The number of gonadotropin-releasing hormone (GnRH) receptors is known to vary throughout the estrous cycle and in other endocrine states in the rat. These changes in receptors parallel closely the concentrations of serum estradiol during the cycle. In the present study, we used two different cytochemical techniques to determine if changes in GnRH receptors represented alterations in the number of GnRH-receptive cells. Furthermore, we tested the effects of estradiol pretreatment on this phenomenon. Dispersed pituitary monolayers taken at different stages of the cycle were stimulated for 3 min with 1 nM biotinylated [D-lys6]-GnRH (bio-GnRH) which was localized using the avidin-biotin-peroxidase complex (ABC) technique and a black peroxidase substrate. Parallel groups were stained, while living, with an avidin-fluorescein conjugate. Some monolayers were pretreated with physiological concentrations of estradiol benzoate (1 nM – 1 pM) prior to bio-GnRH exposure and ABC stains. The resulting stains demonstrated that the percentage of bio-GnRH-receptive cells was 2–3 times greater at 10.00 h proestrus (20.2 ± 4%) when compared to the same time in estrus (8.7 ± 2%), diestrus I (5.2 ± 0.5%), diestrus II (7.6 ± 1%), and at 17.00 h diestrus II (11.4 ± 0.9%) and proestrus (7.4 ± 0.8%). These data correlated well with those obtained from living gonadotropes stained with avidin-fluorescein. Estradiol exerted a biphasic effect dependent upon the stage of the cycle at which the cells were taken.Estradiol (1 pM) treatment of monolayers taken at 10.00 h diestrus I and II increased the percentage of GnRH-receptive cells to that normally observed at 17.00 h diestrus II (11.01 ± 1.5 and 11.9 ± 1%, respectively). However, the same concentration of steroid reduced the number of GnRH-receptive cells taken at 10.00 h proestrus to levels normally observed at diestrus I (6.4 ± 0.7%). These results indicate that changes in GnRH receptors during the estrous cycle represent fluctuations in the percentage of GnRH-bound cells, and that circulating levels of estradiol can mediate this change in number. It is suggested that the recruitment of GnRH-receptive cells, in conjunction with increases in both total and multihormonal gonadotropes, prepares these cells to support the preovulatory gonado