SYNAPTIC TARGETING AND CLUSTERING AND NMDA RECEPTORS

描述：(摘自申请人的摘要) 神经元之间的快速通信是通过以下作用实现的 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。通过监管 谷氨酸门控离子通道神经元的活性和膜密度 改变他们突触输入的强度以响应发育线索 和感官刺激。谷氨酸活性不足和过多 受体被认为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，应该有助于 设计合理的治疗策略治疗神经病学和骨髓炎 精神疾病。 这项提议的目标是了解神经元是如何组装和 调节兴奋性突触的突触后分子机制。 特别是，拟议的研究将确定哪些分子机制 调节N-甲基-D-天冬氨酸的突触靶向性和聚集性 (NMDA)类离子亲性谷氨酸受体。我们最近确认了 N-甲基-D-天冬氨酸受体亚单位NR1的胞质结构域 靶向和受体失活。NR1的这些区域被认为是 通过与细胞骨架蛋白和信号相互作用来调节它们的作用 分子。进一步阐明NMDA受体胞浆功能 结构域，我们将首先识别与NR1相互作用的新分子 亚基使用酵母双杂交筛选和生化方法。 第二，这些相互作用对NMDA受体通道活性和 突触定位将直接通过电生理和 免疫细胞化学方法。第三，NMDA受体靶向基序将是 标记野生型和突变型NR1的荧光定位鉴定 NR2亚单位胞浆结构域在培养神经元中表达。最后， 突触活性对细胞蛋白与天然蛋白相互作用的影响 NMDA受体将使用共价交联技术进行评估。 总之，拟议中的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.