Modification of Vascular Signaling in the Brain by Reactive Oxygen Species

越来越多的数据表明，活性氧和氮中间体参与启动/修改控制血管功能的信号事件。关于活性氧（ROS），最近的报道明确了O2-和H2O2在血管内皮相互作用中的作用。低水平的氧化应激已被证明可以介导增殖事件，而高水平的氧化应激已被证明可以诱导细胞凋亡。该项目的目的是确定ROS在启动或修改调节脑血流量的信号事件中的作用。我们还将探讨ROS修改有丝分裂信号事件的假设，这些信号事件调节毛细血管密度以响应神经元代谢需求。初步数据显示，H202显著提高脑小动脉肌细胞K+通道活性。我们还提供数据表明，细胞色素P450 （P450）酶活性受到氧化应激的抑制。实验计划确定ROS在压力介导的肌张力，脑血流的自动调节和控制肌肉细胞兴奋性的离子通道电导系统中的作用。我们实验室之前的研究表明，星形胶质细胞释放的P450环氧化物响应谷氨酸，通过增强K+通道活性和血管肌肉的超极化，有效地扩张脑小动脉。我们将确定ROS是否通过对肌肉细胞K+通道的作用或通过干扰P450衍生环氧化物的产生来改变功能性充血。初步数据表明，自由基可以改变功能性充血，并且通过向麻醉大鼠脑脊液中注入SOD来清除O2-可以改变基线脑血流量。最后，我们将定义活性氧修饰毛细血管生成的能力。O2-的产生对培养的脑毛细血管内皮细胞胸腺嘧啶掺入有双相作用。短时间（60分钟）暴露于毛细血管内皮O2-会增加胸腺嘧啶的掺入，而长时间（8小时）暴露会减少胸腺嘧啶的掺入。我们将定义氧化应激改变毛细血管增殖和管形成的机制。该项目将使用各种分子、生化和功能技术来定义活性氧在分子、细胞和整个动物水平上改变脑血流量、自我调节和神经活动反应中的作用。与拟建PPG的其他项目有密切的互动，并广泛使用核心资源。

There is a growing body of data demonstrating that reactive oxygen and nitrogen intermediates are involved in initiating/modifying signaling events controlling vascular function. With respect to reactive oxygen species (ROS) recent reports define a role for O2- and H2O2 in vascular endothelial interactions. Low levels of oxidant stress have been shown to mediate proliferative events, while high levels have been shown to induce apoptosis. The purpose of this Project is to define the actions of ROS in initiating or modifying signaling events which regulate cerebral blood flow. We will, also, explore the hypothesis that ROS modify mitogenic signaling events which regulate capillary density in response to neuronal metabolic demand. Preliminary data show that H202 markedly increase K+ channel activity in cerebral arteriolar muscle cells. We also present data demonstrating that cytochrome P450 (P450) enzyme activity is inhibited by oxidant stress. Experiments are planned to define the action of ROS on pressure-mediated myogenic tone, autoregulation of cerebral blood flow and ion channel conductance systems controlling muscle cell excitability. Previous work from our laboratory has shown that P450 epoxides released from astrocytes in response to glutamate potently dilate cerebral arterioles by enhancing K+ channel activity and hyperpolarizing vascular muscle. We will determine if ROS modify functional hyperemia via actions on muscle cell K+ channels or by interfering with P450 derived epoxide production. Preliminary data suggests that free radicals can modify functional hyperemia, and that scavenging of O2- by infusing SOD into the cerebral spinal fluid of anesthetized rats alters baseline cerebral blood flow. Finally, we will define the ability of ROS to modify capillary angiogenesis. Generation of O2- exerts biphasic actions on thymadine incorporation in cultured cerebral capillary endothelial cells. Short (60 minute) exposure of capillary endothelium to O2- increases thymadine incorporation, while longer (8 hours) exposure reduces thymidine incorporation. We will define the mechanisms through which oxidant stress alters capillary proliferation and tube formation. This Project will use a variety of molecular, biochemical and functional techniques to define the action of ROS in modifying cerebral blood flow, autoregulation and responses to neural activity at the molecular, cellular and whole animal level. There is strong interaction with the other Projects of this proposed PPG, and extensive use of Core resources.