Human decidual tissue from uncomplicated term pregnancies was incubated in vitro using a cross-over design of incubation lasting for 72 h. The decidual issue of each membrane was added sequentially to media with the osmolalities 252, 315 and 387 mmol/kg, and the different osmolalities were in 33 experiments induced by changing the concentration either of sodium chloride, potassium chloride, mannitol, sucrose, or choline chloride. At 387 mmol/kg all substances elicited a significant increase in Prl secretion compared with the 315 mmol/kg media (14-27%) or the 252 mmol/kg media (26-46%). When the sodium chloride, potassium chloride, sucrose, and choline chloride media at 387 mmol/kg were compared in another 7 experiments, potassium chloride increased Prl secretion more effectively than the others (10%, P less than 0.05). The incubation cross-over design did not permit determination of the decidual Prl content after incubation in each of the various media, but in 28 experiments the mean decidual Prl content was 12.7 mIU per 100 mg dry tissue (range 4-30) before and 8.7 mIU per 100 mg dry tissue (1-17) after the experiments. The mean amount of Prl secreted during the successive incubations was 152.2 mIU per 100 mg dry tissue (19-672) which showed the secretion to be a result of continued Prl production. The effect of osmolality was independent of the basal production rate. These results indicate that the intracellular ionic concentrations, probably of potassium ion or of chloride ion, are of importance in the regulation of the synthesis and secretion of decidual Prl in vitro.