Role of the Na+/H+ antiport in the regulation of the internal pH of Ehrlich ascites tumor cells in culture.

Ehrlich ascites tumor cells contain a Na+ uptake system, which is activated by internal protons and is inhibited by amiloride with an IC50 of 25 microM and by dimethylamiloride with an IC50 of 0.6 microM at 1 mM external Na+. Decrease of external Na+ or addition of amiloride is followed by a decrease of internal pH. Taken together, these findings suggest the presence of an operative Na+/H+ antiport system, which is involved in the regulation of internal pH. We cannot find a significant contribution of a proton pump activated by glycolysis to the pH gradient. At an external pH between 7.0 and 7.6, quiescent cells are more alkaline than exponentially growing cells (0.1 to 0.17 units). Accordingly, an increase of the affinity of the Na+/H+ antiport for internal protons in quiescent cells is demonstrated by the following findings: The internal pH, at which the half-maximal activation of the amiloride-sensitive Na+ uptake occurs, is shifted from 6.85 to 7.1 at 1 mM external Na+. The threshold value of external pH, below which a pronounced effect of amiloride on steady-state internal pH is observed, is shifted from 7.0 in growing to 7.5 in quiescent cells at physiological Na+ concentrations. Therefore, we conclude that quiescent Ehrlich ascites tumor cells raise their internal pH by increasing the affinity of their Na+/H+ antiporter to internal protons. The Na+/H+ antiport cannot be activated further by addition of serum growth factors to quiescent cells. All experiments were performed at bicarbonate concentrations in the medium which do not exceed 0.5 mM.(ABSTRACT TRUNCATED AT 250 WORDS)