Current-voltage analysis of apical sodium transport in toad urinary bladder: effects of inhibitors of transport and metabolism.

The basal-lateral surface of the epithelium of the urinary bladder of the toad (Bufo marinus) was depolarized by exposure of the serosal surface to 85 mM KCL and 50 mM sucrose. The extent of depolarization appeared to be virtually complete, as evaluated by the invariance in the transepithelial electrical potential difference and conductance on addition of nystatin (a monovalent cation ionophore) to the serosal medium. The Na-specific current (INa) was defined as the current sensitive to the removal of Na from the mucosal medium or inhibitable by addition of amiloride to this medium. In the presence of the high K-sucrose serosal medium, rapid, serial, stepwise clamping of the transepithelial voltage (V) yielded a curvilinear dependence of INa on V; which is taken to represent the I--V curve of the apical Na channels. The constant field equation (Goldman, D.E. 1943; J. Gen. Physiol. 27:37) fits the I--V data points closely, allowing estimates to be made of the permeability to Na of the apical membrane (PNa) and of the intracellular Na activity (Nac). Exposure of the apical surface to amiloride (5 X 10(-7) M) decreased PNa in proportion to the decrease in INa (i.e., approximately 70%) but decreased Nac only 25%. In contrast, an equivalent reduction in INa elicited by exposure of the basal-lateral surface to ouabain was accompanied by only a 20% decrease in PNa and a sixfold increase in Nac. The effects of amiloride on PNa and ouabain on Nac are consistent with the primary pharmacological actions of these drugs. In addition, PNa appears to be under metabolic control, in that 2-deoxyglucose, a specific inhibitor of glycolysis, decreased INa and PNa proportionately, and lowered Nac marginally, effects indistinguishable from those obtained with amiloride.