EXCITOTOXIN EFFECTS ON SPINAL CORD IN VITRO

体外兴奋毒素对脊髓的影响

• 批准号: 3414064
• 负责人: JOHN R DELFS
• 金额: $ 14.87万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-04-01 至 1993-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3414064
• 关键词: acetylcholinesterase; aminoacid; aspartate; cholinergic receptors; degenerative motor system disease; dosage; glutamate receptor; histochemistry /cytochemistry; inhibitor /antagonist; kynurenate; laboratory rat; lactate dehydrogenases; neural degeneration; neurotoxins; neurotransmitter receptor; somatostatin; spinal cord; stimulant /agonist; synapses; tissue /cell culture

Excitatory amino acids (EAAs) acting at specific membrane receptors may promote neuron degeneration or death in many nervous system diseases and have been specifically implicated in pathogenesis of motor neuron diseases. EAA-mediated damage to anatomically and histochemically defined neurons can be examined in organotypic roller tube cultures of postnatal rat spinal cord (OTC-SCs). Our studies show damage to ventral and dorsal horns in OTC-SCs by the excitotoxin, N-methyl-D-aspartic acid (NMDA). This projects examines actions of NMDA and other EAA agonists on cholinergic markers in spinal cord in vitro using the OTC-SC system. First, OTC-SCs will be treated with NMDA, with or without specific antagonists. Damage to spinal cord cholinergic systems will be assessed by histological and morphometric analysis after staining cultures for acetylcholinesterase activity (AChE) and choline acetyltransferase (CHAT) - immunoreactivity, and by biochemical analysis of ACHE and CHAT activities. Lactate dehydrogenase efflux from cultures will be examined to obtain an overall assessment of neuronal damage. Next, dose-response curves and temporal aspects of the effects will be determined. Third, cultures will be treated with the EAA agonists, kainic acid or quisqualic acid. Using specific NMDA-receptor antagonists or non-NMDA- receptor antagonists, the extent to which damage is mediated through different EAA receptor subtypes will be determined. In OTC-SCs, damage by L-glutamate compared with kainate or NMDA is less than in many culture systems, reflecting more closely its effect in vivo. Therefore, the fourth specific aim is to investigate two possible mechanisms for the decreased effect of glutamate: the role of glutamate uptake and the role of kynurenic acid as an endogenous EAA antagonist. Fifth, we will determine whether a potent somatostatin analogue, SMS 201- 995, can alter the toxic effects of NMDA, L-glutamate, or other excitotoxins on neurons in OTC-SCs. The long-term objective of this research is to understand better the role of excitotoxins in spinal neuron degeneration and death.

作用于特定膜受体的兴奋性氨基酸（EAAs）可能 在许多神经系统疾病中促进神经元变性或死亡 特别涉及运动神经元疾病的发病机制。 EAA 介导的对解剖学和组织化学定义的神经元的损伤可以 在出生后大鼠脊髓的器官型滚管培养物中进行检查 线（OTC-SC）。 我们的研究表明，腹角和背角受损 OTC-SCs 由兴奋毒素 N-甲基-D-天冬氨酸 (NMDA) 产生。 这个项目 检查 NMDA 和其他 EAA 激动剂对胆碱能标记物的作用 使用 OTC-SC 系统进行体外脊髓。 首先，OTC-SC 将用 NMDA 进行处理，无论是否有特定的 对手。 对脊髓胆碱能系统的损害将通过以下方式进行评估 培养物染色后的组织学和形态测量分析 乙酰胆碱酯酶活性 (AChE) 和胆碱乙酰转移酶 (CHAT) - 免疫反应性，并通过 ACHE 和 CHAT 活性的生化分析。 将检查培养物中的乳酸脱氢酶流出以获得 神经元损伤的总体评估。 接下来，剂量反应曲线和 将确定影响的时间方面。 第三，培养物将用 EAA 激动剂、红藻氨酸或 使君子酸。 使用特定的 NMDA 受体拮抗剂或非 NMDA 受体拮抗剂 受体拮抗剂，通过介导损伤的程度 将确定不同的 EAA 受体亚型。 在 OTC-SC 中，与红藻氨酸或 NMDA 相比，L-谷氨酸造成的损害较小 比许多培养系统中的效果更接近地反映了其在体内的作用。 因此，第四个具体目标是研究两种可能的 谷氨酸降低作用的机制：谷氨酸的作用 犬尿酸作为内源性 EAA 拮抗剂的摄取和作用。 第五，我们将确定是否是有效的生长抑素类似物，SMS 201- 995，可以改变 NMDA、L-谷氨酸或其他物质的毒性作用 OTC-SC 中神经元的兴奋毒素。 这项研究的长期目标是更好地了解该角色 脊髓神经元变性和死亡中的兴奋毒素。