CHANNEL REGULATION IN NORMAL/HYPERTENSIVE SMOOTH MUSCLE

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The overall objective of this investigation is to characterize ion conductance pathways and elucidate their regulation in smooth muscle cells isolated from normal and hypertensive animal models. Most of the proposed studies will be carried out on single arterial smooth muscle cells isolated from rat and rabbit mesenteric artery or dissected rings of aorta, which can be obtained in small number but which maintain bipolar relaxed morphology and agonist-induced contraction. Patch clamp studies (whole cell and single channel recording) are designed to characterize individual Ca2+, Na+ and K+ channels in smooth muscle cells from normal and hypertensive animals. The regulation of ionic currents by vasoactive receptor agonists and the modulation of channels by receptors, second messengers, G proteins and protein kinases and phosphatases will be elucidated. Flourometric detection and imaging of fluorescent indicators in the same cells will elucidate time courses and homogeneity of cell response with respect to cytosolic Ca2+ and H+. These approaches will be extended to characterize the ion conductance pathways occurring during maintenance of isolated smooth muscle cells under proliferative and non-proliferative tissue culture conditions. The major strength of the proposal is the application of a systematic electrophysiological investigation coupled with biochemical, biophysical and cell biology techniques and a pathological perspective to the study of freshly isolated and tissue cultured arterial smooth muscle cells from normal and hypertensive animal models such as the SHR Kyoto spontaneous hypertensive model and the angiotensin induced secondary hypertensive models. The long term goal of the application is to understand physiological mechanisms and pathological deviations of a) the role of sarcolemma ion channels in regulating vascular smooth muscle tone in situ by multiple input from neuronal, humoral and locally produced signalling elements and b) to assess if phenotypic modulation of ion conductance pathways occurs under hypertension, a condition known to involve hyperplasia of smooth muscle cells similar to those studied.