Characterization of apical membrane Cl-dependent Na/H exchange in crypt cells of rat distal colon

A novel Cl-dependent Na/H exchange (Cl-NHE) has been identified in apical m embranes of crypt cells of rat distal colon. The presence of Cl is required for both outward proton gradient-driven Na uptake in apical membrane vesic les (AMV) and Na-dependent intracellular pH recovery from an acid load in t he crypt gland. The present study establishes that Cl-dependent outward pro ton gradient-driven Na-22 uptake 1) is saturated with increasing extravesic ular Na concentration with a Michaelis constant (K-m) for Na of similar to 24.2 mM; 2) is saturated with increasing outward H concentration gradient w ith a hyperbolic curve and a K-m for H of similar to1.5 muM; 3) is inhibite d by the Na/H exchange (NHE) inhibitors amiloride, ethylisopropylamiloride, and HOE-694 with an inhibitory constant (K-i) of similar to 480.2, 1.1, an d 9.5 muM, respectively; 4) is inhibited by 4,4'-diisothiocyanostilbene-2,2 '-disulfonic acid, an anion exchange inhibitor at low concentration and a C l channel blocker at high dose, and by 5-nitro-2(3-phenylpropylamino) benzo ic acid, a Cl channel blocker, with a K-i of similar to 280.6 and 18.3 muM, respectively; and 5) substantially stimulated Cl-NHE activity by dietary N a depletion, which increases plasma aldosterone and inhibits NHE in surface cell AMV. These properties of Cl-NHE are distinct from those of NHE1, NHE2 , and NHE3 isoforms that are present in colonic epithelial cells; thus thes e results suggest that the colonic crypt cell Cl-NHE is a novel NHE isoform .