The overall goals of this project are aimed at characterizing cerebral vascular muscle so as to improve our understanding and treatment of cerebral vasospasm and other causes of stroke, and to better understand mechanisms involved in treatment of high intracranial pressure by hyperventilation. Single isolated smooth muscle cell will be studied, in both contraction experiments and in patch clamp experiments. Comparative study of membrane Ca2+ and K+ channels will be performed, using smooth muscle cells from the basilar artery, cerebral penetrating arterioles and from the mesenteric artery of the guinea pig. Of particular interest will be comparison of Ca2+ currents in the three preparations, because in basilar artery cells, I have identified a novel current (B-type Ca2+ current) not previously reported in other vascular smooth muscle cells. In addition, I will study the effects of intracellular vs. extracellular changes in ph on the membrane currents and investigate the role second messengers in modulating Ca2+ and K+ currents in these cells. Mechanisms of receptor-activation in basilar artery smooth muscle cells will also be studied, by examining the contractile responses of single cells and effects on ionic currents in response to ATP, serotonin and norepinephrine and PGF2alpha. For each of these agents, I will determine the dependence of the contractile responses on extracellular Ca2+,from those that cause transmembrane flux of Ca2+, and using patch clamp techniques, I will further study those agents that require transmembrane flux of Ca2+, in order to distinguish between three possible mechanisms of activation: a) activation of voltage dependent dihydropyridine-sensitive Ca2+ channels by second messenger; b) inhibition of outward current channels by second messenger, which would result in voltage coupled activation of inward current; c) direct opening of discrete inward current channels by agonist. These experiments on receptor mechanisms will complement my recent finding of dual, opposing effects of serotonin on Ca 2+ currents in basilar artery cells. I expect that this study will lead to a better understanding of the mechanisms regulating cerebrovascular smooth muscle and modulating cerebral blood flow. I anticipate that with such knowledge, diseases leading to stroke can be better understood, and rational therapies can be devised to optimize their treatment.
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