Structure-Function Analysis of the D3 Dopamine Receptor

D3 多巴胺受体的结构功能分析

• 批准号: 6383157
• 负责人: ELDO V KUZHIKANDATHIL
• 金额: $ 30.91万
• 依托单位: UNIV OF MED/DENT OF NJ-NJ MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-15 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6383157
• 关键词: G protein; biological signal transduction; calcium channel; dopamine receptor; electrophysiology; laboratory rat; nucleus accumbens; potassium channel; protein structure function; receptor coupling; receptor expression; tissue /cell culture

DESCRIPTION(Provided by applicant): The dopaminergic system and in particular the D3 dopamine receptor have been implicated in the etiology ofneuropsychiatric disorders such as schizophrenia and drug addiction. Studies using genetic, molecul biological and pharmacological techniques suggest that the D3 dopamine receptor plays an important role locomotor activity and behavioral effects involving reinforcement and reward. However the role of L receptor in neuronal signaling and the molecular mechanisms by which it modulates the physiological and behavioral effects are not well understood. It has been proposed that the D3 receptor exerts its phenotyp effects by modulating ion channel function and consequently neuronal signaling. We have recent demonstrated that the human D3 receptor expressed in the AtT-20 neuroendocrine cell line couples endogenous G-protein activated inward rectifier potassium (GIRK) channels and P/Q-type calcium channel thereby modulating spontaneous secretory activity. In this project, we plan to characterize the properties oft] D3 dopamine receptor and identify the structural features of the receptor that are involved in coupling to o about channels. Given the limited expression profile of the D3 receptor in vivo, the lack of selective pharmacologic ligands, and the need to evaluate properties of mutated D3 receptors, this study will primarily use the AtT-2 heterologous expression system. In addition, to assess physiological relevance, we will also study the I receptor-ion channel coupling in primary cultures of rat nucleus accumbens neurons using a combination electrophysiology and single cell reverse transcriptase-PCR. To define the properties of the D3 dopamine receptor, we will employ methods such as single cell electrophysiology, mutagenesis and expression recombinant and tagged receptors. We will address three specific aims: (1) Investigate the coupling of I receptor to ion channels in primary neuronal cultures from rat nucleus accumbens; (2) Identify the G-proteins and other regulatory proteins that couple the D3 receptor to ion channels and characterize the domains of t D3 receptor that interact with these proteins; and (3) Determine the molecular mechanisms underlying I receptor desensitization. These experiments will provide a better understanding of the properties of the I dopamine receptor and the molecular mechanisms by which it modulates neuronal signaling. In doing so, ti study will make significant contributions toward ongoing efforts to define the role of D3 receptor neurological disorders such as schizophrenia.

描述(由申请人提供)： 多巴胺能系统，特别是D3多巴胺受体 与精神分裂症等神经精神障碍的病因学有关 和吸毒成瘾。利用遗传学、分子生物学和分子生物学的研究 药理学技术表明，D3多巴胺受体在 重要角色运动活动和行为影响涉及 增援和奖励。L受体在神经元信号转导中的作用 以及它调节生理和生理功能的分子机制 行为影响还没有被很好地理解。有人建议，D3 受体通过调节离子通道功能发挥表型效应 因此，神经元信号。 我们最近已经证明，人D3受体在ATT-20中表达 神经内分泌细胞系偶联内源性G蛋白内向激活 整流钾(GIRK)通道及其P/Q型钙通道 调节自发的分泌活动。在这个项目中，我们计划 鉴定D3多巴胺受体的特性并鉴定 参与o偶联的受体的结构特征 频道。鉴于D3受体在体内的有限表达谱， 缺乏选择性的药理配体，需要评估其特性 突变的D3受体，本研究将主要使用ATT-2异源 表达系统。此外，为了评估生理相关性，我们还将 原代培养大鼠核内I受体-离子通道偶联的研究 伏隔神经电生理学与单细胞逆转相结合的研究 转录酶-聚合酶链式反应。为了定义D3多巴胺受体的特性，我们 将采用单细胞电生理学、诱变和 表达重组和标记受体。我们将处理三个具体的问题 目的：(1)研究I受体与离子通道的偶联 大鼠伏隔核神经元培养；(2)鉴定G蛋白和 其他将D3受体偶联到离子通道和 确定与这些蛋白质相互作用的tD3受体的结构域； 以及(3)确定I受体的分子机制 脱敏。这些实验将使我们更好地理解 I型多巴胺受体的特性及其分子机制 调节神经信号。在这样做的过程中，技术研究将具有重要意义 对正在进行的确定D3受体作用的努力的贡献 神经紊乱，如精神分裂症。