gaba receptors gaba receptors
TRANSCRIPT
GABA RECEPTORS
Ian L Martin and Susan M J Dunn
Ian Martin is Professor of Pharmacology inthe School of Life and Health Sciences at theUniversity of Aston, Birmingham B4 7ET, UK.Susan Dunn is a Senior Lecturer in theDepartment of Pharmacology at theUniversity of Bristol, University Walk, BristolBS8 1TD, UK. They have common interestsin GABAergic transmission in the centralnervous system and their research is focusedon the structure-function relationships of theGroup 1a ligand-gated ion channel receptors.
Historical Perspective
GABA is the major inhibitory amino acidtransmitter of the mammalian central nervoussystem and it is present in some 40% of allneurones. Most of the early studies, carried outwith iontophoretic application of GABA in theCNS, indicated that it generally producedinhibitory hyperpolarizing responses onneurones, which were blocked competitively bythe alkaloid bicuculline. The hyperpolarizingresponse is due to an increase in the chlorideconductance of the neuronal membraneallowing chloride ions to flow down theirelectrochemical gradient into the cell.However, in the late 1970s, Bowery and hiscolleagues, in attempts to identify GABAreceptors on peripheral nerve terminals, notedthat GABA application reduced the evokedrelease of noradrenaline in the rat heart and thatthis effect was not blocked by bicuculline. Thisaction of GABA was mimicked by baclofen, 4-amino-3-(4-chlorophenyl) butanoic acid (Figure1), a compound that had no effect on chlorideconductance in central neurones. The newreceptor was named GABA to differentiate itfrom its more familiar cousin, which was termedGABA . The GABA receptor had a rathermore difficult birth. In an attempt to discoverwhich conformation of GABA was responsiblefor activating the receptor, Johnson and hiscolleagues synthesised a number ofconformationally restricted analogues of GABAand noted that -4-aminocrotonic acid (CACA),which has a partially folded conformation (Figure1), depressed the firing of cat spinal neurones ina bicuculline insensitive manner. Thesedepressant effects could not be reproduced bybaclofen, suggesting a pharmacology distinctfrom that of either the GABA or GABA
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receptors. The receptor became known asGABA . The DNAs that encode these receptorproteins have now been identified, providing notonly a facile means for their molecularcharacterisation but also a significant stimulusfor our attempts to understand theirphysiological importance.
The GABA receptors are widely distributedwithin the mammalian CNS and exhibit adifferential topographical distribution.Systematic modification of the natural agonistdemonstrated that GABA receptors can beactivated by a number of compounds (Figure 1)such as muscimol, isoguvacine, 3-aminopropane sulphonic acid, piperidine-4-sulphonic acid and 4,5,6,7-tetrahydro-[5,4-c]-pyridin-3-ol, many of which were subsequentlyused as radioligands. At equilibrium thebinding of GABA agonists is heterogeneous witha high affinity component (K values of 10-20nM) and one or more low-affinity sites withdissociation constants in the range of 100 nM to1 M. The presence of even lower affinity sites(K values about 50 M) has been inferred from
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The GABA ReceptorsA
GABA RECEPTORS
H2N CO2H
GABA (Cat. No. 0344)
Muscimol (Cat. No. 0289)
CACA (Cat. No. 0179)
( )-BaclofenR (Cat. No. 0796)
Gabapentin (Cat. No. 0806)
SKF 97541 (Cat. No. 0379)
3-aminopropyl phosphinic acid
Isoguvacine (Cat. No. 0235)
(1 ,2 )-(+)-2-(aminomethyl)-cyclopropane-1-carboxylate
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H H
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(Bold text denotes compounds available from Tocris)
Figure 1. Structures of selected GABA receptoragonists
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Tocris Cookson Inc., USATel: (800) 421-3701
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The purification of the receptor protein in theearly 1980s provided the opportunity to raisemonoclonal antibodies to the receptor, whichwere later used to study the fine anatomicaldetail of receptor distribution. Subunitpurification, and elucidation of their partial aminoacid sequences was crucial in their eventualmolecular cloning. Only two receptor subunits( and ) were initially identified at the proteinlevel and, in the mid 1980s these were cloned.When co-expressed from their encoding nucleicacids in oocytes, these subunitsproduced a receptor that responded to GABAreceptor agonists and antagonists in theexpected manner.
Subsequent studies have revealed a multiplicityof protein subunits that have been divided intoseven classes, according to similarities in theirdeduced amino acid sequence. Within theseclasses there are further subdivisions intosubunit isoforms, some of which exhibitalternate splice variants. In man, six , threeand three subunit isoforms are presentlyknown, together with single representatives ofthe , , and classes. Additional isoforms inother species are known for some of theseclasses. Within a single subunit class thesequence homology is about 70% but betweenclasses this falls to around 30%. Deducedamino acid sequences of subunits from all of thefamilies indicate that each has a long aminoterminus of about 200 amino acids whichcontains a signature cysteine-cysteine loop,common to the receptor family, prior to the firstof four hydrophobic segments of about 20 aminoacids, which are termed TM1-TM4. TM2 isthought to form a major part of the lining of theion channel as it crosses the cell membrane,while between TM3 and TM4 there is a largeintracellular loop, which is the most divergentpart of the sequence within the sub-family. TheGABA receptors have been purified from thepig brain and imaged, after negative staining, inthe electron microscope. The receptor has adiameter of about 8 nm in the plane of themembrane and consists of five protein subunitsarranged pseudosymmetrically around theintegral ion channel; it appears very similar toimages of the nicotinic acetylcholine receptor.
Despite the plethora of receptor subunits, itappears that there are a limited number ofcombinations expressed . A separategene encodes each subunit andhybridisation, together with immuno-histochemical studies, have revealed a distinctdistribution for these gene products,consistent with the idea that they each serve adefined physiological role. The recognition andfunctional characteristics of the individualGABA receptor subtypes are defined by theirconstituent subunits and while ectopicexpression studies continue to explore thisdiversity, the authentication of particular GABAreceptor subtypes remains a significanttask. However, it is clear that the most commonGABA receptor in the mammalian CNSconsists of two copies each of the 1 and 2subunits together with a single 2 subunit. It isthis receptor that appears to mirror thepharmacological and biophysical characteristics
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the higher concentrations of GABA that arerequired to activate the channel. There has beenmuch speculation about the role of these sites inreceptor function. Whether the sites arephysically distinct or represent allostericconformations of the same site remains anintriguing question.
Electrophysiological studies demonstrated thatthe activation of the receptor resulted inincreased chloride conductance of the cellmembrane with the concentration-responsecurve exhibiting positive cooperativity, consistentwith the presence of at least two agonist bindingsites on the receptor molecule. The agonistinduced current decreased on continuedexposure to high agonist concentrationssuggesting that these receptors undergodesensitisation. Biophysical characterisation ofthe receptor, initially using noise analysis ofneurones in primary culture, provided the firstestimates of mean single channel conductanceand average channel open times. The lattervaried with the nature of the activating agonist.Development of single channel recordingtechniques provided further detail on thestochastic nature of the single channel eventswith the demonstration of multiple singlechannel conductances: 44, 30, 19 and 12 pS.The 30 pS conductance is the most prevalentwith distinct open time states, varying from 0.5to 7.6 ms. The distribution of these states isagonist concentration dependent, with longeropen times predominating at higher agonistconcentration. The competitive antagonistbicuculline appears to reduce conductancethrough the channel by reducing not only theopening frequency but also the mean opentime. Other competitive antagonists, such asthe pyridazinyl GABA derivative SR 95531(Figure 2), are available. In addition, thereceptor can be blocked non-competitively bypicrotoxin and by a number ofbicyclophosphates. Penicillin also decreasesthe channel open probability in a manner that iscompatible with open channel block.
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(Bold text denotes compounds available from Tocris)
Figure 2. Structures of selected GABA receptorantagonists
(+)-Bicuculline (Cat. No. 0130)
Phaclofen (Cat. No. 0178)
CGP 35348 (Cat. No. 1245)
OMe
N
N
NH
O
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SR 95531 (Cat. No. 1262)
SCH 50911 (Cat. No. 0984)
CGP 55845 (Cat. No. 1248)
HN P
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of the GABA receptors characterised previouslyin whole animals and in primary cultures ofcentral neurones.
Weiss and colleagues used site-directedmutagenesis to identify residues in the GABAreceptor that are involved in channel activation.They identified residues in the subunit thatappear to be important for low affinity agonistrecognition. Based on homology with othermembers of the receptor family, and thecurrently available evidence from labelling andmutagenesis studies, it appears likely that theactivation sites for GABA are located at the -interface(s). Recent work has identified residuesin the subunit that may be involved in highaffinity agonist binding and this has led to thesuggestion that these site(s) may lie at the -and - interfaces. The current consensus isthat a benzodiazepine modulatory site lies at theinterface between and subunits. Much ofthe evidence for binding site locations issomewhat indirect, involving mutational studiesto selectively disrupt the sites. However, ourability to interpret these data has recently beensignificantly enhanced by the crystallisation ofan acetylcholine binding protein which bears amarked structural similarity to the extracellulardomain of this receptor family. This protein,though not a ligand-gated ion channel, displaysremarkable homology within its ligand bindingdomains. The crystal structure thus provides avaluable physical template against which themutational information may be interpreted.Figure 3 shows the subunit arrangement of theGABA receptor that is consistent with thecurrently available ligand recognition site data.
The benzodiazepines, because of theirtherapeutic importance, have been a majorfocus of GABA receptor research since thediscovery of saturable, high affinity binding sitesfor [ H]diazepam in the CNS. Agonistactivation of the GABA receptor is augmentedby the anxiolytic benzodiazepines, causing aparallel leftward shift of the GABAconcentration-response curve. All the overteffects of the benzodiazepines: sedative,
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The Benzodiazepines
anxiolytic, anticonvulsant, muscle relaxant andamnesic, are produced via the GABAreceptors. However, not all the GABA receptorsthat exist in the brain recognise thebenzodiazepines. The particular subunitisoform present within an individual GABAreceptor subtype is the primary determinant ofbenzodiazepine recognition (Table 1). If the 1subunit of the most common GABA receptor isreplaced with 4 or 6, the receptor fails torecognise the benzodiazepines. It is now clearfrom both biochemical and mutational analysisthat this insensitivity is due primarily to a singleamino acid substitution: an arginine residue in
4 and 6 subunits replaces histidine 101,which is present in 1, 2, 3 and 5subunits. While receptors containing 1,
2, 3 or 5 together with a and a subunitare all recognised by the ‘classical’benzodiazepines, several agents are able todistinguish the receptors on the basis of their asubunit isoform composition. The first of thesecompounds was the triazolopyridazine CL218,872 , which is related to the recentlyintroduced hypnotic ‘Zaleplon’ (Figure 4), whilecertain -carboline-3-carboxylic acid esters
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Table 1. Approximate binding affinities of benzodiazepine site ligands for GABA receptor subtypes.A
Compound 1 2
Diazepam 16.9
Clonazepam 1.3 1.7 2.0 – – > 10,000
Triazolam 1.8 1.2 3.0 – 1.2 –
Bretazenil 1.2 1.2 1.3 – 2.4 –
1.0 1.1 1.5 107 0.4 90
Ro15-4513 2.6 2.6 1.3 – 0.24 6.5
CL218872 130 1820 1530 > 10,000 490 > 10.000
-CCM 1.7 6.5 4.1 – 27 2050
17 291 357 – > 15,000 –
48.5 27.4 24.5 – 0.45 83.2
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16.1 17.0 > 10,000 14.9 > 10,000
Ro15-1788 (Flumazenil)
Zolpidem
L-655,708
Values are quoted in nM. Information abstracted from references 41, 89 and 90. Note that the determinations were carried out with either 2 or 3subunit isoforms, which do not have a pronounced effect on the affinity of benzodiazepine site ligands.
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Figure 3. Viewing the GABA receptor from theextracellular space, the orientation of thesubunits within the pentamer together with thelocation of the benzodiazepine (Bz) and lowaffinity GABA sites shown is in accord with all themutagenesis data that is available to date
A
(Bold text denotes compounds available from Tocris)
allowed a similar distinction (Figure 4).Zolpidem (Figure 4), currently the most widelyprescribed hypnotic in the USA, is also able todistinguish GABA receptors on the basis oftheir subunit isoform composition: it has ahigh affinity for those receptors which contain
1, a lower affinity for those receptors whichcontain 2 or 3 and a very low affinity forthose receptors which contain 5. Againzolpidem does not recognise receptors whichcontain 4 or 6 subunits. A nomenclaturefor these receptors with distinct affinities for the‘subtype selective’ ligands was developed priorto the cloning era: 1 containing receptorsproved to have the Bz1 phenotype, while thosethat contained 2, 3 or 5 subunits exhibitedBz2 pharmacology. Other nomenclatures haveappeared during the intervening yearsculminating, most recently, in the classificationof GABA receptors by a combination ofmolecular and pharmacologicalcharacteristics; time will tell if the scientificcommunity finds it acceptable.
Perhaps one of the most interestingphenomenological observations to appear fromstudies of the benzodiazepine interaction withthe GABA receptors has been the developmentof the inverse agonist concept. The classicalbenzodiazepines are anxiolytic, sedative/hypnotic, anti-convulsant and muscle relaxantbut one of the first non-benzodiazepine ligandsdiscovered, which was able to displace thebenzodiazepines from their binding sites, wasethyl -carboline-3-carboxylate ( -CCE). Thiscompound has effects which are diametricallyopposed to those of the classicalbenzodiazepines, i.e. it is pro-convulsant. It wastermed an inverse agonist with the classicalbenzodiazepines being then classified as
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agonists. Indeed, electrophysiologicalexperiments with the inverse agonistsshow that they shift the GABA concentration-response curve to the right, thus decreasing thepotency of the natural transmitter. While theagonist benzodiazepine site ligands increasechannel opening frequency, the inverse agonistsdecrease it. It is now clear that within thisseries of -carbolines, whereas the ethyl ester ispro-convulsant and thus a partial inverseagonist, the propyl ester is essentially devoid ofefficacy and thus an antagonist. At the otherend of the scale a methyl ester analogue, 6,7-dimethoxy methyl -carboline-3-carboxylate(DMCM), is overtly convulsant and thus a fullinverse agonist.
Attempts to delineate the functional importanceof individual GABA receptor subunits using thegene knockout technology have proved lessthan fruitful with the 3 and 2 provingneonatally lethal while the 6 knockoutmouse displayed no overt phenotype.However, a knock-in approach, which makesuse of the histidine/arginine exchange of thesubunits mentioned above, has demonstratedthat particular GABA receptor subtypesmediate distinct aspects of benzodiazepinepharmacological profile: those receptorscontaining the 1 subunit are responsible forthe sedative effects while those containing the
2 subunit are responsible for their anxiolyticeffects. This knowledge will undoubtedly provevaluable in the development of agents with arestricted pharmacological profile and a recentreview of the patent literature suggests that thisapproach is well advanced.
The barbiturates also produce many of theireffects by interaction with the GABA receptorsof the mammalian CNS. Like thebenzodiazepines, they shift the GABAconcentration-response curve to the left butunlike the agonist benzodiazepines, thebarbiturates also increase the maximumresponse. They clearly interact with a distinctallosteric site; the barbiturates augment theGABA mediated current by increasing theaverage channel open time but have little effecton channel opening frequency. Whereas thebenzodiazepines require the presence of asubunit within the GABA receptor oligomer toexert their effects this is not the case for thebarbiturates. It is also clear that the barbiturates,at high concentrations, are able to open GABAreceptor channels directly, which alsodistinguishes them from thebenzodiazepines.
In addition to allosteric sites for thebenzodiazepines and barbiturates, the GABAreceptors also exhibit high affinity recognitionsites for certain steroids. The observation thatalphaxalone, the synthetic steroid generalanaesthetic, was able to cause stereoselectivepotentiation of GABA receptor mediatedresponses in cuneate nucleus slices from ratbrain was subsequently confirmed in voltageclamp studies conducted in both neuronal andadrenomedullary chromaffin cells. The
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(Bold text denotes compounds available from Tocris)
Figure 4. Structures of selected benzodiazepinesite ligands
Diazepam
Zopiclone (Cat. No. 1094)
L-655,708 (Cat. No. 1327)
�-CCP (Cat. No. 0612)
Flumazenil (Cat. No. 1328)
Zolpidem (Cat. No. 0655)
Midazolam Zaleplon
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progesterone metabolites 3 -hydroxy-5 -pregnan-20-one (allopregnanolone) and 3 -hydroxy-5 -pregnan-20-one (pregnanolone)together with 3 ,21-dihydroxy-5 -pregnan-20-one (allotetrahydrodeoxycorticosterone) are evenmore potent than alphaxalone (Figure 5). It isnow clear that these steroids facilitate GABAmediated responses via recognition sites distinctfrom both the barbiturates and thebenzodiazepines. Mechanistically the activesteroid analogues appear to produce their effectsby increases in both channel open time andopening frequency; like the barbiturates thesteroids directly activate GABA receptormediated channels at high concentrations. Theneuroactive steroids show limited GABAreceptor subtype specificity. However, it appearsthat in those receptors containing the 4 subunit,GABA currents are inhibited by the naturallyoccurring pregnanolone and alphaxalone whilethey are stimulated in other GABA receptors.The subunit also appears to markedly increasethe efficacy of both pregnanolone andalphaxalone in 4 3 receptors compared tothose comprised of 4 3 2 subunits. Theseobservations are particularly interesting in viewof the plasticity of both 4 and subunitexpression associated with pregnancy,observations which may prove to have a bearingon the mood changes associated not only withpremenstrual stress but also those experiencedin postpartum depression.
There is now general consensus that the GABAreceptor plays a significant role in generalanaesthesia. These receptors fulfil the necessarycriteria being sensitive to clinically relevantconcentrations of the anaesthetic agents
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together with exhibiting the expectedstereospecific effects. The volatileanaesthetics, the halogenated ethers andalkanes together with chloroform and diethylether are differentiated from the intravenousanaesthetics such as the barbiturates, propofoland etomidate by their mechanism of action, butboth groups appear to facilitate GABA mediatedinhibition at the GABA receptor. While thereremains a good deal of work to do, studies overthe past decade have provided a significantbody of evidence to address the sites on thereceptor protein which form the targets forinteraction with the anaesthetics. Notsurprisingly these agents appear to bind tohydrophobic pockets within the protein althoughdifferences have been identified with, forexample, a specific aspartate residue withinTM2 being required for etomidate sensitivity butof no consequence to the activity ofpentobarbitone, propofol or the anaestheticsteroids.
Although the majority of studies have focussedon the GABA receptor it is clear that certainanaesthetics, such as ketamine, nitrous oxideand xenon do not produce their effects throughthis receptor but probably by inhibition of the -methyl- -aspartate receptor. It is also clear thatmany of the anaesthetics interact with otherligand gated ion channels, in addition to theNMDA receptor, with pronounced effects beingseen on the neuronal nicotinic acetylcholinereceptors, particularly those containing the 4subunit, and the 5-HT receptor. Undoubtedlythe potential target population will expand asthese studies progress.
In addition to the GABA receptors there is adistinct class of ligand gated ion channels thatare activated by GABA; referred to as theGABA receptor. The natural agonist GABA isabout an order of magnitude more potent at theGABA receptors than at the most common ofthe GABA receptors. The GABA receptors areactivated by -aminocrotonic acid (CACA),which is not recognised by either the GABA orGABA receptors, suggesting that theyrecognise the partially folded conformation ofGABA (Figure 1). GABA receptors are notblocked by bicuculline and do not recognise thebenzodiazepines, barbiturates or theneuroactive steroids but, like GABA receptorsare blocked by picrotoxin, while 1,2,5,6-tetrahydropyridine-4-yl methyl phosphinic acid(TPMPA; Figure 2) appears to inhibit GABAreceptors selectively. Pharmacologically theyare thus quite distinct. However, molecularcloning studies have revealed that thispharmacological profile is remarkably similar tothat exhibited by the subunits when expressedectopically. Two homologous subunits, 1and 2, have been identified in man and thesecan be expressed as homomers or heteromers,but do not co-assemble with any of the GABAreceptor subunits. The DNAs are encoded onchromosome 6 in man, distinct from the clustersof GABA receptor subunit genes which arefound on chromosomes 4, 5, 15 and X; thesubunits are between 30 and 38% homologous
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Figure 5. Structures of selected steroids,barbiturates and anaesthetics that interact withthe GABA receptorA
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Enflurane
Alphaxolone
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Allopregnanolone3 -hydroxy-5 -pregnan-20-one� �
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to the GABA receptor subunits at the aminoacid level. In the important TM2 region of thesequence, they show greater homology to theglycine subunits than to any of the GABAreceptor subunits. It is assumed that they formpentameric assemblies, similar to the othermembers of the ligand gated ion channel family,which enclose a chloride selective channel. Thesingle channel conductance of the homomeric orheteromeric receptors composed of subunitsis smaller (around 7 pS) than that exhibited bythe GABA receptors (25-30 pS) and the gatingkinetics are quite distinct, with both theactivation and deactivation time constants beingvery slow. These homomers or heteromersalso appear to be remarkably resistant todesensitisation in the presence of highconcentrations of the agonist. While it is clearthat the homomeric and heteromericcombinations of the subunits display distinctbiophysical and recognition characteristicsthey do generally mirror, quite closely, theGABA receptors that have been characterisedin retinal bipolar cells from a number of species.Discussions continue with regard to thenomenclature: do we use sequence homology toclassify proteins or their functional andrecognition characteristics. This question willnot easily be resolved: a man with one watchknows what time it is but a man with twowatches is never quite sure!
GABA also activates metabotropic GABAreceptors, which are widely distributed within thecentral nervous system and also in peripheralautonomic terminals. Their activation causes aninhibition of both basal and forskolin stimulatedadenylate cyclase activity together with adecrease in Ca and an increase in Kconductance in neuronal membranes. Thereceptors are activated by baclofen, used in thetreatment of spasticity, ( )-baclofen being the
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active isomer (Figure 1). There is evidence thatGABA receptor agonists may be useful in thetreatment of pain and to reduce the craving fordrugs of addiction. There is limited informationon the therapeutic potential of GABA receptorantagonists but there is support for the idea thatthey may prove valuable in the treatment ofabsence epilepsy and as cognition enhancers.
GABA receptors have now been identified byexpression cloning using the high affinity ligandCGP-64213. Functional receptors are formedonly after heterodimerization of GABA andGABA (previously known as GBR1 andGBR2) by interaction through their C-termini, thefirst time that this form of 1:1 stoichiometry hasbeen identified within this family. Bothsubunits are members of the 7-transmembranereceptor family that show over 30% sequencehomology to the metabotropic glutamatereceptors. A number of splice variants havebeen identified for both GABA andGABA . The mRNA encoding GABA isfound exclusively in neurones but GABA isfound in both neurones and glia.Immunoprecipitation studies suggest thatGABA and GABA are always found asheterodimers although the paucity of mRNA forGABA in the striatum has led to suggestionsthat further subunits remain to be identified.There have been suggestions that the two mostwidely studied splice variants GABA andGABA my be differentially located within thecell, the former being pre-synaptic while thelatter is found post-synaptically. The largedeletion at the N-terminus which occurs in splicevariant GABA is consistent with a differentialsubcellular localisation and it will be interestingto see the whether or not this is region specific.
Although a significant number of phosphinic acidderivatives have been synthesised as GABAreceptor agonists few exceed the potency of( )-baclofen and none has proved useful in the
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Table 2. Comparative pharmacology of GABA receptors
Compound GABA GABA GABA Reference
Agonist Agonist Agonist
Agonist Inactive Partial agonist 6, 8, 91
P4S
Inactive Inactive Partial agonist 8
Inactive
Antagonist Inactive Inactive 6, 8
Antagonist Inactive Antagonist 6, 8
Inactive Antagonist Inactive 76
Inactive Antagonist
A B C
GABA
Muscimol
Isoguvacine
THIP
TACA
CACA
( )-Baclofen
Bicuculline
Picrotoxin
CGP 35348
CGP 54626
Agonist Inactive Antagonist 6, 8
Agonist Inactive Antagonist 6, 8
Agonist Inactive Antagonist 6, 8
Agonist Inactive Agonist 8
Agonist Inactive 6, 8
Inactive 76
CGP 64213 Inactive Antagonist Inactive 76
Inactive Antagonist Inactive 76
Inactive Inactive Antagonist 8, 86, 87
R
SCH 50911
TPMPA
(Bold text denotes compounds available from Tocris)
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differentiation of the distinct receptor subtypes.However, recently it has been reported thatgabapentin selectively activates the GABAheterodimer coupled to inwardly rectifyingpotassium channels expressed inoocytes but this did not occur with GABA orGABA . While the early specific GABAreceptor antagonists suffered from a limitedpotency with phaclofen, for example, displayingan affinity of only 100 M, a number of selective,high affinity and systemically active antagonistsare now available, although most rely on thephosphinic acid moiety (Figure 2). It isundoubtedly only a matter of time beforesubtype specific antagonists will becomeavailable.
Like the metabotropic glutamate receptors, boththe GABA and GABA subunits possess alarge extracellular N-terminal tail which hasbeen modelled by homology on the bacterialperiplasmic binding protein motif characterisedby two globular domains connected by a hingeregion. It has been suggested that agonistactivation relies on the closure of the twoglobular domains subsequent to agonist binding:the venus fly-trap model. A limited number ofmutagenic studies have been carried out todelineate the residues important in agonist andantagonist recognition but the nature of theinteraction between the two proteins within theheterodimer, in both ligand recognition and theirliaison with the G-proteins integral to receptorfunction remains to be investigated.
The GABA receptors remain somewhat forlorn.Their ubiquitous distribution within the CNS,their proven importance in the modulation oftransmitter release and the late inhibitorypostsynaptic potential promise a good deal astargets for pharmacological intervention.
(B1a,2)
(B1b,2)
(B1c,2)83
B
(B1) (B2)
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References
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1314 15
1617
1819 20
2122 23
2425 26
2728
29
30 3132
3334 35
3637 38
3940 41
4243
44
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Bormann and ClaphamAdams and Brown
AkaikeMathers Barker
Barker and MathersHamill Bormann
MacdonaldTwyman Bowery
TwymanRichards
Stephenson and Barnard
Schofield Hevers andLuddens Nayeem
WhitingWisden Farrar
Amin and WeissSmith and Olsen Brejc
Squires and BraestrupMohler and Okada Sieghart
DuncalfeWieland
Squires
3
226 5
7
71 290
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17 222
249
275 190
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250 391
1
322 212
391
385 410
423
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445 7
328
18
62 38
12
274
366 16
411
266 267
47 271
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PritchettNicoll Simmonds
Harrison and SimmondsCottrell
Harrison Twyman andMacdonald Adkins
SmithSmith Franks and Lieb
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KaupmannWhite Kuner
Billinton NgGalvez
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QuirkHadingham
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However, this has been compromised, untilrecently, by the paucity of potent, systemicallyactive and selective ligands. The molecularcloning of their constituent cDNAs coupled withthe discovery that they function as heterodimershas now engendered a new urgency to researchin this arena and the future promises to beexciting.
Interest in the receptors for GABA, the majorinhibitory transmitter in the CNS, has beendeveloped, with varying degrees of enthusiasm,over the past 40 years. We now have agonistsand antagonists which allow us to differentiate,experimentally at least, between responsesmediated by the three pharmacologically distinctreceptor families with which they interact (Table2). The information base is most extensive forthe GABA receptors, driven largely byobservations that these proteins are the targetsfor a number of drugs with significant clinicalimportance. The expansion continues with theconviction that this almost bewilderingcomplexity can be harnessed for the nextgeneration of pharmacological agents with amore restricted profile of activity. The GABAreceptors remain the rather poor cousins in thesense that their potential is not yet realised. Thedevelopments over the past 7 years or morehave delivered a promissory note that isproducing significant investment and many holdreal conviction in their future. Surveys of theliterature would suggest that the GABAreceptors have generated rather more interestfrom the nomenclaturists than is warranted.However, recent evidence that their distributionis more diverse than previously thought couldprovide a further extension to the potentialinhibitory control of the central nervous system.
A
B
C
88
Conclusions
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GABA Receptors, Tocris Reviews No. 20, March 2002
©2002 Tocris CooksonPublished and distributed by Tocris Cookson, Bristol, UK
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GABA Receptor Compounds
Benzodiazepines and Related Compounds
GABA Receptor Compounds
A
B
Agonists
Antagonists
Other
Agonists
Antagonists
0344 GABA ..............................Endogenous agonist0235 Isoguvacine .....................Specific GABA agonist0289 Muscimol .........................Potent GABA agonist0381 Quisqualamine ................Weak GABA agonist0181 TACA...............................GABA agonist. Also GABA-
substrate and GABA inhibitor0807 THIP ................................GABA agonist0180 ZAPA ...............................Agonist at ‘low affinity’
receptor. More potent than GABA/muscimol
0130 (+)-Bicuculline .................Potent GABA antagonist0109 (-)-Bicuculline .................Water soluble GABA antagonist
0131 (-)-Bicuculline .................Water soluble GABA antagonist
1128 Picrotoxin ........................GABA receptor antagonist1262 SR 95531 .......................Selective, competitive GABA
receptor antagonist
0311 CHEB ..............................Convulsant barbiturate0881 Chlormethiazole ..............Potentiates GABA receptor function0505 Dihydroergotoxine ...........Binds with high affinity to GABA
receptor Cl channel1471 Etomidate ........................GABA mimetic and GABA
modulatory agent
0830 Primidone ........................Potentiates GABA receptor function
0865 1-Amino-5-bromouracil....Agonist at benzodiazepine-GABAsite
0405 -CCB .............................Inverse agonist, putativeendogenous ligand
0612 -CCP .............................Inverse agonist0456 Chlormezanone...............Skeletal muscle relaxant0554 FG-7142 ..........................Inverse agonist0658 FGIN-1-27 .......................Potent, specific ligand for
mitochondrial DBI receptor0659 FGIN-1-43 .......................Potent, specific ligand for
mitochondrial DBI receptor1328 Flumazenil .......................Benzodiazepine antagonist0770 GBLD 345 .......................High affinity agonist
R1327[ H]-L-655,708 ...........Radiolabelled form of (1327)0670 PK-11195.........................Antagonist at peripheral BDZ
receptors0655 Zolpidem * .......................Benzodiazepine agonist1094 Zopiclone.........................Benzodiazepine agonist
0417 ( )-Baclofen..................Selective GABA agonist0796 ( )-Baclofen ....................Active enantiomer0344 GABA ..............................Endogenous agonist0379 SKF 97541 ......................Extremely potent GABA agonist
1245 CGP 35348 .....................Brain penetrant, selective GABAantagonist
1247 CGP 46381 .....................Brain penetrant, selective GABAantagonist
1246 CGP 52432 .....................Potent, selective GABAantagonist
A
3
B
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
A
B
B
B
B
Tuptake
GABA
methobromide
methochloride
1295 Loreclezole......................Subtype-selective GABAreceptor modulator
1327 L-655,708 ........................Selective for 5-containing GABAreceptors
–
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b
RSR
1088 CGP 54626 .....................Potent, selective GABAantagonist
R1088[ H]-CGP 54626 ........Radiolabelled form of (1088)1248 CGP 55845 .....................Potent, selective GABA
antagonist0245 2-Hydroxysaclofen...........Selective GABA antagonist,
more potent than saclofen(0246)
0178 Phaclofen ........................Weak, selective GABAantagonist
0246 Saclofen ..........................Selective GABA antagonist0984 SCH 50911 ......................Selective, competitive, orally
active GABA antagonist
0179 CACA ..............................Partial GABA agonist0344 GABA ..............................Endogenous agonist0815 Imidazole-4-acetic acid....Partial GABA agonist0289 Muscimol .........................Partial GABA agonist0181 TACA...............................GABA agonist
0807 THIP ................................GABA antagonist1040 TPMPA *..........................Selective GABA antagonist0180 ZAPA ...............................GABA antagonist
0206 -Alanine .........................Distinguishes GABAtransporters
1296 CI 966..............................Selective inhibitor of GAT10234 Guvacine .........................Specific GABA uptake inhibitor0236 (±)-Nipecotic acid ............GABA uptake inhibitor0768 Riluzole ...........................GABA uptake inhibitor. Also
glutamate release inhibitor1081 SKF 89976A....................Potent GABA uptake inhibitor.
Penetrates blood brain barrier0181 TACA...............................GABA uptake inhibitor. Also
GABA agonist and substratefor GABA-T
0806 Gabapentin......................Anticonvulsant. Increases brainGABA
0538 -4-Hydroxycrotonic ..GHB receptor ligand
1260 Ivermectin........................Modulates glutamate/GABA-activated CI channels
0386 3-Methyl-GABA ...............Activator of GABA amino-transferase
0780 NCS-382 .........................Antagonist of hydroxybutyricacid
0939 Propofol ...........................Potentiates GABA receptor-mediated inhibition and inhibitsNMDA receptor-mediatedexcitation
0808 Vigabatrin ........................GABA-T inhibitor
*
B
B
B
B
B
B
C
C
C
3C
B
C
C
C
C
A
A
3
C
-
b
Other1513 CGP 7930 .......................Positive modulator at GABA
receptors1514 CGP 13501 .....................Positive modulator at GABA
receptors
R1301[ H]-P4MPA ...............GABA antagonist radioligand0379 SKF 97541 ......................Potent GABA agonist. Also
GABA antagonist
acid
B
B
Agonists
Antagonists
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b
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GABA Receptor Compounds
Inhibitory Amino Acid Uptake Inhibitors
Miscellaneous GABA/Glycine Receptor Compounds
C
GABA[Rev](0302)