1. R 56865 (N-[1-[4-(4-fluorophenoxy)-butyl]-4-piperidinyl]-N-methyl- 2-benzothiazolamine) is a compound known to antagonize cardiac glycoside intoxication. Therefore, the effect of the compound on ouabain binding to intact cardiac tissue as well as cardiac membrane preparations was investigated. 2. The binding of ouabain to highly purified sarcolemmal membranes was not influenced by R 56865 1 x 10(-6) mol l-1 (ouabain: KD = 1.3 x 10(-7) mol l-1, Bmax = 160 pmol mg-1; ouabain + R 56865: KD = 1.4 x 10(-7) mol l-1, Bmax = 168 pmol mg-1). 3. In contrast to the results in purified membranes, the binding of ouabain (10(-8) mol l-1 to 5 x 10(-7) mol l-1) to intact atria was significantly reduced. 4. Ouabain, 5 x 10(-7) mol l-1, led to a transient positive inotropic effect of about 220% followed by a developing negative inotropic effect after 3 h. R 56865, 10(-7) mol l-1, led to a maximal positive inotropic effect of about 290% also followed by a delayed decline of contractile force. A tenfold higher concentration of R 56865 led to sustained positive inotropic effect of about 250% in the same time interval. 5. The different effects of R 56865 on ouabain binding in subcellular preparations and intact tissue do not support the view that R 56865 interferes directly with the action of ouabain on Na/K-ATPase. An indirect effect, which may be mediated by a lowered intracellular sodium load is discussed.