ROS activate KCl cotransport in nonadherent Ehrlich ascites cells but K+ and Cl- channels in adherent Ehrlich Lettré and NIH3T3 cells.

Addition of H(2)O(2) (0.5 mM) to Ehrlich ascites tumor cells under isotonic conditions results in a substantial (22 +/- 1%) reduction in cell volume within 25 min. The cell shrinkage is paralleled by net loss of K(+), which was significant within 8 min, whereas no concomitant increase in the K(+) or Cl(-) conductances could be observed. The H(2)O(2)-induced cell shrinkage was unaffected by the presence of clofilium and clotrimazole, which blocks volume-sensitive and Ca(2+)-activated K(+) channels, respectively, and is unaffected by a raise in extracellular K(+) concentration to a value that eliminates the electrochemical driving force for K(+). On the other hand, the H(2)O(2)-induced cell shrinkage was impaired in the presence of the KCl cotransport inhibitor (dihydro-indenyl)oxyalkanoic acid (DIOA), following substitution of NO(3)(-) for Cl(-), and when the driving force for KCl cotransport was omitted. It is suggested that H(2)O(2) activates electroneutral KCl cotransport in Ehrlich ascites tumor cells and not K(+) and Cl(-) channels. Addition of H(2)O(2) to hypotonically exposed cells accelerates the regulatory volume decrease and the concomitant net loss of K(+), whereas no additional increase in the K(+) and Cl(-) conductance was observed. The effect of H(2)O(2) on cell volume was blocked by the serine-threonine phosphatase inhibitor calyculin A, indicating an important role of serine-threonine phosphorylation in the H(2)O(2)-mediated activation of KCl cotransport in Ehrlich cells. In contrast, addition of H(2)O(2) to adherent cells, e.g., Ehrlich Lettré ascites cells, a subtype of the Ehrlich ascites tumor cells, and NIH3T3 mouse fibroblasts increased the K(+) and Cl(-) conductances after hypotonic cell swelling. Hence, H(2)O(2) induces KCl cotransport or K(+) and Cl(-) channels in nonadherent and adherent cells, respectively.