Regulation of the epithelial Na+ channel by intracellular Na+

The hypothesis that the intracellular Na+ concentration ([Na+](i)) is a regulator of the epithelial Na+ channel (ENaC) was tested with the Xenopus oocyte expression system by utilizing a dual-electrode voltage clamp. [Na+](i) averaged 48.1 +/- 2.2 meg (n = 27) and was estimated from the amiloride-sensitive reversal potential. [Na+](i) was increased by direct injection of 27.6 nl of 0.25 or 0.5 M Na2SO4. Within minutes of injection, [Na+](i) stabilized and remained elevated at 97.8 +/- 6.5 meg (n = 9) and 64.9 +/- 4.4 (n = 5) meg 30 min after the initial injection of 0.5 and 0.25 M Na2SO4, respectively. This increase of [Na+](i) caused a biphasic inhibition of ENaC currents. In oocytes injected with 0.5 M Na2SO4 (n = 9), a rapid decrease of inward amiloride-sensitive slope conductance (g(Na)) to 0.681 +/- 0.030 of control within the first 3 min and a secondary, slower decrease to 0.304 +/- 0.043 of control at 30 min were observed. Similar but smaller inhibitions were also observed with the injection of 0.25 M Na2SO4. Injection of isotonic K2SO4 (70 mM) or isotonic K2SO4 made hypertonic with sucrose (70 mM K2SO4-1.2 M sucrose) was without effect. Injection of a 0.5 M concentration of either K2SO4, N-methyl-D-glucamine (NMDG) sulfate, or 0.75 M NMDG gluconate resulted in a much smaller initial inhibition (<14%) and little or no secondary decrease. Thus increases of [Na+](i) have multiple specific inhibitory effects on ENaC that can be temporally separated into a rapid phase that was complete within 2-3 min and a delayed slow phase that was observed between 5 and 30 min.