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The effects of diethylstilbestrol (DES) treatment and withdrawal on the folliculo-stellate (FS) cells in hyperplastic pituitaries were examined in Fischer 344 (F344) and Wistar Furth (W/F) rats by immunochemistry and electron microscopy. The presence of S-100 protein positive cells was examined by immunostaining in spontaneous and in transplantable rat pituitary tumors. The pituitaries of F344 rats showed a fivefold greater increase in weight in response to 5 weeks of DES treatment compared to pituitaries from W/F rats. S-100 protein (S-100) positive cells were significantly increased in both strains with hyperplastic pituitaries (P less than 0.05) 2 days after DES withdrawal. Ultrastructural studies revealed increased phagocytic activity in FS cells. S-100 positive cells were absent in both MtT/W15 and MtT/F4 transplantable tumors even after treatment with DES. Some spontaneous pituitary tumors in aged female rats had S-100 positive cells within the tumors at the periphery of the tumor nodules (3 of 12 cases), while most of these neoplasms did not contain S-100 positive cells. Incubation of normal dissociated pituitary cells with S-100 protein increased PRL secretion in vitro. The effectiveness of S-100 protein in increasing PRL secretion in vitro was less than thyrotropin releasing hormone (TRH). These results indicate that S-100 positive cells are present in normal and hyperplastic pituitaries, but not in spontaneous or in transplantable rat pituitary tumors and also suggest that the FS cells may exert a paracrine type regulation on PRL secretion.