The distal colon absorbs K+ (JK) and secretes H+ (JH) by what is thought to be an H(+)-K(+)-adenosinetriphosphatase (H(+)-K(+)-ATPase). However, the colonic ATPase differs structurally and functionally from the gastric H(+)-K(+)-ATPase. To evaluate the link between JH and JK, JH and JK were simultaneously measured with ion-specific electrodes in segments of rat distal colon. JH and JK averaged 0.40 +/- 0.03 and 0.30 +/- 0.03 mu eq.h-1.cm-2 (n = 191), but JH and JK did not correlate (r = 0.005, not significant). The gastric H(+)-K+ pump inhibitors SCH-28080 (100 microM) and omeprazole (100 microM), as well as a vacuolar H(+)-ATPase inhibitor, bafilomycin A1 (10 microM), did not affect JH or JK. However, the Na(+)-K(+)-ATPase inhibitors ouabain (1 mM) and N-ethylmaleimide (10 microM) inhibited JK but not JH. Although 1 mM orthovanadate inhibited both JH and JK, at lower concentrations orthovanadate only affected JK. Furthermore, removing K+ from the medium did not affect JH. Secondary hyperaldosteronism increased both JH and JK; however, ouabain (1 mM) reduced JK but not JH. Cl(-)-free medium inhibited voltage-insensitive JH and voltage-sensitive JK. Medium pH affected JH, but that effect was contrary to the effect that pH had on Rb+ flux. These data failed to identify a relationship between JH and JK and appear to suggest that JH and JK occur by separate pathways.