Toxic and nontoxic effects of ouabain were investigated on frog neuromuscular preparation by measuring the mean quantal content of endplate potentials elicited during repetitive nerve stimulation. In the untreated normal muscles, application of 10 microM ouabain gave rise to a slow exponential increase in the transmitter release (toxic ouabain effect) with a certain delay. This delay was increased with either 100 microM amiloride, a Na(+)-Ca2+ exchange blocker, or the intracellular loading of 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA), a specific intracellular Ca2+ chelator. Measurements of frequency augmentation-potentiation (FAP) revealed a specific nontoxic effect of ouabain: 1 microM ouabain pivoted the long-linear FAP relation counter-clockwise without altering the intercept on the ordinate. Contrary to their action in the toxic effect, both 100 microM amiloride and the intracellular loading of BAPTA failed to counteract the nontoxic effect of 1 microM ouabain. The present results suggest that the toxic and nontoxic effects of ouabain are of different entities. The ouabain-sensitive subtype of Na+,K(+)-ATPase, which is abundant in neural tissues, seems to play a specific role in the process of nontoxic potentiation of transmitter release.