Differential expression of estradiol receptors alpha and beta by gonadotropes during the estrous cycle.

This study focused on expression of estradiol receptors (ER) during the estrous cycle. Labeling for ERalpha or beta antigens and luteinizing hormone (LH) or follicle-stimulating hormone (FSH) beta-subunits was done on freshly dispersed pituitary cells. The lowest expression of ERalpha and beta was seen in estrus (23% and 12%, respectively). Expression increased to 42-54% of pituitary cells by diestrus. In males, cells with ERalpha or beta were 37% or 20% of the population, respectively. ERalpha or beta and gonadotropin antigens were in 6-9% of pituitary cells from male rats. Early in the cycle (estrus and metestrus), less than 5% of pituitary cells expressed ERalpha or beta with gonadotropins. These values doubled to reach a peak of 10% during proestrus (just before ovulation). These data show that a rise in expression of both ERalpha and ERbeta is a part of preovulatory differentiation of pituitary gonadotropes.(J Histochem Cytochem 49:665-666, 2001)     

Estrogen and progestin receptors in the reproductive tract of male and female primates

With the aid of monoclonal antibodies specific to the estrogen and progestin receptors, we have examined the cellular localization of these proteins in the reproductive tract of male and female macaques. Two striking findings have resulted from our work with these new reagents. First, these receptors are detectable only in cell nuclei, regardless of hormonal treatment, and second, they are often detectable in stromal, but not epithelial cells when the epithelial cells undergo various estrogen or progestin-dependent events. The latter observation has led us to conclude that stromal cell-epithelial cell interactions may play previously unappreciated roles in the hormonal control of the primate reproductive tract. The lines of evidence that have drawn us to this conclusion will be reviewed.     

Alteration of glutathione reductase expression in the female reproductive organs during the estrous cycle.

The enzyme glutathione reductase (GR) recycles oxidized glutathione (GSSG) by converting it to the reduced form (GSH) in an NADPH-dependent manner. A specific antibody raised against recombinant rat GR was used to localize the protein in the female reproductive organs during the estrous cycle in the rat. In the ovary, the strongest reactivity to the antibody was observed in oocytes, followed by granulosa cells, corpus luteum, and interstitial cells. A strongly positive reaction was also observed mainly in the oviduct epithelia, uterine epithelia, and endometrial gland in the reproductive tract. Oviducts contained the highest GR activity. The GR activity of uterus during metestrus was about twice as high as that for other stages of the cycle. The levels of GR proteins in the tissues roughly matched the activities. The expression of the GR mRNA was highest during metestrus. Because GSH is known to increase gamete viability and the efficiency of fertility, GR, which is expressed in these tissues, is predicted to play a pivotal role in the reproduction process as a source of GSH.     

Estrogen and progesterone concentrations in bovine milk during the estrous cycle

Estrogen and progesterone concentrations in milk during the estrous cycle were estimated in 18 normally cycling Holstein dairy cows. The estrogen and progesterone concentrations in milk during the estrous cycle followed the pattern described for them in blood in the corresponding period. During most of the estrous cycle, estrogen concentration remained at approximately 200 pg/ml and reached a proestrous peak of 360 +/- 127 pg/ml on day 19. The progesterone concentration in milk during the estrous cycle increased to a peak on day 13 (45.5 +/- 6.6 ng/ml) and thereafter declined towards estrus. Estrus detection/prediction based on milk progesterone concentrations appears feasible in view of the significant differences in milk progesterone concentrations between the early luteal (post-ovulatory), luteal and rapid follicular growth periods of the estrous cycle.     

Prolactin receptors in the rat mammary gland. Change during the estrous cycle

Ovine prolactin (o-PRL) binding to mammary gland membranes was studied during the estrous cycle in the rat. Groups of rats were decapitated throughout the 4-day estrous cycle at 10 h00 on the days of diestrus I, diestrus II and estrus and at 10 h00, 12 h00, 16 h00 during the day of proestrus. Daily vaginal smears were taken to determine the stage of the estrous cycle which was also controlled by PRL and LH serum levels. Prolactin receptors were quantified in the 100 000 g pellet. For one Scatchard analysis, mammary gland membranes from 5 animals were pooled. Results given are the mean of 4 or 5 pools. Results obtained showed that the apparent affinity constant (KA) remained unchanged during the days of diestrus II and at all the times studied of proestrus and showed a slight but significant decrease on the days of estrus and diestrus I (or metestrus). The binding capacity did not vary from the day of diestrus II to the proestrus 16h00 (11.3 +/- 2.8 fmoles/mg protein) but sharply increased on the day of estrus (190.4 +/- 35.9 fmoles/mg protein). Binding capacity remained elevated on the day of diestrus I. This increase of PRL receptors on the day of estrous would appear to be an important step in preparing mammary gland for pregnancy and lactation.     

Photoaffinity labeling of pituitary gonadotropin releasing hormone receptors during the rat estrous cycle

A bioactive photoaffinity derivative of gonadotropin releasing hormone was used to identify pituitary gonadotropin releasing hormone receptors at various stages of the rat estrous cycle. Sodium dodecyl sulfate polyacrylamide gel electrophoresis resulted in the identification of a single specific component with an apparent molecular weight of 60,000 daltons throughout the estrous cycle. The amount of radioactivity incorporated into the 60K dalton band in diestrus and proestrus female rats increased 2.5-fold to that of metestrus and estrus female rats. These findings provide additional evidence for the identification of pituitary gonadotropin releasing hormone receptors.     

Myo-inositol in the reproductive tract of the female rat

• 1.1. myo-Inositol concentrations in oviduct, ovary and uterus were many-fold those of blood serum during all four stages of the estrous cycle of the female rat.
• 2.2. Inositol concentration was higher in oviduct than in ovary or uterus and was lower in uterine fluid than in uterus.
• 3.3. Estrus uteri had higher inositol concentrations than uteri in other phases of the cycle.
• 4.4. In order to measure dynamic aspects of the distribution of inositol. the distribution of radioactivity among organs of the reproductive tract of mature female rats was measured 45 min after i.p, injection of [2-³H]myo-inositol.
• 5.5. These organs concentrated inositol from the blood, and the tissue radioactivity (expressed as dpm/mg wet wt of tissue) increased in the sequence: vagina < cervix < uterus < ovary < oviduct.
• 6.6. The uterus and ovary concentrated myo-inositol more strongly during proestrus than during metestrus. diestrus or estrus.
• 7.7. The contents of proestrus follicles were more highly radioactive than was the ovary itself, whereas proestrus uterine fluid was less radioactive than the uterine tissue.

     

Lysosomal hydrolases in reproductive organs during estrous cycle of the hamster

Changes in the total protein content and the activities of lysosomal hydrolases (arylsulfatase, acid phosphatases, β-glucuronidase, β-N-acetylhexosaminidase, α-L-fucosidase, and β-galactosidase) of the hamster genital tract during the 4 days of estrous cycle and in hormonally superovulated hamsters were measured. Levels of lysosomal hydrolases in uteri and uterine fluid changed significantly during the cycle. Similar changes were observed in uterine wet weight and uterine proteins. The pattern of enzyme activities in both the ovary and the oviduct were different from those in uteri. In the ovary, most enzyme activities and the total protein concentration remained elevated after ovulation. Protein concentration and enzyme activities were significantly higher in the ovary, oviduct, and uteri of superovulated hamsters as compared to controls.     

Fluctuations of lactoferrin protein and messenger ribonucleic acid in the reproductive tract of the mouse during the estrous cycle.

The physiological role of lactoferrin (LF), the major estrogen-inducible protein in the murine uterus, is unclear; however, LF may be a useful marker for the study of estrogen action in the uterus. Thus, the expression of LF mRNA and the localization of the protein in genital tract tissues and secretions of female mice (6-8 wk old) at different stages of the estrous cycle were investigated. Uterine luminal fluid (ULF) was analyzed for LF by means of gel electrophoresis and Western blot techniques; LF mRNA and protein were identified in reproductive tract tissues through in situ hybridization and immunocytochemistry. At diestrus, the level of LF mRNA was low, and staining for the protein was very light in uterine epithelial cells; LF was undetectable in ULF. At proestrus, LF mRNA and protein increased in the uterine epithelium and LF was readily detectable in ULF. LF mRNA and protein reached the highest levels at estrus. At early metestrus as compared to estrus, LF mRNA and protein were detected in decreasing amounts in uterine epithelial cells; the protein was undetected in ULF. By late metestrus and diestrus, LF mRNA and protein returned to a low level, and the protein was undetectable in ULF. LF protein was also demonstrated by immunocytochemistry in the epithelium of the oviduct, cervix, and vagina. LF protein fluctuation similar to that observed in the uterus was seen in these tissues; however, the uterus demonstrated the most dramatic changes in the number of epithelial cells involved in LF production during the estrous cycle. In summary, LF mRNA and its expression in uterine epithelial cells of the mouse varied with the stage of the estrous cycle. These results, combined with previously reported findings that LF is a major constituent of mouse ULF under the influence of estrogen, suggest that LF may play an important role in normal reproductive processes.     

Temporospatial changes of kinin b2 receptors during the estrous cycle and pregnancy in the rat uterus

Tissue kallikreins are present in rat uterus during the estrous cycle in luminal and glandular epithelium, in early gestation in the implantation node, and in the last third of pregnancy surrounding the sinusoids in the decidua basalis. The pattern of kinin B2 receptor expression, through which the vasoactive effect of kallikreins is exerted, was studied by in vitro autoradiography and immunohistochemistry. The kinin B2 receptor was observed in the luminal and glandular epithelium, myometrium, endothelial cells of arteries, veins and venules, and smooth muscle cells of endometrial and myometrial arterioles. Immunoblotting of crude membranes revealed a band of 69 kDa that increased in late proestrus and estrus, concordantly with the pattern of immunostaining observed in the tissue. At Day 7 of gestation, the kinin B2 receptor was expressed (binding sites and receptor protein) in the epithelium of the implantation node and decidual cells; these latter cells showed a further increase during gestational Days 9 and 10. From Days 14 to 21, the subplacental decidua became strongly immunoreactive, and on Days 16 and 21 the placental labyrinthine endothelium was intensely stained. During this period, endothelium of arteries and veins, smooth muscular cells of small diameter arterioles, and myometrium also expressed B2 receptors. In unilaterally oil-stimulated pseudopregnancy, the decidual cells and the glandular epithelium show similar immunoreactivity to that during pregnancy. The temporospatial pattern of kinin B2 receptors, coinciding with that of kallikrein or with sites accessible to the generated kinins, further supports an autocrine-paracrine role for the kallikrein-kinin system in the vasoactive changes of implantation and placental blood flow regulation.     

Immunofluorescence study of the hypothalamo-infundibular LRH tract and serum gonadotropin levels in the female squirrel monkey during the estrous cycle

Summary The study of LRH reactive neurons of the medial basal hypothalamic group was made with rabbit antisera to unconjugated synthetic LRH, in normally cycling female squirrel monkeys.The specifically immunoreactive material present along the hypothalamo-infundibular LRH tract shows significant modifications that are particularly distinct in the lateral, posterior and anterior labia of median eminence, with a maximum concentration during the early and middle follicular phases, a sudden fall to a minimum concentration during the late follicular and ovulatory phases and with a progressive increase during the luteal and early follicular phases.Serum FSH and LH levels show a progressive decrease from the luteal phase to the late follicular phase, with differential modifications suggesting that a double regulation might take place in cycling squirrel monkeys.A parallel is suggested between the specifically reactive material along the hypothalamo-infundibular tract, serum gonadotropin levels and variations in the medial basal hypothalamic and infundibular LRH concentrations during the estrous cycle in the squirrel monkey.This work was supported by the D.G.R.S.T. (contrat 76-7-1536) and U.E.R. 3 (Faculty of Medicine). We acknowledge the help of A. Pillez (C.N.R.S.) for sectioning and staining the genital tracts     

Uterine epithelial and eosinophil estrogen receptors in rats during the estrous cycle

Summary In the uterus of the adult female rats, the luminal epithelial cells and the eosinophil leukocytes are rich in cytoplasmic estrogen receptors. During the estrous cycle, the epithelial estrogen receptor concentration reaches its peak level, in proestrus, drops precipitously in estrus, and hits the trough, at metestrus. Repopulation of the cytoplasm with estrogen binding sites occurs during diestrus. This pattern of cyclic change is indicative of a rapid turnover of estrogen receptors in the epithelial cells and its regulation by endogenous estrogens. The concentration of estrogen receptors in the cytoplasm of the eosinophils does not appear to fluctuate during the cycle. But the intrauterine, distribution of these leukocytes is clearly cyclic in pattern, ostensibly influenced by estrogens. While progesterone binding activity is consistently demonstrated in tandem with estrogen receptors in the cytoplasm of the epithelial cells, it has not been observed in the eosinophil leukocytes. These findings support the claim that there are two estrogen receptor systems in the rat uterus, one mediating the intracellular events of the genomic response to estrogens, and the other being concerned with non-genomic responses.Abbreviations BSA Bovine serum albumin - FITC fluorescein isothiocyanate - TMRITC tetramethylrhodamine isothiocyanate - CMO O-carboxymethyl oxime     

Influence of the estrous cycle on hypoxic failure in the female rat heart

Background: Clinical studies have shown that fluctuation in the plasma concentrations of estrogen during the menstrual cycle has an effect on myocardial health in premenopausal women. When estrogen levels are low, the number of ischemic events experienced is increased.Objective: To determine whether the increased ischemic events reported with low plasma estrogen concentrations in women could be reproduced in an animal model, cardiac function was measured during hypoxia in the female rat at different time points of the estrous cycle.Methods: Hearts from female Sprague-Dawley rats were perfused in the working mode at the diestrous (low estrogen; n = 7) and proestrous (high estrogen; n = 6) phases of the estrous cycle, confirmed by plasma estradiol concentrations. Hearts were perfused under aerobic conditions with 5.5 mM glucose, 100 μU/mL insulin, and 1.2 mM palmitate, followed by a 30-minute period of hypoxia with 95% N₂-5% CO₂ gassing.Results: There were no significant differences in heart function between diestrous and proestrous groups prior to hypoxia. However, hypoxia induced perturbations in function that were dependent on the estrous cycle. Reductions in left ventricular systolic and diastolic pressure occurred with hypoxia, but no significant differences in these pressures were observed between groups. Left ventricular pulse pressure and coronary flow also decreased significantly during hypoxia (both, P < 0.05), but hearts from the proestrous group maintained a significantly higher pulse pressure (P < 0.05). Hearts from the proestrous group also maintained significantly higher rates of coronary flow during hypoxia (P < 0.05), compared with hearts from the diestrous group. However, despite the effect of proestrus, correlation coefficients between plasma estradiol concentrations and indices of cardiac function were not significant.Conclusions: Our findings indicate that the estrous cycle of the female rat affects cardiac function during hypoxia. This model may be useful to study the impact of the estrous cycle on metabolic and cardiovascular function.     

Estrogen and LH dynamics during the follicular phase of the estrous cycle in the Asian elephant

Pituitary and corpus luteum hormone patterns throughout the elephant estrous cycle have been well characterized. By contrast, analysis of follicular maturation by measurement of circulating estrogens has been uninformative. This study tested the ability of a urinary estradiol‐3‐glucuronide radioimmunoassay to noninvasively assess follicular development during the nonluteal phase of the elephant estrous cycle, and to determine the relationship between estrogen production and the “double LH surge.” Daily urine and serum samples were collected throughout seven estrous cycles from three Asian elephants, and urine was collected from an additional three females, for a total of 13 cycles. Serum was analyzed for luteinizing hormone (LH), and urine was analyzed for estrogens and progestins. Elephants exhibited a typical LH pattern, with an anovulatory LH (anLH) surge occurring approximately 21 days before the ovulatory LH (ovLH) surge. The urinary estrogen pattern indicated the presence of two follicular waves during the nonluteal phase. The first wave (anovulatory) began 5 days before the anLH surge and reached a maximum concentration the day before the peak. Thereafter, urinary estrogens declined to baseline for 2 weeks before increasing again to peak concentrations on the day of the ovLH surge. Urinary progestins were baseline throughout most of the follicular phase, increasing 2–3 days before the ovLH surge and continuing into the luteal phase. These results support previous ultrasound observations that two waves of follicular growth occur during the nonluteal phase of the elephant estrous cycle. Each wave is associated with an increase in estrogen production that stimulates an LH surge. Thus, in contrast to serum analyses, urinary estrogen monitoring appears to be a reliable method for characterizing follicular activity in the elephant. Zoo Biol 22:443–454, 2003. © 2003 Wiley‐Liss, Inc.