Molecular and Behavioral Analysis of Dopamine Receptor Function

多巴胺受体功能的分子和行为分析

• 批准号: 8259083
• 负责人: KIM Arthur NEVE
• 金额: --
• 依托单位: PORTLAND VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8259083
• 关键词: Acute; Affinity; Agonist; Automobile Driving; Autoreceptors; Behavior; Behavioral; Binding; Binding Proteins; Biochemical; Biological Availability; Calcium-Binding Proteins; Catecholamines; Catechols; Complex; Cyclic AMP; Cytoskeleton; Data; Development; Disease; Distal; Dopamine; Dopamine D1 Receptor; Dopamine D2 Receptor; Dopamine Receptor; Endosomes; Environment; G-Protein-Coupled Receptors; Goals; Hydrogen Bonding; Hydroxyl Radical; Immunohistochemistry; Libraries; Ligand Binding; Ligands; Lipids; Measures; Mediating; Membrane Microdomains; Mental disorders; Methods; Modeling; Molecular; Motor Activity; Mus; Mutant Strains Mice; Mutation; Nerve; Neurons; Nucleus Accumbens; Parkinson Disease; Pattern; Pharmaceutical Preparations; Pharmacologic Substance; Phosphorylation; Positioning Attribute; Property; Protein Binding; Proteins; Proteomics; Quinpirole; RNA Splicing; Receptor Signaling; Regulation; Research; Role; Schizophrenia; Screening procedure; Serine; Signal Pathway; Signal Transduction; Simplexvirus; Testing; Treatment Efficacy; Two-Hybrid System Techniques; Tyrosine 3-Monooxygenase; Ventral Tegmental Area; abstracting; basal forebrain; base; cDNA Library; dopamine system; dopaminergic neuron; human RIPK1 protein; in vivo; member; mutant; nervous system disorder; neurochemistry; neuropsychiatry; neurotrophic protein S100beta; novel; postsynaptic; presynaptic; programs; radioligand; receptor; receptor expression; receptor function; response; therapeutic target

DESCRIPTION (provided by applicant): Project Summary/Abstract Dopamine receptors are the primary therapeutic targets for a variety of neurological and psychiatric disorders, including schizophrenia and Parkinson's disease. The five subtypes of dopamine receptors (D1- D5) are members of the superfamily of G protein-coupled receptors. The D1 and D2 subtypes are the most abundant and mediate most classic dopamine-dependent behaviors. The overall long-term goal of this research program is to delineate and distinguish between D1 and D2 receptor mechanisms, from the initial step of ligand binding to the receptor to the final manifestation of dopamine-dependent behaviors. A secondary aim is to assess the differential functional roles of the alternatively spliced forms of the D2 receptor, D2S and D2L. The first specific aim is driven by the hypothesis that D1 receptor agonists, but not D2 agonists, must have a catechol ring because of distinct residues at two positions in transmembrane helices 6 and 7. We will characterize reciprocal mutants in which one or both of these residues in one receptor subtype is changed to the corresponding residue(s) of the other subtype. We will determine the effect of the mutations on high- and low-affinity binding of and regulation of cyclic AMP accumulation by catecholamine and non-catecholamine agonists. The second specific aim is based on the hypothesis that activation of D1 and D2 receptors regulates their localization in lipid rafts and differentially regulates the abundance of other raft-enriched proteins. We propose to examine the distribution of the receptors in non- raft fractions and the effect of acute or repeated agonist treatment on receptor distribution. In addition, proteomic analyses of raft fractions will be used to identify drug-induced changes in other raft proteins. The third specific aim will test the hypotheses that D2L is the postsynaptic D2 receptor that mediates increased locomotor activity and D2S is the presynaptic autoreceptor that regulates dopamine release and tyrosine hydroxylase activity in the mouse basal forebrain. We will evaluate the behavioral and neurochemical effects of HSV-mediated expression of D2L or D2S in the nucleus accumbens (postsynaptic) or ventral tegmental area (presynaptic) of D2 receptor null-mutant mice. The fourth specific aim is driven by the hypotheses that S100B binding to the D2 receptor is necessary for most efficient function, and that there are D1 and D2 receptor-interacting proteins still to be identified that regulate dopamine receptor signaling and localization. We will carry out additional rounds of cDNA library screening with D1 and D2 receptor intracellular loops to identify new dopamine receptor-interacting proteins and investigate the consequences of the binding of these proteins and S100B to dopamine receptors for the subcellular localization and signaling properties of the receptors. We hope that, by identifying structural features of D1 and D2 receptors that interact with ligands in a subtype-selective manner to determine binding and activation of signaling pathways, dissecting the contributions of dopamine receptor subtypes to biochemical and behavioral responses to dopamine and synthetic agonists, and characterizing novel proteins that regulate dopamine receptor signaling, these studies will contribute to the development of more selective pharmaceutical treatments of neuropsychiatric disorders that involve the dopamine system. PUBLIC HEALTH RELEVANCE: Project Narrative Each year, VA Medical Centers hospitalize about 45,000 veterans for treatment of schizophrenia and treat at least 40,000 Parkinson's disease patients. The causes of the diseases are unknown, but changes in the density of dopamine D2-like receptors have been implicated in the pathophysiology of the schizophrenia, and the primary treatments for the diseases are blockade or stimulation of dopamine receptors. Analysis of endogenous and recombinant dopamine receptors expressed in neurons and in mammalian cell lines, and of the function of dopamine receptors in the behaving rodent, are powerful model systems with which to perform basic research that should contribute to the development of rational strategies for improved treatment of schizophrenia and movement disorders.

描述(由申请人提供)： 项目摘要/摘要多巴胺受体是各种神经和精神疾病的主要治疗靶点，包括精神分裂症和帕金森氏病。多巴胺受体的五个亚型(D1-D5)是G蛋白偶联受体超家族的成员。D_1和D_2亚型是最丰富和最典型的多巴胺依赖行为。该研究计划的总体长期目标是描绘和区分D1和D2受体的机制，从配体与受体结合的初始步骤到多巴胺依赖行为的最终表现。第二个目的是评估D2受体、D2S和D2L选择性剪接形式的不同功能作用。第一个特定目的是假设D1受体激动剂而不是D2受体激动剂必须有一个儿茶酚环，因为在跨膜螺旋6和7的两个位置上有不同的残基。我们将描述在一个受体亚型中的一个或两个残基改变为另一个亚型的相应残基(S)的互易突变体。我们将确定突变对儿茶酚胺和非儿茶酚胺激动剂高亲和力和低亲和力结合和调节环状AMP积累的影响。第二个特定的目的是基于这样的假设，即D1和D2受体的激活调节它们在脂筏中的定位，并以不同的方式调节其他RAFT丰富的蛋白的丰度。我们建议检查受体在非RAFT组分的分布，以及急性或重复的激动剂治疗对受体分布的影响。此外，RAFT组分的蛋白质组学分析将用于识别药物引起的其他RAFT蛋白的变化。第三个特定目标将测试假设，即D2L是突触后D2受体，介导增加的运动活动，而D2S是突触前自体受体，调节小鼠基底前脑中多巴胺的释放和酪氨酸羟基酶的活性。我们将评估单纯疱疹病毒介导的D2L或D2S在D2受体缺失突变小鼠伏核(突触后)或腹侧被盖区(突触前)表达的行为和神经化学效应。第四个特定的目标是假设S100B与D2受体结合是最有效的功能所必需的，并且仍然存在调节多巴胺受体信号和定位的与D1和D2受体相互作用的蛋白。我们将利用D1和D2受体细胞内环进行额外一轮的cDNA文库筛选，以寻找新的与多巴胺受体相互作用的蛋白，并研究这些蛋白和S100B与多巴胺受体结合对受体的亚细胞定位和信号特性的影响。我们希望，通过识别与配体以亚型选择性的方式相互作用的D1和D2受体的结构特征来确定信号通路的结合和激活，剖析多巴胺受体亚型对多巴胺和合成激动剂的生化和行为反应的贡献，以及表征调节多巴胺受体信号的新蛋白质，这些研究将有助于开发更具选择性的药物治疗涉及多巴胺系统的神经精神疾病。 公共卫生相关性： 项目简介每年，退伍军人医疗中心为大约45,000名退伍军人治疗精神分裂症，并治疗至少40,000名帕金森氏症患者。这些疾病的病因尚不清楚，但多巴胺D2样受体密度的变化与精神分裂症的病理生理有关，治疗这些疾病的主要方法是阻断或刺激多巴胺受体。分析神经元和哺乳动物细胞中表达的内源性和重组多巴胺受体，以及行为啮齿动物中多巴胺受体的功能，是进行基础研究的强大模式系统，应有助于开发合理的策略，以改进精神分裂症和运动障碍的治疗。