The role of the circadian transcription factor NPAS2 in the nucleus accumbens to regulate cocaine reward

描述（由申请人提供）：基础和临床研究表明，昼夜节律和成瘾之间存在广泛的双向相互作用。由于环境或遗传干扰而造成的昼夜节律系统紊乱可能会增加成瘾的脆弱性，而长期使用药物会导致昼夜节律紊乱，这种紊乱在戒断期间持续存在，并可能导致复发。虽然这些关系很有趣，但人们对昼夜节律系统与成瘾过渡之间关系的分子机制知之甚少。动物研究表明，长期暴露于可卡因导致特定昼夜节律基因（即，分子钟的组成部分）在中脑边缘多巴胺奖励系统。奖赏回路的一个主要会聚区域和调节药物奖赏和动机的关键底物是丘脑核（NAc）。NAc主要由主要表达多巴胺1或2受体（D1+或D2+）的两种特定亚型的中型多刺神经元（MSN）组成。这两种MSNs亚型在可卡因奖赏行为的调节中具有不同的作用，尽管这些差异背后的分子机制仍不清楚。我们已经确定了一种新的昼夜节律转录因子，神经元PAS结构域蛋白2（NPAS 2）在可卡因奖赏的调节中的作用，可能是通过其与转录因子核因子κ B（NFκB）的相互作用及其在NAc的D1+ MSNs中的选择性表达。K 01的目的如下：1）研究NPAS 2在不同可卡因奖赏行为的调节中的功能作用，（即，条件性位置偏爱和自我给药），通过在NAc的D1+或D2+ MSN中的表达; 2）检查急性和慢性可卡因暴露对Npas 2表达的影响， NFκB沿着NAc中这些转录因子之间的相互作用;以及3）研究NPAS 2是否正调控NAc的D1+和D2+ MSN中NFκ B介导的转录。因此，拟议的K 01奖将表征这些新的细胞类型特异性分子机制在调节可卡因奖赏的NAc中的作用。这些研究还将进一步阐明分子钟在成瘾转变中的作用。这些研究目标与NIDA和NIH BRAIN Initiative的战略目标一致，即通过整合创新的分子和行为方法来确定大脑中特定细胞类型的功能意义。候选人的长期职业目标是开发一个独立的研究项目，调查昼夜节律系统在奖励电路调节中的作用，该电路是向药物成瘾过渡的基础。候选人在昼夜节律研究方面有很强的背景，需要额外的培训来发展转化成瘾研究的专业知识，以及先进的分子方法和复杂的行为测定的结合。拟议的K 01职业发展奖将通过在以下领域提供广泛的概念和方法培训，确保候选人成功过渡到独立调查员：1）基础和转化研究成瘾神经生物学的概念专业知识; 2）熟练掌握先进分子技术的结合方法，包括蛋白质免疫沉淀，免疫组织化学，和脑细胞类型特异性病毒介导的基因转移，与复杂的行为测定，包括可卡因条件性位置偏爱和静脉注射可卡因自我管理的小鼠;和3）整合以前的概念和研究能力与新获得的神经生物学技术和方法，以及成瘾研究的概念专业知识，成为一个独立的研究者。候选人将由一个在成瘾的细胞和分子机制，奖励神经回路和药物诱导的神经可塑性以及昼夜节律方面具有专业知识的教师团队指导和建议。个性化的职业发展计划包括与导师和顾问定期会面、专业技能培训、正式课程、出席会议、正式研究报告以及技术和方法培训。匹兹堡大学和精神病学系提供必要的基础设施，成为一个成功的独立成瘾研究人员，包括特殊的核心和导师研究设施，个性化的职业发展计划和培训机会，以及机构和部门主席长期致力于培养有前途的年轻研究人员。

DESCRIPTION (provided by applicant): Basic and clinical research suggests there are extensive bidirectional interactions between circadian rhythms and addiction. Disruptions to the circadian system, either by environmental or genetic perturbation, may increase the vulnerability to addiction, while chronic drug use leads to circadian disruptions that persist during abstinence and may contribute to relapse. Although these relationships are intriguing, very little is known about the molecular mechanisms underlying the relationship between the circadian system and the transition to addiction. Animal studies have demonstrated that chronic exposure to cocaine leads to alterations in the expression and function of specific circadian genes (i.e., components of the molecular clock) in the mesolimbic dopamine reward system. A major region of convergence for reward circuitry and a key substrate that regulates drug reward and motivation is the nucleus accumbens (NAc). The NAc is comprised of mostly two specific subtypes of medium spiny neurons (MSNs) that predominantly express either dopamine 1 or 2 receptors (D1+ or D2+). These two subtypes of MSNs have distinct roles in the regulation of cocaine reward behaviors, although the molecular mechanisms underlying these differences remain unclear. We have identified a novel role of the circadian transcription factor, neuronal PAS domain protein 2 (NPAS2) in the regulation of cocaine reward, potentially via its interactions with the transcription factor nuclear factor kappa B (NFκB) and its selective expression in D1+ MSNs of the NAc. The aims of the K01 are as follows: 1) to investigate the functional role of NPAS2 in the regulation of different cocaine reward behaviors, (i.e., conditioned place preference and self-administration), via expression in D1+ or D2+ MSNs of the NAc; 2) to examine the effects of acute and chronic cocaine exposure on the expression of Npas2 and NFκB, along with the interactions between these transcription factors in the NAc; and 3) to investigate whether NPAS2 positively regulates NFκB-mediated transcription in D1+ and D2+ MSNs of the NAc. Therefore, the proposed K01 award will characterize the role of these novel cell type specific molecular mechanisms in the NAc that regulate cocaine reward. These studies will also further clarify the role of the molecular clock in the transition to addiction. These research goals are aligned with the strategic goals of NIDA and the NIH BRAIN Initiative to identify the functional significance of specific cell types in the brain by integrating innovative molecular and behavioral approaches. The candidate's long-term career goal is to develop an independent research program investigating the role of the circadian system in the regulation of reward circuitry that underlies the transition to drug addiction. The candidate has a strong background in circadian rhythms research and requires additional training to develop expertise in translational addiction research and the combination of advanced molecular approaches and complex behavioral assays. The proposed K01 Career Development Award will ensure the candidate's successful transition to an independent investigator by providing extensive conceptual and methodological training in the following areas: 1) conceptual expertise on the neurobiology of addiction for basic and translational research; 2) methodological proficiency in the combination of advanced molecular techniques, including protein immunoprecipitation, immunohistochemistry, and brain cell type specific viral-mediated gene transfer, with complex behavioral assays, including cocaine conditioned place preference and intravenous cocaine self-administration in mice; and 3) to integrate previous conceptual and research abilities with newly acquired neurobiological techniques and approaches, and conceptual expertise in addiction research, to become an independent investigator. The candidate will be mentored and advised by a team of faculty with expertise in the cellular and molecular mechanisms of addiction, reward neural circuitry and drug-induced neural plasticity, and circadian rhythms. The individualized career development plan includes a combination of regular meetings with mentors and consultants, grantsmanship skills training, formal coursework, conference attendance, formal research presentations, and technical and methodological training. The University of Pittsburgh and the Department of Psychiatry provides the necessary infrastructure to become a successful independent addiction researcher, including exceptional core and mentor research facilities, individualized career development programs and training opportunities, and a long-standing commitment by the institution and the departmental chair to develop promising young investigators.