The effect of thyroxine on membrane bound (Na⁺, K⁺)-ATPase isolated as a microsomal fraction from rabbit heart and kidney was investigated. In the heart, thyroxine administration produced an increased Ki value (a concentration of ouabain required for half maximal inhibition of (Na⁺, K⁺)-ATPase activity) without alteration of the specific activity of cardiac (Na⁺, K⁺)-ATPase, indicating that the digitalis sensitivity of cardiac (Na⁺, K⁺)-ATPase was decreased. On the contrary, a significant increase of the specific activity of renal (Na⁺, K⁺)-ATPase was observed without change in its digitalis sensitivity. These results suggest that (1) a decreased sensitivity of cardiac (Na⁺, K⁺)-ATPase to digitalis glycosides in thyrotoxic animals may contribute to the decrease in the inotropic and toxic effects of the digitalis glycosides in the hyperthyroid state, and that (2) there may be an organ difference in (Na⁺, K⁺)ATPase.