A new method for determination of relative ion permeabilities in isolated cells

The unidirectional influx of the lipophilic cation tetraphenylphosphonium (TPP+) into isolated epithelial cells is a function of the membrane potential that exists across the cellular plasma membrane. Because of the potential dependence, [14C]TPP+ influx can be used as a qualitative sensor of changes in the membrane potential induced by diffusion of ions after the experimental imposition of transmembrane ion gradients. This report describes a "crossover" procedure in which the influx of [14C]TPP+ during systematic changes in the ionic composition of incubation media is used to identify conditions in which no change in membrane potential occurs. The ion ratio at the crossover provides a measure of the relative permeabilities of the two test ions being compared. By using this approach, the ion permeabilities for intestinal epithelial cells prepared from White Rock chickens can be ranked relative to the permeability of Na+ (PNa), i.e., when PNa is equal to 1.0. The permeability sequence and relative values for ion permeability in this system are tris(hydroxymethyl)aminomethane-gluconate (less than 0.1) less than Li+ (0.3) less than Na+ (1.0) less than Cl- (2.0) less than K+ (6.0) = NO3- (6.0) less than SCN- (18) less than K+ + valinomycin (40). The procedure is general enough in principle to be of broad application to a wide variety of cell or membrane vesicle preparations.