CB1 and DA Receptor Deletion by RNAi in Cocaine Self-Administration

描述（由申请人提供）：本K01指导研究职业发展奖旨在培训维克森林大学生理与药理学系的初级教师Caroline E. Bass，以开发一个独立的假设驱动的研究项目，研究中皮质边缘系统中多巴胺和大麻素受体调节的成瘾过程。Bass博士将运用她的技能开发用于递送RNA干扰（RNAi）转基因的腺相关病毒（AAV）载体，以开发她自己的项目，重点研究在战略定义的脑位点中D1， D2和CB1受体的敲低，这对可卡因强化效果的影响，以及大麻素和多巴胺系统如何在可卡因自我给药模型中相互作用。巴斯博士在大麻素药理学和基因表达变化方面的指导将来自阿林·c·豪利特博士，她在行为模型方面的指导，特别是大鼠自我给药方面的指导将来自大卫·c·s·罗伯茨博士，他们都是这两个领域的杰出科学家。其他指导将涵盖与学术提升和国家认可相适应的所有专业技能。为了实现本课题的目标，我们提出了以下具体目标：1)生成并表征含有RNAi表达盒的AAV载体，该载体可导致大鼠D1、D2和CB1受体mRNA的降解，并确定这种操作导致多巴胺和内源性大麻素神经传递分子成分的代偿性变化；2)通过向纹状体和髓质注射CB1 RNAi-AAV，探讨四氢大麻酚非条件行为效应的作用部位，评价四氢大麻酚诱导抗痛觉、低活性、低体温和猝倒的作用；3)通过对自用可卡因大鼠进行D1或D2 RNAi-AAV的部位特异性注射，确定腹侧纹状体、背侧纹状体和VTA中D1和D2受体在可卡因强化作用中的作用；4)表征腹侧纹状体、背侧纹状体和VTA CB1受体消融对可卡因自我给药的影响。具体而言，将CB1 RNAi-AAV注射到这些区域，并评估可卡因的强化效果。在这些目标的所有阶段，大麻素和多巴胺神经传递的代偿机制将被评估，以确定这些受体系统如何在调节可卡因摄入中相互作用。这些研究将解决我们对多巴胺受体在强化中的功能的认识的基本空白，并作为理解大麻素和多巴胺受体系统如何相互作用以调节成瘾行为的基础。这些发现可能会导致新的和更有效的药物成瘾治疗干预措施的发展。

DESCRIPTION (provided by applicant): This K01 Mentored Research Career Development Award is to train Caroline E. Bass, junior faculty in the Dept. Physiology & Pharmacology at Wake Forest Univ., to develop an independent hypothesis-driven research project to investigate addiction processes that are modulated by dopamine and cannabinoid receptors in the mesocorticolimbic system. Dr. Bass will apply her skills in the development of adenoassociated virus (AAV) vectors for the delivery of RNA interference (RNAi) transgenes to develop her own project focused on the knock down of D1, D2 and CB1 receptors in strategically defined brain loci, the impact this has on the reinforcing efficacy of cocaine, and how the cananbiniod and dopamine systems interact in a cocaine self-administration model of drug abuse. Dr. Bass's mentoring in cannabinoid pharmacology and gene expression changes will come from Dr. Allyn C. Howlett, and her mentoring in the behavioral models, particularly rat self-administration, will come from Dr. David C.S. Roberts, who are distinguished scientists in these respective areas. Other mentoring will cover all aspects of professional skills commensurate with academic promotion and national recognition. The following specific aims are proposed to meet the goals of this proposal: 1) Generate and characterize AAV vectors containing RNAi expression cassettes that cause the degradation of D1, D2 and CB1 receptor mRNA in rats and determine the compensatory changes in molecular components of dopamine and endocannabinoid neurotransmission resulting from this manipulation; 2) Investigate the site(s) of action of non-conditioned behavioral effects of THC by injecting the CB1 RNAi-AAV into the striatum and medulla and assessing the efficacy of THC in inducing antinociception, hypoactivity, hypothermia, and catalepsy; 3) delineate and define the contribution of D1 and D2 receptors in the ventral striatum, dorsal striatum and VTA in the reinforcing effects of cocaine by site specific injection of D1 or D2 RNAi-AAV in rats self-administering cocaine; 4) Characterize the impact of CB1 receptor ablation from the ventral striatum, dorsal striatum and VTA on cocaine self-administration. Specifically, the CB1 RNAi-AAV will be injected into these regions and the reinforcing efficacy of cocaine will be assessed. During all stages of these aims the compensatory mechanisms in cannabinoid and dopamine neurotransmission will be assessed to determine how these receptor systems are interacting in modulating cocaine intake. These studies will address fundamental gaps in our knowledge of dopamine receptor function in reinforcement and serve as a basis for understanding how cannabinoid and dopamine receptor systems interact to regulate addictive behaviors. These findings may lead to the development of novel and more effective therapeutic interventions for drug addiction.