Drug discrimination models in anxiety and depression

Pharmac. Ther. Vol. 47, pp. 267-280, 1990 Printed in Great Britain. All rights reserved

0163-7258/90$0.00+ 0.50 © 1990PergamonPress plc

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D R U G D I S C R I M I N A T I O N MODELS IN A N X I E T Y A N D DEPRESSION J. S. ANDREWS and D. N. STEPHENS Department of Neuropsychopharmacology, Schering AG, Mfillerstrafle 1 70-178, D-1000 Berlin 65, F.R.G. Abstract--Drug discrimination is a technique for investigating the stimulus properties of centrally active drugs. Although many studies have employed animals to investigate the stimulus properties of substances used clinically for the treatment of anxiety and depression, it would be a mistake to consider the internal discriminative stimuli as being related specifically to the anxiolytic or antidepressant properties of these drugs. Rather drug cues are better considered as relating to the pharmacological action of classes of compounds. Thus, benzodiazepine cues generalize to other compounds acting at benzodiazepine receptors, but not to substances (anxiolytic or otherwise) acting at 5-HTtA receptors. Similarly, antidepressants with different pharmacological properties, for example the tricyclic imipramine, or the phenylaminoketone buproprion produce distinct, unrelated discriminative stimuli. For this reason, the limits of drug discrimination techniques for investigating novel anxiolytic or antidepressant drugs should be clearly recognized. Attempts to identify an anxiogenic discriminative stimulus using pentylenetetrazole have also been misguided. In this technique it has proven difficult to separate unequivocally the pharmacological proconvulsant effects of the drug from the psychological construct anxiety. Nevertheless, drug discrimination remains a valuable technique for investigating pharmacological interactions in animals and man.

CONTENTS I. Introduction 2. Anxiolytic Cues 2. I. Benzodiazepine cues 2.2. Nonbenzodiazepine benzodiazepine receptor ligands 2.3. Serotonin cues 2.4. Excitatory amino acids 3. Anxiogenic cues 3.1. Pentylenetetrazole cue 3.2. FG 7142 cue 4. Antidepressant cues 5. Summary and Conclusions References

1. I N T R O D U C T I O N The ability of animals to identify distinct interoceptive states has been inferred from state dependent learning for many years. Drug discrimination paradigms make use of this ability to produce a stable behavioral assay which can be used to investigate similarities and differences within and between classes of psychoactive compounds. Many species have been trained to identify, and respond appropriately to the characteristic interoceptive effects of different drugs. Although a wide range of methods have been used, two procedures with minor variations dominate the field. In the first animals (usually rats) are trained to obtain food (or avoid shock) from one arm or the other of a T-maze depending on which drug has been administered. In the other method, animals are trained to press one lever in a standard operant chamber (again to receive food or avoid shock) in the presence of one drug

267 268 268 269 271 272 273 273 275 276 276 277

and the other lever when administered a second drug or saline. The training drug is alternated with the placebo (or second training drug) in a random m a n n e r until the animal can, on any test or training day, consistently make the appropriate response. Once animals are trained they can be used repeatedly as a behavioral assay for drug effects. Two basic types of experiment are possible: generalization to the stimulus and antagonism of the stimulus. In the first method a test drug is injected in place of the training drug and a dose-response curve generated as to how, and at what dose, the substance substitutes completely for the training drug. In the second procedure, antagonism, the test drug is administered shortly before or after the training drug and a dose-response curve constructed for reduction of drug appropriate responding. A n as yet unresolved question is whether the animals recognize intermediate doses of the training or test drug in a quantal manner. It has been suggested that recognition of the drug is in an 267

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all-or-none fashion, although different animals may be more sensitive to a particular drug dose than others thereby giving rise to a group dose-response curve. On the other hand, others believe that the discriminative stimulus is graded in response to the drug dose. For this reason, some researchers use individual trial-based procedures, or individual percent responding on the appropriate lever (graded effects), and others a simple first lever chosen principle (quantal effect). In practice both procedures appear to function equally well, and therefore, in this paper all drug discrimination procedures will be treated equally, the important point being that animals have learned a particular interoceptive stimulus against which other drugs can be compared. The interoceptive stimulus produced by this technique can be highly specific: animals trained in a two lever discrimination do not learn a simple drug vs no-drug condition. Rather, they learn a 'this-drug'/ 'not-this-drug' discrimination, or more correctly 'this-class-of-compounds' vs 'all-other-substances'. That is an animal trained to distinguish between diazepam and saline will respond on the drug lever only when a similar benzodiazepine (BZ)-like drug is injected and not when substances such as amphetamine, haloperidol or morphine are administered. Multiple drug discriminations are possible (e.g. White and Holtzman, 1981; France and Woods, 1987; Overton and Shen, 1988), even in man (Chait and Johanson, 1988; Preston et al., 1987). Indeed, it has been possible to train animals to distinguish compounds acting at different subtypes of receptor within the same compound class. For example, animals can learn to distinguish between drugs acting at subtypes of opiate receptors (Holtzrnan, 1985). This ability of animals to identify specifically the interoceptive stimulus properties of a substance has led to an increasing interest in defining the limits and uses of the technique. Several postulates have been proposed. If animals are trained to identify a particular drug from saline, then all substances which substitute for the training drug should have similar properties (although not necessarily the identical mechanism of action). It should be noted that saline appropriate responding following a test drug does not mean that the test drug is inactive, only that the interoceptive properties are not the same as the training drug. The conclusions from this can thus be simply stated using one or two examples: all substances thus far identified as heroin-like are potential drugs of abuse; all substances identified as diazepamlike are anxiolytics. The following sections of this review will investigate the usefulness of such statements. This article will attempt to review the important aspects of drug discrimination in psychiatric research. Accordingly, an extensive review of every possible substance used to form a discriminative stimulus will be avoided; instead we will concentrate on the salient aspects of a few representatives from, in particular, anxiolytic research.

2. ANXIOLYTIC CUES The term anxiolytic cue is generally taken to be synonymous with a benzodiazepine agonist cue of

one form or another. However, in recent years other substances have been proposed as, and/or introduced into the market as novel anxiolytics. The most notable of these is buspirone, a partial agonist at the 5-hydroxytryptamine IA (5-HT1A) receptor. As there are also several good reasons to suspect the involvement of 5-HT in anxiolysis, data involving such cues will also be reviewed. 2.1. BENZODIAZEPINECUES Research into the discriminative stimulus properties of BZs was pioneered by Coipaert (Colpaert et aL, 1976; Colpaert, 1977) who demonstrated that rats could be trained in a two lever task to discriminate an oral dose of 5 mg/kg of chlordiazepoxide (CDP) from saline. These studies also suggested that the cue was not exclusively attributable to the sedative or ataxic properties of CDP. Subsequently, the stimulus properties of CDP have been investigated in several animal species, with different routes of administration, but in general have all reached similar conclusions. Further research has shown that all agonistic benzodiazepines can substitute for CDP in several different procedures (e.g. Colpaert, 1977; Stephens et aL, 1987; Sanger, 1988). Accordingly, it has been demonstrated that CDP will itself substitute for other benzodiazepines such as diazepam (Stepbens et al., 1984a; Shannon and Herling, 1983; Young and Glennon, 1987). Other investigators have used not only CDP and diazepam as training drugs, but also many other BZs such as lorazepam (Ator and Griffiths, 1986), oxazepam (Hendry et al., 1983; de la Garza et al., 1987) and midazolam (Gareha et al., 1985; Rauch and Stolerman, 1987). Indeed it is now clear that BZs substitute readily for one another; that is, BZ-trained animals recognize other active BZs as being similar, irrespective of which BZ is used as the training drug. The stimulus properties of the cue are reported as stable in that there are no marked changes in generalization curves over time, or between young and old rats (Amrick and Bennett, 1987a). The discriminative stimulus is specific in that only BZs (or compounds acting at the B Z - G A B A receptor complex, such as barbiturates, e.g. Ator and Griffiths (1983)) generalize; substances from other drug classes, for example stimulants such as amphetamine, do not substitute for the training drug (Colpaert, 1977; Sanger, 1988). Although the generalization between BZs and barbiturates has sometimes been interpreted as meaning the stimulus properties of these behaviorally depressant compounds are identical, there are important differences. BZ cues can be antagonized in a dose-dependent manner by BZ receptor antagonists such as RO 15-1788, Z K 93 426 or CGS 8216 (Herling and Shannon, 1982; Stephens et aL, 1984b; Bennett et al., 1987; Young et al., 1987), whereas barbiturate cues are typically unaffected (Young et al., 1987). In addition, rats can be trained to distinguish BZ and barbiturates in drug-drug discriminations (e.g. Henteleff and Barry, 1989). BZ cues may also show a stereospecific separation of agonistic stimulus properties (J/irbe et al., 1988). Potencies for inducing diazepam-like effects in rats and potencies for inhibiting the binding of

Drug discrimination models radiolabeled diazepam to rat cortex membranes are positively correlated (Shannon and Herling, 1983). Young and Glennon have therefore suggested that a relationship may exist between relative potencies in a BZ drug discrimination procedure and therapeutic potency or a substance's BZ receptor affinity in humans. This suggestion has found some experimental support (Young and Glennon, 1987). As an animal model for identifying compounds with anxiolytic properties it appears then, that the BZ cue has some credibility. However, in recent years it has become apparent for several reasons that drug discrimination cannot be used to identify anxiolytic drugs in any but the most limited situations. The nature of the BZ cue is as yet unclear, but, it would be naive to suggest that it was an anxiolytic cue. Given the broad range of effects of BZs, the discriminative stimulus produced is likely to be extremely complex, and as yet it is unknown which properties of the BZs dominate, if any, in forming a discriminative stimulus. For instance, a correlation exists between the anti° convulsant properties of BZs and their effectiveness as anxiolytics in the clinic (Clody et al., 1983); following Young and Glennon, a positive correlation is therefore to be expected between anticonvulsant efficacy and generalization potencies in BZ cues. Furthermore, convulsant agents such as pentylenetetrazole will antagonize BZ discriminative properties and although the interpretation is extremely unlikely, the evidence is equally persuasive that the BZ cue is in fact an anticonvulsant cue (see also Colpaert et aL, 1976). Despite the obvious complexity of the discriminative stimulus, the cue also suffers paradoxically from being too specific. Using such an approach only BZ receptor ligands with similar properties (apart perhaps from potency) to those already existing would be (and have been) identified and the

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ability to identify novel anxiolytic compounds is lost. Despite this criticism benzodiazepine cues do have an important place in investigating the properties of the BZ-receptor complex and in the development of new anxiolytic drugs. Over the last 12 years the BZreceptor complex has been characterized, subtypes discovered and ligands with unexpected properties identified.

2.2. NONBENZODIAZEPINEBENZODIAZEPINE RECEPTORLIGANDS The discovery of high affinity central binding sites for benzodiazepines (Squires and Braestrup, 1977; Mohler and Okada, 1977) prompted a search for the natural receptor ligand. The //-carboline, //-carboline-3-carboxylic acid-ethyl ester (/~-CCE) was initially thought to be one candidate (Braestrup et al., 1980). Subsequently it was shown that this substance was an artifact of the extraction procedure. Nevertheless, its discovery led to a new appreciation of the complexity of the BZ-receptor complex (see Fig. 1). //-CCE binds specifically to the BZ receptor and is a potent inhibitor of tritiated diazepam or flunitrazepam binding. However, p-CCE was the first of a new class of compounds known as inverse agonists, that is, substances which produce an action opposite to those of the presumed agonist, in this case a benzodiazepine. Depending on their properties, three types of BZ ligand have been identified--agonists, neutral antagonists and inverse agonists. Moreover, in addition to these three main classes, subclasses of partial agonists and inverse agonists have been synthesized (e.g. Jensen et al., 1984; Petersen et al., 1984; Stephens et al., 1987) and tested in the clinic (Dorow et al., 1987). Subsequently derivatives of other BZ

Model of the GABA receptor (GABA-R) benzodlazepine receptor (BZ-FI)-chlodde channel complex Including the barbiturate binding Idte (BARB) CI ®

Chlodde channel protein BZ-R protein

barbiturates picrotoxinin Ro 5-3663

inverse agonists

GABA-R protei

GABA agonlsts GABA antagonists

FIG. 1. Schematic diagram of the GABA-BZ receptor complex illustrating the most important interactions between the various binding sites. All three types of binding sites shown regulate the chloride channel. The BZ and barbiturate binding sites can enhance (or inhibit) the binding of GABA. For discussion of the various discriminative stimulus properties of substances acting at this site see Sections 2.1. and 2.2. Adapted from Pole et al., 1982. Reprinted with permission of the authors and copyright holder, Springer-Verlag, Heidelberg.

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receptor ligands have also been synthesized for each of these ligand classifications. Although /3-carboline and benzodiazepine structures are distinct, the fact that they bind to the same receptor suggests that agonist substances should possess similar discriminative properties with BZs. This has been demonstrated in several studies using CDP or diazepam as the training drug (Stephens et al., 1984a, 1987); in addition, antagonism of BZ cues with/3-carboline and other non-BZ antagonists and inverse agonists has been reported (Bennett et al., 1987; Shannon et al., 1988; Stephens et aL, 1984b). Many of the newer /3-carbolines and other non-BZ BZ ligands are partial as opposed to full agonists, That is they do not show the full range of BZ effects, and may in some cases antagonize the effects of full agonists. Typical partial agonists require a higher receptor occupancy to achieve the same effects as an agonist, and this occupancy rate may differ dramatically from behavior to behavior. Given that there now exist BZ receptor ligands which in many cases show only some of the expected BZ properties e.g. anticonflict but no sedative actions, it becomes possible to research the nature of the BZ cue more thoroughly. Nonsedating BZ receptor ligands such as Z K 91 296 and CGS 9896 generalize readily to BZs (Stephens et al., 1987; Sanger, 1988), suggesting that sedative properties are not essential for the establishment of the cue. A number of non-BZ BZ receptor ligands have been reported recently as producing discriminative stimuli. However, the effects of BZ ligands in substitution experiments has been far from simple. Z K 95 962 is a/3-carboline partial agonist at the BZ receptor (Stephens et aL, 1987). It generalizes to CDP and diazepam, and CDP and diazepam generalize to ZK 95 962 (Andrews and Stephens, 1988). However, further research indicates that the /3-carboline and

BZ discriminative stimuli are not as similar as first thought. Table 1 shows the ED50s for generalization and antagonism of the CDP and Z K 95 962 cues. There are several striking differences, notably the failure of the full agonist Z K 93 423 to generalize to, and RO 15-1788 to antagonize, the Z K 95 962 cue. Similar anomalies have been reported for other partial agonists. For example, zolpidem is an imidazopyridine which binds to central BZ receptors, but its profile of action suggests that in contrast to other BZ ligands the dominant effect is one of sedation (Sanger et al., 1987). In zolpidem trained rats substitution by BZs occurs only at high, extremely sedative doses (Sanger and Zivkovic, 1986; Sanger, 1988). Moreover, generalization to CDP is usually reported as incomplete and again only occurring at very high sedative doses (Sanger, 1988). The same group have also shown that zolpidem interacts with other partial agonists in an atypical manner. The partial agonists CGS 9896 and ZK 91 296 both substituted for, but did not antagonize the CDP discriminative stimulus; in contrast both substances antagonized the zolpidem discriminative stimulus (Sanger, 1987; Sanger and Zivkovic, 1987). In a recent article it was suggested that the anticonvulsant and sedative effects of zolpidem could not be explained by a simple receptor occupancy hypothesis because of zolpidem's unusual interaction with a selective antagonist for sedative effects (the /3carboline 3-(methoxycarbonyl)-amino-/3-carboline, /3-CMC; Perrault et al., 1988). However, an equally plausible interpretation of this data might be that the sedative effects of BZs require a high receptor occupancy so that even a small reduction in the binding of such ligands should result in a loss of sedative effects. Since/3-CMC binds only weakly to BZ receptors it is probably capable of displacing sufficient zolpidem to block its sedative effects while still

TABLE1. The Ability o f Various B Z - G A B A Receptor Ligands to Substitute for, or to Antagonize the Discriminative Stimuli Induced in Rats by the Sedative B Z Chlordiazepoxide and the Nonsedative fl-carboline Z K 95 962

Drug cue Training dose Generalisation Test Drug Chlordiazepoxide ZK 95962 Diazepam CGS 9896 ZK 91296 ZK 93423 Phenobarbital ZK 93426 RO 151788 Antagonism Test Drug RO 151788 FG 7142 ZK 91296 ZK 93426

Chlordiazepoxide (5 mg/kg)

ZK 95962 (10 mg/kg)

2.5 1.6 1.5 2 10 0.1 22 > 20 >40

1.5 1.5 3.8 5 40 > 1.25 >40 20 2.3

1.4 1.8 0.8 0.26 >X

>10 >5 >20 10 Further testing usually not continued as subjects stopped responding

All values represents EDs0s in mg/kg. Data adapted from Andrews and Stephens (1988). See text under Section 2.2 for further discussion.

Drug discrimination models leaving enough attached for anticonvulsant activity which generally requires much lower doses of BZs. The partial agonist CGS 9896 also has an interesting profile of action in comparison to the BZs and zolpidem. This compound is one of a series of pyrazaloquinolines which although active in animal models of anxiety (CGS 9896 is also reported active in the clinic), do not show other effects typical of BZs (Bennett, 1987). Due to the lack of side effects or interactions with substances from other classes such as ethanol or hexobarbital, it has been proposed that CGS 9896 should be labeled an anxioselective substance (Bennett, 1985). Furthermore, Bennett (1985) has suggested that the discriminative stimulus generated by CGS 9896 is dependent on its anxiolytic activity. In keeping with the theme of this paper a specific anxiolytic cue must be regarded with scepticism. It is certainly true that the discriminative stimulus properties of CGS 9896 differ somewhat from that of BZs such as diazepam, but this does not mean that the cue formed is anxiolytic. CGS 9896 displays its activity by an action at the BZ--GABA receptor complex and has the profile of a classic partial agonist at that complex. CGS 9896 will both generalize and antagonize BZ discriminative stimuli, is itself antagonized by RO 15-1788, but not by another type of antagonist--the inverse agonist DMCM (6,7-dimethoxy-4-ethyl-fl-carboline-3-carboxylic acid methyl ester (Bennett et al., 1985; Bennett, 1987; Leidenheimer and Schechter, 1988a), and although interactions with barbiturates in other tests may be minimal, generalization to pentobarbital is readily elicited (Leidenheimer and Schechter, 1988a). The stimulus properties of this and other BZ receptor ligands are however helpful in identifying behavioral differences between the drugs and their interaction with the BZ-GABA receptor complex. Although the effects of BZs are ultimately achieved by an action at the chloride ionophore coupled to the GABA receptor, the discriminative stimulus properties of diazepam apparently cannot be mimicked by GABA itself: GABA agonists and antagonists do not generalize or antagonize the diazepam cue (Haug, 1983). And although GABA agonists also possess muscle-relaxant and sedative properties they are not generally found to possess anxiolytic effects. Again caution must be used in interpreting the results: these findings may suggest that the discriminative stimulus is not dependent entirely on sedative or musclerelaxant effects, but it is not proof that the cue is therefore based on anxiolytic properties. Surprisingly, the GABA agonist SL 75 102 shows a high degree of stimulus similarity with CGS 9896 (Leidenheimer and Schechter, 1988a), and it remains to be seen as to whether or not this substance also exhibits anxiolytic effects. The discriminative stimulus effects of the newer compounds are confusing. There are a range of compounds which both generalize and antagonize BZs, and each other; may interact poorly with barbiturates but may also generalize to the barbiturate discriminative stimuli; are antagonized by some antagonists but generalize to others. In fact these confusing asymmetries can also be observed in the

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BZs themselves. It appears that one little studied phenomena in this substance class is the effect of the training dose on the discriminative stimulus formed. RO 15-1788 has been shown both to antagonize and generalize to the CDP cue depending on the training dose of CDP used (De Vry and Slangen, 1986). In addition the asymmetrical generalization seen between the BZ lorazepam and pentobarbital in appropriately trained animals (Ator and Griffiths, 1989a,b) may also be due to the dose used in training and testing, as well as to the development of tolerance to some of the disruptive effects of the test drugs (although the authors discuss their work as indicating inhomogeneity in the stimulus effects of depressant drugs). Some of the behavioral data may be interpreted to suggest that there are several subtypes of BZ receptor, and that these subtypes may be selectively responsible for specific BZ effects. Clearly drug discrimination studies will play an important part in characterizing these distinct effects. However, it must be noted that although subtypes of the central BZ receptor (the receptor in the CNS GABA-BZ complex as opposed to the 'peripheral' BZ receptor) have been identified by both biochemical pharmacologists (BZ1 and BZ2 receptors; Klepner et al., 1979) and by molecular biologists (e.g. Levitan et al., 1988), any possible distinct functional role or specific ligands are as yet unknown or unavailable (but see Langer and Arbilla, 1988 for a recent proposal on classification). As discussed above, disparities between compounds may not be dependent so much on different subtypes as on dosing and training methods as well as perhaps on pharmacokinetic properties of the compounds. It is apparent however, that there has been a shift in the approach to drug cues based around BZ receptor ligands from if-it-substitutes-its-an-anxiolytic, to the more pragmatic search for behavioral correlates of BZ receptor function. Indeed, it may now be of more interest to seek out active BZ ligands which do not generalize to BZ discriminative stimuli as one approach to identifying novel BZ anxiolytics.

2.3. SEROTONINCUES

The introduction of buspirone into the clinic for the treatment of anxiety has led to a change in how the search for anxiolytics is approached. Buspirone is a 5-HT~A partial agonist which in contrast to the benzodiazepines is reported to be an anxioselective agent (Taylor et al., 1984, 1985); that is buspirone possesses no anticonvulsant or muscle-relaxant effects in addition to its anxiolytic-like properties. Buspirone does not bind to BZ receptors and therefore shows no interaction with BZs, or substances thought to exert effects through the BZ-GABA receptor complex. It has been postulated for many years that the anxiolytic effects of BZ may be due to actions on the limbic 5-HT system (Robichaud and Sledge, 1969; Wise et al., 1972; Gray, 1982). However, it was not until the last few years that intense work on the putative anxiolytic actions of 5-HT iigands began. Undoubtedly this was partly due to the relation between 5-HT and hallucinogenic substances such as d-lysergic acid diethylamide (LSD),

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J.S. ANDREWSand D. N. STEPHENS

and partly to a lack of well defined ligands for the separate receptor subtypes and their association with particular behavioral effects. Classical tests for anxiolytic substances are almost all tests for BZ action. It is therefore not surprising that 5-HT anxiolytics are not consistently identified in such tests (Goldberg et al., 1983; McClosky et al., 1987; Pellow et al., 1987; Taylor et al., 1985) or that 5-HT agents do not substitute in BZ cues (Rauch and Stolerman, 1987). Several new tests have been introduced to help identify 5-HT-like anxiolytics (e.g. Jones et al., 1988), although they too may suffer from being tests of drug action as opposed to animal models of anxiety. In addition, there has been a renewed interest in the discriminative stimulus properties of the new putative anxiolytic class. In part this interest has arisen because of the great proliferation of 5-HT receptor subtypes and the availability of relatively specific ligands for each subtype. The question then is clearly what stimulus properties if any, do the putative 5-HT anxiolytics have in common. Since the first 5-HT anxiolytic in the clinic is reported to be selective for the 5-HT~A subtype, it is not surprising that drug discrimination studies have concentrated on substances acting at this receptor subtype. Buspirone is reported to form a discriminative stimulus in rats (Hendry et al., 1983), but the discrimination was difficult to train because of strong response depressant effects. At the present time it is unclear whether this difficulty was due to the choice of training dose, or the discrimination procedure used. Buspirone does, however, form a potent and specific discriminative stimulus in pigeons (Mansbach and Barrett, 1987), The difference in potency of buspirone in pigeons and rodents has been noted in several studies (e.g. Barrett et al., 1986), but as little is known concerning this difference, or of the general nature of discriminative stimuli in pigeons, it is perhaps better to concentrate on studies involving mammalian species. Ipsapirone, a structural analog of buspirone, but reportedly showing a greater selectivity for the 5-HTIA receptor, has a similar profile of action to buspirone in animal tests and is currently undergoing clinical trials for the treatment of anxiety (Glaser, 1988). An ipsapirone drug cue was established to investigate the pharmacological nature of the interoceptive stimulus (Spencer and Traber, 1987; Cunningham, 1989). The cue appeared to be highly specific: only substances with a high affinity for the 5-HTIA receptor substituted for the training drug (buspirone, 8-OHDPAT (8-hydroxy-2-(din-propylamino)-tetralin), 5-OMe-DMT (5-methoxyN,N-dimethyltryptamine)), whereas nonselective 5HT ligands such as methysergide, 5-HT~B selective ligands or non-5-HT ligands e.g. diazepam and pentobarbital failed to generalize. However, antagonism of the cue by a 5-HT~A antagonist such as NAN-190 (Glennon et al., 1988), has still to be demonstrated, and the ability of each of the substances to cross generalize with one another is still unclear. Ipsapirone substitutes for the discriminative stimuli produced by 8-OHDPAT and 5-OMe-DMT (Cunningham et al., 1987; Spencer et al., 1987), but 5-OMe-DMT does not generally substitute successfully for the 8-OHDPAT discriminative stimulus

(Glennon, 1986; Cunningham et al., 1987), although 8-OHDPAT will substitute for 5-OMe-DMT in trained animals (Spencer et al., 1987). 8-OHDPAT has been reported to label both preand postsynaptic sites (Hall et al., 1985), and it has been suggested that 8-OHDPAT may act as an autoreceptor agonist and a postsynaptic antagonist. It might therefore be expected that the effects of 8-OHDPAT would be difficult to predict and may in fact show opposite effects on behavior depending on the dose. Interestingly, 8-OHDPAT has been reported as both anxiolytic and anxiogenic in animal tests (Carli and Saminin, 1988; Critchley and Handley, 1987). Although 8-OHDPAT does form a discriminative stimulus to which both buspirone and ipsapirone generalize, several researchers have reported that rats trained in the cue are more aggressive or irritable than usual (Winter, 1988). Furthermore, although in general dopaminergic agonists, or non5-HT~A selective substances do not substitute for 8-OHDPAT, yohimbine, an alpha-2 antagonist substitutes (Winter, 1988; Winter and Rabin, 1989), as does lisuride, an ergoline possessing both dopaminergic and serotonergic properties (Cunningham et al., 1987). As both lisuride and yohimbine generalize (Appel et al., 1982; Colpaert, 1984) and ipsapirone partially generalizes (Arnt, 1989) to the LSD discriminative stimulus, this may suggest that all of these compounds also possess 5-HT: activity, around which the LSD cue appears to be based (Cunningham and Appel, 1987). Yohimbine is reported to exert anxiogenic effects (e.g. Davidson and Lucki, 1987), effects opposite to the anxiolytic properties predicted, and apparently observed following ipsapirone. It is then perhaps surprising to note that yohimbine also generalizes to ipsapirone (Winter, 1988; Winter and Rabin, 1989). These results would suggest that unlike other drug classes the 5-HTIA discriminative stimulus is much less specific, or that chronic treatment with 5-HT agents produces behaviorally significant interactions with other systems. These results again illustrate the need to treat drug discrimination data with care. 5-HT discriminative stimuli cannot be simply represented as a short cut to identifying novel anxiolytic agents. As already emphasized, at the present time anxiolytic cues p e r se do not exist, but the discriminative properties of anxiolytic drugs can be used to identify a set of stimuli associated with the drug to discover what properties are related, allowable, or unnecessary for, an anxiolytic compound. 2.4. EXCITATORYAMINOACIDS Recent research has shown that antagonists for excitatory amino acids (EAA, e.g. glutamate, asparate) possess muscle relaxant, anticonvulsant and anticonflict activity (Bennett and Amrick, 1986; Clineschmidt et al., 1982; Stephens et al., 1986; Stephens and Andrews, 1988; Turski et al., 1985). Of the three main receptor subtypes identified, N-methyl-o-aspartate (NMDA) antagonists are the most studied. N M D A can produce convulsions and would appear to be unsuitable for drug discrimination studies. However, rats can be trained to discriminate N M D A from saline at subconvulsant

Drug discrimination models doses (Amrick and Bennett, 1987b). Although this cue has not been fully characterized the discriminative stimulus is blocked completely by the competitive antagonist CPP (3-(2-carboxypiperazin4-yl)propyl-l-phosphonic acid), but only partially by another N M D A antagonist, AP7 (2-amino-7-phosphonoheptanoic acid), and diazepam (Amrick and Bennett, 1987b). The authors suggest that the incomplete antagonism by AP7 is due to the poor penetration of APT into the brain. Unfortunately, despite reports that AP7 exerts anticonflict activity in animals (Bennett and Amrick, 1986; Stephens et al., 1986; Stephens and Andrews, 1988) too little is known about the true nature of the anxiolytic effects. AP7 does generalize to the diazepam discriminative stimulus (Bennett and Amrick, 1986), although whether this is due to an as yet unknown interaction with the BZ~3ABA receptor complex or behavioral similarities such as muscle relaxation or anticonflict activity is unclear. A recent study has suggested that another competitive N M D A antagonist CPP may have a greater affinity for benzodiazepine than N M D A receptors (White et al., 1988). At the present time excitatory amino acid antagonists do not appear to be useful anxiolytics (mainly because it is difficult to evaluate substances with such poor brain penetration). It is therefore too early to consider using EAA antagonist cues to help identify novel anxiolytics. Moreover, one of the most bioavailable compounds MK-801 (Clineschmidt et al., 1982) appears to share a common discriminative stimulus with the hallucinogen phencyclidine (PCP, Tricklebank et al., 1987; Jackson and Sanger, 1988; Koek et aL, 1988; Willets and Balster, 1988a), perhaps not the most desirable property for an anxiolytic to possess. However, the discriminative stimulus properties of noncompetitive NMDA antagonists such as MK 801, may not be the same as those produced by the competitive N M D A antagonists such as AP7 or CPP. Peripheral administration of AP7 or CPP does not produce generalization to PCP (Jackson and Sanger, 1988; Willetts and Balster, 1988b). However, centrally administered AP7 will substitute for PCP in pigeons and rats (Tricklebank et al., 1987; Willetts and Balster, 1988b) albeit at doses which cause a marked response suppression in rats; in contrast CPP only partially generalizes to the PCP cue (Willetts and Balster, 1988b). Barbiturates are also reported to generalize partially to the PCP discriminative stimulus (Colpaert, 1977) and, given that barbiturates also share some similar discriminative stimulus properties with BZs it may be important to further characterize the discriminative properties of EAA using barbiturate cues. To date the interoceptive effects of NMDA antagonists are relatively undefined, but appear to be a combination of not only a specific intrinsic effect but also BZ, PCP and possibly barbiturate-like stimuli. Whether this reflects direct actions at the appropriate receptors, due to nonspecificity of the available ligands, or an indirect effect of N M D A stimulation or antagonism has yet to be determined. In this regard it is interesting to note that Bennett and Amrick (1986) interpreted the AP7 cue as a muscle relaxant cue because of its generalization to diazepam, and the similarity of its effects to other muscle relaxants in conflict tests. JPT 47/2--1

273 3. ANXIOGENIC CUES

Several compounds have been reported to produce anxiogenic effects in man e.g. the convulsant pentylenetetrazole (Rodin and Calhoun, 1970), the fl-carboline partial inverse agonist F G 7142 (Dorow et al., 1983), and the serotonin receptor agonist m-chlorophenyl-piperazine (Charney et al., 1987). These reports, coupled to the ever present need for a relevant animal model of anxiety have led several groups to investigate the discriminative stimulus properties of putative anxiogenic substances. A model which is based on a clear and reproducible state of anxiety would dearly be a great advance over present 'conflict' models in anxiety research, many of which can be easily interpreted as models sensitive to the disinhibitory, or worse, possibly amnestic, effects of benzodiazepines. However, although models based on chemically induced anxiety possess a certain attraction and some face validity, they are not without problems. It is worth taking some time to examine one such model thoroughly and the dangers inherent in interpreting the data produced. 3.1. PENTYLENETETRAZOLECUE Pentylenetetrazole (PTZ) is a well known convulsant drug widely used in both epilepsy and anxiolytic research. Several groups have reported that subconvulsant doses of PTZ can serve as a discriminative stimulus (Shearman and Lal, 1980; Stephens et al., 1987), and it has been suggested that the discriminative stimulus induced by PTZ may be one of anxiety (Shearman and Lal, 1980). Evidence in support of this proposal has arisen from several sources. In humans PTZ has been reported to induce feelings of intense anxiety and impending doom (Rodin and Calhoun, 1970), and in rats the autonomic signs following PTZ administration are similar to those found in stress or anxiety (Emmett-Oglesby et al., 1987). The discriminative stimulus itself is antagonized by known anxiolytics such as diazepam (Shearman and Lal, 1980, 1981; Lal et al., 1980; Lal and Fielding, 1984; Benjamin et al., 1987), whereas other substances reported to induce anxiety in humans generalize to the discriminative stimulus (e.g. FG 7142, Stephens et al., 1984a; cocaine, Shearman and Lal, 1981). Moreover, rats trained to discriminate PTZ and then made 'dependent' on diazepam will select the PTZ-associated lever during diazepam withdrawal in the absence of PTZ or diazepam, a result interpreted as being due to withdrawal-induced anxiety (Emmett-Oglesby et al., 1987; Harris et al., 1988). Interestingly, Velluci et al. (1988) have demonstrated that rats subjected to conspecific defeat also select the PTZ lever even when injected with placebo. Lal and Emmett-Oglesby (1983) have reported that anticonvulsant potency does not correlate well with activity in the cue, and, as several clinically effective antiepileptic drugs do not appear to antagonize the cue, have proposed that the discriminative stimulus is more related to the anxiogenic than to the convulsant action of PTZ. However, there are several flaws with this particular argument and the model in general. In typical tests for anticonvulsant activity a high convulsant dose of

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PTZ is given and the animals tested only once. Repeated injections of subconvulsant doses of PTZ eventually leads to seizures in rats, a process known as kindling. The efficacy of anticonvulsant drugs in preventing kindled seizures sometimes differs markedly from that observed in the acute PTZ model (Ito et al., 1977). In our own lab we have observed that rats trained to discriminate PTZ from saline sometimes exhibit seizures following administration of the training dose. More often the rats display wet-dog shakes or tremors, behaviors similar to those observed by Racine (1972) in his description of the early stages of seizure development in amygdaloidkindled rats. These symptoms have also been reported by other groups working with rats receiving chronic PTZ treatment (Lal and Emmett-Oglesby, 1983), suggesting that kindling may be occurring in these animals. Many, if not all, of the substances reported to substitute for PTZ are themselves convulsants (Lal and Emmett-Oglesby, 1983), but perhaps of more interest, many of the drugs which generalize and are not themselves convulsant on first administration are proconvulsant and induce kindled seizures on repeated administration (e.g. cocaine, Stripling and Ellinwood, 1977; F G 7142, Little et aL, 1987; Corda et al., 1987). Indeed, for some such as cocaine, there is cross-sensitization between itself and PTZ in animals kindled with PTZ (L. Turski, unpublished observations). The potent convulsant strychnine generalizes only partially to the PTZ discriminative stimulus (Shearman and Lal, 1980) and can be seen as going some way to help answer criticisms of the possible convulsant nature of the cue. However, not all convulsants have the same mode of action as PTZ. Strychnine-induced convulsions are primarily spinal in origin and are mediated by glycinergic rather than GABAergic mechanisms. Therefore, strychnine might be expected to produce a rather different internal stimulus from PTZ. In an experiment designed to investigate the relationship between anticonvulsant activity and activity in the PTZ cue, groups of rats were trained to discriminate PTZ from saline or were kindled using a slightly higher, but still subconvulsive dose of PTZ. A series of anxiolytic and anticonvulsant drugs were used to antagonize the cue, and kindled PTZ seizures as well as seizures induced by an acute high dose injection of PTZ (Andrews et al., 1989). In keeping with other reports (Shearman and Lal, 1980; Lal and Emmett-Oglesby, 1983), the results showed that all benzodiazepine receptor ligands antagonized the discriminative stimulus regardless of their activity against acutely induced PTZ seizures. Furthermore, two clinically effective anticonvulsants antagonized the cue (valproate and ethosuximide), whilst two others (carbamazepine and phenytoin) failed to antagonize the cue. Substances which failed to antagonize the cue failed to antagonize PTZ kindled seizures, and several substances which showed extreme potency differences in antagonizing acutely induced PTZ seizures and the cue were also strikingly more effective in antagonizing PTZ kindled seizures (see Table 2). One of the most important criticisms of the PTZ cue as an anxiogenic model is its reliance for validity

on antagonism by benzodiazepine-like anxiolytics; nonbenzodiazepine anxiolytics, such as buspirone, are ineffective (Table 2, Ator et al., 1989). In fact it has been reported that buspirone generalizes to the PTZ discriminative stimulus in baboons (Ator et al., 1989), an effect that may be dependent on buspirone's known effects on the dopamine system, but not consistent with the clinical profile of buspirone. (Interestingly, File (1988) has demonstrated that in some circumstances, such as combination with antidepressants, buspirone can induce convulsions in rats.) Moreover, all benzodiazepine anxiolytics tested in the cue to date also possess anticonvulsant activity, particularly against kindled seizures. It is well known that the anticonvulsant activity of BZs is a good predictor of anxiolytic activity in the clinic (Clody et al., 1983), but this does not automatically make an anticonvulsant an anxiolytic. These observations raise the question as to whether the PTZ discriminative stimulus is related to some form of preseizure activity (aura?). Neither carbamazepine nor phenytoin antagonize the cue (Andrews et al., 1989; Lal and Emmett-Oglesby, 1983), but neither of these substances are effective against PTZinduced seizures. It has been argued that if the PTZ cue was based on a preconvulsive aura, then antiepileptic agents should be effective in antagonizing the cue. Interestingly, although phenytoin is effective in the clinic in several forms of epilepsy, it does not prevent the aura many patients report feeling before seizures (Rall and Schleifer, 1985). It might be expected that since not all antiepileptic drugs are effective in all convulsant models, agents ineffective in the PTZ model of epilepsy would not be active against the PTZ cue. In the experiment cited above (Andrews et al., 1989), only substances which antagonized PTZ-induced seizures antagonized the cue. One of these, valproate, has been reported to possess anxiolytic-like effects (Lal et al., 1980); however, to date we are unaware of any clinical data indicating anxiolytic activity of ethosuximide in the clinic. Indeed, in certain anxious patients anxiety has been reported as a side effect of ethosuximide therapy (Rail and Schleifer, 1985). Studies showing that during withdrawal from chronic BZ treatment, animals previously trained to discriminate PTZ from saline will select the PTZassociated lever in the absence of PTZ, have been suggested as a further validation of the model (Harris et al., 1988). However, although anxiety is one symptom of BZ withdrawal, so too in some circumstances are convulsions (Breir et al., 1984; Levy, 1984). Interestingly, a current topic of some concern is the withdrawal of BZs from epileptic patients who have become tolerant to their potent anticonvulsant properties. This point is of particular relevance to studies purporting to examine withdrawal anxiety using the PTZ cue. These studies assume the validity of the PTZ discriminative stimulus as an anxiety model. Even in this limited context, however, the results are not easy to interpret; it is not clear whether the stimulus detected by the rat is a true anxiety, or a contrast between its no-drug state and the chronic drug state in which it was presumably chronically less anxious. Interpretation is further complicated by the

Drug discrimination models

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TABLE2. The Relationship between Activity in the PTZ Cue of Full and Partial BZ Receptor Agonists and Clinically Effective Antiepileptics and Activity Against Clonic Convulsions Induced by an Acute Injection of PTZ and Against PTZ Kindled Seizures Substance

PTZ cue

Acute PTZ clonic

Kindled PTZ seizure

BZ agonists Diazepam Chlordiazepoxide Lorazepam Lormetazepam Triazolam Midazolam Clonazepam

0.37 1.06 0.05 0.04 0.02 1.25 0.32

1.86 3.1 0.36 0.22 0.25 2.49 0.6

1.84 1.87 0.18 0.24 0.17 0.82 0.24

Non-BZ BZ-agonists ZK 91296 ZK 93423 ZK 95962 Zolpidem CGS 9896 CL 218872

0.83 0.08 0.18 0.96 1.47 2.16

10.8 0.66 12.87 6.81 > 80 4.32

1.86 0.32 0.59 1.83 1.53 4.86

Antiepileptics Valproate Phenytoin Carbamazepine Ethosuximide

126.84 > 60 > 40 107.69

293.6 > 100 > 80 212

99.99 > I00 > 80 176

New/potential anxiolytics Buspirone MK-801 2-APH

> 2.5 >0.32 > 80

> 30 >0.5 --

> 30 >0.5 > 80

All values EDs0s (mg/kg) rat. Adapted from Andrews et al., 1989. Note that a compound's ability to antagonize the PTZ discriminative stimulus is predicted by its ability to prevent kindled seizures, but not necessarily by its action against seizures induced following a single high dose of PTZ. Of the clinically effective antiepileptics, only those with the ability to antagonize PTZ convulsive effects were active in the cue. For discussion see text in Section 3.1. For full details of the experimental procedures see Andrews et al., 1989. Reprinted with permission of the copyright holder, Alan R. Liss, Inc., New York. nature of chronic PTZ and diazepam treatments. Before receiving chronic diazepam the rats have been trained on the PTZ discriminative stimulus. The evidence suggests that the rats become more sensitive to PTZ possibly through kindling. The rats are then treated with diazepam over several days before withdrawal. Both withdrawal from chronic diazepam treatment and chronic treatment with PTZ decrease seizure thresholds (in the case of PTZ permanently). If the PTZ cue is based on a preconvulsant effect it may not be surprising that partial generalization to the PTZ cue is observed in the first few days following withdrawal from diazepam. The alternative view that the rats have chronic anxiety, initially suppressed by diazepam and then returning as rebound anxiety is, in our view, a little too anthropomorphic. Thus, and unfortunately, this model may not be of great help in analyzing the state arising from BZ withdrawal in the clinic as was first believed. 3.2. FG 7142 CUE Recently a cue based around the fl-carboline partial inverse agonist FG 7142 has been established (Leidenheimer and Schechter, 1988b), and although it

is not yet fully characterized it has been suggested that the discriminative stimulus may be anxiogenic in nature. This follows mainly from two lines of reasoning, firstly that F G 7142 has been reported to induce feelings of anxiety in humans (Dorow et al., 1983) and secondly that F G 7142 generalizes to PTZ, an anxiogenic cue. As a model for testing anxiolytic drugs it is open to similar criticisms to those leveled at the PTZ cue. F G 7142, like PTZ induces kindled seizures in rats, therefore, substances active in the cue are likely to be anticonvulsants as opposed to pure anxiolytics. Moreover, FG 7142 is believed to exert its effects through central BZ binding sites (Stephens et al., 1987; Corda et al., 1987). Thus an action at this particular site may be blocked not only by BZ agonists possessing anxiolytic (and anticonvulsant) properties, but also by neutral BZ receptor antagonists with no intrinsic action of their own. These two examples again illustrate the necessity for care when interpreting the nature of discriminative stimuli. Undoubtedly there are many substances which can induce feelings of anxiety when administered acutely to humans. However, many such as PTZ and F G 7142 are also capable of causing seizures in animals following repeated adminis-

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tration. Therefore, as suggested above, apparent anxiolytic activity in cues based on such substances is likely to be confounded with anticonvulsant properties. In this respect it is interesting to note that rats which have been kindled with F G 7142 show no changes in comparison to saline treated animals in the social interaction, plus-maze, Vogel conflict or startle response tests of anxiety (Taylor et al., 1988). In the two examples above, drug effects are probably more parsimoniously explained by convulsant or anticonvulsant properties.

4. A N T I D E P R E S S A N T CUES The discriminative stimulus properties of antidepressants are as poorly understood as the exact mode of action of the antidepressants themselves. Several groups have successfully trained rats to discriminate an antidepressant from saline. Unfortunately, the question as to whether or not the various antidepressants share a common discriminative stimulus has not been adequately addressed as very few generalization studies have been carried out. This is due partly to the difficulty in training, and partly to the high mortality rates in animals used in such tests (Shearman et al., 1978; Jones et al., 1980). The prototype class of antidepressants, the tricyclics, present a particular problem. Using a T-maze shock avoidance paradigm, Overton (1982) has successfully trained rats to discriminate several antdepressant drugs: doxepine, imipramine and amitryptyline. Although cross generalization with these tricyclics could be demonstrated, the overall success of the experiments was low (cited in Jones et al., 1980). Training doses were high in comparison to other behavioral effects observed with such compounds and a low survival rate following repeated administration was observed. Indeed, itappears that only at near toxic doses do the tricyclics produce a discriminative stimulus (Shearman et al., 1978; Overton, 1982; see also Jones et al., 1980). Schechter (1983) compared the effects of differing stress conditions on the ability of rats to recognize the discriminative stimulus induced by imipramine. Only the unstressed animals were able to learn the discrimination successfully, and other tricyclics substituted for the training drug. More recent additions to the antidepressant pharmacy are reported to be less toxic than the prototype antidepressants such as imipramine. One, the phenylaminoketone buproprion although active in the clinic (but see Blackwell and Simon, 1986) has a very different profile of action to classical antidepressants. Although buproprion forms a specific discriminative stimulus in rats (Jones et al., 1980), other antidepressants, with the exception of the putative antidepressant viloxazine, do not substitute for the training drug. In fact the discriminative stimulus appears to be related to a stimulant effect as amphetamine, caffeine and methylphenidate all substitute for buproprion. Another novel antidepressant, the phosphodiesterase (PDE) inhibitor rolipram (Davis, 1984; Wachtel and Schneider, 1988) also forms a discriminative stimulus in rats (Ortmann and Meisburger, 1986; Yamamoto et al., 1987; Andrews

et al., in preparation). In these studies, other specific

and nonspecific phosphodiesterase inhibitors generalized to rolipram but tricyclic antidepressants did not. This would suggest that the discriminative stimulus is based on its phosphodiesterase activity. However, it is unlikely to be so simple: although rolipram inhibits locomotor activity, lisuride, an antiparkinson agent without PDE inhibitory properties, but possessing both dopaminergic and serotonergic activity, has been reported to generalize or partially generalize to the training drug (Andrews et al., in preparation). The few results available using antidepressants as discriminative stimuli would appear to suggest (1) that tricyclics produce a similar discriminative stimulus albeit at near toxic doses and (2) that other clinically active antidepressants can form discriminative stimuli but which are unrelated to other antidepressant drugs. Thus, there is no one common cue for antidepressant drugs, a result in keeping with clinical reports that antidepressants do not produce common subjective effects in normal human subjects (Lehmann and Hopes, 1977). However, it is clear that drug discrimination techniques offer a unique way to investigate the differing interoceptive effects produced by these compounds.

5. S U M M A R Y A N D CONCLUSIONS Drug discrimination techniques are an important method for investigating the interoceptive properties of different psychoactive drugs. However, the discriminative stimuli formed are often complex and occasionally unexpectedly related to other behaviorally active drugs. It appears then that data obtained from such experiments must be treated cautiously: because CDP is anxiolytic and forms a discriminative stimulus it does not automatically follow that the cue is an anxiolytic cue. In this respect results with antidepressants are particularly revealing: although all antidepressants are more or less equally effective in the clinic, the interoceptive stimuli produced by the different compounds are unrelated, and discrimination occurs typically at doses far removed from the therapeutic dose range. Although it may be disappointing to say that there is no anxiolytic cue, anxiogenic cue, antidepressant cue and so on, recognition of this fact is neither a surprising nor a particularly discouraging state of affairs. The specificity of the discriminative stimulus induced by a drug depends on the specificity of the ligand involved as well as on the training dose used. Although several subtypes of BZ receptors are thought to exist, the evidence for their existence as documented by specific agonists and antagonists is at the present time poor. In areas where specific ligands are available there is more certainty in ascribing behavioral effects to a particular receptor. It is apparent that many discriminative stimuli formed by specific receptor ligands are in fact extremely complex and possibly composed of effects at several receptor subtypes. This is especially notable in the 5-HT cues where the further characterization of these cues has suggested either unusual properties or that the specificity of the ligands is not as good as first believed. Separate components of complex multiple

Drug discrimination models stimuli can be identified by animals trained on a mixture of substances (e.g. concurrent nicotine and midazolam, Stolerman et al., 1987) or substances with effects at multiple receptor sites (e.g. lisuride, Appel et aL, 1982; Cunningham and Appel, 1987). Present findings suggest that different training doses may have an important role in uncovering a variety of stimulus effects from a particular compound. One question which has been poorly researched is the stability of the discriminative stimulus over time. Discriminative stimuli from a wide range of psychoactive substances appear to be stable over months or years (see Schechter et aL, 1989). Anecdotal evidence suggests that this is as true for opiates as for BZs, both of which belong to classes of drugs in which the phenomena of tolerance is well documented. The possibility that the stimulus properties of BZ receptor ligands are anxiolytic in nature must again be questioned when tolerance to BZ anxiolytic, as well as sedative and anticonvulsant actions, can be readily observed but not to the BZ discriminative stimulus. One possible explanation for the apparent lack of tolerance to the discriminative stimuli associated with BZs might relate to stimulus fading. In conventional (e.g. visual) discrimination tasks, reducing the intensity of the discriminative stimulus leads to an initial reduction in discriminative performance. However, providing that the stimulus is still detectable, further training at the new intensity leads to a reinstatement of performance to near previous levels. In the case of drug stimuli, the development of tolerance (reduction of stimulus intensity) would be a gradual process with the animal receiving repeated training during stimulus fading. Provided that tolerance is less than complete, it is to be expected that discriminative performance would remain stable. Unfortunately, no data exist in the literature which might allow an assessment of this idea. It has been suggested in the past that discriminative stimuli in animals are analogous to the subjective effects of the drug in man, and therefore may be a measure of the abuse potential of a drug. For the opiates this may be clear, but for the anxiolytics, particularly the newer partial agonist and 5-HT based anxiolytics which do not appear to induce withdrawal symptoms after chronic treatment this may be of little relevance. In conclusion, research into the discriminative stimulus properties of anxiolytics is at a pivotal stage. New ligands are raising the possibility of an even finer dissection of drug effects on behavior, and for drug discrimination research an ever greater number of questions, which for reasons of space have not been adequately discussed. For example what does partial generalization (or asymmetrical generalization) really mean? Overlapping but not identical stimulus properties or substitution for one component of a complex multiple stimulus? How important is the training dose and the pharmacokinetics of the drug to this phenomena? Do nonparallel generalization curves between training and test drugs mean that generalization is due to a different mechanism of action than that of the training drug? And of increasing interest, how important are contextual cues to the development and maintenance of a discriminative stimulus? Drug discrimination procedures cannot be viewed as a

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simple short-cut to the development of new anxiolytics, but to a fuller characterization of the properties of anxiolytic compounds, which may help shed some light on the reasons for success or failure in the clinic.

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