Modification of Vascular Signaling in the Brain by Reactive Oxygen Species

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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.