The overall goal of this program is to understand the role of the renin- angiotensin system (RAS) in controlling blood pressure, especially in hypertension. As part of this effort, we will describe the expression and regulation of the RAS and determine the brain sites at which it acts in transgenic hypertensive rats harboring the mouse Ren-2 gene. This model is exceptional because it allows, for the first time, study of a model of hypertension dependent on the overexpression of the product of a single gene. The overexpression of angiotensin (Ang) peptides will also allow improved resolution in neuroanatomical localization studies. The studies proposed here will examine the role of the increased brain expression of Ang peptides in the maintenance of hypertension in this model. We will test the hypothesis that: """"""""The biological actions of the RAS in the brain are the result of tissue specific enzymatic expression of different combinations of angiotensin peptides and the interaction of these peptides with different angiotensin receptor subtypes."""""""" Our major strategy is to combine complementary techniques to create a highly detailed and comprehensive picture of the brain RAS in both control and transgenic rats. Specifically, we will: 1) map in the brain the locations of Ang peptides and processing enzymes by immunocytochemistry; 2) relate these findings to the locations of specific messenger RNA (mRNA) for angiotensinogen (Aogen), renin and Ang-forming enzymes using complementary (DNA (cDNA) probes; an d3) determine the brain and plasma content of Ang I, Ang II, Ang-(1-7) and other N- and C- terminal fragments of Ang I by RIA after separation by HPLC. Measurements will be performed in tissue punches obtained throughout the brain, in the cerebrospinal fluid (CSF), and in the effluent of push-pull cannulae perfusing discrete regions of the brain. These studies will supply a comprehensive picture of the location of precursors and distribution of bioactive peptides. Other experiments will delineate the processing pathways and metabolic disposition of bioactive peptides. Other experiments will delineate the processing pathways and metabolic disposition of AngI using probes and specific inhibitors of angiotensin converting enzyme (ACE) and neutral endopeptidases 21.26, 24.11 and 24.15. The role of Ang-(1-7) in the pathogenesis of hypertension will be assessed by determining the effect of chronic inhibition of peptide formation on blood pressure, renal sympathetic nerve activity and vascular reactivity. These studies will allow us to research in unprecedented detail the role of the brain RAS in the pathogenesis of this form of hypertension.
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