Modification of the ovarian cycle, ovulation and reproduction in rat by administration of p-chlorophenylalanine in the pre and post ovulation period (author's transl)]

The effect of p-Chlorophenylalanine (PCPA) administered in pre and post-ovulation phases on ovulation reproduction and ovarian cycle was studied in rats. Vaginal smears, number of ovules, mature follicles and ovaric histologic features were evaluated. PCPA administered in the periods after follicular rupture prolongs the diestrus phase from 2 to 10 consecutive days and totally inhibits the reproductive conduct. When administered in the periods just prior to follicular rupture, PCPA prolongs the estrus phase from 2 to 4 consecutive days and does not totally inhibit reproduction. A progressive increase in reproduction took place as treatment time approached ovulation, to such an extent that the number of inseminated and fertilized rats when treated at the ninth hour of the proestrus phase, was superior to the control group. PCPA administered in periods after and before follicular rupture totally inhibits ovulation in all cases. Ovaries present many mature follicles, as many as in the controls sacrificed at the 18 hour of the proestrus, just before ovulation.     

Effect of p-chlorophenylalanine on LH, FSH, prolactin and ovulation in the rat

The effect of p-chlorophenylalanine (PCPA: 300 mg/kg) on the rate of ovulation and plasma LH, FSH and prolactin secretion has been studied in rats at preovulatory periods (18th hour of diestrus) and post-ovulatory periods (9th hour of metaestrus). In both experimental groups, results showed that administration of PCPA caused an increase in both prolactin concentration and number of mature ovarian follicles (p less than 0.001). No changes were observed in FSH levels. LH concentration, however, decreased (p less than 0.001) and ovulation became totally inhibited. Rats treated at the 9th hour of metaestrus exhibited a marked luteinization as well as an increased number of corpus luteum in the ovaric tissue (p less than 0.001), whereas those treated at the 18th hour of diestrus underwent no luteinization and merely showed a greater number of mature ovarian follicles (p less than 0.001). PCPA, therefore, seems not to have a double effect on ovulation, LH, FSH, and prolactin secretion regardless of the pre or post-ovulatory periods. Changes observed in the ovaric tissue might be due to an increase in plasma prolactin concentration which appears earlier in the preovulatory than in the post-ovulatory treated animals. This difference may explain the double effect that has been attributed to the ovaric cycle and reproductive behavior.     

Action of estrogens on pituitary gonadotrophic function and follicular growth during diestrous in the rat (author's transl)

LH and FSH release during the afternoon of diestrus 1 on the one hand, and the rate of follicular growth on the morning of diestrus 1 or diestrus 2, on the other hand, were studied in 4-day cyclic female rats after injection of estradiol benzoate (10 microgram, s.c.) on the morning of estrus. LH and FSH release was observed between 15.00 and 19.00 h during diestrus 1, but did not occur after an injection of pentobarbital (30 mg/kg, i.p.) in diestrus 1 at 13.30 h. No luteinization resulted from an injection of estrogen. Slowed follicular growth was observed on the morning of either diestrus 1 or diestrus 2. These results suggest the existence of a "critical period" for LH and FSH release in diestrus 1 during the afternoon. They indicate that the ovarian response to the endogenous release of gonadotropins is dependent upon the state of development of the ovarian follicles.     

Behavioral, follicular and gonadotropin changes during the estrous cycle in donkeys

Sexual behavior, follicular development and ovulation, and concentrations of circulating gonadotropins during the estrous cycle were studied during the summer in 7 jennies. Mean behavioral estrous length was 6.4 +/- 0.6 days (mean +/- SEM, n=19; 5.6 +/- 0.5 days preovulatory and 0.8 +/- 0.2 days post-ovulatory). Mean diestrous length was 19.3 +/- 0.6 days (n=14). Females in estrus typically showed posturing, mouth clapping, clitoral winking, urinating and tail raising. Mouth clapping began approximately one day sooner and lasted approximately one day longer than winking and tail raising, so that the total duration of clapping was significantly greater than for the other two signs. Follicular changes and concentrations of gonadotropins were determined for 14 estrous cycles (2 per jenny). The follicular end points [diameter of the largest follicle and number of large (>25 mm), medium (20-24 mm), and small follicles (<20 mm)] showed a significant day effect. The diameter of the largest follicle and the number of large follicles began to increase significantly 7 days prior to ovulation with a maximum value the day before ovulation. Medium follicles reached a maximum number 4 days prior to ovulation, and small follicles decreased significantly prior to ovulation. After ovulation, all follicular end points, except the number of small follicles, remained low for the next 12 days. Mean values of FSH were low during estrus and high during diestrus with 2 significant peaks, one 3 days and one 9 days after ovulation. In contrast, mean levels of LH were low during diestrus and high during estrus with a maximum value the day after ovulation. The LH profile showed a more prolonged gradual increase prior to ovulation, than that which has been reported for ponies and horses.     

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)     

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.     

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.     

Twenty-four-hour secretory patterns of cortisol,progesterone and estradiol in heifers during the follicular and luteal phases of the ovarian cycle

Daily variations of plasma cortisol, progesterone and estradiol concentrations were measured by radioimmunoassay in six different normally cycling heifers during estrus (day 1 of the cycle) and diestrus (days 12–15 of the cycle). Each animal was fitted with an indwelling jugular catheter, and blood was withdrawn at 30-min intervals over a 24-h period. Statistical evaluation of the hormonal profiles using time series analysis revealed that all three steroids are secreted episodically with secretory episodes varying in number, magnitude and timing among different heifers. After dividing the 24-h into three 8-h time periods (I, 09.00–17.00 h; II. 17.00–01.00h; III, 01.00–09.00 h) a prominent circadian rhythm was found for cortisol during estrus and diestrus. Diurnal periodicity similar to that of cortisol was noticed for plasma progesterone during estrus but not diestrus when a functional corpus luteum was present. Estradiol secretion during the follicular and luteal phase of the estrous cycle was characterized by intermittent sustained elevations lasting about 9–15 h and marked by a graded rise and fall of hormone levels unrelated to photoperiod.From our results obtained in cycling heifers we conclude the following: (1) Plasma cortisol exhibits a distinct circadian rhythm during estrus and diestrus which is highly correlated with the light–dark cycle. (2) Plasma progesterone during estrus demonstrates a diurnal pattern which is absent in diestrous heifers bearing a corpus luteum. (3) Plasma estradiol lacks circadian rhythmicity but shows a distinct pattern different from that of progesterone, indicating that both steroids are secreted independently and not controlled by a circadian pacemaker.     

间情期与发情期比格犬子宫与卵巢组织形态学的比较

目的比较比格犬的子宫、卵巢在间情期与发情期的组织形态学的差异,为后期研究奠定基础。方法用化学发光法测定了23只经产比格母犬的血清性激素水平,选取经血清学鉴定处于发情期的比格犬2只,间情期的比格犬4只的卵巢和子宫标本,中性多聚甲醛固定,石蜡包埋,常规HE染色镜检,拍照。结果间情期犬子宫及其内膜较薄,间质纤维增生,黄体中以初级卵泡为主,可见1～2个次级卵泡,未见成熟卵泡,卵泡和黄体细胞间纤维较多、血管少。发情期犬子宫及其内膜增厚,内膜腺体腔较大,部分腺体呈分枝状弯曲,腺上皮肿大,胞浆淡染,少数可见核下空泡。卵巢中卵泡数量较多,有初级、次级和1～2个成熟卵泡。黄体细胞数量多,排列规则,境界清楚,间质纤维比较疏松,血管多,未见空泡变性。间情期和发情期犬卵巢中未见明显白体。结论间情期比格犬的性激素维持在一比较低的水平,在发情前期雌激素水平迅速增高,而在排卵后比格犬的孕酮水平较高。随发情周期的变化,卵巢和子宫在形态学上发生相应的变化。     

Effect of a low dose of pregnant mare's serum gonadotropin on follicular recruitment and atresia in cycling rats

Atresia appears to play a central role in selecting the correct number of follicles for ovulation in the rat. A wave of atresia, apparent by noon on metestrus, reduces the number of large healthy follicles to the appropriate quota for ovulation. The purpose of this study was to test the hypothesis that falling levels of gonadotropin on the morning of estrus precipitate the wave of atresia of large follicles seen on metestrus. Endogenous concentrations of follicle-stimulating hormone (FSH) were augmented by a single injection of pregnant mare's serum gonadotropin (PMSG; 0.025 IU/gram body weight) administered at different times during the periovulatory period. Animals given PMSG at 0800 h on estrus (when the endogenous FSH surge was waning) had a supernormal number of large healthy follicles in their ovaries at 1200 h on metestrus. This increase in large healthy follicles was accompanied by a decrease in large atretic follicles in the ovaries. The same dose of PMSG, when administered at other times during the periovulatory period, did not affect any of the parameters measured. These observations suggest that the wave of atresia normally seen in large follicles on metestrus is triggered by the decline in the concentration of FSH during the morning of estrus and can be prevented by prolonging the surge of FSH with administration of PMSG.     

Concentration of prostaglandins F in uterine venous plasma of anesthetized mares during the estrous cycle and early pregnancy

Prostaglandins F were quantitated by radioimmunoassay in uterine venous plasma of anesthetized mares on day 7 of estrus, days 2, 6, 10, 14 or 18 of diestrus and days 10, 14 or 18 of pregnancy. The PGF concentration was greater (P<.01) at day 14 of diestrus than at all other days studied. The concentrations at days 10 and 18 of diestrus and at days 10, 14 and 18 of pregnancy were greater (P<.05) than at day 7 of estrus and days 2 and 6 of diestrus. PGF concentrations at days 10 and 14 were greater (P<.01) for diestrous than for pregnant mares.     

Coexpression of adrenomedullin and its receptors in the reproductive system of the rat: effects on steroid secretion in rat ovary

The present study demonstrates the expression of adrenomedullin (ADM) in the reproductive system of the female rat and its effect on the secretion of estradiol and progesterone. Ovarian ADM and Adm mRNA levels were decreased at estrus, whereas oviductal Adm mRNA levels were low at proestrus. Both tissues were shown to coexpress mRNAs encoding the calcitonin receptor-like receptor and receptor activity-modifying protein 1 (Ramp1), Ramp2, and Ramp3. Gel filtration chromatography of ovarian extracts showed two peaks, with the predominant one eluting at the position of authentic rat ADM (1-50) at estrus and at the position of ADM precursor at diestrus. Positive ADM immunostaining was localized in the granulosa and theca cells of the follicle and corpora lutea of the ovary. Adrenomedullin inhibited FSH-induced estradiol secretion in 2-day-old follicles and also suppressed eCG-stimulated progesterone release in corpora lutea. The inhibitory effect of ADM on the follicles and the corpora lutea was abolished by calcitonin gene-related peptide (8-37) and ADM (22-52), respectively. The presence of ADM and the gene expression of ADM and its receptor components in the female reproductive system suggest a paracrine effect of ADM on ovarian steroidogenesis.     

Evidence for a role of androgens in the growth and maturation of ovarian follicles in rats

Changes in the morphology and steroid content of ovaries were studied after 48 h of intravenous injection of 100 microgram of cyproterone acetate or flutamide to diestrus or estrous rats. Treatment with cyproterone acetate at diestrus caused a decrease in the number of small follicles (less than 200 micrometer), freshly formed corpora lutea and the levels of estradiol-17beta in the ovary, suggesting inhibition of ovulation. Following flutamide administration at diestrus, the number of follicles at all stages of development were reduced with a concomitant decrease in the ovarian levels of the hormones. Thus, flutamide suppressed the growth and maturation of follicles. On administration of these drugs at estrous, the steroid content of ovaries was more pari passu with the increase in the number of mature and medium follicles. The differential effects of the two drugs are discussed in the light of these observations.     

Patterns of follicular growth during the four-day estrous cycle of the rat

Female Sprague-Dawley rats underwent laporatomy during metestrus at 70 to 75 days of age or remained untreated to study the effects of surgical stress on follicular growth. Groups of rats were killed on each day of a 4-day estrous cycle, serial sections of the ovaries were prepared histologically and the number and size of follicles with one or more complete layers of cuboidal granulosa cells were determined. Since no differences due to surgery were found, the data were pooled by day of the estrous cycle (17 or 18 rats/day of cycle) for characterization and comparison of size distribution of follicles on different days of the estrous cycle. Follicles were classified as atretic or healthy and divided into groups by increments of 20 micron of diameter for graphing. Data were analyzed by analysis of variance and least squares means. Significant differences were found in the distribution of both healthy and atretic follicles among days of the estrous cycle. At least 21 follicles/ovary were recruited from less than 260 micron into greater than 260 micron in diameter between proestrus and estrus, and the follicles for ovulation were selected by diestrus. A greater number of growing follicles of 70 to 100 micron in diameter were present at diestrus. From the disappearance of follicles greater than 260 micron between estrus and proestrus, it appears that atresia is a very rapid process.     

Supression by copulation of the inhibitory effect of p-chlorophenylalanine on rat ovulation (author's transl)]

The effect of copulating on reflex ovulation was studied in rat. The effect of PCPA (300 mg/kg, i.p.) on ovulation and reproduction was compared by evaluating number of oocytes in tubes, histologic features of ovaries, vaginal cycle, insemination, fertilization and number of embryos per rat. PCPA, administered on 9th and 16th h of the estrus phase, totally inhibits ovulation, stimulates reproductive behaviour and prolongs the estrogenic phase. When the animals are kept in copulatory conditions for 16 or 46 hours, the inhibition induced in ovulation disappears to the extent that 60% of the rats become pregnant though the number of embryos is under that of the control group. The farther the treatment with PCPA within the same cycle in the ovulatory period, the greater the inhibitory effect on ovulation is and the lesser the neutralizing effect produced by reflexes related to copulation. Administration of PCPA at the 16 hour of the diestrus causes a greater increase in the average number of embryos-as compared to administration at the 9 hour. In periods longer than 48 hours before ovulation, the inhibition brought about by PCPA is not suppressed by copulatory conditions kept for 16 or 24 hours and is only neutralized if they are kept during a complete cycle. Those treated with PCPA in the diestrus phase and maintained in copulatory conditions for 46 hours, present a higher average of embryos than those maintained in similar conditions for 16 hours.     

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。     

In vitro sensitivity of rat ovarian follicles to the direct action of various hormones applied in particular stages of sexual cycle

The direct action of various hormones on the ovary of adult rat ovaries has been investigated. The ovaries were surgically exposed at particular stages of sexual cycle i.e. in estrus, metestrus, diestrus and proestrus, and were injected with various doses of gonadotropic hormones. Injected ovaries were excised 56 hrs after injection and examined histologically. The reactivity of the ovaries to hormone applied was dependent on stage of the cycle. The considerable increase of the amount of t1 and t2 follicles was observed after injection of gonadotropins in estrus stage. In metestrus stage the influence of gonadotropins on the follicles more advanced in development was pronounced. Numerous follicles t3 appeared in diestrus under the influence of all used hormones.     

Ovarian superstimulation after follicular wave synchronization with GnRH at two different stages of the estrous cycle in cattle

The objective of this study was to evaluate superovulatory programs based on synchronization of follicular waves with GnRH at 2 different stages of the estrous cycle. Sixteen Holstein cows were randomly assigned to 1 of 3 groups and administered GnRH (Cystorelin, 4 ml i.m.) between Days 4 and 7 (Groups 1 and 3) or between Days 15 and 18 (Group 2) of the estrous cycle (estrus = Day 0). Four days after GnRH treatment, > or = 7-mm follicles were punctured in Groups 1 (n = 6) and 2 (n = 6) or were left intact in Group 3 (n = 4). All cows were superstimulated 2 d later (i.e., from Days 6 to 10 after GnRH treatment) with a total of 400 mg NIH-FSH (Folltropin-V) given twice daily in decreasing doses. The GnRH treatment caused a rapid disappearance of large follicles (P < 0.005), rapid decrease in estradiol concentrations (P < 0.003), and increase in the number of recruitable follicles (4 to 6 mm; P < 0.04), indicative of the emergence of a new follicular wave within 3 to 4 d of treatment. Between 4 and 6 d after GnRH treatment, the mean number of 4- to 6-mm follicles decreased (4.7 +/- 1.8 to 1.5 +/- 3.3) in the nonpunctured group but increased (3.9 +/- 1.0 to 7.3 +/- 1.9) in the punctured group of cows (P < 0.05). In response to FSH treatment, the increase in the number of > or = 7-mm follicles was delayed by approximately 2 d in the nonpunctured group (P < 0.006). Moreover, the mean number of > or = 7-mm follicles at estrus was higher (16.9 +/- 1.7 vs 11.5 +/- 3.0; P < 0.1) in the punctured than the nonpunctured group. The increase in progesterone concentration after estrus was delayed in the nonpunctured group (P < 0.1) compared with the punctured follicles. Mean numbers of CL as well as freezable (Grade 1 and 2) and transferable (Grade 1, 2 and 3) embryos were similar (P > 0.1) in punctured and nonpunctured groups. Spontaneous estrus did not occur prior to cloprostenol-induced luteolysis in any group, and stage of the estrous cycle during which GnRH was given did not affect (P > 0.1) hormonal and follicular responses in the punctured groups. In conclusion, GnRH given at different stages of the estrous cycle promotes the emergence of a follicular wave at a predictable time. Puncture of the newly formed dominant follicle increases the number of recruitable follicles (4 to 6 mm) 2 d later and, in response to superstimulation with FSH, causes a greater number and faster entry of recruitable follicles into larger classes (> or = 7 mm) and a faster postovulatory increase in progesterone concentrations.     

Concentration of prostaglandins F in uterine venous plasma of anesthetized mares during the estrous cycle and early pregnancy.

Prostaglandins F were quantitated by radioimmunoassay in uterine venous plasma of anesthetized mares on day 7 of estrus, days 2, 6, 10, 14 or 18 of diestrus and days 10, 14 or 18 of pregnancy. The PGF concentration was greater (P less than .01) at day 14 of diestrus than at all other days studied. The concentrations at days 10 and 18 of diestrus and at days 10, 14 and 18 of pregnancy were greater (P less than .05) than at day 7 of estrus and days 2 and 6 of diestrus. PGF concentrations at days 10 and 14 were greater (P less than .01) for diestrous than for pregnant mares.