Neuroprotection after Isoflurane Preconditioning

描述(申请人提供)：中风是美国的主要杀手。脑缺血再灌注性脑损伤是卒中的重要病理生理基础。虽然已经确定了许多神经保护干预措施，但临床上实用的减少缺血性脑损伤的方法还没有很好的确立。我们的长期研究目标是确定有效的缺血性脑损伤干预措施，并了解这些干预措施的机制。我们的初步结果提示了一种新的神经保护方法，在这种方法中，预先短暂地暴露于异氟醚(预适应)可以诱导缺血耐受。初步研究表明，异氟醚预适应(IsoP)诱导的神经保护机制包括急性期谷氨酸转运体功能的改变和延迟期p38丝裂原活化蛋白激酶(MAPK)的激活。拟议的研究将：1)调查哪些谷氨酸转运体参与，以及相关谷氨酸转运体的功能变化如何转化为改善神经预后；2)研究异氟醚如何激活p38；3)确定高迁移率族(HMG)I(Y)转录因子是否位于p38下游，以实现IsoP诱导的神经保护。将使用脑片和体内暂时性大脑中动脉闭塞模型。神经元存活将通过组织学分析进行评估。缺血半暗带的细胞外谷氨酸水平将通过微透析进行量化。反义寡核苷酸将被用来下调选择性蛋白质，如谷氨酸转运体、p38 MAPK激酶和HMG I(Y)。Western分析和免疫组织化学将用于对蛋白质表达进行量化和定位。HMG I(Y)的脑局部表达将通过定量的体外放射自显影来确定。这些研究将深入了解IsoP的神经保护机制，并可能阐明新药可能利用的机制。由于异氟醚已在临床上安全使用多年，异氟醚可能被证明是一种有效的临床方法，以减少缺血性脑损伤。

DESCRIPTION (provided by applicant): Stroke is a major killer in the USA. The underlying pathophysiology for stroke is ischemia and reperfusion brain injury. Although many neuroprotective interventions have been identified, clinically practical methods to reduce ischemic brain injury are not well established yet. Our long-term research goals are to identify effective interventions for ischemic brain injury and to understand the mechanisms of these interventions. Our preliminary results suggest a novel neuroprotective method in which a prior brief exposure to isoflurane (preconditioning) is used to induce ischemic tolerance. Initial studies suggest that the mechanisms for this isoflurane preconditioning (IsoP)-induced neuroprotection involve alteration of glutamate transporter function in the acute phase and activation of p38 mitogen-activated protein kinase (MAPK) in the delayed phase. The proposed studies will: 1) investigate which glutamate transporters are involved and how the functional changes of the involved glutamate transporters translate into improved neurological outcome; 2) examine how p38 is activated by isoflurane; 3) determine whether the high mobility group (HMG) I (Y) transcription factor is downstream of p38 for IsoP-induced neuroprotection. Brain slices and in vivo temporary middle cerebral arterial occlusion models will be used. Neuronal survival will be evaluated by histological analysis. Extracellular glutamate levels in the ischemic penumbra will be quantified by microdialysis. Antisense oligonucleotides will be used to downregulate selective proteins such as glutamate transporters, p38 MAPK kinases and HMG I(Y). Western analysis and immunohistochemistry will be used to quantify and localize protein expression. Brain regional expression of HMG I(Y) will be determined by quantitative in vitro autoradiography. These studies will provide insight into the mechanisms of neuroprotection by IsoP and may elucidate mechanisms that could be exploited by new drugs. Since isoflurane has been used safely in clinical practice for many years, IsoP may prove to be a useful clinical method to reduce ischemic brain injury.