Extracellular histidine residues crucial for Na+ self-inhibition of epithelial Na+ channels

Epithelial Na+ channels ( ENaC) participate in the regulation of extracellular fluid volume homeostasis and blood pressure. Channel activity is regulated by both extracellular and intracellular Na+. The down- regulation of ENaC activity by external Na+ is referred to as Na+ self-inhibition. We investigated the structural determinants of Na+ self-inhibition by expressing wildtype or mutant ENaCs in Xenopus oocytes and analyzing changes in whole-cell Na+ currents following a rapid increase of bath Na+ concentration. Our results indicated that wild-type mouse alphabetagammaENaC has intrinsic Na+ self-inhibition similar to that reported for human, rat, and Xenopus ENaCs. Mutations at His(239) (gammaH239R, gammaH239D, and gammaH239C) in the extracellular loop of the gammaENaC subunit prevented Na+ self- inhibition whereas mutations of the corresponding His(282) in alphaENaC (alphaH282D, alphaH282R, alphaH282W, and alphaH282C) significantly enhanced Na+ self-inhibition. These results suggest that these two histidine residues within the extracellular loops are crucial structural determinants for Na+ self-inhibition.