The role of M1 muscarinic receptor agonism of N-desmethylclozapine in the unique clinical effects of clozapine.

RATIONALE: Clozapine is a unique antipsychotic, with efficacy against positive symptoms in treatment-resistant schizophrenic patients, and the ability to improve cognition and treat the negative symptoms characteristic of this disease. Despite its unique clinical actions, no specific molecular mechanism responsible for these actions has yet been described. OBJECTIVES AND METHODS: To comprehensively profile a large library of neuropsychiatric drugs, including most antipsychotics, at human monoamine receptors using R-SAT, an in vitro functional assay. RESULTS: Profiling revealed that N-desmethylclozapine (NDMC), the principal metabolite of clozapine, but not clozapine itself, is a potent and efficacious muscarinic receptor agonist, a molecular property not shared by any other antipsychotic. To further explore the role of NDMC muscarinic receptor agonist properties in mediating the physiological actions of clozapine, systemically administered NDMC was found to stimulate the phosphorylation of mitogen-activated protein kinase (MAP kinase) in mouse CA1 hippocampal neurons, an effect that was blocked by scopolamine, confirming central M1 muscarinic receptor agonist activity in vivo. Lastly, an analysis of clozapine and NDMC serum levels in schizophrenic patients indicated that high NDMC/clozapine ratios better predicted improvement in cognitive functioning and quality of life than the levels of either compound alone. CONCLUSIONS: The muscarinic receptor agonist activities of NDMC are unique among antipsychotics, and provide a possible molecular basis for the superior clinical effects of clozapine pharmacotherapy.

Assessment of learning and memory using the autoshaping of operant responding in mice.

This unit describes the use of an automated procedure for developing an operant response ("autoshaping") in the mouse. The method has applications in the study of the acquisition of behavior (learning) as well as for the assessment of memory or retention of that task.

5-hydroxytryptamine2A receptor inverse agonists as antipsychotics.

We have used a cell-based functional assay to define the pharmacological profiles of a wide range of central nervous system active compounds as agonists, competitive antagonists, and inverse agonists at almost all known monoaminergic G-protein-coupled receptor (GPCR) subtypes. Detailed profiling of 40 antipsychotics confirmed that as expected, most of these agents are potent competitive antagonists of the dopamine D2 receptor. Surprisingly, this analysis also revealed that most are potent and fully efficacious 5-hydroxytryptamine (5-HT)2A receptor inverse agonists. No other molecular property was shared as universally by this class of compounds. Furthermore, comparisons of receptor potencies revealed that antipsychotics with the highest extrapyramidal side effects (EPS) liability are significantly more potent at D2 receptors, the EPS-sparing atypical agents had relatively higher potencies at 5-HT2A receptors, while three were significantly more potent at 5-HT2A receptors. Functional high-throughput screening of a diverse chemical library identified 530 ligands with inverse agonist activity at 5-HT2A receptors, including several series of compounds related to known antipsychotics, as well as a number of novel chemistries. An analog of one of the novel chemical series, AC-90179, was pharmacologically profiled against the remaining monoaminergic GPCRs and found to be a highly selective 5-HT2A receptor inverse agonist. The behavioral pharmacology of AC-90179 is characteristic of an atypical antipsychotic agent.

Behavioral characterization of Co 134444 (3alpha-hydroxy-21-(1'-imidazolyl)-3beta-methoxymethyl-5alpha- pregnan-20-one), a novel sedative-hypnotic neuroactive steroid.

RATIONALE: Neuroactive steroids have been shown to exhibit a wide range of behavioral activities that are similar but not identical to those of benzodiazepines. These activities include anticonvulsant, anxiolytic and sedative-hypnotic effects. OBJECTIVE: The purpose of the present study was to characterize Co 134444 (3alpha-hydroxy-21-(1'-imidazolyl)-3 -methoxymethyl-5alpha-pregnan-20-one), a novel sedative-hypnotic neuroactive steroid, in a variety of behavioral procedures. METHODS: Anticonvulsant effects were determined by the ability to protect against pentylenetetrazol- and maximal electroshock-induced seizures in mice and rats. Anxiolytic-like effects were determined using a punished drinking procedure in rats. Ataxic effects were determined using a horizontal wire procedure in mice and a rotorod procedure in mice and rats. The discriminative stimulus effects were evaluated in rats trained to discriminate pregnanolone from vehicle. RESULTS: Co 134444 exhibited oral anticonvulsant activity against pentylenetetrazol and maximal electroshock with ED50s of 9.8 and 20.6 mg/kg, respectively, in mice and 23.6 and 25.3 mg/kg, respectively, in rats. Anxiolytic-like efficacy was observed at a dose as low as 3.0 mg/kg, PO, in rats. Ataxic effects were observed with rapid onset and short duration. TD50s were 17.4 and 21.2 mg/kg orally in mice in the horizontal wire and rotorod procedures, respectively, and 39.0 mg/kg in rats using the rotorod. Co 134444 completely substituted for pregnanolone as a discriminative stimulus with little effect on response rate. CONCLUSIONS: Co 134444 exhibits a wide variety of behavioral effects; however, its rapid onset and short duration are consistent with its potential use as a sedative-hypnotic drug.

Characterization of the anxiolytic properties of a novel neuroactive steroid, Co 2-6749 (GMA-839; WAY-141839; 3alpha, 21-dihydroxy-3beta-trifluoromethyl-19-nor-5beta-pregnan-20-one), a selective modulator of gamma-aminobutyric acid(A) receptors.

The purpose of this study was to evaluate the effects of a novel neuroactive steroid, Co 2-6749 (GMA-839; WAY-141839; 3alpha, 21-dihydroxy-3beta-trifluoromethyl-19-nor-5beta-pregnan-20-one), on gamma-aminobutyric acid(A) receptors in vitro and to define its anxiolytic-like effects and side effect profile in vivo. Co 2-6749 fully inhibited [(35)S]t-butylbicyclophosphorothionate binding in rat brain cortical membranes with an IC(50) value of 230 nM and in human gamma-aminobutyric acid(A) receptor subunit combinations of alpha1beta2gamma2L, alpha2beta2gamma2L, alpha3beta2gamma2L, alpha4beta3gamma2L, alpha5beta2gamma2L, and alpha6beta3gamma2L receptors (IC(50) values of 200, 200, 96, 2300, 210, and 2000 nM). Rats were trained in a Geller-Seifter operant conflict paradigm. Co 2-6749 caused a dose-related increase in punished responding with a minimum effective dose of 1.6 mg/kg, p.o., a wide therapeutic index relative to a decrease in unpunished responding and relative to ataxia, and no tolerance. Additionally, ethanol caused less than a 2-fold shift to the left in the dose-response function of Co 2-6749 in the rotorod procedure in rats. In a pigeon conflict paradigm, punished responding was maximally increased to 784% of vehicle control by 30 mg/kg, p.o., with a 2-h duration and no effect on unpunished responding at this dose. Similarly, punished responding in squirrel monkeys was maximally increased to 1774% of control by 10 mg/kg, p.o., with no effect on unpunished responding at this dose. With robust anxiolytic-like activity across species, a large separation between anxiolytic-like effects and sedation/ataxia, a minimal interaction with ethanol, a lack of tolerance, and apparent oral bioavailability, Co 2-6749 makes an ideal candidate for development as a novel anxiolytic drug.

Neuroactive steroids attenuate cocaine-induced sucrose intake in rats, but not cocaine-induced hyperactivity in mice.

RATIONALE: Neuroactive steroids, including the potent anticonvulsants ganaxolone (3alpha-hydroxy-3beta-methyl-5alpha-pregnan-20-one) and Co 2-1068 (3beta-(4acetyl-phenyl)ethynyl-3alpha,21-dihydroxy-5beta+ ++-20-one-21-hemisuccinate), have recently been shown to protect against cocaine-induced seizures. OBJECTIVES: The purpose of the present experiments was to determine whether ganaxolone and Co 2-1068 attenuate acute behavioral effects of cocaine unrelated to seizures. METHODS: In the first experiment, the locomotor effects of Co 2-1068 (10-100 mg/ kg), pentobarbital (10-100 mg/kg) and haloperidol (0.03-0.3 mg/kg), alone or in combination with cocaine (5.6-30 mg/kg), were determined in mice. In the second experiment, the effects on sucrose intake of ganaxolone (4-16 mg/kg), Co 2-1068 (8-64 mg/kg), pentobarbital (4-32 mg/kg), and haloperidol (0.04-0.4 mg/kg), alone or in combination with cocaine (4-16 mg/kg), were determined in rats. RESULTS: Cocaine caused a dose-related increase in locomotor activity in mice, whereas Co 2-1068, pentobarbital and haloperidol caused dose-related decreases. The dopamine antagonist haloperidol, at a dose that had no effect on activity by itself, but not Co 2-1068 or pentobarbital, attenuated the cocaine-induced increase in locomotor activity. Cocaine, ganaxolone, Co 2-1068, and haloperidol produced dose-related decreases in sucrose intake in rats; the effects of pentobarbital on sucrose intake were variable. As with locomotor effects, haloperidol attenuated the cocaine-induced decrease in sucrose intake. In addition, cocaine-induced decreases in sucrose intake were attenuated by ganaxolone and Co 2-1068. Pentobarbital had no statistically significant effect on the cocaine dose-response function. CONCLUSIONS: These results suggest that the interaction of neuroactive steroids with cocaine extends to pharmacologic actions beyond anticonvulsant efficacy, but that the blockade of behavioral effects of cocaine by neuroactive steroids does not apply to all acute behaviors.

Effects of benzodiazepine receptor ligands and ethanol in rats trained to discriminate pregnanolone.

Although GABA(A) receptor positive modulators share many behavioral effects, subtle differences have been detected among their discriminative stimulus effects. The purpose of the present study was to determine the extent of shared discriminative stimulus effects of pregnanolone with various benzodiazepine receptor ligands and with ethanol. Naive male Sprague-Dawley rats were trained to discriminate the endogenous neuroactive steroid pregnanolone (5.6 or 8.0 mg/kg) from vehicle. The benzodiazepine receptor agonists, triazolam and lorazepam, the benzodiazepine receptor partial agonist, bretazenil, the benzodiazepine1 (BZ1) receptor subtype selective agonists, zolpidem and zaleplon and ethanol were tested. Triazolam, lorazepam and bretazenil substituted for pregnanolone. Lorazepam, but not triazolam or bretazenil, decreased response rates at the highest dose tested. Zaleplon completely substituted for pregnanolone with no effect on response rates. Zolpidem substituted for pregnanolone only at a dose that severely disrupted response rates. Ethanol partially substituted for pregnanolone and decreased response rates. The results are consistent with GABA(A) receptor mediation of the discriminative stimulus effects of pregnanolone. The effects on response rates suggest subtle differentiation among the GABA(A) receptor-mediated cues.

Response-rate suppression in operant paradigm as predictor of soporific potency in rats and identification of three novel sedative-hypnotic neuroactive steroids.

Novel neuroactive steroids were evaluated for their effects on operant responding, rotorod motor performance, and electroencephalogram recording in rats. Co 134444, Co 177843, and Co 127501 were compared with the prototypical gamma-aminobutyric acid(A)-positive allosteric modulators triazolam, zolpidem, pentobarbital, pregnanolone, and CCD 3693. Each of the compounds produced a dose-related decrease in response rates under a variable-interval 2-min schedule of positive reinforcement in an operant paradigm. In addition, all compounds produced a dose-related increase in ataxia and significant increases in nonrapid eye movement sleep in this experiment or have been previously reported to do so. Co 134444, Co 177843, and Co 127501 increased nonrapid eye movement sleep at doses that had no effect on rapid eye movement sleep. All of the compounds were more potent at decreasing operant responding than they were at increasing ataxia. Furthermore, the potency of compounds to produce response-rate suppression in an operant paradigm appeared to be a better predictor of soporific potency than did potency in the rotorod assay. The screening for sedative-hypnotic activity resulted in the identification of the novel orally active neuroactive steroids Co 134444, Co 177843, and Co 127501.

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 gamma-amino butyric acidA (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 tryptamine1A (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 GABAA-positive modulators, consistent with their known or putative anxiolytic activity. Further, the 5-HT1A-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.

Interaction of ethanol with excitatory amino acid receptor antagonists in mice.

The purpose of the present study was to determine whether the motor impairment (myorelaxation/ataxia) induced by excitatory amino acid receptor antagonists was exaggerated by pretreatment with ethanol. The results were compared with those of gamma-aminobutyric acid(A) (GABA(A)) receptor positive modulators alone and in combination with ethanol. The excitatory amino acid receptor antagonists, dizocilpine [(+)-MK-801; (5R,1OS)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten+ ++-5,10-imine], (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), LY 326325 [(-)-(3S,4aR,6R,8R)-6-[2-(1(2)H-tetrazol-5-yl)-ethyl]-dec ahydroisoquinaline-3-carboxylic acid], LY 300164 [7,8-methylenedioxy-1-(4-aminophenyl)-4-methyl-3-acetyl-4,5-dihydro-2,3- benzodiazepine], and ACEA 1011 (5-chloro-7-trifluoromethyl-1,4-dihydro-2,3-quinoxalinedione) produced dose-dependent myorelaxation/ataxia in mice as determined using the horizontal wire assay. Their behaviorally toxic doses (TD(50)s) were 0.41, 5.8, 33.0, 5.9, and 31.0 mg/kg, respectively, when administered alone i.p. In the presence of a sub-ataxic dose of ethanol (1.5 g/kg, i.p.), the TD(50)s of the excitatory amino acid antagonists were 0.13, 1.8, 10.4, 1.3, and 14.0 mg/kg, respectively. Similarly, the GABA(A) receptor positive modulators, pregnanolone, chlordiazepoxide, and pentobarbital exhibited TD(50)s of 20.8, 4.6, and 29.7 mg/kg, respectively, when administered alone and 2.7, 0.3, and 11.4 mg/kg, respectively, when administered in the presence of ethanol. Thus, similar to the GABA(A) receptor positive modulators, excitatory amino acid receptor antagonists exhibit the propensity to interact with ethanol and to have their motor side-effects exaggerated.

Positive allosteric modulators of the GABA(A) receptor: differential interaction of benzodiazepines and neuroactive steroids with ethanol.

Endogenous pregnane steroids, such as allopregnanolone (3alpha-hydroxy-5alpha-pregnan-20-one; 3alpha, 5alpha-P) and pregnanolone (3alpha-hydroxy-5beta-pregnan-20-one; 3alpha,5beta-P), allosterically modulate GABA(A) receptor function and exhibit behavioral effects similar to benzodiazepines, though acting at a distinct recognition site. Inasmuch as some positive allosteric modulators of GABA(A) receptor function exhibit profound interactions with ethanol, the effects of 3alpha,5alpha-P and 3alpha,5beta-P were compared to those of two benzodiazepines, triazolam and diazepam, on the motor function of mice and rats when administered either alone or in combination with ethanol. All four test compounds exhibited dose-related impairment of motor function in the horizontal wire task in mice and the rotorod task in rats. Ethanol caused a marked enhancement of triazolamand diazepam-induced motor impairment. In contrast, ethanol enhanced to a lesser extent the motor impairment induced by both neurosteroids in mice and not at all in rats. All four compounds increased ethanol-induced behavioral sleep time in mice, although the benzodiazepines did so at a much smaller fraction of their ataxic doses as compared to the neurosteroids. As one of the undesired side-effects of therapeutic use of benzodiazepines is their interaction with ethanol, development of neuroactive steroids as drugs may offer therapeutic advantages.

An automated learning and memory model in mice: pharmacological and behavioral evaluation of an autoshaped response.

The purpose of the present experiments was to develop and validate pharmacologically an automated, relatively rapid, and reproducible behavioral model of learning and memory using an autoshaping procedure in mice. Nose-poke responses into a recessed area were differentiated by response-dependent reinforcement during two identical consecutive daily sessions. Performance during the first session was considered to be a measure of acquisition and that during the second session a measure of retention. Sensitivity to procedural manipulation, as well as an index of learning under these conditions, was demonstrated, for example, by a decrease in response rate when nose-poke responses did not produce a reinforcer. The sensitivity of the paradigm to pharmacological intervention was examined after drug administration before the first session. Scopolamine (0.1-10.0 mg/kg) had no effect on acquisition but caused a significant dose-related impairment of retention. Dizocilpine (0.01-1.0 mg/kg) impaired both acquisition and retention performance. 8-Hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT; 0.1-1.0 mg/kg) disrupted behavior in general, but failed to have a selective effect on acquisition or retention. Linopirdine (0.1-1.0 mg/kg) showed only a weak enhancement of acquisition, whereas 4-aminopyridine (4-AP; 0.1-1.0 mg/kg) significantly facilitated acquisition. This paradigm offers the potential for a rapid, objective, and reliable indication of whether a drug will affect the acquisition or retention of a positively reinforced response in mice and could be a useful supplement to existing procedures.

Synthesis of 7,8-(methylenedioxy)-1-phenyl-3,5-dihydro-4H-2, 3-benzodiazepin-4-ones as novel and potent noncompetitive AMPA receptor antagonists.

A group of 7,8-(methylenedioxy)-1-phenyl-3,5-dihydro-4H-2, 3-benzodiazepin-4-ones was synthesized and assayed for antagonism of rat brain alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors expressed in Xenopus oocytes. The benzodiazepinones inhibited AMPA-activated membrane current responses in a manner consistent with noncompetitive, allosteric inhibition of the receptor-channel complex. The most potent compound in the series was 1-(4-aminophenyl)-7,8-(methylenedioxy)-3,5-dihydro-4H-2, 3-benzodiazepin-4-one (6), which had an IC50 of 2.7 microM. For comparison, the reference compound GYKI 52466 (2) had an IC50 of 6.9 microM. Compound 6 also had potent anticonvulsant activity in a mouse maximum electroshock-induced seizure (MES) assay: the ED50 was 2.8 mg/kg iv, whereas the ED50 for GYKI 52466 was 4.6 mg/kg iv. In contrast to a previous report, the 7,8-dimethoxy analogue of 6 was a low-potency AMPA antagonist (IC50 >100 microM) and weak anticonvulsant (ED50 >10 mg/kg iv). The benzodiazepinones described herein are potent noncompetitive AMPA receptor antagonists that could have therapeutic potential as anticonvulsants and neuroprotectants.

Effects of AMPA receptor antagonists on dopamine-mediated behaviors in mice.

Current data indicate that dopaminergic and glutamatergic neurotransmitter systems interact. The role of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) glutamate receptor subtypes in modulating dopamine neurotransmission, however, remains unclear. The noncompetitive AMPA antagonists, GYKI 52466 (5-40 mg/kg) and LY300164 (1-6 mg/kg), and the competitive AMPA antagonists, LY326325 (5-80 mg/kg) and NBQX (10-80 mg/kg), were compared to the dopamine antagonist, haloperidol (0.03-1.0 mg/kg), for their ability to inhibit dopamine-mediated behaviors after i.p. administration in mice. The behavioral paradigms included amphetamine- or dizocilpine-induced hyperactivity, amphetamine-induced stereotyped sniffing, and apomorphine-induced climbing and stereotyped sniffing. All four AMPA antagonists and haloperidol attenuated amphetamine- and dizocilpine-induced hyperactivity and decreased spontaneous locomotion. Haloperidol and GYKI 52466 were more potent against amphetamine than against dizocilpine. In contrast, LY326325 was more potent against dizocilpine than against amphetamine. The hyperactivity decreases by LY300164 and NBQX were most likely due to non-specific effects on motor behavior. The AMPA antagonists and haloperidol also attenuated amphetamine- induced stereotypy. Unlike haloperidol, however, GYKI 52466, LY300164, and NBQX failed to attenuate apomorphine-induced climbing and stereotyped sniffing. LY326325, on the other hand, attenuated apomorphine-induced stereotypy, but not climbing. These results indicate that AMPA receptor antagonists can attenuate the behavioral effects of drugs, such as amphetamine and dizocilpine, that increase dopamine neurotransmission. However, the behavioral effects of the direct dopamine agonist apomorphine are not consistently attenuated by AMPA antagonists. The competitive AMPA receptor antagonist LY326325 appears to have a profile distinct from both haloperidol and the other AMPA antagonists tested.

Effects of AMPA receptor positive modulators on amphetamine- and dizocilpine-induced locomotion.

Two allosteric alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor positive modulators, 1-(1,3-benzodioxol-5-ylcarbonyl)piperidine (1-BCP) and 1-(quinoxalin-6-ylcarbonyl)piperidine (CX516), and the antipsychotic drug, haloperidol, were tested for their ability to inhibit hyperactivity induced by amphetamine and dizocilpine in mice. Haloperidol (0.03-1.0 mg/kg) and 1-BCP (20.0-120.0 mg/kg) attenuated hyperactivity induced by both amphetamine and dizocilpine, with higher potency against amphetamine. CX516 (30.0-170.0 mg/kg), however, failed to attenuate amphetamine- and dizocilpine-induced hyperactivity up to a dose which decreased spontaneous locomotor activity. These results indicate that AMPA receptor positive modulators may not be uniform with regard to their effects on dopamine-mediated behaviors and their antipsychotic potential.

Discriminative stimulus effects of the endogenous neuroactive steroid pregnanolone.

Naive male Sprague-Dawley rats were trained to discriminate the endogenous neuroactive steroid pregnanolone (5.6 mg/kg) from saline. Three positive modulators of the GABA(A) receptor complex substituted for pregnanolone: the neuroactive steroid allopregnanolone (1.0-10.0 mg/kg), the barbiturate pentobarbital (3.0-17.0 mg/kg), and the benzodiazepine diazepam (0.3-3.0 mg/kg). In contrast, buspirone, a 5-HT1A-mediated anxiolytic, failed to substitute up to rate-suppressing doses (1.0-5.6 mg/kg). The present experiment demonstrated the ability of an endogenous neuroactive steroid to function as a discriminative stimulus. Moreover, these results suggest that the discriminative stimulus effects of pregnanolone are mediated via positive modulation of GABA(A) receptors.

Behavioral effects of the dopamine autoreceptor agonist PD 128483 alone and in combination with cocaine.

Many of the behavioral effects of cocaine are thought to be mediated via dopamine (DA) reuptake inhibition and a subsequent increase in DA activity. As a DA agonist with selectivity for autoreceptors, PD 128483 has behavioral effects consistent with decreased DA neurotransmission and might be expected to attenuate the behavioral effects of cocaine. The present study was designed to evaluate PD 128483 ((+)-4,5,5a,6,7,8-hexahydro-6-methyl-thiazolo[4,5-f]quinoline-2-amine) alone and in combination with cocaine in three behavioral paradigms. One group of rhesus monkeys (n = 7) was allowed to self-administer cocaine (0.03 or 0.1 mg/kg/injection i.v., fixed-ratio 10, 2 hr/day) in baseline sessions. When behavior was stable, PD 128483 (0.01-0.3 mg/kg/injection i.v.) was made available for self-administration, and failed to maintain responding in three of four monkeys tested. PD 128483 (1.0-10.0 mg/kg i.m.) decreased cocaine self-administration in all four monkeys tested when given before cocaine availability. Another group of monkeys (n = 4) was trained to discriminate cocaine (0.2 or 0.4 mg/kg i.m.) from saline in a two-lever, food-reinforced drug discrimination paradigm. High doses of PD 128483 (3.2-12.8 mg/kg i.m.) substituted for cocaine as a discriminative stimulus in three of four monkeys. Administered in combination with cocaine, PD 128483 had little effect on the cocaine discriminative stimulus up to doses which decreased response rate. PD 128483 (1.0-2.0 mg/kg i.p.) decreased milk intake in rats (n = 9) and, when given in combination with cocaine, shifted the cocaine dose-response function (4.0-16.0 mg/kg i.p.) to the left.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of the discriminative stimulus effects of the novel sedative-hypnotic CL 284,846.

CL 284,846, N-[3-(3-cyanopyrazolo[1, 5-a]pyrimidin-7-yl)phenyl)]-N- ethylacetamide, is a novel non-benzodiazepine sedative-hypnotic with benzodiazepine-like sedative effects, but with less apparent liability for accompanying undesired side effects. In an effort to further characterize its pharmacological activity, CL 284,846 (3.0 mg/kg, IP, 30 min pretreatment) was established as a discriminative stimulus (DS) in rats (n = 7). CL 284,846 (0.3-10.0 mg/kg) showed a dose-related increase in drug-appropriate responding up to the training dose and a dose-related decrease in response rate. The benzodiazepine agonist triazolam (0.1-1.0 mg/kg), the benzodiazepine partial agonist Ro 17-1812 (0.3-3.0 mg/kg) and the triazolopyridazine CL 218,872 (1.0-3.0 mg/kg) substituted for CL 284,846 in all rats, whereas the imidazopyridines zolpidem (3.0-10.0 mg/kg) and alpidem (10.0-30.0 mg/kg), the benzodiazepine partial agonist bretazenil (0.03-10.0 mg/kg) and the novel putative anxiolytic CL 273,547 (10.0-56.0 mg/kg) substituted in most, but not all, rats. Ro 17-1812, bretazenil, and CL 218,872 had no effect on response rate while the other drugs showed a concomitant decrease in rate. The 5-HT1A agonist buspirone (1.0-10.0 mg/kg) and the barbiturate pentobarbital (3.0-17.0 mg/kg) failed to substitute for CL 284,846 up to rate-decreasing doses. The benzodiazepine antagonist flumazenil (3.0-10.0 mg/kg) blocked the DS effects of CL 284,846 in most rats with no effect on response rate. Taken together, these results suggest that the DS effects of CL 284,846 are mediated via benzodiazepine receptors; however, the DS profile of CL 284,846 remains distinct from both benzodiazepine and non-benzodiazepine sedative-hypnotic drugs.

Effects of cholecystokinin antagonists on the discriminative stimulus effects of cocaine in rats and monkeys.

Cholecystokinin (CCK) has been implicated as a modulator of dopamine (DA) neurotransmission in the mesolimbic DA pathway, a primary pathway implicated in the effects of cocaine related to its abuse. The present experiment was designed to examine whether an antagonist that acts at either CCKA or CCKB receptors can modify the discriminative stimulus effects of cocaine in animals. Rats (N = 9) and rhesus monkeys (N = 3) were trained in two-lever drug discrimination paradigms to discriminate cocaine from saline. Lever pressing was maintained by food (all rats, two monkeys) or shock avoidance (one monkey). In rats, the CCKA antagonist MK-329 (1.0-32 mg/kg, i.p.) or the CCKB antagonist CI-988 (1.0-32 mg/kg, i.p.) were administered 30 min before determination of cocaine dose-response functions using a cumulative dosing method. In monkeys, CI-988 (8.0-32 mg/kg, i.m.) was administered 30, 60 or 120 min before the training dose of cocaine. In both species, cocaine produced dose-related increases in the percentage of responses emitted on the cocaine-appropriate lever. In rats, MK-329 shifted the cocaine dose-response function to the right in a dose-related manner. In contrast, CI-988 did not systematically alter the effects of cocaine in either rats or monkeys. Since drug discrimination serves as an animal model of the subjective effects of drugs in humans, these results suggest that CCKA antagonists may decrease the subjective effects of cocaine. It seems unlikely that CCKB antagonists will alter the subjective effects of cocaine.

Evaluation of the reinforcing and discriminative stimulus effects of cocaine in combination with (+)-AJ76 or clozapine.

Because self-administration and discrimination of a drug by animals correlate with its abuse and subjective effects in humans, interventions that modify the reinforcing and discriminative stimulus effects of the drug may be useful in the treatment of its abuse. The present study was designed to evaluate the effects of the putative dopamine autoreceptor antagonist (+)-AJ76 (AJ) or the atypical antipsychotic clozapine (CLZ) on the reinforcing and discriminative stimulus effects of cocaine in monkeys. One group of rhesus monkeys (n = 6) was allowed to self-administer cocaine (0.03 or 0.1 mg/kg/injection i.v. fixed-ratio 10, 2 hr/day). A second group of monkeys (n = 5) was trained to discriminate cocaine (0.2 or 0.4 mg/kg i.m., 10 min presession) from saline in a two lever, food-reinforced, drug discrimination paradigm. When behavior was stable, AJ or CLZ was administered i.m., 15 or 30 min presession. Intermediate doses of both compounds (1.0-3.0 mg/kg of AJ; 0.3-1.0 mg/kg of CLZ) increased cocaine self-administration, while responding remained evenly distributed over the session. A higher dose of CLZ decreased cocaine self-administration in an apparently nonspecific manner. When combined with saline, partial substitution for cocaine was seen in one of three monkeys with AJ and in none with CLZ. In combination with the training dose of cocaine in the discrimination experiment, both AJ and CLZ decreased drug appropriate responding by at least 50% in two of four monkeys, but had little or no effect in the other monkeys up to doses that completely suppressed lever pressing (6.4 mg/kg of AJ; 3.2 mg/kg of CLZ). Taken together, the present findings suggest that any blockade of the reinforcing and discriminative stimulus effects of cocaine by AJ and CLZ was, at best, partial. Furthermore, the stimulant effects of AJ observed in rats were not prominent in monkeys.