Effect of insulin on Na+,K(+)-ATPase in rat collecting duct.

1. The collecting duct is involved in the whole antinatriuretic effect of insulin, as indicated in vitro by the stimulatory effect of the hormone on ouabain-sensitive 86Rb+ uptake. Since Na+,K(+)-ATPase drives Na+ reabsorption, the contribution of the Na+ pump to the effect of insulin was investigated in rat isolated cortical and outer medullary collecting duct. 2. Insulin enhanced ouabain-sensitive 86Rb+ uptake in the absence, as well as in the presence, of either 5 x 10(-4) M amiloride or 10(-3) M hydrochlorothiazide (HCT). Maximal ouabain-sensitive 86Rb+ uptake, measured in Na(+)-loaded tubules, was also enhanced by insulin. The insulin effect persisted both in the absence of external Na+, when the Na+,K(+)-ATPase operates in a Rb(+)-Rb+ exchange mode, and in tubules depolarized by a high external concentration (20 mM) of Rb+ or by addition of 3 mM Ba2+. 3. Insulin treatment did not alter the intracellular Na and K concentrations, the specific binding of [3H]ouabain measured in intact tubules, or the hydrolytic activity of Na+,K(+)-ATPase measured after permeabilization of the tubule cells. 4. In conclusion, in the rat collecting duct, insulin increased Na+,K(+)-ATPase-mediated cation transport independently of Na+ availability, membrane potential and recruitment of pump units. The effect of insulin was lost after cell permeabilization, suggesting the presence of a cytosolic factor which controls the turnover of Na+,K(+)-ATPase.