Ultrasonographic determination of ovarian follicular. Development in superovulated heifers pretreated with FSH-P at the beginning of the estrous cycle

On Day 3 of the estrous cycle (estrus = Day 0), dairy heifers were given either 10 mg i.m. FSH-P (FSH-P primed; n = 9) or a saline vehicle (saline primed; n = 9). On Day 10, all heifers were superovulated with FSH-P (total = 27.7 mg i.m.) in declining doses over 5 d. Heifers were inseminated artificially at estrus. From Day 2 until estrus, the number and size of follicles >2 mm were monitored daily by ultrasonography. The mean (+/- SEM) number of corpora lutea (CL) (6.2 +/- 1.5 vs 10.7 +/- 0.9; P<0.05) and the mean number of recovered embryos and unfertilized ova (3.6 +/- 1.7 vs 8.4 +/- 2.2; P<0.05) were lower in FSH-P-primed than in saline-primed heifers. Prior to initiation of superovulation, follicles >10 mm appeared on Days 6 to 7 in saline-primed heifers but only on Days 8 to 10 in FSH-P-primed heifers (P<0.05). Also, until Day 10, the mean number of follicles 4 to 6 mm and 7 to 10 mm was higher (P<0.05) in FSH-P-primed than in saline-primed heifers. After initiation of the superovulatory treatment (Day 10 to estrus), saline-primed heifers had a greater and faster increase in the mean number of follicles >10 mm (P<0.02) than FSH-P-primed heifers did. Depletion in the number of follicles 2 to 3 mm (P<0.001) between Day 10 and estrus and in the number of follicles 4 to 6 mm (P<0.05) between Day 12 and estrus occurred in both groups of heifers. Decreased superovulatory response and embryo recovery in FSH-P-primed heifers may have been due to the presence of large follicles (>10 mm) prior to the initiation of the superovulatory treatment which reduced the ability of small follicles to grow into larger size classes during superovulatory treatment.     

Effects of FSH commercial preparation and follicular status on follicular growth and superovulatory response in Spanish Merino ewes

Ovarian follicular development was characterized in 24 Spanish Merino ewes to study effects of the follicular status and the FSH commercial product used on follicular growth and subsequent superovulatory response. Estrus was synchronized using 40 mg fluorogestone acetate sponges. The superovulatory treatment consisted in 2 daily i.m. injections of FSH from 48 h before to 12 h after sponge removal. Sheep were assigned randomly to 2 groups treated with 6 decreasing doses (4, 4, 3, 3, 2, 2 mg) of FSH-P or with 6 doses of 1.25 mL of OVAGEN. Growth and regression of all follicles > or = 2 mm were observed by transrectal ultrasonography, and recorded daily from Day 6 before sponge insertion to the first FSH injection, and then twice daily until estrus was detected with vasectomized rams. Differences were detected in follicular development from the first FSH injection to detection of estrus (-48 to 36 h from sponge removal) between groups. Administration of FSH-P increased the appearance of new follicles with respect to OVAGEN (6.3 +/- 0.7 vs 4.8 +/- 0.4; P < 0.05), and the mean number of medium (4 to 5 mm) follicles (8.9 +/- 1.2 vs 6.6 +/- 0.9; P < 0.05). However, the mean number of follicles that regressed in size after sponge removal (5.9 +/- 0.4 vs 3.3 +/- 0.4) and the number of preovulatory sized follicles that did not ovulate (60 vs 42.4%) were also higher in FSH-P treated ewes (P < 0.05). So, finally, there were no differences in ovulation rate, as determined by laparoscopy on Day 7 after sponge removal, between ewes treated with FSH-P or OVAGEN (6.3 +/- 1.9 vs 7.0 +/- 1.7 CL). In all the ewes, the ovulatory response was related (P < 0.05) both to the number of small follicles (2 to 3 mm in diameter) present in the ovaries at the start of treatment with exogenous FSH and to the number of follicles that reached > or = 4 mm in size at estrus, despite differences in the pattern of follicular development when using different commercial products.     

Human menopausal gonadotropin induces ovulation in sheep, but embryo recovery after prostaglandin F2alpha synchronization is compromised by premature luteal regression

Using pregnant mares' serum gonadotropin (PMSG) and follicle stimulating hormone (FSH-P) as conventional gonadotropins, human menopausal gonadotropin (hMG) was tested for its comparative ability to induce multiple ovulations in sheep. Estrous cycles were synchronized using either prostaglandin F(2alpha) (PGF(2alpha)) or progestogen (MAP)-impregnated pessaries. During the mid-luteal phase, control ewes received serial saline injections, whereas test females (which also served as embryo donors) received either a single PMSG injection (1200 IU) or serial injections of FSH-P (total, 21 mg) or hMG (total, 1350 IU) over 3.5 d. These sheep were naturally mated and artificially inseminated (AI) in utero . Number of CL and transferable-quality embryos 5 d after AI was greater (P<0.05) in FSH-P-and hMG-treated donors than in PMSG-treated ewes. The lower number of transferable-quality embryos produced by PMSG-treated donors was attributed to a reduced (P<0.05) fertilization rate compared with that of the other treatment groups. There were no differences (P>0.05) in daily circulating estradiol-17beta and progesterone concentrations among the gonadotropin treatment groups. Gonadotropin-treated ewes demonstrated estrus approximately 24 h earlier than control ewes and, therefore, exhibited an accelerated estradiol-17beta surge and rise in circulating progesterone. Progesterone production in gonadotropin-treated ewes was also more variable than in the controls; this was due, in part, to premature luteal regression which occurred in 4 of 10 PMSG-, 3 of 10 FSH-P- and 6 of 10 hMG-treated ewes also given PGF(2alpha). Ewes with prematurely regressing CL experienced transient luteal tissue development within 4 d of ovulation and produced no embryos. Overall results 1) demonstrate that serial administration of hMG induces multiple ovulations in sheep comparable to FSH-P, and 2) suggest that PGF(2alpha) treatment during ovulation induction adversely affects newly formed luteal tissue compromising subsequent embryo recovery.     

Interrelationships of hormonal and ovarian responses in superovulated heifers pretreated with FSH-P at the beginning of the estrous cycle

The objective of this study was to determine the relationships between follicle stimulating hormone, (FSH), estradiol (E(2)), and progesterone (P(4)) concentrations in peripheral blood samples and the follicular dynamics prior to and during superovulation in heifers pretreated with FSH-P (10 mg, i.m.) (FSH-P-primed; n=9) or not (saline-primed; n=9) on Day 3 (Day 0 = estrus) of the estrous cycle. On Day 10, all heifers were superovulated with FSH-P (27.7 mg i.m.) in declining dosages over 5 days. Prior to and during superovulation, blood samples were collected one to five times daily, and the follicular dynamics were monitored daily by ultrasonography. Prior to superovulation, profiles of P(4) and E(2) did not differ (P>1) between the saline- and FSH-P-primed heifers. The FSH concentrations in saline-primed heifers decreased from 0.43 +/- 0.05 ng/ml to 0.30 +/- 0.04 ng/ml between Days 3 and 7 and then increased progressively to 0.59 +/- 0.04 ng/ml on Day 10. In contrast (P<0.002), FSH concentrations in the FSH-P-primed heifers remained constant between Days 3 and 10 and averaged 0.41 +/- 0.03 ng/ml. Higher increases in E(2) during superovulation (maximum values, 100 vs 46 pg/ml) and in P(4) after superovulation (maximum values, 39 vs 22 ng/ml) in the saline-than in the FSH-P-primed heifers reflected the greater increase in the number of follicles (>10 mm) and in the number of corpora lutea (CL) in the saline-primed heifers. Prior to the preovulatory luteinizing hormone (LH) peak during superovulation, there was a parallel (P>0.1) decrease in FSH concentrations in the saline- and FSH-P-primed groups. Within heifers partial correlations indicated that E(2) was correlated positively with the number of follicles (>/= 7 mm) and the size of the largest follicle during superovulation (r=0.54 to 0.81; P<0.01). Negative correlations were detected (P<0.01) between FSH and the number of follicles >/=7 mm prior to (r=-0.26) and during superovulation (r=-0.37). The results cofirm earlier reports indicating that priming with FSH-P decreases the superovulatory response in cattle. Interrelationships of hormonal and ovarian responses support the concept that the presence of large dominant follicles prior to superovulation limits the superovulatory response.     

Studies of FSH-P induced follicular growth in cows

Because cow ovaries do not contain a dominant follicle before Day 3 of the estrous cycle, we hypothesized that gonadotropin treatment early in the estrous cycle would induce growth of multiple follicles and could be used to induce superovulation. In Experiment 1, when 16 cows were treated with FSH-P beginning on Day 2 of the estrous cycle and were slaughtered on Day 5, all cows responded to gonadotropin treatment by exhibiting a large number ( approximately 19) of estrogenactive follicles >/= 6 mm. In Experiment 2, in response to FSH-P treatment from Day 2 to Day 7, and fenprostalene treatment on Day 6, 11 of 15 cows exhibited estrus and had a mean ovulation rate of 23.7 +/- 1.5. In Experiment 3, an FSH-P treatment regimen identical to that used in Experiment 2 was administered to cows beginning either on Day 2 (Day-2 cows; n=14) or Day 10 (Day-10 cows; n=11) of the estrous cycle. Twelve of 14 Day-2 cows and all Day-10 cows exhibited estrus after fenprostalene treatment. Day-2 cows exhibited 34.3 +/- 7.0 ovulations, which was less (P < 0.05) than that exhibited by Day-10 cows (48.3 +/- 4.4). However, the proportion of embryos recovered per corpus luteum was about 2-fold greater (P < 0.05) for Day-2 cows than for Day-10 cows (0.49 +/- 0.08 vs 0.27 +/- 0.06). These data indicate that beginning gonadotropin treatment early in the estrous cycle, when a dominant follicle is not present, provides an efficacious means to induce growth of multiple follicles and superovulation in cows. However, when FSH was administered for 6 d, beginning the treatment on Day 10 also resulted in a consistent and efficacious response.     

Transfer of superovulated sheep embryos obtained with different FSH-P

Embryo transfer is one way of accelerating genetic improvement in sheep. One of the main obstacles has been the production of good-quality embryos. The use of progestagens and the stimulation of ovulation with follicle stimulating hormone pituitary extract (FSH-P) has permitted the superovulation of donor and recipient ewes and the synchronization of their cycles. The injection of 16 mg FSH-P at the end of progestin treatment gave means of 9 +/- 1.5, 12 +/- 1.5, and 19.5 +/- 2.6 corpora lutea per ewes in the Préalpes, Lacaune, and Romanov x Préalpes breeds respectively (this last breed is particularly prolific). Twenty Préalpes donor ewes produced 133 embryos that were recovered surgically at Day 6 of gestation; of these, 99 morulae were transferable. Forty-five morulae transferred surgically into 24 Préalpes recipient ewes yielded 16 pregnant ewes and 27 lambs (1.7 per ewe). Twenty-two Lacaune ewes yielded 204 embryos, of which 152 morulae were transferable. Of 76 recipients, 58 became pregnant and gave birth to 97 lambs (1.7 per ewe). During anoestrus, the mean ovulation rate decreased from 11.2 to 8.4; 40.6% of the embryos recovered were of transferable quality versus 74.5% during the normal breeding season. An improved superovulation technique, based on the use of FSH-P with a known follicle stimulating hormone to luteinizing hormonal (FSH/LH) ratio, provided us with good-quality embryos. This treatment must be adapted to the season.     

Influence of maternal environment on the number of transferable embryos obtained in response to superovulatory FSH treatments in ewes

In a first experiment, embryo viability was estimated after recovery in the uterus or the oviduct of 70 Manchega ewes following a treatment of superovulation with decreasing doses of OVAGEN. Fewer viable embryos (5.6 +/- 0.9 vs. 8.3 +/- 0.8, P < 0.05) and more degenerative embryos (31.3% vs. 6.8%, P < 0.005) were obtained from the uterus than from the oviduct respectively. In a second experiment performed on 14 ewes, embryo viability was analyzed in relation to the follicular population estimated by ultrasonography (follicles > or = 2 mm) at the first FSH administration. Progesterone (P4) and oestradiol 17beta (E2) concentrations were also determined from the beginning of the superovulation treatment to the recovery of the embryos. The number of viable embryos (4.3 +/- 1.4) was positively correlated (r = 0.824) with of 2-4 mm diameter follicles (P < 0.05), and with E2 concentrations at -12 h (r = 0.891, P < 0.01) , 0 h (r = 0.943, P < 0.0001) and +24 h (r = 0.948, P < 0.05) from estrus detection. Prolonged high levels of E2 up to 72 h with low levels of P4 on days 3 and 4 after estrus had a negative (P < 0.05) effect on embryo viability. These results indicate that ovarian response to superovulatory protocols is related to the individual variations in the number of follicles of 2-4 mm at the start of FSH treatment, and that embryo viability is conditioned by the steroid patterns during the time spent in the genital tract of the super-ovulated ewes.     

Effect of season and duration of FSH treatment on embryo production in sheep

Thirty-six mature Manchega ewes were used in two experiments to determine the effect of season and of 2- or 3-d FSHp treatment on the ovulation rate and number of transferable embryos produced. During the breeding season, estrus was synchronized with FGA (30 mg for 13 d). Begining 48 or 24 h before sponge removal, each ewe received two daily injections of 4-4-3-3-1-1 or 5-5-3-3 mg of FSHp. Concurrently with the two last injections both groups were administered 100 mug of LH. Ewes were tested for estrus and 6 or 7 d later were laparotomized and surgically flushed to recover embryos. The number of corpora lutea (CL), the total number of embryos and of viable embryos were recorded. Six months later (nonbreeding season) the design was repeated, with each ewe receiving the opposite treatment to that received in the fall. Response in ovulation rate and number of viable embryos did not differ between seasons. Mean (SEM) numbers of observed CL and embryos recovered were higher (P<0.001) with the 3-d treatment (8.7+/-5.8 and 7+/-4.8) than with the 2-d treatment (5.8+/-3.2 and 4.4+/-3) when pooled over the two seasons. The mean number of transferable embryos was higher (P<0.01) with the 3-d (4.2+/-3.9) than with the 2-d treatment (2.5+/-2.3).     

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 effects of previous ovarian status on ovulation rate and early embryo development in response to superovulatory FSH treatments in sheep

A total of 64 ewes was used to determine if the changes in superovulatory yields related to the ovarian status at the start of superovulatory treatment are due to differences in the population of gonadotrophin-responsive follicles, alterations in the processes of ovulation or transport of embryos from oviduct to uterus and/or developmental competence of the oocyte/embryo. Ovarian status at the start of a superovulatory FSH step-down treatment, administered coincidentally with a progestagen, was assessed by ultrasonography. On Day 4 after progestagen withdrawal, embryos were recovered from oviduct and their viability was determined by assessing development in vitro culture (IVC) until the hatched blastocyst stage. In all the ewes, the ovulation rate was related positively to the number of 2-3 mm follicles at first FSH injection (P<0.005). However, the total number of embryos and their viability were related to the more limited category of 3 mm follicles (P<0.05), whereas a higher degeneration rate was related to the number of 2mm follicles. The presence of a corpus luteum (CL) at the start of superovulatory treatment exerted a protective effect on embryonic viability, decreasing the degeneration of embryos. On the other hand, the presence of a dominant follicle at first FSH dose affected the mean size of the pool of follicles responding to the superovulation treatment, because ovulation arose from 3 to 5 mm follicles in absence of large follicles (P<0.05), but from 2 to 3 mm follicles when large follicles were present (P<0.005), indicating atresia in medium sized follicles in the presence of a large follicle.     

Induction of the presence of corpus luteum during superovulatory treatments enhances in vivo and in vitro blastocysts output in sheep

This report offers the results of two experiments developed to test possible benefitial effects of the presence of corpus luteum (CL) on in vivo and in vitro sheep embryo production; using two different breeds treated with two different protocols by two different teams at two different centres. In the first trial, estrus was synchronized in 11 ewes with two doses of cloprostenol, 10 days apart. On day 1 after estimated ovulation, sheep were treated with progestagen sponges and superovulated with eight decreasing doses (26.4 units NIH-FSH-S1 x 3, 22.0 units x 2, and 17.6 units x 3) of ovine FSH injected twice daily. Ovulation rate and number of embryos obtained in vivo were compared to those from 12 control ewes without cloprostenol treatment. Presence of a CL improves the number of transferable embryos (7.4+/-0.6 versus 4.1+/-0.6 in control ewes, P < 0.05). The second trial investigated the effects of the presence of CL on embryos produced in vitro from six ewes bearing CL and six ewes without CL at start a superovulatory treatment consisting of 96 units of ovine FSH administered in four equal doses given every 12 h. There were not detected effects of the CL on the number and size of follicles or in the number, morphology and ability to resume meiosis of their oocytes. However, oocytes from ewes with CL showed higher rates of fertilization (73.5 versus 45.5%, P < 0.005), higher development to blastocyst (35.8 versus 19.3%, P < 0.01) and higher hatching rates after vitrification (80.0 versus 25.0%, P < 0.05).     

Ovarian response to gonadotropin treatment initiated relative to wave emergence in ultrasonographically monitored ewes

Follicular recruitment and luteal response to superovulatory treatment initiated relative to the status of the first wave of the ovine estrous cycle (Wave 1) were studied. All ewes (n = 25) received an intravaginal progestagen sponge to synchronize estrous cycles, and ewes were monitored daily by transrectal ultrasonography. Multiple-dose FSH treatment (total dose = 100 mg NIH-FSH-P1) was initiated on the day of ovulation (Day 0 group) in 16 ewes. In the remaining 9 ewes, FSH treatment was started 3 d after emergence of the largest follicle of Wave 1 (Day 3 group). Ewes received PGF(2alpha) with the last 2 FSH treatments to induce luteolysis. Daily blood samples were taken to determine progesterone profiles and to evaluate the luteal response subsequent to superovulation. The ovulation rate was determined by ultrasonography and correlated with direct observation of the ovaries during laparotomy 5 to 6 d after superovulatory estrus when the uterus was flushed to collect embryos. Results confirmed that follicular recruitment was suppressed by the presence of a large, growing follicle. In the Day 0 and Day 3 groups, respectively, mean numbers (+/- SEM) of large follicles (>/= 4 mm) recruited were 6.4 +/- 0.6 and 2.7 +/- 0.7 (P < 0.01) at 48 h after the onset of treatment, and 6.7 +/- 0.5 and 5.1 +/- 0.6 (P = 0.08) at 72 h after the onset of treatment. Ovulation rates were 5.6 +/- 0.8 and 3.3 +/- 0.8 in the respective groups (P < 0.05). The number of transferable embryos was 1.8 +/- 0.5 and 0.3 +/- 0.2 in the respective groups (P < 0.05). Short luteal phases (相似文献   

Follicular dynamics and superovulatory response in Holstein cows treated with FSH-P in different endocrine states

The effect of follicular and/or endocrine environments on superovulatory response was tested. Eighteen nonlactating Holstein cows were superovulated with 32 mg FSH-P given in decreasing doses at 12-h intervals plus two injections of prostaglandin F2-alpha (25 mg each) on the third day of treatment. Cows were assigned randomly to treatments: T1, superovulatory treatment initiated on estrous cycle Day 10.5; T2, CIDR (intravaginal device containing 1.9 g of progesterone) inserted from Days 3 to 9 and superovulation initiated on Day 6.5; T3, identical to T2 but Buserelin (GnRH agonist) was injected (8 mug, i.m.) on Day 3 at the time of CIDR insertion. Embryos were recovered on Day 7 after the superovulatory estrus. Cows were examined daily by ultrasonography and blood was collected for progesterone and estradiol determinations. Mean diameter of the dominant follicle (frequency of first-wave dominant follicle) at the beginning of FSH injections was 13.7 mm (4 6 ), 11.2 mm (6 6 ) and 8.7 mm (6 6 ) (P<0.01) for T1, T2 and T3, respectively. Following initiation of superovulation, follicles moved into larger follicle classes (Class I, <3 mm; Class II, 3 to 4 mm; Class III, 5 to 9 mm; Class IV >9 mm) earliest in T1 (P<0.01). Cumulative follicular diameter and plasma concentrations of estradiol at Day 4 of superovulation were higher (P<0.01) in T1 (200 mm, 82 pg/ml) compared with T2 (123 mm, 24 pg/ml) and T3 (130 mm, 18 pg/ml). Proportion of cows in estrus prior to 12 h vs 12 to 24 h differed (P<0.05) between groups (T1: 5 vs 1; T2: 2 vs 4; T3: 1 vs 5). Mean number of follicles on the last day of superovulation treatment, number of CL and number of embryos plus unfertilized ova recovered were 17.5, 12.2 and 13.3; 13.8, 10 and 8.2 (P<0.1) and 8.7, 4.5 and 2.3 (P<0.05) for T1, T2 and T3, respectively. The developmental stage of the dominant follicle was associated with not only the number of ovulations, but also the size and periestrous concentrations of plasma estradiol associated with the recruited follicles.     

A dominant follicle does not affect follicular recruitment by superovulatory doses of FSH in cattle but can inhibit ovulation

It has been suggested that superovulation in cattle is impaired if FSH injections are initiated in the presence of a dominant follicle, but the results of experiments to test this hypothesis have been contradictory. However, previous experiments were conducted during mid-cycle, when the absence or presence of a dominant follicle is difficult to assess. We took a different approach by comparing the effects of initiating superovulatory injections of FSH (11 equal doses of FSH-P, every 12 h) on Day 1 of the bovine estrous cycle, when a dominant follicle clearly is not present, vs initiation on Day 6, when a dominant follicle clearly is present and actively growing (n = 17 heifers in a "crossover" design). In 8 17 heifers initiation of FSH injections in the presence of a dominant follicle (Day 6 group) caused ovulation of the dominant follicle within 1 to 2 days and formation of a smaller than normal CL. These animals had higher than normal concentrations of plasma progesterone around the time of expected estrus (P < 0.05) and failed to exhibit estrus. Although the mean number and diameter of the follicles recruited in response to FSH injections in heifers that ovulated the dominant follicle prematurely were not different from the other heifers in the Day 6 group, no ovulations were observed, and no embryos or ova were recovered 6 d after insemination. Conversely, when FSH injections were initiated on Day 1 in these 8 heifers, they exhibited estrus, and their plasma progesterone around the time of estrus, mean ovulation rate, and number of total and transferable embryos recovered did not differ from the responses observed in the remaining 9 heifers treated either on Day 1 or on Day 6. Taken together, these results indicate that a dominant follicle does not affect the ability of smaller follicles to be recruited in response to exogenous FSH, but may impair their ovulation. These findings provide an explanation for previous reports of decreased superovulatory responses during times of the cycle when a dominant follicle would be expected to be present.     

Superovulation and embryo quality in beef cows using PMSG and a monoclonal anti-PMSG

The long half-life of pregnant mare serum gonadotrophin (PMSG) reduces its application in the superovulation of cattle; thus, a monoclonal antibody to PMSG (anti-PMSG) was administered at the onset of estrus to increase the number of transferable embryos. Angus, Hereford and Angus x Hereford cows (n = 149) 3 to 9 yr old were assigned randomly to one of three dosages of PMSG (1500, 3000 or 6000 IU) with or without an equivalent dosage of anti-PMSG. Embryos were collected nonsurgically on Day 8 (estrus = Day 0), and all cows were ovariectomized on Day 9. The percentage of cows exhibiting estrus and ovulating decreased (P<0.05) with an increasing dosage of PMSG (82, 76 and 44% for 1500, 3000 and 6000 IU, respectively). Ovarian and total corpora lutea (CL) weight increased (P<0.001) linearly as PMSG dosage increased, but were reduced (P<0.001) curvilinearly by anti-PMSG, resulting in a PMSG by anti-PMSG interaction (P<0.001); the interaction was also significant (P<0.05) for ovulation rate (14.0 vs 14.3, 21.5 vs 24.4 and 29.2 vs 6.6 CL for 1500, 3000 and 6000 IU PMSG, without vs with anti-PMSG, respectively). Anti-PMSG increased (P<0.001) the number of small ovarian follicles (1 to 3 mm diameter) and decreased (P<0.001) the number of large follicles (>10 mm) at ovariectomy; the number of large follicles increased (P<0.001) with PMSG dosage. The number of total and transferable embryos recovered did not differ among PMSG and anti-PMSG dosages; however, the percentage of transferable embryos decreased (P<0.01) with increasing PMSG dosage. In general, neither PMSG dosage nor anti-PMSG influenced embryo quality.     

Superovulation of a low fecundity sheep breed using a porcine gonadotrophin extract with a defined LH content (Pluset)

The aim of this study was to determine the efficiency of a porcine pituitary gonadotrophin extract with a defined pLH content in the superovulation of sheep. Estrus was synchronized in 61 Polish Mountain ewes with intravaginal fluorogestone acetate sponges. Twenty-four hours before the sponges were removed, the ewes underwent different superovulatory treatments: Group I 250 IU of pFSH with 250 IU of pLH (n=19); Group II 500 IU of pFSH with 500 IU of pLH (n=19); and Group III 750 IU of pFSH and 750 IU of pLH (n=18). Gonadotrophine was administered intramuscularly twice a day over a 3-day period in decreasing dosages. A control group of ewes (n=5) was treated with saline. In most of the ewes estrus began about 20 hours after sponges were removed. All the ewes were bred naturally every 12 hours. Superovulation was confirmed in 75% of the treated animals. The ewes receiving 250 IU each of pFSH and pLH produced an average of 7.6 +/- 3.1 corpora lutea (CL), 6.3 +/- 2.4 ova and 4.3 +/- 4.1 transferable embryos. Group II (500 IU of pFSH and pLH) produced 8.5 +/- 4.0 CL, 7.6 +/- 4.1 ova, and 4.1 +/- 2.9 transferable embryos. Group III (750 IU each of pFSH and pLH) produced 8.3 +/- 5.2 CL, 7.5 +/- 5.5 ova and 5.2 +/- 5.1 transferable embryos. The mean embryo recovery rate was 87% for all three groups. Differences in superovulatory response and embryo recovery rate among the groups were not statistically significant (P>0.05).     

Effects of follicular status at treatment on follicular development and ovulation in response to FSH in Spanish merino ewes

Cyclic Spanish Merino ewes were treated on Day 13 of the estrous cycle with 12 mg, i.m., FSH-P in saline (n = 9) or propylene glycol (n = 24), currently with 100 micrograms, i.m., Cloprostenol (Day 0). From Day-6 to Day 0, the ewes were observed daily by transrectal ultrasonography, after Day 0, ultrasonography was performed every 12 h for 72 h. Sizes and locations of > or = 2 mm follicles were recorded at each observation. The ovulation rate was determined by laparoscopy on Day 7 after estrus. The number of ovulations ranged from 0 to 6 in ewes treated with FSH-P in saline and from 0 to 16 in ewes receiving FSH-P in propylene glycol (P < 0.05). In the latter group, the response was bimodally distributed; about half of the females had 1 ovulation, whereas the remainder had > 4 with a mean of 7 ovulations. The ovulation rate was associated with 2 characteristics of the largest follicle present at treatment (Day 0). First, if the largest follicle on Day 0 had not changed in diameter from Day-1 to Day 0, then 7 of 9 ewes had > 3 ovulations; if the largest follicle had either increased or decreased, only 8 of 24 ewes had > 3 ovulations (P < 0.05). Second, there was a linear trend (P < 0.07) for ovulation rate to decrease as the persistence of the largest follicle at treatment increased; no ewe in which the largest follicle on Day 0 remained present for more than 36 h ovulated more than 6 follicles. As with the ovulation rate, the numbers of large follicles on Days 1.5, 2 and 2.5 varied with the interaction of change in diameter of the largest follicle on Day 0 from Day-1 to Day 0 and with vehicle. In summary, the superovulatory response was affected by the change in diameter from Day-1 to Day 0 of the largest follicle on Day 0 and the period required for that follicle to regress after treatment with FSH-P and cloprostenol.     

Ovulation of aged follicles does not affect embryo quality or fertility after a 14-day progestagen estrus synchronization protocol in ewes

The aim was to examine the effect of ovulation of aged follicles on embryo quality and fertility in ewes. In Experiment 1, ewes (n = 39) received a prostaglandin analogue on Day 6 of the cycle and then received either a progestagen sponge from Day 6 to 20 after estrus (Single sponge) or a progestagen sponge on Day 6 that was replaced on Day 11 and 16 and removed on Day 20 (Multiple sponges). In a subgroup of ewes, the growth of ovarian follicles was characterised using ultrasonography. Fertile rams were introduced 48 hours after sponge withdrawal; we slaughtered the ewes on Day 5 of pregnancy and recovered the embryos. The mean age of the ovulatory follicles was greater in ewes that received a single sponge compared with multiple sponges (8.7+/-0.8 days, range 4 to 14, versus 4.5+/-0.7 days, range 3 to 6; P<0.05). However, the groups did not differ (P>0.05) in ovulation rate (2.4+/-0.3 corporal lutea per ewe) or the proportion of good quality embryos recovered (71 to 82%; developed to the early morula stage or further). In Experiment 2, ewes (570 in total) received treatments similar to those in Experiment 1 but were kept until lambing. Ewes that received a single sponge came into heat earlier (P<0.05) than those that received multiple sponges, but > or = 97% of ewes in all groups (P>0.05) were bred by 48 to 72 hours after ram introduction. There was no difference (P>0.05) between groups for the proportion of ewes that lambed to first service (80 to 86%) or the number of lambs per ewe (1.94+/-0.08 lambs). We conclude that when luteolysis occurs at the beginning of progestagen synchronisation, ewes will ovulate aged follicles, but that compared to shorter duration follicles, these follicles produce oocytes that are equally competent to be fertilised and develop into good quality embryos and full-term lambs.     

Superovulatory and endocrine responses in heifers treated with FSH-P at different stages of the estrous cycle

Forty-two Holstein heifers were superovulated with FSH-P (total dose, 30 mg) and cloprostenol. Treatment was initiated on Day 3 (Group D3, n = 11), Day 6 (Group D6, n = 11), Day 9 (Group D9, n = 10) or Day 12 (Group D12, n = 10) of the estrous cycle. Heifers were bled daily for serum progesterone and estradiol-17beta determinations and every 6 h for a 48-h duration at the expected time of estrus for luteinizing hormone (LH) assay. Ova and embryos were flushed from the reproductive tracts and the number of corpora lutea (CL) were recorded after slaughter on Day 7 post-estrus. Mean (+/- SEM) numbers of observed CL were higher (P < 0.05) in Group D9 (33.3 +/- 4.8) than in Group D3 (15.3 +/- 3.8), with Group D6 (17.0 +/- 2.9) and Group D12 (23.9 +/- 7.3) being intermediate. Similarly, mean (+/- SEM) numbers of fertilized embryos were highest (P < 0.05) in Group D9 (13.3 +/- 2.2). There was also a nonsignificant trend for the number of transferable embryos to be greatest in Group D9. Neither serum progesterone concentrations 3 d after the LH peak nor peak serum estradiol 17beta concentrations differed among groups, but both were significantly correlated with numbers of observed CL and total ova and embryos.     

Effect of follicular status on superovulatory response in ewes is influenced by presence of corpus luteum at first FSH dose

The present study was developed to assess possible effects on ovulatory response and embryo yields arising from the presence of a corpus luteum (CL) at the time of initiation of the progestagen treatment used in superovulatory protocols in sheep. In breeding season, estrus was synchronized in 25 Manchega ewes using 40 mg FGA sponges for 14 days, together with a single dose of 125 microg of cloprostenol on Day 12, with Day 0 as day of progestagen insertion. Superovulatory treatment consisted of eight decreasing doses (1.5 x 3 ml, 1.25 x 2 ml, and 1 x 3 ml) of Ovagen twice daily from 60 h before to 24 h after sponge removal. The presence or absence of corpora lutea was assessed by transrectal ultrasonography at progestagen insertion and at first FSH dose. Number and size of all follicles > or = 2 mm were also evaluated at first FSH dose. The number of corpora lutea and the number and viability of recovered embryos in response to the treatment were evaluated 7 days after sponge removal. No significant effect on ovarian response of the presence of a CL at sponge insertion in 21 of the 25 ewes (84%) was detected. However, ewes with a CL at first FSH dose (16 ewes, 64%) yielded a higher number of transferable embryos (7.2 +/- 1.4 versus 2.7 +/- 0.7, P < 0.05), since the embryo degeneration rate was increased in sheep without a CL (42.5% versus 12.7%, P < 0.01). Analysis of possible effects derived from the presence of a large presumptively dominant follicle (> or = 6 mm) at first FSH dose showed that both recovery and viability rates were lowest (P < 0.05) in ewes bearing a large follicle in the absence of a CL (40.5 and 50.6%, respectively), and highest in ewes that did not show a large follicle but in which a CL was present (73.9 and 85.2%). The final number of transferable embryos was very different between groups (10.2 versus 1.8, P < 0.01). These results indicate that the number and quality of embryos obtained from superovulated ewes is affected by the presence of a CL prior to the first FSH dose (i.e. by the stage of the estrous cycle at progestagen insertion) and also by an interaction with suppressive effects from large dominant follicles. This finding suggests the existence of some effects on follicular population prior to the FSH treatment that may compromise follicle and oocyte developmental competence. It seems reasonable to hypothesize that superovulatory yields would be increased by beginning the treatment during the early-luteal phase of the estrous cycle, allowing for the presence of a CL along with the progestagen treatment.