Evidence for direct action of estradiol on Growth Hormone-Releasing Factor (GRF) in rat hypothalamus: Localization of [3H]estradiol in GRF neurons

Abstract

Sex steroids have been shown to influence the secretion of GH. There appears to be no good evidence of the effect of estradiol on the anterior pituitary, while the central site of estradiol action on the regulation of GH secretion is not known. The present investigation was carried out to determine whether some of the GH-releasing factor (GRF) neurons and somatostatin (SRIF) neurons in the hypothalamus and GH cells in the pituitary contain estradiol receptors. Colocalization of [3H]estradiol and antibodies to GRF or SRIF in brain and antibodies to GH in pituitary was studied to show interrelationships between estrogen target cells and peptidergic cells. Eight female Sprague-Dawley rats were ovariectomized, each rat was treated with colchicine, and 24-48 h later the animals were given an iv injection of [2,4,6,7,16,17-3H]estradiol (SA, 166 Ci/mM) at a dose of 0.5 μg/100 g BW. One hour after the injection, the rats were perfused with 4% paraformaldehyde in 0.1 M phosphate buffer (pH 7.4). The hypothalami from the perfused rats and the pituitaries from unperfused rats were frozen in isopentane precooled in liquid nitrogen (-190 C) and processed for autoradiography. The brain autoradiograms were immunostained for GRF, SRIF, and tyrosine hydroxylase [TH; an enzyme for the synthesis of dopamine (DA)], and the pituitary autoradiograms were immunostained for GH by the avidin-biotin peroxidase method. The majority of GRF-containing neurons were found in the arcuate nucleus, with some scattered cells in the lateral region of the ventromedial nucleus and the basal lateral hypothalamus. In the central portion of the arcuate nucleus, 20-30% of GRF-containing neurons showed nuclear concentration of [3H]estradiol. In the anterior portion of the hypothalamus, 10-15% of immunoreactive GRF-containing neurons were labeled with [3H] estradiol. In the lateral basal hypothalamus and the lateral region to the ventromedial nucleus, a few GRF neurons showed nuclear concentration of radioactivity. In contrast, a few SRIF cells in hypothalamic periventricular nucleus showed nuclear labeling with [3H]estradiol. Dual immunostaining with GRF and TH antibodies revealed that the estradiol-labeled GRF neurons did not contain TH immunoreactivity. In addition, 80-90% of GH cells in the anterior pituitary showed nuclear concentration of [3H]estradiol. The present studies demonstrate for the first time that certain populations of GRF neurons are targets for estradiol and indicate that estradiol acts directly on certain hypothalamic GRF neurons. The results suggest that estradiol may have a role in the regulation of GH secretion by modulating GRF release and acting directly on the somatotrophs.