FSH influences follicle viability, oestradiol biosynthesis and ovulation rate in Romney ewes

Injection of steroid-free bovine follicular fluid (bFF; 2 X 5 ml s.c. 12 h apart) into anoestrous ewes lowered plasma FSH concentrations by 70% and after 24 h had significantly (P less than 0.01) reduced the number of non-atretic follicles (greater than or equal to 1 mm diam.) without influencing the total number of follicles (greater than 1 mm diam.) compared to untreated controls. Hourly injections of FSH (10 micrograms i.v. NIH-FSH-S12) for 24 h did not influence the number of non-atretic follicles but did negate the inhibitory effects of bFF on follicular viability. Hourly injections of FSH (50 micrograms i.v., NIH-FSH-S12) + bFF treatment for 24 h significantly increased the total number of non-atretic follicles, and particularly the number of medium to large non-atretic follicles (greater than 3 mm diam.) compared to the untreated controls (both P less than 0.01). The 10 micrograms FSH regimen (without bFF) significantly increased aromatase activity in granulosa cells from large (greater than or equal to 5 mm diam.; P less than 0.01) but not medium (3-4.5 mm diam.) or small (1-2.5 mm diam.) follicles compared to controls. The 10 micrograms FSH + bFF regimen had no effect on granulosa-cell aromatase activity compared to the controls. However, the 50 micrograms FSH plus bFF regimen increased the aromatase activity of granulosa cells from large, medium and small non-atretic follicles 2.6-, 8.3- and greater than or equal to 11-fold respectively compared to that in the control cells. Ewes (N = 11) that ovulated 2 follicles had significantly higher plasma FSH concentrations from 48 to 24 h and 24 to 0 h before the onset of a cloprostenol-induced follicular phase (both P less than 0.01) than in the ewes (N = 12) that subsequently ovulated one follicle. Hourly FSH treatment (1.6 micrograms i.v., NIAMDD-FSH-S15) for 24 h but not for any 6 h intervals between 48 and 24 h or 24 and 0 h before a cloprostenol-induced luteolysis also resulted in significant increases (P less than 0.05) in the number of ewes with 2 ovulations.(ABSTRACT TRUNCATED AT 400 WORDS)     

Superovulation and early embryo development in the adult mouse after prenatal exposure to diethylstilboestrol

Pregnant mice were injected subcutaneously with diethylstilboestrol (DES: 10 micrograms/kg body weight in 0.1 ml corn oil) or corn oil alone on Day 15 or 16 of gestation (Day 1 = day of copulatory plug) and allowed to give birth. Female progeny from control and DES-exposed animals were superovulated with exogenous gonadotrophins at 6-8 weeks of age. In-vivo results indicated that the total number of ovulated ova, 2-cell embryos and blastocysts were significantly increased in DES-exposed progeny but that there was a decline in developmental potential from the ovulated ova stage to the blastocyst stage in these animals. However, there was no significant difference in the in-vitro development of 2-cell embryos to the blastocyst stage between control and DES-exposed animals. These results indicate that the ovaries of mice exposed in utero to DES are capable of responding to exogenous gonadotrophins and that second generation progeny have the potential for normal development to the early postblastocyst stage of embryogenesis. The in-vivo decline in developmental potential may be attributable to reproductive tract abnormalities rather than ova/embryo defects.     

Embryonic mortality during the pre- and post-implantation periods of pregnancy in mature mice after superovulation.

Sexually mature mice were stimulated to superovulate by giving exogenous gonadotrophins at known stages of the oestrous cycle. Untreated animals which ovulated spontaneously served as controls. The number of oocytes ovulated by each female was estimated from counts of the number of CL of pregnancy, and the incidence of embryonic mortality during the pre- and post-implantation stages of pregnancy was assessed from the number of zygotes recovered from the reproductive tract at 2-0 and 4-0 days post coitum and of conceptuses examined at 7-5 and 11-5 days post coitum. The mean number of oocytes ovulated by treated animals was 39-54, compared with 12-80 in controls: in mice which had superovulated, 44% of the ova were lost before implantation compared with about 10% in the controls. Further losses occurred about the time of implantation and at mid-pregnancy and thus the number of embryos classified as normal rarely exceeded the maximum found in controls. Death at mid-pregnancy seemed to be preceded by developmental retardation. The possibility that genetic and environmental factors play a role in embryonic loss after superovulation is discussed.     

Effects of hyperstimulation with gonadotrophins and age of females on oocytes and their metaphase II status in rats

The present study was undertaken to evaluate the effects of hyperstimulation and aging on the number and proportion of oocytes in the metaphase II stage in female Wistar rats. It explored the validity of the hypothesis that a combination of hyperstimulation with pregnant mare serum gonadotrophins (PMSG) and age could compromise, to a greater extent, the oocyte quality as indicated by the proportion of ovulated oocytes in the metaphase II stage. Female Wistar rats were stimulated with varying doses of PMSG and human chorionic gonadotrophins (hCG) and the number and proportion of ovulated oocytes in the metaphase II stage were examined and compared between different groups of young adult (8-10 weeks old) and aging (30-32 weeks old) female rats. While spontaneous ovulation occurred in all young adult rats, only 50% of the aging rats did. The ovulation rate in aging rats was increased from 50 to 93% when non-PMSG-stimulated rats were given a dose of 10 IU of hCG at proestrus. The lower number of ovulated oocytes noted, even in those hyperstimulated with high doses of PMSG/hCG, also indicated a reduction in fertility in aging rats. Under the influence of high doses of PMSG, all aging rats ovulated, but as with the young adult rats, a higher dose of hCG was needed to achieve the maximum number of ovulated oocytes from the PMSG-induced expanded pool of preovulatory follicles. However, the average number of ovulated oocytes in aging rats was, nevertheless, still significantly lower than in young adult rats even when approximation of weight was considered. No consistent significant difference in proportion of normal oocytes was noted within groups and between young adult and aging rats. A lower proportion of ovulated oocytes was arrested at the metaphase II stages when rats, whether they were young adult or aging, were hyperstimulated with 40 IU of PMSG. However, this proportion was restored to normal (about 100%) when a higher dose of hCG, which is a signal responsible for initiating oocyte maturation, was used. Results of the present study showed that there appears to be an age-related reduction of sensitivity of the preovulatory follicles to the ovulation induction signal of hCG and thus higher doses of hCG were needed to ovulate the PMSG-induced expanded pool of dominant follicles. In older rats, apart from the obvious depletion of the pool of follicles, the evidence from the present study suggests that some of these older rats do have follicles, but that these were unable to develop to preovulatory follicles, probably because of the absence of sufficiently high levels of gonadotrophins essential for the initiation of folliculogenesis. PMSG-hyperstimulation can affect nuclear maturation; the proportion of ovulated oocytes not arrested at the metaphase II stage was higher. However, the proportion of ovulated oocytes at the metaphase II was restored to normal by increasing the dose of hCG use. Hence, meiotic aberration in rats is not age-dependent but rather dependent on the amplitude of the luteinizing hormone (LH)/hCG surge present. The results from this study nullified the hypothesis that hyperstimulation in combination with aging would lead to a higher proportion of abnormality in ovulated oocytes with respect to their being at inappropriate meiotic stages.     

Estrous stage- and animal age-independent superovulation in the BrlHan:WIST@Jcl(GALAS) rat.

In most strains of rats, the effects of treatment for the induction of superovulation show major strain differences and are strongly influenced by the stage of the estrous cycle. This study demonstrated, however, that superovulation was easily induced in Wistar strain Brl Han:WIST@Jcl(GALAS) rats by PMSG and hCG administration. To confirm the effects of such treatment, we studied age differences in egg collection efficiency. After superovulation was induced by intraperitoneal administration of 150 IU/kg PMSG and 75 IU/kg hCG given 48 h apart, the mean numbers of oocytes obtained from rats at 4, 8, 12, 20 and 28 weeks of age were 38.9, 33.5, 46.1, 26.9 and 21.3, respectively. No differences caused by the estrous stage at the PMSG administration were observed. In an embryo transfer experiment, fertilized eggs obtained from superovulated rats at each week of age showed equivalent viability until full-term to those from untreated rats. These results suggest that estrous stage-independent superovulation is effective not only in the pre-pubertal stage but also in adult rats.     

The effect of bovine pestivirus infection on the superovulatory response of Friesian heifers

The pathogenesis of reproductive loss associated with bovine pestivirus infection during the preovulatory period was investigated using superovulated heifers. Twenty-five Friesian heifers were selected and randomly assigned to either a control group (n = 12) which did not become infected or to a treatment group (n = 13) which became infected following intranasal instillation of 2 ml of serum inoculum containing 5.5 log(10) TCID(50)/ml non-cytopathic virus, 9 d prior to artificial insemination (AI). Transrectal ultrasonography was used to monitor follicular development and ovulation during the superovulatory period. Animals were superovulated using a standard protocol of twice-daily injections of FSH-P and then were inseminated twice commencing 12 h after the onset of estrus. The intensity of expression of estrus was higher in the control heifers than in the pestivirus-infected heifers. Of 13 pestivirus-infected heifers, only 3 heifers displayed standing estrus compared with that in the control group, in which 10 of 12 heifers exhibited standing estrus. The mean number of ova/embryos recovered from the control group heifers was 5.75 +/-2.31, of which 4.00 +/- 0.72 were evaluated as transferable quality embryos. In comparison, heifers in the pestivirus-infected group yielded only a mean of 0.60 +/-0.34 ova/embryos, of which 0.23 +/- 0.22 were transferable quality embryos. Based on ultrasonographic examination, 24 h after the first AI 82% of the presumptive ovulatory follicles had ovulated in the control group compared with an ovulation rate of only 17% in the treated group. The results of this experiment demonstrated that bovine pestivirus infection during the preovulatory period could adversely affect ovulation, thus leading to a significant reduction in the number of palpable corpora lutea and in the number and quality of embryos recovered.     

Inhibition of sperm transport and fertilization in superovulating ewes

In Experiment 1, all ewes were treated with follicle stimulating hormone (FSH-P) to induce superovulation. Ewes came into natural estrus or were treated with prostaglandin F(2)alpha (PGF(2)alpha) or 6-methyl-17-acetoxyprogesterone (MAP) to regulate the time of estrus. The ewes were mated during estrus and necropsied 3 h after mating. Regulation of estrus with either compound reduced the number of sperm recovered from the cervix, uterus, and oviducts and increased the proportions of sperm recovered from the cervix and uterine body that were immotile, dead, or had disrupted membranes. In Experiment 2, all ewes were in natural estrus. They either ovulated naturally or were superovulated, and ewes in each group were necropsied at 3 or 23 h after mating. Superovulation reduced the number of sperm in oviducts, uterus, and anterior segments of the cervix at both time intervals and increased the proportions of sperm that were immotile, dead, or had disrupted membranes. In Experiment 3, of 3x2 design, ewes were in either natural estrus or estrus regulated with PGF(2)alpha or with MAP; they ovulated naturally or were superovulated. Ewes were necropsied 3 d after mating and ova were examined. Both regulation of estrus and superovulation reduced the proportion of ova that were fertilized and reduced the number of accessory sperm attached to fertilized ova.     

The effect of platelet activating factor on ovulation.

The mechanism of ovulation has been compared to an inflammatory reaction. Platelet activating factor (PAF) is an important mediator of inflammation as it may induce the production of prostaglandins and lysosomal enzyme. We evaluated the potential role of PAF in PMSG-HCG induced ovulation using CV3988, a specific PAF receptor antagonist in a superovulated ICR mice (9-12 weeks old). CV3988 blocked the ovulation in a dose dependent manner, and the significant reduced ovulatory efficiency was observed at more than 500 micrograms dose (p less than 0.001). The ovulatory efficiency reduced by CV3988 was reversed by PAF in a dose dependent manner. In vitro fertilization (IVF) rate of follicular oocytes with treatment of CV3988 was not different from that of ovulated ova without treatment. These results suggest that PAF may be involved in the ovulation process but the presence of PAF may not be essential for the fertilization of the ova as IVF.     

Adrenalectomy: Effects on mating behavior,ovulation, and early gestation in the cycling rat

Adrenalectomy was performed 4 weeks prior to evaluating the effects on mating behavior, ovulation, and the first 10 days of gestation in 4-day cycling female rats. Mating behavior was essentially normal and occurred at times similar to sham-adrenalectomized animals on the afternoon of predicted proestrus. The number of ova ovulated was significantly reduced in mated adrenalectomized subjects in contrast to data reported by Rodgers (1971) for intact females mated at similar times in proestrus. Embryonic swellings were significantly fewer at day 10 of gestation than in the sham-adrenalectomized group providing additional evidence of reduced numbers of ova ovulated. Embryonic weights at day 10 of gestation were not affected by adrenalectomy. Removal of adrenocortical hormones significantly increased uterine weight on the morning of vaginal estrus. The data fail to show a significant delay in sexual receptivity as a result of removing adrenal progesterone, but suggest that adrenalectomy interferes with the neuroendocrine reflex mediating ovulation.     

Effect of estradiol benzoate or GnRH treatment prior to superstimulation in CIDR-treated, Korean native cows (Bos taurus)

The objective of this study was to evaluate the effectiveness of superovulatory protocols by synchronizing the emergence of the follicular wave using estradiol benzoate (EB) or GnRH in CIDR-treated, Korean cows. Sixty-six cows were used in the study and these were divided into three groups. The standard group comprised cows that were between days 8 and 12 of their estrous cycle (n=22). The remaining 44 cows, at all other stages of the estrous cycle, received CIDR and were assigned to two treatment groups that received either 2mg EB (EB-CIDR group, n=22) or 100 microg GnRH (GnRH-CIDR group, n=22) 1 day after CIDR insertion. Gonadotropin treatment began between the 8th and 12th days of the estrous cycle in the standard group, 5 days after EB injection in the EB-CIDR group, and 3 days after GnRH injection in the GnRH-CIDR group. All cows were superovulated with porcine FSH (pFSH) twice daily, with the dose (total 28 mg) decreasing gradually over 4 days. On the 5th and 6th injections of pFSH, 25 and 15 mg doses of PGF(2alpha) were administered. CIDR was withdrawn at the 7th pFSH injection and the cows received 200 microg GnRH at 24h after CIDR withdrawal. Cows were artificially inseminated twice at 36 and 48 h post-CIDR withdrawal and embryos were recovered 7 days after the 1st insemination. The numbers of preovulatory follicles (22.9-28.2), ovulated preovulatory follicles (17.6-21.7) and CL (15.9-17.9) detected by ultrasonography did not differ among groups (P>0.05). Similarly, the numbers of total ova (6.7-10.0), transferable embryos (4.0-6.0), degenerate embryos (1.1-1.8) and unfertilized ova (1.3-4.3) did not differ among groups (P>0.05). Progesterone and estradiol concentrations during superovulation treatments and at embryo recovery were also the same in all groups (P>0.05). We conclude that in CIDR-treated Korean native cows, superovulatory treatments that follow administration of either EB or GnRH (at any stage of the estrous cycle) result in both a superovulatory response and embryo yield comparable to conventional superovulation protocols.     

Estrous cycle stage-independent treatment of PMSG and hCG can induce superovulation in adult Wistar-Imamichi rats.

The estrous cycle influence on the number of ovulated eggs after injection of pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) was investigated in 12, 18, and 24 weeks old adult female Wistar-Imamichi (WI) rats. PMSG (150 IU/kg) was injected at metestrus, diestrus, proestrus, or estrus, followed by hCG (75 IU/kg) 55 h later. Ovulation was induced at all ages and stages of the estrous cycle. The number of ovulated eggs was not affected by stage for similarly aged rats, however, the number of ovulated eggs obtained after treatment decreased with age. These results demonstrate that the PMSG/hCG treatment can induce ovulation at any stage of estrous cycle in WI rats and efficient superovulation at 12 weeks of age.     

外源激素组合控制青春期前奶山羊卵泡超数同步发育及其卵母细胞发育潜能的评价

本研究的目的是探索自青春期前奶山羊获取大量可用于体细胞核移植的卵母细胞的可能性。为此,本研究比较了几种不同组合的激素处理方法(对照、FSH、E2-P4和E2-P4-FSH)对出生39-60日龄的奶山羊卵巢大小、卵泡数量和卵泡大小的影响:同时将出生39-120日龄奶山羊按年龄分成三组来研究年龄对激素处理时招募起始生长卵泡数量的影响:然后,比较了来自E2-P4- FSH和FSH处理的早青春期前奶山羊卵巢上直径大于3mm卵泡中卵母细胞减数分裂能力;最后,通过SCNT方法验证E2-P4-FSH处理的早青春期前奶山羊卵巢上直径大于3mm卵泡中卵母细胞的发育能力。在四组激素处理的早青春期前奶山羊中,E2-P4-FSH处理组的卵巢最大、卵泡(直径大于3 mm)数量最多。在不同的年龄组中,39-60天组奶山羊卵巢上直径大于3mm的卵泡数量显著多于61-90天和91-120天组的。卵母细胞减数分裂能力的分析结果表明,来自E2-P4-FSH处理组的卵母细胞减数分裂能力显著高于FSH处理组的卵母细胞。与E2-P4-FSH处理后的成年奶山羊卵母细胞相比,早青春期前奶山羊卵母细胞发育能力较低:卵母细胞成熟后,作为受体用于体细胞核移植后的克隆囊胚发育率低于成年奶山羊(15.3%versus 22.1%,P<0.01)。然而,早青春期前的奶山羊经E2-P4-FSH处理后,自每头羊卵巢上直径大于3mm的卵泡数显著高于成年奶山羊(108±10.3 versus 28±5.0),因此,每头早青春期前奶山羊产生的克隆囊胚绝对数量显著高于成年奶山羊(7.1±2.7 versus 4.2±1.4)。由此,从本研究可以得出结论:E2-P4-FSH处理的早青春期前奶山羊能够为体细胞核移植研究提供相对多数量的具备一定发育能力的成熟卵。     

Passive immunization with anti-oestradiol antibodies during the luteal phase of the menstrual cycle potentiates the perimenstrual rise in serum gonadotrophin concentrations and stimulates follicular growth in the cynomolgus monkey (Macaca fascicularis)

Three unilaterally ovariectomized cynomolgus monkeys, in which menstrual cycles were driven by pulsatile infusion of synthetic GnRH at a fixed frequency of 1 pulse/h, were provided with a continuous infusion of ovine anti-oestradiol gamma-globulin beginning 13 days after ovulation and continuing for 7 days thereafter. Plasma concentrations of both FSH and LH rose at the start of the antibody infusion and remained elevated throughout the 7-day treatment regimen when compared with control (non-immune gamma-globulin-treated or untreated) animals. Morphometric examination of ovaries at the end of the experimental and control infusions revealed a significant difference (P less than 0.05) in the average size of the largest non-atretic antral follicle in each of the experimental animals when compared with that of the control animals (2.45 +/- 0.23 vs 1.30 +/- 0.53 mm). Collectively, the 3 control animals possessed 9 non-atretic antral follicles greater than 1.0 mm diameter, none of which exceeded a diameter of 2.0 mm. In contrast, the experimental animals had 28 non-atretic follicles of greater than 1.0 mm diameter, 8 of which exceeded 2.0 mm. These observations are consistent with the hypothesis that oestrogen and progesterone are the primary agents responsible for the restraint of gonadotrophin secretion and preovulatory follicular growth during the luteal phase of the primate menstrual cycle.     

Superovulating cows with follicle stimulating hormone and pregnant mare's serum gonadotrophin

A total of 511 embryos was recovered non-surgically from nearly 100 superovulated or untreated donors. Superovulation with FSH-LH resulted in more corpora lutea, recovered ova, and pregnancies (P<.01) than superovulation with PMSG. No differences were observed in numbers of ovulations, embryos recovered, or pregnancies per donor when prostaglandin F_2α was given to donors 2 versus 3 days following gonadotrophin treatment.Pregnancy rates of 12, 31, 58, and 63% were obtained from groups of embryos classified morphologically as poor, fair, good, and excellent (P<.05). Morphologically normal embryos collected and transferred at 5 to 6 days of gestation resulted in more (P<.05) fetuses (75%) than morphologically normal embryos at 8 to 9 days of gestation (56%), but neither was significantly different from morphologically normal embryos at 6.5 to 7.5 days of gestation (61%). There was no difference (P>.05) between pregnancy rates when retarded embryos from untreated donors (12%) were compared to retarded embryos from superovulated donors (22%). However, a higher proportion of morphologically normal embryos from untreated donors developed into fetuses (71%) than did morphologically normal embryos from superovulated donors (59%, P<.05).     

Influence of follicular health on the steroidogenic and morphological characteristics of bovine granulosa cells in vitro

In 24-h cultures, steroid production by cells from non-atretic follicles increased with increasing follicular diameter. Cells from atretic follicles, of all sizes, produced low amounts of oestradiol-17 beta, but very high amounts of progesterone, relative to cells from non-atretic follicles. Increasing the culture period to 72 h caused little change in daily progesterone and oestradiol-17 beta production by granulosa cells from atretic follicles. In contrast, in cells from non-atretic follicles, daily progesterone production increased and daily oestradiol-17 beta production decreased to the levels observed with cells from atretic follicles. Dibutyryl cyclic AMP (1.0 mM) significantly stimulated progesterone production by cells from atretic, but not from non-atretic, follicles. Testosterone (1 microgram/ml) had no effect on progesterone production by cells from atretic follicles, while oestradiol-17 beta, oestrone, testosterone, androstenedione and 5 alpha-dihydro-testosterone (0-1000 ng/ml) each significantly suppressed progesterone production by cells from non-atretic follicles in a dose-dependent manner. Morphometric analysis revealed few subcellular differences between cells from non-atretic and atretic follicles. Mean cell volume was significantly higher for cells from atretic compared to non-atretic follicles, but the mean volumes of the major subcellular components were not influenced by follicle health. The mean surface area of the plasma and nuclear membrane, and granular endoplasmic reticulum was also significantly higher in cells from atretic compared to non-atretic follicles.     

Alteration of follicular dynamics and superovulatory responses by gonadotropin releasing hormone and follicular puncture in cattle: a field trial

A field experiment was conducted to determine the influence of follicular alteration on superovulatory responses. Ultrasonography was performed once daily over 4 d prior to gonadotropin treatment (Day 0), on the day of estrus during superstimulation, and on the day of embryo collection to monitor follicular development. Animals were superstimulated between Days 8 and 12 of the estrous cycle. Follicular status was altered 2 d prior to initiation of superstimulation (Day 0) with GnRH (Cystorelin, 200 micrograms i.m.) administered with (GnRH-puncture group, n = 31) or without (GnRH-no puncture group, n = 52) concomitant removal of the largest follicle by follicular aspiration. Responses were compared with those of an untreated control group superovulated 8 to 12 d after estrus (n = 102). The proportion of animals with a high number (> or = 2) of large follicles (> = 7 mm) on Day 0 was lower (P < 0.001) in the 2 GnRH-treated groups than in the control group, while the increase in the number of medium size follicles (4 to 6 mm) on Day 0 was greater (P < 0.02) in the GnRH-puncture group. During superstimulation, the proportion of superovulatory cycles with a high follicular (> or = 10 follicles) response was similar in the control and GnRH-no puncture groups. Within the GnRH-treated animals, follicular and ovulatory responses were greater in the GnRH-puncture than in the GnRH-no puncture group (P < 0.001 to P < 0.02). Despite these changes in follicular and ovulatory responses, however, the mean number of embryos produced did not differ (P < 0.1) among treatments (4.3 +/- 0.4, 3.7 +/- 0.7, and 5.4 +/- 0.8 in control, GnRH-no puncture, and GnRH-puncture groups, respectively). This was due primarily to an increase in the mean numbers of unfertilized ova (P < 0.005) and in degenerated embryos (P < 0.06) in the GnRH-puncture group. Results indicate that the beneficial effects of treatment with GnRH and follicular puncture 2 d prior to superstimulation on follicular and ovulatory responses were limited by an increase in the number of unfertilized ova and degenerated embryos.     

Superovulation can reduce the developmental competence of bovine embryos

This study was done to determine if different superovulatory regimens could have an effect on the percentage of embryos produced using IVM/IVF/IVC. Cyclic heifers (n = 22) were superovulated between Days 8 and 12 of the estrous cycle with 4, 6 or 8 constant doses of FSH-P (4 mg each, twice daily) +/- the addition of 1 mg prostaglandin 24 h before slaughter. Ovaries from these superovulated cows and from untreated cows were collected and the follicles dissected. Oocytes were classified according to the appearance of their cumulus and cytoplasm. Individual culture as well as group culture were performed but an individual culture reduced the percentage of oocytes developing into embryos for both untreated and superovulated animals. The results indicated that despite the superovulation regimen the developmental competence of the oocytes collected was lower (0 to 15% embryos) than that of oocytes from untreated animals (20 to 34% embryos). Small follicles ( < or = 2.7 mm) yielded mostly oocytes with an incomplete or partially expanded cumulus investment that never developed into an embryo. Differences in the morphology of the oocytes from medium (2.7 to 8 mm) and large ( > or = 8 mm) follicles were apparent, but equal developmental rates were obtained between all classes of oocytes (12 and 8% embryos, respectively). Follicular atresia was reduced significantly after superovulation (81% nonatretic follicles in treated vs 42% nonatretic follicles in untreated animals); however oocytes from atretic and slightly atretic follicles developed similarly to those from nonatretic follicles. These results suggest that although superovulation increases follicular size and decreases atresia, these conditions are not sufficient to confer developmental competence on the oocytes.     

Fertilizability of Superovulated Eggs by Estrous Stage-independent PMSG/hCG Treatment in Adult Wistar-Imamichi Rats

We investigated the fertilization and developmental ability of superovulated eggs obtained from adult Wistar-Imamichi (WI) rats, by using pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) treatment. Female WI rats, 11–13 weeks of age, were divided into four groups by estrous stage (metestrus [ME], diestrus [DE], proestrus [PE], or estrus [E]). PMSG (150 IU/kg) and hCG (75 IU/kg) were injected at an interval of 48 or 55 h and the female rats were mated with mature male rats. The ovulated eggs were collected 20, 24, and 27 h after hCG injection. Regardless of the estrous stage at the time of PMSG injection, the treated rats mated and ovulated similar to the untreated spontaneously ovulated rats (S group). Although the proportion of fertilized eggs in the E- and PE-treated groups was less than the S group 20 h after hCG injection, the proportion was not different among all treated and S groups 24 h after hCG injection. The proportion of fertilized eggs using in vitro fertilization and the proportion of offspring obtained from 2-cell stage embryo transfer did not differ among the treated and S groups. In comparison with PMSG/hCG-treated immature rats, mating and ovulation rate of adult rats were significantly higher. The proportion of fertilized eggs obtained from mated rats did not differ between immature and adult rats. These results demonstrate that adult WI rats are good egg donors for reproductive biotechnological studies using unfertilized or fertilized eggs.     

Ovarian follicle dynamics in immature rats treated with a luteinizing hormone-releasing hormone antagonist (Org. 30276)

The high concentrations of gonadotropins present in immature female rats by the end of the second week of life were suppressed by treatment with an antagonist against luteinizing hormone-releasing hormone (LHRH-A; Org. 30276) on Days 6, 9, 12, and 15 of life. Differential ovarian follicle counts were made on Days 15, 22, 28, and on the day of first estrus of all growing follicles and follicles greater than or equal to 100 x 10(5) microns 3 (mostly antral). In LHRH-A-treated rats, a retardation of follicle growth was noted on Day 15, followed by a gradual loss of growing follicles that amounted to 20% on Day 22 and 40% on Day 28; at first estrus, the total population of growing follicles was only 50% of that present in control rats. Antral follicles, first present at 22 days of age, were lower in number at 28 days of age and at first estrus in LHRH-A-treated rats; this was true for both healthy and atretic follicles. Ovarian weights were significantly reduced in LHRH-A-treated rats at 15 and 28 days of age and on the day of first estrus. However, the numbers of corpora lutea following the first, and normally timed, ovulation were the same in both groups. It was concluded that for early recruitment of follicles to reach a full-sized pool of growing follicles at the age of puberty, high concentrations of gonadotropins early in life have a significant role.     

Timing of the onset and duration of ovulation in superovulated beef heifers

Thirty-two beef heifers were induced to superovulate by the administration of follicle stimulating hormone-porcine (FSH-P). All heifers received 32 mg FSH-P (total dose) which was injected twice daily in decreasing amounts for 4 d commencing on Days 8 to 10 of the estrous cycle. Cloprostenol was administered at 60 and 72 h after the first injection of FSH-P. Heifers were observed for estrus every 6 h and were slaughtered at known times between 48 to 100 h after the first cloprostenol treatment. The populations of ovulated and nonovulated follicles in the ovaries were quantified immediately after slaughter. Blood samples were taken at 2-h intervals from six heifers from 24 h after cloprostenol treatment until slaughter and the plasma was assayed for luteinizing hormone (LH) concentrations. The interval from cloprostenol injection to the onset of estrus was 41.3 +/- 1.25 h (n = 20). The interval from cloprostenol injection to the preovulatory peak of LH was 43.3 +/- 1.69 h (n = 6). No ovulations were observed in animals slaughtered prior to 64.5 h after cloprostenol (n = 12). After 64.5 h, ovulation had commenced in all animals except in one animal slaughtered at 65.5 h. The ovulation rate varied from 4 to 50 ovulations. Approximately 80% of large follicles (> 10 mm diameter) had ovulated within 12 h of the onset of ovulation. Onset of ovulation was followed by a dramatic decrease in the number of large follicles (> 10 mm) and an increase in the number of small follicles (相似文献