Inhibition of estrus and corpora lutea function with Norgestomet

Forty-five nonpregnant, nonlactating, Angus and Brangus cows were utilized to determine how long a Norgestomet ear implant would inhibit estrus when administered at various stages of an estrous cycle. All cows completed a nontreated estrous cycle to ensure normal cyclicity. At the second observed estrus (estrus = Day 1), cows were randomly allotted to be treated at metestrus (Day 3 or Day 4, n = 15); at diestrus (Day 9 or Day 10, n = 14); or at proestrus (Day 15 or Day 16, n = 16). All cows received a 2-ml intramuscular injection of 3 mg of Norgestomet accompanied by a 6-mg Norgestomet ear implant, which remained in situ for 21 days, or until individual cows were observed in estrus. Estrus was inhibited for a mean (+/- SEM) of 18.7 +/- 0.7, 19.9 +/- 0.8, and 17.0 +/- 0.8 days, respectively, when cows were treated at metestrus, diestrus, and proestrus (metestrus and diestrus vs proestrus; P < 0.05). Estrus was inhibited for an entire 21-day implantation period in 27, 50, and 38% of cows treated at metestrus, diestrus, and proestrus, respectively (P > 0.10). Norgestomet inhibited estrus in all cows for 11, 17, and 11 days after implantation when treatment was initiated at metestrus, diestrus, and proestrus, respectively (P > 0.10). These data indicate that a 6-mg Norgestomet ear implant effectively inhibits estrus in all cows for a maximum of 11 days, with some cows exhibiting estrus by Day 12 with the Norgestomet implant in situ.     

酪氨酸对大鼠子宫胞浆雌,孕激素受体的影响

采用放射受体分析法,测定了动情周期不同阶段及去卵巢大鼠子宫胞浆雌二醇和孕酮受体含量,并观察了子宫腔内注射酪、丝、苏三种氨基酸对子宫胞浆雌。醇、孕酮受体含量的影响。结果表明：（１）Ｌ－酪氨酸对动情前期、动情期、间情期大鼠子宫胞浆雌二醇和孕酮受体都具有明显的降低作用。（２）Ｌ－酪氨酸也降低去卵巢大鼠子宫胞浆雌二醇和孕酮含量,即这一作用不是通过影响卵巢激素分泌实现的。（３）Ｌ－苏氨酸仅可降低动情期和间情期大鼠子宫胞浆孕酮受体含量,而对相应周期雌二醇受体没有明显作用。（４）Ｌ－丝氨酸和Ｌ－苏氨酸对去卵巢大鼠子宫胞浆雌二醇和孕酮受体均无影响。     

Variations in concentration of oxytocin and vasopressin in the paraventricular nucleus of the hypothalamus during the estrous cycle in rats

Oxytocin (OT) and arginine-8-vasopressin (AVP) were measured by radioimmunoassay in micropunched hypothalamic neurosecretory nuclei of estrous cycling female Sprague-Dawley rats. In the paraventricular nucleus (PVN): the concentration (pg/microgram protein) of OT was significantly higher in rats in diestrus than during proestrus, estrus, or metestrus, while the concentration during metestrus was significantly greater than in proestrus and estrus; the concentration of AVP was significantly lower in animals in estrus than during the other three stages; because the paraventricular OT levels dropped before proestrus, the AVP/OT ratio was significantly greater in animals in proestrus than in diestrus, metestrus, and estrus. In the supraoptic nucleus (SON) a similar trend was noted: the concentration of OT was highest during diestrus, and AVP was lowest during estrus, though neither was significantly different from other stages. Because the OT and AVP cycles in the SON were asynchronous, the ratio of AVP to OT was significantly higher in proestrus than in metestrus or diestrus and significantly greater in estrus than during diestrus. In contrast to these two areas, peptide concentrations did not vary significantly across the estrous cycle in other sites of nonapeptide synthesis, i.e. the anterior commissural nucleus (ACN) and the suprachiasmatic nuclei (SCN).     

Regulation of uterine epidermal growth factor (EGF) receptors by estrogen in the mature rat and during the estrous cycle

Previous work has shown that the immature rat uterus contains epidermal growth factor (EGF) receptors and that tissue levels of this receptor are increased by the administration of exogenous estrogens. This study was undertaken to determine if estrogen administration also elevated EGF receptor levels in the mature animal and if the growth factor receptor levels varied in concert with endogenous estrogens throughout the estrous cycle. In the mature, castrate rat administration of estradiol, but not non-estrogenic steroids, causes a 2-3-fold elevation of uterine EGF receptors as judged by ligand binding. This increase is maximum in 18 h and is due to an increase in the number of binding sites. In cycling animals EGF receptor levels are low at metestrus, rise at diestrus, reach a maximum (approximately twice metestrus values) at proestrus, and then return at estrus to metestrus levels. These changes in EGF receptor levels parallel changes in plasma estrogens and occupied nuclear estrogen receptor reported by other workers. These results indicate that uterine EGF receptors are increased by exogenous estrogens in both mature and immature animals, and support a physiological role for estrogens in the regulation of this growth factor receptor.     

小熊猫繁殖周期血清雌二醇和孕酮含量变化

为研究小熊猫繁殖周期血清雌二醇、孕酮含量变化规律,采用化学发光免疫分析法连续16 次测定了2只成体雌性小熊猫血清雌二醇和孕酮含量变化,历经发情间期、发情期和两次妊娠期;连续9次测定了7只小熊猫妊娠期的孕酮含量变化。结果:(1)发情间期,小熊猫血清雌二醇的水平一直维持在低水平(基础水平),进入发情前期,血清雌二醇水平明显升高,在发情期一直维持高水平,配种后迅速降至基础水平; (2)小熊猫血清孕酮含量在发情间期和发情期均维持在较低水平,直至发情期过后才出现升高,在妊娠期一直维持高水平,峰值出现在5 月;(3)发情的小熊猫不论妊娠与否,在妊娠期内血清孕酮含量均维持在高水平。研究表明:小熊猫血清雌二醇、孕酮含量变化能直接反映其繁殖规律,雌二醇对启动雌性小熊猫季节性繁殖起重要作用;在妊娠期内小熊猫血清孕酮含量升高不能作为判断小熊猫妊娠的标准;雌性小熊猫在妊娠期有假孕现象。     

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.     

Pain threshold variations in female rats as a function of the estrus Cycle

In this study, the response of female rats in different phases of the estrus cycle to nociceptive stimulation was evaluated using thermal (hot plate and tail immersion) and chemical (formalin) tests. In the hot plate test, the paw licking latency fell significantly (p < 0.05) in the metestrus and diestrus phases compared with the proestrus and estrus phases. The observations in the tail immersion test also followed the same pattern. The significant reductions in the paw licking and tail withdrawal latencies due to a lowered threshold denote an increase in pain sensitivity in the metestrus and diestrus phases. In the formalin test, the licking time fell significantly from the metestrus to the diestrus phase compared with the proestrus and estrus phases, the reduction in this test which was due to an increased threshold connotes a decrease in pain sensitivity. The results therefore seem test dependent. In conclusion, pain threshold in female rats depends on the estrus state. Keywords: Pain threshold, Variation, Estrus cycle.     

Amygdala activation by corticosterone alters visceral and somatic pain in cycling female rats

Irritable bowel syndrome (IBS) is often seen in women, and symptom severity is known to vary over the menstrual cycle. In addition, activation of the hypothalamic-pituitary-adrenal (HPA) axis enhances symptomology and patients with IBS have increased activation of the amygdala, a brain region known to facilitate HPA output. However, little is known about the effects of amygdala activation during different stages of the menstrual cycle. We therefore investigated the effects of amygdala activation on somatic and visceral pain perception over the rat estrous cycle. Female Wistar rats were implanted with either corticosterone (Cort) or cholesterol as a control onto the dorsal margin of the central amygdala. Visceral sensitivity was quantified by recording the visceromotor response (VMR) to colorectal distension (CRD) and somatic sensitivity was assessed via the Von Frey test. In cholesterol controls, both visceral and somatic sensitivity varied over the estrous cycle. Rats in proestrus/estrus responded to CRD with an increased VMR compared with rats in metestrus/diestrus. Somatic sensitivity followed a similar pattern with enhanced sensitivity during proestrus/estrus compared with metestrus/diestrus. Elevated amygdala Cort induced visceral hypersensitivity during metestrus/diestrus but had no effect during proestrus/estrus. In contrast, elevated amygdala Cort increased somatic sensitivity during both metestrus/diestrus and proestrus/estrous. These results suggests that amygdala activation by Cort eliminates spontaneously occurring differences in visceral and somatic pain perception, which could explain the lowered pain thresholds and higher incidence of somatic pain observed in women with IBS.     

Quantitative variations in the number and size of peritoneal mastocytes in the female rat during the normal sexual cycle, following ovariectomy and administration of estrogens

Female rat peritoneal mast cells were found more numerous and of more reduced sizes in metestrus and diestrus than in proestrus and especially estrus where a partial degranulation took place. Ovariectomy caused an important decrease in the number and size of mast cells. A regular treatment of spayed animals by 17 beta estradiol partly inhibited these effects.     

Decreased luteinizing hormone-stimulated progesterone secretion by preovulatory follicles isolated from cyclic rats treated with the progesterone antagonist RU486.

Since administration of the antiprogesterone RU486 to cyclic female rats at metestrus and diestrus results in increased serum levels of LH, estradiol, and testosterone at proestrus, we investigated whether RU486 affects follicular steroidogenesis. Female rats with a 4-day estrous cycle, induced experimentally by a single injection of bromocriptine on the morning of estrus, were given RU486 (2 mg) twice daily (0900 and 1700 h) on metestrus and diestrus. At proestrus the preovulatory follicles were isolated and incubated for 4 h in the absence and presence of LH. In the absence of LH, accumulation of estradiol, testosterone, and progesterone in the medium was not different for RU486-treated rats and oil-treated controls. In contrast, LH-stimulated estradiol, testosterone, and progesterone secretions were significantly lower in RU486-treated rats compared with controls. Addition of pregnenolone to the incubation medium resulted in a significantly lower increase of progesterone in follicles from RU486-treated rats compared with those from oil-treated controls. This suggests that 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD) activity is decreased by administration of RU486 in vivo. Aromatase and 17 alpha-hydroxylase/C17-20 lyase activities were not affected: addition of substrate (androstenedione and progesterone respectively) did not affect differently the amount of product formed (estradiol and testosterone) in RU486- and oil-treated rats. However, LH-stimulated pregnenolone secretion was lower in follicles from RU486-treated rats compared with follicles from oil-treated controls, suggesting that either cholesterol side-chain cleavage activity or LH responsiveness is decreased. At proestrus the preovulatory follicles from RU486- and oil-treated rats were not morphologically different.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormone-sensitive lipase expression and IHC localization in the rat ovary, oviduct, and uterus

Hormone-sensitive lipase (HSL) is a key regulator of cholesterol esters metabolism. The aim of this study was to determine HSL localization in rat female reproductive organs during the ovarian cycle by IHC methods. HSL was located in the ovarian epithelium. The granulosa cells and oocytes of primordial follicles were immunonegative. In mature follicles, HSL was found in oocytes and theca and granulosa cells. However, HSL expression in theca cells and oocytes decreased during follicular atresia. Luteal cells showed HSL staining in cytoplasm during proestrus and estrus, in the nucleus during metestrus, and in cytoplasm and the nucleus during diestrus. In the tubaric ampulla, HSL was located in the epithelial cells nuclei and in the cilia during proestrus and estrus but mainly in the nucleus during metestrus and diestrus. In the isthmus, cells showed HSL immunolabeling in the nucleus and cilia during proestrus, but only in the cilia during estrus, metestrus, and diestrus. In the uterus, HSL was found in the epithelial cells nuclei. HSL-immunoreactive bands at 84, 67, 54, and 43 kDa were found in rat female reproductive organs. HSL labeling in the nucleus of epithelial and germ cells suggests an as yet unknown function for this protein, probably related to oogenesis and cell proliferation.     

Stabilization of estradiol-receptor complexes by elimination of cytosolic factors

Treating mature rat uterine cytosol with dextran coated charcoal (DCC) for 2 hrs at 0-4C in the absence of ligand causes the subsequently formed receptor-estradiol complex to be stable at 37C. Receptor binding is increased by the DCC treatment for uteri excised at metestrus or diestrus but remains nearly unchanged for uteri obtained at proestrus or estrus. Results suggest that the DCC removes or inactivates factor(s) present in the cytosol which render the receptor complex thermolabile.     

Compensatory ovulatory mechanisms operative after first ovulation in rats unilaterally ovariectomized prepubertally

Ovarian steroid contents and serum concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH) and prolactin were measured during the days after first ovulation in rats unilaterally ovariectomized in late prepuberty. In addition, follicle counts were made at second estrus and second metestrus. During the cycle following first ovulation, ovarian estradiol contents in unilaterally ovariectomized (ULO) rats were significantly increased as compared to intact rats on the day of metestrus, on diestrus 1 and on second estrus. Ovarian progesterone was significantly increased on the days of metestrus, on diestrus 1, second proestrus and second estrus, but no differences were seen in ovarian androgen contents. After ULO there was an indication of an augmented FSH surge at the first and the second ovulation. Follicle counts revealed that the total number of healthy as well as of atretic antral follicles on the day of second estrus was significantly increased after ULO, due to increased numbers of the smallest antral follicles. At second metestrus the number of larger antral follicles (350-500 micron 3) and the total number of healthy antral follicles was higher after ULO. It is concluded that the compensatory process after ULO involved increased recruitment of small antral follicles. Activities in the remaining ovary were not simply doubled but a new hormonal balance was established.     

Atrial supersensitivity to noradrenaline in stressed female rats

Stress can change the responses to catecholamines in many tissues. The aim of this study was to investigate the influence of the estrous cycle on the sensitivity of right atria to noradrenaline in female rats subjected to acute swimming stress. Female Wistar rats in proestrus, estrus, metestrus or diestrus were submitted to a 50 min-swimming session. Immediately after the exercise, the rats were killed and their right atria were mounted for isometric recording of the spontaneous beating rate. Concentration-effect curves to noradrenaline were obtained before and after the inhibition of neuronal uptake with phenoxybenzamine (10 microM) and of extraneuronal uptake with estradiol (5 microM). Acute swimming stress did not change the right atrial sensitivity to noradrenaline in rats in estrus, metestrus and diestrus. However, swimming stress produced supersensitivity to noradrenaline in proestrus (pD(2) control: 7.14 +/- 0.03 vs. pD(2) swimming: 7.55 +/- 0.04; p<0.05). This supersensitivity was still observed after uptake inhibition. When catecholamine uptake was inhibited, the concentration-effect curve to noradrenaline was shifted to the left 2.5-fold in the proestrus control group and 1.7-fold in the proestrus stress group (p<0.05). In conclusion, the estrous cycle influenced the acute stress-induced atrial supersensitivity to noradrenaline.     

Probable influence of ova and embryo prostaglandins in the differential ovum transport in pregnant and cycling rats.

In this study we explored the possible underlying mechanism(s) of the differential transport of unfertilized and fertilized ova in cycling and pregnant rats. The number of ova recovered from rat oviducts and uterus was not significantly different in estrus, metestrus and diestrus but dropped sharply at proestrus. When estrus rats were injected with indomethacin (10(-6)), a well known inhibitor of cyclooxygenase, delivered into both ovarian bursae, and sacrificed next day at metestrus, the number of ova in the oviduct was significantly smaller (p less than 0.025) than in controls at metestrus. On the other hand, when diestrus rats were injected with PGE1 (10(-6)) delivered into both ovarian bursae, and sacrificed next day at proestrus, no ova were found in the oviducts, and only a few of them were in the uterus. When fertilized ova were recovered from oviducts and uteri at day 4 of pregnancy (corresponding to proestrus of cycling rats) an average of 4 embryos were still found in the oviducts, proving a differential ovum transport between cycling and pregnant rats. In order to establish if there exists any ova or embryo releasing factor responsible for this difference, the prostaglandins released to the incubation medium by ovum or 3-day embryo were measured. Unfertilized ova produced significantly more PGE1 (p less than 0.05) than PGE2 or PGF2 alpha. The same pattern of PG production was observed with incubated embryos, but in this case the amount of PGE1 released was significantly higher (p less than 0.01) that the PGE1 released by unfertilized ova.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynamics of messenger RNAs encoding inhibin/activin subunits and follistatin in the ovary during the rat estrous cycle

Quantitative changes in ovarian inhibin/activin subunit and follistatin mRNAs during the rat estrous cycle were examined by ribonuclease protection assay using digoxygenin-labeled RNA probes. Levels of ovarian inhibin alpha subunit mRNA remained low throughout estrus, metestrus, and diestrus; abruptly increased on the morning of proestrus; then rapidly decreased when the primary gonadotropin surge occurred. A similar changing pattern was observed in inhibin/activin beta(A) subunit mRNA. On the other hand, inhibin/activin beta(B) subunit mRNA showed a different changing pattern. Levels of beta(B) subunit mRNA remained constant during metestrus and diestrus, abruptly decreased on the afternoon of proestrus, then quickly recovered from the nadir by 1100 h on estrus. Throughout the rat estrous cycle, especially during the periovulatory period, alpha subunit mRNA levels were considerably higher than beta(A) and beta(B) subunit mRNA levels. In addition, changes in plasma concentrations of inhibin A and inhibin B were very similar to that in ovarian beta(A) and beta(B) subunit mRNA levels, respectively, with several-hour delays. These results suggest that levels of beta subunit mRNAs restrict secretion of dimeric inhibins. Levels of follistatin mRNA remained low from the midnight of metestrus to the midnight of diestrus, then increased until initiation of the primary gonadotropin surge. Thereafter, follistatin mRNA decreased, reached the nadir at 0200 h on estrus, then increased abruptly at 1100 h on estrus. Afterward, follistatin mRNA levels remained high until the morning of metestrus. The changing pattern of ovarian follistatin mRNA was similar to, and preceded, the changes in plasma concentrations of progesterone, suggesting that ovarian follistatin may modulate progesterone secretion during the rat estrous cycle.     

Inhibin secretion during the rat estrous cycle: relationships to FSH secretion and FSH beta subunit mRNA concentrations

Serum inhibin and FSH and FSH beta subunit mRNA levels were measured at 3h intervals throughout the 4 day estrous cycle in female rats and hourly between 1000 and 2400 h of proestrus. On proestrus, serum inhibin concentrations fell during the late morning-early afternoon, then increased transiently during the late afternoon gonadotropin surges. Inhibin levels decreased during the late evening of proestrus, coincident with the FSH surge-related rise in FSH beta mRNA levels. Serum inhibin remained relatively stable during estrus and early metestrus, but rose during the late evening of metestrus and remained elevated until early diestrus. FSH beta mRNA levels were elevated on late estrus and early metestrus and declined during the evening of metestrus as serum inhibin levels increased. These data show that concentrations of serum inhibin change during the estrous cycle and that a general inverse relationship exists between serum inhibin and FSH levels and FSH beta mRNA concentrations in the pituitary. This suggests that inhibin may inhibit FSH beta gene expression and FSH secretion during the 4 day cycle in female rats.     

Resynchronization of ovulation and timed insemination in lactating dairy cows, II: assigning protocols according to stages of the estrous cycle, or presence of ovarian cysts or anestrus

Pregnancy rates were compared in lactating dairy cows (n = 1083) assigned to protocols for resynchronization of ovulation based on stages of the estrous cycle, or presence of ovarian cysts or anestrus. Cows were detected not pregnant by ultrasonography 30 d after a previous AI (study day 0) and classified as diestrus, metestrus, proestrus, with ovarian cysts or anestrus. Cows in diestrus (January-May) were assigned to either Ovsynch (GnRH day 0, PGF2alpha day 7, GnRH day 9, and timed-AI [TAI] 16 h later; n = 96), or Quicksynch (PGF2alpha day 0, estradiol cypionate [ECP] day 1, AI at detected estrus [AIDE] on day 2, or TAI on day 3; n = 96). Cows in diestrus (June-December) were assigned to either Ovsynch (n = 156) or Modified Quicksynch (PGF2alpha day 0, ECP day 1, AIDE days 2 and 3, and to Ovsynch on day 4 if not detected in estrus; n = 142). Cows in metestrus were assigned either to Ovsynch (n = 68), Heatsynch (GnRH day 0, PGF2alpha day 7, ECP day 8, AIDE day 9, or TAI day 10; n = 62), or GnRH + Ovsynch (GnRH on day 0, followed by Ovsynch on day 8; n = 64). Cows in proestrus, with ovarian cysts, or anestrus were assigned to either Ovsynch (proestrus n = 89, ovarian cysts n = 97, anestrus n = 8) or GnRH + Ovsynch (proestrus n = 87, ovarian cysts n = 109, anestrus n = 9). Pregnancy rate was evaluated 30, 55 and 90 d after resynchronized AI. For cows in diestrus (January-May), pregnancy rates were higher for Ovsynch (35.9, 29.2 and 26.0%) than for Quicksynch (21.7, 16.7 and 15.6%). For cows in diestrus (June-December), pregnancy rates were similar for Ovsynch (34.4, 24.0 and 23.6%) and Modified Quicksynch (27.1, 26.2 and 21.6%). For cows in metestrus, pregnancy rates were higher for GnRH + Ovsynch (33.3, 24.5 and 20.3%) than for Heatsynch (20.3, 12.9 and 9.8%). For cows with ovarian cysts, pregnancy rates were higher for GnRH + Ovsynch (30.3, 26.6 and 22.9%) than for Ovsynch (20.2, 18.5 and 14.7%). Assignment to resynchronization protocols based on the stages of the estrous cycle, or presence of ovarian cysts improved pregnancy rates.     

Effect of estradiol on the membrane fluidity of the rat vaginal epithelial cells

Changes in the cell surface of vaginal epithelial cells were studied by scanning microscopy and fluorescence spectroscopy. Microvilli which are prominent features of the vaginal epithelial cells in proestrus and diestrus are replaced by sheet-like structures in the estrus phase. Surface morphology of vaginal epithelial cells of estradiol primed rat resembles the vaginal cells from estrus phase rats whereas vaginal cells from control rats resembles the diestrus phase. Measurement of the fluidity of the membranes indicated that the vaginal epithelial cell membrane of estrus rats is more fluid compared to proestrus and diestrus. Similarly, estradiol primed immature rat vaginal epithelial cell membrane was observed to be more fluid than the corresponding control.     

Quantitative differences in immune responses during the various stages of the estrous cycle in female BALB/c mice.

The immune responsiveness of spleens from female BALB/c mice to PHA, Con A, and LPS was greater at proestrus and metestrus as compared with estrus and diestrus. The peaks of responsiveness corresponded to reported elevated levels of estrogen and pregnenolone during these phases of the cycle. Similar results were obtained with the IgM or direct plaque-forming cell responses, which were also increased at proestrus and metestrus. It appears that female hormones may directly or indirectly stimulate immune responsiveness in adult mice.