Structure-Function Analysis of the D3 Dopamine Receptor

D3 多巴胺受体的结构功能分析

• 批准号: 6528602
• 负责人: ELDO V KUZHIKANDATHIL
• 金额: $ 23.55万
• 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/6528602
• 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， 与神经精神疾病如精神分裂症的病因有关 和毒瘾利用遗传学、分子生物学和 药理学技术表明，D3多巴胺受体起作用， 重要的作用运动活动和行为影响， 强化和奖励。然而，L受体在神经元信号传导中的作用 以及它调节生理和 行为效应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其表型效应， 从而产生神经元信号。 我们最近已经证明，在AtT-20中表达的人D3受体， 神经内分泌细胞系偶联内源性G蛋白向内激活 整流钾（GIRK）通道和P/Q型钙通道 调节自发分泌活动。在这个项目中，我们计划 表征D3多巴胺受体的特性并识别 涉及偶联到O的受体的结构特征 渠道鉴于D3受体在体内的有限表达谱， 缺乏选择性的药理学配体，以及需要评估 突变的D3受体，这项研究将主要使用AtT-2异源 表达系统此外，为了评估生理相关性，我们还将 在原代培养的大鼠核团中研究I受体-离子通道的偶联 使用电生理学和单细胞逆转录相结合的方法 转录酶-PCR。为了定义D3多巴胺受体的特性，我们 将采用诸如单细胞电生理学、诱变和 表达重组体和标记的受体。我们将讨论三个具体的 目的：（1）研究I受体与初级支气管平滑肌细胞离子通道的偶联 （2）G蛋白的鉴定和G蛋白的表达; 其他将D3受体与离子通道偶联的调节蛋白， 表征与这些蛋白质相互作用的t D3受体的结构域; （3）确定I受体的分子机制 脱敏这些实验将使我们更好地了解 I多巴胺受体的性质及其分子机制， 调节神经信号。在这样做的时候，这项研究将使 为确定D3受体作用的持续努力做出贡献 神经系统疾病，如精神分裂症。