M-Type K+ Channels: In Vivo Neuroprotective Role during Cerebrovascular Stro

M 型 K 通道：脑血管痉挛期间的体内神经保护作用

• 批准号: 8484755
• 负责人: Sonya Marie Bierbower
• 金额: $ 5.66万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8484755
• 关键词: Action Potentials; Animals; Antiepileptic Agents; Area; Behavioral; Behavioral Assay; Biochemical; Biological Assay; Blood Vessels; Brain; Brain Edema; Brain Infarction; Brain region; Carotid Arteries; Cations; Cell Death; Cells; Cerebrum; Cessation of life; Clinical; Complex; Cytoprotection; Development; Disease; Dominant-Negative Mutation; Dyes; Encephalitis; Endothelium; Equilibrium; Event; Extravasation; Filament; Free Radical Formation; Growth; Hour; Human; Hypoxia; Image; Infarction; Inflammation; Inflammatory Response; Injury; Ipsilateral; Ischemic Stroke; Knock-in Mouse; Lasers; Learning; Lesion; Light; Locomotion; Mediating; Membrane Potentials; Middle Cerebral Artery Occlusion; Modeling; Motor; Motor Skills; Mus; Necrosis; Nervous System Trauma; Neurologic; Neurons; Neurophysiology - biologic function; Neurosciences; Outcome; Parietal Lobe; Patients; Pharmaceutical Preparations; Play; Potassium Channel; Probability; Production; Reactive Oxygen Species; Research; Research Personnel; Role; Rose Bengal; Rotarod Performance Test; Series; Severities; Stroke; Techniques; Testing; Therapeutic Intervention; Thrombosis; Time; Tissues; Training; Wild Type Mouse; Work; acute stroke; career; career development; cell injury; cerebrovascular; channel blockers; design; disability; drug development; flupirtine; improved; in vivo; in vivo Model; innovation; insight; middle cerebral artery; morris water maze; mouse model; neuronal excitability; neuroprotection; novel; novel therapeutic intervention; novel therapeutics; post stroke; research study; response; skills; voltage

DESCRIPTION (provided by applicant): Stroke is the third leading cause of human death in the USA and remains the focus of efforts to define the pathophysiologic mechanisms leading to tissue damage. Neuronal damage and cell loss occurs in numerous brain regions after strokes, thus providing a highly-relevant context for study. This project investigates neuroprotection during ischemic stroke through "M-channels" which are critical K+ channels that stabilize resting potentials by counterbalancing the depolarizing effects of excitatory cation currents. Produced by combinations of KCNQ2-5 subunits, voltage-gated M channels play critical roles in control of neuronal excitability, and action potential firing. We have previously shown that most M-type channels to be up- regulated by reactive oxygen species (ROS), molecules commonly produced during and after ischemic cerebrovascular stroke. This study hypothesizes that M current-mediated neuronal silencing has a neuroprotective role by increasing the open-probability of KCNQ channels, thus decreasing neuronal activity and prolonging activation of cellular cascades resulting in cell death. Two in vivo mouse models are used and the first involves a cerebral infarct within the parietal cortex produced by laser-controlled photothrombosis. When the photo-sensitizing dye, Rose Bengal, is irradiated by laser light, free-radical formation occurs causing endothelium damage, leakage of vascular contents into the parenchyma and vascular thrombosis. The second in vivo model is the middle cerebral artery occlusion (MCAo), in which a filament is introduced non-invasively into the MCA via the carotid artery. The MCAo model produces a catastrophic stroke in the ipsilateral hemisphere, with correspondingly pronounced cerebral deficits. The M-current activity will be pharmacologically altered in both mouse models using the M-channel openers, retigabine (RTG) and flupirtine, and the blocker, XE991, with drug application at various time points post-stroke. Additionally, dominant- negative KCNQ3 knock-in mice will be used in both models to confirm a specific M-channel mechanism. Finally, behavioral assays designed to distinguish degrees of injury and neural function will be used to delineate minute changes in function due to channel activation or blockage. Motor and co-ordination deficits will be assayed with the balance beam, ladder dexterity, open-field locomotion and accelerating rotarod tests. Learning deficits will be assayed using the Morris water maze. Thus, this study uses two strong and exciting stroke models, which may provide novel modes of therapeutic intervention for reducing neuronal damage caused during commonly-occurring ischemic attacks, by modulating the activity of M-type K+ channels.

描述（由申请人提供）：卒中是美国人类死亡的第三大原因，并且仍然是定义导致组织损伤的病理生理机制的工作重点。中风后许多脑区发生神经元损伤和细胞丢失，从而为研究提供了高度相关的背景。该项目通过“M通道”研究缺血性卒中期间的神经保护，M通道是通过平衡兴奋性阳离子电流的去极化作用来稳定静息电位的关键K+通道。电压门控M通道由KCNQ 2 -5亚基组合产生，在控制神经元兴奋性和动作电位放电中起关键作用。我们先前已经表明，大多数M型通道被活性氧（ROS）上调，活性氧是缺血性脑血管卒中期间和之后通常产生的分子。本研究假设M电流介导的神经元沉默具有神经保护作用，通过增加KCNQ通道的开放概率，从而降低神经元活性并延长导致细胞死亡的细胞级联的激活。使用两种体内小鼠模型，第一种涉及由激光控制的光致血栓形成产生的顶叶皮质内的脑梗死。当光敏染料虎红受到激光照射时，会形成自由基，导致内皮损伤、血管内容物渗漏到实质中和血管血栓形成。第二种体内模型是大脑中动脉闭塞（MCAo），其中通过颈动脉将细丝非侵入性地引入MCA。MCAo模型在同侧半球产生灾难性的中风，相应地具有明显的脑功能缺损。在使用M通道开放剂瑞替加滨（RTG）和氟吡汀以及阻断剂XE 991的两种小鼠模型中，M电流活性将在中风后不同时间点应用药物后发生显著改变。此外，显性阴性KCNQ 3基因敲入小鼠将用于两种模型中，以确认特定的M通道机制。最后，旨在区分损伤程度和神经功能的行为测定将用于描绘由于通道激活或阻塞而导致的功能的微小变化。运动和协调缺陷将通过平衡木、梯子灵活性、旷场运动和加速旋转杆测试进行分析。将使用Morris水迷宫测定学习缺陷。因此，本研究使用了两种强大的和令人兴奋的中风模型，这可能会提供新的治疗干预模式，以减少在常见的缺血性发作期间引起的神经元损伤，通过调节M型K+通道的活性。