Neuroprotection after Isoflurane Preconditioning

描述（由申请人提供）：中风是美国的主要杀手。脑卒中的病理生理机制是脑缺血再灌注损伤。虽然已经确定了许多神经保护干预措施，但临床上减少缺血性脑损伤的实用方法尚未得到很好的建立。我们的长期研究目标是确定缺血性脑损伤的有效干预措施，并了解这些干预措施的机制。我们的初步结果提出了一种新的神经保护方法，即事先短暂暴露于异氟醚（预处理）来诱导缺血耐受。初步研究表明，异氟醚预处理（IsoP）诱导的神经保护机制涉及急性期谷氨酸转运蛋白功能的改变和延迟期p38丝裂原活化蛋白激酶（MAPK）的激活。拟开展的研究将：1)研究哪些谷氨酸转运体参与其中，以及相关谷氨酸转运体的功能改变如何转化为神经系统预后的改善；2)研究p38如何被异氟醚活化；3)确定高迁移率组（HMG） I (Y)转录因子是否在p38下游对isop诱导的神经保护作用。采用脑切片和体内暂时性大脑中动脉闭塞模型。通过组织学分析评估神经元存活。细胞外谷氨酸水平在缺血半暗带将定量微透析。反义寡核苷酸将用于下调谷氨酸转运蛋白、p38 MAPK激酶和HMG I(Y)等选择性蛋白。Western分析和免疫组织化学将用于定量和定位蛋白表达。体外放射自显影定量测定HMG I(Y)的脑区表达。这些研究将为IsoP的神经保护机制提供深入的见解，并可能阐明新药物开发的机制。由于异氟醚已在临床实践中安全使用多年，IsoP可能被证明是一种有用的临床方法，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.