Astrocytes and Ischemic Brain Injury

描述（由申请人提供）：心脏骤停幸存者最害怕的残疾之一是神经损伤。全局脑缺血，如心脏骤停所示，导致海马CA1锥体神经元的延迟丢失，而附近的齿状回（DG）则相对抵抗。尽管对海马神经元损伤的研究很多，但对缺血对海马星形胶质细胞的影响知之甚少。这项资助旨在了解星形胶质细胞对海马选择性脆弱性的贡献。星形胶质细胞对许多脑功能至关重要，包括突触传递、代谢和离子稳态、抗氧化防御、炎症、血脑屏障和营养支持。本研究将从三个方面研究海马星形胶质细胞的存活和对缺血和缺血样应激的反应。在目的1中，线粒体功能将通过缺血和再灌注进行评估。线粒体对能量产生和细胞死亡的调节都很重要。再现选择性CA1易感性的海马器官型切片培养和从海马这两个亚区分离的星形胶质细胞将用于电生理研究和实时成像。评估CA1和DG星形胶质细胞再灌注过程中线粒体膜电位、活性氧和细胞内钙的变化时间。比较脆弱地区和抵抗地区将允许对重要变化进行区分。呼吸功能和线粒体复合体活性也将被测量。目的2将关注缺血和再灌注时星形胶质细胞谷氨酸摄取的变化。星形胶质细胞对谷氨酸摄取的早期损害危及邻近神经元，我们发现CA1星形胶质细胞中GLT-1的早期丢失。将进行谷氨酸摄取和转运体表达的详细研究。将监测维持膜电位和谷氨酸摄取所需的ATP储存。Aim 3将利用星形胶质细胞中伴侣蛋白Hsp70的选择性过表达和GLT-1的诱导作为保护策略，评估星形胶质细胞功能的变化以及神经元存活的变化。这些研究将提供海马星形胶质细胞缺血反应的图像，并将星形胶质细胞功能障碍与海马选择性神经元损伤联系起来。维持和改善星形胶质细胞功能为心脏骤停后改善神经功能的治疗提供了新的靶点。

DESCRIPTION (provided by applicant): One of the most feared disabilities in survivors of cardiac arrest is neurological impairment. Global cerebral ischemia, as seen with cardiac arrest, causes delayed loss of CA1 pyramidal neurons in the hippocampus while the nearby dentate gyrus (DG) is relatively resistant. Despite much work on hippocampal neuronal injury, little is known of the effects of ischemia on hippocampal astrocytes. This grant seeks to understand the contribution of astrocytes to hippocampal selective vulnerability. Astrocytes are essential to many brain functions including synaptic transmission, metabolic and ionic homeostasis, antioxidant defense, inflammation, the blood brain barrier, and trophic support. This proposal will study three aspects of hippocampal astrocyte survival and response to ischemia and ischemia-like stress. In aim 1, mitochondrial function will be assessed with ischemia and reperfusion. Mitochondria are important both for energy production and for the regulation of cell death. Hippocampal organotypic slice cultures, which reproduce selective CA1 vulnerability, and astrocytes isolated from these two subregions of the hippocampus will be used for electrophysiological studies and real time imaging. The timecourse of changes in mitochondrial membrane potential, reactive oxygen species, and intracellular calcium during reperfusion in astrocytes in CA1 and DG will be assessed. Comparing the vulnerable and resistant areas will allow discrimination of important changes. Respiratory function and mitochondrial complex activity will also be measured. Aim 2 will focus on changes in astrocyte glutamate uptake with ischemia and reperfusion. Early impairment of glutamate uptake by astrocytes endangers neighboring neurons, and we have found early loss of GLT-1 in CA1 astrocytes. Detailed studies of glutamate uptake and transporter expression will be performed. ATP stores required for maintaining membrane potential and glutamate uptake will be monitored. Aim 3 will use overexpression of the chaperone Hsp70 selectively in astrocytes and induction of GLT-1 as protective strategies and assess changes in astrocyte function as well as changes in neuronal survival. Together these studies will provide a picture of hippocampal astrocyte ischemic response and link astrocyte dysfunction to selective neuronal injury in hippocampus. Maintaining and improving astrocyte function provides a novel target for developing therapies to improve neurological function following cardiac arrest.