The walls of arteries are largely composed of smooth muscle cells. By contracting or relaxing, these cells determine arterial diameter, which in turn regulates blood flow and blood pressure. The concentration of calcium in arterial smooth muscle determines in part the degree of contraction. A major source of calcium entry into these cells is through voltage-dependent L-type calcium channels. The general goal of this proposal is to investigate the poorly understood mechanisms controlling calcium channel function in arterial smooth muscle. More specifically, this research investigates a novel regulatory mechanism where localized oxidant and calcium signaling microdomains functionally converge in arterial smooth muscle cells. This promotes increased L-type calcium channel activity, increased calcium within the smooth muscle cells, and ultimately arterial contraction. Importantly, increased oxidative stress and increased calcium channel activity are thought to be related to vascular dysfunction in obesity-related cardiovascular diseases such as hypertension and stroke. In this application we propose to test a model where the convergence of redox and calcium microdomain signaling requires plasmalemmal caveolae (containing NADPH oxidase and L-type calcium channels) that are closely opposed to peripheral mitochondria. We will also investigate if the resulting coupled redox/calcium signaling contributes to normal arterial functio and to arterial dysfunction in obesity.
Specific Aim 1 tests the hypothesis that NADPH oxidase and L-type calcium channels colocalize in caveolae to produce functionally coupled redox and calcium microdomains in arterial smooth muscle.
Specific Aim 2 tests the hypothesis that a subpopulation of peripheral mitochondria modulate functional coupling of redox and calcium microdomains.
Specific Aim 3 tests the hypothesis that increased functionally coupled redox and calcium microdomains contribute to arterial dysfunction in obesity. The experiments in these Specific Aims will use a combination of voltage-clamp electrophysiology, total internal reflection fluorescence (TIRF) microscopy, molecular biology, transmission electron microscopy, and intact pressurized arteries to examine the structural and functional role of caveolae and mitochondria in redox and calcium microdomain signaling in arterial smooth muscle from healthy and obese animals. The outcome of these experiments will provide mechanistic insights into events underlying arterial dysfunction in obesity and may lead to the development of new rational therapies for managing and preventing cardiovascular disease.
Experiments described in this proposal will examine how oxidant and calcium signals influence each other and cause contraction of blood vessels. This is important because contraction of blood vessels regulates blood flow and blood pressure and increased contraction contributes to cardiovascular diseases such as hypertension,stroke, and coronary artery disease. The ultimate goal of this project is to improve human health by stimulating the development of new pharmaceuticals that reduce over-contraction of blood vessels in order to prevent and manage cardiovascular disease.
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