Lack of correlation between [ 3H]ouabain binding and Na-K ATPase inhibition in rat aorta

Abstract

The binding of [ 3H]ouabain to intact strips of rat aorta was compared with the ability of ouabain to inhibit the uptake of 86Rb by the same preparation. When a cold temperature wash was used to process tissues after binding of [ 3H]ouabain, a class of relatively high affinity binding sites was found (K D = 1.2 × 10 −7 M). Binding was saturable and sensitive to both ATP depletion and elevated potassium. Elevation of cytoplasmic Ca 2+ levels by phenylephrine or c-AMP levels by theophylline and terbutaline had no influence on [ 3H]ouabain binding. Ouabain inhibition of 86Rb uptake progressed to 60% of the total 86Rb uptake at 2 × 10 −3 M from a threshold of about 10 −5 M. Half-maximal inhibition by ouabain occurred at a concentration of 10 −4 M. The disparity between [ 3H]ouabain binding and inhibition of 86Rb uptake indicates that the high affinity binding site in the rat does not contribute to inhibition of Na-K ATPase function.