Dose-dependent effect of cardiotonic steroids on Na,K-ATPase activity from pig and rat kidneys. Dose-dependent inhibition of Na,K-ATPase activity from pig (A) and rat (B) kidneys by different cardiotonic steroids (ouabain, digoxin, marinobufagenin), which were added into incubation media. Dose-dependent “irreversible” inhibition of Na,K-ATPase from pig kidneys in E2P-conformation by ouabain (C) or digoxin (D). Cardiotonic steroids were added into preincubation media and stayed with the enzyme in E2P-conformation for the time that was necessary to reach a steady state. Then aliquots of preincubation medium (20 μL) were transferred into the incubation medium with a volume of 2 mL to assay the enzyme activity. For details, see “Materials and Methods”. Concentration of Na,K-ATPase: 1—20 nM; 2—80 nM; 3—400 nM. Means ± S.D. from 3 experiments performed in triplicates are shown.