Na-K-ATPase localization in teleost urinary bladder by [3H]ouabain autoradiography

Previous studies on the urinary bladder of the seawater-acclimated winter flounder (pseudopleuronectes americanus) demonstrated that active Na and Cl transport were ouabain sensitive. This suggested a relationship between the Na pump and Na-K-ATP-ase. The specific binding of [H]ouabain to Na-K-ATPase provides a means of localizing the site of active Na transport. In isolated bladders, a positive linear correlation (r= 0.89) was found between the active Na transport rate and the Na-K-ATPase activity. Ouabain binding by the bladder surface appeared to be saturable and relatively specific, e.g., was reduced by a high K concentration. When only the mucosal side of the bladder was exposed to 5 muM ouabain, both inhibitory effects and binding were small and are explained by finite permeability of the bladder to ouabain. In contrast, binding and inhibitory effects from the serosal side were much greater. Autoradiographs demonstrated that [3H]ouabain was bound only to the serosal side of the epithelial cells. Ultrastructural examination revealed that the area of ouabain binding coincided with the basal and lateral plasma membranes.