The concentration of immunoreactive luteinizing hormone releasing factor (LH-RF) was measured in pituitary stalk blood collected in the afternoon of expected pro-oestrus from rats ovariectomized and given oil, oestradiol benzoate (OB) or progesterone on the morning of dioestrus. Compared with the values in intact rats, LH-RF concentrations were reduced in ovariectomized animals treated with oil or progesterone but not with OB. Administration of progesterone at 12.30 h of expected pro-oestrus reduced the LH-RF concentrations in OB-treated rats; 5α-pregnan-3β-ol-20-one was not as effective as progesterone. The apparent inhibitory effect of progesterone, which was dose-dependent, was not found in long-term adrenalectomized rats, suggesting that it may be due to the production of an inhibitory substance by the adrenal consequent upon steroid administration. These and other data show that oestradiol triggers the spontaneous LH surge by stimulating the release of LH-RF as well as by enhancing pituitary responsiveness. A different mechanism may underlie the reflex surge of LH in rats exposed to constant light for, in these animals, progesterone was found to stimulate LH-RF release.

The neurotransmitter dopamine (DA) is known to inhibit prolactin (PRL) secretion and the proliferation of lactotropes in the pituitary gland. Dopamine-2 (D2) receptor short (D2S) isoform is expressed in a reduced level while the D2 receptor long (D2L) isoform is expressed in an elevated level during estradiol (E₂)-induced PRL production and cell proliferation in lactotropes. To evaluate the role of these D2 receptor isoforms in E₂-regulated lactotropic cell function, we compared E₂ effects on the level of PRL, cell proliferation, and G proteins in enriched lactotropes and lactotrope-derived PR1 cells containing only D2S isoform (D2S cells), D2L isoform (D2L cells), or no D2 receptor (V cells). Additionally, we determined the effects of G protein blockade on the E₂-induced PRL production and cell proliferation in these cells. We here show that E₂ actions on G proteins, PRL production, and cell proliferation were maximally achieved in D2S cells, oppositely or marginally achieved in D2L cells, and absent in V cells. We also show that the DA and pertussis toxin modulations of E₂ actions on PRL, G proteins, and cell proliferation were maximally achieved in D2S cells compared with in D2L or V cells. Furthermore, we provide evidence for the existence of an inhibitory action of Gi3 on Gs that is under the control of the D2S receptor and is inhibited by E₂. These results suggest that the suppression of D2S-regulated Gi3 inhibition of Gs protein may be one of the mechanisms controlling E₂-activated PRL synthesis and cell proliferation in lactotropes.

The mechanism of the first (pubertal) preovulatory gonadotrophin surge was investigated in Wistar rats by measuring (1) LH releasing factor (LH-RF) in pituitary stalk plasma and in extracts of hypothalamic and preoptic tissue, (2) LH and FSH in peripheral plasma and extracts of anterior pituitary tissue and (3) the LH and FSH response to LH-RF. The mechanism of the first surge appeared broadly to resemble that in the adult. That is, the gonadotrophin surge which occurs on the afternoon of the day before vaginal opening was found to coincide with a surge of immunoreactive LH-RF in pituitary stalk plasma and a significant increase in pituitary responsiveness to LH-RF. The magnitude of the change in pituitary responsiveness was, however, less at puberty than in the adult while the peak of the LH-RF surge was higher at puberty. The surges of LH-RF in stalk plasma and gonadotrophin in peripheral plasma corresponded relatively precisely with a fall in the preoptic and hypothalamic content of LH-RF and in the pituitary content of LH and FSH, suggesting that, in contrast with the adult, the synthesis of LH-RF and gonadotrophin at puberty cannot keep up with their release. A significant increase in stalk plasma LH-RF concentration occurring concomitantly with a surge of LH could be induced on the afternoon of Day 32 of life by administering pregnant mare serum (PMS) on Day 30 (about 10 days before vaginal opening). This, together with other evidence, suggests that the timing of the first gonadotrophin surge depends upon the capacity of the ovary to secrete oestradiol in the form of a surge. The fact that no significant increase in the pituitary responsiveness to LH-RF occurred in PMS-treated rats could account for the fact that the height of the gonadotrophin surge in these animals is only a third that of the spontaneous surge. No significant change in peripheral plasma LH or stalk plasma LH-RF concentrations was found around the time of vaginal opening or cornification in female rats given 1·25 mg testosterone propionate on Day 4 (androgenized female rats). Studies on the effect of ovariectomy and/or adrenalectomy suggested that the ovary and adrenal are involved in the timing of vaginal opening in normal but not in androgenized female animals.

The concentration of LH releasing factor (LH-RF) was measured by radioimmunoassay in blood collected from the cut pituitary stalk of long-term ovariectomized rats anaesthetized with Althesin. Stalk plasma LH-RF concentrations were increased immediately after ovariectomy (carried out at oestrus) and low at 2 and 4 days after operation. The concentrations then began to increase to reach a level at 24–28 days which was significantly higher than the concentrations during the oestrous cycle except for the time of the ovulatory surge at pro-oestrus. This pattern was similar to that of the concentrations of LH in jugular venous plasma taken from the same animals before exposure of the pituitary stalk. Like peripheral plasma LH concentrations, the concentrations of LH-RF in stalk plasma fluctuated and fell significantly and rapidly after the intravenous injection of 1 μg oestradiol-17β. The release of LH-RF in long-term ovariectomized rats, into which had been implanted an oestradiol-containing Silastic capsule, was similar to the diurnal pattern of LH release; the afternoon increase in stalk plasma LH-RF concentration could be blocked by sodium pentobarbitone administered at 13.00 h and augmented by administering this anaesthetic at 13.00 h of the preceding day. The stalk plasma LH-RF concentrations in animals injec[unk]d with oestradiol benzoate (OB) followed 72 h later with either OB or progesterone were lower than the concentrations in animals injected only with oil.

These data show that in the rat (1) ovarian steroids could moderate LH release ('negative feedback') by inhibiting LH-RF release, and that in long-term ovariectomized animals (2) the oestradiol-induced circadian pattern of LH release is due to a circadian pattern of LH-RF release, and (3) the surge of LH produced by administering OB followed by either OB or progesterone is probably due mainly to a massive increase in the responsiveness of the anterior pituitary gland to LH-RF.

Abstract

Transforming growth factor β1 (TGFβ1) has recently been shown to be produced in the prolactin (PRL)-secreting lactotropes of the pituitary gland. TGFβ1 inhibits lactotropic secretion and proliferation, and the production of TGFβ1 in lactotropes is reduced during lactotropic growth following estrogen treatment in ovariectomized rats. In many estrogen-responsive tissues, TGFβ1 has been shown to exert its effect by binding to TGFβ1 type II receptors (TβR II) at the cell surface. In this study, we sought to ascertain whether TβR II is involved in TGFβ1 action on lactotropes by determining the changes of TβR II mRNA and protein levels and specific ¹²⁵I-TGFβ1 binding sites on the lactotropes during estrogen-induced proliferation of lactotropes in Fischer 344 rats. Double immunohistochemical procedures were employed to identify immunoreactive TβR II in PRL-reactive cells. The majority of TβR II-reactive cells in the anterior pituitary were observed to be lactotropes. Dual immunohistochemistry and in situ hybridization procedures also indicated that lactotropes were the major cell types containing TβR II mRNA hybrids. Both the levels of immunoreactive TβR II protein and in situ TβR II mRNA hybrids in the pituitary were significantly decreased in ovariectomized rats after 15 days of estrogen treatment. Determination of ¹²⁵I-TGFβ1 binding sites in lactotropes by double immunohistochemistry and receptor autoradiography also revealed specific binding sites of ¹²⁵I-TGFβ1 in lactotropes in the anterior pituitary. ¹²⁵I-TGFβ1 binding in the anterior pituitary was also reduced following estrogen treatment in ovariectomized rats. These data suggest that down-regulation of TβR II may be an important mechanism of estrogen action on lactotropic cell growth and PRL secretion, and further support the notion that TGFβ1 controls lactotropic function by autocrine/paracrine mechanisms.

Journal of Endocrinology (1996) 149, 19–27