ABSTRACT The role of Na''-H+ exchange in Na ' transport across the apical membrane was evaluated in Necturus gallbladder epithelium by means of intracellular Na ' activity (aNai) and "Na ' uptake measurements. Under control conditions, complete replacement of Na ' in the mucosal solution with tetramethylammonium reduced aNai from 14.0 to 6.9 mM in 2 min (P < 0.001). Mucosal addition of the Na'-H ' exchange inhibitor amiloride (10- ' M) reduced aNa i from 15.0 to 13.3 mM (P < 0.001), whereas bumetanide (10-5 and 10' M) had no effect. Na ' influx across the apical membrane was studied by treating the tissues with ouabain, bathing them in Na-free solutions, and suddenly replacing the mucosal solution with an Na-containing solution. When the mucosal solution was replaced with Na-Ringer's, aNai increased at-I 1 mM/ min. This increase was inhibited by 54 % by amiloride (10-'M, P< 0.001) and was unaffected by bumetanide (10-5 M). Amiloride-inhibitable Na ' fluxes across the apical membrane were also induced by the imposition ofpH gradients. Na' influx was also examined in tissues that had not been treated with ouabain. Under control conditions, "Na ' influx from the mucosal solution into the epithelium was linear over the first 60 s and was inhibited by 40 % by amiloride (10- ' M, P < 0.001) and by 19 % by bumetanide (10-5 M, P < 0.025). We conclude that Nat-H ' exchange is a major pathway for Na ' entry in Necturus gallbladder, which accounts for at least half of apical NO influx both under transporting conditions and during exposure to ouabain. Bumetanide-inhibitable Na ' entry mechanisms may account for only a smaller fraction of Na ' influx under transporting conditions, and cannot explain influx in ouabain-treated tissues. These results support the hypothesis that NaCl entry results primarily from the operation of parallel Na'-H ' and Cl--HC09 exchangers, and not from a bumetanide-inhibitable NaCl cotransporter. Downloaded from jgp.rupress.org on February 21, 201
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