Discriminative stimulus properties of Δ⁹-tetrahydrocannabinol (THC) in C57Bl/6J mice

Abstract

Primarily, rats have served as subjects in Δ⁹-tetrahydrocannabinol's (THC) discrimination studies although other species such as monkeys and pigeons have been used. While the introduction of the knockout and transgenic mice has vastly stimulated the study of the discriminative stimulus effects of drugs there is only a single published report of mice trained to discriminate THC. Thus, this study extended those results by providing a systematic replication that THC serves as an effective discriminative stimulus in mice and by further investigating the mechanisms of action involved in the THC discrimination model in the mouse. Male C57BL/6J mice were trained to discriminate 10 mg/kg THC from vehicle in 2-lever drug discrimination. THC fully and dose dependently substituted for itself. Cannabinoid indoles, except one with low cannabinoid CB₁ receptor affinity, substituted for THC. Anandamide failed to substitute for THC when administered alone but completely substituted when administered with the non-specific fatty acid amide hydrolase inhibitor, phenylmethylsulphonyl fluoride. As expected, nicotine failed to substitute for THC. Lastly, the cannabinoid CB₁ receptor antagonist rimonabant blocked THC's discriminative stimulus effects. Taken together these studies demonstrate THC's ability to produce discriminative stimulus effects as well as demonstrate its pharmacological specificity and mechanism of action in a two-lever drug discrimination mouse model.

Introduction

Mapping of the mouse genome has allowed rapid advances in the development of techniques for mutagenesis of specific genes, as seen in the creation of knockout and transgenic mice (see Picciotto and Wickman, 1998). Although genotype is relatively easy to verify, determination of phenotypic differences is often more difficult, particularly if expression of the mutant phenotype primarily involves behavior. Consequently, recent years have seen increased emphasis on development of novel behavioral techniques for mice as well as alteration of existing procedures (often developed in rats) for use in mice. Since differences in the functional consequences of manipulation of genes that encode for specific receptors or steps in neurotransmission processes may not be evident through use of strictly observational measures, more complicated operant procedures have often been used to measure constructs such as learning and memory. In addition, pharmacological challenge often is employed.

Drug discrimination is increasingly being used in mice as a method to tease out pharmacologically selective differences in behavioral phenotypes. In drug discrimination, a subject is trained to make one response if administered a specific psychoactive drug (i.e., the training drug) and to make another response if administered vehicle or a drug that does not share the same psychoactive properties. In mice, the typical response requirement is a lever press or a head poke on a drug- or vehicle-associated side, respectively, with correct responses resulting in delivery of reinforcement (an appetitive reinforcer such as food pellet or sweetened milk). Previous studies have demonstrated that mice can be trained to discriminate a variety of psychoactive drugs, including clozapine (Philibin et al., 2005), ethanol (Bowen and Balster, 1997), pentobarbital (Balster and Moser, 1987), and nicotine (Varvel et al., 1999). Moreover, knockout mice have been used to gain further insights regarding the discriminative stimulus effects of drugs such as nicotine with alpha 7 knockout mice (Stolerman et al., 2004) and cocaine with dopamine D5 knockout mice (Elliot et al., 2003).

Δ⁹-Tetrahydrocannabinol (THC), the principal psychoactive substituent of the marijuana plant (Cannabis sativa), has also been used as a discriminative stimulus in laboratory animals, including rats (Burkey and Nation, 1997, Järbe et al., 1998, Solinas et al., 2007, Vann et al., 2007, Wiley et al., 1995a), rhesus monkeys (Wiley et al., 1995c), pigeons (Henriksson et al., 1975), and gerbils (Järbe et al., 1975). Collectively, these studies have revealed that the discriminative stimulus effects of THC are pharmacologically selective for psychoactive cannabinoids (Wiley et al., 1995b) and are blocked by administration of rimonabant and other antagonists of cannabinoid (CB₁) receptors in the brain, but not by an antagonist of cannabinoid CB₂ receptors (Järbe et al., 2001, Järbe et al., 2006, Wiley et al., 1995c). Further, centrally active cannabinoids of various structural classes substitute for THC with strong correlation between potency for substitution and affinity for cannabinoid CB₁ receptors (Compton et al., 1993). Moreover, the discriminative stimulus effects of cannabinoids in animals correspond to the subjective effects of cannabinoids that have been assessed in humans (Chait et al., 1988, Lile et al., 2008); hence, THC discrimination represents an animal model of marijuana intoxication (Balster and Prescott, 1992).

Recently, McMahon et al. (2008) reported the first successful training of a 2-nose poke THC versus vehicle discrimination in C57BL/6J mice. In this initial manuscript, the structurally dissimilar cannabinoids, CP 55940 and WIN 55212-2, substituted for THC in mice trained whereas the metabolically stable anandamide analog methanandamide and noncananbinoid drugs (ketamine, cocaine and ethanol) did not. The present study extends the findings of that study and provides additional findings regarding the mechanisms of action of a 2-lever THC versus vehicle discrimination in C57BL/6J mice through a variety of techniques, including co-administration of antagonists or metabolic inhibitors, substitution tests with a non-cannabinoid compound (nicotine), and preliminary assessment of structure–activity relationship with indole-derived cannabinoids (Table 1).