Human decidual explants were incubated in vitro in media containing 10(-5) mol/l ouabain or 10(-4) mol/l ouabain or 0.5 X 10(-4) mol/l bumetanide to change the intracellular ionic concentrations. In isosmotic incubations (osmolality 315 mmol/kg), no significant effect of either 10(-5) mol/l ouabain or 0.5 X 10(-4) mol/l bumetanide was found on the decidual prolactin secretion (D-PRL-s). 10(-4) mol/l ouabain significantly decreased both decidual prolactin production and D-PRL-s (P less than 0.05) and was therefore not used for the following cross-over experiments. Hyperosmotic media (387 mmol/kg) were produced by changing the concentration of either sodium chloride, potassium chloride, or mannitol. All increased D-PRL-s compared with the isosmotic media (315 mmol/kg). 10(-5) mol/l ouabain significantly diminished the increase otherwise elicited by the sodium chloride and the mannitol hyperosmotic media. However, in the hyperosmotic potassium chloride medium with 10(-5) mol/l ouabain, the D-PRL-s remained increased. The hyperosmotic medium (252 mmol/kg) reduced D-PRL-s compared with the isosmotic media (315 mmol/kg) and no significant effect of ouabain was found. Bumetanide did not change the D-PRL-s into any of the hypo- or hyperosmotic media compared with the secretion at 315 mmol/kg. Based on experience from other cell types, the results further indicate that the intracellular ionic concentrations could be of importance to the secretion of decidual prolactin in vitro.