The overall goal of this project is to investigate the underlying mechanisms by which different durations of ischemia and reperfusion (l/R) alter endothelium-dependent vasodilation and vascular tone in middle cerebral arteries (MCA) and parenchymal arterioles (PA) in ways that may affect postischemic reperfusion and stroke outcome. Our preliminary and published studies demonstrate a specific effect of reperfusion on the function of MCA vs. PA. While longer reperfusion durations impair basal tone and myogenic reactivity in MCA, basal tone is considerably greater and well-preserved in PA regardless of l/R. We have hypothesized that the greater basal tone of PA during postichemic reperfusion could contribute to infarct expansion and worsen stroke outcome by limiting reperfusion CBF. In addition, there is an enhanced influence of EDHF on resting tone in PA at short reperfusion durations (30 minutes), but a significantly diminished influence after longer reperfusion (2 hours). The contribution of these vasodilator mechanisms to stroke outcome after different durations of reperfusion will also be investigated. The studies outlined in this project will investigate the effect of l/R on small- and intermediate-conductance calcium-activated potassium channels (SK, IK) in endothelium, and transient receptor potential (TRP) channels and voltage-dependent calcium channels (Cav) in vascular smooth muscle. These channels are important for the control of membrane potential and cell calcium that define vascular function and are a major focus of this Program Project Grant. In addition, we will investigate how peroxynitrite, a reactive nitrogen species produced during postischemic reperfusion, affects smooth muscle membrane potential and cell calcium to cause vasoconstriction, in PA and if treatment to decrease peroxynitrite levels improves stroke outcome and vascular function. The proposed experiments will be in close collaboration with the investigators and Cores of this Program Project Grant. Project 1 investigates calcium signaling in endothelium of PA also with a focus on SK/IK channels under normal conditions and their role in controlling cerebral blood flow. Project 2 is focused on myogenic activity in PA and the role of Cav and TRP channels in mediating this response, a similar focus as in this Project under conditions of l/R. Project 4 will investigate similar changes under conditions of hemorrhagic stroke that could provide valuable insight into both these conditions. We will use a powerful combination of state-of-the-art techniques to study changes in cell calcium, ion channel function, and membrane potential in MCA and PA after focal cerebral ischemia. The considerable expertise of our group of investigators in ion channel function and cell calcium will be utilized fully to provide technical and conceptual support for our Project.
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