Hydrogen peroxide stimulates chloride secretion in primary inner medullary collecting duct cells via mPGES-1-derived PGE₂

We investigated the role and mechanism of H₂O₂ in regulation of NaCl transport in primary inner medullary collecting duct (IMCD) cells. IMCD cells were isolated from wild-type mice and grown onto semipermeable membranes, and short-circuit current (I_sc) was determined by Ussing chamber. Exposure of IMCD cells to H₂O₂ at a range of 100-300 μM caused a rapid increase in I_sc in a time- and dose-dependent manner. This increase was almost abolished by the cystic fibrosis transmembrane conductance regulator (CFTR) Cl^- channel inhibitors diphenylamine-2-carboxylic acid (DPC) and CFTR_inhibitor-172. In contrast, the magnitude of stimulation was unaffected by the epithelial Na ⁺ channel (ENaC) inhibitor amiloride. The H₂O ₂-induced Cl^- secretion was significantly inhibited by indomethacin, as well as by microsomal PGE synthase-1 (mPGES-1) deficiency. Like H₂O₂, PGE₂ treatment induced a twofold increase in I_sc that was reduced by the protein kinase A (PKA) inhibitors H-89 and KT5720. These data suggest that H₂O₂ stimulates CFTR Cl^- channel-mediated Cl^- secretion through cyclooxygenase- and mPGES-1-dependent release of PGE₂ and subsequent activation of PKA.