EXCITOTOXICITY IN CIRCULATORY ARREST--BRAIN INJURY

本项目的目标是确定兴奋的机制。 低温循环中神经递质介导的神经元死亡 逮捕(HCA)，并制定预防它的药理策略。我们 建立了一种模拟临床的犬肝细胞癌存活模型 心血管外科实习。狗只需接受2小时的 在18摄氏度的脑温下循环停止 持续的神经功能缺陷和选择性脑脊髓炎的组织病理类型 神经元坏死和神经递质特异性受体的变化。在……里面 在当前的资助期内，我们发现NMDA拮抗剂， 地佐西平，在20岁之前和20岁以上给药，可以预防大部分伤害 HCA后几个小时。AMPA拮抗剂NBQX也有保护作用，即使 它是在从HCA复温后服用的 与神经元死亡相关的脑电改变 发展起来的。这些观察表明，谷氨酸介导的大部分 HCA的损伤发生在再灌注期，而这些事件 中介损害可以用药物来阻断。 利用活体脑微透析进行的初步实验表明 胞外兴奋性氨基酸的大量增加 神经递质谷氨酸和甘氨酸先于12倍的 精氨酸在一氧化氮合酶(NOS)作用下转化为瓜氨酸 术后再灌注期。使用免疫细胞化学，我们 观察到神经元型一氧化氮合酶(NNOS)活性的表达增加 基本上在脑区再灌流6-18小时后 富含谷氨酸受体，如海马体、基底节和 小脑。伴随着这种酶活性增加的是一种 一氧化氮(NO)代谢产物在血清和尿液中的积聚 被一氧化氮合酶抑制剂有效地减少。初步数据还表明， 肝细胞癌后神经细胞死亡是由细胞凋亡和坏死共同引起的。 在nNOS被激活期间。NO可能直接负责 因为我们可以可靠地抑制细胞的凋亡 使用一氧化氮合酶抑制剂。 探讨HCA术后介导脑损伤的一连串事件 建议：(1)确定神经递质的模式和序列 神经递质受体表达的调节和释放； (2)探讨一氧化氮在HCA后神经毒性中的作用； 确定兴奋性神经递质与一氧化氮的关系 活性；(4)检测细胞凋亡和坏死的发生。 HCA和NOS抑制剂抑制细胞死亡后；(5) 确定如何以及何时操纵NO对神经系统的影响 狗的结局。 这项拟议的研究将加快谷氨酸受体的临床应用 拮抗剂，并可能开发一种新的治疗方法 基于使用异构型特异性一氧化氮合酶抑制剂的神经保护。这个 提供脑保护的能力可能会使患者变得更好 心胸外科手术中循环停止后的结果。

The goal of this project is to define the mechanisms of excitatory neurotransmitter mediated neuronal death during hypothermic circulatory arrest (HCA) and to develop a pharmacologic strategy to prevent it. We established a canine survival model of HCA that closely simulates clinical practice in cardiovascular surgery. Dogs subjected to 2 hours of circulatory arrest at a brain temperature of 18 degrees C suffer a consistent neurologic deficit and histopathologic pattern of selective neuronal necrosis and changes in neurotransmitter specific receptors. In the current period of the grant, we found that the NMDA antagonist, dizocilpine, prevented most of the injury when given before and over 20 hours after HCA. The AMPA antagonist NBQX was also protective even though it was administrated after rewarming from HCA at a time when electroencephalographic (EEG) changes associated with neuronal death had developed. These observations indicate the majority of glutamate mediated damage from HCA evolves in the reperfusion period and that the events mediating damage can be blocked with medications. Preliminary experiments utilizing in vivo brain microdialysis indicate that large extracellular increases in the excitatory amino acid neurotransmitters glutamate and glycine precede a 12-fold rise in the conversion of arginine to citrulline by nitric oxide synthase (NOS) during the postoperative reperfusion period. Using immunocytochemistry, we observed that the expression of neuronal NOS (nNOS) activity increases substantially after 6-18 hours of reperfusion in areas of the brain enriched in glutamate receptors such as the hippocampus, basal ganglia and cerebellum. Concomitant with this increased enzymatic activity is an accumulation of nitric oxide (NO) metabolites in the serum and urine that are effectively reduced by NOS inhibitors. Preliminary data also suggest that neuronal death following HCA occurs due to both apoptosis and necrosis during the period when nNOS is activated. NO may be directly responsible for such programmed cell death since we can reliably inhibit the apoptosis with NOS inhibitors. To explore the cascade of events that mediate brain injury following HCA we propose to: (1) determine the pattern and sequence of neurotransmitter release as well as the regulation of neurotransmitter receptor expression; (2) examine the role of NO in mediating neurotoxicity after HCA; (3) determine the relationship between excitatory neurotransmitters and NO activity; (4) examine the occurrence of both apoptosis and necrosis following HCA and the inhabitation of cell death with NOS inhibitors; (5) determine how and when manipulating NO production affects the neurologic outcome of dogs. The proposed research will expedite the clinical use of glutamate receptor antagonists, and may develop a novel therapeutic approach to neuroprotection based on the use of isoform specific NOS inhibitors. The ability to provide cerebral protection could result in better patient outcomes after circulatory arrest during cardiothoracic procedures.