility.

Reference(s)

[1] Blondel A., Sanger D., Moser P., 2000 Characterisationof the effects of nicotine in five-choice serialreaction time task in rats: antagonists studies.Psychopharmacology, 149: 293–305.


P.2.019 The mismatch hypothesis: a new wayof linking early experiences and adultenvironment to vulnerability to stress

S. Santarelli1 °, K.V. Wagner1, J. Hartmann1, X.D. Wang1,M.V. Schmidt1. 1Max Planck Institute of Psychiatry,Neurobiology of Stress, Munich, Germany

Purpose of the study: Chronic stress is considered one ofthe main risk factors for depression. Interestingly, not allindividuals develop psychopathology after chronic stress.In contrast to the prevailing view that stress effects arecumulative and increase stress vulnerability later in life,the recently formulated mismatch hypothesis proposesthat individuals experiencing high levels of psychosocialstress early in life are programmed for dealing with highpsychosocial stress and are therefore resilient to highstress levels in later life [1]. We here test this hypothesisby comparing the developmental effects of two differentearly life conditions, when tested under two opposite adultenvironments.

Methods used: We used female BALB/c mice thatunderwent either adverse early life conditions (limitednesting material) or a supportive environment (earlyhandling) from PND2 to PND9. At weaning, theanimals of each group were assigned to either grouphousing (supportive environment) or single housing (socialdeprivation, adverse environment). At adulthood, wecompared the molecular and the behavioural effects ofthe interaction between early and adult environment, inparticular on anxiety like and depressive like behaviour.

Table 1. Experimental groups

Group Housed (GH) Single Housed (SH)

Early Handling (EH) EH GH (matched) EH SH (mismatched)Limited Nesting Material (LM) LM GH (mismatched) LM SH (matched)

Summary of results: Animals that underwent twoadverse environments (mismatched) and those thatunderwent two supportive experiences (matched) havea similar behavioural profile. They are less vulnerableto stress when compared to the ones that underwentonly one stressful experience, either during early lifeor during adulthood. Parameters like weight of organs,spontaneous behaviour in an open field or floating inthe forced swim stress were significantly reduced inanimals with mismatched environments compared toanimals with matched environments. Also corticosteronelevels, a measure of hypothalamic–pituitary–adrenal axisresponsiveness, have been significantly affected by theinteraction of the early and adult environment. Incontrast, anxiety like behaviour in the elevated plus

maze was mainly affected by early life experience,whereas sociability behaviour was mainly affected by theadult environment. We also looked for new molecularmarkers for stress vulnerability, and we found that theexpression levels of SLC6A15, a novel candidate fordepression vulnerability, are increased in the hippocampusof mismatched animals.

Conclusions:We conclude that animals placed at adult-hood in a mismatched environmental condition comparedto their early life environment, are more vulnerableto stress in specific behavioural and neuroendocrineparameters. Still further analyses are needed to betterunderstand the effects of this paradigm on other behavioursand to clarify the molecular mechanisms underlying thosebehavioural modifications. Based on these findings, wehypothesize that adult vulnerability to stress could bemodulated by a mismatch between early-life and adultenvironment, in combination with a more resilient orvulnerable genetic make-up.

Reference(s)

[1] Nederhof, E., Schmidt, M.V., 2012. Mismatch orcumulative stress: Toward an integrated hypothesis ofprogramming effects. Physiology & Behavior, 106 (5),691–700.

P.2.020 Disruption of 5-HT7 receptorsaccelerates age-related episodic-likememory decline

G. Beaudet1 °, M. Brehin1, T. Freret1, G. Nee1,V. Delaunay1, M. Boulouard1, E. Paizanis1. 1Universitede Caen Basse-Normandie, GMPc EA4259, Caen, France

Increasing incidence of cognitive impairments includinglearning and memory deficits, with early alterations ofepisodic memory, related to population ageing, is a majorpublic health challenge. Thus it is crucial to identify newtherapeutic targets, not only focussing on symptomaticstrategies, but also aiming at the prevention and healthrecovering. In this way, modulation of serotonergic 5-HT7receptors (5-HT7R) could be an innovative approachsince (1) gene expression of 5-HT7R early decreasesduring senescence in rats (between 3 and 12 months-old),(2) synaptic plasticity of the hippocampus is altered in5-HT7R knocked-out KO) mice, and (3) as we recentlyhave shown in preliminary experiments, pharmacologicalmodulation of 5-HT7 improves episodic-like memoryperformances in mice. However, considering that episodic-like memory assessed in an object recognition task [1]seems not to be altered in 5-HT7 KO mice at 3 months ofage, we hypothesized that a deficit could appear later.


Therefore, in order to identify the implication of5-HT7R on the onset of the age-related memorydecline, memory performances of 7−8 months-old and10−12 months-old KO C57BL/6J mice with homozygousdisruption of the 5-HT7R gene were compared to pairedwild type mice (WT). Recognition memory performanceswere assessed in a Y-maze with a 1-hour delay betweenpresentation and test [2]. Spatial working memoryperformances were assessed by recording spontaneousalternation behaviour in a Y-maze during a 5-minutessession. In addition locomotor activity was assessed in anactivity cage and anxiety-related behaviour was examinedusing the elevated plus-maze (EPM).Our data showed that 10−12 months-old WT mice did

not discriminate the novel object contrary to 7−8 months-old WT mice, which spent more time exploring the novelobject compared to the chance level (univariate t-test:p< 0.05). In addition, disruption of 5-HT7R led to animpaired object discrimination in 7−8 and 10−12 months-old mice. By contrast, ANOVA analysis revealed thatmutant mice exhibited equivalent levels of spontaneousalternation compared to WT mice (around 75%). Inaddition, statistical analysis of locomotor activity did notrevealed any significant effect of genotype or genotype ×age interaction, indicating equivalent basal level oflocomotion. Finally, no differences between mice of bothgenotypes were detected in the global activity or theanxiety-related parameters assessed in the EPM.Our results therefore suggest that in our experimental

conditions, the onset of episodic-like memory declineis situated between 7−8 and 10−12 months-old of agein C57BL/6J mice, and that the lack of 5-HT7R couldaccelerate the onset of this decline, with no relation to anyinhibition of locomotor activity or increased anxiety-likebehaviour. By contrast, working memory performancesseem not to be affected between 7−8 and 10−12 months-old in KO and WT mice. Further pharmacological (usinga selective 5-HT7 antagonist) and molecular (5-HT7Rexpression) studies in WT mice would confirm thisstatement.

Reference(s)

[1] Sarkisyan, G., Hedlund, P.B., 2009. The 5-HT7receptor is involved in allocentric spatial memoryinformation processing. Behavioural Brain Research2002, 26−31.

[2] Leger, M., Quiedeville, A., Paizanis, E., Natkunara-jah, S., Freret, T., Boulouard, M., Schumann-Bard, P.,2012. Environmental enrichment enhances episodic-like memory in association with a modified neuronalactivation profile in adult mice. PLoS One 7, 10.

P.2.021 Role of adenosine (A)2A receptor innicotine addiction − pharmacologicaland genetic aspects

J. Czyzyk1 °, E. Nowak1, M. Bader2, K. Fuxe3,M. Filip1. 1Polish Academy of Sciences, Departmentof Pharmacology, Krakow, Poland; 2Max-Delbruck-Centrum, Molecular Medicine, Berlin, Germany;3Karolinska Institutet, Department of Neuroscience,Stockholm, Sweden

Nicotine is one of the very strong addictive psychoactivesubstance which is commonly available and acceptedby society. The abuse of nicotine is linked to theenhancement of dopaminergic (DA) neurotransmissionand indirect activation of DA D2 receptors in thebrain mesolimbic system. A number of data indicateantagonistic interaction at the molecular, neurochemicaland behavioural levels between D2 and adenosine (A)2Areceptors that occurs in the striatum [1]. This interactionmay have a significance to control behavioural effectsinduced by nicotine [2].We studied the effects of the A2A receptor agonist CGS

21680 and antagonist KW 6002 on the development andexpression of nicotine sensitisation in wild-type (WT) rats.Moreover, we addressed the role of A2A receptors in thelocomotor effects to repeated treatment with nicotine intransgenic (TG) rats overexpressing A2A receptors.Sensitisation was evoked by 5 daily injections of

nicotine (0.4mg/kg, sc) and a challenge dose of the drug(0.4mg/kg, sc) on day 10. During the first 5 days ofdevelopment phase WT animals received the followinginjections: CGS 21680 (0.2−0.4mg/kg, ip) or KW 6002(0.25−0.5mg/kg) in combination with nicotine, whileTGR were treated with vehicle or nicotine (0.4mg/kg,sc), then on 6−9 days the animals remained drug-freein home cages. On day 10, all rats received nicotinechallenge dose. In studies into expression of sensitisation,the WT animals received repeatedly (5 days) vehicleor nicotine (0.4mg/kg, sc), while on day 10 they werepretreated by CGS 21680 (0.05−0.2mg/kg, ip) or KW6002 (0.25−0.5mg/kg, ip) before a challenge dose ofnicotine. Conditioned locomotion was evoked by 5-dailyinjections of nicotine (0.4mg/kg, sc) in experimentalchambers followed by a 4-day drug-free period and avehicle challenge in experimental chambers on day 10.Locomotor activity measurements started immediatelyafter the injection of nicotine and were recorded for60min.During development of sensitisation CGS 21680 in dose

of 0.4mg/kg (but not 0.2mg/kg) given repeatedly withnicotine significantly (P< 0.05) decreased the locomotor


activity to the nicotine challenge dose, while KW 6002was inactive in this procedure. Expression of nicotinesensitisation was reduced by pretreatment with CGS 21680(by 60% in a dose of 0.2mg/kg, and by 55% in a doseof 0.1mg/kg), whereas KW 6002 did not change thelocomotor activity induce by nicotine. TG animals, like theWT rats, showed 2-fold enhancement in locomotor activityto repeated nicotine treatment (days 1−5, and day 10).There was no evidence of conditioned locomotion in TGrats; only WT rats treated repeatedly (1−5 days) withnicotine and challenged with vehicle on day 10 showedenvironment-triggered locomotion.The present results show that stimulation of adenosine

A2A receptors reduces the development and expressionof nicotine sensitisation as well as drug-associatedenvironmental cues in WT animals. The latter effect wasconfirmed in TG rats overexpressing A2A receptors. Thesefindings extent our recent data on cocaine [3] and mayrise the possibility of using A2A receptor agonists in drugaddiction therapy.

Reference(s)

[1] Filip, M., Zaniewska, M., Frankowska, M., Wydra, K.,Fuxe, K., 2012. The Importance of the Adenosine A2AReceptor-Dopamine D2 Receptor Interaction in DrugAddiction. Curr Med Chem. 19(3), 317−55.

[2] Castane, A., Soria, G., Ledent, C., Maldonado, R.,Valverde, O., 2006. Attenuation of nicotine-inducedrewarding effects in A2A knockout mice. Neurophar-macology. 51, 631−40.

[3] Filip, M., Frankowska, M., Zaniewska, M., Prze-galinski, E., Muller, C.E., Agnati, L., Franco, R.,Roberts, D.C., Fuxe, K., 2006. Involvement ofadenosine A2A and dopamine receptors in thelocomotor and sensitizing effects of cocaine. BrainRes. 1077, 67−80.

P.2.022 Oxytocin in the central nucleus of theamygdala mediates social buffering offear

E. Rickenbacher1 °, M. Moita1. 1ChampalimaudFoundation, Champalimaud Neuroscience Programme:Behavioral Neuroscience, Lisbon, Portugal

Fear responses are innate defensive behaviours exhibitedby animals in response to aversive stimuli. During fearconditioning an animal can learn to fear a neutralstimulus (e.g. tone) when it is paired with an aversiveone (e.g. footshock), such that this previously neutralstimulus comes to elicit fear responses. This form oflearning may be crucial as it allows animals to use cues

associated with aversive events to avoid future threats.Interestingly, there is evidence that when animals are inthe presence of a conspecific they show decreased fearresponses to a stimulus previously paired with an aversiveevent; demonstrating that social interactions can bufferfear responses [1].As we are interested in the neural mechanisms of

social buffering, we have developed a paradigm to studysocial buffering in fear conditioned rats. In our experimentrats were conditioned to fear a tone by pairing it withfootshocks. After conditioning, rats were tested for theirfear of the tone through the presentation of the tone in theabsence of shock. Rats quickly learned to fear the tone,displaying a robust fear response, freezing. However, wefound that rats re-exposed to the tone in the presence oftheir naıve cage-mate (n = 8) showed less freezing thanif tested alone (n = 11) (average freezing response acrosstone presentations (p< 0.01)). Moreover, it is known thatexposure to the tone in the absence of footshocks leads toa subsequent decrease in tone-evoked fear responses. Thisdecrement in freezing results either from animals learningthat the stimulus is now safe, while still rememberingthat it was previously predictive of shock, or because theyupdate their memory of the association between tone andshock, losing the initial memory.Thus, we asked whether being re-exposed to the tone in

the presence of the cage-mate affected how rats respondedto the tone in future exposures. When tested subsequently,now alone, rats that were previously re-exposed in thepresence of their cage-mate (n = 17) continued to freezeat a decreased level compared to animals that had beenre-exposed alone (n = 16), showing that social bufferinghas both short term (p< 0.001) and long term effects onfear expression (p< 0.01).Although oxytocin has been implicated in this

process, its underlying neural mechanisms remain poorlyunderstood. We have therefore assessed the role ofoxytocin in the central nucleus of the amygdala (CeA),a major output station that controls several defenceresponses [2,3]. Animals received bilateral CeA cannulaimplants one week prior to fear conditioning and bilateraloxytocin antagonist or vehicle infusions immediately priorto the first tone re-exposure session. Our data show thatthe infusion of an oxytocin antagonist compared to vehicleinto the CeA, prior to re-exposure to the tone, blocks theimmediate and long lasting effect of social buffering onthe elicited fear memory and response (p< 0.01).Our study shows long lasting effects of social

buffering on fear and elucidates the neural mechanismsunderlying the phenomenon. These data contributeimportant knowledge pertaining to underlying causes ofanxiety disorders and possible treatment applications.


Reference(s)

[1] Kiyokawa, Y., Takeuchi, Y., Mori, Y., 2007. Two typesof social buffering differentially mitigate conditionedfear responses. Eur J Neurosci. 26(12): p. 3606−13.

[2] Wilensky, A.E., Schafe G.E., Kristensen M.P., LeDouxJ.E., 2006. Rethinking the fear circuit: the centralnucleus of the amygdala is required for the acquisition,consolidation, and expression of Pavlovian fearconditioning. J Neurosci. 26(48): p. 12387−96.

[3] Ciocchi, S., Herry C., Grenier F., Wolff S.B., LetzkusJ.J., Vlachos I., Ehrlich I., Sprengel R., Deisseroth K.,Stadler M.B., Muller C., Luthi A., 2010, Encodingof conditioned fear in central amygdala inhibitorycircuits. Nature. 468(7321): p. 277−82.

P.2.023 The antagonist of 5-HT7 receptors,SB-269970, and amisulpride bothreverse ketamine-induced cognitiveinflexibility in rats

D. Rafa1 °, A. Nikiforuk1, P. Popik1. 1Polish Academy ofSciences, Department of Behavioural Neuroscience andDrug Development, Krakow, Poland

The prefrontal cortex mediates higher-order executivefunctions, including among other, the cognitive flexibility,i.e., the ability to modify behaviour in response tochanging task demands. This aspect of executive functionis impaired in schizophrenia. Cognitive flexibility mayalso be assessed in rodents in the attentional set-shiftingtask (ASST) [1]. In this paradigm, rats must select abowl containing a food reward based on the ability todiscriminate the odours and the media covering the bait.The ASST requires rats to initially learn a rule and forman attentional ‘set’ within the same stimulus dimensions.At the extradimensional (ED) shift, animals must switchtheir attention to a new, previously irrelevant stimulusdimension and, for example, discriminate between theodours and no longer between the media covering the bait.The animals’ performance at the ED phase is impaired inN-methyl-D-aspartate receptor antagonist (e.g., ketamine)-treated animals [2], regarded as pharmacological model ofschizophrenia-like symptoms.The 5-HT7 (5-hydroxytryptamine 7, serotonin 7)

receptor, one of the most recently identified members ofthe serotonin receptor family, may play an important rolein the pathophysiology and treatment of many psychiatricdisorders. Recent data suggest that the blockade of5-HT7 receptors may exert procognitive effects in animalmodels of schizophrenia [3]. This issue might be ofspecial interest, since several atypical antipsychotics, e.g.,amisulpride, are characterised by a high affinity for 5-HT7

receptors. Nevertheless, little is known about the efficacyof 5-HT7 antagonists the in models of schizophrenia-likecognitive inflexibility.The aim of the present study was to investigate the

role of a potent and selective 5-HT7 receptor antagonist(SB-269970), and amisulpride (an atypical antipsychoticwith high affinity to 5-HT7 receptors) on ketamine-induceddeficits in the ASST task in rats.Ketamine (10mg/kg) was administered to Sprague-

Dawley rats subcutaneously 75min prior to the test. SB-269970 or amisulpride were given intraperitoneally 15minbefore ketamine injection. The number of trials requiredto achieve the criterion of 6 consecutive correct responseswas recorded for each rat and for each discriminationphase. Data were calculated using two-way mixed-designANOVAs followed by the Newman–Keuls post-hoc test.Ketamine administration significantly and specifically

impaired rats’ performance at the ED stage of ASSTas indicated by an increased number of trials tocriterion during this phase. SB-269970 (0.3 and 1mg/kg)and amisulpride (3mg/kg) reversed ketamine-inducedcognitive inflexibility. Additionally, SB-269970 at a doseof 1mg/kg also improved ED performance as compared tocontrol rats. There were no significant drug effects duringany other discrimination stage. Analyses of variancerevealed significant interactions of discrimination phaseand treatment: F[18,204] = 26.93, P< 0.001 (SB-269970)and F[12,156] = 16.84, P< 0.001 (amisulpride).Present study demonstrated the efficacy of the 5-HT7

antagonist, SB-269970, and amisulpride in amelioratingfrontal-like deficits relevant to the psychopathology ofschizophrenia. It thus seems likely that the antagonism of5-HT7 receptors may represent a useful pharmacologicalapproach for cognitive enhancement in schizophrenia.

Reference(s)

[1] Birrell, J.M., Brown, V.J., 2000. Medial frontal cortexmediates perceptual attentional set shifting in the rat.J Neurosci 20, 4320–4324.

[2] Nikiforuk, A., Golembiowska, K. and Popik, P.,2010. Mazindol attenuates ketamine-induced cognitivedeficit in the attentional set shifting task in rats. EurNeuropsychopharmacol 20, 37−48.

[3] Horiguchi, M., Huang, M., Meltzer, H.Y., 2011.The role of 5-hydroxytryptamine 7 receptors in thephencyclidine-induced novel object recognition deficitin rats. J Pharmacol Exp Ther 338, 605−14.


P.2.024 The effects of positive allostericmodulators of a7 nicotinic receptors onrats’ performance in the odour span test

A. Potasiewicz1 °, A. Nikiforuk1, P. Popik1. 1PolishAcademy of Sciences, Department of BehavioralNeuroscience and Drug Development, Krakow, Poland

It is widely accepted that cognitive deficits are a corefeature of schizophrenia. Schizophrenic patients developa wide range of neuropsychological dysfunctions includingdeficits in working memory (WM). Specifically, they sufferfrom shortages in WM span capacity, displaying reducednumber of information kept concurrently in the WM.Nicotinic acetylcholine receptors (nAChRs) play an

important role in the regulation of cognitive processes.Particularly, selective activation of a7-nAChRs representsa promising target for the development of pharmacologicaltreatments for cognitive dysfunctions in schizophrenia.It appears however, that positive allosteric modulators(PAMs) of a7-nAChRs may demonstrate a more favorableprofile than orthosteric agonists [1]. Current data hasdemonstrated a positive influence of nicotine on WMspan capacity assessed using the odour span task (OST)in rats [2,3]. However, while the role a7-nAChRs in thenicotine-induced enhancement in span capacity has beensuggested, the effects of a7-nAChRs PAMs have not beenassessed on that task.The aim of the present study was to investigate the

effects of a7-nAChRs PAMs: CCMI (type I) and PNU-120596 (type II) in comparison to nicotine on the WMcapacity in the OST in rats. Type I PAMs have little orno effect on desensitisation processes, while the actionof type II PAMs is accompanied by a retardation of thekinetics of desensitisation.Male Sprague-Dawley rats were trained to remember an

increasing number of odours according to a nonmatchingto sample rule. Rats had to dig for food rewards inscented cups containing woodchips. Once they retrievedthe reward, a new differently scented cup was added. Ratshad to ignore previous cups and choose a new one inorder to gain a reward. After each correct response a newcup was introduced. The number of consecutive correctresponses served as a measure of the span length. Alsothe number of errors was counted during the session.The effect of nicotine (0.05 and 0.1mg/kg), PNU 120596(1 and 3mg/kg), CCMI (0.3 and 1mg/kg) and vehicle(saline) were evaluated after asymptotic performanceswere obtained. Drugs doses were administered in a Latinsquare design order. Data were calculated using maineffect-design ANOVAs followed by the Newman–Keulspost-hoc test.

Nicotine (0.05 and 0.1mg/kg), administered subcu-taneously (SC) 10 minutes before the test, caused astatistically significant improvement of the span length(p< 0.05). It also significantly decreased a number oferrors (p< 0.05 and p< 0.001 for a dose of 0.05 and0.1mg/kg, respectively). Neither CCMI, nor PNU120596administered SC 30 minutes before the test significantlyaffected the span length. However, a number of errorswas significantly decreased by CCMI at both doses tested(p< 0.05).Although the a7-nAChRs PAMs represent a promising

alternative to orthosteric ligands, their behavioural activityis still poorly understood especially with regard to thedifferences between type I and type II compounds. Hence,despite the limited efficacy of CCMI and PNU120596 onthe span capacity in cognitively unimpaired animals, itseems interesting to further investigate a7-nAChRs PAMsin the models of schizophrenia-like OST deficits.

Reference(s)

[1] Hurst, R.S., Hajos, M., Raggenbass, M., Wall, T.M.,Higdon, N.R., Lawson, J.A. et al., 2005. A novelpositive allosteric modulator of the alpha7 neuronalnicotinic acetylcholine receptor: in vitro and in vivocharacterization. J Neurosci. 25:4396–4405.

[2] Rushforth, S.L., Allison, C., Wonnacott, S.,Shoaib, M., 2010. Subtype-selective nicotinic agonistsenhance olfactory working memory in normal rats:a novel use of the odour span task. Neurosci Lett471:114–118.

[3] Rushforth, S.L., Steckler, T., Shoaib, M., 2011.Nicotine improves working memory span capacityin rats following sub-chronic ketamine exposure.Neuropsychopharmacology 36:2774–2781.

P.2.025 Ventral medial prefrontal cortexinactivation reduces context-inducedreinstatement of nicotine seeking

R.F. Struik1 °, J. Peters1, S. Jonkman-Tielemans1,T.J. De Vries2. 1Free University Medical Centre(VUmc), Anatomy and Neuroscience, Amsterdam, TheNetherlands; 2Free University (VU), Molecular &Cellular Neurobiology Neuroscience Campus AmsterdamCNCR, Amsterdam, The Netherlands

Environments and circumstances strongly contribute torelapse to smoking in humans. In rodents as well,when returned to a drug-paired context, rats willreinstate drug seeking (or relapse), termed context-inducedreinstatement [1]. Preclinical data suggests prefrontalcortex involvement in inhibitory control over drug seeking


behaviour [2]. Inactivation of neurons in the ventralmPFC reduced context-induced reinstatement of heroinseeking [3]. Data on the role of the ventral mPFC innicotine seeking are lacking. Here, in a pharmacologicalinactivation study, we determined the involvement of theventral mPFC in context-induced reinstatement of nicotineseeking.Thirty-two male Sprague Dawley rats were implanted

with intravenous catheters and guide cannulas in theventral mPFC. Animals were trained to self-administernicotine and saline in separate contexts during one-hour sessions per day over 19 days. Contexts consistedof an operant chamber containing a nose-poke hole orlever, a discrete light/tone cue as well as tactile, odour,circadian and sound cues. Responding was extinguishedin a third context. One hour prior to being returned tothe nicotine context, the saline context and the extinctioncontext, animals were infused in the ventral mPFC with0.3 microlitre of either ACSF or a mixture of the GABA-A/GABA-B agonist baclofen (1mM)/ muscimol (0.1mM).Two extinction sessions separated each test session.Responding at the end of self-administration is

higher for nicotine (48.1±3.90, mean±SEM) than saline(8.33±1.26), indicating rats selectively respond fornicotine (one-way ANOVA, p< 0.001). The vehiclegroup reinstated when returned to the nicotine context(17.9±2.8) compared to the extinction context (5.6±2.26;one-way ANOVA, p< 0.01). The baclofen/muscimolgroup exhibited blunted relapse when returned to thenicotine context (8.0±2.4) compared to the extinctioncontext (4.0±1.3). Lower responding in the baclofen/muscimol group in the nicotine context compared to thevehicle group (one-way ANOVA, p< 0.05), indicates thatinactivation of the ventral mPFC reduces context-inducedreinstatement of nicotine seeking. In the saline context,no difference was seen between responding in vehicle(13.2±6.4) versus baclofen/muscimol (6.1±3.6) groups.This seems to indicate ventral mPFC inactivation had noeffect on responding in the saline context.These results suggest that, similar to studies in context-

induced reinstatement of heroin seeking, activity in theventral mPFC is critical for context-induced reinstatementof nicotine seeking. However, ventral mPFC activity is notessential for responding in a saline context and rats are ableto distinguish a nicotine-associated context from a saline-associated context. Since responding in an extinctioncontext is unaffected by ventral mPFC inactivation, thisarea is likely not involved in context-associated inhibitorycontrol over nicotine seeking. Future studies are designedto determine the ventral mPFC neuronal subtypes andprojections involved in context-induced reinstatement ofnicotine seeking.

Reference(s)

[1] Crombag, H.S., Bossert, J.M., Koya, E., Shaham, Y.,2008, Review. Context-induced relapse to drugseeking: a review. Philos Trans R Soc Lond B BiolSci. 363(1507), 3233−43.

[2] Kalivas, P.W., The glutamate homeostasis hypothesisof addiction. Nat Rev Neurosci. 10(8):561−72.

[3] Bossert, J.M., Stern, A.L., Theberge, F.R., Cifani, C.,Koya, E., Hope, B.T., Shaham, Y., 2011. Ventralmedial prefrontal cortex neuronal ensembles mediatecontext-induced relapse to heroin. Nat Neurosci.14(4):420−2.

P.2.026 Pro-inflammatory cytokines induceanhedonia in mice and increasemonoamine transporter activity in thenucleus accumbens

F. van Heesch1 °, J. Prins1, K.G.C. Westphal1,G.A.H. Korte-Bouws1, B. Olivier1, A.D. Kraneveld1,S.M. Korte1. 1Utrecht University, Pharmacology,Utrecht, The Netherlands

A growing body of evidence suggests that pro-inflammatory cytokines contribute to the pathogenesisof depression due to a general medical condition [1].Lipopolysaccharide (LPS), a component of the outermembrane of gram-negative bacteria, binds to toll-likereceptor 4 (TLR4) leading to the rapid release ofpro-inflammatory cytokines. In rats these LPS-inducedpro-inflammatory cytokines induce anhedonia, i.e. theinability to experience pleasure, a core symptom ofmajor depressive disorder. This was shown by increasedthresholds in an intracranial self-stimulation (ICSS)paradigm [2]. The underlying mechanisms are largelyunknown. We hypothesize that peripheral cytokines reachthe brain and affect central monoamine signalling viathe enhancement of monoamine reuptake which leads toanhedonia.To demonstrate that peripheral cytokines induce

anhedonia in mice, C57BL/6 mice were trained in the ICSSparadigm. After establishment of stable ICSS thresholds,mice were treated with saline or LPS (133mg/kg, i.p.).Animals were tested in the ICSS paradigm 1 h, 4 h, 24 h,48 h, 72 h and 96 h after administration. In another group ofmice we measured extracellular monoamine (metabolite)concentrations in the nucleus accumbens (NAc). In the firstmicrodialysis study, mice were treated with saline followedby a saline or LPS injection (133mg/kg, i.p.) 30minlater. To test the involvement of monoamine transporters,in the second study, animals were injected with thetriple reuptake inhibitor DOV 216,303 (5mg/kg, i.p.) 30


minutes before the saline or LPS injection (133mg/kg,i.p.). During the microdialysis studies 30-minute sampleswere collected during a period of 6.5 h.

Results: In the ICSS study, repeated measuresANOVA with time as within factor and treatment asbetween-subjects factor revealed a significant time ×treatment interaction and a significant effect of treatment(F(1.7,24.0) = 6.263, p = 0.009 and F(1,14) = 9.2, p = 0.009respectively). One-way ANOVA per time point revealedsignificant elevations of ICSS thresholds in the LPSgroup at time points 1 h and 4 h after injection.Peripheral LPS injection did not affect serotoninand dopamine levels in the NAc. However, repeatedmeasures ANOVA revealed significant time × treatmentinteractions and effects of treatment for the metabolites(5-HIAA: F(3.6,35.9) = 16.8, p< 0.001 and F(1,10) = 12.6,p< 0.01 respectively; DOPAC: F(2.9,28.9) = 9.4, p< 0.001and F(1,10) = 10.2, p< 0.01 respectively and HVA:(F(3.0,30.0) = 34.5, p< 0.001 and F(1,10) = 16.0, p< 0.01respectively). Pre-exposure to DOV 216,303 reduced oreven prevented the LPS-induced increase in monoaminemetabolite levels (time × treatment interaction and effectof treatment: 5-HIAA: F(2.7,32.0) = 15.0, P< 0.001 andF(1,12) = 8.0, p< 0.05 respectively; DOPAC: p> 0.05 andp> 0.05 respectively and HVA: F(1.9,22.0) = 4.1 andp> 0.05 respectively).Altogether, these results indicate that pro-inflammatory

cytokines play a role in the communication betweenthe immune system and central nervous system. Theincreased elevations in ICSS thresholds after exposureto LPS suggest that pro-inflammatory cytokines induceanhedonia. The microdialysis data suggest that monoaminetransporters in the nucleus accumbens are involved in thisprocess, since pre-exposure to the triple reuptake inhibitorDOV 216,303 reduced or even prevented the increasedreuptake of monoamines by LPS, as reflected by thedecreased extracellular levels of monoamine metabolites.

Reference(s)

[1] Dantzer, R., O’Connor, J.C., Freund, G.G., John-son, R.W., Kelley, K.W., 2008. From inflammationto sickness and depression: when the immune systemsubjugates the brain. Nat Rev Neurosci 1, 46−56.

[2] Borowski, T., Kokkindis, L., Merali, Z., Anisman, H.,1998. Lipopolysaccharide, central in vivo biogenicamine variations, and anhedonia. Neuroreports 17,3797–802.

P.2.027 a7 nicotinic acetylcholine receptor(a7 nAChR) agonists or antagonists aspotential cognition enhancers?

N.P. Van Goethem1 °, J. Prickaerts1. 1MaastrichtUniversity, School for Mental Health and Neuroscience,Maastricht, The Netherlands

a7 nicotinic acetylcholine receptors (a7 nAChRs) areligand-gated ion channels expressed primarily in thebrain. These receptors have been implicated in modulatingmany cognitive functions like attention, episodic memoryand learning. a7 nAChRs are located both pre- andpostsynaptically and modulate glutamate, GABA anddopamine. Furthermore, a7 nAChRs are directly involvedin hippocampal long-term potentiation (LTP), the putativecellular mechanism underlying learning and memory.Activation of a7 nAChRs through selective, partial, orfull agonists and/or modulators, has been shown toimprove cognitive function in both animal and humanstudies. The main cognitive improvement with thesecompounds relate to memory, in accordance with thehigh level of expression of a7 nAChRs in the frontal-cortex and hippocampus. Hence, a7 nAChR agonists/modulators may be attractive drug candidates to improvecognition in Alzheimer’s disease (AD) and schizophreniapatients [1]. The objective of the current study wasto investigate the cognition enhancing properties oflow dose administration of an a7 nAChR antagonistin rats. Memory performance was assessed with theobject recognition task (ORT). The compound used forthese studies was methyllycaconitine (MLA). MLA isa norditerpenoid alkaloid produced by many species ofDelphinium (larkspurs) and is often used because of itsability to potently antagonise the a7 nAChRs. As a potentselective antagonist of the a7 nAChR, MLA is oftenused in animal models to show that putative a7 nAChRagonists indeed work through the a7 nAChR mechanismby blocking the procognitive effects of a7 nAChRagonists by means of MLA administration [1]. In contrast,low doses of MLA have sporadically been reported toimprove cognition in animals. Furthermore, MLA hasbeen shown to facilitate LTP induction in hippocampalregion CA1 in rats. In the current study, it was foundthat MLA significantly improved memory of rats in theORT paradigm (24 h interval ORT, repeated measuresANOVA; MLA (F7,119 = 4.37; P< 0.001) optimal doserange 0.003−0.1mg/kg i.p.). Moreover, it was found thata dose that was too high (1.0mg/kg, i.p.) to improvememory in the natural forgetting paradigm (24 h retentioninterval), was also sufficient to induce a memory deficitin the 1 h retention interval ORT, an interval that normally


leads to good memory performance of rats (1 h intervalORT, repeated measures ANOVA; MLA (F3,51 = 7.33;P< 0.001). This indicates that MLA could act both asa cognition enhancer and as a deficit model in thisparadigm. Among other possibilities, one explanationfor these findings could be that a7 nAChR antagonistspromote a7 nAChR resensitisation. Since a7 nAChRsdesensitise fast, by occupying a subset of a7 nAChRswith selective antagonists without intrinsic value, thesereceptors could have an opportunity to recover or re-sensitise. This hypothesis is currently investigated. Insummary, while the main focus of the a7nAChR as a targetfor cognition enhancement lies on agonists and positivemodulators, the antagonists of these receptors might proveto be a valuable tool for cognition enhancement in AD orschizophrenia.

Reference(s)

[1] van Goethem, N.P., Prickaerts, J., Chesworth, R.,Shapiro, G., Boess, F.G., Methfessel, C., Re-neerkens, O.A.H., Flood, D.G., Hilt, D., Gawryl, M.,Bertrand, S., Bertrand, D., Konig, G., 2012. EVP-6124, a novel and selective a7 nicotinic acetylcholinereceptor agonist, improves memory performance bypotentiating the acetylcholine response of a7 nicotinicacetylcholine receptors. NeuroPharmacology, 62(2),1099–1110.

P.2.028 Enhanced vulnerability to cocainereinforcing effects in female H/Rouenmice selectively bred for depressive-likebehaviour

V. Rappeneau1 °, A.L. Morel1, P.H. Luppi1,J.M. Vaugeois1, M. El Yacoubi1, A. Berod1. 1Centre deRecherche en Neurosciences, CNRS UMR 5292 INSERMU1028, Lyon, France

Cocaine addiction is strongly associated with depression,however the mechanisms underlying this co-occurrenceare not well understood. We hypothesize that depressionstrongly enhances vulnerability for cocaine abuse. Femalesare particularly more vulnerable to depression, and to keyphases of the addiction process. Consequently, we em-ployed a genetically-induced mouse model of depression:the Helpless mice (H/Rouen) versus the Non Helplessmice (NH/Rouen). These lines have been selectively bredfor respectively high versus low spontaneous immobilityin the Tail Suspension Test (TST), a stress paradigmaimed at screening potential antidepressants [1]. The aimof our study was to determine the sensitivity to cocainepsychomotor stimulant and reinforcing effects of female

H/Rouen and NH/Rouen mice, and to shed light onthe neurobiological mechanisms by which depression-likestates enhance addictive behaviours.To determine the sensitivity to acute cocaine psychomo-

tor stimulant effects in H/Rouen and NH/Rouen mice,we compared the locomotor activity to acute cocaineadministration (0−20mg/kg i.p.). As expected, acutecocaine induced a dose-dependent increase in locomotoractivity in both lines. Interestingly, a right shift of the dose-response curve was observed in H/Rouen mice (interactiontreatment × line [F(10,187) = 4.90); p< 0.0001]). We thenevaluated the behavioural sensitisation induced by chroniccocaine treatment (10mg/kg for 5 days). The sensitisationtest was assessed by a cocaine challenge (1mg/kg),after a drug free period of 7 days [2]. We observeda similar motor activation in H/Rouen and NH/Rouenmice (p< 0.0001) pre-treated with cocaine. These resultsindicate similar sensitivity to chronic cocaine psychomotorstimulant effects in both lines, despite differences observedafter acute cocaine treatment.We also investigated the sensitivity to cocaine rewarding

effects in the conditioned place preference paradigm, thepredominant model of drug-seeking behaviour in mice,where animals were trained for 4 days to associatea specific environment to reinforcing properties ofcocaine (10mg/kg i.p.). The time spent in the drug-paired compartment was measured during the preferencetest as an indicator of cocaine rewarding effects [3].Results showed a stronger conditioned place preferencein H/Rouen compared to NH/Rouen mice (interactiontreatment × line [F(1,57) = 3.91; p = 0.05]). The cocainepreference score were interestingly correlated to theimmobility time in the TST (r = 0.42; p = 0.003).The main conclusion that may be drawn from our

experiments is that, compared to NH/Rouen mice,H/Rouen mice were more sensitive to cocaine rewardingeffects. Neuroanatomical studies were consequentlyundertaken to explore the neural substrates that mediatethe increased sensitivity to cocaine rewarding effectsobserved in H/Rouen females. Cocaine conditioned placepreference was associated with an increase in Fos in thenucleus accumbens (Acb), subregions of the prefrontalcortex (PFC), basolateral amygdala (BLA) and dorsalhippocampus (Hc) in H/Rouen and NH/Rouen femalescompared to controls. Moreover, preliminary resultsshowed that in H/Rouen mice, infralimbic and cingulatecortices and the Acb core were less activated whereasBLA and Hc were more activated during expression ofthe cocaine conditioned place preference.

Reference(s)

[1] El Yacoubi M. et al., 2003. Behavioral, neurochemical,and electrophysiological characterization of a genetic


mouse model of depression. Proc Natl Acad Sci USA100(10):6227−32.

[2] Post R.M. et al., 1992. Conditioned sensitization tothe psychomotor stimulant cocaine. Ann N Y AcadSci 654:386−99.

[3] Tzschentke T.M. et al., 2007. Measuring reward withthe conditioned place preference (CPP) paradigm:update of the last decade. Addict Biol 12(3−4):227–462.

P.2.029 Serotonin regulates hippocampalsynaptic plasticity and object memoryin mice

S.P. Fernandez1 °, A. Gruart i Masso2, P. Gaspar1.1Institut du Fer a Moulin, UMR-S 839 INSERM/UPMC,Paris, France; 2Pablo de Olavide University, Division ofNeurosciences, Seville, Spain

Low levels of serotonin (5-HT) have been associated withthe learning and memory deficits seen in Alzheimer’sdisease, autism and major depression [1]. Studiesperformed in healthy volunteers have shown that 5-HTdepletion disrupts consolidation of new information,specifically in tasks involving delayed recall and/orrecognition of visually-presented words, spoken words,pictures or abstract figures [1]. Despite this evidence, themechanism by which 5-HT regulates learning and memoryfunction is not clear.We used a genetic model of 5-HT depletion, Pet1 knock-

out mice (Pet1 KO), to understand the role that 5-HTserves in modulating learning and memory function. InPet1 KO mice, the levels of 5-HT are highly decreased,specifically associative cortical areas and the hippocampalformation are completely depleted of 5-HT axons [2].Our results showed that Pet1 KO mice are capable ofacquiring associative fear conditioning, reward reinforcedassociative learning and motor learning; however, theydo not recall familiarisation with objects. The novelobject recognition test was used to measure hippocampal-dependent declarative memory. The animals are trained tofamiliarise with two identical objects, and then subjected toa test session where one object is replaced by a novel one.Pet1 KO mice consistently failed to spend more time

exploring the novel object, suggesting impairment ofobject recognition memory (67±3% wild-type vs 51±4%KO, P< 0.01). This phenotype was reversed if 5-HT levelswere previously incremented in the brain by administrationof 5-HTP (5-HT precursor), which is without effect incontrol mice (69±3% wild-type vs 70±5% KO, P> 0.05).This observation strongly linked object recognition deficitwith 5-HT depletion; however does not specify the

underlying mechanism. Previously it was shown that theconsolidation of object memory involves the enhancementof synaptic strength across the CA3-CA1 hippocampalconnexion; a phenomenon called as long-term potentiation(LTP). To test the hypothesis that LTP deficit is responsiblefor the memory impairment observed in Pet1KO mice,we performed in vivo recordings of field potentials in theCA1 region of the hippocampus of mice while performingthe object memory task. We found that in Pet1 KO,experience-dependent LTP-like synaptic mechanisms areexaggerated compared to wild-type mice (126±10% wild-type vs 181±12% KO, P< 0.001), possibly explaining theaberration seen in forming correct maps of the objectpresented.Our results from in situ hybridisation studies showed the

inhibitory 5-HT1A receptors are highly expressed in theCA1 region of the hippocampus. We performed pharmaco-logical studies to determine whether the activation of thisreceptor subtype could correct object memory impairment.Pet1 KO mice injected with 8-OHDPAT (0.5mg/Kg), a5-HT1A agonist, spent significantly more time exploringthe novel object, suggesting that memory function wasre-established in these animals.These results established for the first time a direct

link between central 5-HT depletion, memory impairmentand synaptic plasticity deficits. We hypothesize that 5-HTneurons located in the median raphe contact pyramidalneurons in the CA1 region of the hippocampus, andthrough the activation of post-synaptic 5-HT1A regulatesynaptic plasticity and memory function.

Reference(s)

[1] Schmitt JA, Wingen M, Ramaekers JG, Evers EA,Riedel WJ, 2009. 5-HT and human cognitiveperformance. Curr Pharm Des 12, 2473−86.

[2] Kiyasova V, Fernandez SP, Laine J, Stankovski L,Muzerelle A, Doly S, Gaspar P, 2011. A GeneticallyDefined Morphologically and Functionally UniqueSubset of 5-HT Neurons in the Mouse Raphe Nuclei.J Neurosci 31, 2756−68.

P.2.030 Are nutritional variables associated withcognition in stimulant-dependence?

C.F. Whitelock1 °, K.D. Ersche1. 1University ofCambridge, Department of Psychiatry and Behaviouraland Clinical Neuroscience Institute, Cambridge, UnitedKingdom

Introduction: Recently, there has been increasing interestin the role of nutrition in the development of cognitivefunctions in both normal and disordered populations.


For example, diet has been linked to the progression ofcognitive impairments in ageing and to the hyperactive andattentional symptoms characteristic of Attention DeficitHyperactivity Disorder [1]. Alcohol- and drug-dependentindividuals show a variety of cognitive deficits relative tonon-drug using individuals. There is a well-documentedlink between these deficits and nutritional factors inalcoholism: some of the deficits seen in alcoholicsmay be due to their reduced ingestion of the vitaminthiamine for example [2]. However, there has been verylittle investigation into the relationship between nutritionand cognition in drug dependence. Anecdotally, drug-dependent individuals seem to differ in their diets fromnon-drug using individuals. If diet can affect cognitivefunction, a change in diet may alleviate some of thesymptoms associated with stimulant addiction.

Objectives: We hypothesised that dietary differencesbetween stimulant (cocaine and amphetamine)-dependentindividuals and healthy volunteers would be associatedwith differences in cognition.

Methods: Stimulant-dependent individuals (N = 58) andage-matched non-drug using healthy volunteers (N = 63)completed a Food Frequency Questionnaire to assess theirusual food intake and a battery of neuropsychological testson the Cambridge Automated Neuropsychological TestingBattery (CANTAB, www.camcog.com). Participants alsocompleted a selection of clinical questionnaires.

Results: Stimulant-dependent individuals consumedsignificantly higher calories and alcohol than healthyvolunteers. Controlling for these differences, the dietaryintake of drug users was also found to be significantlyhigher in fats, vitamin B6, fibre, magnesium, andmonounsaturated fatty acids, and lower in the consumptionof fruit and fructose. Stimulant-dependent individualsperformed significantly worse on the CANTAB test ofsustained attention than controls. In correlational analyses,nutrients such as fruit (r = 0.400, p< 0.01) and fibre(r = 0.425, p< 0.01) were positively related with reactiontimes on this task. However, this was only true in thehealthy volunteers.

Conclusions: Some of the nutritional differencesbetween stimulant-dependent individuals and healthyvolunteers were associated with performance on a test ofsustained attention in healthy volunteers only. Therefore,although a healthy diet may bestow health benefits, thecognitive impairments of stimulant-dependent individualsmay be such that they are not amenable to dietaryintervention. Alternatively, stimulant use may inhibit theactions of nutrients. Nutrition as having a role in thetreatment of drug-dependence may be an area worthfurther research and consideration. Changing one’s dietmay be a relatively simple and inexpensive, yet important,first step in recovery from stimulant-dependence. For

example, alleviating lack of attention is likely to aidindividuals in achieving optimal treatment outcomes.

Reference(s)

[1] Millichap, J.G., Yee, M.M., 2012. The diet factor inattention-deficit/hyperactivity disorder. Pediatrics 129,330–337.

[2] Kiela, P.R., 2010. Unraveling the pathophysiologyof alcohol-induced thiamin deficiency. Am J PhysiolRenal Physiol 299, F26-F27.

P.2.031 The CBA/J mouse as a genetic modelof visceral hypersensitivity withco-morbid anxiety and depression: roleof glutamate transport

R.D. Moloney1 °, T.G. Dinan1, J.F. Cryan2. 1UniversityCollege Cork, Psychiatry, Cork, Ireland; 2UniversityCollege Cork, Anatomy and Neuroscience, Cork, Ireland

Visceral pain is a pronounced and, at times, dominantfeature of a variety of gastrointestinal disorders. To datethere are no effective pharmacotherapeutic approachesto selectively treat this visceral hypersensitivity asthe underlying molecular mechanisms remain largelyunknown. Responses to painful stimuli differ betweenpopulations, ethnic groups, genders and even amongindividuals of a family. Strain surveys of common inbredand outbred mice are a useful way to gain insight into thegenetic contribution to complex disorders. To date, therehas been a lack of detailed information on the response ofvarious strains in the context of visceral nociception. Thisstudy aimed to redress this issue and investigate straindifferences in visceral nociception and its relationshipwith glutamate transporter (EAAT 1; excitatory aminoacid transporter 1) expression in the spinal cord. Anxietyand depressive-like behaviours were also investigated aspatients suffering from chronic pain disorders commonlydisplay co-morbid anxiety and depression. Furthermore,we investigated the underlying mechanisms (5HT1areceptor and brain derived neurotrophic factor, BDNF)of these comorbidities in the hippocampus, a key brainregion involved in such processes.Adult male mice of the following strains were

used: CBA/J, C3H/HeN, BALB/cOla, C57BL/6JOla,DBA/2JRcc, CAST/EiJ, SM/J, A/JOla, 129P2/Ola,FVB/NHan, Swiss Webster, CD-1. Colorectal distension(CRD) was performed as an indicator of visceralpain [1]. The open field and forced swim test wereperformed to assess anxiety and depressive-like behavioursrespectively. mRNA expression in the lumbar spinal cordand hippocampus was quantified using real time PCR(qRT-PCR).


There was a differential response to CRD in thestrains with the CBA/J and C3H/HeN strains displayinga significantly greatest response to CRD (F(11,117) = 5.358,p< 0.001, post hoc analysis, p< 0.05). Furthermore, thesetwo strains travelled significantly less in the inner zoneof the open field (F(11,116) = 24.300, p< 0.001, post hocanalysis, p< 0.05) and display increased immobility inthe forced swim test compared with C57BL/6J mice(F(11,113) = 34.565, p< 0.001, post hoc analysis, p< 0.05).At a molecular level the viscerally hypersensitive CBA/Jmice have reduced EAAT 1 mRNA expression in thespinal cord compared with the normosensitive C57BL/6Jmice (t(18) = 8.242, p< 0.001). Furthermore CBA/J micedisplay reduced 5HT1a receptor expression (t(18) = 2.615,p< 0.05) and BDNF expression (t(18) = 7.322, p< 0.001)in the hippocampus compared with C57BL/6J mice.This data demonstrates that strain differences occur

in visceral nociception with both CBA/JHsd andC3H/HeNHsd strains exhibiting visceral hypersensitivity.Moreover, these two strains display anxiety and depressive-like behaviours, which are commonly co-morbid withchronic pain disorders. These findings may pave the wayfor future epigenetic studies utilising techniques suchas QTL mapping to identify discrete regions of DNAassociated with a specific phenotypic trait, in particular,regions specific to visceral pain processes. Moreover, it iswell known that inter-individual differences occur in re-sponse to analgesic treatment both clinically and in animalmodels. These variations in efficacy could also be exploredin inbred mouse strains. Taken together this data may aidfuture work aimed at elucidating the mechanisms under-lying disorders where visceral pain and psychopathologyare co-morbid such as irritable bowel syndrome.

Reference(s)[1] Moloney, R.D., O’Leary, O.F., Felice, D., Bettler, B.,

Dinan, T.G., Cryan, J.F., 2012 Early-life stress inducesvisceral hypersensitivity in mice. Neurosci Lett, 512,99–102.

P.2.032 Nanoparticles as disease-modifyingmediators for brain therapy: focus onHuntington’s disease

B.M.D.C. Godinho1 °, J.R. Ogier2, R. Darcy2,C.M. O’Driscoll3, J.F. Cryan1. 1University CollegeCork, Department of Anatomy and Neuroscience, CorkCity, Ireland; 2University College Dublin, Centre forsynthesis and Chemical Biology, Dublin, Ireland;3University College Cork, Pharmacodelivery GroupSchool of Pharmacy, Cork City, Ireland

Neuropsychiatric disorders have been associated withaltered gene expression profiles or mutations within

specific genes which can dramatically influence anddetermine disease progression. Silencing the expressionof such disease-linked genes through short interferingRNA (siRNA) has recently been suggested as a promisingdisease-modifying approach for treating neurodegenera-tive diseases, such as Huntington’s Disease (HD), but alsoto treat depression [1,2]. However, neurons are notoriouslydifficult to transfect and the lack of appropriate and safesiRNA delivery systems has halted progression of RNAinterference (RNAi) technologies. Modified cyclodextrins(CDs) are oligosaccharide-based non-viral siRNA vectorsthat have recently shown to be able to deliver siRNAto neurons in vitro without causing marked toxicity [3].Thus, the aim of this study was to investigate behaviouralimprovements in the transgenic mouse model of HD (R6/2mouse model), after knockdown of the expression of themutant Huntingtin (HTT) gene using a CD-based siRNAdelivery system.Modified CDs (SC12 CD Click Propylamine) and

HTT siRNAs were prepared and complexed as previouslydescribed in O’Mahony et al. 2012 [3]. R6/2 mice weresubjected to stereotaxic surgery at 4−5 weeks of agefor bilateral implantation of cannulas or to performdirect injections into the striatum. Reduction in HTTgene expression was assessed by RT-qPCR. CD.siRNAnanoparticles were injected over 5 weeks (2.5 uL in eachside of the striatum) and behaviour assessment carriedout. Rotarod task, open field task and clasping behaviourwere monitored for up 14 weeks of age. All valuesare represented in average ±SEM and one-way repeatedmeasures ANOVA with Bonferroni’s Post-hoc test wascarried out for statistical analysis.Delivery of CD.siRNA nanoparticles to the R6/2

mouse brain enabled reduction of HTT gene expression.Gene knockdown was found to be sustained for upto 7 days post-injection at the site of injection.Nevertheless, no significant HTT gene knockdown wasobserved in regions of the brain away from the site ofinjection. Behaviour assessment revealed that CD.siRNAnanoparticles significantly delayed rotarod deficits whencompared to untreated R6/2 mice or naked siRNAtreated R6/2 mice (F(2,26) = 3.906, P = 0.033). Moreover,deterioration of rotarod deficits was observed whentreatment was ceased. No significant improvements wereobserved in the open field task or in clasping behaviour.Our data showed that local delivery of CD.siRNA

nanoparticles to the striatum reduces expression of theHTT gene and leads to improvements in the rotarodtask of R6/2 mice. Accumulation of the mutant HTTprotein in other regions of the R6/2 mouse brain mighthave contributed to the selective improvement in motorbehaviour. Using HD as a disease-model, here wedemonstrate that CD-based RNAi delivery can be used


effectively to regulate gene expression in the brain. Finally,the use of this technology is not restricted to HD butapplicable to other neuropsychiatric disorders.Authors acknowledge research funding from Science

Foundation Ireland (Grant no. 07/SRC/B1154) and theIrish Drug Delivery Network.

Reference(s)

[1] Thakker, D.R., Hoyer, D., Cryan, J.F., 2006. Interferingwith the brain: use of RNA interference forunderstanding the pathophysiology of psychiatric andneurological disorders. Pharmacology & therapeutics,109(3), 413–438.

[2] Bortolozzi, A., Castane, A., Semakova, J., San-tana, N., Alvarado, G., Cortes, R., Ferres-Coy, A.,Fernandez, G., Carmona, M.C., Toth, M., Perales, J.C.,Montefeltro, A., Artigas, F., 2012. Selective siRNA-mediated suppression of 5-HT1A autoreceptorsevokes strong anti-depressant-like effects. Molecularpsychiatry 17, 612–623.

[3] O’Mahony, A.M., Godinho, B.M.D.C., Ogier, J., De-vocelle, M., Darcy, R., Cryan, J.F., O’Driscoll, C.M.,2012. Click-Modified Cyclodextrins as NonviralVectors for Neuronal siRNA Delivery. ACS ChemicalNeuroscience 3, (10), 744–752.

P.2.033 Does serotonin depletion augment orcounteract the aggression-provokingeffect of testosterone in mice?

E. Studer1 °, J. Naslund1, L. Westberg1, E. Eriksson1.1Institute of Neuroscience and Physiology, Department ofPharmacology, Goteborg, Sweden

While sex hormones increase aggression in mostmammals [1], the neurotransmitter serotonin has beenreported to exert the opposite effect [2]. Since testosteroneinfluences various indices of serotonergic transmission,one possibility would be that it exerts its pro-aggressiveeffects by reducing a tonic anti-aggressive serotonergicinfluence, either by actions at the level of the raphe nucleiand the serotonergic cellbodies, e.g. affecting serotoninsynthesis or altering autoreceptor function. The hormonecould also directly affect serotonin signalling in terminalareas for serotonergic pathways by alterations in serotoninreceptor function or density. Alternatively, the hormoneand the transmitter may regulate aggression by parallel,independent paths in several brain loci where they haveopposing effects on one or several points of convergence.This study aimed at investigating the validity of

the interaction models proposed above by assessing iftestosterone is capable of enhancing aggression also in the

absence of serotonin. On the assumption that testosteroneenhances aggression by reducing serotonergic output,serotonin depletion would be at least as effective astestosterone in enhancing aggression, and no additionalpro-aggressive effect of testosterone above that inducedby serotonin depletion would be found.Male C57Bl/6 mice were gonadectomised, implanted

with slow release testosterone (T), releasing 250mg ofhormone per day, or blank (B) pellets and housedindividually after 3 weeks of recovery. Following 9 daysof isolation, to allow for establishment of territorialbehaviour, baseline aggression was assayed using thestandard resident intruder paradigm. Briefly, an unfamiliargroup-housed intruder mouse was introduced and be-haviour was recorded for 15 minutes under IR illumination.Starting the next day mice were treated with the serotoninsynthesis inhibitor para-chlorophenylalanine (pCPA) orsaline for 3 days and re-tested 24 hours after the finaldose of pCPA. While both groups of testosterone-treatedanimals displayed enhanced aggression as comparedto hormone-depleted animals serotonin depletion didnot enhance aggression in mice lacking testosterone,and did hence also not diminish the difference be-tween testosterone-treated and hormone-depleted animals(Duration of aggressive behaviour (s); T-pCPA= 75.4SD= 39.9; B-pCPA= 0.0 SD= 0.0; t(14) = 5.7, p< 0.001and T-Saline = 26.4 SD= 31.0; B-Saline = 0.1 SD= 0.1;t(15) = 2.5 p< 0.05). On the other hand, serotonindepletion did enhance aggression further in testosterone-treated mice (t(16) = −2.9, p = 0.011).In conclusion, our data suggest that androgen-induced

aggression is not dependent on intact serotonergic activity,but, on the contrary, is enhanced by serotonin depletion,suggesting that testosterone does not exert its pro-aggressive effect by inhibiting serotonergic output −instead serotonin seems to dampen the influence oftestosterone on aggression via a parallel inhibitorypathway. In addition, our results suggest that presence ofandrogens is an indispensable prerequisite for serotonindepletion to enhance aggressive behaviour, i.e. that theform of aggression dampened by serotonin is an androgen-dependent one.

Reference(s)

[1] Barfield RJ, Busch DE, Wallen K. 1972. Gonadalinfluence on agonistic behavior in the male domesticrat. Hormones and behavior 3, 247−59.

[2] Sanchez C, Arnt J, Hyttel J, Moltzen EK. 1993.The role of serotonergic mechanisms in inhibitionof isolation-induced aggression in male mice.Psychopharmacology 110, 53−59.


P.2.034 Swim-stress exposure effects onnociceptive behaviour and theendocannabinoid system in tworat strains differing in stressresponsivity

E.M. Jennings1 °, B. Okine1, W.M. Olango1, M. Roche2,D.P. Finn1. 1National University Ireland Galway,Pharmacology & Therapeutics College of MedicineNCBES Centre for Pain Research and NeuroscienceCluster, Galway, Ireland; 2National University IrelandGalway, Physiology College of Medicine, Galway, Ireland

Forced swim-stress produces hyperalgesia in Sprague-Dawley (SD) rats [1], but its effects in strains that are stresshyper-responsive (e.g. Wistar-Kyoto rat [WKY]) have notbeen assessed. The endocannabinoid system in the spinalcord and brain plays a key role in both stress and pain [2].The aim of this study was to compare the effects offorced swim-stress exposure on nociceptive responding tonoxious thermal and inflammatory stimuli in two rat strains(SD and WKY) that differ in their stress responsivity, andto determine associated alterations in the endocannabinoidsystem.Adult male SD and WKY rats (n = 8 per group)

were exposed to repeated forced swim sessions overten days; naive rats served as controls (n = 8). Thehot-plate and formalin tests were used to assessnociceptive responding to thermal and inflammatorystimuli, respectively. Mass spectrometry was performed onpost-mortem spinal cord, prefrontal cortex and amygdalato measure endocannabinoid levels and RT-qPCR was usedto measure key genes in the endocannabinoid system. Datawere analysed with repeated measures ANOVA or two-way ANOVA followed by Fisher’s LSD post-hoc analysisor Student’s t-test. P< 0.05 was considered statisticallysignificant and all data are mean±S.E.M.Neither strain displayed stress-induced differences in

thermal responding compared to non-stressed controls.SD rats exposed to ten days of swim-stress exhibited en-hanced late phase formalin-evoked nociceptive behaviour(Composite pain score: 1.1±0.1 vs. 0.6±0.1, P< 0.05)compared with controls. In contrast, WKY rats exposedto swim-stress showed reduced late phase formalin-evokednociceptive behaviour (Composite pain score: 0.1±0.1vs. 0.6±0.3, P< 0.01), versus controls. No significantchanges in endocannabinoid levels or key genes codingfor components of the endocannabinoid system were foundin WKY rat spinal cord, prefrontal cortex or amygdala.Enhanced formalin-evoked nociceptive behaviour in SDrats was not associated with changes in endocannabinoidlevels, CB1 receptor or fatty acid amide hydrolase (FAAH)

expression in the spinal cord. However, swim-stressedSD rats exhibited significantly increased expression ofmonoacylglycerol lipase (MAGL) (61% increase versusnaive, P< 0.05), an enzyme involved in the degradationof the endocannabinoid 2-arachidonoylglycerol (2-AG),in the dorsal horn of the spinal cord ipsilateral toformalin injection. Swim stressed SD rats also exhibiteda significant reduction in anandamide levels in theleft amygdala, compared with non-stressed controls(27.34±1.38 vs 71.37±5.02 nmol/g tissue, P< 0.05).There were no other effects of swim stress on levels ofendocannabinoids, related lipids or genes in the amygdalaor prefrontal cortex, though some within-group differencesbetween concentrations in the left vs right amygdala werenoted.In conclusion, swim-stress in SD rats causes enhanced

formalin-evoked nociceptive behaviour that is associatedwith increased spinal expression of the endocannabinoid-catabolising enzyme MAGL and a reduction in levelsof anandamide in the left amygdala. Further studies arerequired to determine the extent to which these alterationsin the endocannabinoid system may contribute to a stress-induced hyperalgesic phenotype.

Reference(s)

[1] Quintero L., Cardenas R., Suarez-Roca H., 2011.Stress-induced hyperalgesia is associated with areduced and delayed GABA inhibitory control thatenhances post-synaptic NMDA receptor activation inthe spinal cord. Pain 152 1909–1922.

[2] Butler, R.K., Finn, D.P., 2009. Stress-induced analge-sia. Prog Neurobiol 88 184–202.

P.2.035 ‘Anxious’ rats exhibit an enhancedanxiogenic response to acuteSSRI treatment and indices ofheightened serotonergic transmission

J. Naslund1 °, E. Studer1, R. Petterson1, H. Nissbrandt1,E. Eriksson1. 1University of Gothenburg, Department ofPharmacology, Gothenburg, Sweden

Introduction: Acute administration of serotonin reuptakeinhibiting drugs often elicits enhanced anxiety in subjectswith anxiety disorders or anxiety-related personalitytraits but not in non-anxious subjects [1,2], suggestingan enhanced responsiveness to an anxiogenic influenceof serotonin in anxious subjects. This observation, inconjunction with a large number of studies suggestingpolymorphisms in genes encoding serotonin-relatedproteins to be associated with anxiety-related personalitytraits, prompted us to investigate if baseline ‘anxiety’ in


male adult Wistar rats, as assessed using the elevatedplus maze, i) predicts the putatively anxiogenic responseto acute administration of an SSRI and ii) is associatedwith certain biochemical markers of central serotonergicneurotransmission.

Methods: Experiment I: 48 male Wistar rats, aged10 weeks were tested for five minutes on the EPM. Fourweeks later, paroxetine (10mg/kg, i.p.) was administered.The animals were allowed to return to their cages forone hour before being exposed to an open field paradigmfor 40 minutes. Experiment II: 30 male Wistar rats, aged10 weeks were tested for five minutes on the EPM. Sixweeks later, the animals were sacrificed and the brainsdissected. A small block of the brain stem, containing theraphe nuclei was homogenised with protein and mRNAextracted. qPCR was performed using Applied BiosystemsCustom LDA cards with reactions run in duplicate. TPH2levels were assayed using SDS-PAGE and western blotting,run in duplicate.

Results: Data are expressed as mean±S.E.M. andanalyses pertain to differences between the most ‘anxious’third of the animals (according to the EPM) in eachexperiment as compared to the rest. Gene expressiondata is relative to the control group (that is, the non-‘anxious’ animals) whereas protein data is relative to acontrol sample. Mann–Whitney U tests were used in thebehavioural experiment while t-tests were employed in thebiochemical experiment. Experiment I: ‘Anxious’ animalsexperienced a strong anxiogenic-like effect of paroxetinein the open field test (saline: 184.81±44.28, n = 8;paroxetine: 46.88±13.05, n = 8; p = 0.028), an effect absentin other animals (saline: 198.00±33.40, n = 16; paroxetine:230.74±65.33, n = 16; n.s.). Experiment II: ‘Anxious’animals consistently exhibited higher expression of genesrelated to serotonergic transmission in the raphe nuclei;TPH2 (relative expression; ‘anxious’ animals: 1.74±0.16,n = 9; non-‘anxious’: 1.00±0.18, n = 19; p = 0.004), SERT(relative expression; ‘anxious’ animals: 1.70±0.18, n = 9;non-‘anxious’: 1.00±0.20, n = 19; p = 0.011), as well ashigher levels of TPH2 as assayed by western blot (relativeexpression; ‘anxious’ animals: 1.70±0.12, n = 9; non-‘anxious’: 1.39±0.10, n = 20; p = 0.011).

Conclusions: The data obtained support the notionof central serotonergic function being associated withanxiety-like behaviour in rats. Animals assayed asmore ‘anxious’ had a strong response to acute SSRIadministration while also differing in measures ofcentral serotonergic function in a way that would beconsistent with a higher serotonergic activity. This indicatethat further investigation of differences in anxiety-likebehaviour between rats of the same strain and batch couldhelp shed some light over the relationship between geneticvariation in the serotonin system personality differences in

humans, as well as over the nature of the acute, anxiogeniceffect of SSRIs.

Reference(s)

[1] Rammsayer T, Netter P (1990). Personality relateddifferences in response to 5-HT uptake inhibition. IntJ Neurosci 55(2128079): 99–9106.

[2] Ramos RT, Gentil V, Gorenstein C (1993).Clomipramine and initial worsening in panic disorder:beyond the ‘jitteriness syndrome’. J Psychopharmacol7(22290840): 265–269.

S57

Epigenetics: towards new drug targets

Lectures

S.03.01 The epigenetics of brain disorders andtheir inheritance − focus on cognition

I. Mansuy1 °. 1Brain Research Institute, University/ETHZurich, Zurich, Switzerland

The aetiology and expression of behaviours in mammalsare strongly influenced by environmental factors, whetherpositive or negative. Such factors are particularly criticalduring early postnatal periods and can exert their influenceacross the entire life. When positive and favorable, theycan facilitate the proper development and expression ofbehavioural responses, but when adverse and stressful,they can alter behaviour and lead to cognitive disordersand psychopathological symptoms. In particular, traumaticevents in early life are strong risk factors for conditionssuch as attention deficit hyperactivity disorder (ADHD),bipolar depression, and antisocial behaviours. Further,while such disorders can strongly affect the individualswho were directly exposed to the traumatic events, theycan also often be inherited by the offspring and similarlyaffect behavioural responses in first and second generationindividuals. The biological processes responsible for thetransmission of stress-induced symptoms from parentto offspring remain poorly understood but since theyare induced by environmental factors, they have beenpostulated to involve epigenetic mechanisms. This talkwill present an experimental model of traumatic andchronic stress in early life in mice that provides initialevidence for the implication of epigenetic mechanismsin the expression and inheritance of the effects ofearly stress on behavioural responses. This mouse modelshows that chronic and unpredictable maternal separationcombined with unpredictable maternal stress during thefirst two weeks of postnatal life results in the developmentof multiple pathological symptoms in the offspring.It induces cognitive defects, alters social behavioursand social interactions, and leads to depressive-likesymptoms in adult individuals. But at the same time,such traumatic experience also leads to stress resilienceand favours behavioural flexibility in some conditions.These symptoms are strong and persist throughout lifein the animals exposed to stress. Further strikingly, they

are also transmitted to the following offspring across twogenerations, and through both females and males. At amolecular level, the behavioural symptoms are associatedwith persistent changes in several targets, includingcomponents of the HPA pathway, ion channels, neuro-transmitter receptors, and mitochondrial regulators. Initialevidence demonstrates that these molecular alterationscorrelate with persistent changes in DNA methylation atthe promoter-associated CpG island in several genes, andthat these changes are present both, in the brain of theoffspring and the germline of their fathers [1,2]. Further toDNA methylation, other epigenetic mechanisms includingregulation by non-coding RNAs and possibly histoneposttranslational modifications may also be involved.These findings suggest that epigenetic processes largelycontribute to the impact of negative environmental factorsearly in life on adult behaviour and its inheritance [3].

Reference(s)

[1] Franklin, T.B., et al., 2011. Influence of early stresson social abilities and serotonergic functions acrossgenerations in mice. PLoS One 6(7), e21842.

[2] Franklin, T.B., et al., 2010. Epigenetic transmissionof the impact of early stress across generations. BiolPsychiatry 68(5), 408−15.

[3] Bohacek, J., et al., 2012. Transgenerational epigeneticeffects on brain functions. Biol Psychiatry (ahead ofprint).

Posters

P.3.001 Valproic acid treatment preventsthe development of deficit insensorimotor gating in adult prenatallymethylazoxymethanol-treated rats

J. Latusz1 °, E. Bator1, P. Mordalska1, K. Wedzony1,M. Mackowiak1. 1Polish Academy of Sciences,Laboratory of Pharmacology and Brain Biostructure,Krakow, Poland

Prenatal administration of methylazoxymethanol (MAM)at embryonic day 17 (E17) is considered as aneurodevelopmental model of schizophrenia [1]. Althoughthe first symptoms of schizophrenia are seen in adults, the

S58 Epigenetics: towards new drug targets

factors cause the risk of this disease might be presentin early stage of life. The mechanism of this phenomenais still under investigation, however several data indicatethat epigenetic regulation of gene expression duringdevelopment might contribute to behavioural phenotypesof schizophrenia [2]. Thus, the aim of this study wasto investigate whether pharmacological manipulation inepigenetic mechanisms by valproic acid (VA), an inhibitorof histone deacetylases, during pre- and post-pubertymight influence the induction of schizophrenic symptomsin adults. The deficit in sensorimotor gating as a one ofthe psychotic symptoms observed after puberty in MAM-treated animals [1] was chosen to determine which periodof postnatal life is critical for appearance behaviouralsign of psychosis. Pregnant females were injected ip with22mg/kg/ml MAM or saline at E17. The offspring wereweaned 21 days after birth and only males were used inexperiments. VA was given at the dose 250mg/kg, sc,twice a day, from 23rd to 29th day (pre-puberty) or byprepulse induced inhibition of acoustic startle response(PPI) in rats at postnatal days 30, 45, 60, 70, 80, 90 and120 (P30, P45, P60, P70, P80, P90 and P120). Statisticallysignificant decrease in PPI in MAM treated group wasfound since P70 till P120 [(F(3, 66) = 2.8437, p< 0.04)for P70], but not at P30, P45 or P60 [F(3,66) = 0.3971,p = 0.755, F(3, 66) = 1.046, p = 0.38 and F(3,66) = 1.45,p = 0.24, respectively]. Administration of VA in pre-puberty did not influence the sensorimotor gating processat P30 [F(3, 84) = 0.17, p = 0.92], P45 [F(3,84) = 0.713,p = 0.55] and P60 [F(3,84) = 0.52, p = 0.69], but preventedthe appearance of deficit in sensorimotor gating in MAM-treated group measured at P70 [F(3,84) = 2.8, p< 0.4].The above effect was transient, since the decrease inPPI in MAM-treated group was observed again at P80[F(3,84) = 0.76, p = 0.52]. When VA was given in post-puberty, it blocked the reduction in PPI in MAM-treatedgroup detected at P80 [F(3,84) = 3.07, p< 0.04], at P90[F(3,84) = 5.338, p< 0.003] and P120 [F(3,84) = 3.311,p< 0.03]. The obtained data indicate that the deficit insensorimotor gating appears only in post-puberty as itwas reported by Moore et al. [1]. VA given pre-pubertyonly delays appearance of deficit in sensorimotor gatingin MAM-treated rats. In contrast, VA administration inpost-puberty prevents the deficit evoked by MAM. Thus itis conceivable that inhibition of deacetylation process inpost- but not in pre-puberty might block the developmentof some psychotic symptoms such as impairment insensorimotor gating.

Reference(s)

[1] Moore, H., Jentsch, J.D., Ghajarnia, M., Geyer, M.A.,Grace, A.A., 2006. A neurobehavioral systems

analysis of adult rats exposed to methylazoxymethanolacetate on E17: implications for the neuropathologyof schizophrenia. Biol. Psychiatry 60, 253–264.

[2] Dudley, K.J., Li, X., Kobor, M.S., Kippin, T.E.,Bredy, T.W., 2011. Epigenetic mechanism mediatingvulnerability and resilience to psychiatric disorders.Neurosc. & Biobeh. Reviews 35, 1544–1551.

Disclosure statement: This study was supported by grantMNiSzW NN401 066938.

P.3.002 The effect of valproic acid on changesin methylation pattern of histone H3induced by prenatal MAM administrationin mPFC

E. Bator1 °, J. Latusz1, P. Mordalska1, K. Wedzony1,M. Mackowiak1. 1Polish Academy of Sciences,Laboratory of Pharmacology and Brain Biostructure,Krakow, Poland

Several data indicate that epigenetic regulation ofgene expression might be involved in schizophreniaaetiology [1]. Our previous study showed the changesin the methylation pattern of histone H3 in the medialprefrontal cortex (mPFC) in neurodevelopmental modelof schizophrenia [2] based on the prenatal administrationof methylazoxymethanol (MAM) at embryonic day 17(E17) [3]. Thus, the aim of the present study wasto determine whether the repeated valproic acid (VA)administration at pre-puberty can affect the observedchanges of histone H3 methylation at lysine 9 (H3K9)at 30 day old (P30) and at lysine 4 (H3K4) at 60- or 70-day-old rats (P60 or P70) receiving prenatally MAM. VAacting as an inhibitor of histone deacetylases can influencethe epigenetic mechanism and might affect the methylationprocess of histone H3. Pregnant females were injected with22mg/kg/ml MAM or saline ip at E17. The offspring wereweaned 21 days after birth and only males were used inexperiments. VA was given at the dose 250mg/kg, sc,twice a day, from 23rd to 29th day (early adolescence)and the animals were killed at P30, P60 and P70. Atthe same dose VA was injected from 53rd to 59th day oflife (late adolescence) and experiments were performedat P60 and P70. The methylation levels of H3K9me2and H3K4me3 were studied in nuclear fraction of mPFCusing Western blot analysis. It was observed that MAMincreased H3K9me2 level at P30 (30%) [F(1,16) = 6.49,p< 0.03], however VA pretreatment in early adolescencedid not have any effect on observed increase in H3K9me2induced by MAM [F(1,16) = 1.77, p = 0.20]. MAM aswell as VA did not affect the H3K9me2 protein levelat P60 [F(1,16) = 1.2, p = 0.29] or P70 [F(1,16) = 0.15,

Epigenetics: towards new drug targets S59

p = 0.77] for both procedures of VA administration. Incontrast, H3K4me3 methylation level was decreased inpost-puberty at P60 (30%) (F(1,16) = 19.4, p< 0.001) andalso at P70 (35%) [F(1,16) = 5.12, p< 0.04] in MAM-treated group, but not at P30 [F(1,16) = 3.99, p = 0.069].VA given in early adolescence prevented the observeddecrease in H3K4me3 protein in MAM-treated groupat P60 [F(1,16) = 6.084, p< 0.03]. Interaction betweenMAM and VA was insignificant at P70 [F(1,16) = 1.94,p = 0.18], but post hoc analysis revealed the significanteffect of VA administration on MAM-treated group(p< 0.006). VA pretreatment in late adolescence did notalter MAM induced decrease in H3K4me3 protein levelat P60 [F(1,16) = 0.43, p = 0.52], but prevented MAM-evoked reduction of trimethylated form of H3K4 atP70 [F(1,16) = 11.72, p< 0.004]. In addition, the totalhistone H3 level was measured and there were nodifferences between groups at any age points. The obtainedresults indicate that VA administration in adolescencemight prevent changes in gene expression in post-pubertycontrolled by methylation at H3K4.

Reference(s)

[1] Roth, T.L., Lubin, F.D., Sodhi, M., Kleinman, J.E.,2009. Epigenetic mechanism in schizophrenia.Biochimica and Biophysica Acta 1790, 869–877.

[2] Mackowiak, M., Bator, E., Latusz, J., Mordalska, P.,Wedzony, K., 2012. The impairment of lysine methy-lation of histone H3 in the medial prefrontal cortexin the neurodevelopmental model of schizophrenia.FENS Abstract 6, 118.14.

[3] Moore, H., Jentsch, J.D., Ghajarnia, M., Geyer, M.A.,Grace, A.A., 2006. A neurobehavioral systemsanalysis of adult rats exposed to methylazoxymethanolacetate on E17: implications for the neuropathologyof schizophrenia. Biol. Psychiatry 60, 253–264.

P.3.003 Chronic early-life stress programshippocampal neurogenesis andcognitive function: a role forepigenetics?

E.F.G. Naninck1 °, L. Hoeijmakers1, M. Engel1,P.J. Lucassen1, A. Korosi1. 1UvA SILS Center forNeurosciences, Structural and functional plasticity of thenervous system, Amsterdam, The Netherlands

Rationale: Early-life stress (ES) increases vulnerabilityto psychopathologies and impairs cognitive functions laterin life. However, the responsible molecular mechanismsremain elusive. Since ES in humans is common andoften cannot be prevented, a better understanding of the

mechanisms underlying these lasting effects of ES onbrain functioning and disease susceptibility is neededin order to develop therapeutic intervention. Rodentstudies have indicated that ES induces lasting changesin hippocampal neurogenesis and structure, parallel tocognitive deficits [1]. We therefore set out to investigate, inmice, if these lasting abnormalities have an early onset, andassess the acute and lasting effects of ES on neurogenesisand on hippocampal epigenetic modifications. In addition,we investigate the lasting effects of ES on cognition andemotion.

Methods: Chronic ES is induced in C57Bl/6 miceby limiting nesting/bedding material in the cage frompostnatal day (P) 2 till P9 [2]. Levels of postnatal andadult neurogenesis in the hippocampal dentate gyrus (DG)of the offspring are assessed using immunohistochemistryfor proliferating (Ki67+) and differentiating (NeuroD1+/Calretinin+) newborn cells and for their survival (BrdU+).Total histone levels (H3, H2b) as well as active(H3K4me3) and repressive (H3K9me3, H3K27me3)histone modifications are assessed by Western blot of DGtissue. All parameters are measured directly after ES (atP9) and in adulthood (at P150) in ES and control maleand female mice. In addition, performance in the elevatedplus maze, forced swim task, novel object recognition task,novel object location task and Morris water maze wasexamined in ES and control mice from P150 onwards.

Results: ES dams show characteristics of chronic stressas was evident from fragmented maternal care and areduced bodyweight gain. The fragmentation of maternalcare subsequently led to chronic ES in the pups, thatshowed reduced bodyweight gain, reduced thymus weightand elevated basal plasma corticosterone levels at P9.Interestingly, levels of postnatal proliferation (Ki67+)and differentiation (Calretinin+) were increased in theDG of 9-day-old ES mice, indicating an early onset ofthe lasting changes in hippocampal structure after ES.Furthermore, in ES mice, more NeuroD1+ cells were foundin the postnatal migratory stream from the hippocampalsubventricular zone to the DG. Expression levels ofH3/H2b were elevated in ES mice at P9. In addition,behavioural analysis indicates cognitive impairments inadult ES mice. Further characterisation of the lastingeffects of ES on adult neurogenesis and epigeneticregulation is in progress.

Conclusion: Chronic ES alters levels of postnatalneurogenesis already at P9 and impairs hippocampus-dependent cognitive function in adulthood. Unravellingthe molecular mechanisms that underlie these persistentES-induced changes will yield important information onhow stressful early-life experiences program the brain.This may have clinical implications for the developmentof treatments to prevent or repair ES-induced deficits.


Reference(s)

[1] Korosi, A., Naninck, E.F.G., Oomen, C.A.,Schouten, M., Krugers, H., Fitzsimons, C., Lu-cassen, P.J., 2012. Early-life stress mediated modula-tion of adult neurogenesis and behavior. Behav BrainRes 227, 400−9.

[2] Rice, C.J., Sandman, C.A., Lenjavi, M.R., &Baram, T.Z., 2008. A novel mouse model for acuteand long-lasting consequences of early life stress.Endocrinology, 149, 4892–4900.

P.3.004 Possible interplay between BDNF anddynorphin in bipolar disorder: role ofepigenetic mechanisms

A. Di Francesco1 °, C. D’Addario1, B. Dell’Osso2,M.C. Palazzo2, B. Benatti2, D. Galimberti3, B. Arosio4,A.C. Altamura2, M. Maccarrone1. 1University ofTeramo, Biomedical Sciences, Teramo, Italy; 2Universityof Milan, Psychiatry, Milan, Italy; 3University of Milan,Neurological Sciences, Milan, Italy; 4University of Milan,Internal Medicine Geriatric Unit, Milan, Italy

Purpose of the study: Bipolar disorder (BD) is a highlydisabling mood disorder determined by the interplaybetween genes and environmental factors [1]. Amonggenes potentially implicated in the pathophysiology of BD,the brain-derived neurotrophic factor (BDNF) gene hasbeen extensively investigated and we recently suggestedselective changes in DNA methylation of BDNF promoterin subjects with BD type II, highlighting the importanceof epigenetic factors in mediating the onset and/orsusceptibility to BD [2]. To provide further knowledgeon the role of BDNF in the development of BD, wehere studied the epigenetic regulation of prodynorphingene, the precursor of the opioid peptide dynorphin,recently suggested as a downstream effector of BDNFregulation [3].

Methods: We conducted this study on peripheralblood mononuclear cells (PBMCs), accessible cells witha potential use for biomarker discovery in psychiatricdisorders, containing the full complement of epigeneticenzymes found in most tissues, including neurons.Genomic DNA and total RNA were isolated from patientswith BD on stable pharmacological treatment (BD I:n = 48, BD II: n = 41) and from healthy controls (n = 42),with the specific intent of assessing eventual differences interms of methylation between bipolar patients and healthysubjects. To assess prodynorhin mRNA abundances andto quantify prodynorphin promoter DNA methylation,we used Real-Time RT-PCR and Real-Time MethylationSpecific PCR, respectively.

Results: A significant hypermethylation of the pro-dynorphin promoter region was specifically found inBD II patients (CT: 17.14±1.25%; BD I: 18.06±1.26%;BD II: 21.55±1.08%, P< 0.05 vs CT). Consistently, down-regulation of prodynorphin gene expression was observedin BD II but not in BD I patients when compared tocontrols. Of note, higher levels of DNA methylation wereobserved in BD subjects on pharmacological treatmentwith antidepressants (21.01±1.23%) compared with thoseexclusively on mood-stabilising agents (16.89±1.48%;P< 0.05). Moreover, we found a significant correlationbetween BDNF and prodynorphin DNA methylation atgene promoters (P< 0.05, Spearman R= 0.2017).

Conclusions: Present results provide a new clearcorrelation between BDNF and prodynorphin alterationsat epigenetic level, suggesting their possible interactionin the development of BD. This is also supported bythe evidence that mood stabilisers regulate in a similarmanner BDNF and prodynoprhin DNA methylation levelsand thus providing new insight on the specific mechanismof action of these drugs for the treatment of BD. Moreover,we also suggest a possible role for dynorphin as peripheralbiomarker useful for differential diagnosis and, mostly, forprediction of treatment response. Overall, our findings areconsistent with the epigenetic theory of major psychoses,supporting the importance of DNA methylation alterationsin the aetiology of BD.

Reference(s)

[1] Labrie, V., Pai, S., Petronis, A., 2012. Epigenetics ofmajor psychosis: progress, problems and perspectives.Trends Genet. 28, 427–435.

[2] D’Addario, C., Dell’Osso, B., Palazzo, M.C., Be-natti, B., Lietti, L., Cattaneo, E., Galimberti, D.,Fenoglio, C., Cortini, F., Scarpini, E., Arosio, B., DiFrancesco, A., Di Benedetto, M., Romualdi, P., Can-deletti, S., Mari, D., Bergamaschini, L., Bresolin, N.,Maccarrone, M., Altamura, A.C., 2012. SelectiveDNA methylation of BDNF promoter in bipolardisorder: differences among patients with BDI andBDII. Neuropsychopharmacology. 37, 1647–1655.

[3] Logrip, M.L., Janak, P.H., Ron, D., 2008. Dynorphinis a downstream effector of striatal BDNF regulationof ethanol intake. FASEB J. 22, 2393–2404.


P.3.005 Early environment affects activitybased anorexia genetic susceptibilityand epigenetic programming ofneurodevelopmental genes in mice

E. Pjetri1 °, E.V.S. Hessel1, H. Oppelaar1, P.N.E. deGraan1, B. Olivier2, M.J.H. Kas1. 1RudolfMagnus Institute of Neuroscience, Neuroscience andPharmacology, Utrecht, The Netherlands; 2UtrechtUniversity, Department of Pharmacology, Utrecht, TheNetherlands

Background: Human studies have shown that theinteraction between genetic and environmental factorscontribute to adult expression of psychiatric phenotypes,such as anti-social behaviour [1]. Recently, gene–environment interactions have been increasingly reportedfor a wide range of mental disorders such as schizophrenia,depression, and eating disorders [2]. Anorexia nervosais a complex eating disorder associated with the largestmortality rate among psychiatric disorders [3]. Variousgenetic, environmental and developmental risk factors arethought to contribute to this dramatic disease. However, lit-tle is known about the developmental trajectory precedingthis psychiatric disorder. Systematic longitudinal studieswith controlled genetic and environmental backgroundvariation are necessary to identify and understand themechanisms underlying the interplay between environmentand genetic background susceptibility for maladaptivebehaviour. In the present study, the relationship betweenmaternal environment, genetic background and epigeneticmechanisms was studied using low activity basedanorexia (ABA) (C57BL/6J) and high ABA (C57BL/6J-Chr4A/ J/NaJ) (CSS4) susceptible mouse inbred strains.

Methods: Following early life inter- and intra-straincross-fostering of the inbred strains, the mice were exposedto ABA model in adulthood (3−4 months old).The ABA rodent model mimics the paradoxical expres-

sion of behavioural hyperactivity in periods of reducedfood intake that is also observed in anorexia patients.In this rodent model, animals have unlimited voluntaryaccess to a running wheel throughout the experiment,while food is ad libitum available for a limited period at thesame time during several consecutive days. The physicalhyperactivity observed during food restriction leads toaccelerated body weight loss, reminiscent of the featuresof the disease. ABA susceptibility is a ‘time to an eventparameter’ and therefore it was analysed using Kaplan–Meier survival analysis with Mantel-Cox Log-Rank test.To investigate whether fostering was associated with

epigenetic modifications, genome wide DNA methylationlevels were examined in DNA extracted from hippocampal

tissue in cross-fostered and non-cross-fostered CSS4 micea week after weaning the pups from their mother (postnatalday 35).

Results: Independent of the foster mother geneticbackground, fostered CSS4 mice were less susceptibleto ABA when compared to CSS4 mice raised by theirbiological mother (same strain foster mother: c2 = 7.690,p = 0.006; different strain foster mother: c2 = 7.874,p = 0.005). In contrast, no cross-fostering effects wereobserved on ABA susceptibility in the low susceptibleC57BL/6J strain. Global DNA methylation analysisrevealed several differentially hypermethylated genomeregions as a function of cross-fostering in CSS4 strain.These hypermethylated regions included genes, suchas contactin associated protein-like 2 (Cntnap2) geneand a-1C subunit of the L-type voltage-gated calciumchannel (Cacna1c) gene, that have been associated withneurodevelopmental disorders.

Conclusions: Thus, early life events may modulategenetic background susceptibility for adult maladaptivebehaviour via epigenetic modifications and providea basis for studies towards further understanding ofneurodevelopmental processes underlying adult behaviour.

Reference(s)

[1] Caspi, A., McClay, J., Moffitt, T.E., Mill, J., Martin, J.,Craig, I.W., Taylor, A. & Poulton, R., 2002. Roleof genotype in the cycle of violence in maltreatedchildren. Science 297, 851–854.

[2] Campbell, I.C., Mill, J., Uher, R. & Schmidt, U., 2011.Eating disorders, gene–environment interactions andepigenetics. Neurosci Biobehav Rev. 35, 784–793.

[3] Arcelus, J., Mitchell, A.J., Wales, J. & Nielsen, S.,2011. Mortality rates in patients with anorexia nervosaand other eating disorders. A meta-analysis of 36studies. Arch Gen Psychiatry 68, 724–731.

P.3.006 Epigenetic modifications of bdnf geneat hippocampal level induced by chronicethanol intake in C57BL/6J mice

E. Stragier1 °, R. Massart1, M. Hamon1, L. Lanfumey1.1INSERM UMR 894, Centre de Psychiatrie etNeurosciences, Paris, France

Purpose of the study: Although alcohol at high dosage iswell known to induce neurotoxic effects on hippocampalneurogenesis, chronic voluntary intake of alcohol atmoderate dosage has been shown to stimulate dentategyrus (DG) cell proliferation in C57BL/6J mice [1]. Inorder to unveil the molecular mechanisms underlyingalcohol-induced neuroplasticity, we studied the expression


of several markers involved in cell differentiation and plas-ticity in the hippocampus of adult male C57BL/6J micethat had been exposed to free alcohol (10% ethanol versuswater) consumption for three weeks. In addition, alcohol-induced epigenetic modifications were investigated byquantifying the expression of epigenetic markers such asHDACs, which are implicated in gene transcription controland neuronal maturation. Post-translational modificationsof histones (acetylation, methylation), which are knownto modulate gene transcription, were also determined inalcohol- versus water-exposed mice.

Methods: qRT-PCR, chromatin immunoprecipitationand immunohistochemistry approaches were used forrelevant determinations in the hippocampus of 3 month-old male C57BL/6J mice after a 3-week free consumptionof ethanol versus tap water. Paired control mice had onlyaccess to water for the same period.

Results: Voluntary alcohol consumption reached astable level corresponding to 10−12 g of ethanol ingestedper day from the 10th day of exposure to three bottlesof 10% ethanol and a bottle of tap water. After threeweeks of such free ethanol consumption, a significantincrease in the expression of both bHLH activator family(transcription factors implicated in neural cell proliferationand maturation) and BDNF (at both mRNA and proteinlevels), and a concomitant decrease in HDACs, were notedin the hippocampus of ethanol-exposed mice versus pairedmice exposed to water. In addition, MeCP2 expression wasincreased also at both mRNA and protein levels in theDG and CA3 area, and acetylated histone H4 (H4Ac) andtrimethylated histone H3 (H3K4me3) were upregulatedin these hippocampal subareas in alcohol-exposed mice.Specific analysis of bdnf gene showed alcohol-inducedmodifications at transcription level associated with clear-cut changes in H3Ac and H3K4me3 in bdnf promoters(Table 1). Changes in BDNF exons expression occurredin parallel with alcohol-induced increase in BDNF proteinin the hilus and CA3 area of the hippocampus.

Table 1. Alcohol-induced changes in BDNF exons expression andhistone post-translational modifications in bdnf promoters in the mousehippocampus (+, increase; −, decrease)

BDNF exon mRNA expression Promoter

H3Ac H3K4me3

II + = +III + − +VI + + =VIII − − −

Conclusions: Altogether, these results suggest that theincreased hippocampal neurogenesis produced by chronicmoderate alcohol intake under free choice conditions in

C57BL/6J mice [1] is underlain by an up regulation oftranscription factors and BDNF. Epigenetic mechanismsinvolving HDAC and histone modifications at the levelof bdnf gene play probably a key role in alcohol-inducedBDNF upregulation and downstream neurogenesis.

Reference(s)

[1] Paizanis E., Kelaı S., Renoir T., Hamon M., Lan-fumey L. 2007. Life-long hippocampal neurogenesis:environmental, pharmacological and neurochemicalmodulations. Neurochem. Res. 32:1762–1771.

P.3.007 Stress-induced miRNA changes indepression: peripheral biomarker orpathophysiology?

S. Kalman1 °, K. Garbett2, A. Vereczkei2, R.C. Shelton2,K. Mirnics2. 1University of Szeged, Faculty of Medicine,Szeged, Hungary; 2Vanderbilt University, Kennedy Center,Nashville, USA

Introduction: Depression is the second most commonhuman disorder worldwide (WHO). Genetic predispositionand maladaptive stress response considered to be the mostimportant aetiological factors. microRNAs can influencethe metabolism of their target mRNAs, act like fine,‘tuning’ regulators of adaptation, cell cycle, and apoptosisand are proved to be especially important in certain(patho)physiologic processes of the central nervous systemsuch as synaptoplasticity and degeneration [1]. Ouraim was to answer the following questions: (I.) Whatis the difference between the microRNA expressionof dermal fibroblasts originated from control (CNT)and depressed (D) subjects? (II.) How metabolic stresstreatment affects the microRNA expression in the CNTand (III.) D groups? (IV.) How these microRNA profilechanges appear on the level of their target mRNAs?

Methods: Primary fibroblast cultures (passage num-ber = 5−10) were initiated from skin biopsies of 17 controlsand 17 subjects with major depression. Groups werematched for sex and age. The metabolic stress wasevoked by growing subcultured cells in glucose deprived,galactose enriched (GAL); lipid reduced, cholesteroldeficient (NL); and standard (STD) medium for one week.Total- and small-RNAs were isolated and the expressionlevels of more than 1000 miRNAs were measured withmiRNome PCR array. We accepted ddCt � 0.583 andp-valuegrouped �0.05, p-valuepaired �0.05 as a significantchange. The amounts of their suspected target mRNAswere also detected with DNA microarray. The genewas considered to be differentially expressed with theALR � 0.378 and p-value �0.01.


Results: As compared to the CNT fibroblasts (75affected microRNAs), the D samples showed almostthree times more microRNA expression changes (214microRNAs) caused by both GAL and NL metabolic stresstreatments. The microRNA profile alterations were moreremarkable under NL circumstances. Although higheroverlap was detected between the GAL and NL inducedmicroRNA changes in the D group (47 microRNAs, whileonly 4 microRNAs in the CNT), Pearson correlationshowed that the two different stress treatments resultedin very similar microRNA expression pattern in the CNT(r2 = 0.51 and 0.43) but not in the D group. Nearly 10%of the suspected target mRNAs of the most significantlyaffected microRNAs changed due to metabolic stresstreatments in the CNT group.

Conclusion: We suppose that the metabolic stressinduced microRNA changes in dermal fibroblasts ofCNT subjects are part of the normal cell adaptation,while those found in the D group could be the signof a genetically determined, pathologic stress responsepresumed in depression. These results could facilitate theunderstanding of microRNAs’ role in the development ofdepression and contribute to the mapping of the stressrelated physiologic and degenerative gene expressionchanges by setting up a potential new in vitro metabolicstress related depression model and diagnostic tool.

Reference(s)

[1] Serafini, G., Pompili, M., Innamorati, M., Gior-dano, G., Montebovi, F., Sher, L., Dwivedi, Y.,Girardi, P., 2012. The role of microRNAs in synapticplasticity, major affective disorders and suicidalbehavior. Neurosci Res 73(3):179−90.

P.3.008 S-Adenosyl-methionine impairsforced swimming-induced behaviouralimmobility by inhibiting gene expressionin dentate gyrus neurons

E.A. Saunderson1 °, A.F. Trollope1, M. Gutierrez-Mecinas1, A.A. Shaikh1, H. Spiers2, J. Mill2,J.M.H.M. Reul1. 1University of Bristol, School ofClinical Sciences, Bristol, United Kingdom; 2King’sCollege London, Institute of Psychiatry, London, UnitedKingdom

The consolidation of stress-induced adaptive behaviours,such as the learned immobility response in the forced swim(FS) test, depends on specific epigenetic modificationsunderlying gene transcriptional responses in dentate gyrus(DG) granule neurons of the hippocampus. In theseneurons FS evokes the activation of two interacting

signalling pathways, i.e. the glucocorticoid receptor(GR) and the NMDAR/ERK1/2/MSK1-Elk-1 pathways,resulting in phosphorylation of serine10 and acetylationof lysine14 at histone H3 (H3S10p-K14ac) which leadsto induction of immediate early genes (IEGs) c-Fosand Egr-1 [1,2]. These molecular responses are criticalfor the consolidation of the behavioural immobilityresponse [1,2].The drug and endogenous methyl-donor S-adenosyl-

methionine (SAM) impairs the consolidation of thebehavioural immobility response [3] suggesting the in-volvement of histone methylation and/or DNAmethylationin gene transcriptional control underlying the behaviouralresponse but this is unknown. Therefore, to understandthe mechanism of action of SAM and to gain insight intothe involvement of histone/DNA methylation, rats wereinjected with SAM (100mg/kg s.c.) 30 minutes beforeFS (15min, 25ºC). Twenty-four hours later they weresubjected to FS again and immobility behaviour werescored in 10 s bins. SAM had no effect on immobility in theinitial FS test (mean±SEM; vehicle: 15.8±1.6 bins, n = 8;SAM: 15.0±1.9 bins, n = 9; P> 0.05 post-hoc Bonferronitest). However, in the retest, immobility in the SAMgroup was significantly lower (11.4±3.3 bins, n = 9) thanthat in the vehicle group (20.6±2.3 bins, n = 8; P< 0.05),confirming that SAM indeed impairs the consolidation ofthe behavioural immobility response [3].To study whether SAM’s effects on behaviour could

be explained through effects on FS-induced epigeneticand transcriptional responses in DG neurons, rats werepre-treated with SAM or vehicle and submitted to FSor not (baseline control) and killed 1 h later. Comparedto vehicle, SAM evoked a significant decrease in FS-induced c-Fos and Egr-1in DG neurons (Vehicle: c-Fos:127.1±4.6 neurons n = 5, Egr-1: 18.4±2.5 n = 5; SAM:c-Fos 82.6±5.0 neurons n = 6, Egr-1 8.5±1.2 n = 5; bothc-Fos and Egr-1: P< 0.01). However, SAM had noeffect on stress-induced H3S10p-K14ac in DG neuronssuggesting that the drug effect was independent of thisdual epigenetic mark. Next, we investigated the effect ofSAM and FS on the methylation status of histone H3 lysineresidues. Chromatin immuno-precipitation (ChIP) analysisrevealed that SAM and FS had no significant effects onH3K4me3, H3K9me3 and H3K27me3 at the c-Fos andEgr-1 promoters.Recently, we have started investigating DNA methyla-

tion using bisulfite pyrosequencing and methyl-DNA IP(MeDIP) to assess the cytosine methylation status of CpGislands within the c-fos and egr-1 gene promoters. Wefound that cytosine methylation of both promoters wasvery low (<5%) in the DG, the rest of hippocampus, andthe neocortex and did not change after FS. Presently we are


studying the effects of SAM on de novo DNA methylationof the gene promoters.We conclude that SAM impairs the FS-induced

behavioural immobility response through inhibition ofgene transcription in DG neurons. This drug effect appearsto be independent of histone modifications.

Reference(s)

[1] Reul, J.M.H.M., Hesketh, S.A., Collins, A., Gutierrez-Mecinas M., 2009. Epigenetic mechanisms in the den-tate gyrus act as a molecular switch in hippocampus-associated memory formation. Epigenetics 4: 434–439.

[2] Gutierrez-Mecinas, M., Trollope, A.F., Collins, A.,Morfett, H., Hesketh, S.A., Kersante, F., Reul,J.M.H.M., 2011. Long-lasting behavioral responses tostress involve a direct interaction of glucocorticoidreceptors with ERK1/2-MSK1-Elk-1 signaling. ProcNatl Acad Sci USA 108: 13806–13811.

[3] Czyrak, A., Rogoz, Z., Skuza, G., Zajaczkowski, W.,Maj, J., 1992 Antidepressant activity of S-Adenosyl-L-Methionine in mice and rats. J Basic & Clin PhysiolPharmacol 3: 1−16.

P.3.009 Differential effects of prenatal stress onserotonin transporter deficient mice: therole of epigenetic programming

K. Schraut1 °, S.B. Jakob1, G. Kenis2, A.G. Schmitt3,S. Kneitz4, C.J. Scholz4, G. Ortega1, H. Steinbusch2,D.L.A. van den Hove2, K.P. Lesch1. 1University ofWurzburg, Psychiatry Psychosomatics and PsychotherapyMolecular Psychiatry, Wurzburg, Germany; 2Universityof Maastricht, Department of Psychiatry andNeuropsychology, Maastricht, The Netherlands;3University of Wurzburg, Psychiatry Psychosomaticsand Psychotherapy, Wurzburg, Germany; 4Universityof Wurzburg, Laboratory for Microarray ApplicationsInterdisciplinary Centre for Clinical Research (IZKF),Wurzburg, Germany

Prenatal stress (PS) exposure in early life is anenvironmental risk factor that has been shown to affectfetal brain development and to increase the risk for adultpsychopathology. Furthermore, a length-polymorphismin the serotonin transporter gene (5-HTTLPR) hasbeen suggested to modulate the association betweenstress exposure in early life and the development ofpsychiatric disorders in later life by rendering short allelecarriers more vulnerable to stress. The exact molecularmechanisms underlying this gene–environment (G×E)interaction remain to be elucidated though.

Recently, using a maternal restraint stress paradigmof PS in wild-type (WT) and heterozygous (+/−)5-Htt deficient mice, we have shown that the long-term behavioural effects of PS are partly dependent onthe 5-Htt genotype [1]. In our study, 5-Htt+/− mice,particularly females, appeared to be more vulnerable to PSexposure when compared to WT offspring. Additionally,hippocampal gene expression profiles of the femalesindicated that the effects of the 5-Htt+/− genotype, PSexposure, and their interaction were mediated by distinctmolecular mechanisms. More specifically, MAPK andneurotrophin signalling were regulated by both the 5-Htt+/−

genotype and PS exposure, whereas cytokine and Wntsignalling were affected in a 5-Htt genotype × PS manner.Epigenetic mechanisms such as DNA methylation

and histone modifications build the interface betweenenvironment and genome and thus have been recentlyheavily discussed to play a role in the development ofpsychiatric disorders.In light of recent findings concerning the role of

epigenetic mechanism in the regulation of signallingpathways that mediate depression-like behaviour inrodents, the present study aims to examine the role of DNAmethylation in mediating the changes in gene expressionobserved in our 5-Htt×PS paradigm. For this purpose,genome-wide promoter methylation was assessed in thehippocampus using methylated DNA immunoprecipitationfollowed by Affymetrix Mouse Promoter 1.0R Arrayanalysis (MeDIP-on-chip). Probe signals from MeDIP andinput samples were normalised and MeDIP-input signallog2 ratios calculated. A sliding-window approach wasapplied to decrease the noise in the experiment readout.Genomic regions enriched by MeDIP were detected by theCMARRT algorithm.We found that the genotype, the PS exposure and an

interaction of both caused changes in the DNAmethylationof a number of genes that showed also differentialexpression in our previous study indicating a possiblecorrelation between DNAmethylation and gene expressionfor those genes. Among them were genes as Fgfr4,Map3k1, Nos1, Mbp1 and Cabin1. The promoter arrayresults are currently investigated in detail using bisulfitetreatment and pyrosequencing.In conclusion, hippocampal gene expression and

DNA-methylation profiles of female prenatally stressed5-Htt+/− mice suggest that distinct molecular mechanisms,including epigenetic programming, might mediate thebehavioural effects of the 5-Htt genotype, PS exposure,and their interaction.

Reference(s)

[1] Van den Hove DLA, Jakob SB, Schraut KG, Kenis G,Schmitt AG, Kneitz S, Scholz CJ, Wiescholleck V,


Ortega G, Prickaerts J, Steinbusch HWM, Lesch KP(2011) Differential effects of prenatal stress in 5-Httdeficient mice: towards molecular mechanisms of gene× environment interactions. PloS One 6:e22715.

P.3.010 Time-dependent effects ofantidepressant treatments on miRNomeexpression profile in hippocampus ofrats

M. Seguini1 °, D. Tardito1, A. Mallei1, G. Racagni1,M. Popoli1. 1University of Milan, Department ofPharmacological and Biomolecular Sciences, Milan,Italy

In the past few years, it has become clear that beyond thetraditional transcriptional mechanisms, gene expressionis also regulated by microRNAs (miRs), small noncoding RNAs with a key role in post-transcriptionalregulation of gene expression. MiRs have recently beenimplicated in a variety of human diseases, includingneuropsychiatric disorders. Moreover, a role for miRsin the pharmacotherapy of mood disorders has beenrecently suggested. It has been indeed shown that somemiRs and their effectors are modulated by the moodstabilisers lithium and valproate and that fluoxetine (FLX,a selective serotonin reuptake inhibitor-SSRI) modulatesmiR-16 expression [1,2]. Moreover, a recent clinical studyreported changes in blood expression of 30 miRs after12 weeks of treatment with the SSRI escitalopram indepressed patients [3]. Aim of the present study was toverify whether treatment for different time length (3, 7,14 days) with two antidepressants (ADs) characterisedby different primary mechanism of action (FLX, anddesipramine, DMI, a tricyclic AD with predominantlyaction on the noradrenaline reuptake) modulate miRNomeexpression in rat hippocampus.To this aim, total RNA including miRNAs was isolated

from each hemi-hippocampus and reverse transcribed.Quantitative Real Time PCR (qRT-PCR) amplification wascarried out using TaqMan Array rodent MicroRNA A+BCard Set v3.0 using the ddCT method.A total of about 450 miRs were detected in all samples.

The analysis of the miRNome expression showed evidencefor significant effects of AD treatments at the differenttime points assessed. After 3 days of treatment, FLXdown-regulated the expression of 8 miRs and DMI up-regulated the expression of 9 miRs. A more pronouncedeffect was found after 7 days of treatment; indeed, atthis time point FLX modulated 35 miRs (28 up-regulatedand 7 down-regulated), whereas DMI down-regulated theexpression of 15 miRs. Interestingly, 8 of them were

up-regulated by both ADs. Finally, after 14 days oftreatment FLX modulated the expression of 4 miRs (1 up-regulated and 4 down-regulated) and DMI down-regulatedthe expression of 18 miRs. A bioinformatic analysisallowed the identification of putative miRs targets andsignalling pathways modulated by the identified miRs.The predicted target genes of miRs are mainly involvedin pathways related to neuronal brain function, (such assynaptic and neuronal plasticity, neurotransmission, etc)and many of them have been previously associated toboth depression pathophysiology and to the mechanism ofaction of ADs.Following different criteria (i.e., genes involved in

neuroplasticity modulation and candidate genes for mooddisorders according to the more recent evidence available),a number of putative miR target genes have beenselected for validation studies by means of mRNA/proteinexpression studies.The results of this work, showing that AD treatments

induce early and significant modifications in the expressionof hippocampal miRs, could be of help to better clarifythe mode of the action of current ADs and to identify newputative targets for the development of novel drugs withgreater efficacy and a more rapid onset of action.

Reference(s)

[1] O’Connor R.M., Dinan T.G., Cryan J.F., 2012. Littlethings on which happiness depends: microRNAs asnovel therapeutic targets for the treatment of anxietyand depression. Mol Psychiatry 17(4), 359–376.

[2] Dwivedi Y., 2011. Evidence demonstrating role ofmicroRNAs in the etiopathology of major depression.J Chem Neuroanat. 42(2), 142–156.

[3] Bocchio-Chiavetto L., Maffioletti E., Bettinsoli P.,Giovannini C., Bignotti S., Tardito D., Corrada D.,Milanesi L., Gennarelli M., 2012. Blood microRNAchanges in depressed patients during antidepressanttreatment. Eur Neuropsychopharmacol Epub ahead ofprint.

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Clinical neuropsychopharmacology

Lectures

S.04.01 Disrupted cerebral connectivity andbrain disorders: linking circuits to genes

A. Meyer-Lindenberg1 °, H. Tost1, E. Bilek1. 1Universityof Heidelberg, Zentralinstitut fur Seelische GesundheitDepartment of Psychiatry and Psychotherapy, Mannheim,Germany

In the past decade, imaging genetics has evolved to ahighly successful neuroimaging discipline with a varietyof sophisticated research tools. To date, several neuralsystems mechanism have been identified that mediategenetic risk for chronic disabling mental conditionslinked to common candidate and genome-wide-supportedvariants. In particular, the examination of intermediateconnectivity phenotypes has recently gained increasingpopularity. We give an overview of the scientificmethods and evidence that link indices of neural networkorganisation to healthy human behaviour and the geneticsusceptibility for mental illness, following our recentreview [1].The human brain is conceptualised as a complex

network of functionally specialised cell populations thatinteract with each other in a spatially and temporallycoherent fashion, and shape physiological and pathologicalbehaviours, as well as their own coupling patterns, bymeans of neurochemical processes and interconnectingaxonal fibres. The anatomical properties of these networksare highly heritable, and shaped by a multitude of geneticvariants, some of them conferring risk for psychiatricdisorders by affecting the functional integrity of regulatorycircuits controlling complex behavioural phenotypes suchas cognitions and emotions. In the past decade, themassive technological advances in neuroimaging andmolecular genetics facilitated the implementation of anew research strategy (“imaging genetics”) that allowsfor the identification of the neurobiological effects ofsusceptibility genes.Historically, imaging genetics studies have focussed

on the analysis of single genetic variants that werechosen based on candidacy. Candidate gene approachesare hypothesis-driven, and follow a reverse mode ofexamination, as the genes of interest are pre-selected

based on prior evidence that may arise from variousempirical sources such as clinical association studies orbasic neuroscience research. A particularly noteworthycontribution to the functional imaging genetics field isthe work by Egan and coworkers [2], which examinedthe effects of a functional variant in the catechol-O-methyltransferase gene (COMT Val108/158Met, rs4680)on frontal lobe function in patients with schizophrenia,their unaffected siblings, and healthy volunteers. Inall three groups, the carriers of the high dopaminecatabolising variant (Val/Val) were characterised by greateractivity in the prefrontal cortex during working memoryperformance. However, while thousands of imaginggenetics studies have been published to date, only fewreports in psychiatric neuroimaging aimed to characterisethe neural correlates of interaction effects within orbetween genes. In own prior work, we established amethod to extend the reach of imaging genetics to theevaluation of the effects of ambiguous haplotypes onprefrontal cortex function and structure. By applying thismethod to COMT, these analyses showed that the neuraleffects of the Val108/158Met polymorphism are modifiedby other functional variants and haplotypes in COMT in apattern that is consistent with complex non-linear effectsof extracellular dopamine on prefrontal cortex efficiencyand structural integrity. In addition, using fMRI workingmemory data, several other studies provided biologicalvalidation of previously reported effects of gene epistasison risk for schizophrenia (see e.g. the first report of athree-way interaction by Nicodemus and coworkers [3]).In other studies, significant interaction effects betweenCOMT Val108/158Met and other dopamine or glutamateregulating variants such as AKT1 on prefrontal cortexfunction were demonstrated.While candidate gene approaches are facing mounting

criticism in imaging genetics due to the limited evidencefor an association of the variants with the disease pheno-type itself, the investigation of risk variants identified ingenome-wide association studies at appropriate statisticalthresholds has gained much popularity in the recentpast. A particularly interesting case is a genome-widesignificant single nucleotide polymorphism (rs1344706A)in the gene encoding for the zinc finger protein804A (ZNF804A), a genetic variant conferring risk forschizophrenia and possibly bipolar disorder. Using a well-established working memory paradigm, we have recently

S68 Clinical neuropsychopharmacology

demonstrated that healthy carriers of rs1344706 riskgenotypes do not exhibit differences in regional activationper se, but display pronounced gene dosage dependentalterations in the functional coupling of the DLPFC andhippocampus a finding that mirrors the coupling deficitsobserved in schizophrenia patients). Subsequent analysesprovided evidence for task-specific effects of rs1344706Aon prefrontal–hippocampal circuit interaction, therebypointing to a domain-specific configuration of the geneticrisk architecture of schizophrenia. Very recent work bythe Weinberger group has further extended these resultsand provided evidence for this connectivity feature as anintermediate phenotype for schizophrenia.On a more complex level, the structural and functional

composition of neural circuits can be described withgraph theoretical metrics. In graph theory, the topologyof large-scale cortical networks can be quantified by agraph consisting of a number of nodes (i.e., brain regions)that are connected by edges (i.e., interconnecting fibretracts or indices of structural or functional coupling).These phenotypes have been shown to be heritable,and the contribution of individual genetic variants tothese topological features should be examined. Also, theconceptual expansion of imaging genetics to non-geneticrisk factors such as social status and urban environmentwill further expand our knowledge on the systems-level mechanisms of psychiatric disease. Ultimately, theseefforts are expected to pioneer a new generation ofevidence-based diagnosis and treatment strategies for thesechronic disabling mental conditions.

Reference(s)

[1] Tost, H., E. Bilek, Meyer-Lindenberg, A., 2009.Brain Connectivity in Psychiatric Imaging Genetics.Neuroimage 62, 2250−60.

[2] Egan, M.F., T.E. Goldberg, B.S. Kolachana, J.H. Cal-licott, C.M. Mazzanti, R.E. Straub, D. Goldman, andD.R. Weinberger. Effect of Comt Val108/158 MetGenotype on Frontal Lobe Function and Risk forSchizophrenia. [In eng]. Proc Natl Acad Sci U S A98, no. 12 (Jun 5 2001): 6917−22.

[3] Nicodemus, K.K., J.H. Callicott, R.G. Higier, A. Luna,D.C. Nixon, B.K. Lipska, R. Vakkalanka, et al.Evidence of Statistical Epistasis between Disc1,Cit and Ndel1 Impacting Risk for Schizophrenia:Biological Validation with Functional Neuroimaging.[In eng]. Hum Genet 127, no. 4 (Apr 2010): 441−52.

Disclosure statement: Financed by grants from GermanResearch Council, Federal Ministry of Science, EuropeanUnion and NARSAD

Posters

P.4.001 Clinical implications of polarity indexof drugs in maintenance treatment ofbipolar disorder: a naturalistic study

D. Popovic1 °, E. Vieta1. 1Hospital Clinic de Barcelona,Neuroscience, Barcelona, Spain

Purpose of the study: Predominant polarity, defined as atleast twice as many episodes of one pole over the other,is among the strongest predictors of recurrence into aspecific episode in Bipolar Disorder (BD), and should beconsidered when implementing maintenance therapy. Ourgroup has recently developed the Polarity Index (PI), ametric indicating antimanic versus antidepressive potentialof drugs [1]. The purpose of this study was to determinethe role of PI in clinical decision-making. Secondary aimwas to assess differences between predominantly manicand depressed patients, with a special focus on theirpharmacological treatment.

Methods: The study sample was composed of 604patients aged �18, with BD I or II, who signed aninformed consent, enrolled in the systematic prospectivefollow up study of the Bipolar Disorders Program of theHospital Clinic of Barcelona, Spain. Patients who fulfilledcriteria for either Manic (MPP) or depressive (DPP)were compared regarding socio-demographic, clinical andtherapeutic characteristics.The PI, a numeric expression of the efficacy profile of a

given drug, derives from Number Needed to Treat (NNT)for prevention of depression and NNT for prevention ofmania ratio, as emerging from the results of randomisedplacebo-controlled trials [1,2]. Drugs with PI> 1 havestronger antimanic prophylactic properties, while thosewith PI< 1 are more effective for preventing depressiveepisodes than the manic ones. The PI of drugs formaintenance treatment of BD was: 12.09 for risperidone,4.38 for aripiprazole, 3.91 for ziprasidone, 2.98 forolanzapine, 1.39 for lithium, 1.14 for quetiapine, and0.40 for lamotrigine [1]. PI of valproate (0.49) andoxcarbazepine (0.62) may not be reliable due to thefailure of their maintenance trials and therefore were notaccounted for.PI for patients’ current treatment was calculated as mean

value of all prescribed drugs in each patient.Results: 257/604 (43%) of patients with BD I or II

presented predominant polarity. 143 patients (55.6%)fulfilled criteria for DPP and 114 (44.4%) for MPP.Total PI, as well as Antipsychotics’ PI and Mood

Stabilisers’ PI were higher, indicating a stronger antimanicaction, in MPP (Table 1).

Clinical neuropsychopharmacology S69

MPP presented higher prevalence of BD I, male gender,younger age, age at onset and at first hospitalisation, morehospitalisations, primary substance misuse and psychoticsymptoms. DP correlated with BD II, depressive onset,primary life events, melancholia and suicide attempts. Theprescription of first generation antipsychotics and secondgeneration antipsychotics olanzapine and risperidone wassignificantly more frequent among MPP patients, whilstuse of lamotrigine, selective serotonin reuptake inhibitors,serotonin–norepinephrine reuptake inhibitors, tricyclicantidepressants and benzodiazepines was more prevalentamongst DPP patients.

Conclusions: The results of this naturalistic studyconfirm the usefulness of the PI. In this large sample,clinical differences among these groups justify differentialtreatment approach. The PI appears to be a useful way tooperationalise what clinicians do for maintenance therapyin BD.Table 1.

Polarity Index

Manic(n = 114)

Depressive(n = 143)

p MWU

p KSZ

p

Mean SD Mean SD

AP+MS 3.68 3.19 2.22 2.36 0.001* 3385.5 0.000* 2.069 0.000*AP 6.78 4.68 4.77 4.53 0.044* 1281.5 0.006* 1.425 0.035*MS 1.31 0.23 1.14 0.38 0.001* 2679.0 0.001* 1.429 0.034*

AP = Antipsychotics, MS = Mood Stabilisers, MW = Mann–Whitney, KS =Kolmogorov–Smirnov.

Reference(s)

[1] Popovic, D., Reinares, M., Goikolea, J.M., Bon-nin, C.M., Gonzalez-Pinto, A., Vieta, E., 2012.Polarity index of pharmacological agents used formaintenance treatment of bipolar disorder. EurNeuropsychopharmacol. 22(5), 339–346.

[2] Popovic, D., Reinares, M., Amann, B., Salamero, M.,Vieta, E., 2011. Number needed to treat analysesof drugs used for maintenance treatment of bipolardisorder. Psychopharmacology (Berl). 213(4), 657–667.

P.4.002 Genetic differences in drug-metabolisingenzymes: can they be used to predictantidepressant treatment response?

K. Hodgson1 °, K.E. Tansey1, R. Uher1, O.S.P. Daivs1,K.J. Aitchison1, P. McGuffin1. 1King’s College London,MRC SGDP Institute of Psychiatry, London, UnitedKingdom

Purpose of the study: Response to antidepressantmedication varies greatly between patients [1]. One factorwhich may contribute to this variability is the rate at

which the drugs are metabolised into inactive compounds,a process performed by the cytochrome P450 family ofenzymes. Common genetic differences have been observedin two of the key enzymes (CYP2D6 and CYP2C19)involved in the metabolism of many antidepressants, andthese polymorphisms have been linked to variation inrates of drug metabolism. This study examines whethercirculating serum levels of antidepressant are influencedby cytochrome P450 genotype in depressed patients and,furthermore, whether treatment outcomes can be predictedby either cytochrome P450 genotype or circulating levelsof antidepressant.

Methods used: This study used data from GENDEP(Genome-based Therapeutic Drugs for Depression),a multi-centre pharmacogenetic project following a largesample of depressed patients who received one oftwo antidepressants for a period of twelve weeks.The two antidepressants studied were the selectiveserotonin reuptake inhibitor, escitalopram and the tricyclicantidepressant, nortriptyline. These drugs are metabolisedinto inactive compounds via different pathways, andso analyses were performed separately for each drug.Circulating serum levels of antidepressant (escitalopramor nortriptyline) and primary metabolite (desmethylcitalo-pram or 10-hydroxy-nortriptyline) were measured aftereight weeks of treatment using achiral high-performanceliquid chromatography. Data was available from 560patients (314 were receiving escitalopram and 246 werereceiving nortriptyline). Common variants in the genesencoding the cytochrome P450 enzymes CYP2D6 andCYP2C19 were genotyped using the micro-array basedRoche AmpliChip CYP450 assay. Treatment response wasmeasured using percentage change from baseline on theMontgomery-Asberg Depression Rating Scale (MADRS).Information was also available regarding other medicationthat patients were taking during the trial, so the effectof cytochrome P450 enzyme inhibiting medications couldalso be considered.

Summary of results: Cytochrome P450 genotype pre-dicted circulating levels of both escitalopram (b = −4.23,95%CI = −5.67 to −2.79, p = 8.56×10−9) and nortriptyline(b = −32.18, 95%CI = −45.27 to −19.08, p = 1.46×10−6),as well as the primary metabolite of nortriptyline, 10-hydroxy-nortriptyline (b = 16.60, 95%CI = 5.36 to 27.84,p = 0.0038). Other medications known to have an effecton the cytochrome P450 enzymes that were being takenconcurrently with the antidepressants had no impact onthis relationship. Nonetheless, in this sample, cytochromeP450-inhibiting comedication predominantly consisted ofthe use of oral contraceptives, which are weak inhibitors ofthe drug-metabolising enzymes. Neither cytochrome P450genotype, nor circulating levels of drug were predictive ofresponse to antidepressant treatment.


Conclusions: This study investigated the effect ofgenetic variation in cytochrome P450 genotype on rates ofantidepressant metabolism, and the potential relationshipto treatment outcomes for two drugs; escitalopram andnortriptyline. These results suggest that, whilst geneticvariation in cytochrome P450 enzymes does influencecirculating levels of antidepressant for both of these drugs,the variability that is seen in response to treatment isnot the result of individual differences in rates of drugmetabolism.

Reference(s)

[1] Trivedi MH, Rush AJ, Wisniewski SR, Nieren-berg AA, Warden D, Ritz L et al. Evaluationof outcomes with citalopram for depression usingmeasurement-based care in STAR*D: implications forclinical practice. Am J Psychiatry 2006; 163: 28−40.

P.4.003 Publication pressure and burn outamong Dutch medical professors:a nationwide survey

J. Tijdink1 °, Y.M. Smulders1, A.C.M. Vergouwen2.1Free University Medical Centre (VUmc), InternalMedicine/Psychiatrist, Amsterdam, The Netherlands;2St. Lucas Andreas Hospital, Psychiatry, Amsterdam, TheNetherlands

Background: Publication of scientific research papers isimportant for professionals working in academic medicalcentres and is a key activity in academia. There appearsto be increasing focus on quantitative output measures,where quantitative measures of scientific output determinestatus and prestige, and serve to rank universities as wellas individuals. Excessive emphasis on Impact Factors andHirsch indices could therefore generate a type of pressurethat adversely influences quality of science and personalwell-being of scientists [1−3].How this pressure influences professionals’ perception

of science and their personal well-being is unknown.This manuscript addresses publication culture and itsconsequences for science and personal well-being in theNetherlands, as seen through the eyes of our academicleaders.

Methods: We performed an online survey inviting allmedical professors (n = 1206) of the 8 academic medicalcentres in The Netherlands to participate. They were askedto fill out 2 questionnaires, besides their demographicdata; a Scientific Culture Questionnaire and the MaslachBurnout Inventory. The scientific culture questionnairecontained 24 statements, the responses to which werescored on a 5-point Likert scale. The statements of the

scientific culture questionnaire was divided into 3 broaddomains; (1) pressure to publish personally experienced bythe respondent; (2) publication pressure in general terms inthe academic work place, as perceived by the respondent;(3) publication pressure relating to the scientist’s positionand status (e.g. promotion, re-appointment, etc.).

Results: In total, 437 professors completed thequestionnaires. Among them, 54% judge that publicationpressure ‘has become excessive’, 39% believe thatpublication pressure ‘affects the credibility of medicalresearch’ and 26% judge that publication pressure hasa ‘sickening effect on medical science’. The burn outquestionnaire indicates that 24% of medical professorshas signs of burn out. All 3 subdomains of the publicationculture questionnaire were strongly and significantlyassociated with scores on all 3 subscales of the burnoutquestionnaire. Correlations were generally strongest forthe emotional exhaustion subscale.

Conclusion: A substantial proportion of medicalprofessors believe that publication pressure has becomeexcessive, and are developing a cynical view on the validityof medical science. Publication pressure may have adverseeffects on medical science. In the respondents’ opinion,publication pressure can adversely affect validity andreliability of the medical literature. These perceptions arestatistically correlated to emotional exhaustion and otherburn out symptoms. The main limitation is the possibilityof participation bias: Those experiencing high pressurecould either preferentially participate or be reluctant todo so. Among those who participated, there may stillbe a taboo on personal pressure and burn out, causingrespondents to downplay the severity and personal impactof publication pressure. Further research should addressthe effects of publication pressure in more detail andidentify alternative ways to stimulate the quality of medicalscience.

Reference(s)

[1] Fanelli D. Do pressures to publish increase scientists’bias? An empirical support from US States Data. PLoSOne. 2010 5(4): e10271.

[2] West CP, Hubschka MM, Novotny PJ, Sloan JA,Kolars JC, Habermann TM, et al. Association ofperceived medical errors with resident distress andempathy. JAMA. 2006 296(9):1071–1078.

[3] Young NS, Ioannidis JPA, Al-Ubaydli O. Why currentpublication practices may distort science. PLoS Med2008 5(10): e201.


P.4.004 Early onset of lithium prophylaxis aspossible good prognostic factor forstaging bipolar disorder

S. Brioschi1 °, L. Franchini1, C. Colombo1, E. Smeraldi1.1Ospedale San Raffaele, Neuroscienze Cliniche, Milano,Italy

A previous study [1] conducted at our centre, the Clinic forMood Disorders of S. Raffaele Hospital in Milan, reportedthat beginning lithium therapy within the first ten years ofillness predicts better preventive outcomes than beginningprophylaxis later, both in major depression, recurrent andbipolar patients. The aim of the present study was toconfirm these results only considering bipolar patients,and to evaluate clinical markers that may be associatedto the response to the stabilisation therapy. In particular,we analysed the role of the initiation time of maintenancetherapy on the control of recurrence, and, therefore, of theoutcome of Bipolar Disorder.Two hundred fourteen subjects (84 males, 130 females)

affected by Bipolar Disorder receiving a stabilisationtherapy (lithium salts, anticonvulsants, or antipsychotics)were studied. Patients were recruited from the ambulatorycentre of the Mood Disorder Unit of S. Raffaele Hospitalin Milan. Clinical data were collected interviewing the pa-tients through the NIMH Life Chart method. We recordedthe time of onset of the preventive therapy and dividedthe sample into three groups: an ‘Early group’, includingpatients who initiated the preventive therapy within the 5thyear from the onset of illness; a ‘Late group’, includingpatients who initiated the preventive therapy between the5th and 10th year from the onset of illness; and a ‘VeryLate group’, including patients who initiated the preventivetherapy after the 10th year from the onset of illness. Theefficacy of the preventive therapy was evaluated calculatingthe gradient between the recurrence index before andafter starting this treatment. Multiple logistic regressionanalysis was used to determine the factors that influencethe response to the preventive therapy.Patients received, as main stabilisation treatment,

lithium salts (65%). The percentage of responders wasof 77% during a maintenance treatment period of4.5 years. The only variables significantly associatedwith the outcome of preventive therapy and, therefore,with the control of the illness progression were: the useof lithium salts as a first treatment choice (P = 0.02),starting the preventive therapy within 5 years of the illnessonset (P< 0.0001), and the high recurrence index of theillness before treatment (P< 0.0001). The presence ofpsychotic manifestations turned out to be the only factorthat negatively influenced the response to the preventivetherapy (P = 0.03).

The present study confirmed the importance of an earlyintervention in Bipolar Disorder, indicating that startinglithium therapy within the first five years of illness ismore effective than treatments delivered later in the illnesscourse. Finally, our data suggest that the onset time oflithium therapy is a new prognostic element; indeed, anearly onset could improve patient’s long-term prognosis,while a late onset is associated with a progression ofthe Bipolar Disorder. Referring to the staging modelsproposed recently [2], we suggest that the time of initiationof maintenance therapy is a clinic crucial informationfor staging patients with Bipolar Disorder and, therefore,giving an appropriate preventive treatment.

Reference(s)

[1] Franchini, L., Zanardi, R., Smeraldi, E., Gasperini, M.,1999. Early onset of lithium prophylaxis as a predictorof good long-term out come. Eur Arch Psychiatry ClinNeurosci 249, 227–230.

[2] Vieta, E., Reinaires, M., Rosa, A.R., 2011. Stagingbipolar disorder. Neurotox Res 19(2), 279−85.

P.4.005 Effects of cigarette smoking onschizophrenia treatment with olanzapine

K. Zoric1 °, N. Zivkovic1, G. Djokic2. 1NeuropsychiatryHospital “Dr Laza Lazarevic”, acute male psychosis(M department), Belgrade, Serbia; 2NeuropsychiatryHospital “Dr Laza Lazarevic”, Department of ScientificResearch, Belgrade, Serbia

Introduction: Co-morbidity of schizophrenia andcigarette smoking is a well-known phenomenon. Patientswith schizophrenia have elevated rates of cigarettesmoking compared to the general population, and greatdifficulty in smoking cessation. Smoking rates (78−88%)in patients with schizophrenia are much higher than thenormal population. Excessive cigarette smoking, as itwell known, may reduce plasma levels of antipsychotics,including olanzapine up to 50%.

Objective: The goal of this study was to investigate ef-fects of smoking in olanzapine treatment of schizophreniaand to determine does the number of smoked cigarettesduring the day affect olanzapine efficacy.

Methods: This prospective clinical study included 127patients (57.4% males, 42.6% females) with schizophreniadiagnosed by ICD-10 criteria, aged 18−51, which weredivided into four groups. Control group (39 patients) andexperimental groups: group E1, 1−20 cigarettes/24 h (34patients), group E2, 21−40 cigarettes/24 h (28 patients)and group E3, more than 40 cigarettes/24 h (26 patients).Patients were observed for one year period in hospital


and out hospital conditions (as outpatients). All patientswere assessed by Positive and Negative Symptom ScheduleScale (PANSS) at baseline, after six months and afterone year. Data were processed with SPSS for Windows.Kruskal–Wallis and Mann–Whitney tests for independentsamples and Wilcoxon test for relationed samples, wereused in statistical analyses.

Results: No significant differences were found amongfour groups for PANSS score at baseline (p = 0.515).PANSS score reduction at study endpoint, after one year,is statistically significant in all four groups (p = 0.001).Rate of PANSS score reduction after one year ishighest in non-smokers group 63.5%, and lowest ingroup of heavy smokers (40+ cigarettes/24 h) 40.8%.There is a statistically significant difference in PANSSscore reduction between group of non-smokers and allthree smokers groups (p< 0.001). There is a statisticallysignificant difference in PANSS score reduction betweenE1 and E2, and E1 and E3 group (p< 0.001). There was nodifference in PANSS score between group E2 and group E3after one year (p = 0.384). There is a statistically significantdifference in olanzapine average daily dose after one yearbetween group of non-smokers and all three smokersgroups (p< 0.001). There is a statistically significantdifference in olanzapine average daily dose between E1and E2 (p = 0.053), and E1 and E3 group (p< 0.001). Therewas no statistically significant difference in olanzapineaverage daily dose between group E2 and group E3 afterone year (p = 0.179).

Conclusions: The results of our study suggest thatcigarette smoking have significant effect on olanzapinetherapy of schizophrenia. Olanzapine therapy is mostefficient in non-smokers group with most prominentPANSS score reduction. Excessive smoking more than20 cigarettes per day significantly reduces effects ofolanzapine therapy in schizophrenia. Average daily doseof olanzapine is significantly lower in non-smokers groupthan in all three groups of smokers.

Reference(s)

[1] Haselmo, T., Eikeseth, P., Tanum, L., Molden, E.,Refsum, H., 2006. The effect of variable cigaretteconsumption on the interaction with clozapine andolanzapine. European Journal of Clinical Pharmacol-ogy 62 (12): 1049–1053.

[2] Lowe, E.J., Ackman, M.L., 2010. Impact on tobaccosmoking cessation on stable clozapine or olanzapinetreatment. Ann Pharmacother 44 (4): 727–732.

[3] Cole, M.L., Trigoboff, E., Demler, T.L., Opler, L.A.,2010. Imapact of smoking cessation on psychiatricin patients treated with clozapine or olanzapine. JPsychiatr Pract 16 (2): 75−81.

P.4.006 Reward processing in unaffectedsiblings of schizophrenia patients:a functional magnetic resonanceimaging study

M. de Leeuw1 °, M. Vink1, R.S. Kahn1. 1RudolfMagnus Institute of Neuroscience, Psychiatry, Utrecht,The Netherlands

Purpose of the study: Schizophrenia is characterised bya failure in dopaminergic signal transduction. Rewardprocessing is dependent on adequate dopaminergictransduction in mesolimic and mesocortical pathwaysand may therefore be affected in schizophrenia [1,2].Reward processing can be dived into two sub-processes:anticipation and the outcome of reward, whereas rewardanticipation is thought to be primarily mediated bydopaminergic neurons. Despite abnormal dopamine func-tion in schizophrenia, functional MRI studies in patientsduring reward anticipation have shown conflicting results:hypo- and hyperactivation of the ventral striatum have beenreported. This inconsistency may be due to several (illness-related) factors of which the use of dopamine antagonists isprobably the most important one. The goal of the currentstudy is to investigate reward (anticipation) processingusing fMRI in siblings of schizophrenia patients who shareon average 50% of their ill siblings’ genes without havingillness-related confounds such as the use of medication.

Methods used: Twenty-nine unaffected siblings ofschizophrenia patients and 29 matched controls performeda monetary delayed incentive task during scanning. Thetask consisted of 72 trials, each lasting 6 seconds onaverage (range 3−10 s). At the beginning of each trial,a cue was presented signalling a potentially rewarding (asmiling face, ‘reward anticipation’) or nonrewarding (anon-smiling face, ‘neutral anticipation’) trial. Followingthis cue, a target (exclamation mark) was presented towhich subjects had to respond by pressing a button as fastas possible. Feedback on performance was given (either areward or no reward). Subjects could win €1 when theyresponded within the time limit. Subjects were rewarded inonly 50% of the reward trials. fMRI scans were acquiredon a 3.0 T scanner using a 2D-EPI sequence (TR: 1600ms,TE: 23ms) and preprocessed and analysed using SPM5.

Summary of results: Siblings and controls earnedthe same amount (€15). Despite this equal performance,siblings showed hypoactivation of the right anterior insuladuring the reward anticipation (reward anticipation minusneutral anticipation). An ROI analysis, with 11 predefinedregions known to be involved in reward processing,revealed hypoactivation bilaterally in the ventral striatumin siblings compared with controls. During the outcome ofreward (feedback reward minus feedback neutral) the ROI


analysis showed hyperactivation bilaterally in the oribitalfrontal cortices (OFC) in siblings compared to controls.

Conclusions: Our finding of ventral striatum hypoacti-vation during reward anticipation and OFC hyperactivationduring reward outcome in siblings is consistent with thenotion of impaired dopamine functioning in the fronto-striatal network typically associated with schizophrenia.Moreover, this study supplies further evidence thatdeficiencies in fronto-striatal pathways mediated bydopminargic signal transmission may be a core deficiencyunderlying cognitive processes in schizophrenia [3]. Futureexperiments should clarify what genes may be responsiblefor these phenotypic anomalies.

Reference(s)

[1] Ziauddeen, H. & Murray, G.K., 2010. The relevanceof reward pathways for schizophrenia. Current opinionin Psychiatry 23, 91−6.

[2] Barch, D.M. & Dowd, E.C., 2010. Goal represen-tations and motivational drive in schizophrenia: therole of prefrontal–striatal interactions. SchizophreniaBulletin 36, 919−34.

[3] Vink, M., Ramsey, N.F., Raemaekers, M., Kahn, R.S.,2006. Striatal dysfunction in schizophrenia andunaffected relatives. Biological Psychiatry 60, 32−9.

P.4.007 Associations of antiepileptic drugs withanxiety and depressive symptoms inpaediatric epilepsy: a pilot study

D. Stevanovic1 °, J. Jancic2. 1General Hospital Sombor,Department of Psychiatry, Sombor, Serbia; 2Clinicfor Neurology and Psychiatry for Children and YouthUniversity of Belgrade School of Medicine Serbia,Department of Neurology, Belgrade, Serbia

Purpose: Psychopathology in epilepsy is generally consid-ered either to be caused by the epilepsy (ictal, post-ictal orinter-ictal disorders) or to be a comorbidity [1]. Although itis widely accepted that psychopathological manifestationsin a person with epilepsy are multifactorial − even a two-way relationship between epilepsy and psychopathologyhas been proposed, the role of antiepileptic drugs(AEDs) has attracted significant research attention.This has led to the discovery of both beneficial andadverse effects of AEDs on emotion, cognition, andbehaviour [2]. Different mechanisms of action havebeen proposed for the psychotropic effects of AEDs.In particular, the enhancement of gamma-aminobutyricacid (GABA) inhibitory neurotransmission has sedating/anxiolytic/depressiogenic effects, and blocking glutamateexcitatory neurotransmission has activating/anxiogenic/antidepressive effects [2]. Studies of psychopathology

in paediatric epilepsy in relation to AEDs are scarce,especially with respect to specific symptoms such asanxiety and depressive symptoms. The aim of this studywas to investigate associations of AEDs with anxiety anddepressive symptoms among children with epilepsy.

Methods: The present analysis was performed ondata from 54 children with epilepsy (mean age14.06±2.74 years) who had participated in our previousstudy [3]. Anxiety symptoms were assessed using theScreen for Child Anxiety Related Emotional Disorders(SCARED), while depressive symptoms were assessed us-ing the Mood and Feeling Questionnaire (MFQ). Consid-ering that psychopathology in epilepsy emanates from aninterplay between various factors, we created a generalisedlinear model to evaluate associations of AEDs with anxietyand depressive symptoms while taking into account othermajor factors as well (i.e., age, type of epilepsy, seizurefrequency, and epilepsy duration). Separate models weretested for anxiety and depressive symptoms.

Results: Nine children were taking carbamazepine(CBZ), 25 valproate (VPA), 5 lamotrigine (LTG), and15 were taking two or three AEDs (VPA, LTG, CBZ,oxcarbamazepine, primidone, clobazam, and clonazepam).Age, type of epilepsy, seizure frequency, epilepsy duration,AED, depressive symptoms, and interaction among thesevariables accounted for 66.9% of the variations inanxiety symptoms (F = 6.99, p< 0.0001). The same set ofvariables, with anxiety instead of depressive symptoms,accounted for 60.7% of the variations in depressivesymptoms (F = 5.28, p< 0.0001). Only taking two/threeAEDs was associated with significantly lowered anxietysymptoms (t = −3.06, p = 0.004), accounting for 18.6% ofadditional variance.

Conclusions: Although it has previously been reportedthat VPA could be associated with increased depressivesymptoms and LTG with increased anxiety symptoms [2],this preliminary study found no association between CBZ,VPA or LTG monotherapy and either type of symptomsamong children with epilepsy. However, taking two orthree AEDs was associated with significantly loweredanxiety symptoms; this could be an indirect effect ofgreater sedation by AEDs enhancing GABA inhibitoryneurotransmission.Considering that this cross-sectional study included a

small number of children, data from prospective studiesincluding more children − especially with newly diagnosedepilepsy − are needed to investigate all associations ofAEDs with anxiety and depressive symptoms.

Reference(s)

[1] Krishnamoorthy, E.S., Trimble, M.R., Blumer, D.2007. The classification of neuropsychiatric disordersin epilepsy: a proposal by the ILAE Commission


on Psychobiology of Epilepsy. Epilepsy behav 10,349−53.

[2] Mula, M., Monaco, F. 2009. Antiepileptic drugs andpsychopathology of epilepsy: an update. EpilepticDisord 11, 1−9.

[3] Stevanovic, D., Jancic, J., Lakic, A. 2011. The impactof depression and anxiety disorder symptoms onthe health-related quality of life of children andadolescents with epilepsy. Epilepsia 52, e75−8.

P.4.008 Lithium-regulated genes inlymphoblastoid cells from bipolar-affective patients

S. Kittel-Schneider1 °, M. Hilscher1, S. Schreck1,C.J. Scholz1, A. Reif1. 1University of Wurzburg, Deptof Psychiatry, Wurzburg, Germany

Introduction: Bipolar affective disorder affects up to 1%of the population worldwide. The underlying molecularpathomechanisms however still remain unclear and theexact mode of action of the current pharmacologicaltreatment options is still elusive. Lithium still is consideredto be the first-line mood stabiliser. However, responseis not uniform across the board, as only ca. 10%of all bipolar patients can be considered full lithiumresponders [1]. Unfortunately, there is no possibility topredict the lithium response in the individual patient beforeinitiating treatment. The aim of our study was thereforeto investigate molecular mechanisms of lithium action bystudying the expressional profile of lymphoblastoid celllines generated from bipolar patients.

Methods: EBV-immortalised lymphoblasts from bipo-lar patients (n = 10) and healthy controls (n = 10) wereincubated with either lithium chloride or vehicle (LiCl,c = 1mMol/l) for 3 weeks. After 1, 2 and 3 weeks oftreatment cells were harvested and mRNA was isolated.We conducted a microarray study to detect expressionalchanges upon lithium treatment. Next, we selected themost promising differentially regulated genes in termsof lithium-associated or disorder-associated pathways onthe basis of the microarray results. Also, we added‘circadian genes’ based on literature and peripheral geneexpression profiles (http://www.ebi.ac.uk/arrayexpress/).The microarray results were then verified via quantitativereal time PCR. The PCR results were normalised againsttwo housekeeping genes (HPRT1, TBP1) using LinRegand GeNorm software. The statistical analysis of the datawas carried out using SPSS statistical software.

Results: AIF1 and ANP32E expression were decreasedafter treatment for 3 weeks in bipolar patients. EPHB1expression was reduced in bipolar patients on baselinelevel compared to healthy controls, but not influenced by

lithium-treatment. PLEKHA2 expression was significantlyincreased after 3 weeks of lithium incubation, yet moreprominently in bipolar patients. KCNK1 showed a lowerbaseline level in the bipolar cells and lithium treatmentlead to an increase of the expression. Regarding the geneswhich are part of the molecular clockwork, DBP wassignificantly decreased in expression by lithium treatment,less prominent expressional changes were seen in PER1,PER2 and NR1D2.

Discussion: These results suggest that there is acomplex interaction of immuno-modulatory effects (AIF1)of lithium and effects regarding ion channels (KCNK1).Furthermore, genes which seem to play a role in synapto-(ANP32E) und neurogenesis (EPHB1) were identified aslithium-regulated in this peripheral cell model. Addition-ally, we could show that lithium regulates the expression ofgenes of the molecular circadian clockwork (DBP, PER1,PER2 and NR1D2), which are in part regulated via GSK-3beta. In summary, we describe interesting new lithium-regulated genes and we could also identify differencesin baseline gene expression between bipolar patient andhealthy control cells. These findings could prove useful inthe future development of biomarkers for diagnosis andtreatment response in bipolar affective disorder.

Reference(s)

[1] Kessing, L.V., et al., 2011, Valproate v. lithium inthe treatment of bipolar disorder in clinical practice:observational nationwide register-based cohort study.Br J Psychiatry. 199(1): p. 57−63.

P.4.009 Age at onset and cognitive impairmentin schizophrenia: an ecological cross-sectional study with stabilised patients

A. Caldiroli1 °, M. Buoli1, E. Caletti1, R.A. Paoli1,S. Zago2, A.C. Altamura1. 1Ospedale MaggiorePoliclinico Fondazione IRCCS Ca’ Granda, Psychiatry,Milano, Italy; 2Ospedale Maggiore Policlinico FondazioneIRCCS Ca’ Granda, Neurosciences, Milano, Italy

Purpose of the study: It is widely accepted that schizo-phrenic patients have global cognitive impairment whichis influenced by different clinical variables [1]. Amongthem, a longer duration of untreated psychosis (DUP)/duration of illness (DUI) and substance abuse show moreevidence to be involved in cognitive deficits; longer DUIwas found to correlate with impairment on psychomotorprocessing speed, verbal fluency and verbal learning [2].Furthermore, an interesting study has shown a relationshipbetween an early age at onset and pronounced deficits onDigit Symbol Coding and Tower of London [3]. Purpose ofthe present study is to detect if some clinical variables can


be predictive of cognitive impairment in schizophrenicswith particular attention on age at onset.

Methods: Thirty-five clinically stabilised schizophrenicpatients (twenty-nine males and six females) wererecruited from community services afferent to theDepartment of Psychiatry (University of Milan). Allpatients included into the study had to be diagnosed asschizophrenics by SCID-I, to show a Positive and NegativeSyndrome Scale (PANSS) score <50 and to be treated withan antipsychotic monotherapy or a combined antipsychotictreatment. They were assessed by the Brief Assessmentof Cognition in Schizophrenia (BACS) and an executive/social cognition battery including the Multiple ErrandsTest for Use in Hospital Settings (MET-HV), the HotelTask, the Iowa Gambling task, the Reading The Mind inthe Eyes Test and the Faux Pas Test. Informations aboutclinical variables derived from clinical charts and from in-terviews with the patients and their relatives. Binary logis-tic models were performed to find an eventual associationbetween continuous clinical variables and cognitive testfailures. In addition the total sample was divided in groupsaccording to dichotomous variables (gender, diagnosticsub-type and substance abuse) and the presence of cogni-tive deficits was compared between groups by c2 tests.

Summary of results: An earlier age at onset was foundto be predictive of frontal cognitive impairment (Tower ofLondon p = 0.022, OR= 0.709). In addition female genderwas more probably associated with mistakes at MET-HV(c2 = 4.80, p = 0.05, phi = 0.40) and Hotel Task (c2 = 5.25,p = 0.04, phi = 0.4) than male one. Finally, cannabisabusers showed more frequently deficits on verbal fluency(c2 = 9.35, p = 0.04, phi = 0.52) and executive functioning(Tower of London) (c2 = 11.67, p = 0.02, phi = 0.58) thanalcohol/cocaine abusers.

Conclusions: Female patients with an early age at onsetand cannabis abuse seem to have the worst cognitiveprofile among schizophrenics. These data illustrate theimportance of age at onset on the course of thedisorder, as happens for other psychiatric disorders (e.g.obsessive–compulsive disorder) and confirm that earlyonset population need a higher intensity of care in lightof the more severe cognitive impairment. Limits of thestudy include the small sample size and the differences inthe pharmacological treatment between the patients, whichcould be confounding factors in the interpretation of thedata. Prospective studies with larger samples are warrantedto confirm the result of this naturalistic study.

Reference(s)

[1] Gur, R., 2011. Neuropsychiatric aspects of schizo-phrenia. CNS Neuroscience & Therapeutics 17,45−51.

[2] Bajs, M., Janovic, S., Bajs, M., Dordervic, V., Jevtovic,S., Radonic, E., Kalember, P., 2011. Correlation ofcognitive functions with some aspects of illness, treat-ment and social functioning in recurrently hospitalizedschizophrenic patients. Coll Antropol 35, 39−44.

[3] Rajji, T.K., Ismail, Z., Mulsant, H., 2009. Age at onsetand cognition in schizophrenia: meta-analysis. BJP195, 286–293.

P.4.010 Association of personality features withlithium prophylactic response

D. Dembinska-Krajewska1 °, S. Kliwicki1, M. Chlopocka-Wozniak1, J. Rybakowski1. 1Poznan University ofMedical Sciences, Department of Adult Psychiatry,Poznan, Poland

Purpose of the study: Association of personality featureswith response to given psychotropic drug has not beenfrequently studied so far. Lithium is still a cornerstone forthe long-term therapy of bipolar disorder. Clinically, thebest response to lithium is associated with episodic clinicalcourse, complete remission, bipolar family history and lowpsychiatric co-morbidity, however, a specific personalityprofile for the best lithium response has not been estimated.Such a possibility has recently occurred with an adventof new temperamental scales, specifically designed forbipolar illness, and an ability to quantitatively assesslithium prophylactic response. The aim of the presentstudy was to find a possible relationship between theeffectiveness of prophylactic lithium efficacy measured byAlda scale [1] and the scores of the Temperament Scale ofMemphis, Pisa, Paris, and San Diego Autoquestionnaire(TEMPS-A), as well as the scores of the Oxford-LiverpoolInventory of Feelings and Experiences (O-LIFE) scale, ina group of long-term lithium-treated patients.

Methods used: The study was performed on 70 patientswith bipolar mood disorder (21 male, 49 female), aged31−82 (59±12) years, recruited from the outpatientsin Department of Adult Psychiatry, Poznan Universityof medical Sciences. They have been receiving lithiumfor at least 5 years (5−37, mean 15+8) years. Thecourse of illness was assessed retrospectively basedon the results of Alda scale and scored in the rangeof 0−10. The TEMPS-A questionnaire 110 questionsversion has been used, assessing five temperamentdomains: depressive, cyclothymic, hyperthymic, irritableand anxious. Previously, the TEMPS-A scale has beenvalidated in a group of 521 Polish college students [2]. Theshort version of O-LIFE scale has been used measuringsuch dimensions of schizotypy as unusual experiences,cognitive disorganisation, introvertive anhedonia, and im-pulsive nonconformity. Using this scale we have previously


found that bipolar patients obtained significantly higherscores on schizotypy compared with control subjects andwe found an association between the features of schizotypyand creativity in bipolar patients [3].

Summary of results: The mean score on the scaleAlda was in the whole group 5.9+2.7, similar in male andfemale patients. As to five temperaments of TEMPS-A, theresponse to lithium significantly positively correlated withhyperthymic temperament scores (r = 0.31, p = 0.009), andnegatively with anxious (r = −0.27, p = 0.022), cyclothymic(r = −0.26, p = 0.032), and depressive (r = −0.23, p = 0.052)temperament scores. As to the O-LIFE scales, the responseto lithium demonstrated a significant negative correlationwith cognitive disorganisation (r = −0.24, p = 0.025).Correlation with unusual experiences and introversion andanhedonia other was also negative, and with impulsivenonconformity positive in men and negative in women,but not statistically significant.

Conclusions: The results of the study indicatethat temperamental features of hypomania (hyperthymictemperament) and lack of cognitive disorganisation predictthe best results of lithium prophylactic response. They areconsistent with primarily antimanic lithium efficacy, itsweaker effect in rapid cycling and co-morbid anxiety anda lack of antipsychotic effect.

Reference(s)

[1] Grof, P., Duffy, A., Cavazzoni, P., Grof, E., Garnham,J., MacDougall, M., O’Donovan, C., Alda, M. 2002.Is response to prophylactic lithium a familial trait?J Clin Psychiatry 63, 942–947.

[2] Borkowska, A., Rybakowski, J.K., Drozdz, W.,Bielinski, M., Kosmowska, M., Rajewska-Rager, A.,Bucinski, A., Akiskal, K.K., Akiskal, H.S. 2010.Polish validation of the TEMPS-A: the profileof affective temperaments in a college studentpopulation. J Affect Disord 123, 36−41.

[3] Rybakowski, J.K., Klonowska, P. 2011. Bipolar mooddisorder, creativity and schizotypy: An experimentalstudy. Psychopathology 44, 296–302.

P.4.011 Clinical features and drug-inducedside effects in early versus lateantidepressant responders

C. Fabbri1 °, A. Marsano1, M. Balestri1, A. Serretti1.1University of Bologna, Department of Biomedical andNeuroMotor Sciences, Bologna, Italy

Early antidepressant response has been demonstrated asthe result of a true antidepressant effect [1] and a predictorof subsequent stable response [2], but few data about the

features of this pattern of response compared to the lateone were reported.With the purpose to study the clinical profile of

early response/remission compared to late response/remission, two independent major depressive disorder(MDD) samples (the Sequenced Treatment Alternativesto Relieve Depression or STAR*D level 1 n = 1922and an Italian sample n = 171) were investigated. MDDdiagnosis was ascertained according to DSM-IV-TRcriteria in both samples and depressive symptomatologywas assessed biweekly by the Quick Inventory ofDepressive Symptomatology Clinician Rated (QIDS-C) inthe STAR*D while weekly by the Hamilton DepressiveRating Scale (HDRS) in the Italian sample. Patients wereall treated with citalopram in the STAR*D while in anaturalistic setting in the Italian sample (66.67% SSRIs,23.40% SNRIs, 2.92% TCAs, 2.92% NaSSAs, 2.92%SARIs, 1.17% other antidepressants). Anxiolytics, sedativehypnotics, and other medications for concomitant generalmedical conditions were the only additional medicationsallowed in both samples. Antidepressant induced sideeffects were evaluated according to the Patient-RatedInventory of Side Effects (PRISE) in the STAR*D andDosage Record & Treatment Emergent Symptom scale(DOTES) in the Italian sample at each postbaseline visit.The primary outcome was to identify clinical-

demographic predictors of early (2nd week) versus late(4th–6th weeks) antidepressant response (reduction inHDRS or QIDS-C score of at least 50%) and remission(HDRS� 7 or QIDS-C� 5). Given the hypothesis thatearly and late response may be determined by differencesat pharmacokinetic/pharmacodynamic level, the secondaryoutcome was to evaluate possible differences in antidepres-sant induced side effects.The distribution of variables of interest was compared

between early versus late responders/remitters by logisticregression models with baseline symptom severityas covariate. A Bonferroni correction for multiplecomparison was applied. The R software served for theanalysis (cran.r-project.org/).In the STAR*D, higher levels of baseline core depressive

symptoms according to the Bech subscale [3] wereassociated with early response (p = 0.00017), as well aslower baseline insomnia (p = 0.003) and higher functioning(p = 0.001) according to the Work and Social AdjustmentScale (WSAS). The absence of anxious MDD showed atrend of association with early response (p = 0.038) andremission (p = 0.026). In the Italian sample a trend ofhigher anxious MDD rate (p = 0.19) and lower baselinequality of life (p = 0.078) according to the World HealthOrganization Quality of Life Scale (WHOQOL) wereobserved in late remitters.


In the STAR*D late responders reported higher levelsof antidepressant induced side effects, especially difficultyin sleeping (p = 5.68×10−13), with a similar trend in theItalian sample (p = 0.09).Thus, early antidepressant response may represent

a phenotype characterised by higher core depressivesymptoms, lower insomnia, absence of anxious MDD,higher functioning at baseline and lower antidepressantinduced side effects. The identification of early versus lateantidepressant responders at the beginning of the treatmentmay be useful in order to guide therapeutic choices inclinical settings.

Reference(s)

[1] Taylor, M.J., Freemantle, N., Geddes, J.R., Bhagwagar,Z. 2006. Early onset of selective serotonin reuptakeinhibitor antidepressant action: systematic review andmeta-analysis. Arch Gen Psychiatry 63, 1217−23.

[2] Szegedi, A., Jansen, W.T., van Willigenburg, A.P., vander Meulen, E., Stassen, H.H., Thase, M.E. 2009.Early improvement in the first 2 weeks as a predictorof treatment outcome in patients with major depressivedisorder: a meta-analysis including 6562 patients. JClin Psychiatry 70, 344−53.

[3] Bech, P., Gram, L.F., Dein, E., Jacobsen, O., Vitger, J.,Bolwig, T.G. 1975. Quantitative rating of depressivestates. Acta Psychiatr Scand 51, 161−70.

P.4.012 Effects of second generationantipsychotics on cognitive domainsmeasured with the Matrics ConsensusCognitive Battery in early psychoses

I. Montalvo1 °, M. Creus1, A. Gutierrez-Zotes1,L. Ortega1, R. Monseny1, T. Feliu1, J. Franch1, E. Vilella1,J. Labad1. 1HPU Institut Pere Mata IISPV UniversitatRovira i Virgili, Early Psychosis Unit & ResearchDepartment, Reus, Spain

Introduction: Cognitive impairments, which are presentat early stages of psychoses, are related to low functionaloutcomes [1]. Second generation antipsychotics may havedifferent effects on cognition, although this remainsuncertain [2]. The purpose of the present study wasto compare the effect of different second generationantipsychotics on the cognitive performance in patientswith a first episode of psychosis.

Methods: We included 54 subjects (46.3% women;mean age 24.5 years) with a psychotic disorder witha duration of illness <5 years, attending the EarlyPsychosis Program from Reus (Tarragona, Spain). Severityof psychotic symptoms was assessed with the PANSS.Cognition was assessed with the MATRICS Consensus

Cognitive Battery (MCCB) [3], that explores sevencognitive domains: Speed of Processing (SOP), Attention/Vigilance (AV), Working Memory (WM), Verbal learning,Visual Learning, Reasoning and Problem Solving (RPS)and Social Cognition (SC), and the Overall CompositeScore. Current antipsychotic treatment was requested.Of all patients, 10 were not receiving antipsychoticdrugs, 36 were on monotherapy (17 risperidone/paliperidone, 13 olanzapine, 6 aripiprazole) and 8 onpolytherapy. Antipsychotic drug doses were transformedinto equivalents of chlorpromazine (mg/day). Biperdenand benzodiazepine (diazepam equivalents) doses wereregistered.Statistical analyses were performed with SPSS v.17.0.

ANOVA was used to compare MCCB T-scores foreach cognitive domain by antipsychotic groups. Amultiple linear regression analysis was used to explorethe relationship between antipsychotic doses and neu-rocognitive domains after excluding those subjects onpolytherapy. For each cognitive domain, T-scores (age andgender corrected) were considered the dependent variable.Antipsychotic drugs were included in the equationas three independent variables (risperidone/paliperidone,aripiprazole and olanzapine), as well as biperden anddiazepam doses. All these variables were included ineach model with the enter procedure. Standardised betacoefficients will be shown.

Table: MCCB Cognitive Domains (T-scores) by antipsychotics

1.No anti-psychotics

2.Risperidone/Paliperidone

3.Olanzapine

4.Aripiprazole

5.Polytherapy

ANOVAP-value

Significantpost-hoc tests(Bonferroniadjustment)

SOP 37.8 (14.2) 33.4 (7.7) 44.1 (9.2) 33.8 (10.1) 19.9 (11.3) <0.001 1> 5; 3> 5AV 36.3 (8.6) 33.0 (7.4) 43.8 (9.8) 38.0 (8.2) 30.0 (6.2) 0.004 3> 2; 3> 5WM 35.7 (8.9) 38.4 (9.0) 37.5 (7.5) 37.8 (8.1) 32.9 (13.3) 0.696Verbal 45.1 (10.2) 42.6 (5.5) 43.6 (9.5) 39.7 (7.3) 35.5 (3.9) 0.089Visual 34.8 (13.0) 40.4 (9.3) 45.9 (8.5) 33.3 (9.9) 22.1 (11.2) <0.001 2> 5; 3> 5RPS 37.5 (8.9) 42.9 (10.1) 47.7 (9.6) 42.5 (6.3) 34.8 (6.0) 0.031SC 39.4 (11.4) 37.3 (11.1) 39.0 (13.0) 43.8 (11.4) 42.8 (5.6) 0.747Composite 29.6 (11.8) 31.7 (7.3) 37.7 (8.4) 30.0 (10.9) 25.5 (4.5) 0.117

Data are Mean (SD).

Results: The mean (SD) positive, negative and generalPANSS scores of the sample were 10.2 (3.8), 14.5 (5.4)and 24.9 (7.4) respectively. There were no significantdifferences in PANSS scores between groups.MCCB T-scores by antipsychotic groups are described

in Table. Subjects with olanzapine performed better thanthose subjects on polytherapy in SOP, AV and visuallearning. In AV, those subjects on olanzapine performedbetter than those with risperidone/paliperidone.In the multiple regression analyses, adjusted by

diazepam and biperden, olanzapine was significantlyassociated to a better performance in SOP (B= 0.383,p = 0.032), visual learning (B = 0.394, p = 0.029), andRPS (B= 0.477, p = 0.007). Risperidone/paliperidone wereassociated with a poorer cognitive performance inAV (B=−0.400, p = 0.041). We found no significant


differences related to the Overall Composite Score,although olanzapine showed a trend towards significance(B = 0.356, p = 0.062).

Conclusions: Olanzapine shows a better cognitiveprofile than other second generation antipsychotics. SOP,AV, visual learning and RPS seem to be more clearlyaffected by antipsychotic treatment.

Reference(s)

[1] Fett AK, Viechtbauer W, Dominguez MD, Penn DL,Van os J, Krabbendam L., 2011. The relationshipbetween neurocognition and social cognition withfunctional outcomes in schizophrenia: a meta-analysis.Neurosci. Biobehav. Rev. 35 (3), 573–588.

[2] Woodward ND, Purdon SE, Meltzer HY, Zald DH.,2007 A meta-analysis of cognitive change withhaloperidol in clinical trials of atypical antipsychotics:dose effects and comparison to practice effects.Schizophrenia Research. 89, 211–224.

[3] Rodriguez-Jimenez R, Bagney A, Garcia-Navarro C,Aparicio AI, Lopez-Anton R, Moreno-Ortega M,Jimenez-Arriero MA, Santos JL, Lobo A, Kern RS,Green MF, Nuechterlein KH, Palomo T. 2012. TheMATRICS consensus cognitive battery (MCCB): Co-norming and standardization in Spain. SchizophreniaResearch. 134(2−3), 279−84.

P.4.013 Modulation of brain structure bycatechol O-methyltransferase Val158Metpolymorphism in chronic cannabis users

A. Batalla1 °, C. Soriano-Mas2, M. Torrens3, J.A. Crippa4,S. Bhattacharyya5, L. Blanco-Hinojo6, B.J. Harrison7,M. Farre3, J. Pujol6, R. Martın-Santos1. 1HospitalClinic IDIBAPS CIBERSAM, Department of Psychiatry,Barcelona, Spain; 2Bellvitge University Hospital-IDIBELL CIBERSAM, Department of Psychiatry,Barcelona, Spain; 3IMIM (Hospital del Mar MedicalResearch Institute)-INAD-Parc de Salut Mar, HumanPharmacology and Clinical Neuroscience and Disordersby Use of Substance, Barcelona, Spain; 4University of SaoPaulo, Neuroscience and Cognitive Behavior Department,Ribeirao Preto, Brazil; 5King’s College London Instituteof Psychiatry, Department of Psychosis Studies, London,United Kingdom; 6Institut d’Alta Tecnologia-PRBB,CRC Hospital del Mar, Barcelona, Spain; 7MelbourneNeuropsychiatry Centre, Department of Psychiatry,Melbourne, Australia

Background: Neuroimaging studies have shown thatchronic consumption of cannabis may result in alterationsin brain morphology [1]. Recent work focussing on the

relationship between brain structure and the catechol-O-methyltransferase (COMT) gene polymorphism suggestthat functional COMT variants may affect brain volumein schizophrenia patients, subjects at risk for psychosis andin healthy individuals [2]. Furthermore, epidemiologicaland experimental studies have shown that val allelecarriers may be more sensitive to the longer termeffects of cannabis as well as the acute effects ofdelta-9-tetrahydrocannabinol [3]. Nevertheless, there areno previous studies published that have examined theinfluence of COMT polymorphism on brain morphologyin subjects chronically exposed to cannabis.

Aims: The aim of the present study was to explore theinfluence of COMT Val158Met functional polymorphismon four key regions: the prefrontal cortex, neostriatum(caudate-putamen), anterior cingulate cortex (ACC), andthe hippocampus–amygdala complex, in a group ofearly-onset chronic cannabis users compared with non-using control subjects. We hypothesized that COMTVal158Met functional polymorphism would be associatedwith brain morphological deficits in cannabis users relativeto healthy controls, with dose-dependent associationsbetween volume brain variations and val allele dosage.

Methods: We selected 29 chronic cannabis users whobegan use cannabis before 16 years of age and matchedthem to 28 healthy volunteers in terms of age, educationallevel and intelligence quotient. Participants were male,Caucasians aged between 18 and 30 years. All wereassessed by a structured psychiatric interview (PRISM) toexclude any lifetime Axis-I disorder according to DSM-IV.COMT genotyping was performed and structural magneticresonance imaging data was analysed by voxel-basedmorphometry (VBM).

Results: The results showed a significantly influenceof the COMT polymorphism in bilateral ventral caudatenucleus volume in both groups but in an opposite direction:more copies of val allele was associated with lesser volumein chronic cannabis users and more volume in controls.An opposite pattern was observed for the left amygdala;the greater number of copies of val allele was associatedwith increased volume in chronic cannabis users anddecreased volume in controls. There were no effects ofCOMT genotype on volumes of the whole brain or theother selected regions. Irrespective of genotype, we alsoidentified a significant positive correlation between caudalrectal gyrus-subgenual cingulate cortex volume and thenumber of joints consumed. Finally, we reported an almostsignificant grey matter volume increase in the postcentralgyrus of the left hemisphere in chronic cannabis users.

Conclusions: Our findings support recent reports ofneuroanatomical changes associated with cannabis useand, for the first time, reveal that these changes may beinfluenced by the COMT genotype. Further prospective,


longitudinal research is needed to examine the gene–environment influence and the mechanisms of long-termcannabis related brain impairment.Grants: Ministerio de Sanidad y Consumo: PNSD

PI101/2006, PNSD PI041731/2011; DIUE de la Gener-alitat de Catalunya: SGR 2009/1435.

Reference(s)

[1] Martin-Santos, R., Fagundo, AB., Crippa, J.A.,Atakan, Z., Bhattacharyya, S., Allen, P., Fusar-Poli, P.,Borgwardt, S., Seal, M., Busatto, G.F., McGuire, P.,2010. Neuroimaging in cannabis use: a systematicreview of the literature. Psychol Med 40, 383−98.

[2] Honea, R., Verchinski, B.A., Pezawas, L., KolachanaB.S., Callicott, J.H., Mattay, V.S., Weinberger, D.R.,Meyer-Lindenberg, A., 2009. Impact of interactingfunctional variants in COMT on regional gray mattervolume in human brain. Neuroimage 45, 44−51.

[3] Estrada, G., Fatjo-Vilas, M., Munoz, M.J., Pulido, G.,Minano, M.J., Toledo, E., Illa, J.M., Martın, M., Mi-ralles, M.L., Miret, S., Campanera, S., Bernabeu, C.,Navarro, M.E., Fananas, L., 2011. Cannabis useand age at onset of psychosis: further evidence ofinteraction with COMT Val158Met polymorphism.Acta Psychiatr Scand 123, 485−92.

P.4.014 Fear conditioning and context learningin relation to treatment outcome: a studyin patients with panic disorder andsocial phobia

P. Duits1 °, J.M.P. Baas2, I.M. Engelhard1, M.A. vanden Hout1, D.C. Cath1. 1Utrecht University, Clinicaland Health Psychology, Utrecht, The Netherlands;2Utrecht University, Experimental Psychology, Utrecht,The Netherlands

Anxiety disorders are among the most prevalentpsychiatric disorders. Results from single-cue fearconditioning studies point to modest increases in fearacquisition and fear extinction in patients with anxietydisorders [1]. However, results from discrimination studieshave provided inconsistent support so far. First purpose ofour study was to further investigate differences betweenpatients with anxiety disorders in fear conditioning. Weadded a safety cue and a safe context to our fearconditioning paradigm to measure context learning andinhibition of fear as well. The second purpose of ourstudy relates to the state of the art treatment in anxietydisorders: exposure with response prevention. Within thisbehavioural treatment, fear extinction mechanisms are atthe core of the procedure. However, the exact relationship

between speed of fear acquisition and extinction andsubsequent treatment effect in patients with anxietydisorders is still unclear. Therefore, our second studyaim was to investigate the predictive value of fearacquisition and extinction mechanisms in patients withanxiety disorders on treatment outcome.Patients with panic disorder (n = 15) and social phobia

(n = 15) who finished their exposure treatment in thepast years, were included in the present study. An age-,gender- and education-matched control group (n = 20) wasalso included. Fear learning mechanisms were studied inan experimental laboratory test consisting of three phases.Within the task, participants watched movie clips in whichtwo different environments (contexts) were visited forevery experimental block. During the first phase, onespecific cue (the increase in background illumination) wasbe followed by the administration of a mild electrical shockto the wrist in one of the contexts, but not the other.After six blocks in which participants may spontaneouslylearn the association between the light in the shockcontext and the shock, specific instructions about whenthe shock can in fact be administered were provided (thissecond phase consisting of 5 blocks). The last phase ofthe experiment consisted of four more blocks withoutthe shock reinforcements. During all phases, subjectivefear, awareness of the light-shock association and fearpotentiated startle to all conditions of the experiment (i.e.,the two contexts and the presence / absence of the lightcue therein) were assessed. Treatment outcome had beenassessed at time of treatment by means of widely usedgeneral and disorder specific questionnaires.Contrary to our expectations, the conditioning task

demonstrated no differences in fear acquisition, extinctionand context learning between patients and healthy controls.These results suggest that pathological fear mechanismsrepresent a state characteristic at time of an anxietydisorder being present rather than an underlying trait, sincethe patients already finished their anxiety treatment at thetime of participation in the present study.

Reference(s)

[1] Lissek, S., Powers, A.S., McClure, E.B., Phelps, E.A.,Woldehawariat, G., Grillon, C., & Pine, D.S. (2005).Classical fear conditioning in the anxiety disorders: Ameta-analysis. Behaviour Research and Therapy 43,1391–1424.


P.4.015 Long term therapy with methylphenidateinduces modest effects on growth inADHD children

C. Balia1 °, A. Anedda1, F. Granitzio1, S. Carucci1,A. Zuddas1. 1Cagliari University Hospital, DeptBiomedical Science, Cagliari, Italy

Background: Although stimulants are the most effectivemedication for Attention Deficit Hyperactivity Disorder(ADHD), poor growth is a common concern, especiallywith children already on the lower growth percentiles.Studies providing longitudinal data indicate a reductionin both height and weight gain: these effects are usuallyminimal, but there is substantial variability with somechildren completely unaffected, whereas others showssignificant growth suppression [1].

Objectives: To evaluate whether long term immediaterelease methylphenidate (IR-MPH) therapy (one or twoyears) interferes with the growth of ADHD children andto assess whether the effects on growth are related to thelength of the treatment or to the daily dose.

Methods: Growth parameters were collected from 90ADHD aged 6 to 14, enrolled at one of the site of theItalian National Register for ADHD. All patient were onIR-MPH and with a minimum follow-up of 12 months.65 were Drug Naıve (DN), 25 were already on MPHsince 1−3 years prior to enrollment in the Registry (PR).Weight, height, BMI, height Z-score and BMI Z-scorewere recorded at each follow-up visit (baseline and after6, 12, 18, 24 months). Growth velocity SDS and heightdeficit were calculated after 12 and 24 months.

Data Analysis: Categorical data were analysed usingcontingency tables (c2), continuous variables werecompared by one-way ANOVA. Repeated measuresANOVA was performed for height and BMI z scores atbaseline, 6, 12, 18, 24 month follow up and for heightvelocity SDS at 12 and 24 months.

Results: At baseline Height Z-scores of the entiresample was −0.33±0.98, BMI Z-score was equal to0.19±1.14. During the 24 months in the study, subjectsgained in absolute values of height and weight. HeightZ-score showed a significant decrease only from T12 toT24 (p = 0.05). BMI Z-score decreased significantly atT12 (p< 0.001) remaining essentially unchanged at T24.Height deficit was about 0.5 cm at 12 months and1.3 cm after 24 months. MPH dose/kg/day changed from0.49±0.21mg when starting medication, to 0.68±0.24 atT12 and to 0.75±0.25 at T24. No significant differenceswere found on growth parameters at baseline whenstratifying between DN and PR. As in the total sample, inboth groups a significant decrease in BMI Z-score frombaseline to T12 (p< 0.001) and in height Z-score between

T12 and T2 (p = 0.05) was found. No changes in growthvelocity from baseline to the different times considered,were observed, neither in DN nor in PR.

Discussion: The findings of the present study suggestthat the effects of MPH on growth are relatively smalland unlikely to be of clinical concern for this population.Expected and actual deficit in growth should be consideredin the context of the benefits the patient receives from themedication. In the present sample the height deficit appearsto be more related to the maximum pro/die dose ratherthan to the length of therapy. More research is neededto better elucidate the mechanism of growth suppressionand to implement specific treatment strategies for ADHDchildren.

Reference(s)

[1] Pliszka, S.R., Matthews, T.L., Braslow, K.J., Wat-son, M.A., 2006. Comparative Effects of Methyl-phenidate and Mixed Salts Amphetamine on Heightand Weight in Children With Attention-Deficit/Hyperactivity Disorder. J Am Acad Child AdolescPsychiatry 45, 520–526.

P.4.016 Regional differences of SERT occupancyin major depression: an in vivo PETstudy using [11C]DASB

P. Baldinger1 °, G.S. Kranz1, M. Savli1, W. Wadsak2,D. Haeusler2, A. Hahn1, M. Mitterhauser2, C. Philippe2,S. Kasper1, R. Lanzenberger1. 1Medical University ofVienna, Dept of Psychiatry and Psychotherapy, Vienna,Austria; 2Medical University of Vienna, Dept of NuclearMedicine − PET Center, Vienna, Austria

Objective: The blockage of serotonin transporters (SERT)responsible for serotonin reuptake from the synaptic cleftinto the presynaptic neuron is the primary mechanism ofaction of selective serotonin reuptake inhibitors (SSRIs).One might assume that the selective affinity of an SSRIfor SERT and hereby its antidepressant effectivenessmight be similar throughout the brain. However, SERTactivity mediated via various factors may differ betweenregions as SERT is a priori not equally distributed in thebrain. Furthermore, as SERT internalisation process andthereby its availability in a distinct area depends on itsactivity, SERT occupancy by an SSRI might equally varythroughout the brain [1]. Here, we investigated whetherSERT occupancy by escitalopram/citalopram is equallydistributed in brain areas known to play a role in majordepression using positron emission tomography (PET).

Methods: 19 outpatients (13 females; age 42.3±7.8years (mean±SD)) suffering from major depressivedisorder (17-item HAMD � 16, no pharmacological


treatment 3 months prior scanning) were included in thispooled longitudinal study. Subjects received oral doses ofeither escitalopram (10mg/day, 10 subjects) or citalopram(20mg/day, 9 subjects) and underwent three [11C]DASBPET scans: before treatment (PET1), 6 h following thefirst SSRI dose (PET2) and 6 h after the last dose (PET3),which was administered daily for a minimum of 3 weeks(24.73±3.3 days) as described previously [2]. Quantifi-cation of SERT binding potential (BPND) was performedusing MRTM2. PET images were spatially normalised toa template in MNI space using SPM8. SERT BPND wascomputed using as regions of interest (ROI) approach,where ROIs were selected from a customised templatebased on the AAL atlas. Cerebellar grey was used as refer-ence region. Using SPSS median SERT occupancy valuesacross subjects for each of the 27 ROIs were tested againstoverall occupancy (= median of all 27 ROIs). One-sampleWilcoxon Signed-Rank Tests were performed usingmedian = 68.13 for PET2 and median = 71.50 for PET3.

Results: Injected doses, masses, and specific activ-ities for [11C]DASB did not differ between groupsand time points. Regarding PET2, SERT occupanciessignificantly differ from overall median values inmiddle temporal gyrus (T* = −3.59, p< 0.001), subgen-ual cingulate cortex (T* = 3.78 p< 0.001), amygdala(T* = 3.82, p< 0.001), nucleus accumbens (T* = 3.66,p< 0.001), median (T* = 3.78, p< 0.001) and dorsalraphe nucleus (T* = 3.82, p< 0.001). For PET3 thiswas the case for inferior temporal gyrus (T* = −3.37,p = 0.001), supramarginal gyrus (T* = −3.37, p = 0.001),middle temporal gyrus (T* = −3.59, p< 0.001), subgen-ual cingulate cortex (T* = 3.58, p< 0.001), amygdala(T* = 3.82, p< 0.001), nucleus accumbens (T* = 3.54,p< 0.001), median (T* = 3.82, p< 0.001) and dorsalraphe nucleus (T* = 3.82, p< 0.001). All results surviveBonferroni correction for multiple comparisons.

Conclusions: SERT occupancy was shown to varythroughout the cortex in several subcortical and corticalbrain areas, e.g. the subgenual cingulate cortex andthe amygdala, brain regions known to be involved inthe pathogenesis of depression. This is in accordancewith previous preclinical studies showing that SSRIconcentrations differ between brain regions and mighttherefore impact on occupancy values in a variousdegree [3] cortically and subcortically. This region-specificmodulation by escitalopram and citalopram might beof major clinical relevance in the treatment of majordepression.

Reference(s)

[1] Meyer H.J., Wilson A.A., Sagrati S., Hussey D.,Carella A., Potter W.Z., Ginovart N., Spencer E.P.,Cheok A., Houle S., 2004. Serotonin Transporter

Occupancy of Five Selective Serotonin ReuptakeInhibitors at Different Doses: An [11C]DASB PositronEmission Tomography Study. Am J Psychiatry161:826–835.

[2] Lanzenberger R., Kranz G.S., Haeusler D., AkimovaE., Savli M., Hahn A., Mitterhauser M., SpindeleggerC., Phillipe C., Fink M, Wadsak W., KaranikasG., Kasper S., 2012. Prediction of SSRI treatmentresponse in major depression based on serotonin trans-porter interplay between median raphe nucleus andprojection areas. Neuroimage Nov 1;63(2):874−81.

[3] Kugelberg F.C., Apelqvist G., Carlsson B., Ahlner J.,Bengtsson F., 2001 In vivo steady-state pharmacoki-netic outcome following clinical and toxic of racemiccitalopram to rats. British Journal of Pharmacology132, 1683–1690.

P.4.017 Post training noradrenaline reuptakeinhibition modulates fear memoryconsolidation and abolishes long termfear responses in humans

J. Almeida1 °, J. Tulen1, F. van der Veen2, S. Kushner1.1Erasmus MC, Department Psychiatry, Rotterdam,The Netherlands; 2Erasmus University, DepartmentPsychology, Rotterdam, The Netherlands

Background: Although memory enhancement for signif-icant events such as life threatening experiences is animportant factor in survival, traumatic events can lead topsychological disturbances, including posttraumatic stressdisorder (PTSD). After a given event, formation of its cor-respondent memory undergoes a process termed consoli-dation [1]. This raises the possibility that pharmacologicalintervention in the aftermath of trauma could forestall thedevelopment of PTSD and other anxiety disorders [2].Here we aimed to delineate the precise effects of

acute differential modulation of noradrenaline (NA)and serotonin in memory consolidation processes. Forthis purpose we combined a differential pavlovian fearconditioning (FC) paradigm with the administration of asingle dose of the selective NA reuptake inhibitor (SNRI)atomoxetine and the selective 5-HT reuptake inhibitor(SSRI) citalopram.

Methods: Ninety healthy volunteers were recruited andassigned to a differential pavlovian FC protocol, involvingtwo conditional stimuli (CS+ and CS−) where only theformer was paired with a shock. Afterwards subjectstook a pill (atomoxetine vs citalopram vs placebo) ina double-blind RCT design. Blood samples were takenbefore conditioning and at t = 1 h and t = 3 h. A day laterparticipants underwent extinction training in which the twoCS were repeatedly presented without the shock.


Six months later participants were submitted toreinstatement followed by re-extinction training toexamine the long term effects of the intervention. Skinconductance responses were obtained at all occasions forfear memory measure.

Results: Sixty-two subjects showed successful fearacquisition on day 1 and were included in the analysis(placebo = 23; citalopram = 21; atomoxetine = 18). Nodifferences were found between the 3 groups (two-wayANOVA regarding main effects of group and time; non-significant). Testing on day 2 revealed no differencesconcerning CS+ responses between treatment groups(F2,61 = 1.93). However there was a significant maineffect of group concerning CS− responses (F2,61 = 10.07;p< 0.05). Follow-up t-tests of the first two CS− trialsshowed significantly lower responses for the atomox-etine group (t = 2.43, t = 2.14; both p< 0.05). MHPG(3-methoxy-4-hydroxyphenylglycol, a NA metabolite)absolute change levels correlated significantly with meanCS+ and CS− difference in the first three trials forthe atomoxetine group (R2 = 0.28; p< 0.05), with nocorrelation for the placebo group.Six months afterwards, 37 participants (placebo = 14;

citalopram = 10; atomoxetine = 13) were located andreassessed. Average first CS+ responses between groupsshowed significance (Kruskal–Wallis; p< 0.05). Follow-uptesting revealed significantly lower CS+ responses for theatomoxetine group.

Conclusion: This study shows that memory consoli-dation can be influenced by SNRI administration afteracquisition, lowering CS− responses on the short term,with an apparent similar effect in the CS+ on the longterm.The findings suggest that the emotional salience of a

noxious stimuli conferred by NA could in part stem fromthe relative downgrading of the concomitant/temporaladjacent stimuli rather than elevation of its absolute value.As generalisation of abnormal responses to similar yetunrelated trauma cues is one of the cardinal featuresof PTSD this study might offer new insights into itsprophylaxis in the aftermath of trauma. Additionally itadds weight to the assumption that memory formation isan ongoing dynamic process that continues long after theevents that led to its generation.

Reference(s)

[1] Pitman, R.K., Rasmusson, A.M., Koenen, K.C.,Shin, L.M., Orr, S.P., Gilbertson, M.W., et al., 2012.Biological studies of post-traumatic stress disorder.Nat Rev Neurosci. 11, 769−87.

[2] Bush, D.E., Caparosa, E.M., Gekker, A., Ledoux, J.,2010. Beta-adrenergic receptors in the lateral nucleus

of the amygdala contribute to the acquisition but notthe consolidation of auditory fear conditioning. FrontBehav Neurosci. 4, 154.

P.4.018 General error monitoring systemdysfunction in obsessive compulsivedisorder patients: an event-relatedpotential study

L. Carmi1 °, U. Alyagon2, A. Zangen2, R. Dar3, J. Zohar1.1Sheba Medical Center, Pychiatry, Ramat Gan, Israel;2Ben Gurion University, Life Science–Neuroscience,Beer-Sheva, Israel; 3Tel Aviv University, Psychology, TelAviv, Israel

Introduction: Enhanced Error Related Negativity (ERN)signal after making mistakes is more frequent in OCDpatients compared to healthy participants and was foundto be generated by theta band [1]. Nevertheless, it isnot clear whether this hyperactive ERN signal reflectsa hyperactive monitoring system that is over-sensitive topunishment cues or a dysfunction of a more general andless affective monitoring system. The latter manner wouldimply a system which continuously seeks out erroneousinformation in the inner or outer environment, regardlessof a mistake being made. Indeed, normal participantshave shown enhanced theta activity merely by identifyingerroneous content [2].To test the hypothesis of a general hyperactive

monitoring system we tested whether OCD patients showenhanced theta activity after identifying erroneous contentas compared to controls. As the ERN enhancement iscorrelated with OC symptom severity, we hypothesizedthat reduction of the hyper theta activity in OCD patients,via Deep Trans Cranial Magnetic Stimulation (dTMS),will be correlated with a decrease in symptom severity.

Method: We used the Stroop task to elicit ERNand an arithmetic task [2] to elicit theta activity inresponse to erroneous content. In our interim analysis,Nine OCD patients and nine healthy controls were tested.In the arithmetic task, participants were requested only toindicate whether a solution of a simple equation is corrector incorrect. The OCD group went through five weeks ofinhibitory blind dTMS treatment (active vs. sham). EEGmeasurements were taken at baseline, after the first dayof treatment and at the end of treatment course. The OCsymptom severity was evaluated by YBOCS.

Results: A non-phase locked wavelet analysis re-vealed a significant interaction of group and condition[F(2,32) = 5.23 (p< 0.01), p.e = 0.24]. A relative powerincrease in low frequency bands (delta, theta and alpha)were shown in the OCD group compared to healthy controlafter merely identifying wrong solutions. In contrast,


no differences were found when correct solutions werepresented. Activity had a midline frontal locus (Afzelectrode), similar to previous findings, suggesting ACCas a possible source [3].The YBOCS scores (Table 1) revealed that 40% of the

real dTMS treatment group (5 patients) reached remissioncriteria (reduction of 35% from baseline score) after5 weeks of treatment. No response was obtained in thesham group.

Table 1. YBOCS scores at baseline, middle and end of treatment in OCDgroup

Subject Baseline End (5th week)

Active 1 24 28Active 2 30 31Active 3 27 17*Active 4 29 27Active 5 24 8*Sham 1 27 26Sham 2 29 26Sham 3 22 22Sham 4 22 20

*Indicates remission.

Conclusion: Over-monitoring condition is expressedin OCD patients not only when an error is committed(ERN), but also upon observing an error. Moreover, areduction of this hyper activity (via dTMS) was associatedwith reducing OC symptom severity (as measured inthe YBOCS scores). These interim findings may implya cognitive endophenotype basis for the bias attributed toOCD patients in particular and OCD spectrum in general.

Reference(s)

[1] Cohen, M.X., Elger, C.E., Ranganath, C., 2007.Reward expectation modulates feedback-related neg-ativity and EEG spectra. Neuroimaging 35, 968–978.

[2] Tzur, G., Berger, A., 2007. When things look wrong:theta activity in rule violation. Neuropsychologia 45,3122–3126.

[3] Luu, P., Tucker, D.M., Makeig, S., 2004. Frontalmidline theta and the error related negativity:Neuro physiological mechanisms of action regulation.Clinical Neurophysiology 115, 1821–1835.

P.4.019 Insight and recovery in schizophrenicpatients: an observational study

D. Cannavo1 °, C. Concerto1, E. Battaglia1, O. Bianchini1,E. Aguglia1. 1University of Catania, Psychiatry, Catania,Italy

Purpose of the study: Progress in therapeutic options forschizophrenia has revived long-term expectations for re-searchers, practitioners and patients. At present, definitions

of therapeutic outcome include both maintained symp-tomatic remission and appropriate functioning in a concep-tual framework that targets patient’s recovery as the ulti-mate goal. Paradoxically, insight in schizophrenic patientsis associated with positive outcomes such as better compli-ance and recovery, negative outcomes such as depression,hopelessness, low self-esteem and quality of life [1,2]. Theaim of this study was to investigate the correlation betweeninsight and recovery in schizophrenic patients accordingto criteria for both symptomatic and functional remission.

Methods used: We designed an observational study of24 months of duration; visits were scheduled at baseline,12 and 24 months. We included 70 patients (15 patientstreated with olanzapine, 15 with risperidone, 15 witharipiprazole, 15 with haloperidol and 10 with ziprasidone)affected by paranoid schizophrenia according to DSM IVTR criteria who changed pharmacological therapy fordifferent reasons (inefficacy, presence of side effects orcontraindications) or naive patients who started pharmaco-logical treatment. Rating scales used were SAI (Schedulefor the Assessment of Insight) for the assessment ofinsight, PANSS (Positive and Negative Syndrome Scale)for psychopathological picture, Short Form questionnaire(SF-36) for quality of life, Psychological GeneralWell-Being Index (PGWBI) for well-being and GlobalAssessment of Functioning scale (GAF) to rate the social,occupational and psychological subjective functioning.

Results: After 2 years, 50% of the subjects achievedsymptom remission and 25.5% had adequate socialfunctioning for 2 years or more. Only 12% of subjectsmet full recovery criteria for 2 years or longer. Patients inthe remission group showed a significant better outcomeduring follow-up on all PANSS subscales (positive,negative, and general symptoms subscales), a significanthigher level in social functioning and an improvement inthe subjective perception of well-being. No differenceswere found in terms of quality of life between groups. Forwhat concerns pharmacological antipsychotics treatment,although there were no significant differences in patientsdistribution about symptomatic remission/functioninglevels, patients treated with second generation antipsy-chotics (olanzapine, risperidone, aripiprazole, ziprasidone)that met the symptomatic remission/functioning criteriawere higher compared to those treated with typicalantipsychotics (haloperidol) in monotherapy. all patientsin recovery showed an improvement in insight levels,especially for patients treated with second generationantipsychotics. Recovery after 2 years was predicted byfemale sex, higher age, pre-morbid social adaptation andlow level of negative symptoms at baseline.

Conclusions: The results indicate that only a smallproportion of patients affected by schizophrenia achieverecovery and suggest that social functioning, medication


adherence, type of antipsychotic and improvement ininsight levels are important predictors for recovery [3].Therefore more sensitive instruments and a larger sampleare necessary to confirm these results.

Reference(s)

[1] Cavelti M, Kvrgic S, Beck EM, Rusch N, Vauth R.2012. Self-stigma and its relationship with insight, de-moralization, and clinical outcome among people withschizophrenia spectrum disorders. Compr Psychiatry.53(5):468−79.

[2] Kurtz MM, Tolman A., 2011. Neurocognition, insightinto illness and subjective quality-of-life in schizo-phrenia: What is their relationship? SchizophreniaResearch 127, 157–162.

[3] Stefanopoulou E, Lafuente AR, Saez Fonseca JA,Huxley A., 2009. Insight, global functioning andpsychopathology amongst in-patient clients withschizophrenia. Psychiatr Q 80, 155–165.

P.4.020 Allopurinol for mania − a randomisedtrial of allopurinol vs. placebo as add-ontreatment in manic bipolar patients

S. Burshtein1 °, M. Weiser1, A.A. Gershon1, G. Marian2,N. Vlad3, I.G. Grecu4, E. Tocari5, A. Tiugan6,M. Hotineanu7, J.M. Davis8. 1Sheba Medical Center,Psychiatry C, Ramat Gan, Israel; 2Spitalul Clinic dePsihiatrie Obregia, Psychiatry, Bucuresti, Romania;3Spitalul de Psihiatrie Botosani I, Psychiatry, Botosani,Romania; 4Spitalul Clinic Judetean Mures-Sectia ClinicaPsihiatrie I, Psychiatry, Targu Mures, Romania; 5SpitalulJudetean Vrancea Sectia Psihiatrie Focsani, Psychiatry,Focsani, Romania; 6Sp. Clinic de Urgenta Militar“Dr. Stefan Odoblegea”, Psychiatry, Craiova, Romania;7Spitalul Clinic de Psihiatrie Chisinau, Psychiatry,Chisinau, Moldova Rep. of; 8University of Illinois,Psychiatry, Chicago, USA

Objective: An emerging body of evidence supports a rolefor dysfunctional purinergic neurotransmission in mooddisorders. Adenosine agonists have been shown to haveproperties similar to those of dopamine antagonists; thereis a well characterised interaction between adenosine anddopamine receptors in the ventral striatum, and increasingadenosinergic transmission has been demonstrated toreduce the affinity of dopamine agonists for dopaminereceptors. Allopurinol increases adenosine levels in thebrain, hence is hypothesized to reduce the symptoms ofmania.Two randomised, placebo-controlled trials administering

add-on allopurinol to manic patients showed significantlygreater improvements in YMRS scores for drug compared

to placebo [1,2], while a more recent, relatively small, add-on study showed negative results [3]. Based on these data,our objective was to examine the efficacy of allopurinol asadd-on treatment to mood stabilisers and/or antipsychoticsin manic bipolar patients.

Methods: We performed a large, well powered, multi-centre, 6-week randomised, placebo-controlled trial ofallopurinol added to mood stabilisers and/or antipsychoticsin 180 bipolar patients in an acute manic episode.

Results: Of the 180 participants enrolled in the study,90 were randomised to allopurinol and 90 to placebo.Baseline YMRS scores were 24.6 in the placebo group(SD= 5.6), and 25.4 in the allopurinol group (SD= 5.3)(t = −1.02, p = 0.31). Completion rates were similar, with74/90 (82.2%) completing the 6 week study in theallopurinol group and 75/90 (83.3%) in the placebo group(c2 = 0.04, p = 0.84).Overall, both the treatment and placebo groups

improved significantly. Mean total YMRS score improvedby 11.0 points in the allopurinol group (SD= 8.65,t = −11.83, p< 0.001, effect size 1.612) and 10.6 pointsin the placebo group (SD= 8.56, t = −11.48, p< 0.001,effect size 1.497) by the end of the study. Other clinicaloutcome measures such as CGI-BP and PANSS showedsimilar improvements. However, no significant between-groups differences were detected in change of YMRS score(t = 0.28, p = 0.78), or in the secondary outcome measuresat study’s end.There was no difference in the percent of patients who

responded to treatment (defined as showing at least 50%improvement in YMRS score) between the two groups,with 37.8% responders in the allopurinol group and 38.6%responders in the placebo group (p = 0.92).The frequency of adverse events was also similar

between the two groups (38.9% for the placebo groupand 41.1% for the allopurinol group, p = 0.88).

Limitations: None of our patients received lithium.However, the side effects of lithium and its’ narrowtherapeutic index, made the use of lithium less commonand therefore our study results reflect the common currentclinical practice. Another limitation of our study is thevariety of antipsychotic treatments on which we addedon allopurinol. This is also an outcome of the currentclinical practice for acute in which there is no single ‘goldstandard’ medication.

Conclusions: The findings of this large, well-poweredstudy do not support allopurinol as a treatment for acutemania.

Reference(s)

[1] Akhondzadeh S, Milajerdi MR, Amini H, Tehrani-Doost M. Allopurinol as an adjunct to lithium and


haloperidol for treatment of patients with acute mania:a double-blind, randomized, placebo-controlled trial.Bipolar Disord. 2006; 8:485−9.

[2] Machado-Vieira R, Soares JC, Lara DR, LuckenbaughDA, Busnello JV, Marca G, et al. A double-blind,randomized, placebo-controlled 4-week study on theefficacy and safety of the purinergic agents allopurinoland dipyridamole adjunctive to lithium in acute bipolarmania. J Clin Psychiatry. 2008; 69:1237−45.

[3] Fan A, Berg A, Bresee C, Glassman LH, Rapa-port MH. Allopurinol augmentation in the outpatienttreatment of bipolar mania: a pilot study. BipolarDisord. 2012; 14:206−10.

P.4.021 Genome-wide association analysisand pathway analysis in predictingantidepressant treatment response

N. Antypa1 °, A. Drago1, A. Serretti1. 1University ofBologna, Department of Biomedical and NeuroMotorSciences, Bologna, Italy

Pharmacogenetic research seems to be a promisingpath towards achieving personalised treatment, however,the link between genetic risk variants and complexmental disorders such as depression is far fromdirect. Genomewide association (GWAS) studies onantidepressant efficacy have yielded poor results that failto reach genomewide significance [1] and are hard toreplicate. A possible reason is that antidepressant responsecan be influenced by a multiplicity of sociodemographicand environmental factors, which possibly interact withgenetic variation [2]. For example, the patient’s qualityof life has been previously found to be an independentpredictor of acute antidepressant treatment response [3].The aim of the present study was twofold. Firstly,

we included the patients’ level of quality of life as apsychosocial predictor of response in a GWAS modelof treatment response to citalopram, in a large groupof depressed patients from the Sequenced TreatmentAlternatives to Relieve Depression (STAR*D) trial.Secondly, since a number of candidate genes have emergedfrom prior gene–environment (G×E) interaction studieson depression, we were interested in investigating whethersuch possibly putative cluster of genes is likely to representa causal pathway in predicting treatment response.We investigated a sample of 1426 depressed patients

from the STAR*D trial: 774 responders and 652 non-responders and a subset of 418,865 single nucleotidepolymorphisms (SNPs) were assessed. We examinedwhether genetic variations interact with the patients’levels of Quality of Life (QoL) to predict antidepressant

response, after controlling for demographic characteristics,depression severity and population stratification. As asecond step, we conducted a pathway analysis, whichexplored whether candidate genes (that have emergedfrom prior G×E interaction studies) represent a biologicalpathway in predicting treatment response.The GWAS model yielded one SNP associated with

response, the rs520210 SNP of the NEDD4L gene(p = 3.64×10−8; OR: 0.76). We found that responderswere most likely to be carriers of the minor allele (A),compared to non-responders, especially those reportinglow QoL upon entering the trial. Other genes among thetop findings include FKBP1A, SLC37A1, ASTN1 andTNFRSF10B. The pathway analysis showed that SNPsassociated with treatment response at a p level <0.001 weremore frequently found within the pathway in comparisonto elsewhere in the genome. The HTR2A gene containedthe most significant markers that predicted response.Our findings point to one possible target gene,

namely, the NEDD4L (18q21.31), which is an ubiquitinprotein ligase, thereby playing a role in the proteinrecycling and house keeping activities of the cell.Other genes among our top hits are also proposedfor further independent replication. Our pathway resultsprovide further support on the role of the HTR2Agene in influencing antidepressant response. Despitemethodological and statistical challenges, taking intoaccount distinctive predictors of response, along withgenetic factors, could be a promising avenue for futureresearch.

Reference(s)

[1] Garriock HA, Kraft JB, Shyn SI, Peters EJ,Yokoyama JS, Jenkins GD et al. 2010. A genomewideassociation study of citalopram response in majordepressive disorder. Biol Psychiatry 67, 133–138.

[2] Malhotra AK. 2010. The pharmacogenetics ofdepression: enter the GWAS. Am J Psychiatry 167,493–495.

[3] Pyne JM, Bullock D, Kaplan RM, Smith TL, Gillin JC,Golshan S et al. 2001. Health-related quality-of-lifemeasure enhances acute treatment response predictionin depressed inpatients. J Clin Psychiatry 62, 261–268.


P.4.022 Neural correlates of generalised anxietydisorder without comorbid depression:preliminary data from a functional MRIstudy

K. Hilbert1 °, U. Lueken1, K. Beesdo-Baum1.1Technische Universitaet Dresden, Institute of ClinicalPsychology and Psychotherapy, Dresden, Germany

Background and Aims: Few functional neuroimagingstudies on Generalised Anxiety Disorder (GAD), whichis characterised by excessive and uncontrollable worriesabout a variety of situations accompanied by physicalsymptoms, are available so far. These studies reportedperturbations in the amygdala, ventrolateral prefrontalcortex (VLPFC), anterior cingulate cortex and middlefrontal gyrus. But as GAD samples mostly include patientswith comorbid depression, it remains unclear whether thereported neural correlates result from GAD alone [1].Preliminary data from an ongoing study is reported. Weaimed to validate the neural correlates of GAD reportedso far by investigating emotional processing under twodifferent attentional states in a GAD sample withoutcomorbid depression.

Methods: N=22 medication-free subjects (n = 10 GADpatients without depression and n = 12 healthy controls),diagnosed by a standardised clinical interview andcarefully matched on age (mean = 32.09, range: 21-57),sex (17 female), and level of education (16 high-schoolequivalent), participated in an event-related functionalmagnetic resonance imaging (fMRI) session. Using anemotional processing paradigm that has been appliedsuccessfully in a more complex form before [1], subjectsviewed 16 fearful and 16 happy faces, each presented forfour seconds and repeated four times in randomised order.Attention was varied by instructing the subjects to rateeither their internal fear or the nose-width of the facesonline during the scanning on a five-point likert scale.Blood oxygen level dependent (BOLD) fMRI data werecollected on a 3-T scanner and preprocessed and analysedwith SPM8 (whole-brain analysis and region-of-interest,p< 0.001 uncorrected).

Results: For the contrast fearful versus happy faces,GAD patients compared to healthy controls showedincreased activation in the parietal cortex and decreasedactivation in the ventrolateral prefrontal cortex, middlefrontal gyrus and middle cingulate gyrus during ratingsof internal fear. When rating the nose-width of thefaces, increased activation in the parietal cortex, temporalcortex, orbitofrontal cortex and ventrolateral prefrontalcortex and decreased activation in the inferior frontaloperculum was found in GAD patients for the samecontrast.

Conclusions: Present results indicate a failure ofbrain areas such as the prefrontal cortex, middle frontalgyrus and middle cingulate gyrus to regulate emotionalprocessing in GAD. These processes seem to be modulatedby the attentional focus of the subjects. Results thusconfirm previous findings and indicate the existence ofGAD-specific neural alterations. Furthermore, a robusthyperactivation of the parietal cortex was found inGAD patients, possibly reflecting emotional memoryprocesses during stimulus perception [2] present in GADsamples. However, no differential activation was foundin the amygdala, possibly due to limited statisticalpower. If replicated, these results may prove relevantfor the validator-based classification of the disorderand the ongoing nosological discussion. Future studiesshould employ comparisons of non-comorbid GADsamples with depression and other anxiety disorders torefine knowledge on GAD-specific neural correlates anddetermine similarities and differences between GAD andrelated disorders.

Reference(s)

[1] Beesdo, K., Lau, J.Y.F., Guyer, A.E., McClure-Tone, E.B., Monk, C.S., Nelson, E.E., Fromm, S.J.,Goldwin, M.A., Wittchen, H.-U., Leibenluft, E.,Ernst, M., Pine, D.S., 2009. Common and distinctamygdala-function perturbations in depressed vsanxious adolescents. Arch Gen Psychiatry 66(3), 275–285.

[2] Murty, V.P., Ritchey, M., Adcock, R.A., LaBar, K.S.,2011. Reprint of: fMRI studies of successful emotionalmemory encoding: a quantitative meta-analysis.Neuropsychologia, 49(4), 695–705.

P.4.023 Habenular nuclei in different phasesof major depressive disorder:a magnetic resonance imagingvolumetric study

J. De Diego-Adelino1 °, M. Carceller1, M. Serra-Blasco1,Y. Vives-Gilabert2, B. Gomez-Anson3, D. Puigdemont4,E. Alvarez4, V. Perez4, M.J. Portella4. 1Hospital SantPau CIBERSAM UAB, Dept. of Psychiatry, Barcelona,Spain; 2UAB, Port d’Informacio Cientıfica (PIC),Barcelona, Spain; 3Hospital Sant Pau CIBERNEDUAB, Neuroradiology, Barcelona, Spain; 4Hospital SantPau CIBERSAM UAB, Psychiatry, Barcelona, Spain

Purpose of the study: Habenular nucleus (HbN) isan epithalamic structure involved in stress-response,anxiety and reward-processing. It receives frontolimbicand hippocampal afferences and also projects inhibitory


fibres to the brainstem monoaminergic nuclei, thus it mightplay an important role in the pathophysiology of MajorDepressive Disorder (MDD) [1]. Prior studies [1−3] havereported functional and volumetric abnormalities in MDD,however, habenular volume has been hardly comparedbetween different stages of depressive disease leaving apartprocesses related with chronicity and treatment resistance.We aimed to investigate whether habenular volume

differed between patients with MDD in distinct stagesof the illness and healthy controls. We hypothesizedthat chronic/treatment-resistant depression would associatesmaller habenular volumes.

Methods: A cross-sectional structural neuroimagingwith a 3-T Philips Achieva scanner was conducted. Samplewas composed by 61 outpatients with MDD (DSM-IV) indifferent stages of illness (First-episode, n = 21; Remitted-recurrent MDD, n = 20; Chronic/treatment-resistant MDD,n = 20) and 34 healthy controls. All participants wereright-handed and groups were comparable in age, sexand educational level. HbN were manually segmented bytwo researchers blinded for the clinical data. Inter-raterkappa coefficients were 0.83 (p = 0.006) and 0.8 (p = 0.01)for right and left habenula, respectively. Total, greymatter (GM) and white matter (WM) habenular volumeswere calculated for each participant.

Results: Total habenular volume did not showsignificant differences among groups. A significant groupeffect was observed in right habenular WM volume(F = 2.895; df = 3,91; p = 0.039), but not for the rest ofcomparisons. Post-hoc t-test showed that patients with afirst-episode had higher WM volume in the right habenulaas compared to healthy controls (p = 0.046) and to chronicpatients (p = 0.03). Similar findings were seen for theleft HbN, but the differences did not reach statisticalsignificance.

Conclusions:Although no differences in total habenularvolumes were found across the groups, our findings ofabnormalities in WM volume provide some evidencefor the potential involvement of the habenula in MDD.Particularly, we observed higher habenular WM volumesin patients with a first-episode than in healthy controlsand patients with chronic/treatment-resistance MDD. ThisWM hypertrophy could be related with the habenularhyperactivity described among patients with depressionby functional MRI studies [2] and might be part of theneural substrate of MDD in early stages. By contrast,the smaller WM volumes observed in later phases of theillness, especially in chronic patients, suggest that changesin WM volumes within this structure might play a rolein the treatment response. In fact, HbN has been targetedfor deep brain stimulation in treatment-resistant MDD [1].Because of the cross-sectional design of the present study,it would remain unclear whether small WM habenular

volume would be a vulnerability marker for treatment-resistance or whether a progressive atrophy would occuralong the course of the illness (perhaps due to an overuseof the structure).The conclusions should be taken cautiously because pa-

tients were on medication. Additionaly, though reliabilityinter-rater coefficients were good, habenular nucleus is alittle structure highly attached to surrounding nuclei, whichcould reduce the likelihood of finding further differences.

Reference(s)

[1] Henn, F.A., 2012. Circuits, cells, and synapses: towarda new target for deep brain stimulation in depression.Neuropsychopharmacology 37, 307–108.

[2] Roiser, J.P., Levy, J., Fromm, S.J., Nugent, A.C.,Talagala, S.L., Hasler, G., Henn, F.A., Sahakian, B.J.,Drevets, W.C., 2009. The effects of tryptophandepletion on neural responses to emotional words inremitted depression. Biol Psychiatry 66, 441−50.

[3] Savitz, J.B., Nugent, A.C., Bogers, W., Roiser, J.P.,Bain, E.E., Neumeister, A., Zarate, C.A. Jr,Manji, H.K., Cannon, D.M., Marrett, S., Henn, F.,Charney, D.S., Drevets, W.C., 2011. Habenula volumein bipolar disorder and major depressive disorder:a high-resolution magnetic resonance imaging study.Biol Psychiatry 69, 336–343.

P.4.024 Using positron emission tomographyto investigate microglial activationin alcohol dependence: preliminaryfindings

N.J. Kalk1 °, Q. Guo2, R. Cherian3, D. Erritzoe4,A. Waldman5, K. Dar3, R.N. Gunn2, D.J. Nutt1,E.A. Rabiner2, A.R. Lingford-Hughes1. 1ImperialCollege London, Centre for Neuropsychopharmacology,London, United Kingdom; 2Imanova Limited, Imaging,London, United Kingdom; 3Central North West LondonNHS Foundation Trust, Addiction Psychiatry, London,United Kingdom; 4Imperial College, Centre forNeuropsychopharmacology, London, United Kingdom;5Imperial College, Medicine, London, United Kingdom

There is pre-clinical evidence for microglial activationin alcohol dependence and withdrawal which may relateto alcohol-induced cognitive impairment [1]. An increasein microglia was found in human brain tissue fromalcoholics [2]. Microglial activation can be measured inhumans in vivo using Positron Emission Tomography(PET) tracers that bind to the 18kDa Translocator Protein(TSPO) which has relatively low constitutive expression inthe brain but is richly expressed in activated microglia. Weare investigating whether there is microglial activation in


male human alcoholics, shortly after detoxification, using[C-11]PBR28 PET in a group comparison study.13 healthy controls and 7 alcoholics were recruited.

[11-C]PBR28’s affinity for the TSPO varies according tothe presence of a single nucleotide polymorphism on oneor both alleles (3). Therefore, participants were genotypedprior to scanning to facilitate interpretation of PET data.Dynamic PET data were acquired for 90 minutes followingbolus injection of 331.9±15.27 MBq of [11-C]PBR28.Arterial plasma activity was measured and corrected forthe presence of metabolites.PET data were co-registered to the participant’s

structural MRI scan which facilitated the application ofan in-house anatomical atlas via spatial normalisation todefine regions of interest (ROIs): cortical grey matter,the anterior cingulate cortex, hippocampus, thalamus andmidbrain. ROIs were applied to the dynamic PET data togenerate time activity curves. A two tissue compartmentalmodel with a fixed blood volume (5%) was used toestimate regional volumes of distribution (VT). Two-wayANCOVAs were performed to examine the effect ofgenotype and disease status (alcohol dependent vs healthy)in each ROI. Age was included as a co-variate, as TSPObinding is known to increase with age.Data from 13 healthy control participants and 5 patients

are reported. The average age of the alcoholic group was49±10 (n = 5) and the healthy group was 54±10 (n = 13).The average time from detoxification in the patient groupwas 25±9 days. Four healthy controls, and four alcoholics,were dependent on nicotine. Two alcoholics reported ahistory of cocaine use, but none met the criteria fordependence on cocaine or other illicit drugs.Mean cortical VT in the healthy control group

was 5.0±1.35 vs. 4.7±2.0 in the alcoholic group(CC genotype), and 3.31±0.79 in the healthy group vs3.3±0.36 in the alcoholic group (CT genotype). TheANCOVA found a significant effect of age across allregions studied (cortical grey matter F = 7.331, p = 0.018;anterior cingulate cortex F = 5.297, p = 0.040; thalamusF = 5.972 p = 0.030, hippocampus F = 6.151 p = 0.028;midbrain F = 5.811, p = 0.031) and an effect of genotypeapproaching or reaching significance in the cortexand thalamus (cortical grey matter F = 3.617, p = 0.080;thalamus F = 5.740, p = 0.032) findings which replicateprevious work. No significant effect of group was found,but this may reflect insufficient power. Further recruitmentis ongoing.

Reference(s)

[1] Obernier, J.A., White, A.M., Swartzwelder, H.S.,Crews, F.T., 2009. Cognitive deficits and CNS damageafter a 4-day binge ethanol exposure in rats. PharmacolBiochem Behav 72(3): 521−32.

[2] He, J., Crews, F.T., 2008. Increased MCP-1 andmicroglia in various regions of the human alcoholicbrain. Exp Neurol 210(2): 349−58.

[3] Owen, D.R., Yeo, A.J., Gunn, R.N., Song, K.,Wadsworth, G., Lewis, A., Rhodes, C., Pulford, D.J.,Bennacef, I., Parker, C.A., StJean, P.L., Cardon, L.R.,Mooser, V.E., Matthews, P.M., Rabiner, E.A., Ru-bio, J.P., 2012. An 18-kDa translocator protein (TSPO)polymorphism explains differences in binding affinityof the PET radioligand PBR28. J Cereb Blood FlowMetab 32(1): 1−5.

P.4.025 Effects of deep brain stimulation ofprelimbic and infralimbic areas of theprefrontal cortex of the rat

L. Jimenez-Sanchez1 °, L. Perez-Caballero2,A. Castane1, X. Lopez-Gil1, L. Campa1,M. Galofre1, E. Berrocoso3, A. Adell1. 1IIBB-CSIC-CIBERSAM, Department of Neurochemistry andNeuropharmacology, Barcelona, Spain; 2University ofCadiz-CIBERSAM, Neuropsychopharmacology ResearchGroup Department of Neuroscience (Pharmacologyand Psychiatry), Cadiz, Spain; 3University of Cadiz-CIBERSAM, Neuropsychopharmacology Research GroupPsychobiology Area Department of Psychology, Cadiz,Spain

Neuroimaging studies have shown a hyperactivity ofsubgenual cingulate region (Cg25) in major depression [1].Further, normalisation of this activity has been asso-ciated with effective clinical response to a variety ofantidepressant treatments. Deep brain stimulation (DBS)also reduced the hyperactivity of Cg25, and has provedefficacious for treatment-resistant depression patients [1].In spite of its therapeutic action, little is known about theprecise mechanism of DBS. In the present work, we haveused in vivo microdialysis to examine the effects of 1-hourDBS (130 Hz frequency, 200mA amplitude and 90mspulse width) applied to prelimbic (PrL) or infralimbic(IL) areas of the prefrontal cortex (PFC) on the releaseof serotonin (5-HT), dopamine (DA), noradrenaline (NA)and glutamate (Glu) in the entire PFC of male Wistar rats.Neuronal activation throughout the brain after PrL or ILDBS was also examined by c-Fos immunohistochemistry.The antidepressant-like effect of DBS was measured usingthe forced swim test (FST).The results show that DBS of the IL region exerts

an antidepressant-like action in the FST (measured bya decrease in immobility and an increase in swimmingand climbing) that is associated with increased releases of5-HT, DA, NA and Glu in the PFC. In addition, the effects


of IL DBS on cortical glutamate (but not on 5-HT), as wellas its antidepressant effect in the FST, are dependent uponthe activation of AMPA receptors, inasmuch as they wereprevented by the systemic administration of the AMPAreceptor antagonist, NBQX. However, DBS of the PrLdisplayed a depressogenic-like action in the FST measuredas increased immobility and decreased swimming andclimbing. Further, PrL DBS reduced the release of DA,NA and Glu in the PFC. On the other hand, importantdifferences in the immunolabelling of c-Fos were observedafter 1-h DBS of PrL or IL regions. Thus, after IL DBS,c-Fos expression was higher in the basolateral nucleus ofthe amygdala, and mediodorsal and centromedial thalamicnuclei, and lower in the habenula and shell of the nucleusaccumbens compared with DBS PrL.To determine whether the actions of IL DBS were due

to a local effect in the IL subdivision of the PFC or to theconnections between IL and subcortical areas of the brain,a cortical transection was performed. The results showedthat IL DBS potentiated the antidepressant-like effectsof the cortical transection in the FST, thus suggestingthat a local action of DBS is responsible of its rapidantidepressant-like effect.We conclude that the early antidepressant-like action

of DBS of the IL region of the PFC would depend onthe activation of AMPA receptor-dependent glutamatergictransmission. The increase of monoaminergic transmissionin the PFC and the activity of subcortical projectionareas would play an important role in the long-termantidepressant-like effect of IL DBS.

Reference(s)

[1] Mayberg, H.S., Lozano, A.M., Voon, V., McNeely,H.E., Seminowicz, D., Hamani, C., Schwalb, J.M.,Kennedy, S.H., 2005. Deep brain stimulation fortreatment-resistant depression. Neuron 45, 651–660.

P.4.027 Six-month follow-up study of repetitivetranscranial magnetic stimulation in thetreatment of resistant major depressivedisorder

C. Concerto1 °, D. Cannavo1, F. Magnano SanLio1,R. Ricceri2, G. Lanza2, E. Aguglia1. 1University ofCatania, A.O.U. Policlinico-Vittorio Emanuele U.O.P.I.of Psychiatry, Catania, Italy; 2University of Catania,A.O.U. Policlinico-Vittorio Emanuele Department ofNeuroscience, Catania, Italy

Purpose of the study: Over the last years TranscranialMagnetic Stimulation (TMS) has been investigated tostudy motor pathways and motor cortical excitability

in healthy subjects and in patients with neurologicaldisorders. Several clinical studies have been focussedon the efficacy of Repetitive Transcranial MagneticStimulation (rTMS) in the treatment of Resistant MajorDepression which is also the psychiatric field whererTMS has received more indications and approvalsworldwide. In the treatment of pharmaco-resistant majordepression, interventions strategy include combinationof antidepressant drugs with different mechanisms ofaction and augmentation of somatic treatments topharmacological therapy. In this regard, the use of rTMSas add-on treatment truly represents one of the major fociin clinical research.

Objectives: Object of this study was to evaluate thelong-term effect on depressive symptoms and frontal lobeabilities of rTMS as add-on therapy in the treatment ofdrugs resistant major depression [1].

Methods: A sample of fifteen drug resistant depressedoutpatients meeting the Diagnostic and Statistical Manualof Mental Disorders, Fourth Edition, Text Revision criteriafor nonpsychotic major depressive disorder were assignedto receive fifteen sessions of high frequency rTMSstimulation of the left dorsolateral prefrontal cortex. ActiverTMS was performed five days per week for four weeksconsecutively. Each session of rTMS was delivered asfollows: 10 Hz, four-second-train duration, a 26-secondinter-train interval, for a total of 3000 pulses per sessionlasting 37.5 minutes. The intensity of the magneticstimulus was set at 120% of the resting motor threshold.During the course of rTMS, the patients were taking theirmedication which included Selective Serotonin ReuptakeInhibitors (SSRI), Tricyclic Antidepressant and AtypicalAntipsychotic Drugs. Their doses were required to remainstable in the four weeks preceding the trial and for itsentire duration. Patients were followed up for six months todetermine the long-term effect of rTMS as add-on therapy.Outcome measures for the evaluation of depressivesymptoms consisted of Hamilton Depression RatingScale 21-item (HAM-D) and the Montgomery AsbergDepression Rating Scale (MADRS). A neuropsychologicalbattery for the evaluation of different frontal lobe abilitiesincluded the Frontal Assessment Battery and the StroopColor Word Test interference. Effectiveness data wasgathered at baseline (T0), at the end of treatment (T1)and 6 months after the end of treatment (T2) [2].

Results: Results from the present study showedstatistically significant mood improvements as indexedby a reduction of more than 40% on the HamiltonRating Scale for Depression and on Montgomery AsbergDepression Rating Scale at the end of treatment and sixmonths after treatment. Frontal Assessment Battery andthe Stroop Color Word Test interference outcome scores


didn’t significantly differ at treatment end and after sixmonths.

Conclusion: Our findings demonstrate the antidepres-sant effect of adjunctive Repetitive Transcranial MagneticStimulation treatment. Moreover this study confirmsgood tolerability of rTMS and the maintenance of theantidepressant effect six months after the treatment, asshown by a decrease in HAM-D and MADRS [3].

Reference(s)

[1] Sackeim, H.A., 2001. The definition and meaning oftreatment-resistant depression. J Clin Psychiatry 62,10−17.

[2] Giupponi, G., Pycha, R., Dell’Osso, B., Pompili, M.,Walpoth, M., Hausmann, A., Di Pauli, J., Erfurth, A.,Conca, A., 2009. Neurophysiological and neuropsy-chiatric aspects of transcranial magnetic stimulation.Clinical Neuropsychiatry 6, 234–235.

[3] Avery, D.H., Holtzheimer, P.E., Fawaz, W., Russo, J.,Neumaier, J., Dunner, D.L., Haynor, D.R., Clay-poole, K.H., Wajdik, C., Roy-Byrne, P., 2010. Acontrolled study of repetitive transcranial magneticstimulation in medication-resistant major depression.Biol Psychiatry 59, 187–194.

P.4.028 Neural correlates of stress-relatedhypothalamic–pituitary–adrenal axisactivity in neuroticism

A. Boehringer1 °, F. Lederbogen1, H. Tost1, L. Haddad1,A. Meyer-Lindenberg1. 1Central Institute of MentalHealth, Systems Neuroscience in Psychiatry, Mannheim,Germany

Neuroticism, the stable tendency to react with negativeemotional responses to threat, frustration, or loss, isassociated with blunted hypothalamic–pituitary–adrenal(HPA) axis responses to stress [1]. Dysfunctionalinteractions between the perigenual anterior cingulatecortex (pACC) and the amygdala have been identifiedas a neural correlate of neuroticism related personalitytraits [2]. The amygdala is implicated in HPA axisactivation and the pACC inhibits amygdala function.Changes in pACC–amygdala brain circuitry may thereforeaccount for the relationship between neuroticism andHPA axis dysfunction. Here, we tested this hypothesisin a sample of healthy individuals using an establishedsocial stress paradigm and functional magnetic resonanceimaging (fMRI).Thirty-two (16 males, mean±SD 43.6±14.4 years)

healthy participants were exposed to a psychosocialstress task (Montreal Imaging Stress Task, MIST) while

their brains were scanned using fMRI. During thistask, participants were asked to solve mental arithmetictask under time pressure and negative social evaluativefeedback from the study investigators. HPA axis responses(as assessed by salivary cortisol) as well as cardiovascularand subjective stress responses to the MIST were measuredrepeatedly. At the neural level, effects of stress on brainactivity as well as functional connectivity between thepACC and the amygdala were assessed.At the group level, the MIST induced significant

increases in HPA axis and cardiovascular activity, as wellas subjective feelings of stress (all P< 0.001). None ofthese stress measures was significantly associated withneuroticism. In line with previous work, variability inthe endocrine stress response was high, allowing tosplit the study group into participants who reacted witha substantial increase in cortisol (cortisol responders,n = 18) and others who did not respond (cortisol non-responders, n = 13) using a cut-off of 2.5 nmol/l increasein cortisol. Cortisol responders to the stressor showedlower stress related activity in the perigenual anteriorcingulate cortex (pACC) than cortisol non-responders,suggesting a role of the pACC in HPA axis inhibition(P< 0.05, family-wise error corrected (FEW) for pACCand amygdala volume). Deactivation of the pACC withinthe cortisol responder group was less pronounced inindividuals high in neuroticism (P< 0.05, FEW for pACCand amygdala volume), indicating that high neuroticismwas associated with less release of inhibitory tone of thepACC on the HPA axis. Moreover, rostral pACC–amygdalafunctional connectivity was higher in high neuroticismcortisol responders (p< 0.05 FEW corrected for amygdalavolume), suggesting that changes in pACC–amygdalabrain circuitry may contribute to altered HPA axis controlin individuals high in neuroticism.Here, we identify a neural correlate of altered HPA

axis control in cortisol responders high in neuroticism.The presented data provide further evidence for a specificrole of pACC in the neural processing of social stressand propose a neural mechanism by which altered pACC–amygdala functional interaction translates into an increasein inhibitory control of the HPA axis activity in individualswith maladaptive emotional personality characteristics.These findings may improve our understanding of thewell-established association between neuroticism, stress,and various mental diseases [3].

Reference(s)

[1] Oswald et al., Neuropsychopharmacology 2006; 31:1583−91.

[2] Pezawas et al., Nat Neurosci 2005; 8: 828−34.[3] Lahey American Psychologist 2009; 64: 241−56.


P.4.029 Antioxidant status and fatty acids inadolescents with Asperger syndromeand first episodes of early-onsetpsychosis

M.G. Moron-Nozaleda1 °, M. Alvarez-Blazquez1,C.M. Dıaz-Caneja1, E. Dulin2, C. Moreno1,M.C. Guisasola3, C. Arango1, M. Parellada1. 1HospitalGeneral Universitario Gregorio Maranon, Child andAdolescent Psychiatry Department, Madrid, Spain;2Hospital General Universitario Gregorio Maranon,Biochemistry Department, Madrid, Spain; 3HospitalGeneral Universitario Gregorio Maranon, ExperimentalMedical and Surgery Unit, Madrid, Spain

Purpose of the study: Evidence suggests that oxidativestress-mediated cell membrane pathology and impairmentof lipid metabolism may be involved in the physiopathol-ogy of autism spectrum disorders and schizophrenia [1,2].Several studies have reported significantly reduced levelsof polyunsaturated fatty acids (PUFAs) in plasma and redblood cell membranes from patients with schizophreniaand autism [3]. The aim of this study was to explorethe differences in oxidative status and plasma PUFAsas indicators of cell membrane integrity in two groupsof adolescents, one with Asperger syndrome (AS) andanother with a first episode of early-onset psychosis, andcompare them with healthy controls.

Methods: Twenty-four adolescents with AS (mean age12.7±2.5 years, 95.8% male), 24 with a first episode ofpsychosis (mean age 15.9±1.2 years, 66.7% male) and 23controls (mean age 13.1±3 years, 87% male) participatedin the study. Inclusion criteria: age 7−17 years; diagnosisof first episode early onset psychosis or AS. Exclusioncriteria: mental retardation, neurological disorders, historyof head trauma and pregnancy. Fasting venous bloodsamples were collected into EDTA evacuated tubes.After immediate centrifugation, plasma and whole bloodaliquots were transferred into cryogenic tubes and storedfrozen at −80ºC. PUFAs were analysed in plasma usinga gas chromatograph. Plasma Total Antioxidant Status(TAOS) was measured with a TAOS Assay Kit. Forthe statistical analysis, because a number of measuredvariables did not come for normal distribution, they wereanalysed using non-parametric tests (Kruskal–Wallis andMann–Whitney).

Summary of results: PUFA composition in plasmaand TAOS levels are shown in Table 1. The plasmaTAOS was statistically significantly lower in the AS groupcompared with the healthy controls and with the psychosispatients. No differences were found in total PUFAsor any PUFA individually, except for eicosapentaenoic

acid in psychosis. However, significant differences werefound in total phospholipids both in AS and psychosisas compared with the controls. The AA/DHA ratio(arachidonic acid n = 6/docosahexaenoic acid n = 3) wassignificantly higher in AS.

Conclusion: Reduced antioxidant defence, pro-oxidantimbalance and abnormal cell membrane composition arefound in AS. Abnormal lipid composition may alsocontribute to the pathophysiology of psychosis. Furtherresearch will assess the role of drugs that act at theoxidative pathway in these disorders.

Table 1. PUFA composition in plasma and TAOS levels

Asperger Psychosis Control p

Betweengroups

AspergervsControl

AspergervsPsychosis

Psychosisvs Control

Total PUFAs 519.28±182.9 559.05±206.43 490.57±206.21 0.183 0.088 0.584 0.158Total PL 0.56±0.20 0.68±0.48 0.39±0.39 0.036 0.041 0.184 0.030AA/EPA 0.98±1.58 0.79±0.71 0.57±0.36 0.591 0.596 0.389 0.399AA/DHA 5.6±2.16 4.68±2.47 3.86±1.12 0.016 0.004 0.162 0.184AA 4.94±1.05 4.58±1.35 4.42±1 0.484 0.259 0.360 0.853EPA 7.48±1.93 6.89±2.05 8.72±2.27 0.109 0.239 0.340 0.038DHA 1.03±0.53 1.32±1.00 1.22±0.47 0.156 0.099 0.097 0.797TAOS 1.13±0.21 1.28±0.28 1.26±0.25 0.017 0.043 0.007 0.312

Results are shown as mean±SD. PL: Phospholipids; AA: Arachidonic acid; EPA: Eicosapentaenoic acid;DHA: Docosahexaenoic acid.

Reference(s)

[1] Parellada M, et al., Plasma antioxidant capac-ity is reduced in Asperger syndrome, Journalof Psychiatric Research (2011), doi: 10.1016/j.jpsychires.2011.10.004.

[2] Akyol O, Herken H, Uz E, Fadillioglu E, Unal S,Sogut S, et al. The indices of endogenous oxidative andantioxidative processes in plasma from schizophrenicpatients. The possible role of oxidant/antioxidantimbalance. Progress in Neuropsychopharmacology &Biological Psychiatry 2002; 26: 995e1005.

[3] Ming X, Stein TP, BrimacombeM, JohnsonWG, Lam-bert GH, Wagner GC. Increased excretion of a lipidperoxidation biomarker in autism. Prostaglandins,leukotrienes and essential fatty acids 2005; 73:379e84.


P.4.030 Bright light dose correlates with changein striatal serotonin transporter bindingin healthy Scandinavians

B. Mc Mahon1 °, A.S. Andersen1, L. Feng1, K.K. Holst1,M.K. Madsen1, S. Lehel2, M.M. Herth2, P. Iversen3,L. Hasholt4, G.M. Knudsen1. 1Copenhagen UniversityHospital Rigshospitalet, Neurobiology Research Unitand Center for Integrated Molecular Brain Imaging,Copenhagen, Denmark; 2Copenhagen University HospitalRigshospitalet, PET and Cyclotron Unit, Copenhagen,Denmark; 3Copenhagen University Hospital HvidovreHospital, Center for Integrated Molecular Brain Imagingand Danish Research Centre for MR, Copenhagen,Denmark; 4Copenhagen University, Department ofcellular and Molecular Medicine, Copenhagen, Denmark

Background: Lack of daylight is prominent at highlatitudes and this may be perceived as an environmentalstressor associated with a high frequency of seasonalaffective disorder that can be treated with brightlight therapy. We and others have in cross-sectionalstudies observed season dependent fluctuations in theserotonin transporter (SERT), with high striatal bindingaround winter solstice and low binding around summersolstice. In addition, we have identified a genotype-dependent interaction with these environmental stressorswith SERT binding being dependent on the carrierstatus of the 5-HTTLPR promoter polymorphism [1].This gene*environment paradigm predicts the SERTfluctuations with a negative correlation between SERTbinding and daylight minutes in carriers of the short5-HTTLPR allele (S-allele), but less so in homozygotecarriers of the long allele (L-allele). The aim of thepresent study was to examine the correlation between theintensity of bright light and change in striatal bindingpotential (BPND) and if any such change was dependingon 5-HTTLPR carrier status. We hypothesize that there is anegative correlation between change in striatal SERT BPNDand lux dose and that this correlation is more prominentin carriers of the S-allele.

Method: Seventy healthy males were pre-screenedfor inclusion and based on their 5-HTTLPR trialleliccarrier status they were invited for participation. Ofthese, 24 healthy male volunteers (mean age: 24±4.5,range 18−36 years and mean minutes of daylight atbaseline: 449±25, range 421–504min) participated in thestudy. Exclusion criteria were significant medical history,known retinal pathology and the use of photosensitisingmedications. Eight La-homozygotes and 16 S-allelecarriers was exposed to a bright light intervention atvarying degrees (mean intensity: 4112±2851, range 131–

10.971 lux) for three weeks. Post intervention all subjectsreceived a visit where the distance and angle to thelight source was measured. In addition, a semi-structuredinterview was conducted to assess compliance, subjectiveeffects and daily outdoor activities. The intensity of allbiolamps was individually determined with respect todistance and angle with an Elma 1335 luxmeter. Theintensity and spectrum of the biolamps will be furthercharacterised by the Department of Photonics EngineeringDTU and the data are considered preliminary.

Results: Participants were investigated with MRIand [11C]DASB on an HRRT PET scanner (meaninj. dose: 589±29MBq, range 421–609) before and afterintervention. Quantification was done with MRTM2. Asignificant positive correlation was found in striatum(slope 2.501e-005 ±1.027e-005, R square: 0.2203) forall subjects. When subdividing subjects with respectto 5-HTTLPR genotype status we saw a tendencyfor a positive correlation in the S-allele carrier group(slope: 2.328e-005 ±1.224e-005, R square: 0.21) butnot in the L-allele homozygote group (slope: 3.221e-005±2.296e-005, R square: 0.28).

Conclusion: Contrary to the summer down regulationof SERT, we found that a smaller amount of bright lightup regulated the SERT suggesting that the relationshipbetween bright light dose and SERT change might not beexplained by a linear model.

Reference(s)

[1] Kalbitzer, J., Erritzoe, D., Holst, K.K., Nielsen, F.A.,Marner, L., Lehel, S., Arentzen, T., Jernigan, T.L.,Knudsen, G.M., 2010. Seasonal changes in brainserotonin transporter binding in short 5-HTTLPR-allele carriers but not in long-allele homozygotes. BiolPsychiatry 67, 1033−9.

P.4.031 Effects of mental distress on cognitivefunctioning in patients admitted forcardiac rehabilitation after acutecoronary events

J. Burkauskas1 °, J. Brozaitiene1, R. Bunevicius1.1Lithuanian University of Health Sciences, Institute ofBehavioral Medicine, Palanga, Lithuania

Introduction: Impaired cognition increases mortality inCAD patients, even in cases of mild impairment [1].Thus, it is important to investigate and determine factorsassociated with cognitive functioning in this particularpopulation. Our recent findings in patients indicate thatpatient-oriented outcomes including health related qualityof life [2] and fatigue [3] are associated with mental


distress factors such as depression and anxiety. Thequestion of how mental distress might be specificallyassociated with cognitive functioning received less ofattention. The aim of our study was to investigate anassociation of cognitive functioning with mental distressin CAD patients.

Methods: This study was conducted over a 3-yearperiod examining CAD patients two weeks after acutemyocardial infarction or unstable angina attending cardiacrehabilitation programme. 539 patients participated in thestudy; 386 (72%) men and 153 (28%) women with a meanage of 58 years (SD= 9). Participants were evaluated forheart functional class, following the guidelines providedby the New York Heart Association. Mini Mental StateExamination (MMSE) was used to asess global cognitivefunctioning. Digit Span Test and Digit Symbol Testwere used to assess auditory attention, mental flexibility,psychomotor performance and incidental learning. TrailMaking Test A and B was used to measure perceptualspeed, task switching and executive control. Mentaldistress factors were evaluated using the Beck DepressionInventory-II (BDI-II) to measure depressive symptoms,the State-Trait Anxiety Inventory (STAI) to measurestate anxiety (STAI-S) and trait anxiety (STAI-T), andType-D Scale (DS-14) to measure Type-D personalitycharacteristics.

Results: In univariate regression analysis almost allmental distress, socio-demographic and clinical character-istics were significantly associated with different cognitivefunctions. After adjusting for gender, age, education, NewYork Heart Association functional class, MMSE scoresremained associated with DS-14 (b =−0.144, p< 0.01);Digit Span Test Backward recall of digits, with STAI-S(b =−0.120, p< 0.01). Digit symbol test raw scores wereassociated with BDI-II (b =−0.115, p< 0.01), as well aswith STAI-S (b =−0.084, p = 0.03). Similarly, time whichindividuals took to complete the Digit Symbol Test wasassociated with BDI-II (b = 0.145, p< 0.01), and STAI-S(b = 0.110, p< 0.01). Significant associations remainedbetween Trail Making Test A scores and STAI-S scores(b = 0.113, p< 0.01) as well as between Trail MakingTest B scores and STAI-S scores (b = 0.103, p = 0.04),while Trail Making Test B-A scores remained associatedwith STAI-T scores (b =−0.119, p< 0.03).

Conclusions: In non-demented CAD patients twoweeks after acute cardiac events cognitive functioning isassociated with mental distress independently from cardiacfunctional class, gender, age and education. Specifically,Type-D personality worsens global cognitive functioning.The presence of symptoms of depression and higher levelsof situational anxiety negatively affects test completiontime and psychomotor performance. Higher levels ofsituational anxiety also results in decrease of perceptual

speed, mental flexibility and task switching while higherlevels of trait anxiety improves executive control.

Reference(s)

[1] Shavelle, R., Paculdo, D., Strauss, D., Kush S.,2009 Cognitive Impairment and Mortality in theCardiovascular Health Study. J Insur Med. 41(2), 110–116.

[2] Staniute, M.J., Brozaitiene, J., Bunevicius, R., 2011Effects of Social Support and Stressful Life Eventson Health-Related Quality of Life in Coronary ArteryDisease Patients. J Cardiovasc Nurs. [Epub ahead ofprint].

[3] Bunevicius, A., Stankus, A., Brozaitiene, J., Girdler,S.S., Bunevicius, R., 2012 Relationship of fatigue andexercise capacity with emotional and physical statein patients with coronary artery disease admitted forrehabilitation program. Am Heart J. 162(2), 310–316.

P.4.032 Neuroinflammation in temporal cortex inschizophrenia patients

T.F. van der Doef1 °, M. Yaqub2, M.G. Bossong1,R. Boellaard2, A.D. Windhorst2, A.A. Lammertsma2,R.S. Kahn1, B.N.M. van Berckel2. 1University MedicalCenter Utrecht, Psychiatry, Utrecht, The Netherlands;2VU University Medical Center, Nuclear Medicine andPET Research, Amsterdam, The Netherlands

Background: There is increasing evidence that neuroin-flammation is associated with schizophrenia. Neuroinflam-mation is characterised by the activation of microglialcells. Increased expression of the translocator protein is abiomarker for microglial activation and can be measuredin vivo using the positron emission tomography ligand(R)-[11C]PK11195. The purpose of this study was tocompare the regional distribution of (R)-[11C]PK11195binding in patients with schizophrenia to that in healthycontrols.

Methods: (R)-[11C]PK11195 binding potential (BPND)was assessed in ten patients with schizophrenia andten age-matched healthy controls. Psychopathology wasmeasured using the Positive and Negative Syndrome Scale(PANSS). All patients received atypical antipsychoticdrugs. Dynamic (R)-[11C]PK11195 scans were acquiredusing an ECAT EXACT HR+ scanner.PET data were analysed using receptor parametric

mapping (RPM), a basis function implementation of thesimplified reference tissue model (SRTM). Supervisedcluster analysis (SCVA) algorithm was used to extractthe reference tissue input function from the dynamicPET scans [1,2]. We used a supervised cluster method with


four kinetic classes (SCVA4), which is a revised versionof the supervised cluster analysis approach of Turkheimeret al. [1,2]. The outcome measure was (R)-[11C]PK11195binding potential.Subsequently, grey matter regions of interest (ROIs)

were delineated on a T1-weighted structural MRI scanusing an automatic procedure, resulting in the followingregions: frontal, temporal, parietal and occipital cortex,and cerebellum. Regional values of BPND were obtained byprojecting the ROIs onto the parametric (R)-[11C]PK11195BPND images.Multivariate analysis of variance (MANOVA) was used

to test for differences in binding potential between patientsand controls with group as the between-subjects factor andregion of interest as the within-subjects factor.

Results: MANOVA showed an overall significant effectof group (F(5) = 3.7, p = 0.03). Schizophrenia patientsshowed increased (R)-[11C]PK11195 binding potential inthe temporal cortex (F(1) = 6.1, p = 0.02) (Table 1). Therewere no significant differences in mean (R)-[11C]PK11195binding potential in the other areas tested (Table 1).Patients with schizophrenia had minimal to moderatesymptoms of the disease at the time of PET scanning(PANSS total score = 52.5±9.5).Table 1. Regional (R)-[11C]PK11195 BPND values

Region Mean BPND ±SD General Linear Model

Controls Schizophrenia F p

Frontal 0.11±0.09 0.10±0.06 0.05 0.82Temporal 0.03±0.08 0.11±0.08 6.06 0.02*Parietal 0.15±0.11 0.13±0.06 0.20 0.66Occipital 0.33±0.13 0.35±0.14 0.10 0.75Cerebellum 0.16±0.08 0.15±0.13 0.06 0.82

*p� 0.05.

Conclusions:An increase in (R)-[11C]PK11195 bindingpotential was found in the temporal cortex in schizophreniapatients, suggesting focal microglial activation. This inturn provides preliminary evidence for neuroinflammationin the temporal cortex in schizophrenia during the firstyears of the illness. This may provide an explanationfor progressive tissue loss in this area in schizophrenia.Further studies are warranted to assess anti-inflammatorytreatment in this disease.

Reference(s)

[1] Turkheimer, F.E., Edison, P., Pavese, N., Roncaroli, F.,Anderson, A.N., Hammers, A., Gerhard, A., Hinz, R.,Tai, Y.F. and Brooks, D.J., 2007. Reference and targetregion modeling of [11C]-(R)-PK11195 brain studies.J Nucl Med. 48, 158–167.

[2] Yaqub, M., van Berckel, B.N., Schuitemaker, A., Hinz,R., Turkheimer, F.E., Tomasi, G., Lammertsma, A.A.

and Boellaard, R., 2012. Optimization of supervisedcluster analysis for extracting reference tissue inputcurves in (R)-[(11)C]PK11195 brain PET studies. JCereb Blood Flow Metab. 32, 1600–1608.

P.4.033 Cerebrospinal fluid biomarkers of braininjury in bipolar disorder

J. Jakobsson1 °, E. Palsson1, C.J. Ekman2, C. Sellgren3,A.G.M. Johansson2, H. Zetterberg1, K. Blennow1,M. Landen1. 1The Sahlgrenska Academy Universityof Gothenburg, Department of Psychiatry andNeurochemistry, Goteborg, Sweden; 2KarolinskaInstitutet, Department of Clinical Neuroscience,Stockholm, Sweden; 3Karolinska Institutet, Departmentof Medical Epidemiology and Biostatistics, Stockholm,Sweden

Purpose of the study: Bipolar disorder is a commonpsychiatric disorder characterised by recurrent episodesof mania/hypomania and depression. Structural imagingstudies suggest that bipolar disorder is associated withmorphological abnormalities of the brain. The mostrecurrent finding is that patients with bipolar disorder havelateral ventricular enlargement and increased rates of deepwhite matter hyperintensities [1]. Furthermore, severalstudies suggest that structural brain changes are associatedwith a decline in cognitive function. We previouslyinvestigated a neurodegenerative component of bipolardisorder by analysing several cerebrospinal fluid (CSF)biomarkers for neurodegenerative processes in bipolarpatients and healthy controls [2]. No major alterations wereobserved between patients and controls thus excludingneurodegenerative processes related to these biomarkers(T-tau, P-tau, and amyloid beta). There are, however,other established CSF biomarkers reflecting injuriesin different cells/structures in brain. These biomarkersinclude: neurofilament light chain (NF-L), Heart-typefatty acid binding protein (H-FABP), calcium bindingprotein S100B, and myelin basic protein (MBP) [3]. Inthis study the relation between CSF biomarkers of braininjuries and bipolar disorder were investigated. The aimwas to test the suggested neuroprogressive component inbipolar disorder and to investigate possible associationsbetween CSF biomarkers and disease severity, ongoingmedications, and/or cognitive functions.

Methods: 137 patients were recruited from theSt. Goran bipolar project, enrolling patients from thebipolar unit at the Northern Stockholm Psychiatric Clinic,Stockholm, Sweden. 86 age and gender matched healthy,population-based controls were randomly selected byStatistics Sweden (SCB). Cerebrospinal fluid sampling


(lumbar puncture) was performed when the participantswere in a stable euthymic mood. The cerebrospinal fluidconcentrations of the different biomarkers were measuredusing ELISA. Linear regression (using age and gender ascovariates) was used to analyse patient–control differencesin CSF biomarker concentrations. P-values less than 0.05were regarded as significant (two-tailed).

Results: The CSF concentrations of NF-L and MBPwere higher in patients when compared with healthycontrols (NF-L: b = 0.166, t = 3.578, df = 1, p< 0.001;MBP: b = 0.102, t = 12.023, df = 1, p = 0.044). There wereno differences in the concentration of S100B and h-FABPbetween patients and controls (S100B: b = 0.046, t = 0.742,df = 1, p = 0.459; h-FABP: b = 0.095, t = 1.584, df = 1,p = 0.115). We also observed several associations betweenCSF biomarkers and medications at the time of CSFsampling and performances at neuropsychological tests.

Conclusions: We conclude that the large subjectpopulation in the current study is suitable for findingpotential biomarkers for bipolar disorder as well asidentifying different subgroups of bipolar disorder. Thus,the findings in this study have the potential of increasingthe understanding of neuroprogression in bipolar disorderas well as giving indication of the biological effects ofdifferent medications. Future studies will give emphasisto the identification of associations between cerebrospinalfluid biomarker concentrations and alterations in brainstructures, genetic variations, and cognitive functions.

Reference(s)

[1] Kempton MJ, Geddes JR, Ettinger U, Williams SC,Grasby PM. Meta-analysis, database, and meta-regression of 98 structural imaging studies inbipolar disorder. Archives of general psychiatry.2008;65(9):1017−32. Epub 2008/09/03.

[2] Jakobsson J, Zetterberg H, Blennow K, Ekman C,Johansson AGM, Landen M. Altered concentrationsof amyloid precursor protein metabolites in thecerebrospinal fluid of patients with bipolar disorder.Neuropsychopharmacology. 2012;In press (acceptedOctober 2012).

[3] Olsson B, Zetterberg H, Hampel H, Blennow K.Biomarker-based dissection of neurodegenerativediseases. Prog Neurobiol. 2011. Epub 2011/04/29.

P.4.034 Glucose abnormalities in newlydiagnosed, medication-naıve patientswith bipolar disorder, mania, andpsychosis

C. Garcia-Rizo1 °, B. Kirkpatrick2, E. Fernandez-Egea3,C. Oliveira4, I. Grande4, J. Undurraga4, E. Vieta4,M. Bernardo4. 1Hospital Clinic of Barcelona, Dept.of Psychiatry, Barcelona, Spain; 2College of Medicineand Scott & White Healthcare, Dept. of Psychiatry,Texas, USA; 3Cambridgeshire and Peterborough NHSFoundation Trust, Department of Psychiatry, Cambridge,United Kingdom; 4Hospital Clinic of Barcelona,Department of Psychiatry, Barcelona, Spain

Severe mental disorders, such as schizophrenia and majordepression have been associated with increased mortality,which is due in part to an increased risk of diabetesand cardiovascular disease. Some evidence suggests thatpatients with schizophrenia and major depression havean increased risk of these disorders independently ofmedication and other confounding factors. The associationof these problems with bipolar disorders has receivedless attention. The prevalence of Type 2 diabetes mellitus(T2DM) in bipolar disorders ranges from 8% to 17% [1],a threefold increase compared to the general population.Bipolar patients with co-morbid T2DM may have a moresevere course of the psychiatric illness and refractoriness totreatment. However pharmacological treatment, includingboth antipsychotic agents and mood stabilisers, mayconfound this relationship.We test the hypothesis that abnormal glucose tolerance

is present in patients with bipolar disorder at the onsetof the disease. Newly diagnosed drug-naıve patients withDSM-IV bipolar I disorder were recruited at the timeof their first clinical contact for psychotic symptom andunderwent a two-hour glucose tolerance test. We recruiteda control group that was matched to these patients withrespect to age, gender, ethnicity, body mass index (BMI),smoking habit (average number of cigarettes per day),cortisol values and socioeconomic status of the family oforigin. The two matched groups were compared using thenon-paired Student’s t-test, or the c2 test for comparisonsof proportions. Smoking and BMI were log-transformedin order to approximate a normal distribution. Baselineglucose values were compared with the Mann Whitney U,due to a skewed distribution.Fasting measures of glucose metabolism were very

similar for the two groups. Baseline glucose concentrations(mg/dL[SD]) were 92.6[17.4] for the bipolar group and85.5[6.5] for the control group (p = 0.275). Baseline insulinconcentrations (mU/L[SD]) were 8.9[4.0] for the bipolargroup and 10.8[5.9] for the control group (p = 0.295).


Cortisol values were also similar in both groups, meanvalue 18.8mg/dL for patients and 19.1mg/dL for controls(p = 0.906).In contrast, the bipolar patients had a higher mean

two hour glucose (2HG) value compared with matchedcontrols (respective means, mg/dL[SD], of 145.9[16.9]vs. 84.8[27.8]; p< 0.001). Two patients (29%) but onlyone control subject (2%) met criteria for impairedfasting glucose (p = 0.037). Six patients (86%) and twocontrols (4%) met criteria for impaired glucose tolerance(p< 0.001).We found that newly diagnosed, medication-naıve

patients with bipolar disorder, manic, with psychoticfeatures had higher 2HG concentrations than did matchedcontrol subjects. These differences could not be attributedto confounding by BMI, gender, age, psychotropicmedications, cortisol concentration, socioeconomic status,ethnicity, smoking or drugs that affect glucose tolerance.Our results are consistent with the proposals that bipolardisorder is a multi-systemic disease [2], or a syndrome ofaccelerated aging, and with the concept of allostatic loadas applied to bipolar illness [3].

Reference(s)

[1] Newcomer, J.W., 2006. Medical risk in patients withbipolar disorder and schizophrenia. J Clin Psychiatry67 Suppl 9, 25−30; discussion 36−42.

[2] Leboyer, M., Soreca, I., Scott, J., Frye, M., Henry, C.,Tamouza, R., Kupfer, D.J., 2012. Can bipolar disorderbe viewed as a multi-system inflammatory disease? JAffect Disord 141, 1−10.

[3] Vieta, E., Popovic, D., Rosa, A.R., Sole, B.,Grande, I., Frey, B.N., Martinez-Aran, A., Sanchez-Moreno, J., Balanza-Martinez, V., Tabares-Seisdedos,R., Kapczinski, F., 2012. The clinical implicationsof cognitive impairment and allostatic load in bipolardisorder. Eur Psychiatry Apr 23 [Epub ahead of print].

P.4.035 Clinical improvement and plasmaticconcentrations of fluoxetine in majordepressive disorder (MDD), obsessive–compulsive disorder (OCD) andgeneralised anxiety disorder (GAD)

A. Blazquez1 °, S. Mas2, M.T. Plana1, A. Lafuente2,I. Mendez1, L. Lazaro1. 1Hospital Clinic of Barcelona,Department of Child and Adolescent Psychiatry andPsychology, Barcelona, Spain; 2University of Barcelona,Department of Anatomic Pathology Pharmacology andMicrobiology, Barcelona, Spain

Introduction: Fluoxetine is a useful drug in thetreatment of major depression disorder (MDD), obsessive

compulsory disorder (OCD) and generalised anxietydisorder (GAD) in child and adolescent populations [1].Despite its efficacy, 30−40% of the patients do not

respond to treatment [2].Aim: To evaluate if clinical improvement is related to

fluoxetine plasma levels at 8 and 12 weeks after staringtreatment in a sample of adolescents diagnosed of MDD,OCD or GAD.

Methods: The study was conducted at the Childand Adolescent Psychiatry and Psychology Service ofthe Institute of Neurosciences at the Hospital Clinic inBarcelona. The period of recruitment was from June 2011to October 2012. All the subjects met DSM-IV diagnosticcriteria for MDD, OCD or GAD made by an experiencedchild psychiatrist. All subjects began fluoxetine treatmentat the initial phase of the study (week 0). Patients and theirparents were interview at week 0 with the Spanish versionof the semi-structured diagnostic interview K-SADS-PL (Schedule for Affective Disorders and Schizophreniafor School-Age Children-Present and Lifetime Version)in order to assess current and past psychopathology.Information about severity of illness was obtained atweek 0 through the CDI (children’s depression inventory),OCI/CV (obsessive–compulsory inventory-child version),SCARED (the screen for child anxiety related emotionaldisorders), CGI (the clinical global impression) andGAF/CGAS (global assessment of functioning) scales.In patients with OCD, CYBOCS (children’s Yale-Brownobsessive compulsive scale) was also administered. Toassess clinical improvement CDI, SCARED, OCI, CGIand GAF/CGAS scales were administered at weeks 8 and12. UKU (Udvalg for Klinske Undersogelser) scale wasadministered to assess side effects of treatment.All data were analysed using SPSS 18.0 for Windows.

Spearman’s rank correlation coefficient was used to assessrelationship between two variables. Statistical significancewas set at p< 0.05.

Results: The sample included 50 patients (34% malesand 66% females). The mean age was 14.72 years (DS1.86). A 76% of the patients were diagnosed with MDD,12% OCD, 12% of GAD.At week 8 the mean dose of fluoxetine was 23.44mg/day

(DS= 10.37) and the mean of plasmatic concentrations offluoxetine was 82.38 ng/ml (DS= 83.443). No significantlinear correlation between plasmatic concentrations offluoxetine and side effects (measured by UKU scale) wasfound (Spearman’s rank correlation coefficient = 0.27;p = 0.852> 0.05).At week 12 the mean dose of fluoxetine was

25.32mg/day (DS= 13.32) and the mean of plas-matic concentrations of fluoxetine was 109.90 ng/ml(DS= 129.029). A significant linear correlation betweenplasmatic concentrations of fluoxetine and side effects was


found (Spearman’s rank correlation coefficient = 0.311;p = 0.028< 0.05). The most common side effects weresomnolence and asthenia. Also was found a significantcorrelation between plasmatic concentrations of fluoxetineand punctuation at CGI scale (Spearman’s rank correlationcoefficient = 0.294; p = 0.038< 0.05).

Conclusions: At week 8 no relation between clinicalimprovement/side effects and plasmatic concentrations offluoxetine was found, but at week 12 side effects wererelated to plasmatic concentrations of fluoxetine and alsoto clinical improvement measured by CGI scale.

Reference(s)

[1] Wilens TE, Cohen L, Biederman J, Abrams A,Neft D, Faird N et al. Fluoxetine Pharmacokineticsin pediatric patients. J Clin Psychopharmacol. 2002;22 (6):568−75.

[2] Findling RL, McNamara NK, Stansbrey RJ, Feeny NC,Young CM, Peric FV, et al. The relevance ofpharmacokinetic studies in designing efficacy trialsin juvenile major depression. J Child AdolescPsychopharmacol. 2006; 16 (1−2):131−45.

P.4.036 Corticotropin-releasinghormone and serotonin − theneuropsychopharmacology offear learning deficit from rodentsto humans

I. Heitland1 °, L. Groenink2, E.Y. Bijlsma2, R.S. Oosting2,J.L. Kenemans1, J.M.P. Baas1. 1Utrecht University,Experimental Psychology & Psychopharmacology,Utrecht, The Netherlands; 2Utrecht Institute ofPharmaceutical Sciences, Psychopharmacology, Utrecht,The Netherlands

Background: From an evolutionary perspective, theacquisition of fear responses enables organisms to respondappropriately to predictors of aversive events. In thelaboratory, this process is often modelled by classical fearconditioning procedures in which an originally neutralconditioned stimulus (CS; e.g., a light) is repeatedlypaired with an unconditioned aversive stimulus (UCS; e.g.an electrical shock). During the course of this learningprocess, conditioned fear responses develop towards thethreat cue. As a consequence, absence of the CS may cometo signal periods of safety. However, if this contingencyis not learned, threat remains unpredictable. This canresult in chronic states of maladaptive anxiety in thecontext in which the CS is presented [1]. Literaturesuggests that this learning deficit plays a crucial rolein the pathogenesis of human anxiety disorders [2], andlarge interindividual variability in fear learning deficits has

been reported [1]. As of yet, however, it remains largelyunknown which neurotransmitter systems are involvedin human fear learning deficits. Recently, preclinicalstudies have shown that the corticotropin-releasing factor(CRF) [3] and serotonin (5HT) are interactively involvedin adaptive fear learning. Here, we aim to translate thispreclinical evidence to the human realm.

Methods: 150 healthy medication-free human subjectscompleted a well-established cue and context fearconditioning procedure in a virtual reality environment [1].As in preclinical studies, fear potentiation of the startlereflex (FPS) was measured to assess fear conditioning.During the paradigm, we assessed both uninstructed fearacquisition and instructed fear expression. Due to thepaucity of pharmacological administrations with regard tothe CRF system, we chose to investigate innate variabilityin both the corticotropin releasing hormone system andthe serotonin system. Therefore, all participants weregenotyped for polymorphisms located within regulatoryregions of the corticotropin releasing hormone receptor 1(CRHR1 − rs878886) and the serotonin transporter(5HTTLPR). These polymorphisms have previously beenlinked to panic disorder (CRHR1 [rs878886] G-allele) andanxious symptomology and personality (5HTTLPR shortallele), respectively.

Results:G-allele carriers of CRHR1 (rs878886) showedno acquisition of fear conditioned responses (FPS) to thethreat cue in the uninstructed phase, whereas fear acquisi-tion was present in C/C homozygotes. Moreover, carryingthe risk alleles of both rs878886 [G-allele] and 5HTTLPR[short allele] was associated with increased FPS to thethreat context during this phase. After explicit instructionsregarding the threat contingency were given, the cue FPSand context FPS normalised in all genotype groups.

Conclusion: The present results indicate that geneticvariability in the corticotropin-releasing hormone recep-tor 1, especially in interaction with the 5HTTLPR, isinvolved in the acquisition of fear in humans. Fearexpression in the contrary was not associated with geneticvariability in both systems. These results translate prioranimal findings to the human realm.

Reference(s)

[1] Baas, J.M., van Ooijen, L., Goudriaan, A., Kenemans,J.L., 2008. Failure to condition to a cue is associatedwith sustained contextual fear. Acta Psychol (Amst)127, 581–592.

[2] Grillon, C., 2002. Associative learning deficitsincrease symptoms of anxiety in humans. BiolPsychiatry 51, 851–858.

[3] Bijlsma, E.Y., van Leeuwen, M.L., Westphal, K.G.,Olivier, B., Groenink, L., 2011. Local repeatedcorticotropin-releasing factor infusion exacerbates


anxiety- and fear-related behavior: differential in-volvement of the basolateral amygdala and medialprefrontal cortex. Neuroscience 173, 82−92.

P.4.037 HMGB1 signalling alters T-cellfunctioning in response to redoxstatus in depressed patients: effect onglucocorticoid receptor function

J. Rybka1 °, A. Cattaneo2, K. Kedziora-Kornatowska3,D. Kupczyk1, J. Kedziora1. 1Nicolaus CopernicusUniversity Collegium Medicum, Department ofBiochemistry, Bydgoszcz, Poland; 2FatebenefratelliGiovanni di Dio, Genetics Unit, Brescia, Italy; 3NicolausCopernicus University Collegium Medicum, Departmentand Clinic of Geriatrics, Bydgoszcz, Poland

It is well known that inflammation plays a pivotalrole in depression pathogenesis and treatment response,also interacting with other biological systems, includingglucocorticoid receptor (GR) functionality and redox-oxidative functions [1]. One molecule, named High-Mobility Group protein B1 (HMGB1), is an importantmediator of redox status and also an effective stimulustriggering inflammation and GR function.In this study we aimed to investigate more in detail the

role of HMGB1 as a mediator of inflammatory responsein depression. In addition to measuring HMGB1 westudied pro- and antioxidant status alongside with T cellsphenotype and also proinflammatory cytokine profiles indepressed patient. Moreover, we have also investigatedthe GR response in relation to oxidative and inflammatorystatus.

Methods: Blood samples were collected from 15patients diagnosed with recurrent depressive disorder(rDD) and from 19 healthy controls. We measuredseveral markers related to oxidative-redox status includingplasma hydrogen peroxide (H2O2) and malondialdehyde(MDA) levels in red blood cells. Furthermore, activitiesof antioxidant enzymes superoxide dismutase (SOD1),glutathione proxidase (GPx1), glutathione reductase(GSHR) in red blood cells and heme oxygenase (HO1)levels in plasma were assessed in this study.IL-2, IL-6, IL-8, IFN-g, IL-17, HMGB1, neopterin

and cortisol were assayed immunoenzymatically insera. Surface phenotype expression of T regulatoryand T effector cells were investigated by means offlow cytometry. The GR response was measured bydexamethasone-induced inhibition of IL-6 produced byLPS-stimulated peripheral blood.

Results: We observed pro- and antioxidant imbalancein depressed patients expressed by significant changes in

the enzyme activities. In particular we found increasedlevels of H2O2 (p< 0.01), increased concentrationsof lipid peroxidation product MDA (p< 0.001) andaltered activities of antioxidant enzymes as expressed bydecreased activities of SOD1 (p< 0.05), GPx1 (p< 0.01)and HO1 (p< 0.01) and increased activity of GSHR(p< 0.001).Inflammatory status of depressed patients was reflected

by increased concentration of IL-6 (p< 0.05) and de-creased concentration of IL-17A (p< 0.01). Interestingly,alterations in cytokine levels where accompanied byincreased Treg/Teffector cells ratio (p< 0.01). Moreoverimmune activation and oxidative stress in depressedpatients were confirmed by elevated neopterin levels(p< 0.001).We also revealed a distinct role of oxidative potential

of the blood as measured by H2O2, MDA, SOD1, GPx1,HO1, GSHR in modulating HMGB1 levels (p< 0.01) andHMGB1 proinflammatory activity (p< 0.001). Further-more, HMGB1 signalling was observed to alter cytokineprofile, specifically IL-6/IL-17A balance (p< 0.01) andTreg/Teffector cells ratio (p< 0.05) in depressed patients.Finally, we also found in depressed patients significant

correlations between glucocorticoid responses and redoxand inflammatory parameters such as MDA, IL-17A andIFN-g (all p< 0.05).

Conclusions: Our data suggest that the immunomod-ulatory role of HMGB1 might be related to the changeof its activity induced by redox imbalance in depressedpatients. Moreover, the shifted immune balance alongsidewith prooxidative environment modulates glucocorticoidresponse in these patients.

Reference(s)

[1] Rybka, J., Kedziora-Kornatowska, K., Kedziora, J.,Kucharski, R., 2009. Immunosenescence and late lifedepression. Centr Eur J Immunol 34 (4), 271–275.

P.4.038 Effect of neuregulin-1 gene functionalvariant and environmental factors onalcohol use disorder

M. Vaht1 °, E. Kiive1, K. Laas1, J. Parik2, T. Veidebaum3,J. Harro1. 1University of Tartu, Department ofPsychology, Tartu, Estonia; 2University of Tartu, Instituteof Molecular and Cell Biology, Tartu, Estonia; 3NationalInstitute for Health Development, Tallinn, Estonia

Neuregulin-1 is a signalling protein that affects neuronaldevelopment, synaptic plasticity, neuronal survival, glu-tamatergic neurotransmission, and glial functioning. Afunctional promoter polymorphism in the neuregulin-1


gene (NRG1) − SNP8NRG243177/rs6994992; (C/T) inthe type IV promoter region of NRG1 − has been foundto affect brain structure [1]. Subjects with C/C genotypewere found to have higher white and also grey mattervolume in several brain regions, for example inferior,middle and superior frontal gyri. Thus, the effects of NRG1on psychiatric disorders are of interest.

Purpose of the study:We aimed at examining the effectof NRG1 genotype and different environmental factors onpsychiatric disorders in population-representative sampleof young adults.

Methods: We used data of the older cohort of thelongitudinal Estonian Children Personality Behavior andHealth Study that at age 25 went through M.I.N.I.interview for lifetime psychiatric disorders. Data ondiagnosis were available on 501 subjects, 221 of themmale, 280 female. The rationale and procedure ofsample formation have been described elsewhere [2].NRG1 genotype frequencies were in HWE: C/C − 38%,T/T − 12%, C/T − 50%.

Table 1. NRG1 × environment effects on AUD in females

NRG1 × AUD

C/C T-allele

No AUD Aud No AUD Aud

Stressful life events, 18 yrs 2.5±2.4 6.3±3.8 2.5±2.5 5.0±2.2Victim of rape/attempted rape, 18 yrs 18% 58%*** 14% 0%Stressful life events, 25 yrs 3.8±2.7 6.0±2.6* 4.2±2.8 3.6±2.7Relationship problems, 25 yrs 58% 94%* 60% 60%Depressiveness, MADRS, 18 yrs 11.1±6.1 19.5±7.7* 11.3±6.5 10.5±5.7Depressiveness, MADRS, 25 yrs 8.0±6.1 12.1±7.8 8.6±6.4 8.2±5.0Aggressiveness, 18 yrs 2.2±1.2 1.5±0.8** 1.8±1.0 3.3±1.0*p< 0.05, **p< 0.01, ***p< 0.001 − difference from T-allele carriers with AUD.

Results: The NRG1 genotype had no effect onprevalence of anxiety or affective disorders. By age 25,74 (33%) males and 21 (8%) females had suffered fromalcohol use disorder (AUD). In females, subjects withC/C genotype were approximately 5 times more likelyto have AUD by the age of 25 than T-allele carriers(df = 2, c2 = 14.2, p = 0.001): 15% of females with C/Cgenotype and 3% of T-allele carriers had had AUD. Inmales, no effect of NRG1 genotype on lifetime AUD wasfound (df = 2, c2 = 0.1, p = 0.946). Neuroticism was notaffected by NRG1 genotype either, however, it was higherin females with AUD. Females with C/C genotype andAUD had experienced more stressful life events, especiallyrelationship problems by the age of 25 (Table 1). Alreadyby the age 18 they had more likely been victims of rape orattempted rape, and they had higher depressiveness scores.Interestingly, they had also exhibited less aggressivebehaviour as reported by teachers. The effect of NRG1genotype on AUD was not affected by the age of drinkingthe first half a dosage of alcohol. However, there was a

tendency towards more frequent use of alcohol in femaleswith C/C genotype and AUD at the age of 18.

Conclusion: NRG1 genotype significantly affects therisk of developing alcohol use disorder in females, andthe relationship is affected by environmental conditions.

Reference(s)

[1] Barnes, A., Isohanni, M., Barnett, J.H., Pietilainen, O.,Veijola, J., Miettunen, J., Paunio, T., Tanskanen, P.,Ridler, K., Suckling, J., Bullmore, E.T., Jones, P.B.,Murray, G.K., 2012. Neuregulin-1 genotype isassociated with structural differences in the normalhuman brain. Neuroimage 59, 2057–2061.

[2] Tomson, K., Merenakk, L., Loit, H.-M., Maestu, J.,Harro, J., 2011. The relationship between serotonintransporter gene promoter polymorphism and serumlipid levels at young age in a longitudinal population-representative study. Prog NeuropsychopharmacolBiol Psychiatry 35, 1857–1862.

S101

Author Index

Abedat, S.; S17 (P.1.018)Adan, R.A.; S11 (P.1.010)Adell, A.; S88 (P.4.025)Aghajanov, M.; S22 (P.1.023)Aguglia, E.; S83 (P.4.019), S89 (P.4.027)Aguilera, G.; S7 (P.1.006)Aitchison, K.J.; S69 (P.4.002)Almeida, J.; S81 (P.4.017)Almeida, R.; S37 (P.2.013)Altamura, A.C.; S60 (P.3.004), S74(P.4.009)

Alvarez, E.; S86 (P.4.023)Alvarez-Blazquez, M.; S91 (P.4.029)Alyagon, U.; S82 (P.4.018)Andersen, A.S.; S92 (P.4.030)Anedda, A.; S80 (P.4.015)Antypa, N.; S85 (P.4.021)Arango, C.; S91 (P.4.029)Arosio, B.; S60 (P.3.004)Artigas, F.; S6 (P.1.004), S40 (P.2.017)

Baas, J.M.P.; S79 (P.4.014), S97 (P.4.036)Bader, M.; S43 (P.2.021)Baldinger, P.; S80 (P.4.016)Balestri, M.; S76 (P.4.011)Balia, C.; S80 (P.4.015)Batalla, A.; S78 (P.4.013)Bator, E.; S57 (P.3.001), S58 (P.3.002)Battaglia, E.; S83 (P.4.019)Beaudet, G.; S42 (P.2.020)Beesdo-Baum, K.; S86 (P.4.022)Belelli, D.; S16 (P.1.017)Benatti, B.; S60 (P.3.004)Benfenati, F.; S21 (P.1.022)Bennett, K.A.; S4 (P.1.002)Bernardo, M.; S95 (P.4.034)Berod, A.; S49 (P.2.028)Berrocoso, E.; S88 (P.4.025)Bespalov, A.; S41 (P.2.018)Bhattacharyya, S.; S78 (P.4.013)Bianchini, O.; S83 (P.4.019)Biggio, F.; S7 (P.1.005)Biggio, G.; S7 (P.1.005)Bijlsma, E.Y.; S97 (P.4.036)Bilek, E.; S67 (S.04.01)Blanco-Hinojo, L.; S78 (P.4.013)Blazquez, A.; S96 (P.4.035)Blennow, K.; S94 (P.4.033)Bloomfield, P.S.; S20 (P.1.021)Bockaert, J.; S8 (P.1.007)

Boehringer, A.; S90 (P.4.028)Boellaard, R.; S93 (P.4.032)Bortolozzi, A.; S6 (P.1.004)Bossong, M.G.; S93 (P.4.032)Bouet, V.; S36 (P.2.012)Boulay, V.; S9 (P.1.008)Boulouard, M.; S36 (P.2.012), S42(P.2.020)

Branski, P.; S10 (P.1.009)Brehin, M.; S42 (P.2.020)Brioschi, S.; S71 (P.4.004)Brozaitiene, J.; S92 (P.4.031)Bunevicius, R.; S92 (P.4.031)Buoli, M.; S74 (P.4.009)Burkauskas, J.; S92 (P.4.031)Burnat, G.; S10 (P.1.009)Burshtein, S.; S84 (P.4.020)

Caldiroli, A.; S74 (P.4.009)Caletti, E.; S74 (P.4.009)Campa, L.; S88 (P.4.025)Cannavo, D.; S83 (P.4.019), S89(P.4.027)

Cano-Colino, M.; S37 (P.2.013)Carceller, M.; S86 (P.4.023)Carmi, L.; S82 (P.4.018)Carucci, S.; S80 (P.4.015)Castane, A.; S88 (P.4.025)Cath, D.C.; S79 (P.4.014)Cattaneo, A.; S98 (P.4.037)Celada, P.; S40 (P.2.017)Celikel, T.; S35 (P.2.011)Chaumont-Dubel, S.; S8 (P.1.007)Cherian, R.; S87 (P.4.024)Chlopocka-Wozniak, M.; S75 (P.4.010)Chruscicka, B.; S10 (P.1.009)Cohen, H.; S17 (P.1.018)Colle, R.; S18 (P.1.019)Collier, D.A.; S11 (P.1.010)Colombo, C.; S71 (P.4.004)Compte, A.; S37 (P.2.013)Concerto, C.; S83 (P.4.019), S89(P.4.027)

Corbani, M.; S9 (P.1.008)Corruble, E.; S18 (P.1.019)Creus, M.; S77 (P.4.012)Crippa, J.A.; S78 (P.4.013)Cryan, J.F.; S51 (P.2.031), S52 (P.2.032)Czyzyk, J.; S43 (P.2.021)

D’Addario, C.; S60 (P.3.004)Daivs, O.S.P.; S69 (P.4.002)Dar, K.; S87 (P.4.024)Dar, R.; S82 (P.4.018)Darcy, R.; S52 (P.2.032)David, D.J.; S18 (P.1.019)Davis, J.M.; S84 (P.4.020)Dazzi, L.; S7 (P.1.005)De Diego-Adelino, J.; S86 (P.4.023)de Graan, P.N.E.; S61 (P.3.005)de Leeuw, M.; S72 (P.4.006)de Paola, V.; S20 (P.1.021)de Visser, L.; S33 (P.2.009)de Vries, E.; S11 (P.1.010)De Vries, T.J.; S46 (P.2.025)Defrancesco, M.; S32 (P.2.007)Deisenhammer, E.A.; S32 (P.2.007)Delaunay, V.; S42 (P.2.020)Dell’Osso, B.; S60 (P.3.004)Dembinska-Krajewska, D.; S75 (P.4.010)Di Francesco, A.; S60 (P.3.004)Diaz, A.; S13 (P.1.013)Dıaz-Caneja, C.M.; S91 (P.4.029)Dinan, T.G.; S51 (P.2.031)Dine, J.; S12 (P.1.011)Djokic, G.; S71 (P.4.005)Doboszewska, U.; S19 (P.1.020)Dorofeikova, M.; S41 (P.2.018)Drago, A.; S85 (P.4.021)Dravolina, O.; S41 (P.2.018)Duhr, F.; S8 (P.1.007)Duits, P.; S79 (P.4.014)Dulin, E.; S91 (P.4.029)Dupuis, D.; S8 (P.1.007)Dziedzicka-Wasylewska, M.; S15(P.1.015)

Eder, M.; S12 (P.1.011)Ekman, C.J.; S94 (P.4.033)El Yacoubi, M.; S49 (P.2.028)Engberg, G.; S14 (P.1.014)Engel, M.; S59 (P.3.003)Engelhard, I.M.; S79 (P.4.014)Erhardt, S.; S14 (P.1.014)Eriksson, E.; S53 (P.2.033), S54(P.2.035)

Erritzoe, D.; S87 (P.4.024)Ersche, K.D.; S50 (P.2.030)Espinoza, S.; S21 (P.1.022)

S102 Author Index

Fabbri, C.; S76 (P.4.011)Faron-Gorecka, A.; S15 (P.1.015)Farre, M.; S78 (P.4.013)Feliu, T.; S77 (P.4.012)Fellig, Y.; S17 (P.1.018)Feng, L.; S92 (P.4.030)Fernandez, S.P.; S50 (P.2.029)Fernandez-Egea, E.; S95 (P.4.034)Ferres-Coy, A.; S6 (P.1.004)Filip, M.; S43 (P.2.021)Finn, D.P.; S54 (P.2.034)Fone, K.C.F.; S25 (S.02.02), S27(P.2.002)

Franch, J.; S77 (P.4.012)Franchini, L.; S71 (P.4.004)Freret, T.; S36 (P.2.012), S42 (P.2.020)Freudenberg, F.; S35 (P.2.011)Fumagalli, F.; S21 (P.1.022)Fuxe, K.; S43 (P.2.021)

Gainetdinov, R.; S21 (P.1.022)Galimberti, D.; S60 (P.3.004)Galindo, M.F.; S3 (P.1.001)Galofre, M.; S88 (P.4.025)Garbett, K.; S62 (P.3.007)Garcia-Rizo, C.; S95 (P.4.034)Gardier, A.M.; S18 (P.1.019)Gaspar, P.; S50 (P.2.029)Gass, N.; S28 (P.2.003)Gelegen, C.; S11 (P.1.010)Gershon, A.A.; S84 (P.4.020)Giannotti, G.; S21 (P.1.022)Godinho, B.M.D.C.; S52 (P.2.032)Goelman, G.; S17 (P.1.018)Gojkovic-Bukarica, L.; S30 (P.2.005)Gomez-Anson, B.; S86 (P.4.023)Grande, I.; S95 (P.4.034)Granitzio, F.; S80 (P.4.015)Grecu, I.G.; S84 (P.4.020)Gressier, F.; S18 (P.1.019)Groenink, L.; S97 (P.4.036)Gruart i Masso, A.; S50 (P.2.029)Gruca, P.; S15 (P.1.015)Guiard, B.P.; S18 (P.1.019)Guillon, G.; S9 (P.1.008)Guisasola, M.C.; S91 (P.4.029)Gunn, R.N.; S87 (P.4.024)Guo, Q.; S87 (P.4.024)Gutierrez-Mecinas, M.; S63 (P.3.008)Gutierrez-Zotes, A.; S77 (P.4.012)

Haddad, L.; S90 (P.4.028)Haeusler, D.; S80 (P.4.016)Hahn, A.; S80 (P.4.016)Hamon, M.; S61 (P.3.006)Harrison, B.J.; S78 (P.4.013)Harro, J.; S98 (P.4.038)

Hartmann, J.; S42 (P.2.019)Hasholt, L.; S92 (P.4.030)Heitland, I.; S97 (P.4.036)Herrmann, L.; S12 (P.1.011)Herth, M.M.; S92 (P.4.030)Hessel, E.V.S.; S61 (P.3.005)Hilbert, K.; S86 (P.4.022)Hilscher, M.; S74 (P.4.008)Hinterhuber, H.; S32 (P.2.007)Hodgson, K.; S69 (P.4.002)Hoeijmakers, L.; S59 (P.3.003)Holsboer, F.; S12 (P.1.011)Holst, K.K.; S92 (P.4.030)Hotineanu, M.; S84 (P.4.020)Howes, O.D.; S20 (P.1.021)

Ionescu, I.A.; S12 (P.1.011)Iversen, P.; S92 (P.4.030)

Jaako, K.; S33 (P.2.008)Jain, R.K.; S33 (P.2.008)Jakob, S.B.; S64 (P.3.009)Jakobsson, J.; S94 (P.4.033)Jancic, J.; S73 (P.4.007)Jennings, E.M.; S54 (P.2.034)Jimenez-Sanchez, L.; S88 (P.4.025)Joels, M.; S33 (P.2.009)Johansson, A.G.M.; S94 (P.4.033)Jonkman-Tielemans, S.; S46 (P.2.025)Jordan, J.; S3 (P.1.001)Jurek, B.; S7 (P.1.006)

Kahn, R.S.; S72 (P.4.006), S93 (P.4.032)Kalk, N.J.; S87 (P.4.024)Kalman, S.; S62 (P.3.007)Kas, M.J.H.; S11 (P.1.010), S33(P.2.009), S61 (P.3.005)

Kasper, S.; S80 (P.4.016)Kayukova, E.; S41 (P.2.018)Kedziora, J.; S98 (P.4.037)Kedziora-Kornatowska, K.; S98 (P.4.037)Kemmler, J.; S32 (P.2.007)Kenemans, J.L.; S97 (P.4.036)Kenis, G.; S64 (P.3.009)Kiive, E.; S98 (P.4.038)King, M.V.; S25 (S.02.02), S27 (P.2.002)Kirkpatrick, B.; S95 (P.4.034)Kittel-Schneider, S.; S74 (P.4.008)Kliwicki, S.; S75 (P.4.010)Klugmann, M.; S35 (P.2.011)Knapp, S.; S27 (P.2.002)Kneitz, S.; S64 (P.3.009)Knudsen, G.M.; S92 (P.4.030)Kolasa, M.; S15 (P.1.015)Koricka, S.; S33 (P.2.009)Korosi, A.; S59 (P.3.003)Korte, S.M.; S47 (P.2.026)

Korte-Bouws, G.A.H.; S47 (P.2.026)Kostrzewa, E.; S11 (P.1.010)Kraneveld, A.D.; S47 (P.2.026)Kranz, G.S.; S80 (P.4.016)Kupczyk, D.; S98 (P.4.037)Kushner, S.; S81 (P.4.017)Kusmider, M.; S15 (P.1.015)

Laas, K.; S98 (P.4.038)Labad, J.; S77 (P.4.012)Lafay-Chabassier, C.; S39 (P.2.016)Lafuente, A.; S96 (P.4.035)Lallai, V.; S7 (P.1.005)Lambert, J.; S16 (P.1.017)Lammertsma, A.A.; S93 (P.4.032)Landen, M.; S94 (P.4.033)Landgraf, R.; S12 (P.1.011)Lanfumey, L.; S61 (P.3.006)Langmead, C.J.; S4 (P.1.002)Lanza, G.; S89 (P.4.027)Lanzenberger, R.; S80 (P.4.016)Larsson, M.; S14 (P.1.014)Latusz, J.; S57 (P.3.001), S58 (P.3.002)Layer, L.E.; S35 (P.2.011)Lazaro, L.; S96 (P.4.035)Lederbogen, F.; S90 (P.4.028)Lehel, S.; S92 (P.4.030)Lerer, B.; S17 (P.1.018)Lesch, K.P.; S64 (P.3.009)Licheri, V.; S7 (P.1.005)Lifschytz, T.; S17 (P.1.018)Lignani, G.; S21 (P.1.022)Linge, R.; S13 (P.1.013)Lingford-Hughes, A.R.; S87 (P.4.024)Liu, Y.; S7 (P.1.006)Loi, M.; S33 (P.2.009)Lopez-Gil, X.; S88 (P.4.025)Lory, O.; S17 (P.1.018)Lotan, A.; S17 (P.1.018)Lucassen, P.J.; S33 (P.2.009), S59(P.3.003)

Lueken, U.; S86 (P.4.022)Luppi, P.H.; S49 (P.2.028)Lutzu, S.; S7 (P.1.005)

Maccarrone, M.; S60 (P.3.004)Macchi, F.; S5 (P.1.003)Mack, V.; S35 (P.2.011)Mackowiak, M.; S57 (P.3.001), S58(P.3.002)

Madsen, M.K.; S92 (P.4.030)Maggi, S.; S21 (P.1.022)Maggio, R.; S16 (P.1.016)Magnano SanLio, F.; S89 (P.4.027)Makshakov, G.; S41 (P.2.018)Mallei, A.; S65 (P.3.010)

Author Index S103

Mannoury la Cour, C.; S8 (P.1.007), S16(P.1.016)

Mansuy, I.; S57 (S.03.01)Marian, G.; S84 (P.4.020)Marin, P.; S8 (P.1.007)Marksteiner, J.; S32 (P.2.007)Maroteaux, L.; S37 (P.2.014)Marsano, A.; S76 (P.4.011)Marshall, F.; S2 (S.01.02), S4 (P.1.002)Marshall, K.M.; S38 (P.2.015)Martın-Santos, R.; S78 (P.4.013)Marx, V.; S35 (P.2.011)Mas, S.; S96 (P.4.035)Massart, R.; S61 (P.3.006)Mathe, A.A.; S34 (P.2.010)Mc Mahon, B.; S92 (P.4.030)McGuffin, P.; S69 (P.4.002)McIntosh, A.; S25 (S.02.02)Medrihan, L.; S21 (P.1.022)Melis, M.; S22 (P.1.024)Mendez, I.; S96 (P.4.035)Meyer-Lindenberg, A.; S28 (P.2.003),S67 (S.04.01), S90 (P.4.028)

Mill, J.; S63 (P.3.008)Millan, M.J.; S8 (P.1.007), S9 (P.1.008),S16 (P.1.016)

Mion, J.; S9 (P.1.008)Mirnics, K.; S62 (P.3.007)Miszkiel, J.; S31 (P.2.006)Mitsogiannis, M.; S11 (P.1.010)Mitterhauser, M.; S80 (P.4.016)Mlyniec, K.; S19 (P.1.020)Moita, M.; S44 (P.2.022)Moloney, R.D.; S51 (P.2.031)Molteni, R.; S5 (P.1.003)Monseny, R.; S77 (P.4.012)Montalvo, I.; S77 (P.4.012)Mordalska, P.; S57 (P.3.001), S58(P.3.002)

Morel, A.L.; S49 (P.2.028)Moreno, C.; S91 (P.4.029)Moron-Nozaleda, M.G.; S91 (P.4.029)Musazzi, L.; S13 (P.1.012)

Naninck, E.F.G.; S59 (P.3.003)Naslund, J.; S53 (P.2.033), S54 (P.2.035)Nava, N.; S13 (P.1.012)Nee, G.; S42 (P.2.020)Negm, O.; S27 (P.2.002)Neill, J.C.; S38 (P.2.015)Neumann, I.D.; S7 (P.1.006)Nicolas, C.; S39 (P.2.016)Niederstatter, H.; S32 (P.2.007)Nikiforuk, A.; S45 (P.2.023), S46(P.2.024)

Nissbrandt, H.; S54 (P.2.035)Nowak, E.; S43 (P.2.021)

Nowak, G.; S19 (P.1.020)Nutt, D.J.; S87 (P.4.024)Nyengaard, J.R.; S13 (P.1.012)

Obradovic, D.I.; S30 (P.2.005)O’Driscoll, C.M.; S52 (P.2.032)Ogier, J.R.; S52 (P.2.032)Okine, B.; S54 (P.2.034)Olango, W.M.; S54 (P.2.034)Oliveira, C.; S95 (P.4.034)Olivier, B.; S47 (P.2.026), S61 (P.3.005)Oosting, R.S.; S97 (P.4.036)Oppelaar, H.; S61 (P.3.005)Ortega, G.; S64 (P.3.009)Ortega, L.; S77 (P.4.012)

Paizanis, E.; S42 (P.2.020)Palazzo, M.C.; S60 (P.3.004)Palsson, E.; S94 (P.4.033)Panetta, K.; S16 (P.1.017)Paoli, R.A.; S74 (P.4.009)Papp, M.; S5 (P.1.003), S15 (P.1.015)Parellada, M.; S91 (P.4.029)Parik, J.; S98 (P.4.038)Parson, W.; S32 (P.2.007)Pazos, A.; S13 (P.1.013)Perez, V.; S86 (P.4.023)Perez-Caballero, L.; S88 (P.4.025)Peters, J.; S46 (P.2.025)Petit, A.C.; S18 (P.1.019)Petterson, R.; S54 (P.2.035)Philippe, C.; S80 (P.4.016)Pilc, A.; S10 (P.1.009)Pistis, M.; S22 (P.1.024)Pitychoutis, P.M.; S37 (P.2.014)Pjetri, E.; S61 (P.3.005)Plana, M.T.; S96 (P.4.035)Popik, P.; S45 (P.2.023), S46 (P.2.024)Popoli, M.; S13 (P.1.012), S65 (P.3.010)Popovic, D.; S68 (P.4.001)Portella, M.J.; S86 (P.4.023)Potasiewicz, A.; S46 (P.2.024)Powell, S.B.; S14 (P.1.014)Price, J.; S1 (S.01.01)Prickaerts, J.; S48 (P.2.027)Prins, J.; S47 (P.2.026)Przegalinski, E.; S31 (P.2.006)Puigdemont, D.; S86 (P.4.023)Pujol, J.; S78 (P.4.013)

Quesseveur, G.; S18 (P.1.019)Quiedeville, A.; S36 (P.2.012)

Rabiner, E.A.; S87 (P.4.024)Racagni, G.; S5 (P.1.003), S65 (P.3.010)Rafa, D.; S45 (P.2.023)Rappeneau, V.; S49 (P.2.028)

Reif, A.; S74 (P.4.008)Reul, J.M.H.M.; S63 (P.3.008)Ricceri, R.; S89 (P.4.027)Rickenbacher, E.; S44 (P.2.022)Riga, M.S.; S40 (P.2.017)Risterucci, C.; S28 (P.2.003)Riva, M.A.; S5 (P.1.003)Rocchi, C.; S16 (P.1.016)Roche, M.; S54 (P.2.034)Rybakowski, J.; S75 (P.4.010)Rybka, J.; S98 (P.4.037)

Saez-Atienzar, S.; S3 (P.1.001)Sagheddu, C.; S22 (P.1.024)Samardzic, J.; S30 (P.2.005)Sanna, E.; S7 (P.1.005)Santarelli, S.; S42 (P.2.019)Sartorius, A.; S28 (P.2.003)Saunderson, E.A.; S63 (P.3.008)Savli, M.; S80 (P.4.016)Schenker, E.; S28 (P.2.003)Schintu, N.; S34 (P.2.010)Schmidt, M.V.; S42 (P.2.019)Schmidt, U.; S12 (P.1.011)Schmieg, N.; S16 (P.1.016)Schmitt, A.G.; S64 (P.3.009)Scholz, C.J.; S64 (P.3.009), S74 (P.4.008)Schraut, K.; S64 (P.3.009)Schreck, S.; S74 (P.4.008)Schwarz, A.J.; S28 (P.2.003)Schwieler, L.; S14 (P.1.014)Seeburg, P.H.; S35 (P.2.011)Seguini, M.; S65 (P.3.010)Sellgren, C.; S94 (P.4.033)Serra-Blasco, M.; S86 (P.4.023)Serretti, A.; S76 (P.4.011), S85 (P.4.021)Seveno, M.; S8 (P.1.007)Shaikh, A.A.; S63 (P.3.008)Shelton, R.C.; S62 (P.3.007)Slattery, D.A.; S7 (P.1.006)Slonimsky, A.; S17 (P.1.018)Smeraldi, E.; S71 (P.4.004)Smulders, Y.M.; S70 (P.4.003)Solesio, M.E.; S3 (P.1.001)Solinas, M.; S39 (P.2.016)Sonn, K.; S33 (P.2.008)Soriano-Mas, C.; S78 (P.4.013)Sowa-Kucma, M.; S19 (P.1.020)Spedding, M.; S28 (P.2.003)Spiers, H.; S63 (P.3.008)Sprengel, R.; S35 (P.2.011)Steinbusch, H.; S64 (P.3.009)Stepan, J.; S12 (P.1.011)Stevanovic, D.; S73 (P.4.007)Stragier, E.; S61 (P.3.006)Struik, R.F.; S46 (P.2.025)Studer, E.; S53 (P.2.033), S54 (P.2.035)

S104 Author Index

Sukhanov, I.; S21 (P.1.022)Svenningsson, P.; S34 (P.2.010)Szewczyk, B.; S19 (P.1.020)

Talani, G.; S7 (P.1.005)Tansey, K.E.; S69 (P.4.002)Tardito, D.; S65 (P.3.010)Tighe, P.; S27 (P.2.002)Tijdink, J.; S70 (P.4.003)Tiugan, A.; S84 (P.4.020)Tocari, E.; S84 (P.4.020)Tomlinson, A.; S38 (P.2.015)Torrens, M.; S78 (P.4.013)Tost, H.; S67 (S.04.01), S90 (P.4.028)Trollope, A.F.; S63 (P.3.008)Tucci, V.; S21 (P.1.022)Tulen, J.; S81 (P.4.017)

Uher, R.; S69 (P.4.002)Undurraga, J.; S95 (P.4.034)Utzeri, C.; S7 (P.1.005)

Vaht, M.; S98 (P.4.038)van Berckel, B.N.M.; S93 (P.4.032)van den Burg, E.H.; S7 (P.1.006)van den Hout, M.A.; S79 (P.4.014)van den Hove, D.L.A.; S64 (P.3.009)van der Doef, T.F.; S93 (P.4.032)

van der Veen, F.; S81 (P.4.017)van Gestel, M.; S11 (P.1.010)Van Goethem, N.P.; S48 (P.2.027)van Heesch, F.; S47 (P.2.026)van Lith, H.A.; S11 (P.1.010)Vaugeois, J.M.; S49 (P.2.028)Veidebaum, T.; S98 (P.4.038)Vereczkei, A.; S62 (P.3.007)Vergouwen, A.C.M.; S70 (P.4.003)Verhagen, L.A.W.; S11 (P.1.010)Verstuyft, C.; S18 (P.1.019)Vieta, E.; S68 (P.4.001), S95 (P.4.034)Vilella, E.; S77 (P.4.012)Vink, M.; S72 (P.4.006)Vives-Gilabert, Y.; S86 (P.4.023)Vlad, N.; S84 (P.4.020)

Waddington, J.; S25 (S.02.01)Wadsak, W.; S80 (P.4.016)Wagner, K.V.; S42 (P.2.019)Waldman, A.; S87 (P.4.024)Wang, X.D.; S42 (P.2.019)Watson, D.J.G.; S25 (S.02.02)Weaver, A.; S4 (P.1.002)Weber-Fahr, W.; S28 (P.2.003)Wedzony, K.; S57 (P.3.001), S58(P.3.002)

Wegener, G.; S13 (P.1.012)

Weinstock, M.; S27 (P.2.001)Weiser, M.; S84 (P.4.020)Westberg, L.; S53 (P.2.033)Westphal, K.G.C.; S47 (P.2.026)Whitelock, C.F.; S50 (P.2.030)Wigmore, P.; S27 (P.2.002)Windhorst, A.D.; S93 (P.4.032)Wotjak, C.T.; S12 (P.1.011)

Yaqub, M.; S93 (P.4.032)Yen, Y.C.; S12 (P.1.011)Yenkoyan, K.; S22 (P.1.023)

Zago, S.; S74 (P.4.009)Zangen, A.; S82 (P.4.018)Zaru, A.; S29 (P.2.004)Zecchillo, C.; S5 (P.1.003)Zetterberg, H.; S94 (P.4.033)Zhang, X.; S34 (P.2.010)Zharkovsky, A.; S33 (P.2.008)Zheng, L.; S28 (P.2.003)Zivkovic, N.; S71 (P.4.005)Zohar, I.; S27 (P.2.001)Zohar, J.; S82 (P.4.018)Zoric, K.; S71 (P.4.005)Zuddas, A.; S80 (P.4.015)Zurawek, D.; S15 (P.1.015)Zvartau, E.; S41 (P.2.018)

S105

Keyword Index

Acetylcholine; S41 (P.2.018), S48(P.2.027)

Ageing; S42 (P.2.020), S95 (P.4.034)Aggression:– basic; S53 (P.2.033)Alcoholism; S22 (P.1.024), S61(P.3.006), S98 (P.4.038)

Alzheimer’s disease:– basic; S22 (P.1.023), S33 (P.2.008)Animal models; S5 (P.1.003), S6(P.1.004), S11 (P.1.010), S13 (P.1.013),S17 (P.1.018), S19 (P.1.020), S25(S.02.01, S.02.02), S27 (P.2.002),S29 (P.2.004), S30 (P.2.005), S31(P.2.006), S34 (P.2.010), S38 (P.2.015),S39 (P.2.016), S40 (P.2.017), S41(P.2.018), S45 (P.2.023), S46 (P.2.025),S49 (P.2.028), S51 (P.2.031), S52(P.2.032), S54 (P.2.034), S61 (P.3.005)

Antidepressants:– basic; S5 (P.1.003), S6 (P.1.004), S18(P.1.019), S27 (P.2.001), S28 (P.2.003),S54 (P.2.035), S65 (P.3.010), S80(P.4.016), S88 (P.4.025)

– clinical; S18 (P.1.019), S69 (P.4.002),S76 (P.4.011), S85 (P.4.021), S89(P.4.027), S96 (P.4.035)

Antipsychotics; S77 (P.4.012)Anxiety; S12 (P.1.011), S17 (P.1.018),S27 (P.2.001), S42 (P.2.019), S44(P.2.022), S54 (P.2.035), S73 (P.4.007),S79 (P.4.014), S92 (P.4.031), S97(P.4.036)

Anxiety disorders; S12 (P.1.011), S79(P.4.014), S81 (P.4.017), S86 (P.4.022)

Attention-deficit/hyperactivity disorder;S80 (P.4.015)

Behavioural pharmacology; S25(S.02.02), S30 (P.2.005), S31(P.2.006), S33 (P.2.009), S36 (P.2.012),S37 (P.2.013), S39 (P.2.016), S41(P.2.018), S46 (P.2.024), S48 (P.2.027),S50 (P.2.029), S54 (P.2.035), S57(P.3.001)

Biological markers; S58 (P.3.002), S82(P.4.018), S86 (P.4.023), S91 (P.4.029),S94 (P.4.033), S97 (P.4.036), S98(P.4.037)

Biological rhythms; S92 (P.4.030)

Bipolar disorders; S60 (P.3.004), S68(P.4.001), S71 (P.4.004), S74 (P.4.008),S75 (P.4.010), S84 (P.4.020), S94(P.4.033), S95 (P.4.034)

CB1 receptors; S7 (P.1.005)Cell culture; S1 (S.01.01), S3 (P.1.001),S74 (P.4.008)

Childhood disorders; S1 (S.01.01), S80(P.4.015), S91 (P.4.029), S96 (P.4.035)

Cognition; S77 (P.4.012)Cognitive enhancing drugs; S36(P.2.012), S38 (P.2.015), S45 (P.2.023),S48 (P.2.027)

Computational model; S37 (P.2.013)

Depression:– basic; S13 (P.1.012, P.1.013), S27(P.2.001), S35 (P.2.011), S42 (P.2.019),S47 (P.2.026), S49 (P.2.028), S62(P.3.007), S64 (P.3.009), S92 (P.4.031)

– clinical; S32 (P.2.007), S73 (P.4.007),S76 (P.4.011), S86 (P.4.023), S89(P.4.027), S98 (P.4.037)

Diagnoses & classification; S76(P.4.011), S86 (P.4.022)

Dopamine; S7 (P.1.005), S15 (P.1.015),S16 (P.1.016), S21 (P.1.022), S22(P.1.024), S37 (P.2.014), S72 (P.4.006)

Drug dependence & abuse:– basic; S22 (P.1.024), S29 (P.2.004),S31 (P.2.006), S39 (P.2.016), S43(P.2.021), S46 (P.2.025), S49 (P.2.028),S50 (P.2.030)

– clinical; S74 (P.4.009), S78 (P.4.013),S87 (P.4.024)

Drug development; S2 (S.01.02), S57(P.3.001)

Drug monitoring; S20 (P.1.021), S80(P.4.015), S83 (P.4.019)

Early psychoses; S77 (P.4.012)Eating disorders; S11 (P.1.010), S29(P.2.004), S61 (P.3.005)

Epigenetics; S58 (P.3.002), S63 (P.3.008)Epilepsy; S73 (P.4.007)

G-proteins; S10 (P.1.009)Gene expression; S1 (S.01.01), S5(P.1.003), S7 (P.1.006), S10 (P.1.009),S52 (P.2.032), S54 (P.2.034), S60

(P.3.004), S61 (P.3.006), S62 (P.3.007),S63 (P.3.008)

Genetics/Molecular genetics; S11(P.1.010), S32 (P.2.007), S57(S.03.01), S61 (P.3.005), S67(S.04.01), S69 (P.4.002), S78(P.4.013), S85 (P.4.021), S92 (P.4.030),S97 (P.4.036), S98 (P.4.038)

Glutamate; S4 (P.1.002), S13 (P.1.012),S14 (P.1.014), S27 (P.2.002), S35(P.2.011), S37 (P.2.014), S51 (P.2.031),S88 (P.4.025)

Immediate early genes; S34 (P.2.010)Inhibitory neurotransmitters; S7(P.1.005)

Lithium & other mood stabilisers; S68(P.4.001), S71 (P.4.004), S74 (P.4.008),S75 (P.4.010)

Memory & cognitive disorders; S3(P.1.001), S42 (P.2.020), S46 (P.2.024),S50 (P.2.029), S59 (P.3.003), S74(P.4.009), S81 (P.4.017), S92 (P.4.031)

Methodology; S2 (S.01.02), S10(P.1.009), S33 (P.2.008), S84 (P.4.020)

Microglia; S20 (P.1.021)MicroRNA; S65 (P.3.010)Miscellaneous:– basic; S70 (P.4.003)– clinical; S71 (P.4.004)Molecular neurobiology; S19 (P.1.020),S57 (S.03.01), S61 (P.3.006), S65(P.3.010), S94 (P.4.033)

Neuroanatomy; S46 (P.2.025)Neuroendocrinology; S9 (P.1.008), S53(P.2.033), S59 (P.3.003)

Neuroimaging:– functional; S28 (P.2.003), S67(S.04.01), S72 (P.4.006), S80(P.4.016), S86 (P.4.022), S87 (P.4.024),S90 (P.4.028), S93 (P.4.032)

– structural; S67 (S.04.01), S78(P.4.013), S86 (P.4.023)

Neuroimmunology; S47 (P.2.026), S93(P.4.032), S98 (P.4.037)

Neuroleptics & antipsychotics:– basic; S40 (P.2.017)

S106 Keyword Index

– clinical; S68 (P.4.001), S71 (P.4.005),S83 (P.4.019), S84 (P.4.020)

Neuropeptides; S7 (P.1.006), S12(P.1.011), S44 (P.2.022), S60 (P.3.004)

Neurophysiology:– clinical; S82 (P.4.018)Neuroprotection; S22 (P.1.023)New research; S3 (P.1.001), S38(P.2.015), S50 (P.2.030), S70 (P.4.003),S89 (P.4.027)

Nociception; S51 (P.2.031)Noradrenaline; S22 (P.1.023), S81(P.4.017)

Nutrients/Antioxidants; S19 (P.1.020),S30 (P.2.005), S50 (P.2.030), S91(P.4.029)

Obsessive compulsive disorders; S82(P.4.018)

Other anxiolytics; S44 (P.2.022)

Panic disorders; S79 (P.4.014)Personality disorders; S75 (P.4.010)

Pharmacokinetics; S69 (P.4.002), S96(P.4.035)

Psychoneuroendocrinology; S90(P.4.028), S95 (P.4.034)

Psychoneuroimmunology; S14 (P.1.014),S87 (P.4.024)

Publication pressure; S70 (P.4.003)

Receptors; S2 (S.01.02), S4 (P.1.002), S8(P.1.007), S9 (P.1.008), S13 (P.1.013),S15 (P.1.015), S16 (P.1.017), S28(P.2.003), S33 (P.2.009), S43 (P.2.021)

Schizophrenia:– basic; S14 (P.1.014), S16 (P.1.016),S17 (P.1.018), S20 (P.1.021), S21(P.1.022), S25 (S.02.01, S.02.02), S27(P.2.002), S37 (P.2.014), S40 (P.2.017),S46 (P.2.024), S57 (P.3.001), S58(P.3.002), S72 (P.4.006)

– clinical; S71 (P.4.005), S74 (P.4.009),S83 (P.4.019), S93 (P.4.032)

Serotonin; S6 (P.1.004), S8 (P.1.007),S16 (P.1.017), S18 (P.1.019), S32(P.2.007), S34 (P.2.010), S36 (P.2.012),S37 (P.2.013), S42 (P.2.020), S45(P.2.023), S47 (P.2.026), S50 (P.2.029),S53 (P.2.033), S64 (P.3.009), S80(P.4.016), S85 (P.4.021), S88 (P.4.025),S92 (P.4.030)

Signal transduction; S4 (P.1.002), S8(P.1.007), S9 (P.1.008), S16 (P.1.016)

Sleep disorders; S16 (P.1.017)Smoking; S71 (P.4.005)Stress; S7 (P.1.006), S13 (P.1.012), S15(P.1.015), S33 (P.2.009), S42 (P.2.019),S54 (P.2.034), S57 (S.03.01), S59(P.3.003), S62 (P.3.007), S63 (P.3.008),S64 (P.3.009), S90 (P.4.028), S98(P.4.038)

Transgenic models; S21 (P.1.022),S25 (S.02.01), S33 (P.2.008), S35(P.2.011), S43 (P.2.021), S52 (P.2.032)

This book has been produced electronically by Elsevier B.V.

Every effort has been made to faithfully reproduce the papers as submitted. However, no responsibility isassumed by the organisers for any injury and/or damage to persons or property as a matter of products liability,negligence or otherwise, or from any use or operation of any methods, products, instructions or ideas containedin the material herein. Because of the rapid advances in the medical sciences, we recommend that independentverification of diagnoses and drug dosages should be made.

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