The CRF(1) receptor antagonist DMP696 produces anxiolytic effects and inhibits the stress-induced hypothalamic-pituitary-adrenal axis activation without sedation or ataxia in rats

Abstract Rationale. CRF₁ antagonists may be effective in the treatment of anxiety disorders while having fewer side effects compared with classical benzodiazepines. Objectives. The effects of a small molecule selective CRF₁ antagonist DMP696 on anxiety-like behaviors and stress-induced increases in corticosterone in rats exposed to a novel environment and on locomotor activity and motor coordination were determined in rats. These effects of DMP696 were compared with those produced by the classical benzodiazepine chlordiazepoxide (CDP). Methods. DMP696 or CDP were administered PO, 60 minutes before behavioral testing in rats. Their effects on latency to exit a dark chamber and stress-induced increase in corticosterone in the Defensive Withdrawal test (an animal model of anxiety), locomotor activity, and rotorod performance (measure of ataxia) were determined. Results. DMP696 significantly reduced exit latency and reversed the stress-induced increase in corticosterone in the Defensive Withdrawal test at doses of 3.0–10 mg/kg and higher. In contrast, CDP significantly decreased exit latency at 10 and 30 mg/kg, but not at 100 mg/kg, due to concurrent non-specific side effects. Unlike DMP696, CDP had no effect on the stress-induced increase in corticosterone at lower doses, but resulted in a significant increase at higher doses. DMP696 did not reduce locomotor activity or impair motor coordination at doses up to 30-fold higher than doses effective in the Defensive Withdrawal model. In contrast, CDP produced significant sedation and ataxia at the same doses that were effective in reducing exit latency. Conclusions. These data suggest that the CRF₁ antagonist DMP696 might retain the therapeutic benefits of classical benzodiazepines but have fewer motoric side effects. Electronic Publication     

Effects of CRF<Subscript>1</Subscript> receptor antagonists and benzodiazepines in the Morris water maze and delayed non-matching to position tests

Rationale Benzodiazepines continue to be widely used for the treatment of anxiety, but it is well known that benzodiazepines have undesirable side effects, including sedation, ataxia, cognitive deficits and the risk of addiction and abuse. CRF₁ receptor antagonists are being developed as potential novel anxiolytics, but while CRF₁ receptor antagonists seem to have a better side-effect profile than benzodiazepines with respect to sedation and ataxia, the effects of CRF₁ receptor antagonists on cognitive function have not been well characterized. It is somewhat surprising that the potential cognitive effects of CRF₁ receptor antagonists have not been more fully characterized since there is some evidence to suggest that these compounds may impair cognitive function.Objective The Morris water maze and the delayed non-matching to position test are sensitive tests of a range of cognitive functions, including spatial learning, attention and short-term memory, so the objective of the present experiments was to assess the effects of benzodiazepines and CRF₁ receptor antagonists in these tests.Results The benzodiazepines chlordiazepoxide and alprazolam disrupted performance in the Morris water maze and delayed non-matching to position at doses close to their therapeutic, anxiolytic doses. In contrast, the CRF₁ receptor antagonists DMP-904 and DMP-696 produced little or no impairment in the Morris water maze or delayed non-matching to position test even at doses 10-fold higher than were necessary to produce anxiolytic effects.Conclusions The results of the present experiments suggest that, with respect to their effects on cognitive functions, CRF₁ receptor antagonists seem to have a wider therapeutic index than benzodiazepines.     

Effect of SB 200646A,a 5-HT2C/5-HT2B receptor antagonist,in two conflict models of anxiety

SB 200646A is the first selective 5-HT_2C/5-HT_2B receptor antagonist and has previously been observed to have anxiolytic-like properties in the rat social interaction test. In the present study the effects of the compound in two conflict models of anxiety, the rat Geller-Seifter and marmoset conflict test, were examined. In the rat Geller-Seifter test, suppressed responding was increased by all doses of SB 200646A between 5 and 40 mg/kg PO when given 1 h pretest. Unsuppressed responding was slightly increased only at 10 mg/kg PO. Suppressed responding was also increased by the benzodiazepine anxiolytic, chlordiazepoxide, at 1, 2.5 and 5 mg/kg PO 1 h pretest. Unsuppressed responding was modestly increased by chlordiazepoxide only at 5 mg/kg PO. In the marmoset conflict test marmosets were trained to lever press for a palatable food reward. Lever pressing was subsequently suppressed by air puffs. In this procedure suppressed responding was increased by both the benzodiazepine anxiolytic diazepam at 2 and 5 mg/kg PO and SB 200646A after 10 and 20 mg/kg PO. Both treatments caused small increases in unsuppressed responding at 2 and 20 mg/kg PO respectively. Taken together with the previous effects of SB 200646A in the rat social interaction test, this is compelling evidence that 5-HT_2C/2B receptor antagonists may possess anxiolytic properties.     

Naloxone blocks anxiolytic-like effects of benzodiazepines in Swiss but not in Balb/c mice

 The ability of naloxone to block the effects of the benzodiazepines chlordiazepoxide and diazepam was evaluated in Swiss and Balb/c mice subjected to the light/dark choice test of anxiety or to a choice paradigm for measuring spontaneous exploratory behaviour. In Swiss mice, naloxone (5 or 10 mg/kg) completely or partially suppressed the anxiolytic-like effects of chlordiazepoxide (5 mg/kg) and diazepam (1 mg/kg) in the light/dark test. Naloxone alone was ineffective. None of these compounds affected locomotion in the free exploratory test. In Balb/c mice, naloxone did not reduce the anxiolytic-like action of benzodiazepines in the light/dark test. Moreover, naloxone did not antagonize the decrease in neophobia observed after anxiolytic treatment in Balb/c mice in the free exploratory paradigm. In this strain, benzodiazepines produced an increase of locomotor activity, whereas naloxone decreased it. The stimulant effects of benzodiazepines on locomotor activity were abolished by naloxone. As naloxone (2 mg/kg) reversed the morphine-induced hyperthermia both in Swiss and in Balb/c mice, differences in possible pharmacokinetic factors between the two strains can be ruled out as an explanation for the failure of naloxone to antagonize anxiolytic-like effects in Balb/c mice. Therefore, the ability of naloxone to reverse anxiolytic effects does not hold for all strains of mice. Received: 19 April 1996 / Final version: 22 November 1996     

Differential EEG effects of the anxiolytic drugs, deramciclane (EGIS-3886), ritanserin and chlordiazepoxide in rats

The influence of serotonergic and benzodiazepine type anxiolytic drugs on the cortical activation and sleep-wakefulness cycle were compared by evaluating the effects of ritanserin and deramciclane (EGIS-3886), two 5-HT₂ receptor antagonists, and chlordiazepoxide on the electroencephalogram (EEG) in freely moving rats. Following drug administration (1, 3, and 10 mg/kg, PO for all drugs), EEG was continuously sampled for 6 h and power spectra were calculated for every 5 s to assess changes in slow wave activity and sleep phases. In a separate test, anticonvulsant effects of the drugs were examined in mice. Both deramciclane and ritanserin slightly increased total time spent in deep sleep (DS) and lengthened sleep episodes. In contrast, chlordiazepoxide had a strong inhibitory action on DS, sleep time being shifted to more superficial light sleep (LS). The incidence and length of the high voltage spindle (HVS) episodes characteristic for the motionless, awake rat were increased at the highest dose of both deramciclane and ritanserin, while it was decreased by chlordiazepoxide. In mice, chlordiazepoxide had a marked anticonvulsant effect, while deramciclane was moderately effective and ritanserin ineffective. In conclusion, the 5-HT₂ receptor antagonist anxiolytic drugs seem to be superior compared to the benzodiazepine type anxiolytic drug, chlordiazepoxide, as ritanserin and deramciclane improved sleep quality by increasing sleep episode length and time spent in DS, while chlordiazepoxide enhanced sleep fragmentation and decreased DS. Received: 1 July 1998/Final version: 4 September 1998     

Effects of 5-HT1A receptor agonists,partial agonists and a silent antagonist on the performance of the conditioned emotional response test in the rat

In the present study, the effects of 5-HT_1A receptor ligands with varying degrees of intrinsic activity at the 5-HT_1A receptor were examined in the conditioned emotional response (CER) test and their effects compared to those of the benzodiazepine receptor agonists, diazepam and chlordiazepoxide. Diazepam (3.0 mg/kg) and chlordiazepoxide (3.0 mg/kg), and the 5-HT_1A receptor partial agonists, ipsapirone (10.0 mg/kg) and gepirone (3.0 mg/kg), alleviated conditioned suppression of lever pressing. The 5-HT_1A receptor partial agonist, buspirone (0.1–1.0 mg/kg), the 5-HT_1A receptor agonist, 8-OH-DPAT (0.01– 0.10 mg/kg), and the 5-HT_1A receptor antagonist, WAY-100635 (0.03–3.0 mg/kg), had no effects on conditioned fear. Neither enhancing the level of food deprivation nor pretreatment with the amnesic agent scopolamine induced anxiolytic-like effects in the present CER test. The anxiolytic-like effects of ipsapirone in this test were completely reversed by WAY-100635. These results indicate that 5-HT_1A agonist, but not antagonist actions, induce an anxiolytic effect in the CER test in rats. Received: 13 March 1996/Final version: 8 July 1996     

The pharmacology of DMP696 and DMP904, non-peptidergic CRF1 receptor antagonists

CRF(1) antagonists DMP696 and DMP904 were designed as drug development candidates for the treatment of anxiety and depression. Both compounds display nanomolar affinity for human CRF(1) receptors, and exhibit >1000-fold selectivity for CRF(1) over CRF(2) receptors and over a broad panel of other proteins. DMP696 and DMP904 block CRF-stimulated adenylyl cyclase activity in cortical homogenates and cell-lines expressing CRF(1) receptors. Both compounds inhibit CRF-stimulated ACTH release from rat pituitary corticotropes. Binding and functional studies indicate that DMP696 and DMP904 behave as noncompetitive full antagonists. DMP696 and DMP904 exhibit anxiolytic-like efficacy in several rat anxiety models. In the defensive withdrawal test, both compounds reduce exit latency with lowest effective doses of 3 and 1 mg/kg, respectively. The anxiolytic-like effect is maintained over 14 days of repeated dosing. In the context of a novel environment used in this test, DMP696 and DMP904 reverse mild stress-induced increases in plasma CORT secretion but at doses 3-4-fold greater than those required for anxiolyticlike efficacy. DMP696 and DMP904 are ineffective in three depression models including the learned helplessness paradigm at doses up to 30 mg/kg. At lowest anxiolytic-like doses, DMP696 and DMP904 occupy >50% CRF(1) receptors in the brain. The in vivo IC(50) values (plasma concentrations required for occupying 50% CRF(1) receptors) estimated based upon free, but not total, plasma concentrations are an excellent correlation with the in vitro IC(50) values. Neither compound produces sedation, ataxia, chlordiazepoxide-like subjective effects or adverse effects on cognition at doses 10-fold higher than anxiolytic-like doses. Neither compound produces physiologically significant changes in cardiovascular, respiratory, gastrointestinal or renal functions at anxiolytic-like doses. DMP696 and DMP904 have favorable pharmacokinetic profiles with good oral bioavailabilities. The overall pharmacological properties suggest that both compounds may be effective anxiolytics with low behavioral side effect liabilities.     

Benzodiazepine/GABA(A) receptors are involved in magnesium-induced anxiolytic-like behavior in mice

Behavioral studies have suggested an involvement of the glutamate pathway in the mechanism of action of anxiolytic drugs, including the NMDA receptor complex. It was shown that magnesium, an NMDA receptor inhibitor, exhibited anxiolytic-like activity in the elevated plus-maze test in mice. The purpose of the present study was to examine interaction between magnesium and benzodiazepine/GABA(A) receptors in producing anxiolytic-like activity. We examined behavior of mice treated with magnesium and benzodiazepine/GABA(A) receptor ligands, in the elevated plus maze. The anxiolytic-like effect of magnesium (20 mg/kg) was antagonized by flumazenil (10 mg/kg) (benzodiazepine receptor antagonist) while combined treatment with the non-effective doses of magnesium (10 mg/kg) and benzodiazepines (diazepam (0.5 mg/kg) or chlordiazepoxide (2 mg/kg)) produced synergistic interaction (increased time in open arms and number of open arm entries) in this test. The obtained data indicate that benzodiazepine receptors are nvolved in the anxiolytic-like effects of magnesium.     

Behavioral pharmacology of the pyrazoloquinoline CGS 9896, a novel putative anxiolytic

CGS 9896, a pyrazoloquinoline that displaces [³H]benzodiazepines from their CNS receptors, has been claimed to be a nonsedative anxiolytic in animals. However, conflicting results have been obtained with this compound in animal tests of anxiety and seizures. The present study reports the effects of CGS 9896 in three animal tests of anxiety, against seizures induced by picrotoxin and pentylenetetrazole, and on the holeboard test in rats. CGS 9896 (injected intraperitoneally) has an anxiolytic-like activity in the social interaction test (5--10 mg/kg) and in the elevated plus-maze (10--20 mg/kg) but not in the Vogel punished drinking procedure (5--10 mg/kg). CGS 9896 had potent anticonvulsant effects against pentylenetetrazole (total protection at 5 mg/kg) but was considerably less potent against picrotoxin-induced seizures (only 50% reduction at 40 mg/kg). CGS 9896 (10 mg/kg) caused significant reductions in spontaneous exploratory and locomotor behavior in the holeboard test in rats. In all test procedures, CGS 9896 was considerably less potent than the benzodiazepines diazepam or chlordiazepoxide. In conclusion, these data support the evidence suggesting that CGS 9896 possesses anxiolytic and anticonvulsant properties in rodents but suggest that caution should be observed in concluding that a compound is nonsedative on the basis of one or two measures.     

Behavioural profiles in the mouse defence test battery suggest anxiolytic potential of 5-HT1A receptor antagonists

  Rationale: Compounds varying in selectivity as 5-HT_1A receptor antagonists have recently been reported to produce anxiolytic-like effects comparable to those of benzodiazepines in the mouse elevated plus-maze procedure. Objective: In view of the potential clinical significance of these findings, the present experiments compared the behavioural effects of diazepam (0.5–3.0 mg/kg) with those of several non-selective 5-HT_1A receptor antagonists [NAN-190, 0.1–3.0 mg/kg, MM-77, 0.03–1.0 mg/kg, (S)-UH-301, 0.3–3.0 mg/kg and pindobind-5-HT_1A, 0.03–1.0 mg/kg], and three selective 5-HT_1A receptor antagonists (WAY100635, 0.01–3.0 mg/kg, p-MPPI, 0.1–3.0 mg/kg and SL88.0338, 0.3–3.0 mg/kg) in the mouse defence test battery (MDTB). Methods: In this well-validated anxiolytic screening test, Swiss mice are directly confronted with a natural threat (a rat) as well as situations associated with this threat. Primary measures taken during and after rat confrontation were flight, risk assessment (RA), defensive threat/attack and escape attempts. Results: Diazepam significantly decreased flight reactions after the rat was introduced into the runway, reduced RA activities of mice chased by the rat, increased RA responses displayed when subjects were constrained in a straight alley and reduced defensive upright postures and biting upon forced contact. All the selective 5-HT_1A receptor antagonists and NAN-190 also reduced flight, RA in the chase test, and defensive threat and attack behaviours. (S)-UH-301 and pindobind-5-HT_1A reduced RA in the chase test, but only partially modified defensive threat and attack. Unlike the other drugs tested, MM-77 produced significant effects only at doses which also markedly reduced spontaneous locomotor activity, suggesting a behaviourally non-specific action. In contrast to diazepam, the 5-HT_1A receptor ligands failed to affect RA in the straight alley test. Following removal of the rat from the test area, only diazepam and (S)-UH-301 reduced escape behaviour (contextual defence) at doses which did not decrease locomotion. Overall, the present findings indicate that except for one RA behaviour and escape responses, the 5-HT_1A receptor ligands studied modified the same defensive behaviours as diazepam, suggesting potential therapeutic efficacy in the management of anxiety disorders. However, the magnitude of the effects of the 5-HT_1A compounds on defence was generally smaller than that of the benzodiazepine. Conclusion: As all of the 5-HT_1A compounds tested in this series share antagonistic activity in models of postsynaptic 5-HT_1A receptor function, it is proposed that this action accounts for their effects on defence. Received: 29 June 1998 / Final version: 16 December 1998     

Corticotropin releasing factor antagonist, α-helical CRF<Subscript>9–41</Subscript>, reverses nicotine-induced conditioned,but not unconditioned,anxiety

Rationale. Unconditioned anxiogenic effects of nicotine have been observed in the social interaction (SI) test 5 min after injection of a low dose and both 5 min and 30 min after injection of a high dose. Conditioned anxiety has also been observed 24 h after testing in the SI with a high dose of nicotine. Objectives. In order to determine whether these three anxiogenic effects shared a common mechanism, we investigated the role of corticotropin releasing factor (CRF). We therefore examined whether the CRF antagonist α-helical CRF_9–41 could block these three anxiogenic effects of nicotine. Methods. To test the unconditioned anxiogenic effects, pairs of male rats were tested in SI 5 min after s.c. vehicle or nicotine (0.1 mg/kg) or 30 min after s.c. vehicle or nicotine (0.45 mg/kg), and 30 min after i.c.v. artificial cerebrospinal fluid (aCSF) or α-helical CRF_9–41. To test conditioned anxiety, rats were exposed to the SI test on day 1, 5 min after vehicle or nicotine (0.1 mg/kg). On day 2, they were re-tested in SI 30 min after i.c.v. aCSF or α-helical CRF_9–41 (5 μg). Results. α-Helical CRF_9–41 did not block the unconditioned anxiogenic effect of either dose of nicotine. Nicotine (0.1 mg/kg, 5 min) elicited a conditioned anxiogenic response that was significantly reversed by α-helical CRF_9–41. The CRF antagonist alone had no effect. Conclusions. CRF is an important mediator of the conditioned anxiety to nicotine, but may not play a role in mediating the acute anxiogenic effects. Electronic Publication     

Chronic treatment with antidepressants affects glycine/NMDA receptor function: behavioral evidence

Like the clinically effective benzodiazepine anxiolytic, chlordiazepoxide, the glycine/NMDA receptor antagonist L-701,324 (3, 7.5 and 10 mg/kg), produces dose-related increases in the percentage of time spent in the open arms and the percentage of entries into the open arms of an elevated plus maze in mice. Consistent with its proposed mechanism of action, these anxiolytic effects of L-701,324 (7.5 mg/kg) are reversed by pretreatment with glycine (500 and 800 mg/kg). Chronic treatment with citalopram (20 mg/kg for 21 days), imipramine (15 mg/kg for 21 days) and electroconvulsive shock (ECS, for 8 days), produced a reduction in the anxiolytic-like actions of L-701,324 (7.5 mg/kg) such that they could not be reversed by glycine. In contrast, the anxiolytic effects of L-701,324 and reversal of these effects by glycine were unaffected by acute treatment with imipramine, chronic administration with placebo or the neuroleptic chlorpromazine, or sham ECS. Further, imipramine administered for 21 days did not affect the anxiolytic effect of 5 mg/kg of chlordiazepoxide. The apparent reduction in the anxiolytic-like actions of a specific glycine/NMDA receptor antagonist following chronic treatment with a variety of antidepressants is consistent with previous neurochemical and molecular studies indicating that chronic antidepressant treatment can affect NMDA receptor function.     

Stereoselective discriminative stimulus effects of zopiclone in rhesus monkeys

Abstract Rationale. The behavioral effects of racemic zopiclone are similar to those of benzodiazepines that positively modulate GABA at the GABA_A receptor complex; however, it is not clear how enantiomers or metabolites of zopiclone contribute to the benzodiazepine-like behavioral effects of racemic zopiclone. Objectives. Racemic zopiclone, its (R)- and (S)- enantiomers, and the (S)-N-desmethyl metabolite, were evaluated for discriminative stimulus effects in untreated and diazepam treated rhesus monkeys. Methods. One group of monkeys discriminated the benzodiazepine midazolam and another group, treated daily with the benzodiazepine diazepam (5.6 mg/kg, PO), discriminated the benzodiazepine antagonist flumazenil. Results. (RS)-Zopiclone (0.32–17.8 mg/kg) and (S)-zopiclone (0.1–10 mg/kg) substituted with similar potencies for midazolam (≥80% midazolam-appropriate responding). The midazolam-like discriminative stimulus effects of (RS)-zopiclone were antagonized by flumazenil (pK _B=7.52). (R)-Zopiclone occasioned a maximum 45% midazolam-appropriate responding at a dose of 100 mg/kg; (S)-desmethylzopiclone produced saline-appropriate responding up to a dose of 100 mg/kg. All four test compounds occasioned predominantly vehicle-appropriate responding in diazepam treated monkeys discriminating flumazenil. (RS)-Zopiclone (10 mg/kg) attenuated the discriminative stimulus effects of flumazenil in diazepam treated monkeys. Conclusions. These results clearly demonstrate that in rhesus monkeys the discriminative stimulus effects of zopiclone are stereoselective and qualitatively similar to those of midazolam. These results fail to show any benzodiazepine-like or benzodiazepine antagonist-like discriminative stimulus effects for (S)-N-desmethylzopiclone, suggesting that any behavioral (e.g. anxiolytic) effects of this compound are not the result of actions at benzodiazepine receptors. Electronic Publication     

Anxiolytic effects of valproate and diazepam in mice are differentially sensitive to picrotoxin antagonism

Although it is widely believed that the anxiolytic effects of benzodiazepines are mediated through facilitation of GABA(A) receptor function, behavioural studies have to date provided rather weak support for this hypothesis. In particular, considerable inconsistency has been noted both for the effects of GABAergic manipulations in animal models of anxiety and the ability of GABA(A) receptor antagonists to block the anxiolytic effects of diazepam (DZ) and chlordiazepoxide. In view of the sensitivity of the murine plus-maze to the anxiety-modulating effects of GABAergic agents as well as classical benzodiazepines, the current study examined the extent to which the anxiolytic actions of valproic acid (VPA) and DZ in this test involve picrotoxin (PX)-sensitive receptor mechanisms. Subjects were male DBA/2 mice, test duration was 5 min, and ethological scoring methods were employed. Our results show that, while devoid of intrinsic behavioural effects under present test conditions, PX (0.25-0.5 mg/kg) selectively antagonised the anxiolytic-like (but not other) effects of VPA (400 mg/kg). In contrast, the same doses of PX failed to block any of the behavioural changes induced by DZ (1.5 mg/kg), including disinhibition of open arm exploration. These data suggest that the plus-maze anxiolytic effects of DZ in DBA/2 mice are not mediated through PX-sensitive GABA(A) receptors. Further studies will be required to assess the generality of present findings to other mouse strains, species and behavioural paradigms.     

Naloxone blocks the effects of chlordiazepoxide on acquisition but not performance of differential reinforcement of low rates of response (DRL)

The opiate antagonist naloxone can completely or partially reverse the effects of the benzodiazepines on appetitive behaviours and conflict tasks involving electric shook. If naloxone changes the anxiolytic action of the benzodiazepines it should, theoretically, be effective in tasks employing nonreward as well as those employing shock.We tested naloxone and chlordiazepoxide on acquisition and performance of a nonreward task, DRL. With both continuous administration during acquisition of DRL, and intermittent administration during stable performance, chlordiazepoxide (5 mg/kg IP) increased burst responding and shifted the peak of the inter-response time (IRT) distribution curve to shorter IRTs. Naloxone (3 mg/kg IP) blocked the effects of chlordiazepoxide on acquisition of DRL. Naloxone (3 mg/kg and 10 mg/kg IP) did not change the effects of chlordiazepoxide on well-learned performance of the DRL schedule.These results show that endogenous opiates could mediate some but not all of the actions of the benzodiazepines. They also suggest that state-dependent and truly anxiolytic effects of the benzodiazepines (McNaughton 1985) may have different pharmacological substrates.     

Evaluation of cannabinoid agonists using punished responding and midazolam discrimination procedures in squirrel monkeys

Rationale  Clinical studies have suggested that marijuana and nabilone have anxiolytic effects in humans, yet studies of anxiolytic-like effects of cannabinoid agonists in mice and rats have yielded mixed results. Objective  The purpose of the present study was to compare the effects of cannabinoid agonists and clinically used anxiolytic drugs in monkeys using punished responding and midazolam discrimination procedures. Methods  Monkeys were trained to discriminate an i.m. injection of 0.3 mg/kg midazolam from saline or, in a separate group, to respond under a multiple schedule of food reinforcement composed of punished and nonpunished components. Effects of the cannabinoid agonists Δ⁹-tetrahydrocannabinol (Δ⁹-THC; 0.01–3 mg/kg), WIN 55,212-2 (0.03–1 mg/kg) and CP 55,940 (0.003–0.03 mg/kg), and the benzodiazepine midazolam (0.01–1 mg/kg) and the barbiturate pentobarbital (1–18 mg/kg) were evaluated. Results  Δ⁹-THC and CP 55,940 did not have antipunishment effects and Δ⁹-THC and WIN 55,212-2 did not produce midazolam-like discriminative stimulus effects up to doses that substantially decreased response rate. In contrast, pentobarbital, like midazolam, increased punished responding at doses comparable to those that substituted for the midazolam discriminative stimulus. Conclusion  Cannabinoid agonists do not have anxiolytic-like effects in behavioral procedures commonly used to characterize benzodiazepines and other drugs in squirrel monkeys.     

Similar effects of buspirone and chlordiazepoxide on a fixed interval schedule with long-term, low-dose administration

Buspirone is a novel anxiolytic which does not share the muscle relaxant, anticonvulsant and sedative properties of classical anxiolytics such as the benzodiazepines. Its effects on behavioural tests of anxiolytic action generally match those of classical anxiolytics provided a low dose is used. However, in a previous experiment, buspirone appeared to affect fixed interval responding in a way which differed qualitatively as well as quantitatively from the classical anxiolytic chlordiazepoxide. It takes as much as 2 weeks for the clinical effects of anxiolytics to develop, during which time the side effects of benzodiazepines undergo tolerance. We, therefore, decided to compare long-term pre-administration (60 days, three injections/day) of buspirone and chlordiazepoxide on learning of a fixed interval 60-s schedule. The doses were based on previous acute dose-response tests of hippocampal theta rhythm in freely moving animals. Buspirone (0.1 mg/ kg i.p.) and chlordiazepoxide (0.4 mg/kg i.p.) produced similar increases in responding, especially in the middle of acquisition of the fixed interval schedule. Consistent with our acute electrophysiological tests, the effects of 0.4 mg/kg chlordiazepoxide were somewhat larger than those of 0.1 mg/kg buspirone. These results suggest that the acute effects of buspirone, but probably not chlordiazepoxide, on fixed interval responding are contaminated by side effects which do not seriously affect the results with long-term administration. The effects of both novel and classical anxiolytics on control of hippocampal theta rhythm appear to predict the magnitude of their common anxiolytic effects and to be unrelated to their different side effects.     

Buspirone, chlordiazepoxide and diazepam effects in a zebrafish model of anxiety

Zebrafish are becoming more widely used to study neurobehavioral pharmacology. We have developed a method to assess novel environment diving behavior of zebrafish as a model of stress response and anxiolytic drug effects. In a novel tank, zebrafish dwell in the bottom of the tank initially and then increase their swimming exploration to higher levels over time. We previously found that nicotine, which has anxiolytic effects in rodents and humans, significantly lessens the novel tank diving response in zebrafish. The specificity of the diving effect was validated with a novel vs. non-novel test tank. The novel tank diving response of zebrafish was tested when given three anxiolytic drugs from two different chemical and pharmacological classes: buspirone, chlordiazepoxide and diazepam. When the test tank was novel the diving response was clearly seen whereas it was significantly reduced when the test tank was not novel. Buspirone, a serotonergic (5HT_1A receptor agonist) anxiolytic drug with some D₂ dopaminergic effect, had a pronounced anxiolytic-like effect in the zebrafish diving model at doses that did not have sedative effects. In contrast, chlordiazepoxide, a benzodiazepine anxiolytic drug, which is an effective agonist at GABA-A receptors, did not produce signs of anxiolysis in zebrafish over a broad dose range up to those that caused sedation. Diazepam another benzodiazepine anxiolytic drug did produce an anxiolytic effect at doses that did not cause sedation. The zebrafish novel tank diving task can be useful in discriminating anxiolytic drugs of several classes (serotonergic, benzodiazepines and nicotinic).     

Pharmacological evaluation of a modified conflict procedure: punished drinking in non-water-deprived rats

Rationale: Conflict procedures used to detect anxiolytic-like activity of drugs often rely on maintaining strict schedules of water or food availability. It is ethically and practically desirable to reduce such states of deprivation in animal testing. Objective: The purpose of the present experiment was to develop and pharmacologically characterize a conflict drinking procedure that did not require the use of water-deprived animals. Methods: Rats were tested during daily sessions with alternating unpunished drinking (no tone: lick=sucrose solution) and signaled punished drinking (tone: lick=sucrose+shock) components, and developed individual steady baselines over a brief training period (approximately 3–4 weeks). The drugs tested i.p. were the positive allosteric modulators of γ-amino butyric acid_A (GABA)_A receptors, diazepam (0.03–30 mg/kg), chlordiazepoxide (0.03–30 mg/kg), lorazepam (0.03–10 mg/kg), zolpidem (0.3–10 mg/kg), pentobarbital (1–30 mg/kg), pregnanolone (1–30 mg/kg), and bretazenil (0.03– 10 mg/kg); the 5-hydroxy tryptamine_1A (HT)_1A-mediated anxiolytics, buspirone (1–10 mg/kg) and ipsapirone (1–17 mg/kg); and the negative controls d-amphetamine (0.3–3 mg/kg), haloperidol (0.01–0.3 mg/kg), morphine (0.3–17 mg/kg), and imipramine (0.3–30 mg/kg). Results: The experimental procedure was sensitive to increases in punished drinking by the GABA_A-positive modulators, consistent with their known or putative anxiolytic activity. Further, the 5-HT_1A-mediated anxiolytics increased punished drinking, although to a lesser extent and over a more narrow dose range than did the GABAergic drugs. In contrast, d-amphetamine, haloperidol, morphine, and imipramine failed to increase punished drinking up to doses that decreased unpunished drinking. Conclusions: The present results indicate that water deprivation is not a necessary condition to engender drinking conflict behavior or to obtain pharmacological effects similar to those obtained with other classical conflict procedures. Received: 23 November 1998 / Final version: 15 March 1999     

Combined discriminative stimulus effects of midazolam with other positive GABA<Subscript>A</Subscript> modulators and GABA<Subscript>A</Subscript> receptor agonists in rhesus monkeys

Rationale Interactions among compounds at GABA_A receptors might have important implications for the therapeutic and other effects of positive GABA_A modulators (e.g. benzodiazepines).Objectives This study examined whether a midazolam discriminative stimulus is modified by GABA_A agonists that act at sites other than benzodiazepine sites.Methods Rhesus monkeys discriminating midazolam (0.32 mg/kg SC) received direct-acting GABA_A receptor agonists (e.g. muscimol and gaboxadol), an indirect-acting GABA_A receptor agonist (progabide), ethanol, another benzodiazepine (triazolam), a barbiturate (pentobarbital), or a neuroactive steroid (pregnanolone) alone and in combination with midazolam.Results When administered alone, triazolam (0.1 mg/kg), pentobarbital (17.8 mg/kg) and pregnanolone (5.6 mg/kg) occasioned high levels of midazolam lever responding, ethanol (1–3 g/kg) occasioned intermediate levels of midazolam lever responding, and muscimol (0.32–1 mg/kg), gaboxadol (3.2–10 mg/kg) and progabide (10–32 mg/kg) occasioned low levels of midazolam lever responding. When combined with less-than-fully effective doses of midazolam, progabide (32 mg/kg) and ethanol (1 g/kg), but not muscimol and gaboxadol, enhanced the midazolam discriminative stimulus. Triazolam, pregnanolone and pentobarbital increased the potency of midazolam to occasion midazolam lever responding and the effects of these combinations were additive.Conclusions Direct-acting GABA_A receptor agonists are qualitatively different from positive GABA_A modulators in rhesus monkeys trained to discriminate midazolam. Although GABA_A receptor agonists and modulators can enhance the actions of benzodiazepines at the GABA_A receptor complex, the same drugs do not necessarily modify the discriminative stimulus effects of benzodiazepines. These results underscore the importance of the mechanism by which drugs alter Cl^– flux at the GABA_A receptor complex as a determinant not only of drug action but also of drug interaction and whether any particular drug enhances the behavioral effects of a benzodiazepine.