SYNAPTIC TARGETING AND CLUSTERING AND NMDA RECEPTORS

描述：（来自申请人摘要） 神经元之间的快速通讯是通过以下动作完成的 突触后受体上的神经递质。在兴奋性突触，这些 受体属于离子型谷氨酸受体家族。通过调节 谷氨酸门控离子通道、神经元的活性和膜密度 根据发育线索改变突触输入的强度 和感官刺激。谷氨酸盐活性不足和过量 受体与精神疾病和神经缺陷有关。 确定谷氨酸受体调节的分子特征对于 我们对正常兴奋性突触功能的理解， 设计合理的治疗策略， 精神病 这项提议的目的是了解神经元如何组装， 调节兴奋性突触的突触后分子机制。 特别是，拟议的研究将确定分子机制， 调节N-甲基-D-天冬氨酸的突触靶向和聚集 （NMDA）类离子型谷氨酸受体。我们最近发现 NMDA受体亚基NR 1的胞质结构域对受体 靶向和受体失活。NR 1的这些区域被认为是 通过与细胞骨架蛋白和信号传导相互作用来介导它们的作用 分子。为了进一步阐明NMDA受体胞浆内的功能， 结构域，我们将首先确定与NR 1相互作用的新分子 使用酵母双杂交筛选和生物化学方法来鉴定亚基。 第二，这些相互作用对NMDA受体通道活性的影响， 突触定位将直接通过电生理学测试， 免疫细胞化学方法。第三，NMDA受体靶向基序将是 使用标记的野生型和突变型NR 1的荧光定位鉴定 和NR 2亚基胞质结构域在培养的神经元中表达。最后 突触活动对细胞蛋白质与天然蛋白质相互作用的影响 将使用共价交联技术评估NMDA受体。 这些实验将阐明基本的分子机制 潜在的突触传递和突触可塑性。

DESCRIPTION: (from applicant's abstract) Rapid communication between neurons is accomplished by the action of neurotransmitters on postsynaptic receptors. At excitatory synapses, these receptors belong to the family of ionotropic glutamate receptors. By regulating the activity and membrane density of glutamate-gated ion channels, neurons alter the strength of their synaptic inputs in response to developmental cues and sensory stimuli. Both inadequate and excessive activity of glutamate receptors has been linked to psychiatric disease and neurological deficit. Determining molecular features of glutamate receptor regulation is crucial for our understanding of normal excitatory synapse function, and should facilitate the design of rational strategies for the treatment of neurologic and psychiatric disease. The objective of this proposal is to understand how neurons assemble and regulate the postsynaptic molecular machinery present at excitatory synapses. In particular, the proposed research will determine molecular mechanisms which regulate the synaptic targeting and clustering of the N-methyl-D-aspartate (NMDA) class of ionotropic glutamate receptors. We have recently identified cytoplasmic domains of the NMDA receptor subunit NR1 important for receptor targeting and receptor inactivation. These regions of NR1 are thought to mediate their effects by interacting with cytoskeletal proteins and signaling molecules. To further elucidate the function of NMDA receptor cytoplasmic domains, we will first identify novel molecules which interact with NR1 subunits using both yeast two-hybrid screening and biochemical approaches. Second, the effects of these interactions on NMDA receptor channel activity and synaptic localization will be directly tested by electrophysiological and immunocytochemical methods. Third, NMDA receptor targeting motifs will be identified using fluorescent localization of tagged wild-type and mutant NR1 and NR2 subunit cytoplasmic domains expressed in cultured neurons. Finally, the effect of synaptic activity on the interaction of cellular proteins with native NMDA receptors will be assessed using covalent crosslinking techniques. Together, the proposed experiments will elucidate basic molecular mechanisms underlying synaptic transmission and synaptic plasticity.