The objective of this revised proposal is to elucidate the expression and role of bradykiritin (BK) B2 and B receptors in blood pressure and cardiovascular regulation. Kinins are potent vasodilating peptides that are released from precursor kininogens by kallikreins. Kinins bind to BK receptors and mediate a broad spectrum of biological effects including vasodllation, ion transport, smooth muscle contraction and relaxation, pain and inflammation. There are two types of kinin receptors, B2 and B1 which differ in biological properties and primary structures. This project is based on the hypothesis that the BK receptor plays a role in blood pressure regulation which is supported by our encouraging preliminary results that antisense inhibition of B2 receptors in rats causes an increase of blood pressure while transgenic mice over-expressing human B2 receptors are hypotensive. Since the last submission of this application, we continue to make progress in molecular cloning, expression and locallzation of BK receptors, developed essential biochemical and molecular probes and established technologies which form the basis for the current application.
The specific aims of this proposal are: (l) to clone and characterize the BK receptor genes from human and rat, and to create DNA constructs with and without adenoviral vectors for somatic gene delivery and transgenic mouse development, (2) to analyze elements which regulate the BK receptor genes' transcription in a-cell culture system by deletion analysis, site-directed mutagensis, gel retardation and footprinting assays, and in vivo by deletion analysis via direct gene delivery, (3) to analyze the role of BK receptors in central and peripheral regulation of blood pressure by antisense inhibition and by combination gene delivery in normotensive and hypertensive rats, (4) to develop and characterize transgenic mice over-expressing BK receptor genes and to analyze phenotypic alterations such as blood pressure changes, renal function and smooth muscle contraction. The proposed studies could provide fundamental information about the structure, regulation and function of BK receptors in order to facilitate the design of therapeutic agents for treating hypertension, cardiovascular disease and inflammation.
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