Carreño Diana, Lotfipour Shahrdad
Department of Pharmaceutical Sciences, University of California, Irvine, Irvine, CA, United States.
Department of Pathology and Laboratory Medicine, University of California, Irvine, Irvine, CA, United States.
Front Psychiatry. 2023 Jan 10;13:1064211. doi: 10.3389/fpsyt.2022.1064211. eCollection 2022.
Large-scale human candidate gene studies have indicated that a genetic variant (rs2304297) in the alpha(α)6 nicotinic acetylcholine receptor (nAChR) subunit, encoded by the 6 gene, may play a key role in adolescent nicotine addictive behavior. We hypothesized that the polymorphism selectively enhances nicotine + cue-primed reinstatement, but not nicotine- or cue-reinstatement in α6 (risk) vs. α6 (non-risk) allele carriers, without having baseline effects on natural rewards.
Using CRISPR-Cas9 genomic engineering, we developed a humanized rat line with the human gene variant of the 6 3'-UTR polymorphism in Sprague-Dawley rats. Genetically modified adolescent male and female rats were food trained under a fixed-ratio (FR)1 schedule of reinforcement and progressively increased to FR5. Animals were implanted with catheters and began nicotine self-administration (15 μg/kg/infusion) at FR5. Upon reaching stable responding, reinforced behavior was extinguished by removal of drug and cues. Reinstatement testing began for cue only, nicotine only, and nicotine + cue in a Latin Square Design. Animals were returned to extinction conditions for 2 days minimum between testing.
For natural food rewards, nicotine self-administration, progressive ratio, and extinction, adolescent male and female (α6 and α6 ) rats exhibited equivalent behaviors. Male α6 rats show enhanced nicotine + cue-primed reinstatement when compared with male α6 rats. This genotype effect on reinstatement was not seen in female rats.
Our findings support the functional role of the human 6 3'-UTR SNP genetic variant in sex-dependently enhancing nicotine seeking behavior in adolescent rats. Overall, the findings support clinical and preclinical data highlighting a role of α6 nAChRs mediating sex heterogeneity in substance use and related phenotypes.
大规模人类候选基因研究表明，由CHRNA6基因编码的α6烟碱型乙酰胆碱受体（nAChR）亚基中的一个基因变异（rs2304297）可能在青少年尼古丁成瘾行为中起关键作用。我们假设，在α6（风险）与α6（非风险）等位基因携带者中，该多态性选择性增强尼古丁+线索引发的复吸，但不增强尼古丁或线索引发的复吸，且对自然奖赏没有基线影响。
利用CRISPR-Cas9基因组工程技术，我们在斯普拉格-道利大鼠中培育出了携带人类CHRNA6 3'-UTR多态性基因变异的人源化大鼠品系。对经过基因改造的青春期雄性和雌性大鼠进行食物训练，采用固定比率（FR）1强化程序，并逐步增加到FR5。给动物植入导管，并在FR5时开始尼古丁自我给药（15μg/kg/输注）。在达到稳定反应后，通过去除药物和线索来消除强化行为。采用拉丁方设计对仅线索、仅尼古丁和尼古丁+线索进行复吸测试。在测试之间，动物至少要回到消退状态2天。
对于自然食物奖赏以及尼古丁自我给药、渐进比率和消退，青春期雄性和雌性（α6和α6）大鼠表现出相同的行为。与雄性α6大鼠相比，雄性α6大鼠表现出增强的尼古丁+线索引发的复吸。在雌性大鼠中未观察到这种基因型对复吸行为的影响。
我们的研究结果支持人类CHRNA6 3'-UTR SNP基因变异在性别依赖性地增强青春期大鼠尼古丁寻求行为中的功能作用。总体而言，这些结果支持临床和临床前数据，突出了α6 nAChR在介导物质使用和相关表型中的性别异质性方面的作用。
