EXCITOTOXIC MECHANISMS OF ETOH--NMDA RECEPTOR FUNCTION

乙醇的兴奋性毒性机制--NMDA受体功能

• 批准号: 2615897
• 负责人: MARY LOU VALLANO
• 金额: $ 7.54万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-06-01 至 2000-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2615897
• 关键词: NMDA receptors; biological signal transduction; brain mapping; calmodulin dependent protein kinase; dentate gyrus; drug withdrawal; enzyme activity; ethanol; glutamates; hippocampus; laboratory rat; mature animal; neurotoxins; polymerase chain reaction; tissue /cell culture; western blottings

DESCRIPTION: Evidence from several independent laboratories using different experimental approaches indicates that acute ethanol selectively inhibits N-methyl-D-aspartate receptor (NR) function at pharmacologically relevant concentrations, whereas chronic ethanol produces NR supersensitivity due, at least in part, to receptor upregulation. As a consequence, excessive activation of NRs during alcohol withdrawal is thought to contribute to the associated seizures, autonomic instability and neurotoxicity. Pharmacologic treatment with available NR antagonists is undesirable due to generalized inhibition of these ubiquitous receptors. However, NRs are functionally diverse due to differences in subunit composition and associated signal transduction systems. This diversity may account for the selective effects of ethanol on NRs in different brain regions, and in different neuronal populations within a particular region. By exploring the mechanistic basis for this regional selectivity, it should be possible to design and utilize in the alcoholic patient more selective antagonists that are targeted to vulnerable neuronal populations. To further define the role of NRs in chronic ethanol-induced neurotoxicity in hippocampus, the following hypotheses will be tested in adult rats: (I) increased levels of specific NR subunits, and possibly mRNAs, will be observed in hippocampal subfields (CA1, CA3, DG) of ethanol-treated rats, relative to pair-fed control rats, in a manner that corresponds to development of ethanol dependence. Moreover, increased levels of NR subunits will be positively correlated with increased expression or activity of a type II Ca2+/calmodulin-dependent protein kinase (CaM KII) that is co-localized with NRs at excitatory synapses and has been implicated in other forms of excitotoxicity; (ii) chronic ethanol ingestion will differentially sensitize CA1, CA3 and DG neurons to NR-mediated excitotoxicity, relative to pair-fed controls, in a reversible manner. This effect will be mediated by CaM KII. To test these hypotheses, ethanol will be administered to rats for 1-12 weeks and, in some cases, will include a withdrawal period. In ai-m 1, hippocampal subfields from ethanol-treated and control rats will be compared using NRI, NR2 and CaM KII immunoblotting and RT-PCR assays. In aim 2, hippocampal slices will be compared for regional differences in vulnerability to glutamate- or NMDA-mediated excitotoxic damage using a "live-dead" assay in conjunction with confocal imaging analysis.

描述：使用不同的几个独立实验室的证据 实验方法表明急性乙醇有选择地抑制 N-甲基-D-天冬氨酸受体（NR）功能在药理学相关 浓度，而慢性乙醇在应有的 至少部分是受体上调。 结果，过多 据认为，戒酒期间NR的激活有助于 相关的癫痫发作，自主不稳定和神经毒性。 药理学 可用的NR拮抗剂的治疗由于广义而不受欢迎 抑制这些无处不在的受体。 但是，NR在功能上是 由于亚基组成和相关信号的差异而导致的多样 转导系统。 这种多样性可能解释了选择性效果 在不同大脑区域和不同神经元中NRS上的乙醇的 特定区域内的种群。 通过探索机械基础 对于这种区域选择性，应该可以设计和利用 在酒精患者中，更多的选择性拮抗剂针对 脆弱的神经元种群。 进一步定义NRS在 慢性乙醇诱导的海马神经毒性，以下 假设将在成年大鼠中进行检验：（i）特异性水平提高 NR亚基，可能是mRNA，在海马子场中观察到 （CA1，CA3，DG）的乙醇处理大鼠，相对于配对的对照大鼠， 以与乙醇依赖性发展相对应的方式。 此外，NR亚基的水平增加将与 II型Ca2+/钙调蛋白依赖蛋白的表达或活性增加 蛋白激酶（CAM KII）在兴奋性时与NRS共定位 突触并与其他形式的兴奋性毒性有关； （ii） 慢性乙醇摄入将差异化CA1，CA3和DG 在A中，神经元与NR介导的兴奋性毒性相对于配对对照组 可逆的方式。 这种效果将由CAM KII介导。 测试这些 假设，将乙醇给大鼠施用1-12周，在某些人中 案件将包括撤回期。 在AI-M 1中，海马子场 使用NRI，NR2和 CAM KII免疫印迹和RT-PCR分析。 在AIM 2中，海马切片将 比较在谷氨酸或 NMDA介导的兴奋毒性损伤，使用“活死”测定 通过共聚焦成像分析。