The prolactin (PRL) gene is known to be expressed not only in the anterior pituitary but also in the decidualized human endometrium. This study was designed to detect the site of synthesis of PRL during pregnancy by in situ hybridization histochemistry. Decidual and trophoblast tissues from early pregnancy were obtained from patients undergoing therapeutic abortion at 8-10 weeks of gestation. Term placentae were obtained from patients with uncomplicated deliveries at 38-40 weeks. Sections of these tissues were hybridized with 35S-labeled RNA probe complementary to human PRL mRNA. Specific hybridization signals were distributed over the decidual cells in early and term pregnancy. In the decidua capsularis of early pregnancy, labeled cells were concentrated close to the amniotic cavity, although decidual cells were distributed evenly. In the decidua parietalis, almost all decidual cells were labeled, but no specific labeling was seen in the endometrial glands or capillary endothelium. In the decidua basalis, greater signals were always detected over the decidual cells in early pregnancy than in term pregnancy, when sections, which were hybridized with the same probe and exposed simultaneously, were compared. No specific hybridization was detected in the trophoblast cells. These results not only confirm that PRL is specifically synthesized in the decidual cells but also indicate that there are regional and periodical differences in PRL gene expression in the decidual cells during pregnancy.