The long term objective of this project is to define the mechanisms responsible for the changes that occur in the reactivity of arteries with aging.
The specific aims of this application are designed to test several hypotheses concerning the possible mechanisms including: 1) the function of arterial wall Na+-K+-ATPase (Na-pump); 2) the intrinsic reactivity of the contractile system; 3) Beta-adrenergic relaxation of contracted arteries; and 4) membrane cholesterol and phospholipids. Studies will be conducted using several isolated arteries from the NIA's colony of Fischer 344 rats at ages of 1 to 33 months. Studies of Na-pump function will be performed under conditions of maximal activity in Na+ loaded arteries and of resting activity in arteries using the Rb86-uptake procedure. Studies of Na-pump activity will include: a) dependence on intracellular (Na+) and extracellular (Rb+); b) dose-dependence of ouabain inhibition of Na-pump activity; c) effect of cell Na+ increase on agonist dose-response relations to norepinephrine, secrotonin and K+. Activation with Ca+2 following detergent (saponin) increased membrane permeability will be performed to determine the intrinsic properties of the contractile apparatus. The effects of substrates Mg+2, MgATP-2 and calmodulin on these responses will be determined. Relaxation responses of precontracted arteries will be tested using forskolin (adenyl cyclase activation), isoproterenol (Beta-receptor), and acetylcholine (endothelium mediated). The contributiion of membrane cholesterol and phospholipid content to altered arterial wall function (vascular reactivity, Na-pump function and 42K efflux) will be stuided by: a) determining the aging changes of various lipid classes in arteries; b) altering membrane cholesterol content with cholesterol acceptor or donor particles; c) measuring NE activated phosphotidylinositol (PI) hydrolysis and d) altering membrane PI content with PI loading; and depleting procedures. Schild analysis will be applied to test for the occurrence of conformational changes in Alpha- and Beta-adrenergic receptors. These studies will help direct attention to several arterial smooth muscle membrane and cellular mechanisms that may be responsible for the altered vascular reactivity that occurs in aging and will further serve to focus future research efforts.
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