Dynorphin Modulation of N-Methyl-D-Aspartate (NMDA) Receptor Function

描述(申请人提供)：脊髓损伤是一种灾难性的事件，极大地改变了受害者的生活方式，使个人终身行动不便，反复出现的医疗问题和慢性神经性疼痛。损伤的最终严重程度不仅取决于蹦床最初造成的损伤，还取决于一系列生化事件，这些事件在几天的过程中进一步侵蚀脊髓的完整性。其中一条途径会导致阿片肽强啡肽的浓度和释放急剧增加。强啡肽是一种神经毒肽，通过激活兴奋性氨基酸受体的N-甲基-D-天冬氨酸(NMDA)亚类来损伤脊髓中的神经元。强啡肽造成的损害可以在未受创伤的脊髓中产生症状，从慢性异位痛(疼痛到通常不痛的刺激)到完全瘫痪。因此，强啡肽可能在创伤后脊髓损伤中起重要作用。这个项目的最终目标是了解强啡肽是如何导致脊髓损伤的。 强啡肽激活NMDA受体的机制目前尚不清楚。在这项提议中，将要检验的假设是强啡肽激活低活性或静止的NMDA受体，该受体缺乏NR1亚单位(NR1a)的外显子-5编码区。全细胞膜片钳实验将在培养的脊髓神经元、脊髓切片和已知的NMDA受体亚单位组合转染的细胞系中进行。这些实验将证明：1)强啡肽增强NMDA受体亚群的活性，2)NR1a亚单位是强啡肽增强NMDA受体功能的关键，3)含有NMDA受体的低活性NR1a是强啡肽增强脊髓突触活性的原因。这些实验将确定强啡肽的神经毒性机制，并确定在脊髓损伤的关键创伤后时期进行干预的潜在药理靶点。

DESCRIPTION (provided by applicant): Spinal cord injury is a catastrophic event that dramatically alters the lifestyle of the victim and subjects the individual to a lifetime of reduced mobility, recurrent medical problems and chronic neuropathic pain. The ultimate severity of the injury is not only determined by the initial damage produced by the tramna, but also by a cascade of biochemical events that further erodes the integrity of the spinal cord over the course of several days. One of these pathways leads to a dramatic increase in the concentration and release of the opioid peptide dynorphin. Dynorphin is a neurotoxic peptide that damages neurons in the spinal cord by activating the N-methyl-D-aspartate (NMDA) subclass of excitatory amino acid receptors. The damage produced by dynorphin can produce symptoms in non-traumatized spinal cord ranging from chronic allodynia (pain to normally non-painful stimuli) to full paralysis. Thus, it is likely that dynorphin contributes significantly to the post trauma spinal cord damage. The ultimate goal of this project is to understand how dynorphin contributes to spinal cord injury. The mechanism by which dynorphin activates NMDA receptors is currently not known. In this proposal the hypothesis that will be tested is that dynorphin activates low activity or quiescent NMDA receptors that lack the exon-5 coded region of the NR1 subunit (NRla). Whole cell patch clamp experiments will be conducted in cultured spinal cord neurons, spinal cord slices, and cell lines transfected with known combinations of NMDA receptor subunits. These experiments will demonstrate that 1) dynorphin enhances the activity of a subset of NMDA receptors, 2) the NRla subunit is critical for dynorphin enhancing NMDA receptor function, and 3) the low activity NRla containing NMDA receptors are responsible for dynorphin's enhancement of synaptic activity in the spinal cord. These experiments will identify the mechanism for dynorphin's neurotoxicity as well as identify potential pharmacological targets for intervention in the critical post trauma period of spinal cord injury.