The effects of prostaglandins E1, E2, F1alpha, and F2alpha on prolactin and growth hormone synthesis and secretion were studied in cultured rat pituitary cells. Total extracellular accumulation of prolactin was measured by radioimmunoassay. When hormone synthesis was studied, the cells were cultured in the presence of [3H]leucine for the last hour of each treatment period. The intra- and extracellular radioactive hormones were determined in the same microsample by a specific and quantitative immunoprecipitation method, employing polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate. The results show that all prostaglandins stimulated the extracellular accumulation of prolactin at 3-30 nM. Prostaglandin E1 was more effective in stimulating extracellular accumulation of prolactin than prostaglandin E2 and the F compounds. Prostaglandin F2alpha at 3 nM doubled the rate of prolactin synthesis, but had no effect on growth hormone or total cell protein synthesis after 24 h of treatment. The first effect of prostaglandin F2alpha was observed after 5 h of incubation, and the time-course of effect on prolactin synthesis was similar to that of thyrotropin-releasing hormone.