Mechanism(s) of chloride transport in human distal colonic apical membranevesicles

Previous studies from our laboratory have demonstrated the presence of an e lectroneutral Cl-/HCO3- exchange process across the human proximal colonic apical membrane vesicles (AMV). However, very little is known about the mec hanism(s) of chloride transport in the apical membrane of the human distal colon. Utilizing AMV purified from organ donor distal colonic mucosa and a rapid Millipore filtration technique, the mechanisms of Cl-36(-) uptake int o these vesicles were examined. Outwardly directed OH- and HCO3- gradients markedly increased the uptake of Cl-36(-) into these vesicles, demonstratin g a transient accumulation over the equilibrium uptake. Voltage clamping in the presence of K+/valinomycin reduced the OH- and HCO3- gradient-stimulat ed Cl-36(-) uptake into these vesicles by similar to 30% indicating that th e conductive Cl- uptake pathway was present in these vesicles along with th e electroneutral exchange process. Under voltage-clamped conditions, the in hibitors the bicarbonate transporters, DIDS and SITS (1 mM), inhibited OH- and HCO3- gradient-stimulated Cl-36(-) uptake by similar to 50%. Acetazolam ide showed small but significant inhibition of chloride uptake. Amiloride, bumetanide, and furosemide failed to inhibit Cl-36(-) uptake. Chloride upta ke into these vesicles exhibited saturation kinetics with an apparent K-m f or chloride of 16.7 mM and a V-max of 5.9 nmol/mg/15 sec. Chloride, acetate , nitrate, but not sulfate (50 mM each), inhibited 5 MM Cl-36(-) uptake. In wardly directed gradients of Na+, K+ or both together did not stimulate chl oride uptake into these vesicles indicating that the uptake of Cl- and Nain human distal colonic AMV does not involve Na-Cl or Na-K-2Cl cotransport. In conclusion, these studies demonstrate that Cl- transport across the api cal membranes of human distal colon involves both conductive pathway and el ectroneutral Cl-/HCO3- (OH-) exchange processes. In view of our previous de monstration of a Na+/H+ exchange process in these AMV, we propose that the operation of dual ion exchange mechanisms of Na+/H+ and Cl-/HCO3- is the pr imary mode of electroneutral NaCl absorption across the apical membranes of the enterocytes of the human distal colon.