Cell Nuclear Estrogen Receptor Research Articles

Raloxifene as Treatment for Various Types of Brain Injuries and Neurodegenerative Diseases: A Good Start.

Recent studies have shown that the selective estrogen receptor modulator (SERM) raloxifene had pronounced protective effects against progressing brain damage after traumatic brain injury (TBI) in mice. These studies, indicating beneficial effects of raloxifene for brain health, prompted the study of the history and present state of knowledge of this topic. It appears that, apart from raloxifene, to date, four nonrelated compounds have shown comparable beneficial effects—fucoidan, pifithrin, SMM-189 (5-dihydroxy-phenyl]-phenyl-methanone), and translocator protein (TSPO) ligands. Raloxifene, however, is ahead of the field, as for more than two decades it has been used in medical practice for various chronic ailments in humans. Thus, apart from different types of animal and cell culture studies, it has also been assessed in various human clinical trials, including assaying its effects on mild cognitive impairments. Regarding cell types, raloxifene protects neurons from cell death, prevents glial activation, ameliorates myelin damage, and maintains health of endothelial cells. At whole central nervous system (CNS) levels, raloxifene ameliorated mild cognitive impairments, as seen in clinical trials, and showed beneficial effects in animal models of Parkinson’s disease. Moreover, with stroke and TBI in animal models, raloxifene showed curative effects. Furthermore, raloxifene showed healing effects regarding multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS) in cell culture. The adverse biological signals typical of these conditions relate to neuronal activity, neurotransmitters and their receptors, plasticity, inflammation, oxidative stress, nitric oxide, calcium homeostasis, cell death, behavioral impairments, etc. Raloxifene favorably modulates these signals toward cell health—on the one hand, by modulating gene expression of the relevant proteins, for example by way of its binding to the cell nuclear estrogen receptors ERα and ERβ (genomic effects) and, on the other hand (nongenomic effects) by modulation of mitochondrial activity, reduction of oxidative stress and programmed cell death, maintaining metabolic balance, degradation of Abeta, and modulation of intracellular cholesterol levels. More specifically regarding Alzheimer’s disease, raloxifene may not cure diagnosed Alzheimer’s disease. However, the onset of Alzheimer’s disease may be delayed or arrested by raloxifene’s capability to attenuate mild cognitive impairment. Mild cognitive impairment is a condition that may precede diagnosis of Alzheimer’s disease. In this review, relatively new insights are addressed regarding the notion that Alzheimer’s disease can be caused by bacterial (as well as viral) infections, together with the most recent findings that raloxifene can counteract infections of at least some bacterial and viral strains. Thus, here, an overview of potential treatments of neurodegenerative disease by raloxifene is presented, and attention is paid to subcellular molecular biological pathways that may be involved.

GPER1 Signaling Initiates Migration of Female V-SVZ-Derived Cells.

SummaryIn the rodent ventricular-subventricular zone (V-SVZ) neurons are generated throughout life. They migrate along the rostral migratory stream (RMS) into the olfactory bulb before their final differentiation into interneurons and integration into local circuits. Estrogen receptors (ERs) are steroid hormone receptors with important functions in neurogenesis and synaptic plasticity. In this study, we show that the ER GPER1 is expressed in subsets of cells within the V-SVZ of female animals and provide evidence for a potential local estrogen source from aromatase-positive astrocytes surrounding the RMS. Blocking of GPER1 in Matrigel cultures of female animals significantly impairs migration of V-SVZ-derived cells. This outgrowth is accompanied by regulation of phosphorylation of the actin-binding protein cofilin by GPER1 signaling including an involvement of the p21-Ras pathway. Our results point to a prominent role of GPER1 in the initiation of neuronal migration from the V-SVZ to the olfactory bulb.

Effects of ovariectomy, estrogen and soy isoflavones in rats submandibular glands

Objective: A decrease in granular convoluted tubule (GCT) cells and acini occurs in the submandibular glands of castrated female rats, while in rats submitted to hormone replacement and phytotherapy with soy isoflavones, this effect is reversed. This study aimed to elucidate the mechanisms through which these changes occur. Material and Methods: Rats (n=84) were ovariectomized and 21 were sham-operated. Ovariectomized rats were randomly subdivided and orally administered the following: 17 ?-estradiol (OVX-E; n=21), 15 mg/kg/day of soy isoflavone extract (OVX-I; n=21), 17 ?-estradiol + soy isoflavone extract (OVX-A; n=21); and water as placebo (OVX; n=21). The rats were euthanized three, five and eight weeks after ovariectomy. The submandibular salivary glands were submitted to histological processing with HE stain and immunohistochemistry was performed using the streptavidin-biotin-peroxidase complex. The cell area and the expression of proliferating cell nuclear antigen and estrogen receptor ? were evaluated. Results: The results were statistically analyzed by ANOVA and Tukey test. A decrease in the area of GCT cells in the OVX, was observed, in contrast with an increase in the OVX-E. PCNA in the acinar cells and estrogen receptors were elevated in the OVX-I group. Conclusion: Castration exerts an immediate reductive effect on the volume of GCT cells. Estrogens, soy isoflavones and their combination have different mechanisms of action on the homeostasis of the gland. Estrogens cause an increase in GCT cells area, while isoflavones enhance cell proliferation and the expression of estrogen receptor-?. Their association showed no additional increase in the effect studied.KeywordsEstrogen; Morphometry; Salivary glands; Soy isoflavones.

A morphological study of uterine alterations in mice due to exposure to cadmium

We investigated the morphologic and molecular effects of exposure to cadmium (Cd) for 30 and 60 days on the uteri of mice. We assessed uterine morphometric measurements, eosinophilia, mast cell numbers, endometrial apoptosis, proliferation and estrogen receptor alpha (ERα) immunoreactivity. We examined vaginal smears that reflected the hormonal alterations in the female reproductive tract. Because the female reproductive tract exhibits different morphology at each stage of the estrous cycle, we sacrificed all animals at estrus to make appropriate comparisons. Female BALB/c mice were exposed to 200 ppm Cd in their drinking water for either 30 or 60 days. Cd exposure caused significant decreases in endometrial thickness and number of glands in estrus phase uteri. The endometrial eosinophilia in the groups exposed to Cd also decreased compared to controls. Cd exposure increased the number of mast cells. Luminal and glandular epithelia were examined using the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and by immunostaining proliferating cell nuclear antigen (PCNA) and estrogen receptor α (ERα). Compared to controls, the apoptotic index increased with time in both Cd exposed groups, while the proliferation index decreased. ERα immunoreactivity was decreased in both Cd exposed groups compared to controls; the decrease was most apparent in the 30 day Cd group. We found that 60 day Cd exposure increased apoptosis in the endometrium, which may affect the receptivity of the uterus for implantation.

Orbital Cavernous Haemangiomas: Immunohistochemical Study of Proliferative Capacity, Vascular Differentiation and Hormonal Receptor Status

Background/Aims: Immunohistochemical characterisation of orbital cavernous haemangiomas (CHs) with respect to proliferative capacity, hormone receptor status and vascular differentiation.Methods: Eleven cases of orbital CHs were reviewed. Immunohistochemical stains for Mib-1, proliferating cell nuclear antigen (PCNA), Bcl-2, estrogen and progesterone receptors (ER & PR), CD31, D2-40, and VEGF were investigated in 11 specimens.Results: Immunohistochemical staining revealed positivity for PCNA in ten of the 11 cases (91%). Bcl-2 was positive in 8 cases (73%). VEGF and PR were each weakly positive in 3 cases. All cases were negative for Mib-1, ER and D2-40. The staining was localized around the endothelium.Conclusion: This is the first study to characterise in detail the immunohistochemical features of orbital CHs. The proliferative markers PCNA and Mib-1 show discordant expression in these lesions and the expression of PCNA and Bcl-2 in the absence of Mib-1 is indicative of low proliferative potential. Small subsets of these tumors express PR and VEGF, which may partly explain the proliferative capacity of some orbital CHs.

Dietary tocopherols inhibit cell proliferation, regulate expression of ERα, PPARγ, and Nrf2, and decrease serum inflammatory markers during the development of mammary hyperplasia

Previous clinical and epidemiological studies of vitamin E have used primarily α-tocopherol for the prevention of cancer. However, γ-tocopherol has demonstrated greater anti-inflammatory and anti-tumor activity than α-tocopherol in several animal models of cancer. This study assessed the potential chemopreventive activities of a tocopherol mixture containing 58% γ-tocopherol (γ-TmT) in an established rodent model of mammary carcinogenesis. Female ACI rats were utilized due to their sensitivity to 17β-estradiol (E2 ) to induce mammary hyperplasia and neoplasia. The rats were implanted subcutaneously with sustained release E2 pellets and given dietary 0.3% or 0.5% γ-TmT for 2 or 10 wk. Serum E2 levels were significantly reduced by the treatment with 0.5% γ-TmT. Serum levels of inflammatory markers, prostaglandin E2 and 8-isoprostane, were suppressed by γ-TmT treatment. Histology of mammary glands showed evidence of epithelial hyperplasia in E2 -treated rats. Immunohistochemical analysis of the mammary glands revealed a decrease in proliferating cell nuclear antigen (PCNA), cyclooxygenase-2 (COX-2), and estrogen receptor α (ERα), while there was an increase in cleaved-caspase 3, peroxisome proliferator-activated receptor γ (PPARγ), and nuclear factor (erythroid-derived 2)-like 2 (Nrf2) in γ-TmT-treated rats. In addition, treatment with γ-TmT resulted in a decrease in the expression of ERα mRNA, whereas mRNA levels of ERβ and PPARγ were increased. In conclusion, γ-TmT was shown to suppress inflammatory markers, inhibit E2 -induced cell proliferation, and upregulate PPARγ and Nrf2 expression in mammary hyperplasia, suggesting that γ-TmT may be a promising agent for human breast cancer prevention.

The expression of PCNA, c-erbB-2, p53, ER and PR as well as atypical hyperplasia in tissues nearby the breast cancer

To examine the proper margin for breast conservative surgery in Chinese women population. 40 breast cancer specimens were collected and each sample was dissected into several groups: primary tumor group, 1cm paracarcinoma, 2cm paracarcinoma, 3cm paracarcinoma and excessive 3cm paracarcinoma groups. The immunohistochemistry staining was performed to measure the expression levels of proliferating cell nuclear antigen (PCNA), c-erbB-2, p53, estrogen receptor (ER) and progesterone receptor (PR). The gene expressions of PCNA, c-erbB-2, and p53 gradually decreased with the increased distance from primary tumor (P<0.05). The 1 and 2cm paracarcinoma group (no differences between the two, P>0.05) showed higher risk factors (c-erbB-2, p53) than the 3cm and excessive 3cm paracarcinoma groups (P<0.05). The expression of PCNA, ER, and PR showed no correlation with cancer progression (P>0.05). Beyond the paracarcinoma 2cm distance, the tissues showed significant decreases in tumor gene expression, which could represent the appropriate region for breast conservative surgery.

Effects of the levonorgestrel-releasing intrauterine system on cell proliferation, Fas expression and steroid receptors in endometriosis lesions and normal endometrium

The objectives of this study were: (i) to evaluate the effects of the levonorgestrel-releasing intrauterine system (LNG-IUS) on both proliferation and apoptosis markers and hormone receptors of the eutopic and ectopic endometrium of women experiencing pain related to endometriosis and (ii) to compare the results with those obtained with GnRH agonist (GnRHa) injections. Pre- and post-treatment endometrium and endometriosis specimens were obtained from 22 women experiencing pain related to endometriosis who were treated with LNG-IUS (n = 11) or GnRHa (n = 11) for 6 months. Changes in the expression of proliferating cell nuclear antigen, Fas, progesterone receptor (PRA) and estrogen receptor alpha (ER-alpha) were analyzed by immunohistochemistry. The cell proliferation index was significantly reduced in the epithelium and stroma of both the eutopic and the ectopic endometrium after treatment with the LNG-IUS and GnRHa. Only LNG-IUS users showed an increased H-score for Fas in the epithelium of the eutopic and ectopic endometrium (P < 0.05). Expression of ER-alpha and PRA by the glandular epithelium was lower in the eutopic endometrium after both treatments, but this reduction was noted in the ectopic endometrium only after LNG-IUS treatments (P < 0.05). No difference was detected between groups for any of the markers. LNG-IUS reduced both cell proliferation and the expression of PRA and ER-alpha and increased Fas expression in the eutopic and ectopic endometrium of patients with endometriosis. Some of these actions were not observed with GnRHa.

Progesterone and Progestin Receptors in the Brain: The Neglected Ones

Progesterone and Progestin Receptors in the Brain: The Neglected Ones

Expression of Tumor Antigens and Heat-Shock Protein 70 in Breast Cancer Cells After High-Intensity Focused Ultrasound Ablation

Previous results have shown that high-intensity focused ultrasound (HIFU) ablation can potentially activate a host antitumor immunity. The goal of this study was to investigate whether the tumor antigens expressed on breast cancer cells may be preserved after HIFU treatment, and to explore the potential mechanisms regarding the enhanced antitumor response. The primary lesion in 23 patients with biopsy-proven breast cancer were treated with HIFU, then submitted to modified radical mastectomy. By using biotin-streptavidin-peroxidase immunohistochemical technology, a variety of cellular molecules expressed on breast cancer cells, including tumor antigens and heat-shock protein 70 (HSP-70), were stained in all breast specimens. A complete absence of staining was recorded as negative, and immunoreaction of the tumor cells was considered to be positive for antigen expression. Nuclear positivity of breast cancer cells for proliferating cell nuclear antigen, estrogen receptor, and progesterone receptor was detected in 0%, 9%, and 9% of the treated samples, respectively. The positive rate of cytoplasmic staining for matrix metalloproteinase 9, carbohydrate antigen 15-3, vascular endothelial growth factor, transforming growth factors beta1 and beta2, interleukin 6, and interleukin 10 was 0%, 52%, 30%, 57%, 70%, 48%, and 61% in the treated cancer cells, respectively. The positive rate of cellular membrane staining for epithelial membrane antigen, CD44v6, and HSP-70 was 100%, 0%, and 100% in the zones of treated cancer cells, respectively. After HIFU ablation, some tumor antigens remained in the tumor debris. This could provide a potential antigen source to stimulate antitumor immune response.

17beta-estradiol enhances the production of granulocyte-macrophage colony-stimulating factor in human keratinocytes.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is effective for impaired wound repair. Estrogen is known to enhance wound repair. We examined if 17beta-estradiol (E2) may in vitro enhance GM-CSF production in human keratinocytes. E2 and membrane-impermeable bovine serum albumin-conjugated E2 increased GM-CSF secretion, mRNA stability, and promoter activity. The element homologous to activator protein-1 (AP-1) on the promoter was responsible for the activation. E2 enhanced transcriptional activity and DNA binding of AP-1. E2 transiently generated c-Fos protein, and shifted AP-1 composition from c-Jun homodimers to c-Fos/c-Jun heterodimers in keratinocytes. E2-induced enhancement of GM-CSF secretion, mRNA stability, and promoter activity were not suppressed by estrogen receptor antagonist ICI 182,780, however, suppressed by conventional protein kinase C inhibitor Gö6976 and PD98059, an inhibitor of mitogen-activated protein kinase kinase (MEK). Gö6976 and PD98059 suppressed E2-induced c-Fos expression and enhancement of DNA-binding and transcriptional activity at AP-1. E2 induced membrane translocation of protein kinase Calpha, which was suppressed by phosphatidylinositol (PI)-specific phospholipase C (PLC) inhibitor U73122. E2 stimulated the phosphorylation of extracellular signal-regulated kinase (ERK), which was suppressed by PD98059, Gö6976, and U73122. E2 transiently generated inositol 1,4,5-triphosphate in keratinocytes, which was suppressed by U73122 and guanine nucleotide-binding protein inhibitor. These results suggest that E2 may enhance GM-CSF production via guanine nucleotide-binding protein-coupled membrane receptors and signaling cascade of PI-specific PLC/protein kinase Calpha/MEK/ERK.

Smad 3 regulates proliferation of the mouse ovarian surface epithelium.

Smad 3 is a signaling intermediate for the transforming growth factor beta (TGFbeta) family; however, little is known about the role this protein plays in the regulation of the ovarian surface epithelium (OSE). Using a transgenic mouse model, we found that in the absence of Smad 3 there was a distinct morphological alteration of OSE cells. Wild-type (WT) OSE was flat with thin cells, while Smad 3-deficient (Smad 3 -/-) OSE was thick with plump cuboidal cells. WT OSE had less immunostaining for proliferating cell nuclear antigen (PCNA) and estrogen receptor alpha (ERalpha) than Smad 3 -/- OSE. However, there were no differences in the number of apoptotic cells or Bax and Bcl-2 levels between WT and Smad 3 -/- OSE. Although WT mice had higher levels of serum estradiol than Smad 3 -/- mice, WT and Smad 3 -/- mice had similar levels of progesterone. These data suggest that Smad 3 regulates OSE morphological appearance and proliferation in the absence of high serum estradiol levels or alterations in progesterone levels.

Signal Transduction Events Elicited by Natural Products that Function as Cancer Chemopreventive Agents

Cancer chemoprevention is a rapidly emerging paradigm for slowing, stopping or reversing the carcinogenic process. A multiplicity of biological mechanisms, functioning as complex interactions of gene products and regulatory pathways, are stimulated or inhibited by cancer chemopreventive agents. Several natural products and dietary components have been shown to function as chemopreventive agents. Screening chemical libraries of natural products for properties associated with cancer chemoprevention provides important insight into structural motifs driving rational design of more effective agents. In this review, the signal transduction pathways that are modulated by chemopreventive natural products are summarized. The activation of several signal transduction pathways triggered by polyphenolic antioxidants, flavonoids, retinoids, phytoestrogens and isothiocyanates are illustrated. These natural products may disrupt many signaling pathways, including transduction of cell surface (epidermal growth factor) or nuclear (estrogen) receptors via inhibition of their associated tyrosine kinase activities that regulate mitogenic signaling cascades (e.g., c-Raf-1). Alternatively, cytoprotective signal transduction pathways may be activated in a concentration- and time-dependent manner. The consequences of the modulation of signal transduction pathways by administration of these cancer chemopreventive compounds, singly or in combination, are the inhibition of cell cycle activating cascades and induction of cell survival / protection genes. Understanding the activation of signal transduction events elicited by various drugs and chemicals may yield insights into the regulation of the expression of genes coordinating cell proliferation, death, as well as, drug metabolizing and other cytoprotective enzymes. Monitoring the activation or inhibition of these pathways may serve as endpoints for rapid screening of effective cancer chemopreventive agents.

Cell Membrane and Nuclear Estrogen Receptors (ERs) Originate from a Single Transcript: Studies of ER and ER Expressed in Chinese Hamster Ovary Cells

Cell Membrane and Nuclear Estrogen Receptors (ERs) Originate from a Single Transcript: Studies of ER and ER Expressed in Chinese Hamster Ovary Cells

Cell membrane and nuclear estrogen receptors (ERs) originate from a single transcript: studies of ERalpha and ERbeta expressed in Chinese hamster ovary cells.

The existence of a putative membrane estrogen receptor (ER) has been supported by studies accomplished over the past 20 yr. However, the origin and functions of this receptor are not well defined. To study the membrane receptor, we transiently transfected cDNAs for ERalpha or ERbeta into Chinese hamster ovary (CHO) cells. Transfection of ERalpha resulted in a single transcript by Northern blot, specific binding of labeled 17beta-estradiol (E2), and expression of ER in both nuclear and membrane cell fractions. Competitive binding studies in both compartments revealed near identical dissociation constants (K(d)S) of 0.283 and 0.287 nM, respectively, but the membrane receptor number was only 3% as great as the nuclear receptor density. Transfection of ERbeta3 also yielded a single transcript and nuclear and membrane receptors with respective Kd values of 1.23 and 1.14 nM; the membrane receptor number was only 2% compared with expressed nuclear receptors. Estradiol binding to CHO-ERalpha or CHO-ERbeta activated Galphaq and G(alpha)s proteins in the membrane and rapidly stimulated corresponding inositol phosphate production and adenylate cyclase activity. Binding by 17-beta-E2 to either expressed receptor comparably enhanced the nuclear incorporation of thymidine, critically dependent upon the activation of the mitogen-activated protein kinase, ERK (extracellular regulated kinase). In contrast, c-Jun N-terminal kinase activity was stimulated by 17-beta-E2 in ERbeta-expressing CHO, but was inhibited in CHO-ERalpha cells. In summary, membrane and nuclear ER can be derived from a single transcript and have near-identical affinities for 17-beta-E2, but there are considerably more nuclear than membrane receptors. This is also the first report that cells can express a membrane ERbeta. Both membrane ERs activate G proteins, ERK, and cell proliferation, but there is novel differential regulation of c-Jun kinase activity by ERbeta and ERalpha.

Cellular proliferation, estrogen receptor, progesterone receptor, and bcl-2 expression in GnRH agonist-treated uterine leiomyomas

Gonadotropin-releasing hormone (GnRH) agonists are commonly used in the treatment of uterine leiomyomas, but little is known about their histological and cellular effects on these neoplasms. We examined a cellular proliferation index as determined by the nuclear antigen Ki-67 and proliferating cell nuclear antigen (PCNA) expression, estrogen receptor (ER), and progesterone receptor (PR) expression in 27 leiomyomas from patients treated with the GnRH agonist leuprolide acetate (LA) and compared them with 33 untreated controls. All leiomyomas were removed by myomectomies from premenopausal woman after 2 to 6 months of LA treatment or in the follicular phase of the menstrual cycle in the untreated controls. Histological features examined included cellularity, nuclear atypia, vascular changes (dilated, thickened, or thrombosed vessels), edema, calcification, hemorrhage, necrosis, hyalinization, and mitotic activity. Although no difference was found between GnRH-treated and non-treated groups with respect to most histological features examined, immunohistochemical studies showed a significant decrease in the cellular proliferation index, ER, and PR expression in the LA-treated cases compared with nontreated controls. The cellular proliferation index, ER, and PR expression decreased by 85%, 49%, and 36%, respectively, in the LA-treated group as compared with controls ( P < .001). A subset of cases from the LA-treated and nontreated groups were also analyzed with respect to bcl-2 (an inhibitor of apoptosis) expression, and no significant difference between the LA-treated and nontreated groups was observed with both groups showing a strong (>75% of cells) cytoplasmic staining pattern. Results of this study show that LA treatment of leiomyomas results in a decrease in number of cycling cells.

European Musculoskeletal Oncology Society Amsterdam, October 6-7, 1994

A retrospective EMSOS study to assess risk factors for local recurrence in desmoid tumors was initiated in 1991. Paticnts and methods. 1.51 patients treated at 1 1 EMSOS institutions were studied. The average age was 33 (1-80) years and 82 were women Tumor locations were: upper extremity 39, lower extremity 40, trunk 49, neck 5 , other sites 13, and multiple sites 5. The average tumor size was 7 (1340) cm. 145 patients were operated on. 2 patients received preand 6 patients postoperative radiotherapy. 5 patients received medical treatment ( 1 tamoxifen, 4 chemotherapy). Histopathology review, DNA-cytometry, proliferating cell nuclear antigen (PCNA) and estrogen receptor determination were performed. Re.sdt.s. The diagnosis was confirmed in all cases. All primary tumors were diploid and estrogen receptor negative. A Cox regression analysis showed a 4.0 relative risk for upper extremity location and a 2.5 relative risk for marginal excision, respectively, for occurrence of first local recurrence. Tumor depth, vascular pattern, cellularity and PCNA index were of no prognostic importance.

Up-regulation of the uterine estrogen receptor and its messenger ribonucleic acid during the mouse estrous cycle: the role of estradiol.

Uterine estrogen receptor (ER) and ER mRNA were measured in cycling and ovariectomized (OVX) estrogen-treated mice to probe the physiological regulation of the intracellular distribution and biosynthesis of ER. On proestrus, when plasma estradiol (E2) levels are highest, the cell nuclear ER concentration was 2.4-fold greater than on metestrus. This increase was primarily attributable to an increase in total cellular ER (cytosolic plus nuclear ER) and only secondarily to an activation of ER, as measured by its redistribution from the cytosolic (i.e. nuclear-extractable) to the nuclear (nonextractable) fraction. Total cellular ER concentration was 1.8-fold higher on proestrus than on metestrus, whereas the fraction of total ER in the nuclear compartment (i.e. the percentage activated) was only 1.3-fold higher. The concentration of cellular ER mRNA was 3-fold greater on proestrus than on the other days of the estrous cycle, suggesting that the increased concentration of ER on proestrus was a consequence of increased ER gene expression. In OVX mice, physiological and, to a lesser extent, supraphysiological levels of E2 increased cell nuclear ER. As in proestrous mice, the increased ER content contributed more than ER activation to the increased cell nuclear ER concentration. Physiological, but not supraphysiological, concentrations of E2 increased ER mRNA in OVX mice. Together, these results suggest that up-regulation by E2 of ER mRNA and ER accounts for most of the increased nuclear binding of ER on proestrus. E2-dependent activation and consequent DNA binding of ER presumably initiate this process, but quantitatively account for only a small fraction of the increased nuclear binding of ER.

Sex differences in estrogen receptor binding in the rat hypothalamus: effects of subsaturating pulses of estradiol

The effects of subsaturating pulses of estradiol on cell nuclear retention of estrogen receptors in brain regions of male and female rats was determined. In the first experiment, age-matched adrenalectomized/gonadectomized (ADX/GDX) rats were killed 1 h after i.v. injection of a subsaturating dose of free estradiol (1.0 μg/kg b. wt.) and cell nuclear estrogen receptor binding was measured in microdissected brain areas by an in vitro exchange assay. As we have previously reported with saturating doses of estradiol (3.6–36.0 μg/kg b. wt.), a greater concentration of receptors was measured in the ventromedial nucleus (VMN), periventricular preoptic area (PVPOA), and medial preoptic area (mPOA) of the female than in the same regions of the male brain. Binding levels did not differ between the sexes in the bed nucleus of the stria terminalis, arcuate nucleus, or the corticomedial amygdala. In the second experiment, GDX/ADX male and female rats received either a single or double pulse of 0.5 μg estradiol/kg b. wt. administered 6 h apart. Rats were killed 1 h after the second pulse. In animals treated with a single pulse of estradiol 1 h before sacrifice, higher concentrations of estrogen binding were measured in the female VMN and PVPOA than in the male. Except in the mPOA, the concentration of estrogen receptor binding measured was not different from that measured in animals that had received an additional, earlier pulse of estradiol. In this area, receptor concentrations were lower in male rats treated with two pulses than in males treated with one pulse. Thus, receptor dynamics in the mPOA of the male may differ from that of the female and from that of other brain regions. In the third experiment, cytosol estrogen receptor concentrations measured in untreated male and female GDX/ADX rats were compared. Consistent with our data on cell nuclear binding after exposure to estradiol, cytosol receptor concentrations in the female VMN, PVPOA and mPOA were greater than those in the male. These results extend our previous findings of regional sex differences in maximal cell nuclear estrogen binding capacity by demonstrating that these differences exist in the presence of subsaturating pulses of estradiol and that they reflect differences in cytosol estrogen receptors.

Development of progesterone-facilitated lordosis in female guinea pigs: relationship to neural estrogen and progestin receptors

Ovariectomized (OVX), 20-day-old female guinea pigs did not exhibit lordosis following treatment with estradiol benzoate (EB) and progesterone. Behavioral responsiveness to EB and progesterone developed abruptly; by 30 days of age, responses typical of adult females (>9weeks of age) were observed. In vitro assays of neural steroid receptors were performed to test the hypothesis that a deficiency in the concentration of hypothalamic and/or preoptic area estrogen and/or progestin receptors contributes to the lack of progesterone-facilitated lordosis in juvenile (20-day-old) females. Assay of cytosol progestin receptors in the mediobasal hypothalamus (MBH) and preoptic area (POA) of immature and adult females 40 h after injection of EB confirmed a previous report that the concentration of estradiol-induced cytosol progestin receptors in the MBH of juvenile females was slightly lower than that observed in adults. Furthermore, MBH cell nuclear accumulation of progestin receptors after progesterone injection was markedly lower (i.e. 42%) in immature, as compared to adult females. The concentrations of cytosol estrogen receptors in the MBH and POA did not differ between immature and adult OVX guinea pigs. Also, cell nuclear estrogen receptor accumulation in the MBH after estradiol injection was comparable in immature and adult females. These data suggest that a deficiency in cell nuclear progestin receptor accumulation in the MBH may contribute to the absence of progesterone-facilitated lordosis in estradiol-primed, immature female guinea pigs. This age-related difference in cell nuclear progestin receptor accumulation may be due, in part, to reduced concentrations of estradiol-induced cytosol progestin receptors. However, the deficient response to estradiol in immature females does not appear to be the result of reduced levels of cytosol estrogen receptors or to aberrant cell nuclear estrogen receptor accumulation in the MBH.