Abstract 184: Demonstration Of An Aldosterone-Response Pathway In Human Adrenal
Aldosterone (aldo) stimulates Na+ reabsorption by upregulation of the amiloride-sensitive epithelial Na+ channel (ENaC). In many ENaC-expressing tissues, Nedd4L and the kinase Sgk1 determine the number of ENaC channels at the membrane and responsiveness to aldo. 11β HSD type II (11β hydroxysteroid dehydrogenase type II) confers mineralocorticoid specificity. We have unexpectedly found expression of ENaC in human adrenal zona glomerulosa (ZG) cells and have investigated the presence and possible function of a complete aldo response pathway. Tissue was obtained from patients undergoing adrenalectomy and used to generate primary cultures of ZG cells. The effects of amiloride analogues on aldo release from immortalised (H295R) cells were determined by RIA. RT-PCR showedα, β and γ subunits of ENaC, the mineralocorticoid receptor (MR), Nedd4L, Sgk1 and 11βHSD type II. Using Western blotting and cell fractionationα, β and γ ENaC were demonstrated in plasma membrane and cytosol and upon aldo stimulation of primary cultured human ZG cells, the levels of β and γ ENaC expression at the cell surface increased. Concentration-response studies for effects of Na+, K+ and amiloride analogues on 24h aldo release from H295R cells showed stimulation by K+, suppression by both Na+ and EIPA (blocker of Na+ −H+ exchange, NHE) and no change with benzamil (ENaC blocker). Our experiments suggest that an entire aldo response pathway is present in human ZG. Since NHE blockade or Na+ both suppress aldo release, NHE is unlikely to be the route by which the adrenal senses Na+. We speculate that ENaC is the route by which physiological changes in Na+ suppress aldo secretion and provides aldo-mediated auto feedback.