CALCIUM HOMEOSTASIS IN HYPOXIC NEURONS

缺氧神经元中的钙稳态

• 批准号: 2259449
• 负责人: Mark Paul Goldberg
• 金额: $ 8.56万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-01-01 至 1996-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2259449
• 关键词: NMDA receptors; alternatives to animals in research; calcium; calcium flux; cell death; cerebral ischemia /hypoxia; confocal scanning microscopy; electrophysiology; fluorescence microscopy; homeostasis; imaging /visualization /scanning; inhibitor /antagonist; laboratory mouse; microelectrodes; neocortex; neural degeneration; neurons; neuropharmacology; tissue /cell culture; video recording system; voltage /patch clamp

Hypoxic-ischemic brain injury from stroke and other cerebral vascular diseases is the leading cause of neurologic disability and mortality. Although no clinical therapy has yet been proven effective in limiting neuronal loss from of acute cerebral ischemia, improved understanding of the cellular mechanisms underlying hypoxic-ischemic brain injury may provide opportunities for rational therapy. Recent evidence supports the hypothesis that a failure of intracellular calcium homeostasis may be central to the development of hypoxic-ischemic neuronal degeneration. The present proposal utilizes two research tools to examine this hypothesis and its consequences. First, a cortical cell culture model of hypoxic-ischemic neuronal loss has been characterized in the sponsor's laboratory; preliminary observations indicate this injury is associated with intracellular calcium accumulation. Second, ratio imaging techniques using the fluorescent calcium indicator, fura-2, will allow direct observation of cytosolic free calcium alterations in hypoxic neurons. Calcium measurements will be paired with assessments of neuronal survival under hypoxic conditions to identify differentially vulnerable cell populations. Pharmacological studies will assess major routes of intracellular calcium entry and clearance, to determine how these are altered by energy depletion and to examine potential therapeutic interventions. Confocal fluorescence microscopy will be utilized to examine the subcellular distribution of calcium changes in hypoxic neurons. Finally, attention will turn to cellular ionic events which occur in concert with calcium alterations, using whole cell patch-clamp and perforated-patch microelectrode physiology and fluorescent indicators for pH, magnesium, and other ions. In addition to research training in the neurosciences, the CIDA program will provide focused experience in teaching, general neurology, and clinical investigation in cerebral vascular disease.

中风和其他脑血管的缺氧 - 缺血性脑损伤 疾病是神经系统残疾和死亡率的主要原因。 尽管尚未证明尚无临床疗法有效限制 急性脑缺血因神经元丧失，对 缺氧 - 缺血性脑损伤的基础机制可能 提供理性治疗的机会。 最近的证据支持了细胞内失败的假设 钙稳态可能是发展的核心 缺氧缺血性神经元变性。 目前的建议使用 两种研究工具，用于检查这一假设及其后果。 首先，低氧缺氧神经元丧失的皮质细胞培养模型 在赞助商的实验室中已被描述；初步的 观察结果表明这种损伤与细胞内有关 钙积累。 其次，使用比率成像技术使用 荧光钙指示剂Fura-2将直接观察 低氧神经元中的无胞质钙改变。 钙测量将与神经元的评估配对 在低氧条件下生存以识别差异脆弱 细胞种群。 药理学研究将评估主要途径 细胞内钙的进入和清除，以确定它们是如何的 通过能量耗尽的改变并检查潜在的治疗性 干预措施。 共聚焦荧光显微镜将用于 检查低氧钙变化的亚细胞分布 神经元。 最后，注意将转向细胞离子事件 使用全细胞贴片钳一致地发生与钙改变发生 和穿孔的微电极生理和荧光 pH，镁和其他离子的指标。 除了神经科学的研究培训外，CIDA计划还 将在教学，一般神经病学和 脑血管疾病的临床研究。