In the rat, two distinct angiotensin type 1a receptor (AT1aR) mRNAs are synthesized from a single AT1aR gene. These transcripts are comprised of exons 1 and 3 (E1,3) and exons 1, 2 and 3 (E1,2,3). These 2 transcripts code for identical receptor proteins and differ only in the lengths of their 5' leader sequence (5'LS). During our previous funding period, we established that the E1,2,3 transcript possesses RNA cis elements within exon 2 that inhibit the efficiency of translation. These splice variant differences in translational efficiency result in higher AT1aR densities and signaling activity in cells expressing the E1,3 transcript compared to those expressing the E1,2,3. Our new studies suggest that the translational efficiency of AT1aR splice variants is diminished in aged rats. We have also found that the response time - i.e., the time it takes to up and down regulate adrenal cortical AT1R densities in response to altered sodium intake is significantly longer in aged rats compared to young animals. This sluggish response time in aged animals is also associated with diminished adrenal responses; i.e., the magnitude of change in plasma aldosterone induced by altered sodium intake is significantly less in aged rats compared to young animals. These observations support clinical and experimental studies that indicate aging is associated with reduced tissue responsiveness to Ang II and have led us to the following general hypothesis: Reduced tissue responsiveness to Ang II that is associated with aging is due to impaired translational regulation of AT1aR transcripts via RNA cis acting elements within exon 2; this dysregulation leads to attenuation in the rapidity, magnitude and threshold sensitivity of the AT1R response to Ang II and thereby contributes to the well known age-associated impairments in fluid and electrolyte homeostasis. We plan to test this hypothesis in young and aged rats under four manipulations of the renin angiotensin system including in Aim 1: response to a low sodium (LS) diet and Ang II infusion at a subpressor dose that mimics the levels achieved by sodium restriction in rats maintained on a NS diet; and in Aim 2: during re-equilibration to a normal sodium (NS) diet after sodium restriction and during re- equilibration to reduced levels of Ang II by challenge with an angiotensin converting enzyme inhibitor in rats maintained on a LS diet for 2 weeks.
In Aim 3, we will investigate the mechanisms of RNA cis acting elements within exon 2 that contribute to Ang II-mediated AT1R regulation in adrenal glomerulosa and vascular smooth muscle cells. The regulation of AT1R expression and activity by altered sodium intake varies in different tissues. Sodium restriction up-regulates the density of AT1Rs on adrenal glomerulosa cells and increases adrenal AT1R- mediated aldosterone secretion. In contrast, sodium restriction down-regulates vascular AT1Rs and reduces vascular contractility to Ang II. AT1Rs are also differentially regulated in specific regions of the kidney and brain by altered sodium intake. In this proposal, we plan to focus on the adrenal and mesenteric resistance arteries because the AT1R is reciprocally regulated in these tissues by altered sodium intake. These studies will determine the post-transcriptional mechanisms of AT1R regulation in these reciprocally regulated tissues and how these mechanisms are impaired in aged rats in response to altered sodium intake. ? ? ?
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