We have recently reported that, in the rat pituitary, prolactin (PRL) is synthesized and stored in two different cell types: the typical PRL cell characterized by the presence of large irregular secretory granules and another cell type containing small round secretory granules. Since the secretion of PRL is stimulated by estrogens, we decided to investigate the effect of gonadectomy and estradiol (E2) replacement therapy in female rats on both cell types using a combination of in situ hybridization and immunostaining. Gonadectomy induced a marked decrease in hybridization signal and an almost complete disappearance of the classical PRL cells. The most abundant PRL cells were the small-granule-containing cells and other PRL cells characterized by the presence of both small and large granules. These changes in the PRL cell population were completely prevented by E2 administration to castrated animals. These studies indicate that E2 cannot only regulate PRL mRNA but also induce important morphological changes in PRL cell types.