the consequences of uncontrollable stress are sensitive to duration of prior wheel running

The consequences of uncontrollable stress are sensitive to duration of

prior wheel running

Benjamin N. Greenwood, Teresa E. Foley, Dan Burhans, Steven F. Maier, Monika Fleshner

• Chronic exposure to uncontrollable stressors (UcS)– behavioral depression– learned helplessness

(LdH)

LdH

• LdH– psychological condition in which a human being/animal

has learned to believe that a situation is helpless– tail shock

• impaired escape performance• exaggerated fear conditioning

• Prevented/reversed by – Antidepressant/anxiolytic drugs– Physical activity

• 6+ weeks of voluntary access to running wheels

Prior works

• Activation of 5-HT neurons in the DRN is both necessary and sufficient to produce LH behaviors.

• dorsal raphé nucleus (DRN) and serotonin (5-HT) play a role in – stress-related affective disorders (depression/anxiety)– pharmacological action of antidepressant and

anxiolytic drugs– the modulation of the behavioral effects of

uncontrollable stressors

Stress

Controllable• Increase in 5-HT activity in

the DRN – involved in protection against

LH provided by wheel running– 5-HT neurons in the DRN can

be hyper-activated and sensitized

uncontrollable• greater c-Fos induction in the

DRN 5-HT neurons – exaggerated 5-HT efflux

(during stress and later behavioral testing)• within the DRN• in DRN projection sites

• manipulations that decrease 5-HT neural activity in the DRN during uncontrollable stress eliminate LH behaviors

c-Fos• … is a cellular proto-oncogene belonging to the immediate

early gene family of transcription factors.

• …transcription is upregulated in response to many extracellular signals, e.g. growth factors.

• …phosphorylation by MAPK, PKA, PKC or cdc2 alters the activity and stability of c-Fos.

• …dimerises with Jun to form the AP-1 transcription factor, – which upregulates transcription of a diverse range of genes

involved in everything from proliferation and differentiation to defense against invasion and cell damage.

corticotropin-releasing hormone (CRH)

• Release is influenced by – stress– cortisol – sleep/wake cycle

• Acts as a NT and a hormone– CRHRs in the …

• Central nucleus of the amygdala(CeA) • Bed nucleus of the stria terminalis (BNST)

5-HT & SSRIs• citalopram

– (Celexa, Cipramil, Emocal, Sepram, Seropram)

• escitalopram oxalate – (Lexapro, Cipralex, Esertia)

• fluoxetine – (Prozac, Fontex, Seromex, Seronil,

Sarafem, Fluctin (EUR)) • fluvoxamine maleate

– (Luvox, Faverin) • paroxetine

– (Paxil, Seroxat, Aropax, Deroxat, Rexetin, Xetanor, Paroxat)

• sertraline – (Zoloft, Lustral, Serlain)

CRH

• …is critical for mediation of behavioral consequences of UcS

• …has excitatory influences on 5-HT neurons located in the caudal DRN (dorsal raphé nucleus)– Caudal DRN CRH-R2 ant. injection prevent LdH

– No effect if injected rostral

• LdH behaviors are dependent upon hyperactivity of DRN 5-HT neurons during exposure to uncontrollable stress.– 6 weeks of wheel running prevents LdH and attenuates

uncontrollable stress-induced activity of DRN 5-HT neurons.

• Possible reasons:– 5-HT1A (autoreceptor)

• Increases in 5-HT1A inhibition can result in increased DRN 5-HT activity

– Other regions• Regions afferent to DRN could play their own game

• Physically active and sedentary rats have equal corticosterone responses during exposure to uncontrollable tail shock

• c-Fos is not globally attenuated in all stress reactive brain regions

Candidates regions

• Intra-DRN microinjections of CRH-R2/EAA antagonists prevent symptoms of UcS

• CRH and NMDA agonists can influence DRN 5-HT neurons– Afferent regions containing

• CRH• EAAs (excitatory amino acids) [eg. Glutamate or aspartate]

BNST• Activation of BNST

– produces neuroendocrine and behavioral responses that resemble those produced by stress

– inactivation of the BNST decreases neuroendocrine and behavioral responses to stress

• Lesion:– eliminate typical

behavioral conseq. of UcS

BLA & CeA• Basolateral amygdala (BLA):

– Info about environmental signals are associated w/ aversive unconditioned stimuli in the BLA

• CeA– stimulation produces

behavioral, cardiovascular and neuroendocrine components of fear

• Lesions– reduce or eliminate fear

LHb

• Electrical stimulation of the LHb excites DRN neurons

• Lesions– eliminate stress-induced

increases in 5-HT in the DRN.

– eliminate typical behavioral conseq. of UcS

• Anxiogenic and fear responses elicited by BNST and AMG stimulation are similar to stimulation of the DRN

• Take them out!

Goals

• effects of 3 weeks and 6 weeks of wheel running on LdH behaviors and uncontrollable stress-induced – c-Fos in the BNST, AMG and LHb– 5-HT & c-Fos in the DRN

Animals

• Adult, male Fischer F344 rats – 12:12 h light/dark cycle

(lights on 6am–6pm)– individually housed with

attached running wheel– 2 weeks acclimatization

(locked wheels)

Activity

2 groups

• No stress (home cage)• Stressed (tail shock)– Plexiglas tube + 100, 5 s tail shocks (1.5 mA) on a

1-min variable-interval schedule. • (8 am – 10 am (light)

– Returned to home cage

human equivalent

conditioned fear/shuttle box (24h post stress)

1. Freezing (Every 8s): 4 paws on floor + no movement

2. Foot shock: auto off after 30 s

Immuno

• Rats (n = 8/group) – Terminally sampled 90 min after tail shock (c-Fos

optimum)

• 35 m coronal sections (cryostat) of– DRN– BNST– AMG– LHb

c-Fos labeling

c-Fos/5-HT double labeling (DRN)

c-Fos + 5HT 5-HT

c-Fos

c-Fos immunoreactivity

Activity and body weights

Running Distance/week

Escape

Freezing

5-HT

c-Fos

# double labeled

c-Fos expression in the BNST

c-Fos Expression in the AMG

c-Fos Expression in the LHb

Summary

• Run long!– More sensitive to

duration (less to distance)

– Burghardt (12) • + maze

– 8 weekies yes– 4 weekies no

Summary

• Attenuation of 5-HT activity in the DRN contributes to the prevention of LdH by wheel running.

• BNST has a role in mechanism by which wheel running prevents LdH– But not the AMG or LHb