NMDA RECEPTORS

NMDA受体

• 批准号: 6563124
• 负责人: MICHAEL D BROWNING
• 金额: $ 18.46万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-12-01 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6563124
• 关键词: NMDA receptors; behavioral /social science research tag; biological signal transduction; ethanol; laboratory mouse; neuropharmacology; nitric oxide; posttranslational modifications; protein structure function; psychopharmacology; receptor expression; sleep disorders

Component 6 will focus on the role of one type of glutamate receptor, the N-methyl-D-aspartate receptor (NMDAR), in initial sensitivity to ethanol. Ethanol inhibits NMDAR function, and NMDARs play an important role in brain function. We have observed that long-Sleep (LS) and Short-Sleep (SS) mice, which differ in their initial sensitivity to the hypnotic effects of ethanol, show marked behavioral differences to the NMDAR antagonists MK- 801 and CPP. SS mice exhibited greater locomotor activation than LS mice at lower drug doses; and LS, but not SS, mice were sedated at higher drug doses. We have also found line differences in the density of [3H]MK-801 binding sites and ratio of NMDAR subunits (NR2A:NR2B) in LS and SS mouse brains. Therefore, this proposal focuses on the NMDAR as a candidate gene for differential ethanol sensitivity in the inbred lines of LS (ILS) and SS (ISS) mice. They hypothesis being tested is that: Differences between ILS/ISS in initial sensitivity to ethanol may be due to differences in NMDAR expression, composition, post-translational processing and/or signal transduction. Quantitative autoradiographic analyses of [3H]MK-801 and [3H]CPP binding and quantitative western analysis of NR1, NR2A, NR2B and NR2C subunits will be used to determine whether differences in NMDAR expression are localized to specific regions of the ILS and ISS mouse brains, particularly those involved in locomotor activity. In order to assess whether ILS and ISS mouse brain NMDARs contribute to the differential ethanol sensitivity of these mice, behavioral studies with MK-801 and CPP will be conducted using the LSxSS recombinant inbred strains. Next, in vivo electrochemical measures of NMDA-induced nitric oxide generation in intact brain and in vitro measures of NMDA induced electrophysiological activity in brain slices will be used to determine whether differences also exist in NMDAR signal transduction and in ethanol's inhibition of NMDAR activity in ILS and ISS mouse brain. Lastly, the contribution of potential differences in NMDAR subunit composition and/or phosphorylation state in specific regions of ILS and ISS brain will be investigated using co-immunoprecipitation assays, quantitative western analyses and front phosphorylation assays. The results of these experiments will increase our understanding of how excitatory glutamate systems contribute to genetic differences in initial sensitivity to ethanol. Understanding the factors mediating initial ethanol sensitivity is important because low initial sensitivity has been found to be associated with high risk for alcoholism.

组件6将集中在一种类型的谷氨酸受体的作用， N-甲基-D-天冬氨酸受体（NMDAR），对乙醇的初始敏感性。 乙醇抑制NMDAR功能，并且NMDAR在以下方面发挥重要作用： 大脑功能我们观察到长睡眠（LS）和短睡眠（SS） 小鼠，它们对催眠作用的初始敏感性不同， 乙醇，表现出显着的行为差异，以NMDAR拮抗剂MK- 801和CPP。SS小鼠比LS小鼠表现出更大的运动激活 在较低的药物剂量下; LS，但不是SS，小鼠在较高的药物剂量下被镇静 剂量我们还发现了[3 H]MK-801密度的谱线差异 LS和SS小鼠中NMDAR亚基（NR 2A：NR 2B）的结合位点和比率 大脑因此，本建议将NMDAR作为候选基因 对于LS（ILS）的近交系中的差异乙醇敏感性， SS（ISS）小鼠。他们的假设正在测试的是： ILS/ISS对乙醇的初始敏感性可能是由于 NMDAR表达、组成、翻译后加工和/或信号 转导[3 H]MK-801和 [3 H]CPP结合和NR 1、NR 2A、NR 2B和 NR 2C亚基将用于确定NMDAR的差异是否 表达定位于ILS和ISS小鼠的特定区域 大脑，特别是那些参与运动活动的大脑。为了 评估ILS和ISS小鼠脑NMDAR是否有助于 这些小鼠的不同乙醇敏感性， MK-801和CPP将使用LSxSS重组近交系进行 菌株接下来，NMDA诱导的一氧化氮的体内电化学测量 完整脑中的氧化物生成和NMDA诱导的体外测量 脑切片中的电生理活动将用于确定 是否在NMDAR信号转导和 乙醇对ILS和ISS小鼠脑中NMDAR活性的抑制。最后， NMDAR亚基组成中电位差异贡献 和/或ILS和ISS脑的特定区域中的磷酸化状态将 使用免疫共沉淀试验、定量Western 分析和前磷酸化测定。的结果予以 实验将增加我们对兴奋性谷氨酸 系统有助于遗传差异的初始敏感性， 乙醇了解介导初始乙醇敏感性的因素 是重要的，因为已经发现低初始灵敏度 与酗酒的高风险有关。