ZINC-DEPENDENT APPARENT DESENSITIZATION NMDA RECEPTORS

谷氨酸是脊椎动物中枢神经系统中应用最广泛的神经递质。NMDA受体是谷氨酸受体的一个亚类，在多种形式的突触可塑性中发挥关键作用，奠定了学习和记忆的基础。NMDA受体还参与多种脑病理过程，包括中风诱导的细胞死亡、癫痫和退行性神经系统疾病。因此，关于NMDA受体功能调控的详细知识不仅对我们理解突触可塑性很重要，而且具有广泛的临床意义。在谷氨酸持续存在的情况下，NMDA受体（特别是NR1/NR2A受体）经历一个称为脱敏的过程，这被认为是其功能的重要调节机制。本实验室的初步数据表明，环境锌除了作用于细胞外高亲和力位点引起电压无关抑制外，还有助于NR1/NR2A受体的明显脱敏。本研究旨在系统地研究锌依赖性脱敏在人工表达系统和神经元中的作用，以便更好地了解这一现象的潜在机制和生理意义。锌依赖性脱敏的一种可能机制是谷氨酸结合位点和细胞外锌位点之间的正变构相互作用。这一假设将通过在HEK 293细胞中表达的重组NR1/NR2A受体进行几项生物物理实验来验证。我们还假设锌依赖性脱敏和甘氨酸非依赖性脱敏是由不同的分子决定因素介导的。一系列嵌合受体和点突变体将被用来验证这一假设。最后，我们还将研究锌对培养神经元中天然NMDA受体脱敏的影响。本研究可能揭示NMDA受体功能调控的新机制，并为锌在脑中的功能作用提供新的认识。此外，它可能为开发新的治疗中风、癫痫和其他神经系统疾病的药物提供有用的信息。

Glutamate is the most widely used neurotransmitter in the vertebrate central nervous system. NMDA receptors, a subclass of receptors for glutamate, play a critical role in multiple forms of synaptic plasticity that lay the foundation for learning and memory. NMDA receptors are also implicated in a variety of pathological processes of the brain, including stroke-induced cell death, epilepsy, and degenerative neurological diseases. Thus, detailed knowledge about regulation of NMDA receptor function not only is important for our understanding of synaptic plasticity, but also has broad clinical implications. In the continuous presence of glutamate, NMDA receptors (in particular NR1/NR2A receptors) undergo a process called desensitization that is thought to be an important regulatory mechanism for their function. Preliminary data from this laboratory suggest that ambient zinc, in addition to acting at extracellular high-affinity site to cause voltage-independent inhibition, contributes to apparent desensitization of NR1/NR2A receptors. The goal of this study is to investigate the zinc- dependent desensitization systematically in artificial expression systems and neurons, so we could better understand the underlying mechanism and physiological implication of this phenomenon. One possible mechanism for zinc-dependent desensitization is a positive allosteric interaction between the glutamate-binding site and the extracellular zinc site. This hypothesis will be tested with several biophysical experiments, using recombinant NR1/NR2A receptors expressed in HEK 293 cells. It has also been postulated by us that the zinc-dependent desensitization and glycine-independent desensitization are mediated by distinct molecular determinants. A series of chimeric receptors and point mutants will be used to test this hypothesis. Finally, the effects of zinc on desensitization of native NMDA receptors in cultured neurons will also be investigated. The proposed study may delineate a new mechanism in the regulation of NMDA receptor function, and provide new insight on the functional roles of zinc in the brain. Furthermore, it may provide useful information for the development of new therapeutic agents for stroke, epilepsy and other neurological diseases.