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Dietary sodium depletion has multiple diverse effects on ion transport in the rat colon, including both the induction and inhibition of electroneutral NaCl absorption in proximal and distal colon of rat, respectively. To establish the mechanism of the differential regulation of Na⁺ absorption by sodium depletion, this study utilized 1) HOE-694, a dose-dependent inhibitor of Na⁺/H⁺exchanger (NHE) isoforms, in studies of proton gradient-driven²²Na uptake (i.e., Na⁺/H⁺exchange) by apical membrane vesicles (AMV);2) Northern blot analyses of NHE isoform-specific mRNA abundance; and3) Western blot analyses of NHE isoform-specific protein expression. HOE-694 inhibition studies establish that 25 μM HOE-694-sensitive (NHE2) and 25 μM HOE-694-insensitive (NHE3) Na⁺/H⁺exchange activities are present in AMV of both proximal and distal colon of normal rats. In proximal colon, dietary sodium depletion enhanced …