Effect of potassium depletion of Hep 2 cells on intracellular pH and on chloride uptake by anion antiport

Abstract

The effect of K⁺ depletion of Hep 2 cells on ion fluxes, internal pH, cell volume, and membrane potential was studied. The cells were depleted of K⁺ by incubation in K⁺‐free buffer with or without a preceding exposure to hypotonic medium. Efflux of K⁺ in cells not exposed to hypotonic medium was inhibited by furosemide or by incubation in Na⁺‐free medium, indicating that in this case at least part of the K⁺ efflux occurs by Na⁺/K⁺/Cl⁻ cotransport. After exposure to hypotonic medium, K⁺ efflux was not inhibited by furosemide, whereas it was partly inhibited by 4,4′‐diisothiocyano‐2,2′‐stilbene‐disulfonic acid (DIDS). Exposure to hypotonic medium induced acidification of the cytosol, apparently because of efflux of protons from intracellular acidic vesicles. When isotonicity was restored, a rebound alkalinization of the cytosol was induced, because of activation of the Na⁺/H⁺ antiporter. While hypotonic shock and a subsequent incubation in K⁺‐free buffer rapidly depolarized the cells, depolarization occurred much more slowly when the K⁺ depletion was carried out by incubation in K⁺‐free buffer alone. The cell volume was reduced in both cases. K⁺ depletion by either method strongly reduced the ability of the cells to accumulate ³⁶Cl⁻ by anion antiport, and K⁺‐depleted cells were unable to increase the rate of ³⁶Cl⁻ uptake in response to alkalinization of the cytoscl.