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Behavioral effects of GABA agonists in relation to anxiety and benzodiazepine action
Life Sciences, Vol. 40, pp. 2429-2436 Printed in the U.S.A.
BEHAVIORAL
Pergamon
Journals
EFFECTS OF GABA AGONISTS IN RELATION TO ANXIETY AND BENZODIAZEPINE ACTION R A Shephard
Behavioral Analysis and Behavioral Biology Research Centre and Department of Psychology, University of Ulster at Jordanstown, N e ~ o w n a b b e y , BT37 OQB N. Ireland, UK. (Received
in final
form April
7, 1987)
Summary A considerable body of biochemical and neurophysiological evidence implicates GABA in anxiety and in benzodiazepine action. The present article surveys the behavioral effects of GABA agonists and their interactions with drugs acting at the benzodiazepine receptor in animal anxiety paradigms. Certain GABA agonists, notably valproate, simulate many behavioral actions of benzodiazepines. Moreover, several behavioral studies of the interaction of GABA agonists with benzodiazepines support the hypothesis of a benzodiazepine receptor complex with one or more GABA, benzodiazepine and probably other binding sites. However, there are also a number of anomalous findings of GABA agonist action alone and in combination with benzodiazepines. It is argued that these paradoxical results can better be accounted for in terms of the receptor complex and the distribution of the drugs, rather than by suggesting that the anxiolytic actions of benzodiazepines are not mediated by GABA systems. The potential clinical usefulness of GABA agonists in anxiety is commented upon. Introduction In both phylogenetic and neuroanatomical terms, the neurotransmitter GABA ranks as amongst the most widely distributed and it is currently estimated to account for about a third of all neurotransmission in the central nervous system (i). In view of this, it is unsurprising that drugs affecting GABA systems produce pronounced effects on the central nervous system, including behavioral actions. GABA agonists generally induce sedation and anticonvulsant effects; actions which are also characteristic of benzodiazepine tranquillizers. This parallel has, for some time, been used to support speculation that benzodiazepines might elicit their pharmacological effects, including their anxiolytic actions, by facilitating GAHAergic transmission. More recently, direct support for this hypothesis has been derived from neurophysiological and biochemical experiments (I, 2, 3, 4, 5, 6, 7). Possibly the most salient argument is the evidence for GABA involvement in benzodiazepine receptor binding, which is facilitated by a number of GABA agonists, but inhibited by its antagonists (8, 9). Since the anxiolytic effects of benzodiazepines are related to activation of benzodiazepine receptors (I0), it would be expected that GABA agonists should potentiate benzodiazepine action and/or simulate their behavioral effects, whilst GABA antagonists should induce essentially opposite effects. A number of studies, reviewed elsewhere (ii, 12) have shown GABA antagonists to elicit apparent anxiogenic actions or to attenuate benzodiazepine effects and, whilst some of these experiments are imperfectly designed (Ii, 12) and it is difficult to draw specific conclusions from in-
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supported.
The main p u r p o s e of this review, however, is to e x a m i n e the extent to w h i c h GABA a g o n i s t s have been shown to reproduce the e f f e c t s of benzodiazepine~ in animal a n x i e t y m o d e l s (or c o n f l i c t behaviors) or to p o t e n t i a t e such benzodia z e p i n e actions, as w o u l d be p r e d i c t e d from biochemical and n e u r o p h y s i o l o g i c a l evidence. The nature of these a n x i e t y m o d e l s has f r e q u e n t l y been r e v i e w e d e l s e w h e r e (eg 12, 13, 14, 15) and it should be noted that, logically speaking, these two q u e s t i o n s are i n d e p e n d e n t (12). Thus, in theory G A B A systems c o u l d have a role in the p h y s i o l o q y of anxiety which is entirely i n d e p e n d e n t of the s y s t e m s involved in b e n z o d i a z e p i n e action w h i c h might result in f u l f i l m e n t of the " s t i m u l a t i o n " but not " p o t e n t i a t i o n " prediction. C o n v e r s e l y , and again t h e o r e t i c a l l y , if the role of G A B A in a n x i e t y systems was c o n f i n e d to actions at the b e n z o d ] a z e p i n e receptor complex, then GABA a g o n i s t s might not elicit a n t i - c o n f l i c t e f f e c t s themselves, but could enhance the actions of substances a f f e c t i n g the b e n z o d i a z e p i n e site. Effects
of GABA a ~ o n i s t s
on conflict
behavior
A number of c o m p o u n d s which inhibit the a c t i v i t y of the enzyme GABA t r a n s a m i n a s e are available. Since this enzyme is important in the inactivation of G~BA, i n h i b i t i n 9 it p r o d u c e s i n c r e a s e s in brain GI~BA c o n c e n t r a t i o n s and G A B A t r a n s a m i n a s e inhibitors can be r e g a r d e d as i n d i r e c t l y - a c t i n g G A B A agonists. Drugs e x e r t i n g this action do not, however, g e n e r a l l y elicit benzodiaze p i n e - l i k e a n t i - c o n f l i c t effects in a p p r o p r i a t e behavioral tests, negative findings being r e p o r t e d for a m i n o - o x y a c e t i c acid (16, 1 7 ) , ~ -vinyl GABA (16) and e t h a n o l a m i n e - O - s u l f a t e (18). The last has been c o n t r a d i c t e d by s u b s e q u e n t i n v e s t i g a t i o n s (19), but the balance of e v i d e n c e at p r e s e n t s u g g e s t s that GABA t r a n s a m i n a s e i n h i b i t o r s have no a n t i - c o n f l i c t actions. It may be, of course, that the m o d e s t increases in total brain G A B A c o n c e n t r a t i o n induced by this type of dru 9 simply fail to result in a p p r e c i a b l e increases in functional GABA a c t i v i t y at the synapses relevant to c o n f l i c t or anxiety systems. Rather more c o m p l e x and c o n t r o v e r s J a ! are the e f f e c t s of drugs thought to have direct a g o n i s t action at GABA r e c e p t o r s on c o n f l i c t behavior. Again, lack of b e h a v i o r a l a c t i v i t y has been r e p o r t e d for a nun~0er of agents; namely m u s c i m o l (20, 21), p r o g a b i d e (20) and THIP (16). Apart from the p o s s i b i l i t y that C~BA may not be involved in the p h y s i o l o g y of anxiety (discussed later), two e x p l a n a t i o n s of this lack of e f f e c t have been advanced. The first was simply that these c o m p o u n d s do not r e a d i l y enter the brain following the systemic a d m i n i s t r a t i o n s used and t h e r e f o r e that no a n t i - c o n f l i c t e f f e c t s could be expected. This is supported by d e m o n s t r a t i o n s of a n t i - c o n f l i c t actions of m u s c i m o l (3) and of GABA itself (22, 23), when given intracerebrally. Against this, it has been r e p o r t e d that muscimol a f f e c t s n e u r o p h y s i o l o g i c a l p a r a m e t e r s followin 9 systemic a d m i n i s t r a t i o n s of doses equal to, or even o r d e r s of magnitude below, those which fail to elicit a n t i - c o n f l i c t a c t i o n s (24), and suggested that the a p p a r e n t lack of a c t i v i t y of many GABA a g o n i s t s may be because they affect only the GABA A receptor (24). In support of this hypothesis, it was shown that c o m b i n e d systemic a d m i n i s t r a t i o n of a p p r o p r i a t e doses of G A B A A a g o n i s t (muscimol) and a GABA a g o n i s t (baclofen) together p r o d u c e d antiB . c o n f l i c t e f f e c t s when neither c o m p o n e n t of thls treatment was e f f e c t i v e when given alone. At present, it w o u l d be d i f f i c u l t to accept or reject "~lequivotally either the route of a d m i n i s t r a t i o n or the receptor s u b - t y p e s e x p l a n a t i o n of the a p p a r e n t inactivity of some GABA receptor agonist in conflict tests. However, it should De noted that both a r g u m e n t s m i l i t a t e against the rejection of the G A B A h y p o t h e s i s of anxiety. There is at least one G A B A a g o n i s t which r e p r o d u c i b l y s i m u l a t e s the e f f e c t s of b e n z o d i a z e p i n e s in c o n f l i c t and r e l a t e d b e h a v i o r a l paradigms. Thus,
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v a l p r o a t e a t t e n u a t e s the s u p p r e s s i o n of o p e r a n t r e s p o n d i n g (25, 26) a n d of w a t e r d r i n k i n g in d e p r i v e d rats (16, 27, 28), induced by e l e c t r i c shock. Valproate also r e d u c e s neophobia, b e h a v i o r a l i n h i b i t i o n i n d u c e d by n o v e l t y (29, 30). Other b e h a v i o r a l e f f e c t s of b e n z o d i a z e p i n e s also seem to be shared by valproate; including a n t a g o n i s m of the s t i m u l u s p r o p e r t i e s of p e n t y l e n e t e t r a z ole (26) and e n h a n c e m e n t of saline d r i n k i n g in n o n - d e p r i v e d rats (Shephard and Johnston, unpublished) and both have w e l l - k n o w n m u s c l e relaxant and a n t i c o n v u isant p r o p e r t i e s . The a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e occur f o l l o w i n g systemic a d m i n i s t r a t i o n , and this drug seems most e f f e c t i v e in doses of 100-400 mg/kg. A l t h o u g h the o c c u r r e n c e of such e f f e c t s w o u l d seem u n c o n t r o v e r s i a l , their precise m e c h a n i s m is u n f o r t u n a t e l y less clear. A l t h o u g h v a l p r o a t e can inhibit G A B A t r a n s a m i n a s e (31) it p r o d u c e s no a p p r e c i a b l e e l e v a t i o n of forebrain GABA levels at a n t i - c o n f l i c t doses (16), w h e r e a s a g e n t s which p r o d u c e this b i o c h e m i c a l effect do not modify c o n f l i c t behavior, as s u r v e y e d earlier. The p o s s i b i l i t y that v a l p r o a t e may have direct a g o n i s t actions at the G A B A receptor is a more likely e x p l a n a t i o n of its a n t i - c o n f l i c t p r o p e r t i e s . That v a l p r o a t e d i s p l a c e s r e c e p t o r bound t r i t i a t e d d i h y d r o p i c r o t o x i n i n (32), w o u l d seem to be e v i d e n c e of such action at the receptor level, as w o u l d n e u r o p h y siological studies (33, 34). Moreover, two studies have shown a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e to be a n a t a g o n i s e d by p i c r o t o x i n (28, 30), the latter being c l e a r l y c o m p e t i t i v e antagonism, s u g g e s t i n g that the drugs are acting at the same site (12, 30). B i c u c u l l i n e has also been r e p o r t e d to a n t a g o n i s e such v a l p r o a t e a c t i o n s (16), though there is also a n e g a t i v e report (28). The int e r a c t i o n s of v a l p r o a t e with drugs a c t i n g at b e n z o d i a z e p i n e r e c e p t o r s are disc u s s e d in the following section. If the a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e are due to a g o n i s t a c t i o n s at G A B A receptors, then why is it e f f e c t i v e f o l l o w i n g systemic a d m i n i s t r a t i o n whilst the other c o m p o u n d s r e v i e w e d above are not? Clearly, its clinical efficacy in e p i l e p s y (31) together w i t h e x p e r i m e n t a l data leave no doubt that it p e n e t r a t e s the b l o o d - b r a i n b a r r i e r and it may be an a g o n i s t at both GABA~ and G A B A B receptors, though this does not appear to have been tested directl~. The e f f i c a c y of valproate, of i n t r a c e r e b r a l muscimol or G A B A and of combinations of m u s c i m o l and b a c l o f e n given s y s t e m i c a l l y in a t t e n u a t i n g c o n f l i c t behavior strongly support a role for G A B A in a n x i e t y systems. A l t h o u g h there are a p p a r e n t l y a n o m a l o u s findings, these can be a c c o u n t e d for and should not be a l l o w e d to o b s c u r e this conclusion. B e n z o d i a z e p i n e s can elicit b e h a v i o r a l e f f e c t s which are s e e m i n g l y unrelated to anxiety. Amongst these are s t i m u l a t i o n of c o n s u m p t i o n of p a l a t a b l e fluids and of water in d e p r i v e d rats (35). Moreover, food c o n s u m p t i o n may be e n h a n c e d by b e n z o d i a z e p i n e s (II, 35), even when no c o m p o n e n t of u n f a m i l i a r i t y or n e o p h o b i a is involved in the test procedure, though there is less evidence for this than seems to be g e n e r a l l y s u p p o s e d (12, 36). The unclear parallel between this last action of b e n z o d i a z e p i n e s and the e f f e c t s of G A B A e r g i c drugs on feeding has also been used to q u e s t i o n the G A B A h y p o t h e s i s of anxiolytic drug action (11). In general both d i r e c t l y (37, 38) and i n d i r e c t l y (38, 39, 40) acting GABA a g o n i s t s induce a n o r e c t i c e f f e c t s following systemic adm i n i s t r a t i o n an action which is a c t u a l l y o p p o s i t e to that r e p o r t e d for benzodiazepines. When G A B A a g o n i s t s and a n t a g o n i s t s are given i n t r a c e r e b r a l l y , their e f f e c t s on feeding d e p e n d upon the site of a d m i n i s t r a t i o n (40, 41, 42, 43), for example, G A B A w o u l d appear to be an inhibitor of feeding in the lateral h y p o t h a l a m u s (42, 43, 44, 45), a c o n c l u s i o n w h i c h is the reverse of that o b t a i n e d from studies of systemic a d m i n i s t r a t i o n s . However, when one c o n s i d e r s the w i d e s p r e a d i n h i b i t o r y role of G A B A in the brain, such r e s u l t s are hardly paradoxical. Indeed, they are no more s u r p r i s i n g than the w e l l - k n o w n fact that a p p l i c a t i o n of lesions or, on the other hand, e l e c t r i c a l s t i m u l a t i o n to d i f f e r e n t brain regions can elicit o p p o s i t e b e h a v i o r a l effects. Since it is clear from studies of b o t h the p h y l o g e n e t i c and n e u r o a n a t o m i c a l d i s t r i b u t i o n of
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benzodiazepine receptors that these only occur at some GABA synapses (12), these generally opposite effects of GABA agonists and benzodiazepines may simply be due to the latter drugs predominantly affecting C~kBA systems which stimulate feeding, for example by reducing the activity of inhibitory systems. Interactions
of GABA agonists
with dru9s acting at benzodiazepine
receptors
Both neurophysiological (2, 6, 7) and receptor binding (8, 9) studies suggest that the effects of GABA agonists and benzodiazepines should be at least additive, if not potentiative, of each other. In view of these findings, a number of investigations have assessed the extent to which this is the case for behavioral effects. One study of this question (21) using a number of anxiety paradigms failed to show enhancement of the anti-conflict actions of a marginally-effective dose of diazepam (0.5 mg/kg) with muscimol. However, since muscimol alone was inactive in this work, possibly for the reasons surveyed in the previous section, the significance of this is questionable. More paradoxical is the mutual antagonism evident between chlordiazepoxide and valproate on neophobia, even though both exert anti-neophobic actions when given alone (29). Neither are such apparent anomalies confined to anxiolytic effects; benzodiazepines have also been found to block muscimol-induced myoclonic jerks in mice (46). Whilst these studies do suggest some relationship between GABA and benzodizepines, they clearly do not conform with the above prediction of mutual synergism. However, the GABA transaminase inhibitor ethanolamine-Osulfate potentiates some effects of chlordiazepoxideon conflict behaviors (18, 19), although variously reported to be effective or ineffective in attenuating conflict when given alone. A related approach to linking GABA and benzodiazepine systems in behavioral experiments has been to examine the capacity of benzodiazepine antagonists to attenuate anti-conflict effects of GABA agonists. The actions of valproate on punished drinking (28) are antagonised by 5 mg/kg, though not 25 mg/kg of RO 15-1788. Moreover, RO 15-1788 (10 mq/kg) antagonises the effects of valproate on neophobia (30), apparently a non-competitive antagonism since it is not overcome by increasing the valproate dose and therefore consistent with there being GABA and benzodiazepine sites on a receptor complex. In contrast, RO 151788 does not seem very effective in attenuating actions of valproate on behaviors which are not anxiety-related (28), but rather RO 5-3663, which interacts with the picrotoxinin-sensitive site on the receptor complex, is a more effective antagonist. Interestingly, the G A B A r e c e p t o r antagonist bicuculline was not very effective in antagonising any b ~ a v i o r a l actions of valproate in one report (28), but blocks the effects of valproate on punished drinking in another (16). To conclude this section, although behavioral studies of GABA/benzodiazepine interactions recently lagged behind biochemical and neurophysiological ones to a considerable degree, there is now abundant behavioral evidence, including some from conflict studies, supporting the notion of a GABA/benzodiazepine receptor complex. However, the nature of the interactions is not simple and they certainly cannot be understood in terms of the comparatively early finding of enhanced benzodiazepine receptor binding with GABA agonists and inhibited binding with GABA antagonists. For even a tentative analysis of the mechanism of the behavioral pharmacological interactions surveyed here, we must consider the still-developing understanding of the receptor complex itself. The GABA/benzodiazepine receptor complex: Implications for behav!qral pharmacology The idea that GABA and benzodiazepine receptors are somehow coupled together has been with us almost since the discovery of the latter. To this basic
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proposition have from time to time been added hypotheses of multiple GABA receptors (eg 47), multiple benzodiazepine receptors (eg 48), endogenous ligands acting at benzodiazepine receptors (see 49 for review), a picrotoxinin/barbiturate/RO 5-3663 site (I), plus the idea that the entire complex might be coupled with a chloride channel protein (I, 3). It would be beyond the scope of this article to evaluate the evidence for or against these claims, except to state in passing that the peculiar efficacy of valproate in conflict procedures tempts one to add a valproate site to the already complex complex! However, since there seems to be no direct biochemical evidence for the latter, it would be more parsimonious to suppose valproate's effectiveness to be due to a multiple action at more than one previously-postulated site. Given the multiple structures involved in the GABA/benzodiazepine receptor complex and our incomplete understanding of the relationships between these structures, it is perhaps unsurprising that behavioral studies of interactions between GABA agonists and drugs acting at the benzodiazepine site have, as noted above, not generally confirmed the mutual synergism between GABA and benzodiazepine agonists which would be predicted from certain biochemical and neurophysiological work. Many of these apparently paradoxical findings have been obtained with valproate; indeed since most other GABA agonists fail to modify conflict behavior alone, it might be expected that they do not affect benzodiazepine action (21). Similarly, the greater effectiveness of a lower (5 mg/kg), as opposed to a higher (25 mg/kg) dose of RO 15-1788 in antagonising certain behavioral effects of valproate, including those on conflict behavior (28), can readily be accounted for in terms of intrinsic actions of RO 15-1788 at higher doses (28, 50). More problematically, the actions of valproate on neophobia (29, 30) and in eliciting "wet-dog shakes" (51, 52) can each be antagonised not only by RO 15-1788, but also by anxiolytic benzodiazepine agonists as well. Such findings, however, would be explained if we suppose that behavioral effects of valproate depend upon there being an optimal and sub-maximal level of stimulation at the benzodiazepine site, as has been proposed previously (30). Thus, RO 15-1788 in lower doses, reduces the activity at the benzodiazepine site (possibly occasioned by endogenous ligands) below the optimum and thus prevents valproate actions. Higher doses of RO 15-1788, because of intrinsic agonist actions (50, 53), lack such effects (28). Appropriate doses of anxiolytic benzodiazepine agonists, however, increase activity at the benzodiazepine site above the optimum for behavioral effects of valproate. It should be noted that what is proposed here is not merely a "ceiling effect" since combinations of anti-neophobic doses of valproate and chlordiazepoxide are significantly less effective than either given alone (29). A better analogy might be the effect of acetylcholine at the neuromuscular junction where either receptor blockade with tubocurarine or overstimulation with anticholinesterase drugs can induce paralysis. Clinical
implications
A significant part of the rationale for trying to discover the mechanism of action of therapeutic agents such as the benzodiazepines is the hope that understanding this will facilitate the development of new treatments which are more effective, lack some of the disadvantages of the old, or both. Regarding the possible use of GABA agonists clinically as anxiolytics, it has to be admitted that their efficacy in humans has yet to be established and that any possible advantage they might have over benzodiazepines is even more speculative. It is evident from the animal experimental studies reviewed here that valproate exerts apparent anxiolytic effects in such work. It would also appear to be of comparable efficacy, though lower potency, with benzodiazepines, unlike serotonergic agents which are generally less effective (12). Coincident-
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ally, v a l p r o a t e is also the only G A B N a g o n i s t r e v i e w e d here to be in wides p r e a d t h e r a p e u t i c use, not as an anxiolytic, but in e p i l e p t i f o r m d i s o r d e r s (31). It might, therefore, be e x p e c t e d that a n x i o l y t i c e f f e c t s of v a l p r o a t e in humans w o u l d have been w i d e l y d e t e c t e d as a sort of "side-effect" of its use as an a n t i c o n v u l s a n t . That this "side-effect" does not appear to have been reported, w o u l d seem to be a p o w e r f u l c r i t i c i s m of animal anxiety models, almost all of which have been shown to be v a l p r o a t e - s e n s i t i v e . However, a l t e r n a t i v e hypo t h e s e s can be advanced. In the first place, a benign and (at least for nona n x i o u s patients) subtle s i d e - e f f e c t such as a n x i o l y s i s might not be readily d e t e c t a b l e or thought worth r e p o r t i n g by either p a t i e n t or physician. Of course, some p a t i e n t s will be s u f f e r i n g b o t h severe anxiety and c o n v u l s i v e disorders; might this s u b - g r o u p not result in d e t e c t i o n of a n x i o l y t i c effects of valp r o a t e were they to be given this drug? The p r e d i c t i o n from animal work would seem to be not if they were also r e c e i v i n g b e n z o d i a z e p i n e s (for either conditions) since these drugs p r o d u c e mutual a n t a g o n i s m (29). Secondly, g e n e r a l i z i n g over a n u m b e r of studies and test p r o c e d u r e s , v a l p r o a t e exerts a n t i c o n v u i s a n t e f f e c t s at lower doses than those n e c e s s a r y to a t t e n u a t e e x p e r i m e n t a l conflict in rats. A s s u m i n g that this d i f f e r e n c e in p o t e n c y also holds true for humans, then it p e r h a p s u n s u r p r i s i n g that p e r s o n s r e c e i v i n g the lower dose of v a l p r o a t e a p p r o p r i a t e to c o n v u l s i v e disorders, do not e x p e r i e n c e striking anxiolysis. ~lly two e x p l i c i t l y - d e s i g n e d studies have i n v e s t i g a t e d the a c t i o n s of G A B N a g o n i s t s on human anxiety, one e x a m i n i n g THIP (54) and the other p r o g a b i d e (55). B o t h d e t e c t e d some a n x i o l y t i c actions, though, over the dose ranges used, these e f f e c t s were r e p o r t e d l y w e a k e r than those of the b e n z o d i a z e p i n e s . Moreover, ~ I P e l i c i t e d severe s i d e - e f f e c t s at a n x i o l y t i c doses. A l t h o u g h these s t u d i e s cannot be said to be s u p p o r t i v e of a p o t e n t i a l use of GABA a g o n i s t s in clinical anxiety, it shou]d be p o i n t e d out that n e i t h e r of these drugs have c o n s i s t e n t l y p r o v e d e f f e c t i v e in animal c o n f l i c t studies, as r e v i e w e d above, and t h e r e f o r e s e l e c t i n g them for i n v e s t i g a t i o n s in humans was questionable.
Conclusions The G A B A h y p o t h e s e s of a n x i e t y and of b e n z o d i a z e p i n e action have been w i t h us for some time and seem w e l l - s u p p o r t e d by b i o c h e m i c a l and n e u r o p h y s / o logical studies. B e h a v i o r a l evidence, and p a r t i c u l a r l y that r e l a t i n g to animal a n x i e t y models, has been more equivocal and has led some to p o s t u l a t e a dual m e c h a n i s m of action for b e n z o d i a z e p i n e s . Thus, it has been s u g g e s t e d that w h i l s t a n t i c o n v u l s a n t and sedative e f f e c t s of b e n z o d i a z e p i n e s might be m e d i a t e d through G A B A systems, a n x i o l y t i c a c t i o n s may not (11, 56, 57). Crucial to the p l a u s i b i l i t y of this h y p o t h e s i s is the e x i s t e n c e of m u l t i p l e b e n z o d i a z e p i n e r e c e p t o r types, since, if there were but one, it w o u l d seem highly unlikely that those c o n c e r n e d with sedative or a n t i c o n v u l s a n t a c t i o n s would just so happen to be a s s o c i a t e d with G A B A systems, w h e r e a s those c o n c e r n e d with a n x i o l y t i c a c t i o n s are not. The major e v i d e n c e for sub-types of b e n z o d i a z e p i n e receptor d e r i v e s from studies with CL 218872 for w h i c h a specific a n x i o ] y t i c action has been c l a i m e d (48), but also d i s p u t e d (58). Even if one regards the case for these s u b - t y p e s to be e s t a b l i s h e d , this does not e l i m i n a t e the p o s s i b i l i t y that b o t h s u b - t y p e s are linked with GABA systems. There have been a n u m b e r of animal studies of the c a p a c i t y of GABA agonists to simulate the a c t i o n s of b e n z o d i a z e p i n e s which have p r o v e d negative and the clinical e v i d e n c e is not c o m p e l l i n g as it stands. However, many of the drugs used may not s t i m u l a t e GABA systems in the most a p p r o p r i a t e fashion by only a f f e c t i n g some GABA r e c e p t o r s or by e l e v a t i n g whole brain G A B A w i t h o u t a f f e c t i n g the s y n a p t i c a l l y - a c t i v e component. Some may not reach the brain at all and n e g a t i v e findings are f a v o r e d by statistical procedures. In view of the several studies s h o w i n g a n t i c o n f l i c t e f f e c t s of valproate, of intracerebral G A B A a g o n i s t s and of muscimol/baclofen c o m b i n a t i o n s r e v i e w e d above, to discount
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the GABA h y p o t h e s i s of a n x i e t y w o u l d seem p r e m a t u r e in the extreme. Improved u n d e r s t a n d i n g of the p h y s i o l o g y and p h a r m a c o l o g y of the G A B A / b e n z o d i a z e p i n e r e c e p t o r c o m p l e x will e n a b l e us to make b e t t e r p r e d i c t i o n s of b e h a v i o r a l p h a r m a c o l o g i c a l i n t e r a c t i o n s b e t w e e n drugs a c t i n g at this c o m p l e x and to u n d e r s t a n d better the r e s u l t s a l r e a d y obtained. Clearly, there is rather little clinical e v i d e n c e at p r e s e n t and the design of such studies should be informed by animal investigations. References I.
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BEHAVIORAL
Pergamon
Journals
EFFECTS OF GABA AGONISTS IN RELATION TO ANXIETY AND BENZODIAZEPINE ACTION R A Shephard
Behavioral Analysis and Behavioral Biology Research Centre and Department of Psychology, University of Ulster at Jordanstown, N e ~ o w n a b b e y , BT37 OQB N. Ireland, UK. (Received
in final
form April
7, 1987)
Summary A considerable body of biochemical and neurophysiological evidence implicates GABA in anxiety and in benzodiazepine action. The present article surveys the behavioral effects of GABA agonists and their interactions with drugs acting at the benzodiazepine receptor in animal anxiety paradigms. Certain GABA agonists, notably valproate, simulate many behavioral actions of benzodiazepines. Moreover, several behavioral studies of the interaction of GABA agonists with benzodiazepines support the hypothesis of a benzodiazepine receptor complex with one or more GABA, benzodiazepine and probably other binding sites. However, there are also a number of anomalous findings of GABA agonist action alone and in combination with benzodiazepines. It is argued that these paradoxical results can better be accounted for in terms of the receptor complex and the distribution of the drugs, rather than by suggesting that the anxiolytic actions of benzodiazepines are not mediated by GABA systems. The potential clinical usefulness of GABA agonists in anxiety is commented upon. Introduction In both phylogenetic and neuroanatomical terms, the neurotransmitter GABA ranks as amongst the most widely distributed and it is currently estimated to account for about a third of all neurotransmission in the central nervous system (i). In view of this, it is unsurprising that drugs affecting GABA systems produce pronounced effects on the central nervous system, including behavioral actions. GABA agonists generally induce sedation and anticonvulsant effects; actions which are also characteristic of benzodiazepine tranquillizers. This parallel has, for some time, been used to support speculation that benzodiazepines might elicit their pharmacological effects, including their anxiolytic actions, by facilitating GAHAergic transmission. More recently, direct support for this hypothesis has been derived from neurophysiological and biochemical experiments (I, 2, 3, 4, 5, 6, 7). Possibly the most salient argument is the evidence for GABA involvement in benzodiazepine receptor binding, which is facilitated by a number of GABA agonists, but inhibited by its antagonists (8, 9). Since the anxiolytic effects of benzodiazepines are related to activation of benzodiazepine receptors (I0), it would be expected that GABA agonists should potentiate benzodiazepine action and/or simulate their behavioral effects, whilst GABA antagonists should induce essentially opposite effects. A number of studies, reviewed elsewhere (ii, 12) have shown GABA antagonists to elicit apparent anxiogenic actions or to attenuate benzodiazepine effects and, whilst some of these experiments are imperfectly designed (Ii, 12) and it is difficult to draw specific conclusions from in-
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supported.
The main p u r p o s e of this review, however, is to e x a m i n e the extent to w h i c h GABA a g o n i s t s have been shown to reproduce the e f f e c t s of benzodiazepine~ in animal a n x i e t y m o d e l s (or c o n f l i c t behaviors) or to p o t e n t i a t e such benzodia z e p i n e actions, as w o u l d be p r e d i c t e d from biochemical and n e u r o p h y s i o l o g i c a l evidence. The nature of these a n x i e t y m o d e l s has f r e q u e n t l y been r e v i e w e d e l s e w h e r e (eg 12, 13, 14, 15) and it should be noted that, logically speaking, these two q u e s t i o n s are i n d e p e n d e n t (12). Thus, in theory G A B A systems c o u l d have a role in the p h y s i o l o q y of anxiety which is entirely i n d e p e n d e n t of the s y s t e m s involved in b e n z o d i a z e p i n e action w h i c h might result in f u l f i l m e n t of the " s t i m u l a t i o n " but not " p o t e n t i a t i o n " prediction. C o n v e r s e l y , and again t h e o r e t i c a l l y , if the role of G A B A in a n x i e t y systems was c o n f i n e d to actions at the b e n z o d ] a z e p i n e receptor complex, then GABA a g o n i s t s might not elicit a n t i - c o n f l i c t e f f e c t s themselves, but could enhance the actions of substances a f f e c t i n g the b e n z o d i a z e p i n e site. Effects
of GABA a ~ o n i s t s
on conflict
behavior
A number of c o m p o u n d s which inhibit the a c t i v i t y of the enzyme GABA t r a n s a m i n a s e are available. Since this enzyme is important in the inactivation of G~BA, i n h i b i t i n 9 it p r o d u c e s i n c r e a s e s in brain GI~BA c o n c e n t r a t i o n s and G A B A t r a n s a m i n a s e inhibitors can be r e g a r d e d as i n d i r e c t l y - a c t i n g G A B A agonists. Drugs e x e r t i n g this action do not, however, g e n e r a l l y elicit benzodiaze p i n e - l i k e a n t i - c o n f l i c t effects in a p p r o p r i a t e behavioral tests, negative findings being r e p o r t e d for a m i n o - o x y a c e t i c acid (16, 1 7 ) , ~ -vinyl GABA (16) and e t h a n o l a m i n e - O - s u l f a t e (18). The last has been c o n t r a d i c t e d by s u b s e q u e n t i n v e s t i g a t i o n s (19), but the balance of e v i d e n c e at p r e s e n t s u g g e s t s that GABA t r a n s a m i n a s e i n h i b i t o r s have no a n t i - c o n f l i c t actions. It may be, of course, that the m o d e s t increases in total brain G A B A c o n c e n t r a t i o n induced by this type of dru 9 simply fail to result in a p p r e c i a b l e increases in functional GABA a c t i v i t y at the synapses relevant to c o n f l i c t or anxiety systems. Rather more c o m p l e x and c o n t r o v e r s J a ! are the e f f e c t s of drugs thought to have direct a g o n i s t action at GABA r e c e p t o r s on c o n f l i c t behavior. Again, lack of b e h a v i o r a l a c t i v i t y has been r e p o r t e d for a nun~0er of agents; namely m u s c i m o l (20, 21), p r o g a b i d e (20) and THIP (16). Apart from the p o s s i b i l i t y that C~BA may not be involved in the p h y s i o l o g y of anxiety (discussed later), two e x p l a n a t i o n s of this lack of e f f e c t have been advanced. The first was simply that these c o m p o u n d s do not r e a d i l y enter the brain following the systemic a d m i n i s t r a t i o n s used and t h e r e f o r e that no a n t i - c o n f l i c t e f f e c t s could be expected. This is supported by d e m o n s t r a t i o n s of a n t i - c o n f l i c t actions of m u s c i m o l (3) and of GABA itself (22, 23), when given intracerebrally. Against this, it has been r e p o r t e d that muscimol a f f e c t s n e u r o p h y s i o l o g i c a l p a r a m e t e r s followin 9 systemic a d m i n i s t r a t i o n s of doses equal to, or even o r d e r s of magnitude below, those which fail to elicit a n t i - c o n f l i c t a c t i o n s (24), and suggested that the a p p a r e n t lack of a c t i v i t y of many GABA a g o n i s t s may be because they affect only the GABA A receptor (24). In support of this hypothesis, it was shown that c o m b i n e d systemic a d m i n i s t r a t i o n of a p p r o p r i a t e doses of G A B A A a g o n i s t (muscimol) and a GABA a g o n i s t (baclofen) together p r o d u c e d antiB . c o n f l i c t e f f e c t s when neither c o m p o n e n t of thls treatment was e f f e c t i v e when given alone. At present, it w o u l d be d i f f i c u l t to accept or reject "~lequivotally either the route of a d m i n i s t r a t i o n or the receptor s u b - t y p e s e x p l a n a t i o n of the a p p a r e n t inactivity of some GABA receptor agonist in conflict tests. However, it should De noted that both a r g u m e n t s m i l i t a t e against the rejection of the G A B A h y p o t h e s i s of anxiety. There is at least one G A B A a g o n i s t which r e p r o d u c i b l y s i m u l a t e s the e f f e c t s of b e n z o d i a z e p i n e s in c o n f l i c t and r e l a t e d b e h a v i o r a l paradigms. Thus,
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v a l p r o a t e a t t e n u a t e s the s u p p r e s s i o n of o p e r a n t r e s p o n d i n g (25, 26) a n d of w a t e r d r i n k i n g in d e p r i v e d rats (16, 27, 28), induced by e l e c t r i c shock. Valproate also r e d u c e s neophobia, b e h a v i o r a l i n h i b i t i o n i n d u c e d by n o v e l t y (29, 30). Other b e h a v i o r a l e f f e c t s of b e n z o d i a z e p i n e s also seem to be shared by valproate; including a n t a g o n i s m of the s t i m u l u s p r o p e r t i e s of p e n t y l e n e t e t r a z ole (26) and e n h a n c e m e n t of saline d r i n k i n g in n o n - d e p r i v e d rats (Shephard and Johnston, unpublished) and both have w e l l - k n o w n m u s c l e relaxant and a n t i c o n v u isant p r o p e r t i e s . The a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e occur f o l l o w i n g systemic a d m i n i s t r a t i o n , and this drug seems most e f f e c t i v e in doses of 100-400 mg/kg. A l t h o u g h the o c c u r r e n c e of such e f f e c t s w o u l d seem u n c o n t r o v e r s i a l , their precise m e c h a n i s m is u n f o r t u n a t e l y less clear. A l t h o u g h v a l p r o a t e can inhibit G A B A t r a n s a m i n a s e (31) it p r o d u c e s no a p p r e c i a b l e e l e v a t i o n of forebrain GABA levels at a n t i - c o n f l i c t doses (16), w h e r e a s a g e n t s which p r o d u c e this b i o c h e m i c a l effect do not modify c o n f l i c t behavior, as s u r v e y e d earlier. The p o s s i b i l i t y that v a l p r o a t e may have direct a g o n i s t actions at the G A B A receptor is a more likely e x p l a n a t i o n of its a n t i - c o n f l i c t p r o p e r t i e s . That v a l p r o a t e d i s p l a c e s r e c e p t o r bound t r i t i a t e d d i h y d r o p i c r o t o x i n i n (32), w o u l d seem to be e v i d e n c e of such action at the receptor level, as w o u l d n e u r o p h y siological studies (33, 34). Moreover, two studies have shown a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e to be a n a t a g o n i s e d by p i c r o t o x i n (28, 30), the latter being c l e a r l y c o m p e t i t i v e antagonism, s u g g e s t i n g that the drugs are acting at the same site (12, 30). B i c u c u l l i n e has also been r e p o r t e d to a n t a g o n i s e such v a l p r o a t e a c t i o n s (16), though there is also a n e g a t i v e report (28). The int e r a c t i o n s of v a l p r o a t e with drugs a c t i n g at b e n z o d i a z e p i n e r e c e p t o r s are disc u s s e d in the following section. If the a n t i - c o n f l i c t e f f e c t s of v a l p r o a t e are due to a g o n i s t a c t i o n s at G A B A receptors, then why is it e f f e c t i v e f o l l o w i n g systemic a d m i n i s t r a t i o n whilst the other c o m p o u n d s r e v i e w e d above are not? Clearly, its clinical efficacy in e p i l e p s y (31) together w i t h e x p e r i m e n t a l data leave no doubt that it p e n e t r a t e s the b l o o d - b r a i n b a r r i e r and it may be an a g o n i s t at both GABA~ and G A B A B receptors, though this does not appear to have been tested directl~. The e f f i c a c y of valproate, of i n t r a c e r e b r a l muscimol or G A B A and of combinations of m u s c i m o l and b a c l o f e n given s y s t e m i c a l l y in a t t e n u a t i n g c o n f l i c t behavior strongly support a role for G A B A in a n x i e t y systems. A l t h o u g h there are a p p a r e n t l y a n o m a l o u s findings, these can be a c c o u n t e d for and should not be a l l o w e d to o b s c u r e this conclusion. B e n z o d i a z e p i n e s can elicit b e h a v i o r a l e f f e c t s which are s e e m i n g l y unrelated to anxiety. Amongst these are s t i m u l a t i o n of c o n s u m p t i o n of p a l a t a b l e fluids and of water in d e p r i v e d rats (35). Moreover, food c o n s u m p t i o n may be e n h a n c e d by b e n z o d i a z e p i n e s (II, 35), even when no c o m p o n e n t of u n f a m i l i a r i t y or n e o p h o b i a is involved in the test procedure, though there is less evidence for this than seems to be g e n e r a l l y s u p p o s e d (12, 36). The unclear parallel between this last action of b e n z o d i a z e p i n e s and the e f f e c t s of G A B A e r g i c drugs on feeding has also been used to q u e s t i o n the G A B A h y p o t h e s i s of anxiolytic drug action (11). In general both d i r e c t l y (37, 38) and i n d i r e c t l y (38, 39, 40) acting GABA a g o n i s t s induce a n o r e c t i c e f f e c t s following systemic adm i n i s t r a t i o n an action which is a c t u a l l y o p p o s i t e to that r e p o r t e d for benzodiazepines. When G A B A a g o n i s t s and a n t a g o n i s t s are given i n t r a c e r e b r a l l y , their e f f e c t s on feeding d e p e n d upon the site of a d m i n i s t r a t i o n (40, 41, 42, 43), for example, G A B A w o u l d appear to be an inhibitor of feeding in the lateral h y p o t h a l a m u s (42, 43, 44, 45), a c o n c l u s i o n w h i c h is the reverse of that o b t a i n e d from studies of systemic a d m i n i s t r a t i o n s . However, when one c o n s i d e r s the w i d e s p r e a d i n h i b i t o r y role of G A B A in the brain, such r e s u l t s are hardly paradoxical. Indeed, they are no more s u r p r i s i n g than the w e l l - k n o w n fact that a p p l i c a t i o n of lesions or, on the other hand, e l e c t r i c a l s t i m u l a t i o n to d i f f e r e n t brain regions can elicit o p p o s i t e b e h a v i o r a l effects. Since it is clear from studies of b o t h the p h y l o g e n e t i c and n e u r o a n a t o m i c a l d i s t r i b u t i o n of
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benzodiazepine receptors that these only occur at some GABA synapses (12), these generally opposite effects of GABA agonists and benzodiazepines may simply be due to the latter drugs predominantly affecting C~kBA systems which stimulate feeding, for example by reducing the activity of inhibitory systems. Interactions
of GABA agonists
with dru9s acting at benzodiazepine
receptors
Both neurophysiological (2, 6, 7) and receptor binding (8, 9) studies suggest that the effects of GABA agonists and benzodiazepines should be at least additive, if not potentiative, of each other. In view of these findings, a number of investigations have assessed the extent to which this is the case for behavioral effects. One study of this question (21) using a number of anxiety paradigms failed to show enhancement of the anti-conflict actions of a marginally-effective dose of diazepam (0.5 mg/kg) with muscimol. However, since muscimol alone was inactive in this work, possibly for the reasons surveyed in the previous section, the significance of this is questionable. More paradoxical is the mutual antagonism evident between chlordiazepoxide and valproate on neophobia, even though both exert anti-neophobic actions when given alone (29). Neither are such apparent anomalies confined to anxiolytic effects; benzodiazepines have also been found to block muscimol-induced myoclonic jerks in mice (46). Whilst these studies do suggest some relationship between GABA and benzodizepines, they clearly do not conform with the above prediction of mutual synergism. However, the GABA transaminase inhibitor ethanolamine-Osulfate potentiates some effects of chlordiazepoxideon conflict behaviors (18, 19), although variously reported to be effective or ineffective in attenuating conflict when given alone. A related approach to linking GABA and benzodiazepine systems in behavioral experiments has been to examine the capacity of benzodiazepine antagonists to attenuate anti-conflict effects of GABA agonists. The actions of valproate on punished drinking (28) are antagonised by 5 mg/kg, though not 25 mg/kg of RO 15-1788. Moreover, RO 15-1788 (10 mq/kg) antagonises the effects of valproate on neophobia (30), apparently a non-competitive antagonism since it is not overcome by increasing the valproate dose and therefore consistent with there being GABA and benzodiazepine sites on a receptor complex. In contrast, RO 151788 does not seem very effective in attenuating actions of valproate on behaviors which are not anxiety-related (28), but rather RO 5-3663, which interacts with the picrotoxinin-sensitive site on the receptor complex, is a more effective antagonist. Interestingly, the G A B A r e c e p t o r antagonist bicuculline was not very effective in antagonising any b ~ a v i o r a l actions of valproate in one report (28), but blocks the effects of valproate on punished drinking in another (16). To conclude this section, although behavioral studies of GABA/benzodiazepine interactions recently lagged behind biochemical and neurophysiological ones to a considerable degree, there is now abundant behavioral evidence, including some from conflict studies, supporting the notion of a GABA/benzodiazepine receptor complex. However, the nature of the interactions is not simple and they certainly cannot be understood in terms of the comparatively early finding of enhanced benzodiazepine receptor binding with GABA agonists and inhibited binding with GABA antagonists. For even a tentative analysis of the mechanism of the behavioral pharmacological interactions surveyed here, we must consider the still-developing understanding of the receptor complex itself. The GABA/benzodiazepine receptor complex: Implications for behav!qral pharmacology The idea that GABA and benzodiazepine receptors are somehow coupled together has been with us almost since the discovery of the latter. To this basic
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proposition have from time to time been added hypotheses of multiple GABA receptors (eg 47), multiple benzodiazepine receptors (eg 48), endogenous ligands acting at benzodiazepine receptors (see 49 for review), a picrotoxinin/barbiturate/RO 5-3663 site (I), plus the idea that the entire complex might be coupled with a chloride channel protein (I, 3). It would be beyond the scope of this article to evaluate the evidence for or against these claims, except to state in passing that the peculiar efficacy of valproate in conflict procedures tempts one to add a valproate site to the already complex complex! However, since there seems to be no direct biochemical evidence for the latter, it would be more parsimonious to suppose valproate's effectiveness to be due to a multiple action at more than one previously-postulated site. Given the multiple structures involved in the GABA/benzodiazepine receptor complex and our incomplete understanding of the relationships between these structures, it is perhaps unsurprising that behavioral studies of interactions between GABA agonists and drugs acting at the benzodiazepine site have, as noted above, not generally confirmed the mutual synergism between GABA and benzodiazepine agonists which would be predicted from certain biochemical and neurophysiological work. Many of these apparently paradoxical findings have been obtained with valproate; indeed since most other GABA agonists fail to modify conflict behavior alone, it might be expected that they do not affect benzodiazepine action (21). Similarly, the greater effectiveness of a lower (5 mg/kg), as opposed to a higher (25 mg/kg) dose of RO 15-1788 in antagonising certain behavioral effects of valproate, including those on conflict behavior (28), can readily be accounted for in terms of intrinsic actions of RO 15-1788 at higher doses (28, 50). More problematically, the actions of valproate on neophobia (29, 30) and in eliciting "wet-dog shakes" (51, 52) can each be antagonised not only by RO 15-1788, but also by anxiolytic benzodiazepine agonists as well. Such findings, however, would be explained if we suppose that behavioral effects of valproate depend upon there being an optimal and sub-maximal level of stimulation at the benzodiazepine site, as has been proposed previously (30). Thus, RO 15-1788 in lower doses, reduces the activity at the benzodiazepine site (possibly occasioned by endogenous ligands) below the optimum and thus prevents valproate actions. Higher doses of RO 15-1788, because of intrinsic agonist actions (50, 53), lack such effects (28). Appropriate doses of anxiolytic benzodiazepine agonists, however, increase activity at the benzodiazepine site above the optimum for behavioral effects of valproate. It should be noted that what is proposed here is not merely a "ceiling effect" since combinations of anti-neophobic doses of valproate and chlordiazepoxide are significantly less effective than either given alone (29). A better analogy might be the effect of acetylcholine at the neuromuscular junction where either receptor blockade with tubocurarine or overstimulation with anticholinesterase drugs can induce paralysis. Clinical
implications
A significant part of the rationale for trying to discover the mechanism of action of therapeutic agents such as the benzodiazepines is the hope that understanding this will facilitate the development of new treatments which are more effective, lack some of the disadvantages of the old, or both. Regarding the possible use of GABA agonists clinically as anxiolytics, it has to be admitted that their efficacy in humans has yet to be established and that any possible advantage they might have over benzodiazepines is even more speculative. It is evident from the animal experimental studies reviewed here that valproate exerts apparent anxiolytic effects in such work. It would also appear to be of comparable efficacy, though lower potency, with benzodiazepines, unlike serotonergic agents which are generally less effective (12). Coincident-
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ally, v a l p r o a t e is also the only G A B N a g o n i s t r e v i e w e d here to be in wides p r e a d t h e r a p e u t i c use, not as an anxiolytic, but in e p i l e p t i f o r m d i s o r d e r s (31). It might, therefore, be e x p e c t e d that a n x i o l y t i c e f f e c t s of v a l p r o a t e in humans w o u l d have been w i d e l y d e t e c t e d as a sort of "side-effect" of its use as an a n t i c o n v u l s a n t . That this "side-effect" does not appear to have been reported, w o u l d seem to be a p o w e r f u l c r i t i c i s m of animal anxiety models, almost all of which have been shown to be v a l p r o a t e - s e n s i t i v e . However, a l t e r n a t i v e hypo t h e s e s can be advanced. In the first place, a benign and (at least for nona n x i o u s patients) subtle s i d e - e f f e c t such as a n x i o l y s i s might not be readily d e t e c t a b l e or thought worth r e p o r t i n g by either p a t i e n t or physician. Of course, some p a t i e n t s will be s u f f e r i n g b o t h severe anxiety and c o n v u l s i v e disorders; might this s u b - g r o u p not result in d e t e c t i o n of a n x i o l y t i c effects of valp r o a t e were they to be given this drug? The p r e d i c t i o n from animal work would seem to be not if they were also r e c e i v i n g b e n z o d i a z e p i n e s (for either conditions) since these drugs p r o d u c e mutual a n t a g o n i s m (29). Secondly, g e n e r a l i z i n g over a n u m b e r of studies and test p r o c e d u r e s , v a l p r o a t e exerts a n t i c o n v u i s a n t e f f e c t s at lower doses than those n e c e s s a r y to a t t e n u a t e e x p e r i m e n t a l conflict in rats. A s s u m i n g that this d i f f e r e n c e in p o t e n c y also holds true for humans, then it p e r h a p s u n s u r p r i s i n g that p e r s o n s r e c e i v i n g the lower dose of v a l p r o a t e a p p r o p r i a t e to c o n v u l s i v e disorders, do not e x p e r i e n c e striking anxiolysis. ~lly two e x p l i c i t l y - d e s i g n e d studies have i n v e s t i g a t e d the a c t i o n s of G A B N a g o n i s t s on human anxiety, one e x a m i n i n g THIP (54) and the other p r o g a b i d e (55). B o t h d e t e c t e d some a n x i o l y t i c actions, though, over the dose ranges used, these e f f e c t s were r e p o r t e d l y w e a k e r than those of the b e n z o d i a z e p i n e s . Moreover, ~ I P e l i c i t e d severe s i d e - e f f e c t s at a n x i o l y t i c doses. A l t h o u g h these s t u d i e s cannot be said to be s u p p o r t i v e of a p o t e n t i a l use of GABA a g o n i s t s in clinical anxiety, it shou]d be p o i n t e d out that n e i t h e r of these drugs have c o n s i s t e n t l y p r o v e d e f f e c t i v e in animal c o n f l i c t studies, as r e v i e w e d above, and t h e r e f o r e s e l e c t i n g them for i n v e s t i g a t i o n s in humans was questionable.
Conclusions The G A B A h y p o t h e s e s of a n x i e t y and of b e n z o d i a z e p i n e action have been w i t h us for some time and seem w e l l - s u p p o r t e d by b i o c h e m i c a l and n e u r o p h y s / o logical studies. B e h a v i o r a l evidence, and p a r t i c u l a r l y that r e l a t i n g to animal a n x i e t y models, has been more equivocal and has led some to p o s t u l a t e a dual m e c h a n i s m of action for b e n z o d i a z e p i n e s . Thus, it has been s u g g e s t e d that w h i l s t a n t i c o n v u l s a n t and sedative e f f e c t s of b e n z o d i a z e p i n e s might be m e d i a t e d through G A B A systems, a n x i o l y t i c a c t i o n s may not (11, 56, 57). Crucial to the p l a u s i b i l i t y of this h y p o t h e s i s is the e x i s t e n c e of m u l t i p l e b e n z o d i a z e p i n e r e c e p t o r types, since, if there were but one, it w o u l d seem highly unlikely that those c o n c e r n e d with sedative or a n t i c o n v u l s a n t a c t i o n s would just so happen to be a s s o c i a t e d with G A B A systems, w h e r e a s those c o n c e r n e d with a n x i o l y t i c a c t i o n s are not. The major e v i d e n c e for sub-types of b e n z o d i a z e p i n e receptor d e r i v e s from studies with CL 218872 for w h i c h a specific a n x i o ] y t i c action has been c l a i m e d (48), but also d i s p u t e d (58). Even if one regards the case for these s u b - t y p e s to be e s t a b l i s h e d , this does not e l i m i n a t e the p o s s i b i l i t y that b o t h s u b - t y p e s are linked with GABA systems. There have been a n u m b e r of animal studies of the c a p a c i t y of GABA agonists to simulate the a c t i o n s of b e n z o d i a z e p i n e s which have p r o v e d negative and the clinical e v i d e n c e is not c o m p e l l i n g as it stands. However, many of the drugs used may not s t i m u l a t e GABA systems in the most a p p r o p r i a t e fashion by only a f f e c t i n g some GABA r e c e p t o r s or by e l e v a t i n g whole brain G A B A w i t h o u t a f f e c t i n g the s y n a p t i c a l l y - a c t i v e component. Some may not reach the brain at all and n e g a t i v e findings are f a v o r e d by statistical procedures. In view of the several studies s h o w i n g a n t i c o n f l i c t e f f e c t s of valproate, of intracerebral G A B A a g o n i s t s and of muscimol/baclofen c o m b i n a t i o n s r e v i e w e d above, to discount
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the GABA h y p o t h e s i s of a n x i e t y w o u l d seem p r e m a t u r e in the extreme. Improved u n d e r s t a n d i n g of the p h y s i o l o g y and p h a r m a c o l o g y of the G A B A / b e n z o d i a z e p i n e r e c e p t o r c o m p l e x will e n a b l e us to make b e t t e r p r e d i c t i o n s of b e h a v i o r a l p h a r m a c o l o g i c a l i n t e r a c t i o n s b e t w e e n drugs a c t i n g at this c o m p l e x and to u n d e r s t a n d better the r e s u l t s a l r e a d y obtained. Clearly, there is rather little clinical e v i d e n c e at p r e s e n t and the design of such studies should be informed by animal investigations. References I.
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25. 26. 27. 28 29. 30. 31. 32. 33. 34.
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