Involvement of cannabinoid CB2 receptor in alcohol preference in mice and alcoholism in humans

The Pharmacogenomics Journal (2007) 7, 380–385 & 2007 Nature Publishing Group All rights reserved 1470-269X/07 $30.00 www.nature.com/tpj ORIGINAL ARTICLE Involvement of cannabinoid CB2 receptor in alcohol preference in mice and alcoholism in humans H Ishiguro1, S Iwasaki1, L Teasenfitz2, S Higuchi3, Y Horiuchi1, T Saito4, T Arinami1 and ES Onaivi2 1 Department of Medical Genetics, Doctoral Program in Social and Environmental Medicine, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan; 2Department of Biology, William Paterson University, Wayne, NJ, USA; 3National Hospital Organization, Kurihama Alcoholism Center, Nobi, Yokosuka, Kanagawa, Japan and 4 Department of Psychiatry, Sapporo Medical University, School of Medicine, Chuuouku, Sapporo, Hokkaido, Japan Correspondence: Dr H Ishiguro, Department of Medical Genetics, University of Tsukuba, 1-1-1 Tennoudai, Tsukuba, Ibaraki 305-8575, Japan. E-mail: We tested if cannabinoid type 2 receptor (CB2) in the central nervous system plays a role in alcohol abuse/dependence in animal model and then examined an association between the CB2 gene polymorphism and alcoholism in human. Mice experiencing more alcohol preference by drinking showed reduced Cb2 gene expression, whereas mice with little preference showed no changes of it in ventral midbrain. Alcohol preference in conjunction with chronic mild stress were enhanced in mice treated with CB2 agonist JWH015 when subjected to chronic stress, whereas antagonist AM630 prevented development of alcohol preference. There is an association between the Q63R polymorphism of the CB2 gene and alcoholism in a Japanese population (P ¼ 0.007; odds ratio 1.25, 95% CI, (1.06–1.47)). CB2 under such environment is associated with the physiologic effects of alcohol and CB2 antagonists may have potential as therapies for alcoholism. The Pharmacogenomics Journal (2007) 7, 380–385; doi:10.1038/sj.tpj.6500431; published online 26 December 2006 Keywords: CMS; alcoholism; cannabinoid; reinforcement Introduction Alcohol is one of the oldest substances abused by human, and significant advances have been made towards understanding the neurobiological effects of alcohol. However, the exact mechanism that underlies alcohol addiction is not completely understood. Although classical genetic studies, such as twin and adoption studies, estimated the genetic contribution to alcoholism as approximately 0.5,1–3 few family-based linkage studies have yielded consistent linkages at specific loci.4–7 The genetic polymorphisms in the alcohol metabolizing pathway, such as aldehyde dehydrogenase 2 (ALDH2)8 and alcohol dehydrogenase (ADH)9–12 appear to be associated with alcoholism vulnerability. Aside from the genes encoding the metabolizing enzyme, there appear to be no single gene that plays a significant role in alcoholism, and small functional gene effects may act in conjunction with environmental factors to promote alcoholism. However, such gene–environment interaction underlie alcoholism have not been revealed yet, although a study reported that forced swim stress led to an increase of ethanol consumption in mice.13 Several lines of experimental evidence support roles for the endocannabinoid system in alcoholism. The endocannabinoid system consists of cannabinoid receptors, endocannabinoids, enzymes for the synthesis and degradation of endocannabinoids, and also perhaps endocannabinoid transporters, which have Received 25 July 2006; revised 30 August 2006; accepted 24 September 2006; not been identified. There are two well-characterized cannabinoid receptors (CNRs), CB1/CNR1 and CB2/CNR2, that mediate the effects of endocannabinoid published online 26 December 2006 CB2 and alcoholism H Ishiguro et al 381 and exocannabinoid from marijuana use. CNR1 is expressed primarily in the central nervous system (CNS) and peripheral tissues, whereas CNR2 is expressed mainly in some peripheral and immune cells and has therefore been traditionally referred to as the peripheral cannabinoid receptor.14,15 Studies in rodent models have suggested that CNR1 is involved in the neural circuitry regulating alcohol consumption and motivation to consume alcohol. For example, although CNR1 agonists stimulate alcohol intake and the motivational properties of alcohol, the CNR1 antagonist rimonabant suppresses acquisition and maintenance of alcohol drinking behavior, relapse-like drinking, and the motivational properties of alcohol in rats.16 Ethanol self-administration and ethanol-conditioned place preference were reduced in mice lacking Cnr1, and treatment with rimonabant reduced ethanol intake in heterozygotes but had little or no effect in the Cnr1 mutant mice, suggesting that cannabinoid system is an essential component of the molecular pathways that underlie the reinforcing effects of alcohol.17 In CNS, cannabinoids and ethanol activate the same reward pathways, and recent advances in understanding of the neurobiological basis of alcoholism suggest that the CNR1 system may play a key role in the reinforcing effects of ethanol and in modulating ethanol intake.18 CNR1 polymorphisms were found to be associated with polysubstance abuse, including alcoholism.19 Thus, cannabinoids, which are the main component of marijuana, act with exocannabinoids on the endocannabinoid system, which plays a significant role in vulnerability to development of addiction and other mental disturbances.20 CNR2 may also be associated with addiction vulnerability as a modulator of the reward system. CNR2 has been observed in the brainstem,21 cerebellum22 and several other regions of the brain.23 We also found that expression of Cnr2 is altered in response to cocaine and heroin (manuscript in preparation). In view of our findings that CNR2 is expressed in the mammalian brain, and that this expression is altered in response to addictive drugs, we hypothesized that genetic variants of CNR2 may have a significant effect on alcohol dependency. Recently, the polymorphism which makes the substitution of glutamine at amino acid position 63 by arginine (Q63R: two base pairs replacement polymorphism, although registered as single base polymorphism, rs2501432 in NCBI SNP database) was reported to be associated with autoimmune disease,24 and human osteoporosis.25 Sipe et al.24 reported its functional change from the polymorphism in the immune system by in vitro study that may suggest a differential function in CNS. Therefore, we examined a possible role of Cnr2 in alcohol preference/abuse/addiction in mice and association between the Q63R polymorphism in the CNR2 gene and alcoholism in a Japanese population. Results Expression of Cnr2 is regulated by ethanol exposure in mice After 15 days of free access to alcohol drink, there were differences in development of reinforcement for ethanol under the same experimental conditions. Although there was little difference in alcohol consumption during the days using 2–8% alcoho (...truncated)