PROJECT 1 - MOLECULAR ANALYSIS OF NMDA RECEPTOR MODULATORY SITES

项目I - Zhang & Chen项目总监/首席研究员（最后，第一，中间）：Lipton，Stuart A. 项目总结（见说明）： 谷氨酸受体的N-甲基-D-天冬氨酸亚型（NMDAR）对于中枢神经系统（CNS）的正常功能是必需的。然而，NMDAR的过度激活，特别是与突触受体相对的突触外受体的过度激活，至少部分地介导许多神经障碍中的神经元或突触损伤，例如缺氧缺血性脑损伤，以及如最近提出的唐氏综合征。必须在不干扰其正常脑功能的情况下实现对过度NMDAR活性的阻断。我们已经采取了两种方法对NMDAR进行临床耐受的药理学和遗传干预。一种方法是使用美金刚以及NO物质通过S-亚硝基化进一步下调NMDAR。 美金刚胺和NO样物质作用的NMDAR的结构决定因素将在该资助的支持下进一步表征。另一种方法是利用由NRSA和NRSB组成的NMDAR亚基的新家族的抑制作用，通过影响通道通透性来下调NMDAR，在某种意义上模拟也在这里开发的NMDAR拮抗剂药物的作用。我们将研究NRS亚基的MS结构域下调NMDAR活性的作用，并设计基于NRS配体结合结构域（LBD）的筛选试验，以发现调节含NRS受体的新化合物。这些试剂将用于表征含NRS的受体，并可能用于神经保护。 因此，本研究的具体目的如下：1）通过电生理学研究NMDAR的氨基末端结构域（ATD）和LBD之间的松散连接区的S-亚硝基化/氧化还原调节的作用; 2）开发LBD衍生的筛选分析以筛选NMDAR的NRS亚基的选择性配体。这些配体将通过二次测定、化学修饰和共结晶进一步表征和精制; 3）研究来自NRS亚基的外前庭（MS）区的肽对NMDAR渗透性的抑制作用。

Project I - Zhang & Chen Program Director/Principal Investigator (Last, First, Middle): Lipton, Stuart A. PROJECT SUMMARY (See instructions): The N-methyl-D-aspartate subtype of gluatmate receptor (NMDAR) is essential for normal function of the central nervous system (CNS). However, excessive activation of NMDARs, particulariy of extrasynaptic as opposed to synaptic receptors, mediates, at least in part, neuronal or synaptic damage in many neurological disorders, such as hypoxic-ischemic brain injury and, as recently suggested, in Down syndrome. Blockade of excessive NMDAR activity must be achieved without interference with its normal brain function. We have taken two approaches for clinically-tolerated pharmacological and genetic intervention on NMDARs. One approach is to use Memantine but also NO species to further down regulate the NMDAR by S-nitrosylation. The structural determinants on NMDARs for the action of Memantine and NO-like species will be characterized further under the auspices of this grant. Another approach is to utilize the inhibitory effect of a novel family of NMDAR subunits, composed of NRSA and NRSB, to downregulate NMDARs by affecting channel permeability, in a sense mimicking the effect of the NMDAR antagonist drugs that are also being developed here. We will study the role of the MS domain of NRS subunits that downregulate activity of NMDARs and also design NRS ligand-binding domain (LBD)-based screening assays to discover new compounds that modulate NRS-containing receptors. These agents will be useful for characterizing NRS-containing receptors, and possibly for neuroprotection. Accordingly, the Specific Aims of this proposal are as follows: 1) To study the effect of S-nitrosylation/redox modulation of the loose linker region between the amino-terminal domain (ATD) and the LBD of NMDARs by electrophysiology; 2) To develop LBD-derived screening assays to screen for ligands selective for the NRS subunit of the NMDAR. These ligands will be further characterized and refined by secondary assays, chemical modification, and co-crystallization; 3) To study the inhibitory effect of peptides derived from the out vestibule (MS) region of NRS subunits on NMDAR permeability.