The renin-angiotensin-aldosterone system plays important roles in the regulation of blood pressure and salt and water balance. Thus, the regulation of renin and aldosterone release has important implications in diseases as hypertension and congestive heart failure. While AII, ACTH and potassium are the major regulators of aldosterone release, other factors may modulate their actions. Several lines of evidence suggest that other cells present in the adrenal gland may contribute to steroidogenesis. Of particular interest are endothelial cells (ECs) since these cells are in close anatomical relationship to steroidogenic cells. In this regard, we found that the release of aldosterone was grater when zona glomerulosa (ZG) cells were incubated in the presence of ECs than when ZG cells were incubated alone. This stimulatory effect did not occur with smooth muscle cells, pericytes or fibroblast. The steroidogenic factor was present in the media of ECs and could be transferred to ZG cells and promote aldosterone release. The factor was found to be a protein but was not a known steroidogenic peptide as AII, bradykinin or endothelin. The proposed studies will test the hypothesis that ECs, which are in close anatomical proximity to ZG cells in the adrenal gland, release a factor that modulates or regulates aldosterone release. We propose to purify and identify this endothelial derived steroidogenic factor (EDSF) and determine how its release and actions are regulated. These studies will be conducted in cultured bovine adrenal ZG cells and ECs, the perfused rat adrenal and anesthetized rats. The hypothesis will be tested by the following specific aims; (1) We will further characterize the influence of ECs on aldosterone release from ZG cells. Using co-incubations of ZG cells and ECs and EC conditioned medium, we will determine the effect of various agonists on the release of EDSF from ECs and on the action of EDSF on ZG cells. (2) We will purify and determine the amino acid composition and sequence of EDSF. (3) We will study the release and the regulation of the synthesis of EDSF by various agonists. (4) We will study the mechanism of action of EDSF on ZG cells. The studies will compare the site of action of synthetic EDSF will be investigated using specific assays and inhibitors. Additionally, we will determine if AG cells have specific binding sites for EDSF. (5) We will determine the role of the endothelial factor in the regulation of steroidogenesis in vitro in the in situ perfused adrenal gland and in vivo in anesthetized rats. The contribution of endothelial factor to flow- induced steroidogenesis will also be examined. These studies should provide new insights into the contribution of EC to the regulation of aldosterone release.
