Development of estradiol-induced progestin receptor immunoreactivity in the hypothalamus of female guinea pigs

The inability of young female guinea pigs to display progesterone-facilitated lordosis has been attributed, in part, to a deficiency in the concentration of hypothalamic estradiol-induced progestin receptors, as measured by in vitro binding assays. An immunocytochemical technique was used to ascertain where, within the mediobasal hypothalamus, estradiol-induced progestin receptor levels are lower in immature than in adult females. Adult (greater than 7 weeks) and juvenile (3 weeks) ovariectomized females received 10 micrograms estradiol benzoate, a dose that primes adult, but not immature females to respond behaviorally to progesterone. Progestin receptor-immunoreactive (PR-IR) cells were counted in the arcuate nucleus (ARC) and ventrolateral hypothalamus (VLH), the two regions containing the densest populations of estradiol-induced progestin receptors in the mediobasal hypothalamus. There was no age difference in the number of PR-IR cells in the rostral or caudal VLH, but immunostaining was darker in the rostral VLH of juveniles as compared to adults. We found similar numbers of PR-IR cells in the rostral and mid-ARC, but 35% fewer immunostained cells in the caudal ARC of immature, as compared to adult females. Furthermore, staining intensity was weaker in the mid- and caudal ARC of the juvenile females. These data suggest that the ARC, not the VLH, is a site of fewer estradiol-induced progestin receptors in immature females.     

Progestin receptors in substance P-immunoreactive neurons in the hypothalamus of male guinea pigs after behaviorally effective estradiol pulse treatment.

Pulsatile administration of estradiol effectively primes orchidectomized (ORCH) male guinea pigs to display progesterone-facilitated lordosis. In contrast, a single injection of estradiol benzoate (EB) is not behaviorally effective. In ovariectomized female guinea pigs, estradiol pulses induce progestin receptors selectively in substance P neurons in the ventrolateral hypothalamus (VLH), a site at which estradiol primes females to respond behaviorally to progesterone. To test the hypothesis that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P neurons in the VLH in males, ORCH animals received a single injection of EB 40 h before, or two pulses of estradiol-17 beta, 39 and 11 h before perfusion. Colchicine was administered intracerebroventricularly prior to perfusion. The only difference found between the two estradiol treatment groups was a higher number of progestin receptor-immunoreactive (PR-IR) cells in the rostral VLH of estradiol pulse-treated males. There were no significant differences in the number of PR-IR cells in the mid- or caudal VLH, nor in the number of substance P-immunoreactive (SP-IR) neurons in the VLH/ventromedial hypothalamus (VMH) of animals receiving the two estradiol treatments. Furthermore, the percentage of PR-IR cells in the VLH also immunoreactive for SP did not differ between the estradiol pulse- (22%-25%) and the EB-injected animals (22%-32%). These data do not support the hypothesis that administration of behaviorally effective estradiol pulses, as compared to behaviorally ineffective EB injections, induce progestin receptors selectively in substance P neurons in the VLH of male guinea pigs.     

Estradiol pulses induce progestin receptors selectively in substance P-immunoreactive neurons in the ventrolateral hypothalamus of female guinea pigs

Low doses of estradiol, administered as pulses, are as effective as higher doses for priming ovariectomized (OVX) guinea pigs to display progesterone-facilitated lordosis. High doses of estradiol, administered by constant-release implants, induce progestin receptors in many substance P-immunoreactive (SP-IR) neurons in the ventrolateral hypothalamus (VLH), a site at which estradiol primes OVX guinea pigs to respond behaviorally to progesterone. To test the hypothesis that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P-containing neurons in the VLH, OVX females received estradiol implants 1 week prior to perfusion, or two pulses of estradiol- 17β, injected 39 and 11 h before perfusion. Colchicine was administered intracerebroventricularly prior to perfusion. No significant differences were observed in the total number of progestin receptor-immunoreactive (PR-IR) or substance P-immunoreactive cells in the VLH and VLH/ventromedial hypothalamus (VMH), respectively, of females receiving the two estradiol treatments. However, the percentage of PR-IR cells in the VLH also immunoreactive for SP was significantly higher in the estradiol pulse-treated (53%), than in the estradiol capsule-implanted animals (36%). These data suggest that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P-containing neurons in the VLH and are consistent with the hypothesis that substance P is involved in progesterone-facilitated lordosis in guinea pigs.     

Estradiol pulses induce progestin receptors selectively in substance P-immunoreactive neurons in the ventrolateral hypothalamus of female guinea pigs.

Low doses of estradiol, administered as pulses, are as effective as higher doses for priming ovariectomized (OVX) guinea pigs to display progesterone-facilitated lordosis. High doses of estradiol, administered by constant-release implants, induce progestin receptors in many substance P-immunoreactive (SP-IR) neurons in the ventrolateral hypothalamus (VLH), a site at which estradiol primes OVX guinea pigs to respond behaviorally to progesterone. To test the hypothesis that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P-containing neurons in the VLH, OVX females received estradiol implants 1 week prior to perfusion, or two pulses of estradiol-17 beta, injected 39 and 11 h before perfusion. Colchicine was administered intracerebroventricularly prior to perfusion. No significant differences were observed in the total number of progestin receptor-immunoreactive (PR-IR) or substance P-immunoreactive cells in the VLH and VLH/ventromedial hypothalamus (VMH), respectively, of females receiving the two estradiol treatments. However, the percentage of PR-IR cells in the VLH also immunoreactive for SP was significantly higher in the estradiol pulse-treated (53%), than in the estradiol capsule-implanted animals (36%). These data suggest that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P-containing neurons in the VLH and are consistent with the hypothesis that substance P is involved in progesterone-facilitated lordosis in guinea pigs.     

Immunocytochemical colocalization of progestin receptors and β-endorphin or enkephalin in the hypothalamus of female guinea pigs

Double-label immunocytochemistry was used to determine whether estradiol-induced progestin receptors and either β-endorphin or leucine-enkephalin are colocalized in female guinea pig brain. Ovariectomized, adult guinea pigs were implanted with capsules containing estradiol-17β to induce high levels of progestin receptors, and injected intracerebroventricularly with co chicine to improve visualization of the opiate peptides. Sections through the hypothalamus and preoptic area were processed for progestin receptor, followed by β-endorphin or leucine-enkephalin immunocytochemistry. As reported previously, high concentrations of progestin receptor-immunoreactive (PR-IR) cells were found in the preoptic area (medial and periventricular portions, medial preoptic nucleus) and hypothalamus (anterior hypothalamic and arcuate nuclei, ventrolateral area). Many β-endorphin-IR cells contained PR-IR in the arcuate nucleus and its surroundings (33%) and in the dorsomedial area of the hypothalamus (64%). Scattered enkephalin-IR cells were found in the septal nucleus, medial and lateral preoptic area, bed nucleus of the stria terminalis, and the arcuate nucleus. The ventromedial nucleus of the hypothalamus and dorsolateral magnocellular nucleus, respectively, contained moderate and heavy concentrations of enkephalin-IR cells. Although some of these areas also contained PR-IR, enkephalin-IR was colocalized consistently with PR-IR only in a small number of cells in the arcuate nucleus and ventromedial/ventrolateral area of the hypothalamus. These data, taken together with earlier observations that virtually all cells containing estradiol-induced PR-IR also contain estrogen receptor-IR, provide neuroanatomical evidence that hypothalamic actions of progesterone and estradiol may be mediated by β-endorphin and/or enkephalin.     

Immunocytochemical colocalization of progestin receptors and beta-endorphin or enkephalin in the hypothalamus of female guinea pigs

Double-label immunocytochemistry was used to determine whether estradiol-induced progestin receptors and either beta-endorphin or leucine-enkephalin are colocalized in female guinea pig brain. Ovariectomized, adult guinea pigs were implanted with capsules containing estradiol-17 beta to induce high levels of progestin receptors, and injected intracerebroventricularly with colchicine to improve visualization of the opiate peptides. Sections through the hypothalamus and preoptic area were processed for progestin receptor, followed by beta-endorphin or leucine-enkephalin immunocytochemistry. As reported previously, high concentrations of progestin receptor-immunoreactive (PR-IR) cells were found in the preoptic area (medial and periventricular portions, medial preoptic nucleus) and hypothalamus (anterior hypothalamic and arcuate nuclei, ventrolateral area). Many beta-endorphin-IR cells contained PR-IR in the arcuate nucleus and its surroundings (33%) and in the dorsomedial area of the hypothalamus (64%). Scattered enkephalin-IR cells were found in the septal nucleus, medial and lateral preoptic area, bed nucleus of the stria terminalis, and the arcuate nucleus. The ventromedial nucleus of the hypothalamus and dorsolateral magnocellular nucleus, respectively, contained moderate and heavy concentrations of enkephalin-IR cells. Although some of these areas also contained PR-IR, enkephalin-IR was colocalized consistently with PR-IR only in a small number of cells in the arcuate nucleus and ventromedial/ventrolateral area of the hypothalamus. These data, taken together with earlier observations that virtually all cells containing estradiol-induced PR-IR also contain estrogen receptor-IR, provide neuroanatomical evidence that hypothalamic actions of progesterone and estradiol may be mediated by beta-endorphin and/or enkephalin.     

Progestin receptors in substance P-immunoreactive neurons in the hypothalamus of male guinea pigs after behaviorally effective estradiol pulse treatment

Pulsatile administration of estradiol effectively primes orchidectomized (ORCH) male guinea pigs to display progesterone-facilitated lordosis. In contrast, a single injection of estradiol benzoate (EB) is not behaviorally effective. In ovariectomized female guinea pigs, estradiol pulses induce progestin receptors selectively in substance P neurons in the ventrolateral hypothalamus (VLH), a site at which estradiol primes females to respond behaviorally to progesterone. To test the hypothesis that behaviorally effective estradiol pulses induce progestin receptors selectively in substance P neurons in the VLH in males, ORCH animals received a single injection of EB 40 h before, or two pulses of estradiol-17β, 39 and 11 h before perfusion. Colchicine was administered intracerebroventricularly prior to perfusion. The only difference found between the two estradiol treatment groups was a higher number of progestin receptorimmunoreactive (PR-IR) cells in the rostral VLH of estradiol pulse-treated males. There were no significant differences in the number of PR-IR cells in the mid- or caudal VLH, nor in the number of substance P-immunoreactive (SP-IR) neurons in the VLH/ventromedial hypothalamus (VMH) of animals receiving the two estradiol treatments. Furthermore, the percentage of PR-IR cells in the VLH also immunoreactive for SP did not differ between the estradiol pulse- (22%–25%) and the EB-injected animals (22%–32%). These data do not support the hypothesis that administration of behaviorally effective estradiol pulses, as compared to behaviorally ineffective EB injections, induce progestin receptors selectively in substance P neurons in the VLH of male guinea pigs.     

Heterogeneous Regulation of Steroid Hormone Receptors in the Brain

SYNOPSIS. The ovarian steroid hormones, estradiol and progesterone,act in the guinea pig brain to regulate the expression of sexualbehavior. In studies of the cellular mechanisms of steroid hormoneaction, we have used an immunocytochemical technique to studythe regulation of these receptors in different neuroanatomicalregions. We have observed that progestin receptor-immunoreactivityin cells in certain neuroanatomical regions are more responsiveto particular steroid hormone treatments than are cells in otherregions. Similarly, we have observed selective regulation ofprogestin receptor-immunoreactivity in neurons identified onthe basis of their neuropeptide content. Finally, in the rostralpart of the ventrolateral hypothalamus, a site involved in hormonalregulation of female sexual behavior, estrogen receptor-immunoreactiveneurons that have dopamine-ß-hydroxylase varicositiesclosely-associated have higher levels of immunostaining forestrogen receptors than neurons without this relationship. Takentogether, these studies demonstrate the possibility of studyingthe microregulation of steroid hormone receptors in subsetsof neurons defined by neuroanatomical location, neuropeptide/neurotransmittercontent, afferent input and projection sites. The ability tostudy interactions among different systems at the cellular levelmay help us to understand more clearly the cellular processesinvolved in hormonal regulation of fundamental neuroendocrineprocesses, including the neuroendocrine regulation of sexualbehavior     

Evidence of sex hormone binding globulin binding sites in the medial preoptic area and hypothalamus.

We have demonstrated a high density of both radiolabeled progesterone and estradiol conjugated to bovine serum albumin binding sites in the medial preoptic area and hypothalamus. Infusions of sex hormone binding globulin into the medial preoptic area of rats increased their female sexual receptivity similarly to the effect of estradiol conjugated to bovine serum albumin, suggesting sex hormone binding globulin acts at binding sites for estradiol conjugated to bovine serum albumin. In this study sex hormone binding globulin was used to displace radiolabeled progesterone conjugated to bovine serum albumin from plasma membrane fractions from the medial preoptic area-anterior hypothalamus and medial basal hypothalamus of ovariectomized rats injected with either 5 microg estradiol benzoate or sesame oil vehicle. We found that sex hormone binding displaced radiolabeled progesterone conjugated to bovine serum albumin in both areas and that in vivo estradiol treatment greatly increased the relative displacement by sex hormone binding globulin in the medial preoptic area-anterior hypothalamus. We interpret these data as indicating the presence of sex hormone binding globulin receptors in brain plasma membranes and further suggest that endogenous steroid conditions may alter these receptors.     

Progesterone, but not progesterone-independent activation of progestin receptors by a mating stimulus, rapidly decreases progestin receptor immunoreactivity in female rat brain

Recent studies suggest that progestin receptors may be activated in vivo by neurotransmitters in the absence of ligand. More specifically, vaginal-cervical stimulation (VCS) can influence sexual behavior by activating progestin receptors in the absence of progesterone. Another way to test if progestin receptors are influenced by particular stimuli is to examine progestin receptor immunostaining. We report that progestin receptor immunoreactivity is decreased in the forebrain of estradiol-primed ovariectomized (OVX) rats within 1 h after a subcutaneous injection of progesterone, a time by which rapid down-regulation of progestin receptors does not seem to have occurred. In estradiol-primed OVX rats, VCS also decreased progestin receptor immunoreactivity within 1 h in the medial preoptic area, but not in any other area examined. To determine if the decrease in immunoreactivity by VCS was due to adrenal secretions or by ligand-independent activation of progestin receptors, we repeated the experiment in estradiol-primed OVX/adrenalectomized rats. Prior removal of the adrenal glands blocked the rapid decrease in progestin receptor immunoreactivity, even though data from other experiments suggest that progestin receptors are activated by VCS at this time. These studies suggest the possibility that progestin receptors may be affected differentially by progesterone-dependent or by progesterone-independent pathways. This raises the possibility that activation of progestin receptors by these two distinct pathways may lead to different neuronal consequences.     

Estrogenic effects of zearalenone on the expression of progestin receptors and sexual behavior in female rats

Zearalenone is a resorcylic acid lactone compound that is produced by fungal infection of edible grains and is believed to influence reproduction by binding to estrogen receptors. In order to study the potential estrogenic effects of this compound in the brain, we examined the effects of zearalenone on the expression of neuronal progestin receptors and feminine sexual behavior in female rats. Ovariectomized rats were treated with zearalenone (0.2, 1.0, or 2.0 mg), estradiol benzoate, or vehicle daily for 3 days. They were then either perfused, and progestin receptors visualized by immunocytochemistry, or injected with progesterone and tested for sexual receptivity with male rats. Progestin receptor-containing cells were counted in the medial preoptic area and ventromedial hypothalamus. The two highest doses of zearalenone increased the concentration of neuronal progestin receptors, as did 10 microg of estradiol. The highest dose of zearalenone (2 mg) also induced progestin receptor staining density comparable to that of 10 microg of estradiol benzoate. In behavioral tests, ovariectomized animals treated with 2 mg of zearalenone followed by progesterone showed levels of sexual receptivity comparable to females treated daily with estradiol benzoate (2 microg) followed by progesterone. These studies suggest that, although structurally distinct and less potent than estradiol, zearalenone can act as an estrogen agonist in the rat brain.     

Antagonism of estrogen-induced prolactin release by progesterone

Previous work from our laboratory has shown that during the process of nuclear occupancy of the progesterone receptor complex (1-2 h), nuclear estradiol receptors of the anterior pituitary are depleted. The purpose of this study was to determine whether the depletion of nuclear estradiol receptors by progesterone had functional biological significance. The ovariectomized (26 days of age) immature rat was used as the model for analysis of this question. The ability of estradiol to release prolactin from the anterior pituitary was the function chosen to determine the biological significance of the progesterone and estradiol interactions. In response to estradiol exposure (2 micrograms/rat), prolactin release reached peak values from 8 h to 12 h and returned to control levels by 24 h. A second injection of estradiol 13 h after the initial injection stimulated a second increase in serum prolactin at 25 h. This model of two injections of estradiol 13 h apart served to provide adequate levels of anterior pituitary progesterone receptors and elevated serum prolactin levels upon which superimposed progestin modulation could be examined. A single injection of progesterone (0.8 mg/kg BW) 1 h before the second estradiol injection blocked the increase in serum prolactin. This action was a receptor-mediated event because progesterone had no effect without estrogen priming or when the progesterone antagonist RU486 was used. Finally, when the interval between the progesterone and second estradiol injection was extended to 4 h, a time period when progesterone does not deplete pituitary nuclear estrogen receptors, the estrogen-induced increase in serum prolactin was not blocked.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cytoplasmic progesterone receptors in the hypothalamus-preoptic area of the mouse: effect of estrogen priming

A synthetic progestin, R5020, was used to identify cytoplasmic progestin receptors in the hypothalamuspreoptic area (HPOA) of ovariectomized mice. These high-affinity receptors exhibited an apparent dissociation constant of approx. 1 nM. The receptors were specific for progestins. [3H]R5020 binding was inhibited by more than 50% with a 50-fold excess of either radioinert R5020 or progesterone. 5 alpha-Dihydroprogesterone inhibited binding to a lesser extent. 3 alpha-Hydroxy-5 alpha-pregnane-20-one and cortisol did not compete for [3H]R5020 binding. Administration of estradiol benzoate (10 micrograms), 48 h prior to death, resulted in a 54% increase in the HPOA progestin receptor concentration when compared to oil-injected controls. These data demonstrate that there are specific and saturable cytoplasmic progestin receptors in the mouse HPOA and that the concentration of these receptors is increased after estrogen treatment.     

Long-term ovariectomy and hormone-induced sexual behavior, progestin receptors, and hypothalamic morphology in female rats

Long-term ovariectomy reduces the ability of estradiol and progesterone treatment to induce sexual receptivity in female rats. Previous researchers suggested that this effect may be due to a decreased induction of neural progestin receptors by estradiol in the long-term ovariectomized rats. The present study was designed to replicate and extend this finding, and to search for neuroanatomical correlates by measuring the volume of the ventromedial nucleus (VMN) of the hypothalamus, a putative site of action of estradiol and progesterone for the induction of female sexual behavior. Long-term ovariectomy (5 to 6 weeks) as compared to short-term ovariectomy (1 week) reduced the ability of estradiol-17 beta and progesterone treatment to induce sexually receptive and proceptive behaviors. Consistent with previous reports, our data show that the reduced levels of cytosol progestin receptors after long-term ovariectomy and estradiol treatment are related to a reduced ability of estradiol to induce the receptors. Long-term ovariectomy did not affect the concentration of cytosol progestin receptors in the preoptic area, suggesting a neuroanatomical specificity to this effect. Contrary to our predictions, long-term ovariectomy did not affect the volume of the VMN. In fact, estradiol treatment, while blocking the effect of long-term ovariectomy on sexual behavior, decreased the volume of the VMN. Therefore, the measurement of the volume of the VMN is not a good predictor of the responsiveness to steroid hormone induction of sexual behavior.     

Induction of Cytosolic Progestin Binding Sites by Catecholestrogens in Rat Pituitary Gland and Uterus: Different Potencies of 2- and 4-Hydroxyestradiol

The ability of catecholestrogens to induce cytosolic progestin binding sites in the hypothalamus, pituitary gland, and uterus of ovariectomised-adrenalectomised rats was demonstrated by the increase in high-affinity [3H]promegestone binding sites (KD 1.39, 0.50, and 0.54 nM, respectively) following a single subcutaneous injection (26.4 micrograms/animal) of the 3.4-dibenzoate ester of 4-hydroxyestradiol. The affinity and the time course of induction of these binding sites were very similar to those after a single injection of an equivalent dose (20 micrograms/animal) of estradiol 3-benzoate, exhibiting maximal receptor levels after 44 h. Widely differing efficacies in the induction of progestin binding sites were observed between the dibenzoate esters of 2- and 4-hydroxyestradiol. 2-Hydroxyestradiol 2,3-dibenzoate was ineffective in the pituitary gland up to a dose of 132 micrograms/animal, whereas 4-hydroxyestradiol dibenzoate was equipotent to estradiol benzoate, showing a maximal induction of progestin binding sites at single doses in the range of 13.2-26.4 micrograms/animal (equivalent to 10-20 micrograms of estradiol benzoate). As compared to the pituitary gland, the uterus was much more sensitive to the systemic administration of estrogen benzoates. At single doses in the range of 1.32-6.6 micrograms/animal (equivalent to 1-5 micrograms of estradiol benzoate), 4-hydroxyestradiol dibenzoate induced maximal levels of progestin receptors, and even 2-hydroxyestradiol dibenzoate, when given at a high dose (132.4 micrograms/animal, equivalent to 100 micrograms of estradiol benzoate), produced a slight increase in progestin binding sites.     

Neuroendocrine changes in female rats born from streptozotocin-diabetic mothers

In adult female rats born from Streptozotocin-diabetic mothers, blood glucose measured under basal conditions or 30 min after glucose administration was similar to controls; however at 180 min 50% of offspring from diabetics was moderately hyperglycemic whereas 100% of controls were normoglycemic. The time of vaginal opening, and after maturity, the number of rats with regular estrous cycles was in the range of controls. After ovariectomy, control rats receiving estradiol showed a sharp increase of serum LH at 4 pm following progesterone treatment at 10 am, while rats born from diabetic mothers failed to modify serum LH. Estradiol receptors in cell nuclei and cytosolic progestin receptors were determined in anterior pituitary, hypothalamus and preoptic area of rats subjected to a 4-day estradiol treatment. Changes were statistically significant in the hypothalamus only, in that rats born from diabetic mothers showed reduced induction of progestin receptors coupled to increased binding of (3H)-estradiol in cell nuclei. These findings bring support for a hypothalamic defect in rats born from diabetic mothers, the reduction of hypothalamic progestin receptors being reflected in the reduced sensitivity to the positive feedback action of progesterone to release LH.     

Steroid-monoamine feedback interactions in discrete brain regions using as a model the monosodium glutamate (MSG)-lesioned rat

The present studies examine the effects of neonatal treatment with monosodium glutamate (MSG) on dopamine (DA), 5-hydroxytryptamine (5-HT) and norepinephrine (NE) metabolism in discrete brain regions and correlate them with steroid receptor kinetics in the anterior pituitary (PIT), preoptic hypothalamus (POA) and caudal hypothalamus (HYP), and with steroid negative and positive feedback effects on luteinizing hormone (LH) secretion. Substantial decreases in the neuronal activity of all three amines in the arcuate nucleus, decreased DA and 5-HT metabolism in the suprachiasmatic nucleus and, surprisingly, increased metabolism of 5-HT and NE in the median eminence was observed in adult ovariectomized (OVX), MSG-treated versus OVX, vehicle-treated litter mate controls. Measurement of estradiol receptors in the nuclear and cytosolic fractions of the POA, HYP and PIT from MSG- and vehicle-treated rats killed during diestrus or 2 weeks after OVX revealed no differences. Similarly, no differences in cytosolic progestin receptors between control and MSG unprimed or estradiol-primed, OVX rats or on progestin receptor translocation induced by progesterone in Eb-primed rats were observed. Negative and positive feedback effects of estradiol or the positive feedback of progesterone on LH secretion were not significantly impaired in MSG rats, and indeed, MSG animals actually were hyper-responsive to the administration of the steroids or of luteinizing hormone-releasing hormone. These results indicate that the MSG-induced damage to DA, 5-HT and NE elements observed within several preoptic and hypothalamic nuclei does not impair estrogen and progestin receptor kinetics, nor does it prevent adequate negative or positive steroid feedback responses, if appropriate steroid regimens are employed, and that the impaired gonadal function reported in these animals does not result primarily from inadequate steroid feedback mechanisms.     

Intracellular progesterone receptors are differentially regulated by sex steroid hormones in the hypothalamus and the cerebral cortex of the rabbit

The aim of this study was to examine the role of sex steroid hormones in the regulation of intracellular progesterone receptors (PR) in the rabbit central nervous system. We determined PR concentration in cytosol preparations from the hypothalamus, the frontal, tempo-parietal and occipital cortex, by using the specific binding of the synthetic progestin [³H]ORG 2058. PR concentration was higher in the hypothalamus of intact adult females than in that of adult males and prepubertal females, whereas no significant differences were observed in the cerebral cortex of these animals. PR concentration was similar in the three cortical regions analyzed, indicating a homogeneous distribution of PR in the cerebral cortex. The administration of estradiol to ovariectomized animals increased PR concentration in the hypothalamus but not in the cortex. The administration of progesterone to ovariectomized rabbits did not modify PR concentration in any region, however when progesterone was administered after estradiol, it induced a significant diminution in hypothalamic PR concentration without effects in the cortex. These findings suggest that in the rabbit, PR are estrogen regulated in the hypothalamus but not in the cerebral cortex. In the latter, PR are not regulated by progesterone, whereas in the former the estrogen-induced PR are down-regulated by progesterone. Interestingly, hypothalamic PR constitutively expressed in ovariectomized animals are progesterone-insensitive.     

Regulation of estrogen-stimulated lordosis behavior and hypothalamic progestin receptor induction by antiestrogens in female rats

Two estrogen antagonists, CI-628 (CI) and tamoxifen (TX), were used to examine the relationship between estrogen priming of lordosis behavior and progestin receptor induction in the hypothalamus-preoptic area (HPOA) of ovariectomized female rats. Lordosis behavior was assessed by measuring lordosis quotients (LQ) in response to injection of 2 micrograms of estradiol benzoate (EB) followed 48 hr later by 500 micrograms of progesterone (P). Behavior testing began 4 hr after P injection. The effects of antiestrogens were assessed by injecting CI and TX (1-2 mg) from 0 to 48 hr prior to EB. Levels of cytosol progestin receptor in the HPOA were determined by quantifying the specific binding of 0.5 nM [3H]R5020 to cytosols from animals receiving the same EB and antiestrogen treatments used in behavioral testing. TX given concurrently with or CI given 2 hr before EB abolished both lordosis behavior and induction of HPOA progestin receptors. In contrast, CI given 12 hr prior to EB abolished lordosis but permitted a 95% elevation in the concentration of progestin binding sites in the HPOA. TX or CI given 48 hr before EB resulted in moderate levels of lordosis (mean LQs from 56 to 69) and induction of HPOA progestin receptors from 85 to 130% above noninjected controls. However, CI given 24 hr prior to EB produced less than a 40% increase in brain R5020 binding even though lordosis behavior was equivalent to that seen in the 48-hr animals (mean LQ = 53). These data indicate that the effects of antiestrogens on female sexual behavior and on the synthesis of brain progestin receptors depend on which antiestrogen is used and the time interval between administration of estrogen and antiestrogen. They also demonstrate that under some conditions estrogen induction of cytosol progestin receptors in the HPOA can be dissociated from estrogen priming of lordosis behavior in rats.     

Progesterone in high doses may overcome progesterone's desensitization effect on lordosis by translocation of hypothalamic progestin receptors

The cellular mechanism by which progesterone desensitizes the central nervous sytem to further facilitation of lordosis by progesterone was studied. In the first experiment guinea pigs were injected with doses of estradiol benzoate and progesterone that result in refractoriness to further stimulation of lordosis by moderate doses of progesterone. Confirming previous reports, a large dose of progesterone was effective in overcoming this desensitization, resulting in lordosis responses. A lower dose of progesterone was ineffective in this regard. In subsequent experiments, cytosol and nuclear progestin receptors were measured in the hypothalamus-preoptic area and cerebral cortex with an in vitro exchange assay using [³H]R 5020 as the ligand. A high, effective dose of progesterone resulted in 46–48% higher levels of nuclear progestin receptors assayed in the hypothalamus-preoptic area than did a lower, ineffective dose. These results are consistent with the idea that progesterone injection in estradiol-primed rodents causes a subsequent desensitization to itself by decreasing the concentration of cytoplasmic progestin receptors in hypothalamus-preoptic area. A subsequent progesterone injection then results in a lower level of nuclear progestin receptors. Large doses of progesterone may overcome this desensitization by causing accumulation of levels of nuclear progestin receptors sufficient to facilitate lordosis.