Apical NH 4 Cl acts to reduce intracellular Na concentration in mTAL via cellular depolarization (1098.6)

Abstract

We have recently demonstrated that both cellular acidification and low intracellular [Na] promote NADPH oxidase activity in mTAL via activation of the voltage‐gated proton channel Hv1. NH 4 + is normally present in the luminal fluid and its concentration increases during acidosis. It is known that apical uptake of NH 4 + promotes intracellular acidification in mTAL, however the effect of apical NH 4 Cl on intracellular [Na] has not been studied. The goal of the current study was to determine the effect of increasing apical [NH 4 Cl] on intracellular [Na] in mTAL. We hypothesize that increasing apical NH 4 Cl concentration reduces intracellular [Na]. Measurements were performed using a FluoStar Omega fluorescent plate reader in cultured mTAL cells loaded with the Na sensitive dye CoroNa Green (n=6). Addition of NH 4 Cl to the media (HBSS pH 7.40) dose dependently reduced intracellular [Na] in cultured mTAL. Both the NKCC2 inhibitors furosemide and bumetanide lowered intracellular [Na] but did not prevent the action of NH 4 Cl to further reduce intracellular [Na]. Depolarization of mTAL using KCl reduced intracellular [Na] and greatly blunted the response to 8mM NH 4 Cl (‐62%, P<0.0002). We conclude that physiologically relevant increases in apical [NH 4 Cl] (4‐8mM) lower intracellular [Na] in mTAL and that these changes are mediated predominately by cellular depolarization. We speculate that during acidosis, increased NH 4 Cl delivery to the mTAL promotes NADPH oxidase activity in part by lowering intracellular [Na]. Grant Funding Source : AHA SDG 10SDG4150061