Cation Permeability of a Cloned Rat Epithelial Amiloride‐Sensitive Na + Channel

1 Conductance of heterotrimeric rat epithelial Na + channels (α,β,γ‐rENaCs) for Li + and Na + in planar lipid bilayers was a non‐linear function of ion concentration, with a maximum of 30.4 ± 2.9 pS and 18.5 ± 1.9 pS at 1 M Li + and Na + , respectively. 2 The α,β,γ‐rENaC conductance measured in symmetrical mixtures of Na + –Li + (1 M) exhibited an anomalous mole fraction dependence, with a minimum at 4:1 Li + to Na + molar ratio. 3 Permeability ratios P K / P Na and P Li / P Na of the channel calculated from the biionic reversal potentials were dependent on ion concentration: P K / P Na was 0.11 ± 0.01, and P Li / P Na was 1.6 ± 0.3 at 50 m m ; P K / P Na was 0.04 ± 0.01 and P Li / P Na was 2.5 ± 0.4 at 3 M, but differed from the ratios of single‐channel conductances in symmetrical Li + , Na + or K + solutions. The permeability sequence determined by either method was Li + > Na + > K + ≫ Rb + > Cs + . 4 Predictions of a model featuring two binding sites and three energy barriers (2S3B), and allowing double occupancy, developed on the basis of single ion current–voltage relationships, are in agreement with the observed conductance maximum in single ion experiments, conductance minimum in the mole fraction experiments, non‐linearity of the current–voltage curves in biionic experiments, and the concentration dependence of permeability ratios. 5 Computer simulations using the 2S3B model recreate the ion concentration dependencies of single‐channel conductance observed for the immunopurified bovine renal amiloride‐sensitive Na + channel, and short‐circuit current in frog skin, thus supporting the hypothesis that ENaCs form a core conduction unit of epithelial Na + channels.