Novel Regulation of the Epithelial Na+ Channel by Soluble Adenylyl Cyclase in Kidney Collecting Duct Cells

Abstract

Alkalosis impairs natriuresis, but the mechanisms involved are unclear. In contrast to transmembrane adenylyl cyclases, soluble adenylyl cyclase (sAC) mediates HCO3−‐induced elevation of cAMP in intracellular microdomains. We thus hypothesized that sAC may regulate the epithelial Na+ channel (ENaC) in the kidney. Confocal images of rat kidney revealed co‐immunolocalization of sAC and α‐ENaC in collecting duct principal cells. Mouse polarized kidney collecting duct (mpkCCDc14) cells, which express sAC by immunoblot, were used as an in vitro model system. The specific sAC inhibitor KH7 (60 μM) inhibited amiloride‐sensitive short‐circuit currents (Isc) by >50% within 30 min. Similar ENaC inhibition was found with a catechol estrogen, an alternative sAC inhibitor. Aldosterone (1 μM) doubled ENaC‐dependent Isc over 4 h, an effect that was abolished by KH7. Conversely, 8‐Br‐cAMP overrode the inhibition of ENaC by KH7. Moreover, KH7 fully blocked forskolin‐induced ENaC apical recycling, suggesting that sAC activity is required for this process. Consistent with this idea, neither sAC over‐expression nor KH7 treatment modulated ENaC currents expressed in Xenopus oocytes, where significant ENaC recycling has been shown not to occur. In summary, these results suggest that sAC is a novel regulator of ENaC. sAC activity is necessary for both aldosterone stimulation of ENaC currents and ENaC recycling in CCD cells.