The psychoactive plant cannabinoid, Delta9-tetrahydrocannabinol, is antagonized by Delta8- and Delta9-tetrahydrocannabivarin in mice in vivo.

BACKGROUND AND PURPOSE: To follow up in vitro evidence that Delta(9)-tetrahydrocannabivarin extracted from cannabis (eDelta(9)-THCV) is a CB(1) receptor antagonist by establishing whether synthetic Delta(9)-tetrahydrocannabivarin (O-4394) and Delta(8)-tetrahydrocannabivarin (O-4395) behave as CB(1) antagonists in vivo. EXPERIMENTAL APPROACH: O-4394 and O-4395 were compared with eDelta(9)-THCV as displacers of [(3)H]-CP55940 from specific CB(1) binding sites on mouse brain membranes and as antagonists of CP55940 in [(35)S]GTPgammaS binding assays performed with mouse brain membranes and of R-(+)-WIN55212 in mouse isolated vasa deferentia. Their ability to antagonize in vivo effects of 3 or 10 mg kg(-1) (i.v.) Delta(9)-tetrahydrocannabinol in mice was then investigated. KEY RESULTS: O-4394 and O-4395 exhibited similar potencies to eDelta(9)-THCV as displacers of [(3)H]-CP55940 (K (i)=46.6 and 64.4 nM, respectively) and as antagonists of CP55940 in the [(35)S]GTPgammaS binding assay (apparent K (B)=82.1 and 125.9 nM, respectively) and R-(+)-WIN55212 in the vas deferens (apparent K (B)=4.8 and 3.9 nM respectively). At i.v. doses of 0.1, 0.3, 1.0 and/or 3 mg kg(-1) O-4394 and O-4395 attenuated Delta(9)-tetrahydrocannabinol-induced anti-nociception (tail-flick test) and hypothermia (rectal temperature). O-4395 but not O-4394 also antagonized Delta(9)-tetrahydrocannabinol-induced ring immobility. By themselves, O-4395 and O-4394 induced ring immobility at 3 or 10 mg kg(-1) (i.v.) and antinociception at doses above 10 mg kg(-1) (i.v.). O-4395 also induced hypothermia at 3 mg kg(-1) (i.v.) and above. CONCLUSIONS AND IMPLICATIONS: O-4394 and O-4395 exhibit similar in vitro potencies to eDelta(9)-THCV as CB(1) receptor ligands and as antagonists of cannabinoid receptor agonists and can antagonize Delta(9)-tetrahydrocannabinol in vivo.

Interactions between THC and cannabidiol in mouse models of cannabinoid activity.

RATIONALE: Interest persists in characterizing potential interactions between Delta(9)-tetrahydocannabinol (THC) and other marijuana constituents such as cannabidiol (CBD). Such interactions may have important implications for understanding the long-term health consequences of chronic marijuana use as well as for attempts to develop therapeutic uses for THC and other CB(1) agonists. OBJECTIVES: We investigated whether CBD may modulate the pharmacological effects of intravenously administered THC or inhaled marijuana smoke on hypoactivity, antinociception, catalepsy, and hypothermia, the well characterized models of cannabinoid activity. RESULTS: Intravenously administered CBD possessed very little activity on its own and, at a dose equal to a maximally effective dose of THC (3 mg/kg), failed to alter THC's effects on any measure. However, higher doses of CBD (ED(50)=7.4 mg/kg) dose-dependently potentiated the antinociceptive effects of a low dose of THC (0.3 mg/kg). Pretreatment with 30 mg/kg CBD, but not 3 mg/kg, significantly elevated THC blood and brain levels. No interactions between THC and CBD were observed in several variations of a marijuana smoke exposure model. Either quantities of CBD were applied directly to marijuana, CBD and THC were both applied to placebo plant material, or mice were pretreated intravenously with 30 mg/kg CBD before being exposed to marijuana smoke. CONCLUSIONS: As the amount of CBD found in most marijuana strains in the US is considerably less than that of THC, these results suggest that CBD concentrations relevant to what is normally found in marijuana exert very little, if any, modulatory effects on CB(1)-receptor-mediated pharmacological effects of marijuana smoke.

Delta9-tetrahydrocannbinol accounts for the antinociceptive, hypothermic, and cataleptic effects of marijuana in mice.

Although it is widely accepted that delta9-tetrahydrocannabinol (delta9-THC) is the primary psychoactive constituent of marijuana, questions persist as to whether other components contribute to marijuana's pharmacological activity. The present experiments assessed the cannabinoid activity of marijuana smoke exposure in mice and tested the hypothesis that delta9-THC mediates these effects through a CB1 receptor mechanism of action. First, the effects of delta9-THC on analgesia, hypothermia, and catalepsy were compared with those of a marijuana extract with equated delta9-THC content after either i.v. administration or inhalation exposure. Second, mice were exposed to smoke of an ethanol-extracted placebo plant material or low-grade marijuana (with minimal delta9-THC but similar levels of other cannabinoids) that were impregnated with varying quantities of delta9-THC. To assess doses, delta9-THC levels in the blood and brains of drug-exposed mice were determined following both i.v. and inhalation routes of administration. Both marijuana and delta9-THC produced comparable levels of antinociception, hypothermia, and catalepsy regardless of the route of administration, and these effects were blocked by pretreatment with the CB1 antagonist SR141716 [N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide HCl]. Importantly, the blood and brain levels of delta9-THC were similar in mice exhibiting similar pharmacological effects, regardless of the presence of non-delta9-THC marijuana constituents. The present experiments provide evidence that the acute cannabinoid effects of marijuana smoke exposure on analgesia, hypothermia, and catalepsy in mice result from delta9-THC content acting at CB1 receptors and that the non-delta9-THC constituents of marijuana (at concentrations relevant to those typically consumed) influence these effects only minimally, if at all.

Delta(9)-THC-induced cognitive deficits in mice are reversed by the GABA(A) antagonist bicuculline.

RATIONALE: The results of recent in vitro studies have underscored the important role that activation of CB(1) receptors has on GABAergic activity in brain areas associated with memory. OBJECTIVES: The primary purpose of this study was to test the hypothesis that the memory disruptive effects of Delta(9)-tetrahydrocannabinol (Delta(9)-THC) in vivo are mediated through GABAergic systems. Conversely, we also evaluated whether blocking CB(1) receptor signaling would alter memory deficits elicited by GABA agonists. METHODS: The GABA(A) antagonist bicuculline and GABA(B) antagonist CGP 36742 were evaluated for their ability to ameliorate Delta(9)-THC-induced deficits in a mouse working memory Morris water maze task. Mice were also assessed in a T-maze task, as well as non-cognitive behavioral assays. Additionally, the effects of GABA(A) and GABA(B) agonists were assessed in either CB(1) (-/-) mice or wild type mice treated with the CB(1) antagonist SR 141716. RESULTS: Memory deficits resulting from 10 mg/kg Delta(9)-THC in the Morris water maze were completely reversed by bicuculline, though unaffected by CGP 36742. Bicuculline also blocked the disruptive effects of Delta(9)-THC in the T-maze, but failed to alter non-mnemonic effects of Delta(9)-THC. Although CB(1) (-/-) mice exhibited supersensitivity to muscimol-induced water maze deficits compared with wild type control mice, muscimol elicited virtually identical effects in SR 141716-treated and vehicle-treated wild type mice. CONCLUSIONS: This is the first demonstration of which we are aware showing that GABA(A) receptors may play a necessary role in Delta(9)-THC-induced memory impairment in whole animals.

Disruption of CB(1) receptor signaling impairs extinction of spatial memory in mice.

RATIONALE: A growing body of in vitro and in vivo evidence indicates that a central endocannabinoid system, consisting of CB(1) receptors and endogenous cannabinoids, modulates specific aspects of mnemonic processes. Previous research has demonstrated that either permanent or drug-induced disruption of CB(1) receptor signaling interferes with the extinction of a conditioned fear response. OBJECTIVES: In the present study, we evaluated whether the endocannabinoid system also plays a role in extinguishing learned escape behavior in a Morris water maze task. METHODS: CB(1) (-/-) mice and mice repeatedly treated with 3 mg/kg of the CB(1) receptor antagonist SR 141716 (Rimonabant) were trained to locate a hidden platform in the Morris water maze. Following acquisition, the platform was removed and subjects were assigned to either a massed (i.e., five consecutive sessions consisting of four 2-min trials/session) or a spaced (a single, 1-min trial every 2-4 weeks) extinction protocol. RESULTS: Strikingly, both 3 mg/kg SR 141716-treated mice and CB(1) (-/-) mice continued to return to the target location across all five trials in the spaced extinction procedure, while the control mice underwent extinction by the third or fourth trial. In contrast, both the 3-mg/kg SR 141716-treated and CB(1) (-/-) mice exhibited extinction in the massed extinction trial procedure. CONCLUSIONS: These findings indicate that disruption of CB(1) receptor signaling impairs extinction processes in the Morris water maze, thus lending further support to the hypothesis that the endocannabinoid system plays an integral role in the suppression of non-reinforced learned behaviors.

Endocannabinoids in cognition and dependence.

Cannabis use is associated with a wide range of pharmacological effects, some of which have potential therapeutic benefit while others result in negative outcomes. Acute cannabinoid intoxication has been well documented to produce deficits in cognitive functioning with concomitant changes in glutamatergic, GABAergic, and cholinergic neurochemical systems in the hippocampus, each of which has been implicated in memory. Additionally, cannabis-dependent individuals abstaining from this drug can undergo a constellation of mild withdrawal effects. The use of the CB(1) cannabinoid receptor antagonist SR141716A and transgenic mice lacking the CB(1) receptor are critical tools for investigating the role of the endocannabinoid system in cognition, drug dependence, and other physiological processes. Converging evidence in which performance in a variety of memory tasks is enhanced following either SR141716A treatment or in CB(1) receptor knockout mice indicates that this system may play an important role in modulating cognition. There are also indications that this system may function to modulate opioid dependence. The purpose of this review is to describe recent advances that have furthered our understanding of the roles that the endocannabinoid system play on both cognition and drug dependence.

Evaluation of CB1 receptor knockout mice in the Morris water maze.

The endocannabinoid system has been proposed to modulate a variety of physiological processes, including those that underlie cognition. The present study tested whether this system is tonically active in learning and memory by comparing CB(1) receptor knockout mice (CB(1)(-/-)) to wild-type mice (CB(1)(+/+)) in several Morris water maze tasks. Also, the effects of three cannabinoid agonists, Delta(9)-tetrahydrocannabinol (Delta(9)-THC), R-(+)-[2,3-dihydro-5-methyl-3[morpholinyl)methyl]-pyrrolo[1,2,3-de]-1, 4-benzoxazinyl]-(1-naphthalenyl)methanone mesylate (WIN 55,212-2), and methanandamide, were evaluated in a working memory procedure. Both genotypes exhibited identical acquisition rates in a fixed platform procedure; however, the CB(1)(-/-) mice demonstrated significant deficits in a reversal task in which the location of the hidden platform was moved to the opposite side of the tank. This phenotype difference was most likely due to an increased perseverance of the CB(1)(-/-) mice in that they continued to return to the original platform location, despite being repeatedly shown the new platform location. In addition, Delta(9)-THC (ED(50) = 1.3 mg/kg), WIN 55,212-2 (ED(50) = 0.35 mg/kg), and methanandamide (ED(50) = 3.2 mg/kg) disrupted the performance of CB(1)(+/+) mice in the working memory task at doses that did not elicit motivational or sensorimotor impairment as assessed in a cued version of the task. Furthermore, doses of each drug that were maximally disruptive in CB(1)(+/+) mice were ineffective in either N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide HCl (SR 141716A)-treated CB(1)(+/+) or CB(1)(-/-) mice. These results provide strong evidence that cannabinoids disrupt working memory through a CB(1) receptor mechanism of action, and suggest that the endocannabinoid system may have a role in facilitating extinction and/or forgetting processes.

Effects of antipsychotic drugs on operant responding after acute and repeated administration.

RATIONALE: The current generation of atypical antipsychotic drugs represents an improvement over traditional ("typical") antipsychotics in many respects. However, a theoretical framework and adequate preclinical models have not yet been developed to predict or explain differences among the atypical antipsychotics, a necessary component of future development. OBJECTIVES: The purpose of the present set of experiments was to identify differences between the acute and subchronic effects of several atypical antipsychotic drugs and the typical antipsychotic haloperidol on operant responding in rats. METHODS: The effects of haloperidol and the atypical antipsychotics clozapine, olanzapine, risperidone, sertindole, quetiapine, remoxipride, and thioridazine were determined in rats trained to respond for food reward under a multiple fixed ratio 30/fixed interval 60 s schedule. A profile of the acute effects of each drug on response rates, response durations, and within-session effects were determined. Next, the dose of each drug that produced 75% suppression of response rates was administered for 16 consecutive days to determine whether or not tolerance would develop to the rate-suppressing effects of that dose. RESULTS: All drugs produced dose-related decreases in response rates. Only haloperidol and risperidone produced significant increases in response duration, while only haloperidol and remoxipride displayed within-session response decrements. Tolerance was evident for clozapine and to a lesser extent thioridazine. CONCLUSIONS: These results illustrate that the current generation of atypical antipsychotics are a heterogeneous group and that operant procedures may be useful for identifying differences preclinically. Specifically, clozapine appears to possess properties that distinguish it from other atypical antipsychotics, particularly after repeated dosing.

Differential effects of delta 9-THC on spatial reference and working memory in mice.

RATIONALE: Marijuana remains the most widely used illicit drug in the U.S., and recent attention has been given to putative therapeutic uses of marijuana and cannabinoid derivatives. Thus, developing a better understanding of delta9-THC (tetrahydrocannabinol)-induced mnemonic deficits is of critical importance. OBJECTIVES: These experiments were conducted to determine whether delta9-THC has differential effects on spatial reference and working memory tasks, to investigate its receptor mechanism of action, and to compare these effects with those produced by two other compounds--scopolamine and phencyclidine--known to produce mnemonic deficits. In addition, the potency of delta9-THC in these memory tasks was compared with its potency in other pharmacological effects traditionally associated with cannabinoid activity. METHODS: Two different versions of the Morris water maze were employed: a working memory task and a reference memory task. Other effects of delta9-THC were assessed using standard tests of hypomotility, antinociception, catalepsy, and hypothermia. RESULTS: delta9-THC disrupted performance of the working memory task (3.0 mg/kg) at doses lower than those required to disrupt performance of the reference memory task (100 mg/kg), or elicit hypomotility, antinociception, catalepsy, and hypothermia. These performance deficits were reversed by SR 141716A. The effects of delta9-THC resembled those of scopolamine, which also selectively disrupted the working maze task. Conversely, phencyclidine disrupted both tasks only at a dose that also produced motor deficits. CONCLUSIONS: These data indicate that delta9-THC selectively impairs performance of a working memory task through a CB, receptor mechanism of action and that these memory disruptions are more sensitive than other pharmacological effects of delta9-THC.

Clozapine discrimination with a low training dose distinguishes atypical from typical antipsychotic drugs in rats.

RATIONALE: Previous drug discrimination studies with clozapine have not reliably distinguished between atypical and typical antipsychotics. OBJECTIVES: The present study was conducted to determine whether low-dose clozapine drug discrimination could distinguish atypical from typical antipsychotics. METHODS: Rats were trained to discriminate 1.25 mg/kg clozapine from vehicle in a two-lever drug discrimination procedure. RESULTS: Generalization testing revealed full substitution with the atypical antipsychotics olanzapine (90.3% maximum generalization), sertindole (99.8%), and risperidone (87.1%) and partial substitution for quetiapine (seroquel, 66.4%) and the typical antipsychotics haloperidol (56.8%) and thioridazine (74.3%). Remoxipride (23.1%) and the typical antipsychotics chlorpromazine (27.9%) and fluphenazine (29.5%) did not reliably substitute for clozapine. CONCLUSIONS: In contrast to previous clozapine drug discrimination studies with higher training doses, the atypical antipsychotics olanzapine, sertindole, and risperidone reliably substituted for clozapine while typical antipsychotics did not. These results suggest that low-dose clozapine drug discrimination may be a more sensitive assay for distinguishing atypical from typical antipsychotic drugs.

The discriminative stimulus properties of the atypical antipsychotic olanzapine in rats.

RATIONALE: Analysis of the preclinical behavioral effects of atypical antipsychotic agents will provide a better understanding of how they differ from typical antipsychotics and aid in the development of future atypical antipsychotic drugs. OBJECTIVES: The present study was designed to provide information about the discriminative stimulus properties of the atypical antipsychotic olanzapine. METHODS: Rats were trained to discriminate the atypical antipsychotic olanzapine (either 0.5 mg/kg OLZ or 0.25 mg/kg OLZ, i.p.) from vehicle in a twolever drug discrimination procedure. The atypical antipsychotic clozapine fully substituted for olanzapine in both the 0.5-mg/kg OLZ group (99.3% drug lever responding [DLR]) and the 0.25-mg/kg OLZ group (99.9% DLR). The typical antipsychotic chlorpromazine also substituted for olanzapine in both the 0.5-mg/kg OLZ group (87.5% DLR) and in the 0.25-mg/kg OLZ group (98.9% DLR); whereas, haloperidol displayed partial substitution for olanzapine in the 0.5-mg/kg OLZ group (56.1% DLR) and in the 0.25-mg/kg OLZ group (76.4% DLR). The 5.0-mg/kg dose of thioridazine produced olanzapine-appropriate responding in the 0.5-mg/kg OLZ group (99.6% DLR), but only partial substitution was seen with the 0.25-mg/kg OLZ training dose (64.0% DLR). The atypical antipsychotics raclopride (53.9% DLR) and risperidone (60.1% DLR) displayed only partial substitution in the 0.5-mg/kg OLZ group. Both the muscarinic cholinergic antagonist scopolamine (90.0% DLR) and the 5-HT2A/2C serotonergic antagonist ritanserin (86.0% DLR) fully substituted for olanzapine in the 0.5-mg/kg OLZ group. CONCLUSIONS: In contrast to previous discrimination studies with clozapine-trained rats, the typical antipsychotic agents chlorpromazine and thioridazine and the serotonin antagonist ritanserin substituted for olanzapine. These results demonstrate that there are differences in the mechanisms underlying the discriminative stimulus properties of clozapine and olanzapine. Specifically, olanzapine's discriminative stimulus properties appear to be meditated in part by both cholinergic and serotonergic mechanisms.

Discriminative stimulus (DS) properties of nicotine in the C57BL/6 mouse.

Previous research conducted in this and other laboratories has examined the role of genetic factors in determining sensitivity to (-)-nicotine in a variety of behavioral and physiological measures in the rat. More recent research further indicates that genetic factors can also influence the level of sensitivity to (-)-nicotine when serving as a discriminative stimulus (DS) in different rat strains. However, there has been little work examining the influence of genotype on the discriminative stimulus (DS) properties of (-)-nicotine in mice, a species that has played a major role in understanding the relationship between genetics and (-)-nicotine pharmacological effects. To further our understanding of the role of genetics and the ability of (-)-nicotine to exert DS control of behavior in the mouse, a group of C57BL/6 mice was trained to discriminate 0.4 mg/kg (-)-nicotine from saline using a two-lever operant procedure. (-)-Nicotine's discriminative stimulus in C57BL/6 mice appears to be similar to that generated in the rat. Results from behavioral tests with other drugs indicated that d-amphetamine exhibited a partial generalization, while (+)-nicotine fully generalized with nicotine. Tests of antagonism with mecamylamine and scopolamine further showed the cholinergic specificity of the (-)-nicotine DS in the mouse; mecamylamine but not scopolamine completely antagonized the (-)-nicotine DS. This work lays the groundwork for future comparisons of different mouse strain's sensitivities to (-)-nicotine's discriminative stimulus as well as using this behavioral model to search for new nicotinic receptor agonists and antagonists.

Effects of modulation of NMDA neurotransmission on response rate and duration in a conflict procedure in rats.

N-Methyl-D-aspartate (NMDA) antagonists and gamma-aminobutyric acid agonists share a number of common pharmacological properties, including motor and anticonvulsant effects. In the present study, site-selective NMDA antagonists were evaluated for potential anxiolytic efficacy and motor impairment in a modified Geller-Seifter conflict procedure, an animal model widely used to screen drugs for anxiolytic effects. Male Sprague-Dawley rats were trained to respond for food reward under a multiple FI 30 s (food only), FR 10 (food + shock) operant schedule. Consistent with the results of previous studies, the benzodiazepines chlordiazepoxide and diazepam selectively increased punished responding and increased response durations at higher doses. The competitive NMDA antagonist CGP 37,849 increased punished responding at some doses, though not selectively, and also increased response duration in both schedule components. The glycine-site modulators milacemide, ACEA 1011 and ACEA 1021, the NR2B-selective polyamine site antagonist eliprodil and NMDA did not produce anticonflict effects at any dose and had inconsistent effects on response durations. These results suggest that the anticonflict effects of NMDA antagonists are not as reliable as those of the benzodiazepines. Further research is needed to clarify the experimental conditions under which the anxiolytic potential of NMDA antagonists is most evident.

Mianserin as a discriminative stimulus in rats: asymmetrical cross-generalization with scopolamine.

The present study was conducted to determine if the tetracyclic antidepressant mianserin could be established as a discriminative stimulus in rats. One group of rats was trained to discriminate mianserin (4.0 mg/kg, IP) from saline in a two-lever drug discrimination procedure, and a second group of rats was trained to discriminate the muscarinic cholinergic antagonist scopolamine (0.25 mg/kg, IP) from saline. Generalization testing with the training drugs yielded an ED50 of 0.502 mg/kg for the mianserin-trained rats and an ED50 of 0.048 mg/kg for the scopolamine-trained rats. Asymmetrical cross-generalization between mianserin and scopolamine was observed, because scopolamine produced mianserin-appropriate responding, but mianserin did not produce scopolamine-appropriate responding. This study is the first demonstration that rats can be trained to discriminate mianserin from saline and that antagonism of muscarinic cholinergic receptors is sufficient to produce mianserin-appropriate responding.