Localization of sodium pump sites in frog urinary bladder.
[3H]Ouabain binding in frog and toad urinary bladder was investigated by short-circuit current (SCC), scintillation counting and autoradiographic techniques. SCC data and analysis of tissue digests following serosal exposure to ouabain showed that ouabain binding and inhibition of Na+ transport was completely reversible in toad bladder whereas, in frog bladder, [3H]ouabain was tightly bound and Na+ transport remained suppressed even after a 60-min washout. Mucosal exposure of frog bladder to [3H]ouabain or serosal exposure after preincubation with unlabeled ouabain led to a marked reduction in binding. Specificity of binding was assessed further  by adjusting the concentration of certain (Na+ -K+)-ATPase ligands(K+, ATP) to levels known to reduce ouabain binding. High K+ concentrations and depletion of endogenous ATP by incubation under anoxic conditions resulted in a significant drop in [3H]ouabain binding. Autoradiographic analysis showed that grains are localized primarily to the basolateral plasma membranes of the granular cells, providing direct morphological evidence for the location of Na+ pumps at these sites. Although autoradiographs did not provide sufficient resolution to rule out unequivocally ouabain binding to the mitochondria-rich cell, morphological evidence suggests that grain densities are significantly higher between adjacent granular cells than between granular cell-mitochondria-rich cell interfaces.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D001693:Biological Transport, Active; D002024:Bufo marinus; D010042:Ouabain / Q000378:metabolism; D011892:Rana catesbeiana; D011955:Receptors, Drug; D012964:Sodium / Q000378:metabolism*; D001743:Urinary Bladder / Q000378:metabolism* / Q000648:ultrastructure
D011955:Receptors, Drug; D010042:Ouabain; D012964:Sodium; D000251:Adenosine Triphosphatases