AJP - Cell Physiology, Vol 248, Issue 5 399-C405, Copyright © 1985 by American Physiological Society
A new method for determination of relative ion permeabilities in isolated cells
G. A. Kimmich, J. Randles, D. Restrepo and M. Montrose
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.
