Mitochondrial dynamics in astrocytic processes after transient ischemia

短暂性缺血后星形胶质细胞过程中的线粒体动力学

基本信息

批准号：
8921078
负责人：
John Charles O'Donnell
金额：
$ 3.73万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-01 至 2016-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8921078
关键词：
3-methyladenine Acute Antioxidants Area Astrocytes Attenuated Autophagocytosis Autophagosome Binding Biolistics Brain Buffers Calcium Cell Death Cells Cessation of life Confocal Microscopy Cyclosporine DNA Data Development Dominant-Negative Mutation Dyes Employee Strikes Enzymes Extracellular Space Failure Fluorescent Dyes Genetic Glucose Glutamate Transporter Glutamates Grant Hippocampus (Brain)Image Individual Injury Intervention Ions Ischemia Knowledge Label Lead Light MK801 Measures Medicine Membrane Potentials Mentors Metabolic Mitochondria Modeling Monitor N-Methyl-D-Aspartate Receptors N-Methylaspartate National Institute of Neurological Disorders and Stroke Neurodegenerative Disorders Neurons Neurotransmitters Oxygen Pathology Pennsylvania Process Production Prosencephalon Ranvier&apos s Nodes Reactive Oxygen Species Resources Role Scientist Signal Transduction Site Slice Stroke Synapses TO-PRO-3 Time Training Transfection Ubiquitination Universities cell type central nervous system injury density deprivation excitotoxicity experience extracellular in vivo induced hypothermia inhibitor/antagonist mitochondrial dysfunction mitochondrial membrane mutant natural hypothermia neuron loss new therapeutic target novel parkin gene/protein prevent public health relevance research study response therapeutic target ubiquitin-protein ligase uptake

项目摘要

DESCRIPTION (provided by applicant): Astrocytes are the most abundant cell type in brain. They are responsible for clearing extracellular glutamate, the predominant excitatory neurotransmitter, from the synapse to maintain crisp signaling and prevent excitotoxicity. In forebrain, the astrocytic glutamate transporter, GLT1, is responsible for the vast majority of glutamate uptake. Mitochondria are invested throughout fine astrocytic processes where they colocalize with GLT1. We recently discovered physical and functional interactions between GLT1, multiple glycolytic enzymes and mitochondria. In completing Aim 1 of my original grant submission, we concluded that mitochondria in astrocytic processes are retained near glutamate transporters and synapses. Our data suggest that this distribution is regulated by neuronal glutamate release, astrocytic glutamate uptake, and reversal of the Na+/Ca2+ exchanger. Mitochondria can support glutamate uptake by providing ATP and buffering ions, and there is growing evidence suggesting that a portion of transported glutamate is oxidized in mitochondria to generate energy in these compartments. Mitochondrial dysfunction and excitotoxicity from failure of astrocytic glutamate uptake are at the core of the delayed cell deat that persists after an ischemic insult. Aside from inducing hypothermia, this pathology is currently untreatable. I have observed a loss of mitochondrial density in astrocytic processes in response to oxygen glucose deprivation (OGD) that precedes the delayed neuronal cell death that is common to this ex vivo model and to stroke in vivo. In the first aim I will characterize th OGD-induced loss of mitochondria from astrocytic processes, and determine if it is preceded by changes in mitochondrial membrane potential and reactive oxygen species. I will pharmacologically block or activate NMDA receptors to investigate the relationship between excitotoxicity and reduced mitochondrial occupancy of astrocytic processes. In a preliminary study, I found that cyclosporin A reduced cell death and attenuated the loss of mitochondrial density in astrocytic processes after OGD. Cyclosporin A increases mitochondrial capacity for calcium buffering. Treatment with cyclosporin A will be evaluated as mechanism of intervention for attenuating the loss of mitochondria from processes. I have also observed increased mitophagy (a mechanism for degradation of dysfunctional mitochondria) in astrocytic processes after OGD. In aim two I will characterize changes in mitophagy after OGD. I will also evaluate the effects of pharmacological and genetic inhibition of mitophagy on mitochondrial occupancy of astrocytic processes and delayed neuronal death after OGD. By providing the first ever examination of the role of mitochondrial dynamics in astrocytic processes during ischemic injury, execution of this project could help lead to new therapeutic targets for a field of medicine that desperately needs them.

描述（由申请人提供）：星形胶质细胞是大脑中最丰富的细胞类型。它们负责从突触中清除细胞外谷氨酸（主要的兴奋性神经递质），以维持清晰的信号传导并防止兴奋性毒性。在前脑中，星形胶质细胞谷氨酸转运蛋白 GLT1 负责绝大多数谷氨酸的吸收。线粒体分布在星形胶质细胞的整个过程中，与 GLT1 共定位。我们最近发现了 GLT1、多种糖酵解酶和线粒体之间的物理和功能相互作用。在完成我最初提交的资助的目标 1 时，我们得出的结论是，星形胶质细胞过程中的线粒体保留在谷氨酸转运蛋白和突触附近。我们的数据表明，这种分布受到神经元谷氨酸释放、星形细胞谷氨酸摄取和 Na+/Ca2+ 交换器逆转的调节。线粒体可以通过提供 ATP 和缓冲离子来支持谷氨酸的吸收，并且越来越多的证据表明，部分运输的谷氨酸在线粒体中被氧化，从而在这些区室中产生能量。线粒体功能障碍和星形细胞谷氨酸摄取失败引起的兴奋性毒性是缺血性损伤后持续存在的延迟性细胞死亡的核心。除了引起体温过低之外，这种病理学目前无法治疗。我观察到星形胶质细胞过程中线粒体密度的损失是对氧糖剥夺（OGD）的反应，这种损失发生在延迟性神经元细胞死亡之前，这对于这种离体模型和体内中风来说是常见的。在第一个目标中，我将描述 OGD 诱导的星形胶质细胞过程中线粒体的损失，并确定其之前是否发生线粒体膜电位和活性氧的变化。我将通过药理学阻断或激活 NMDA 受体，以研究兴奋性毒性与星形胶质细胞过程中线粒体占用减少之间的关系。在一项初步研究中，我发现环孢素 A 减少了 OGD 后星形胶质细胞过程中的细胞死亡并减轻了线粒体密度的损失。环孢素 A 增加线粒体的钙缓冲能力。将评估环孢菌素 A 治疗作为减少过程中线粒体损失的干预机制。我还观察到 OGD 后星形胶质细胞过程中线粒体自噬（功能失调的线粒体降解的一种机制）增加。在目标二中，我将描述 OGD 后线粒体自噬的变化。我还将评估线粒体自噬的药理和遗传抑制对星形胶质细胞过程的线粒体占用和 OGD 后延迟神经元死亡的影响。通过首次检查线粒体动力学在缺血性损伤期间星形胶质细胞过程中的作用，该项目的执行可能有助于为迫切需要它们的医学领域带来新的治疗靶点。