Aminergic neurones, phenolamine pathways and octopamine

TIP.¥ - Ma.v 1981

113

maceutical industry which require urgent answers; helping to find some of them offers a challenging and exciting future!

Readinglist l W e l l s • N . ( 1 9 R n ) . ~ e d i c i n e ~ : 5 0 year~ ofpro.¢re~s

1930+1980. Officeof ttcahh Economic',,I,ondon 2 I,aurence, D. R. alld Black, J, W. (1978) rtle medicineyou take. Font;ma,Glasgow 3 Steward, F. and Wibberley. G. (19811)Nan+re' (London) 284. II 8-12{1 4 Dixon.B. (1980) Trend~Phannacol.Sci. 1. I-ll 5 Weathcrall.M. {1979) Pharm.J. 223. 345-347

l)r +~t. D. Day graduated in P h a r m a c y ]rom the ~t lu,,d o]'Pharmacv, Univergity o [ L o n d o n tn lOttO Itt" ~pv,tr o n e year a~ a re~earch pltar~na(olo.~t~l t~ittt Sht~ &

Aminergic neurones, phenolamine pathways and octopamine

Ita~er t ld arid Illt'n rl'lt4¢tled lt~ ~ ~qltare' Io ol+ltatl a I'h D. u n d e r the +hr,'tflon ,~l l'e,,, "~or ~t. J. R a n d I t , ~pent the ),ear 196 t 454 on po~fih~ toral fl'llo~ ~hq~ at the Uh'~ehmd ('hlllt. ()ht~t, I .~ t aPtd reo~rnol to I ' K to It'a(h Phartn,t~ ~," ~[lidt'llls al I¢rtghloPt l%l*tcchnt( l l g h 4 .fiTI. l i e mt,~ vd to the lh'partmvnr • it Pharlna 0 . [:'llt['P¥~ll~ O] .t ~tetll field ~l,l~ s14(¢ ¢'~Sli t'lY ,tl,lnanted ~ll*nltlt l.t't tl~r('¢ ¢ I q 7 0 t ;rod I'Ira~fi,r 119 "21 tie ~Oqll lhe lwr~od 1975 l q ~" ,t~ t, e n t , , t I e , / u , , ' r in Pharma~olo¢~ + ,a the [ t e d u a l '~(hool. t ~n+erstt~' ot ~,'oftlrlk,llattt. In / O 7 7 I)r Ihax r e , t e d itlto rll~. Pharma,,'utt+,d l~ldlt~l.'~ as t h'a,i oJ H:oblglt ~d R,'~+',*r( h It tdl Rt t l~at t~ ( obn,:n I h l . Ihdl, and bet a m c / b e e , It*r '+.l R<'xt',zr, If ot 1~7q.

mammalian li~su~s. Subsequently, lahorato D animals and man x~,'re ~,hox~n to excrete increased Io, cb, o f p-h~drox~mandelcic acid. a m a l o r metabt,lite ,,f

oetopamine, after the admini,,trali(,n of MAO inhibitor~,. The uptake. -ttwage and relea,,e o f oelopanline ffoln c(ffc-

Donald F. H. Dougan and Denis N. Wade

cholaminergic ncr,,c tt.'rminal,, atter inhibition o! M A O led to the amine b¢,ng characterized as a "fal,,c tran,,mittcr m

Department o f Clinical Pharmacology. St. I/incent'~ Ho.wital. C!ni~er.~it) ' o f .Ve~ South Wale~, Kemington 2033. N.S. W., Australia

mammalian nep, ous tissue. Until recentl.,., stud~ of the amine ~a~, limited b.x a~ailable methodolog.,,. Further.

One o f the most intriguing clues in the study o f n aropsychiatric disorders" is the ability o f amphetamine to induce psychotic episodes in man. Recent studies indicate that the production ofhydro,~vlated metabolites o f amphetamine during psychotic episodes in man is correlated with the severity o f t he amphetamine-induced psychosis. Further. two malor hvdroxylated metabolites o f amphetambte are alpha-methylated attalogues of the namralty occurring brain phenolamines, octopamine attd O,ramine. The tae o f the G C / M S method of estimating brain phenolamines has provided the means to invest(~ate the rehttionship between brain aminergic neurones, the phenolambws, oclopamine and tvramb~e and neurop~ychiatric disorders directl.v. Specific neurotransmitters have been associated with relatively few neurones in the mammalian central nervous system (CNS), even within the relatively well studied amine-containing areas• The brain phenolamines, octopamine and tyramine, accumulate in aminergic neurones but only in nanogram quantities, unlike the catccholamines, dopamine and noradrenaline, which are present in microgram quantities. Octopamine has been thought of as an unimportant end product of tyrosine metabolism which accumulates in catecholaminergic nerve endings after inhibition of the enzyme monoamine oxidase (MAO). However, in recent years, good indirect evidence has accumulated indicating that octopamine has a neurotransmitter role in some invertebrate species*, and mo+e recently, specific receptors for octopamine have been found in the central nervous system of the rat.

Octopamine is present in rat brain in early stages of development at molar concentrations higher than that of the catecholamines. In addition to ph3logenetic implications, this raises the possibilit.~ that octopamine may have a function of its own in the fetal brain z. This article will review recent data ~ hich indicate that octopamine in the mammalian CNS deserves consideration as an aminergic neurotransmitter in its ov, n right rather than being characterized as a 'false or "substitute transmitter'. Octopamine was first discovered in the salivary ghmds of the octopus in 1938 b~ Vialli and Erspamer during a search for the active principle in the ~enom of this invertebrate. After chemical and pharmacological investigations, Erspamer identified the structure of octopaminc as 1p-hydroxyphenylethanolamine and described its sympathomimetic effects in

aminergic path~ays containin~ catecholamines could be ~isualizcd ~ith the aid ol fluorescent histochcmical technique,. ~hcrea:, pmati~e neurones containing oclopamine or other phenolanline~ Ldonc could not be demonqrated t, phenolamines form relativel} ~eak fluorofors, Hov, e~er, it appears, that the technique', of expcrimentall.,.-induced, di',crete, brain lesions combined v.ith ~a,, chronlatograph)/mass spcclromet~ (tiC/MS) to detect and measure pheno amine, can bc used to demonstr:,te indircctt', lhc presence of aminergic path~a~.s in the I~rain ~ hich sD.'cifically contain octopanline. l h e presence of independent octopanlinecontainin,2 neuronal s + , . M e n t s in the n l a n l malian brain ~ould xtrongl.~ ,upport ,t functional role for octt~palnine a'~ at tlcurotransmitter.

Me&suringoctopamine Detection of the phcnokmunc, octopamine {OCT} and its related analogues, progressed rapidl} during the t,:trlx lqT0', v,ith the development of sens ti,,e radioenzymatic methods of assay. Yhese techniques, combined v, ith variou,, solvent extractions or paper chromatoT, r:tphx to increase the specificity, resulted in estimates of OCT in ~arious areas of the 't i~¢~l~t N,oith H~qland Birth ¢,]1~.t1 Prt,~ 1,~i

T I P S - ,~da)' 1981

114 mammalian brain which varied between 0 and I0 ng g-' wet weight of tissue. This variation between values recorded in the literature may reflect early technical problems, particularly with the specificity of the assay in the hands of various workers. For example, ihc enzyme used for the assay procedure, phe~yle([mii~,l.tr.~.L t,methyl transferase (PNMT) (EC 2.2.1-) accepts both meta-octoparaine and paraoctopamine as well as noradrenaline, normetanephrine and phenylethanolamine as alternative substrates. In this respect, Daniclson and others a have rightly indicated that any interpretation of the effects of denervation or pharmacological manipulation are rendered suspect or even invalid if a large amount of an unknown raetaholite is included in the endogenous value~. This may be of particular importance ia explaining the paradc~xical effects of ~ m e psychoactive drugs which are recorded using the radio-enzymatic technique to reduce the noradrenaline concentration of various brain areas but to produce little change in octopamine concentrations. Measurement of phenolamines by a directly and highly specific method of assay such as GC/MS avoids this "ahernative substrate" problem and overcomes many of the disadvantages a.~socia:ted with radioenzymatic methods. Furthermore. mass fragmentography can be used to measure simultaneousliy a nwi~ber of naturally occumng and synthetic phenolaraines. Additionally. metabolic transformation of pheno[amines can readily be studied and drug- or lesion-induced changes easily followed. The gas chromatograph serves to resolve the various phenolaraines from each o~her while the mass spectrometer acts as a detector for quantitation by measuring the ion densiD' of the specific characteristic fragments (ma~ to charge ratio) ,of each compound. Prior to estimation, volatile phenolamine derivatives are prepared by reacting the phenolamines with pentafluoropropionic anhydride. The first use of the mass spectrometer to mea,ure OCT in nervous tissue was made by ~uek and others" during 1976, who identified OCT in the rat pineat organ at a

concentration of approximately 0.07 ng per pineal compared to their estimate of 0.14 ng when determined by the radioenzTmatic assay. The sensitivity of the ma.~s spectrometer as a detector in the chemical ionization (CI) mode was increased by Buck and others who introduced purificauon of the extracted phenotamines on a precolumn containing an ion-exchange resin. The direct use of ethyl acetate extraction without the use of ion-exchange purificalion has not proved satisfactory. Thus Karoum and others s failed to detect OCT in concentrations of more than 0.5 ng g-~ in the hypothalamus, an area estimated by other laboratories to contain 3-4 ng g-~ wet weight of tissue. Octopamine synthesis A summary of our present understanding of the biosynthetic pathways for p-oetopamine is shown in F'ig. 1. Administration of the amino acids. phcnylalanine or tyrosine produces a small increase in brain OCT as does the administration of phenylethylamine (PEA). Prelreatment of rats with MAO inhibitors greatly enhances t h e ~ changes. The studies of Harmer and Horn 6 demonstrating that the increase in brain OCT concenlration was greater after the administration of PEA than that resulting from the administration of phenylalanine or tyrosine suggested that two major pathways for OCT biosynthesis exist in the mammalian brain. One pathway begins with the decarboxylation of tyrosine to tyramine (TYR) which, in turn, is /3-hydroxylated to OCT. The other starts with the decarboxylation of phenylalanine to PEA which, in turn. is converted to OCT via the intermediates TYR or phenethamilamine. The marked increases in OCT levels found after the administration of precursors and MAO inhibilors is indicative of the high rate of turnover of OCT in rat brain. Further. the administration ot a-methylated precursors of OCT which are not substrates for MAO also leads to significant accumulation of a-methylated

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analogue:,.The ability Io use c~-me~hylated precursors of OCT analogues in animals not pretreated with MAO inhibi~ors has provided valuable informatkm on the rclalive imporlance of the two pt~stulated phenolaraine pa[hways. Fluoromelric raethods of assay 7 have detccled the presence oft,-methylt~, raminc (c~-MTYR) and ~-melhylocmpaminc (t~-MOCT) in rat striatum suggesting that these compounds also accumulate after administration of ~t-methyltyrosinc (aMTYRO)- The,~ findings have hcen confirmed by direct GC/MS measurement. However, the formation of a-MTYR and a-MOCT from d-amphetamine (D-AMP) confirms thai the alternative PEA-pathway for OCT synthesis also exists. Further, ot-MOCT arid its immediate precursor a-M-I'YR have been shown by GC/MS methods to accumulate in the hypothalamus and striatum after administration of D-AMP. The relatively impol rant role of this ~a~tler pathway is indicated by the relative amounts of t~-MTYR and ot-MOCT formed from D-AMP and a-MTYRO which have been measured by the GC/MS technique in our laboratory Table Ilt~. Surprisingly. the administrat+on of a-methylphenylalanine (a-MPA) did not lead to the formation of para-hydroxylated compounds but to meta-hyroxylated t~-methylated raetabolites of OCT and T $ R (Table I). The precise reason for the metahydroxylation of G-MPA is not known. Until recently, the effect of tyrosine hydroxylase inhibitors on the accumulation of OCT in the brain has not been clea~. On the one hand. Brandau and Axelrod g, using a radio-enzymatic technique, reported an unexpected reduction in ~he accumulation of OCT after the +dministration of a tyrosine hydroxylase ,nhibitor. This finding was unexpected as an increase in tyrosine and TYR concentrations in the brain could be predicted. On the other hand, these data may be explained by tyrosine hydroxylase inhibilors releasing OCT in the CNS. However, experiments in our laboratory indicate that after single dose administration of a-MTYRO, the accumulation of ot-methylated metabolites of a-MTYRO is accompanied by a raoderate increase in the levels of OCT and TYR in rat striatum (Table I). It is generally assumed that a-MOCT is formed in the brain within noradrenergic granules in which dopamine/~-hydroxylase is located. Accordingly. by destroying noradrenergic nerve terminals, the fl-hydroxylation of c~-MTYR formed from

TIPS -May 1981

I !5

T A B L E i. l)eteelion by GC/MS of ,~-methylt~:lopamin¢ and ¢,-mclhyltyramine tn rat striatum idler admmiqratum ol .-meth~tated pr¢cup,~,t~ . t ,~ct,,pammc

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Meanvaluesare expressedin ng g ' (numberof experimentsin bra, keis). All drugs~,ereadmlni,,lered intrapernmncall) 211h beforethe amm4Nv,crc ~a~.tlhccd D-AMP is prevented. This interpretation is based on two important assumptions; ( ! ) the enzyme dopamine ,8-hydroxylase (DBH) is contained solely in noradrenergic nerve terminals and (2) 6-hydroxydopamine (6HD) selectively destroys noradrenergic nerve terminals and not other aminergic terminals which may contain DBH. Recent studies on the intraneuronal location of the enzymes involved in the synthesis of eateeholamines and on the specificity of the degenerative effects of 6HD on aminergie neurones in the brain indicate that the above assumptions may not be totally correct. There is evidence, for example, that possible adrenaline containing neurones may exist in lhe brain which not only contain the enzymes DBH and PNMT involved in the synthesis of adrenaline but also that these neurones seem to be resistant to the neurotoxic action of 6HD. The nature of the aminergic terminals which accumulate OCT and TYR in the striatum is not clear. The effects of brain lesions severing the ascending dopaminergic projection to the striatum on OCT and TYR concentrations in the striatum have recently been reported from this laboratory. The mass fragmentographie technique was used to determine striatal concentrations of both OCT and TYR after cutting unilaterally the major monoaminergic pathways to the striatum. Striatai concentrations of dopamine were also measured by GC/MS. All unilesioned rats were given pargyline 75 mg kg -~ i.p. 2 h before decapitation. In preliminary experiments t°, these lesions reduced striata~ concentrations of dopamine (DA) and TYR by 84 and 80%, respectively, on the ipsilateral side but, unexpectedly, increased striatal levels of OCT by 115%. These data suggest that OCT is not accumulated in dopaminergic nerve terminals in the 5triatum, although a large proportion of TYR may be accumulated. Although these data strongly suggest the presence of specific octopaminergic neurones in the striatum, an alternative explanation is possible. Small quantities of noradrenaline

(NA) have been found in the striatum indicating the presence of noradrenergic terminals in which OCT may accumulate. Experiments are in progress to determine if the increase in OCT levels in the slriatum after the above lesions is accompmfied by a concomitant increase or decrease in the concentration of NA. The function of these small amounts of NA, OCT and TYR in the striatum is not clear.

Octopaminereceptors lontophoretie studies of t h e ~ amines on caudate neurones show that the above amines all have depressant effects on sDmtaneously active and glutamate-excited neurones similar to the depressant effects of DA. The powerful depressant effects on neuronal firing rhat follow from an application of trace amines could be due to a direct effect on receptors or to the release of neurotransmitters'. Other work involving micro-iontophoretic application of phenolamines and catecholamines has indicated the pre~ncc of specific OCT receptors in rat brain. Hicks and McLennan n compared the actions of OCT, NA and DA on single cortical neurones of the rat. OCT was found to both excite and depress neurones of the cortex. Frequently, cells depressed by NA were excited by OCT, the time course of action also being different. Further. DAelicited excitations were observed but were not correlated with OCT-elicited effects. As OCT-mediated effects do not correlate wi~h either DA or NA, it seems reasonable to ascribe a direct action for OCT at itsmut specific receptors. The results with pharmacological antagonists also offers compelling evidence th:.~t OCT receptors do exist and that they arc distinct from those for the cateeholamines. For example, propranolol and a-flupen,. thixol, respectively, prevent NA- and DA-induced changes in firing rate. Neither of these compounds reduced tile effects elicited by OCT suggesting that the receptors mediating the actions of OCT are pharmacologically distinct from those of eatccbolamines.

Although stereosdecli~,e receptop, for OCF have been described in both the mollusc Tapes wadmgi ~3 and the rat brain TM, the.,,+appear to have diflerent pharmacological specificity. In the spontaneously beating ventricle of Tapes. metoclopramide and clozapine have been shov.n to be c f f e c l l s c antagonist,~ at the stereo-selective receptors for OCT. but these comt~mnd', apparently are ineffective as antagonist,, of OCT or the catecholamin¢:, I)A and NA in the corlex or the spinal cord of the ra. In this respect, mammalian OCT receplor', do not re~'mble those in the moltu,,c ,,~here clozapme and metociopramide bl4ck lhe cardio-excitatoD OCT receptor,, and not receptors mediating the cardio-excitator) effects of DA. To date. no ,pecific ::ntagtmist has been found for putafi*e m tmmali m OCT receptors. Hinge*or. a, ,nammali m OCT receptors resemble specific receptt,r, for OCT in the Tapes ~entricle in their stereoseleetivit), the ",trier structur:d ,,pecificily of the molluscan receptor max be rueful in ~,ecking more ,,elective conltxmndcapable of blocking O C I in the marqmalian bruin. Further, until a selecti~ e OCT an,'.agon ,t is found t\)r mammalian receptors,, tl:e search t\~r ~,pecific OCT receptor* in the mammalian brain ~ill be complicated b~ tile pre~nce of rcccptor, f,'~l catecholamines. Clinical role It is gencrall) a:,,,umcd that di,.ordcrs of amine metabolism occur in the affi..cti~e disorders, in schizophrenia and in ~onlc metabolic disorder,, including hepanc encephalopath). In the pa>t. the Io~ conc e n t r a l l o n o f p h e n o I a m l n c ~ , in t h e b r a i n h:|r,

ted to tile x ie~ that their occurrence i~ incidental to the s%nthesis of other a m i n e : , s u c h as catecholamincs and serotonin, Ho~ever, Io¢, conccntrationsper ~e do not den} the role of putative neurotransmittcr phenolamines in ps.vchmtric disorders a~ it is the turnover and release of phenolamines at specific nerve terminals which ix functionall~ more important. Several authors ha~c su&~ested that mctabt~lites ef amphetamine may be

TIPS-May

i 16 resl:,0,nsible for the psychotic symptoms afte: prolonged use of high doses. Alphamethyloctopamine is a known metabolite of amphetamine in man while amet hyi-p-methoxyoctopamine (p-methoxy amphetamine) has, on the basis of alleged psychomimetic properties in the rat, been implicated in amphetamine psychosis in man. Further, the studies of Anggard and Gunne ~5 indicate that the hydroxylated metabolites of amphetamine may be associated with amphetamine-induced psychosis. The relationship of the hydroxylatcd metabolites to the' occurrence of psychotic symptoms is further strengthened by the correlation between the intensity of the psychotic manifestations and the urinary output of hydroxylated metabolites. The identity of these compounds was established by gas chromatography, radioactivity monitoring and mas~ spectrometry as p-hydroxya mphetamine, a-methyloctopamine and norephedrine. No pmethoxy-

3 Danielum. T. J.. Boulton. A. A. and Robertsnn, H. A. ( 1977) J. Neurocheot. 29. I 131- I 135 4 Buck. S, H., Murphy, R. C, and M~llinoff,P. B. (1977) Brain Res. 122, 281-2t~7 5 Karoum,F, Nasrallah, 11.,Potkin,S., Chuang, L.. Moyer-Sehwing, J., PhiLlips, L and Wyatt, R.J. (1979) 1. Neurochem. 33, 201-212 6 Harmer, A.J. and Horn, A.S. (1976) 1. Neuroehem. 26, 987-993 7 Doteuehi.M., Wans,C. and Costa, E. (1974) MoL Pharma¢ol. 10, 225-234 8 Duffield, P. D., Dougan, D. F. H., Wade. D.N. and Duffield. A.M. Clin, E W, Pharmacol. Physiol. 7. No. 6. 657--658 9 Brandau, K. and Axelrod. J. (1972)Arch. Pharmacok 273. 123-133 I0 Dougan, D. F. H.. Dul'field,P. H.. Wade. D N.. Duffield, A. M. and Paxinos,G. Clin. Exp. P,harmaeol. Physiol. 7. No. 6,, 658 I1 Henwood. R.W., Boulton. A.A. and PhilLis, J. W. (1979) Brain Res. 164.347-351 12 Hicks. T. P. and McLennan11978) Bt. J. Pharmacol. 64. 485-491 13 Wade, DN. and Dougan, D. F. H. {1979) in Dopaminergie Ergot Derivatives and Motor Func-

1981

lion (Fuxe. I(. and Calne. D. B.. edsL pp 85-ItX). Perga,aoo P, ess. Oxft~rd and New York 14 I)ao. W. P, C. ;rod Walker. R. J. (l'.;7,J) Io, J, PharmacoL 67.132 15 Anggard. E. and Gunne, t.. A. (1975) in Neurop,s'~who.Pharmaeoh~gy (Boissier, J.R.. Hippius. H and Pichol. P.. eds). pp. 253-261,

Excerpta Medica,Amsterdam Denis IV. Wade is I"tamdation Profi,ssor of Clinical Pharmacology' at tile Univers'ity o f New South Wales, Sydney. Australur and Director o f the Department o f Clinical Pharmacology, St. Vincem's Hospital, Sydney. His res~lrch interests have included a long-term interest in Parkinson's Di,sease, levodopa and the control o f dopamincrgic nearones. Donald F. H. Dougan is a National Healdt and Medical Research Council Senior Research Officer at the UniversiD. o f New South Wales. Sydney. Australia. He is currently the recipient o f several NH and MRC grants. His re,~earch interesa relate to the characterizaaon o f multiple receptors" for dopamine and octopamine artd the mass fragmentographic determination o f the e/leers o f psy,choactive drugs on the biosynthesis o f brain phenohvnines.

Benzodiazepine - r e c e p t o r s : a r e t h e re e n d o g e n o u s l i g a n d s in the brain ?

,. p r e c u r ~ r of a - M O C T and possibly p-metho~yamphetamine, although this last conversion has not yet be~n de monstrated. These da~a may indicate a role for one or more of these phenolamine metabolites in the production of psychotic symptoms after chronic use of amphetamine, the closest human model of chronic paranoid schizophrenia. AmFhetamine is chemically similar to the endogenous sympathomimetic a m i n e , ,'EA. As PEA is a precursor of OCT, abnormalities leading to the increased Froductlon of this type of phenolamine may b¢ involved in the ~enesis of some psycho,ses. Further. there is increasing evidence that r!he neuropsychiatrie complications of hepatic failure are closely associated with an increase in brain phenolamine concen~:rations. Increased concentrations of the phenolamines, TYR and OCT, are found m the brain of animals after e~ perimental portal-systemic shunting and in animals and hvmans with hepatic encephalopathy, ,the clinical severity of the encephalopathy correlates closely with the OCT concentra':ion in the plasma and urine. The possibility that naturally occurring phenolamines have an important role in the pathogenesis of psychiatric disorders cannot be. ig~aored.

H. M6hler Pharmaceutical f@search Department, F. Hoffmann.La Roche & Co,, Ltd., CH-4002 Basle, Switzerland.

In analogy to the identification of morphine receptt~rs in the brain and the subsequent isolatioa of opioid peptides, the discovery of benzodiazepine receptors ~'2 has prompted a search for endogenous brain constituents, which may interact with benzodiazepine receptors and thus clarify their possible physiological role. It was hoped that such compounds would provide new molecular terms for the description of brain function and of those diseases which are ameliorated by benzodiazepine treatment like insomnia, anxiety states, muscle spasms or convulsions. This article reviews the attempts to isolate such ligands.

Possible physiological roles of benzodiazepine receptors

Benzodiazepine receptors are the primary target structures in the CNS to which benzodiazepines are bound. A sequence of events is thereby triggered which ultiReading .st mately results in the pharmacological and 1 "Saaved.~a..LM., Brownstein, M.l., Carpender, D.O. an~ Aselrod. J. {1974) Science 185, therapeutic effects of benzodiazepines. The benzodiazepine receptors arc mem364-3:~Y 2 Sa,avedra. I.M.. Coyle. J.T. and Axelrod. 3. brane proteins localized in synaprsesa (I974J1 N,.,uroctlem. 23. 511-515 which, at least in part, could be identified

: ~ ElSg'nF~'I~Irl 1-H!r~a[l~ B.~n~d~al Pro ~s 1981

immunocytochemically as GABAergicL Although a presynaptic localization of these receptors on G A B A neurons cannot be excluded, there is evidence that they are part of the postsynaptic GABA-receptor unit which is operative in the ger, eration of the synaptic potential. The G A B A recepIor unit can be pictured as a supramolecular complex containing functionally related macromolecules such as the G A B A receptor, its associated CI- ionophore, the benzodiazepine receptor and other proteins. The GABA-dependent activation of the CI- ionophore may be modulated allosterically through activation of the benzodiazepine receptor. Such a molecular concept is in line with ~he proposal that the main central effects of benzodiazepines result from an enhancement of GABAergic synaptic transmission in the CNSE However, it should be kept ia mind that benzodiazepine receptors might, in addition, occur in non-GABAergic synapses. Depending or~ the assumptions on the possible physiological role of the receptor, two types of endogenous ligands, if any,