Mechanism and physiology of NMDA receptor desensitization

NMDA受体脱敏机制和生理学

• 批准号: 9765414
• 负责人: Kelvin Chan
• 金额: $ 3.44万
• 依托单位: STATE UNIVERSITY NEW YORK STONY BROOK
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-27 至 2021-09-26
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9765414
• 关键词: AMPA Receptors; Acute; Address; Adopted; Affect; Agonist; Binding; Biophysical Process; Brain; Brain Diseases; Charge; Chronic; Clinical; Cysteine; Data; Development; Electrophysiology (science); Event; Frequencies; Gated Ion Channel; Glutamate Receptor; Glutamates; Glycine; Hippocampus (Brain); Image; Individual; Ion Channel; Ion Channel Gating; Kainic Acid Receptors; Kinetics; Knowledge; Learning; Ligand Binding Domain; Ligands; Light; Long-Term Depression; Long-Term Potentiation; Mediating; Memory; Mental disorders; Modality; Molecular Conformation; Mus; Mutation; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; NMDA receptor A1; Nervous system structure; Neurotransmitters; Pathway interactions; Patients; Physiological; Physiology; Play; Process; Property; Recombinants; Recovery; Role; Schizophrenia; Signal Transduction; Site; Slice; Structure; Synapses; Synaptic plasticity; System; Techniques; Technology; Testing; Transfection; Transmembrane Domain; base; crosslink; desensitization; dimer; experimental study; gene gun; glutamatergic signaling; hippocampal pyramidal neuron; insight; mutant; neuropsychiatric disorder; neuropsychiatry; novel; novel therapeutics; patch clamp; protein function; receptor; tool; two-photon; unnatural amino acids

PROJECT SUMMARY NMDA receptor hypofunction has been implicated in neuropsychiatric disorders such as schizophrenia, a disabling mental disorder that affects over 2.2 million patients in the US. One modality for NMDA receptor hypofunction is aberrant gating, the process where the receptor converts glutamate binding into opening of the associated ion channel. One pathway to regulate hypofunction is NMDA receptor desensitization, a gating configuration where the receptor is ligand-bound, but the ion channel is non-conducting. However, its physiological and mechanistic basis is poorly defined in part because of the lack of tools to study it. In this proposal I will take advantage of new tools – single site mutations that selectively alter specific features of NMDA receptor desensitization recently discovered in the Wollmuth lab – as well as cutting edge technologies to address the mechanism and physiology of NMDA receptor desensitization. Whereas desensitization in non-NMDA receptors depends almost exclusively on the rearrangement of the ligand-binding domain (LBD) dimer interface, such a mechanism is less significant in NMDA receptors and all domains, most notably the transmembrane domain (TMD), have been implicated in NMDA receptor desensitization. In Aim 1, I will test the general hypothesis that the mechanism of NMDA receptor desensitization is fundamentally different from that of non-NMDA receptors, depending strongly on the conformation of the ion channel. To test this hypothesis, I will take advantage of newly identified single-site mutations, patch clamp electrophysiology as well as techniques to regulate the subunit composition of NMDA receptors and light-activated unnatural amino acids. These experiments will help define the structural mechanisms that govern NMDA receptor desensitization, aiding in the development of novel therapeutics that can selectively modulate NMDA receptor activity by specifically targeting desensitization. In Aim 2, I will test the hypothesis that NMDA receptor desensitization leads to decreased excitatory signaling at synapses during high activity. Indeed, my preliminary data show that fast applications of glutamate caused a higher degree of current decay in these mutants, suggesting a potential role of desensitization in NMDA hypofunction. To test this hypothesis more rigorously, I will express mutant NMDA receptors that have altered desensitization properties in organotypic hippocampal slice cultures to address how they change synaptic dynamics including Ca2+ influx and synaptic plasticity. These experiments will address how NMDA receptor desensitization contributes to synaptic physiology. The information gained by the experiments in this proposal will provide insight into the mechanism of NMDA receptor desensitization and its role in synaptic dynamics. My experiments will aid in the development of novel therapeutics that selectively modulates NMDA receptor activity by specifically targeting desensitization.

项目摘要 NMDA受体功能障碍已在神经精神疾病（如精神分裂症）中隐含 在美国影响超过220万患者的精神障碍。 NMDA接收器的一种方式 功能障碍是异常的门控，接收器将谷氨酸结合到开口的过程 相关的离子通道。一种调节功能功能功能功能的途径是NMDA受体脱敏，A 接收器与配体结合的门控配置，但离子通道是无导体的。但是，它 生理和机械基础的定义很差，部分原因是缺乏研究它的工具。在这个 建议我将利用新工具 - 单个站点突变，这些突变有选择地改变 NMDA受体脱敏最近在Wollmuth实验室中发现的 - 解决NMDA受体脱敏的机制和生理的技术。 而非NMDA接收器中的脱敏几乎完全取决于重排 在配体结合域（LBD）二聚体界面中，这种机制在NMDA接收器中不那么重要 所有域，最著名的是跨膜域（TMD），已在NMDA接收器中暗示 脱敏。在AIM 1中，我将测试NMDA接收器机制的一般假设 脱敏与非NMDA接收器的脱敏性不同，具体取决于 离子通道的构象。为了检验这一假设，我将利用新确定的单位点 突变，斑块夹电生理学以及调节亚基组成的技术 NMDA受体和光激活的非天然氨基酸。这些实验将有助于定义结构 控制NMDA受体脱敏的机制，有助于开发新的治疗 这可以通过特异性靶向脱敏来选择性地调节NMDA受体活性。 在AIM 2中，我将检验以下假设：NMDA受体脱敏导致兴奋信号降低 高活动期间的突触。确实，我的初步数据表明，谷氨酸的快速应用引起 这些突变体的当前衰变程度更高，表明脱敏在NMDA中的潜在作用 功能障碍。更严格地检验该假设，我将表达具有 有机海马切片培养物中的脱敏特性的改变，以解决它们如何改变 突触动力学，包括CA2+影响和突触可塑性。这些实验将解决NMDA 受体脱敏有助于突触生理。 实验在本提案中获得的信息将提供有关机制的洞察力 NMDA受体脱敏及其在突触动力学中的作用。我的实验将有助于 开发新型疗法，该疗法有选择地调节NMDA受体活性 靶向脱敏。