Brain Injury and Intercelluar Calcium Waves

描述(由申请人提供)：当前提案的目标是 评估创伤性脑损伤改变细胞间质的假说 星形胶质细胞之间的沟通，一个潜在的关键和经常被忽视的 大脑处理的一个方面。星形胶质细胞曾被视为被动细胞，但 现在被描述为大脑中信号处理的积极贡献者。 然而，创伤对钙介导的星形胶质细胞间质的影响 尚未对信号进行评估。因此，该提案评估了 创伤性损伤改变细胞内钙信号的假说 胶质合胞体中的星形胶质细胞使用以下特定目标： 特定目标1：皮质星形胶质细胞损伤改变起源和 细胞间钙波的传播。我们会比较速度和速度 星形胶质细胞损伤后细胞内钙波距星形胶质细胞的距离 未受伤的星形胶质细胞。(方法：使用定量荧光显微镜 评估细胞间钙波)。 具体目标2：损伤改变缝隙连接偶联。我们会比较一下 荧光染料通过缝隙连接转移，连接蛋白43的表达 蛋白和连接蛋白43蛋白的磷酸化 损伤的星形胶质细胞。方法：用荧光黄转移法测量细胞的偶联。 用免疫印迹法检测缝隙连接蛋白的表达和磷酸化。 连接蛋白43蛋白)。 具体目标3：细胞内钙信号将维持在 IP3介导的星形胶质细胞内药物调控 钙信号。(方法：使用元素激动剂和拮抗剂。 确定细胞间钙波变化的信号通路 可以衰减)。 这些拟议的实验第一次评估了 复杂而广泛的钙信号网络上的创伤性脑损伤 在星形细胞中。目的#1直接检查机械损伤对 星形胶质细胞内的细胞内钙信号。Aim#2检查潜在的 参与损伤诱导的改变的机制(即缝隙连接) 细胞内钙信号。目标#3检查潜在的治疗方法 可以恢复细胞间信号的干预。评选结果 拟议中的实验不仅可能为病理生理学提供新的见解 但也有可能导致新的治疗方法的发展 脑外伤的治疗方法。

DESCRIPTION (provided by applicant): The goal of the current proposal is to evaluate the hypothesis that traumatic brain injury alters intercellular communication between astrocytes, a potentially crucial and often overlooked aspect of brain processing. Astrocytes were once viewed as passive cells, but are now characterized as active contributors to signal processing in the brain. Yet, the effect of traumatic injury on calcium-mediated astrocyte intercellular signaling has not been evaluated. Therefore, the proposal evaluates the hypothesis that traumatic injury alters intercellular calcium signaling between astrocytes in the glial syncytium using the following specific aims: Specific Aim 1: Injury of cortical astrocytes alters origination and propagation of intercellular calcium waves. We will compare the velocity and distance of intercellular calcium waves in injured astrocytes to that of uninjured astrocytes. (Method: Use quantitative fluorescent microscopy to evaluate intercellular calcium waves). Specific Aim 2: Injury alters gap junction coupling. We will compare fluorescent dye transfer through gap junctions, expression of Connexin 43 protein, and phosphorylation of Connexin 43 protein between uninjured and injured astrocytes. (Method: Use lucifer yellow transfer to measure coupling of gap junctions; use Western Blots to measure expression and phosphorylation of Connexin 43 protein). Specific Aim 3: Intercellular calcium signaling will be maintained in astrocytes by pharmacological manipulation of IP3-mediated intracellular calcium signaling. (Method: Use agonists and antagonists of elements in the signaling pathway to determine if alterations in intercellular calcium waves can be attenuated). These proposed experiments evaluate, for the first time, the effect of traumatic brain injury on the intricate and extensive calcium signaling network among astrocytes. Aim #1 directly examines the effects of mechanical injury on intercellular calcium signaling in astrocytes. Aim #2 examines the potential mechanisms (i.e. gap junctions) involved in injury-induced alterations in intercellular calcium signaling. Aim #3 examines potential therapeutic interventions which could restore intercellular signaling. The results of the proposed experiments may provide not only new insights into the pathophysiology of TBI, but also potentially lead to the development of novel therapeutic approaches for the treatment of the traumatically injured brain.