Ultrastructural and quantitative immunohistochemical changes induced by nonsteroid antiandrogens on pituitary gonadotroph population of prepubertal male rats.

Specific blockade of the androgen receptor by the nonsteroid antiandrogens flutamide and Casodex has proven to be a useful tool for studying androgens in vivo. The aim of the present study was to investigate the effect of antiandrogen administration at the pituitary level by evaluating the ultrastructural changes in gonadotrophs, in correlation with the quantitative immunohistochemical findings, and by comparing these alterations with the effect of androgen deprivation by castration either with or without subsequent androgen replacement. Male Sprague-Dawley rats (23 days old) were grouped as follows: (1) controls, (2) flutamide-injected (10 mg/rat/day), (3) Casodex-injected (10 mg/rat/day), (4) castrated, and (5) castrated plus androgen-replaced (dihydrotestosterone propionate; 40 microg/rat/day). Groups were sacrificed after 10 days of maintenance under each condition. Pituitaries were processed for both light and electron microscopy. Serial sections (4 microm) were obtained at different levels and immunostained by means of the primary murine monoclonal antibodies anti-FSH and anti-LH and a peroxidase-mediated EnVision System (Dako). Volume density, cell density and mean cell area were measured with an image analysis system (Imaging Technology, Software Optimas 5.2). The mean cell area (p < 0.001) and the volume density (p < 0.05) increased significantly in the flutamide- and Casodex-treated groups as well as the castrated group of FSH and LH cells. On the other hand, androgen replacement in the castrated rats, however, reduced in both parameters related to control animals. The cell density of FSH-secreting cells was increased (p < 0.05) in the Casodex and flutamide treatment as well as castrated group. The cell density of LH-secreting cells was augmented (p < 0.05) in the Casodex-treated group, while there was no increase in such parameter with flutamide and castration. The ultrastructure of all groups showed two types of gonadotrophs. Type I cells contained large (300-500 nm) and small (150-200 nm) secretory granules, while type II cells were smaller, and exhibited only small granules (100-200 nm). Flutamide-treated, Casodex-treated and castrated groups presented a decreased number of secretory granules with some exocytotic profiles, well-developed rough endoplasmic reticulum and an expanded Golgi complex of both types of cells. The gonadotrophs from the castrated group exhibited numerous mitochondria with electron-dense ring-shaped laminar figures, while in the castrated plus androgen-replaced rats only a few mitochondria had similar changes to those observed in castrated animals, as a possible residual alteration. Finally, the gonadotrophs from flutamide-treated rats showed mitochondrial alterations with clear areas and isolated electron-dense laminar figures. In summary, we conclude that lack of androgen reaction through the effects of nonsteroid antiandrogens and castration on prepubertal rats produced a hypertrophia-hyperplasia of the FSH cells, and hypertrophia of LH-secreting cells, with marked alterations at the ultrastructural level suggestive of a hyperstimulation stage.