Regulation of Expression of Epidermal Growth Factor Receptors in Gonadotropes by Epidermal Growth Factor and Estradiol : Studies in Cycling Female Rats *

Changes in expression of epidermal growth factor (EGF) receptors by gonadotropes parallel those of GnRH receptors. Gonadotropes increase their expression of EGF receptors (EGFR) during diestrus to reach a peak on the morning of proestrus. This is followed by a decline in expression to reach a nadir by estrus. We hypothesized that regulatory factors that stimulate changes in GnRH receptors might mediate the same changes in EGFR. To test this hypothesis, pituitary cells were collected from cycling rats and grown overnight in media with or without serum, 100 pM estradiol, or 60 ng/ml activin. On the next day, some of the cultures were further stimulated with 1 nM GnRH (4 h). The cells were then dual-labeled for EGFR and LHb or FSHb antigens and analyzed for their content of EGFR and gonadotropins. Neither activin nor estradiol increased percentages of cells with gonadotropin antigens and EGFR. Estradiol decreased percentages of cells with EGFR and LH in proestrous rats and those with EGFR and FSH in diestrous rats. The estradiol-mediated decline in EGFR expression during proestrus is similar to that seen when GnRH receptors are studied. Serum containing media alone increased percentages of LH and FSH cells with EGFR in populations from estrous or metestrous rats. Therefore, further experiments were conducted to learn if serum factors or EGF might be a regulator. Removal of serum from the growth media did not prevent the increase in percentages of LH cells with EGFR over the 18-h growth period. However, removal of serum did prevent the increased percentages of FSH cells with EGFR. Similarly, adding 1:100 anti-EGF to the serum containing media did not affect expression of EGFR by LH cells. However, it did cause a 27% decrease in percentages of FSH cells with EGFR. Finally, when 10 ng/ml EGF was added to metestrous populations in serumfree media there was a 1.41. 5-fold increase in percentages of LH or FSH cells with EGFR. Collectively, these studies show that EGF receptors are not stimulated in gonadotropes by the same hormones that up-regulate GnRH receptors. Furthermore, EGF itself may be among the factors that up regulate EGFR in gonadotropes. EGF receptors may be down-regulated by estradiol during proestrus, but the effect is limited to LH cells. Finally, EGF’s differential effects on LH and FSH cells suggests that it may selectively act on monohormonal gonadotropes. EGF receptors may be a marker for a unique subset of developing gonadotropes. (Endocrinology 138: 5434–5441, 1997) E growth factor (EGF) may be an important regulatory factor in the pituitary. It stimulates TSH (1) PRL (2), ACTH (3, 4) or GH (3) release. In addition, it appears to be involved with the regulation of the reproductive system. Miyake et al. (5) found that perifusion of pituitary tissue with EGF stimulates the release of LH. Their results showed that EGF regulates the secretion of gonadotropins both by stimulating the hypothalamus and by increasing pituitary responsiveness to estradiol. Radford et al. (6) demonstrated direct effects of EGF on LH release. EGF may regulate pituitary cells via paracrine or autocrine mechanisms. Previously, we demonstrated that EGF messenger RNA (mRNA) is produced by subsets of all of the pituitary cell types, including gonadotropes (7). Another group has shown that subsets of all pituitary cells will release EGF in reverse hemolytic plaque assays (8, 9). In a study of immature pituitary cells, Mouihate et al. (10) showed that most (72%) of the EGF-secreting cells were LH antigen-bearing gonadotropes and that the secretion of EGF could be enhanced by 10 nm GnRH. Collectively, these data suggest that EGF is a paracrine or autocrine mediator that may be regulated by key reproductive hormones. To further support the hypothesis for autocrine mechanisms, Mouihate and Lestage reported data that showed that most (73%) of the EGF secreting cells also express EGF receptors (11). Recent studies in this laboratory have characterized the distribution of GnRH or EGF receptors and changes in their expression during the estrous cycle (7, 12–15). Dual-labeling studies showed that subsets of all pituitary cell types expressed GnRH or EGF receptors (7, 12–16). Furthermore, the changes in EGF receptor expression by gonadotropes were similar to those reported previously for GnRH (13–16). There is an increase in gonadotropes that bind EGF (12) or GnRH (13–16) from metestrus to the morning of proestrus. The increase in GnRH receptive cells agreed with results from radioreceptor assays (reviewed in Refs. 13–16). After the proestrous peak, there is a decline in EGF or GnRH receptor expression by gonadotropes to reach a nadir by estrus