Aldosterone Rapidly Activates Na/H~+ Exchange in M-1 Cortical Collecting Duct Cells via a PKC-MAPK Pathway

摘要

Background: In this study, the mechanism of the rapid non-genomic effect of aldosterone on Na~+/H~+ exchanger (NHE)-mediated intracellular pH (pH|) recovery from an acid load in murine M-1 cortical collecting duct cells was assessed. Methods: Spectrofluorescence microscopy and Western blot analysis was carried out and NH_4CI was used to induce the acid load. Results: Aldosterone (10 n/W) induced a rapid (<5 min) concentration-dependent increase in pHi recovery in M-1 cells, an effect mimicked by its precursor deoxycorticosterone (1 n/V7). This response was unaffected by the mineralocorticoid receptor (MR) antagonist spironolactone (10muM) but was significantly reduced by the NHE antagonists 5'-(N-ethyl-N-isopropyl)amiloride (EIPA) (20muM) and cariporide (1 muM). The PKC inhibitor chelerythrine chloride (1muM) significantly attenuated the aldosterone-induced increase in NHE1 activity. HBDDE (80muM), a PKC_a inhibitor, inhibited the rapid aldosterone effect whereas rottlerin (15muM), a PKC§ antagonist did not. The glucocorticoid receptor agonists hydrocortisone (1 muM) and dexamethasone (100muM) decreased NHE activity, whereas the synthetic mineralocorticoid fludrocortisone (1 n/W) had no significant effect. MAPK inhibition using PD98059 (25muM) significantly attenuated the rapid aldosterone effect; Western blot analysis showed that aldosterone activation of ERK 1/2 was unaffected by pretreatment with spironolactone but was inhibited following chelerythrine chloride. Conclusion: Aldosterone causes a rapid non-genomic increase in NHE1 activity in M-1 cells via a PKC_a/MAPK pathway independent of the classical MR.