Molecular regulation of endothelin‐1 synthesis by renal collecting duct

Collecting duct (CD)‐derived endothelin‐1 (ET‐1) modulates systemic blood pressure and Na excretion, however little is known about control of CD ET‐1 synthesis. To address this, rat inner medullary CD (IMCD) were studied in primary culture. ET‐1 release and mRNA levels were unchanged by blockade of NO synthase, cyclooxygenase, ERK, JNK or p38 MAPK, but were reduced (by 40%) after inhibition of PKC. ET‐1 release and mRNA levels were decreased by 60–80% by chelation of intracellular (BAPTA) or extracellular (EDTA) Ca2+, inhibition of calmodulin (W‐7) or CaMKII (KN‐93), and decreased by 35% by nifedipine. Transfection of IMCD with rat ET‐1 promoter‐luciferase constructs revealed maximal activity within 1.9 kb 5′ to the transcription start site; 10, 30, and 80% of this activity were in the 0.25, 0.56 kb, and 3.2 kb promoter regions, respectively. W‐7 markedly inhibited activity of the 1.9 kb, but not 0.56 kb, promoter region. Transfected rat aortic endothelial cells had maximal activity in the 0.56 kb region (as compared to the 1.9 and 3.2 kb regions). In summary, IMCD ET‐1 synthesis is regulated by PKC and a Ca2+/calmodulin‐dependent pathway. The Ca2+/calmodulin‐sensitive pathway is active in IMCD, but not endothelial cells. This suggests that that IMCD‐specific enhancer elements exist within the ET‐1 promoter that confer unique calcium responsiveness.