Changes in blood flow induce vascular remodeling and changes in vascular reactivity, that are mediated by the endothelium. An increase in blood flow induces the endothelium to release paracrine substances that alter vascular smooth muscle tone and growth (i.e., endothelium-derived relaxing factor (EDRF), transforming growth factor-beta (TGFbeta), and platelet- derived growth factor). We and others have accumulated evidence that potassium and calcium channels in the endothelial cell appear to be involved in the mechanotransduction of hemodynamic (i.e., flow) and humoral signals. We propose to clone and to characterize, using molecular and cellular techniques, the endothelial and calcium channels that play a role in the flow response, as well as the response to other stimuli. For this purpose, molecular genetic techniques will be used to clone and to sequence endothelial potassium and calcium channels. The channels will be expressed in Xenopus oocytes and in mammalian cells, so as to characterize their electrophysiologic and pharmacologic properties. To determine the importance of specific channels in the mechanotransduction of humoral and hemodynamic stimuli, the effect of molecular manipulations of these channels on endothelial physiology will be examined. We will attenuate or augment the expression of these channels using antisense and overexpression strategies. The effect of these manipulations on the response of endothelial cells to flow or pharmacologic stimuli will be investigated using molecular, biochemical, and bioassay techniques to detect the synthesis and release of EDRF and TGFbeta. This project will also determine whether the ability to transduce the stimulus of flow is an intrinsic property of the channels, or is conferred by a separate mechanoreceptor. Experiments to clone such a receptor are proposed. In sum, this project will provide a new understanding of the role of calcium and potassium channels in endothelial physiology, and may identify potential targets for therapeutic intervention.
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