CENTER FOR CNS INJURY STUDIES

本申请提出了四个项目和核心设施进行调研 中枢神经系统损伤的机制。方案1脊髓实验治疗 受伤。我们最近发现环孢素A(CsA)可以减少反应， 释放血管扩张和炎症物质，并调节 神经递质受体和释放。我们建议研究一下 钙调神经磷酸酶在脊髓损伤中的作用这些实验将比较 CsA、FK506(另一种钙调神经磷酸酶抑制剂)单独和在 与雷帕霉素(K506(另一种钙调神经磷酸酶)的拮抗剂)联合 抑制剂)，单独或与雷帕霉素(FK506的拮抗剂)联合使用 钙调神经磷酸酶抑制)，对脊髓离子移位、损伤体积、 运动恢复、血流和5-羟色胺、GABA和谷氨酸诱导 脊椎兴奋性和损伤。项目2.大脑中的酸碱平衡 受伤。损伤组织中pH变化的机制和后果 都没有被很好地理解。我们将测量pH、二氧化碳、HCO3和CO32 脑缺血区和损伤脊髓的缓冲作用 酸中毒、酸中毒和碳酸氢酶在pH值中的变化。我们 认为pH变化调节NMDA受体和间质 缓冲会影响扩散抑郁的速度和程度，并 脑片缺血性损伤。使用导通选择性微电极， 电压钳和分光光度的方法，我们将检验这一假设 电压相关的Na：HCO3共转运控制着H+、Na+和HCO3- 正常血糖和高血糖患者星形胶质细胞膜的分布 缺血症。 项目3：中枢神经系统损伤中的抗坏血酸和谷胱甘肽。我们假设 抗坏血酸(ASC)和谷胱甘肽(GSH)的丢失对脑细胞有贡献 易受氧化损伤。细胞外ASC将在 缺氧性脑片及中脑缺血后大脑皮层 动脉闭塞。细胞内ASC将从细胞外估计 体积和总组织水平。初步研究表明，ASC和 GSH分别定位于神经元和星形胶质细胞，ASC和 GSH随年龄增长而下降，女性较低。我们会确认这些 结果，确定低氧和年龄是否损害ASC动态平衡， 评估ASC和GSH对缺氧性脑片的保护作用，确定 缺血时ASC和GSH丢失的机制及其相关的组织损伤 男性和女性均有ASC丢失。 项目4.缺血中的水区划。这样做的目的是 该项目是为了了解水是如何在细胞外和细胞内转移的 缺血脑组织中的脑室。水的变化将被评估为 细胞外示踪剂、离子选择性微电极和称重干燥 方法。我们将操纵细胞外离子水平并阻止离子 运输和谷氨酸受体。水的运动将在#年进行调查。 上了年纪的动物的切片。Na：HCO3共转运的作用将是 评估过了。光散射和Na-K变化将与干湿法进行比较 体重和示踪剂测量。

This application proposes four projects and core facilities to investigate mechanisms of CNS injury. Project 1 Experimental Therapy of Spinal Cord Injury. We recently discovered that cyclosporin A (CsA) reduces responses, releases vasodilatory and inflammatory substances, and modulates neurotransmitter receptors and release. We propose to study the role of calcineurin in spinal cord injury. The experiments will compare the effects of CsA, FK506 (another calcineurin inhibitor), alone and in combination with rapamycin (an antagonist of K506 (another calcineurin inhibitor), alone and in combination with rapamycin (an antagonist of FK506 inhibition of calcineurin), on spinal cord ionic shifts, lesion volumes, motor recovery, blood flow, and serotonin-,GABA- and glutamate-induced spinal excitability and injury. Project 2. Acid-base homeostasis in brain injury. The mechanisms and consequences of pH changes in injured tissues are not well-understood. We will measure pH, PCO2, HCO3, and CO32 in ischemic brains and injured spinal cords, to determine the role of buffer capacity, lactoacidosis, and carbonic anhydrase in the pH changes. We propose that pH shifts modulate NMDA receptors and that interstitial buffering influences the rate and magnitude of spreading depression and ischemic injury in brain slices. Using on-selective microelectrodes, voltage clamp, and spectrophotometric methods, we will test the hypothesis that voltage-dependent Na:HCO3 co-transport governs H+, Na+, and HCO3- distribution across astrocytic membranes in normo- and hyperglycemic ischemia. Project 3. Ascorbate and glutathione in CNS Injury. We hypothesize that ascorbate (Asc) and glutathione (GSH) loss contributes to brain cell vulnerability to oxidative damage. Extracellular Asc will be measured in hypoxic brain slices and in ischemic cerebral cortex after middle cerebral artery occlusion. Intracellular Asc will be estimated from extracellular volumes and total tissue levels. Preliminary studies suggest that Asc and GSH are localized in neurons and astrocytes respectively, that both Asc and GSH decline with age and are lower in females. We will confirm these results, determine whether hypoxia and age compromises Asc homeostasis, assess protective effects of Asc and GSH in hypoxic brain slices, identify mechanisms of Asc and GSH loss in ischemia, and correlate tissue damage with Asc loss in male and females. Project 4. Water compartmentalization in Ischemia. The objective of this project is to understand how water shifts between extra- and intracellular compartments in ischemic brain tissues. Water shifts will be assessed with extracellular tracers, ion-elective microelectrodes, and weighing-drying method. We will manipulate extracellular ionic levels and block ionic transport and glutamate receptors. Water movements will be investigated in slices from aged animals. The role of the Na:HCO3 cotransport will be assessed. Light scattering and Na-K changes will be compared with wet-dry weight and tracer measurements.