Superovulatory response of Chios sheep to PMSG during spring and autumn

To study the superovulatory response of Chios sheep to pregnant mares' serum gonadotrophin (PMSG), two experiments were carried out; one in spring and one in autumn. Four doses of PMSG (1500 IU, Group 1; 1000 IU, Group 2; 750 IU, Group 3; 500 IU, Group 4; controls, Group 5) were tested on 46 ewes. Oestrus was synchronised by means of MAP intravaginal sponges and PMSG was injected i.m. at the time of sponge withdrawal. When in oestrus, ewes were naturally mated. On Day 7 after sponge removal, mid-ventral laparotomy was performed and the uterine horns and/or oviducts were flushed with 20–40 ml Dulbecco's phosphate-buffered saline supplemented with 15% foetal bovine serum (FBS). The embryos were examined under a dissecting microscope and were evaluated according to morphological criteria.The interval from sponge removal to the onset of oestrus was significantly (P < 0.001) shorter in autumn than in spring in all groups. No significant differences regarding superovulatory response, collection and fertilisation rate or numbers of ova and embryos collected were found between spring and autumn. The clinical signs of oestrus started earlier (P < 0.001) in all PMSG treated animals than in the controls, both in spring and in autumn. The highest ovulation rate was recorded in Group 2 (5.9 ± 1.0), followed by Groups 1 (5.0±0.9), 3 (3.9±0.5), 4 (26±0.4) and 5 (1.3±0.1). The increase observed in total ovarian response (corpora lutea + large anovulated follicles) parallelled the increase of PMSG dose (10.7 ± 1.6, 7.7 ± 0.9, 4.5 ± 0.6, 3.4 ± 0.5 and 1.8 ± 0.2 for Groups 1, 2, 3, 4 and 5, respectively). The highest mean number of ova was collected from Group 3 (3.4±0.5), followed by Groups 2 (2.6 ± 0.4), 4 (2.2 ± 0.3), 1 (1.6 ± 0.5) and 5 (1.1 ± 0.1). The higher doses of PMSG (1500 and 1000 IU) significantly increased the mean number of anovulated follicles and significantly decreased recovery rate. Mean number of high viability embryos collected per ewe treated (0.9 ± 0.6, 1.5 ± 0.4, 2.2 ± 0.5, 1.5 ± 0.4, 0.9 ± 0.1 for Groups 1, 2, 3, 4 and 5, respectively) was not improved by PMSG dose.It is concluded that Chios sheep can be superovulated in autumn and in spring with similar results. Clinical signs of oestrus are initiated earlier in autumn than in spring. PMSG treatment shortens the interval from sponge removal to the onset of oestrus. Although PMSG does not seem to be the most suitable hormone for the induction of superovulation in Chios sheep, a dose of 750–1000 IU PMSG gives satisfactory results; higher doses are associated with side effects in a significant number of animals (many anovulated follicles, low recovery rate).     

Relationship of length of uterus in prepubertal pigs and number of corpora lutea and fetuses at 30 days of gestation

Relationships of the length of the uterus at one reproductive stage to the length at other stages and the effect on potential litter size were determined. In Experiment 1, the length of the uterus was measured in situ at 20, 60, or 100 days of age at laparotomy with 20 gilts in each of the three age groups. Forty days after the initial measurement, the uterus was again measured in situ, gilts were ovariectomized and hysterectomized, and associations among uterine measurements at the two different stages were determined. Correlations between uterine length in situ and between uterine length and weight 40 days later were all greater than 0.75 (P < 0.001). The length of one uterine horn increased from 13.9 cm at 20 days of age to 36.7 cm at 140 days of age (P < 0.01). In Experiment 2, 66 gilts were unilaterally hysterectomized and ovariectomized (UHOX) at 150 days of age and the ovary was weighed. Length of the one horn was measured in 36 gilts. At 10 days after first estrus, the length of the remaining uterine horn was measured at laparotomy and corpora lutea were counted in 53 gilts. In 15 of the 53 gilts the remaining uterine horn was removed to obtain uterine weights. At the second or third estrus, 38 gilts were mated and at Day 30 of gestation, 31 gilts were pregnant. The gilts were killed, and the length of the uterus measured and corpora lutea (CL) and fetuses were counted. The length of one uterine horn at 150 days of age was 70 cm with a range of 47–110 cm. At 10 days after first estrus, length had increased to a mean of 141 cm with a range of 86–194 cm and at 30 days of gestation the mean was 244 cm with a range of 186–311 cm. There were 12.2 CL at first estrus, which was not different from 12.4 CL at the second or third estrus. The mean number of fetuses in one horn at 30 days of gestation was 9.5 with 77% prenatal survival. Length of uterine horn at 150 days of age was correlated with uterine horn length (r = 0.56, P < 0.001) at 10 days after first estrus and number of live fetuses (r=0.39, P<0.05) at 30 days of gestation. At Day 10 after first estrus, uterine length was not correlated with the number of CL, whereas at Day 30 of gestation, the number of CL and uterine length were correlated with the number of live fetuses in those gilts with below the mean number of live fetuses, but not in those gilts with above the mean of live fetuses. The number of live fetuses (r=0.66, P < 0.001) and fetal survival (r=0.63; P < 0.001) were correlated with uterine horn length in pregnant UHOX gilts. Length of the prepubertal uterus gives an indication of postpubertal length and the potential litter size in pigs.     

Use of progestagen-gonadotrophin treatments in estrus synchronization of sheep

The main objective of this study was to investigate the effectiveness of certain progestagen-gonadotrophin treatments on synchronization of estrus in sheep. In Experiment I, 30 Chios ewes were treated at the beginning of the breeding season with medroxyprogesterone acetate (MAP) intravaginal sponges for 12 days and a single i.m. treatment of either FSH (Group 1,10 IU, n = 8; Group 2, 5 IU, n = 8; Group 3, 2.5 IU, n = 8) or eCG (Group 4, 400 IU, n = 6) at the time of sponge removal. Ten days after sponge removal laparotomy was performed to record ovarian response. Clinical estrus was observed in more (though not at a significant level) FSH treated than eCG treated sheep (62.5% versus 33.3%). Administration of 400 IU eCG resulted in the highest mean number of CL perewe ovulating (2.8 +/- 0.2), with administration of 10 IU FSH producing the next best results (2.1 +/- 0.3). Statistically significant differences in the mean number of CL per ewe ovulating were found only between ewes in Group 3 (1.7 +/- 0.4) and Group 4 (2.8 +/- 0.2) (P < 0.05). In Experiment II, 53 Chios and 30 Berrichon ewes were treated during the mid-breeding season with MAP intravaginal sponges for 12 days and a single i.m. treatment of either 10 IU FSH (27 Chios and 16 Berrichon ewes) or 400 IU eCG (26 Chios and 14 Berrichon ewes), at the time of sponge removal. Ewes that were in estrus on Days 2-4 and 19-23 after sponge removal were mated to fertile rams. No significant differences were recorded between treatment or breed groups in the proportions of ewes observed in estrus after treatment. In the Berrichon breed, FSH administration resulted in higher lambing rates (93.8% versus 57.1%, P < 0.05) and higher mean number of lambs born per ewe exposed to rams (1.4 +/- 0.2 versus 0.8 +/- 0.2, P < 0.05) than that of eCG. After treatment with eCG, the mean number of lambs born per ewe exposed to rams was higher in the Chios than the Berrichon breed (1.4 +/- 0.2 versus 0.8 +/- 0.2, P < 0.05). After treatment with FSH, the lambing rate was higher in the Berrichon than the Chios breed (93.8% versus 63.0%, P < 0.05). In conclusion, a single FSH treatment (5 or 10 IU) at the end of progestagen treatment appears to be more effective than eCG for the induction of synchronized estrus in sheep at the beginning of the breeding season, with no cases of abnormal ovarian response observed. During the mid-breeding season FSH (10 IU) appears to be equally as effective as eCG (400 IU) in respect of lambing rate and mean number of lambs born per ewe.     

Ovarian morphology,plasma progesterone concentrations and early embryo survival in the sow

Sixty-five Large White sows were used to examine relationships between ovarian morphology and embryo survival at 30 days gestation and plasma progesterone concentration before and after service.The total mass of luteal tissue was positively correlated with the number of corpora lutea on the ovaries (r = 0.68), and formed a fairly constant proportion of ovarian mass at 30 days gestation. The mean number of embryos per sow was 11.2 ± 0.76, and embryo survival rate was estimated to be 76.5%. There was a positive correlation between ovulation rate and number of embryos at 30 days of pregnancy (r = 0.39). The survival rate of embryos was inversely related (r = ?0.66) to the mean distance between embryos in the uterus. The means of plasma progesterone levels on days 11, 12 and 13 after service were positively correlated with the means of progesterone levels on days 9, 10, 11 and 12 of the cycle before service, the number of corpora lutea, the total mass of luteal tissue and the total mass of the ovaries, but not to numbers of embryos.     

A histological study of corpora lutea from superovulated beef heifers

There is great variability between animals in the number of viable embryos produced following different superovulation regimens. It is not clear if all the follicles that ovulate produce healthy oocytes and form normal corpora lutea (CL) following superovulation. The objective of this study was to assess and compare CL from heifers undergoing three superovulatory regimes with CL from unstimulated heifers on the basis of morphology and morphometric analysis of luteal cells.Beef heifers were superovulated using either: (a) 24 mg porcine follicle stimulating hormone (pFSH) given twice daily over a 4 day period in decreasing doses commencing on day 10 of the oestrous cycle; (b) a single injection of 2000 IU pregnant mare serum gonadotrophin (PMSG) given on day 10 of the cycle; (c) as in (b) but followed by 2000 IU anti-PMSG (IgG to neutralise endogenous PMSG) at the time of the first insemination which was 12–18 h after the onset of oestrus (n = 33 per treatment). Luteolysis was induced 48 h after initial gonadotrophin administration and CL were collected on day 7 of the subsequent cycle and from ten unstimulated heifers (controls) at the same stage of the oestrous cycle. CL morphology was studied at light and electron microscopy levels. Morphometric analysis was performed on luteal cells. Subcellular morphology was similar in heifers from all groups. However, CL from superovulated heifers had more connective tissue than CL from control heifers; the connective tissue content of CL in the anti-PMSG-treated group was particularly marked. Both large and small luteal cells in the heifers receiving anti-PMSG had significantly smaller (P < 0.001) area and sphere volume than similar cells from CL of heifers in the other groups.     

Superovulatory responses to eCG in llamas (Lama glama )

Llamas are copulation-induced single-ovulators, and multiple ovulation and embryo transfer (MOET) methods have not yet been developed for this species. Superovulatory responses to eCG given during an induced (Group A) or simulated (Group B) luteal phase were investigated using ultrasound to observe ovarian follicles and corpora lutea (CLs) and plasma progesterone was used to assess luteal function. Embryos were recovered nonsurgically. Group A (n = 19): donors were given 8 microg, im GnRH analogue (Day 0) to induce ovulation of a mature follicle, 1000 IU, im eCG (Day 7), and 250 microg PGF(2alpha) analogue (Day 9). Group B (n = 17): donors were given a subcutaneous progestagen implant (3 mg Norgestomet) at Days 0 to 7) and 1000 IU, im eCG (Day 5). When most (>65%) of the follicles in both Groups A and B had matured at 5 to 11 d post eCG, the donors were given 8 microg, im GnRH and mated once (n = 26) or twice within a 24-h interval (n = 10); embryos were recovered 6 to 9 d post ovulation. More follicles and corpora lutea were induced in Group B than in Group A, but a similar mean number of embryos were recovered (1.3 vs 1.6), and a similar proportion of donors yielded multiple embryos (35 vs 32%). The embryo recovery rate was similar for Groups A and B (39 and 37%), but it was higher (P < 0.001) with 2 (72%) rather than 1 (22%) mating, and it was negatively correlated with CL number (P < 0.05). Overall, 80% of the llamas had a precocious CL and elevated plasma progesterone concentrations when multiple follicles reached maturity. This was associated with increased subsequent superovulation and embryo recovery (P < 0.01). Peak plasma progesterone was positively correlated with the CL number (P < 0.05). From these results we conclude that superovulation may be achieved with eCG given during either an induced or a simulated luteal phase, that embryo recovery is improved following 2 matings rather than 1, and that MOET may indeed be feasible for use in the llama.     

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.     

Relationships between ovulation rate and embryonic and placental characteristics in multiparous sows at 35 days of pregnancy

The objective of this study was to investigate relationships between ovulation rate (OR) and embryonic and placental development in sows. Topigs Norsvin^R sows (n=91, parity 2 to 17) from three different genetic backgrounds were slaughtered at 35 days of pregnancy and the reproductive tract was collected. The corpora lutea (CL) were counted and the number of vital and non-vital embryos, embryonic spacing (distance between two embryos), implantation length, placental length, placental weight and embryonic weight were assessed. The difference between number of CL and total number of embryos was considered as early embryonic mortality. The number of non-vital embryos was considered as late mortality. Relationships between OR and all other variables were investigated using two models: the first considered parity as class effect (n=91) and the second used a subset of sows with parities 4 to 10 (n=47) to analyse the genetic background as class effect. OR was significantly affected by parity (P<0.0001), but was not affected by the genetic background of the sows. Parity and genetic background did not affect embryonic and placental characteristics at 35 days of pregnancy. OR (varying from 17 to 38 CL) was positively related with early embryonic mortality (β=0.49±0.1 n/ovulations, P<0.0001), with late embryonic mortality or number of non-vital embryos (β=0.24±0.1 n/ovulations, P=0.001) and with the number of vital embryos (β=0.26±0.1 n/ovulations, P=0.01). However, dividing OR in four classes, showed that the number of vital embryos was lowest in OR class 1 (17 to 21 CL), but not different for the other OR classes, suggesting a plateau for number of vital embryos for OR above 22. There was a negative linear relationship between OR and vital embryonic spacing (β=−0.45±0.1 cm/ovulation, P=0.001), implantation length (β=−0.35±0.1 cm/ovulation, P=0.003), placental length (β=−0.38±0.2 cm/ovulation, P=0.05) and empty space around embryonic-placental unit (β=−0.4±0.2 cm/ovulation, P=0.02), indicating uterine crowding. Further analyses showed that effects of OR on embryonic and uterine parameters were related with the increase in late mortality and not early embryonic mortality. Therefore, we conclude that a high OR results in an moderate increase in the number of vital embryos at day 35 of pregnancy, but compromises development in the surviving embryonic/placental units, suggesting that the future growth and survival of the embryos might be further compromised.     

Superovulatory response and quality of embryos recovered from anestrus ewes after a single injection of porcine FSH dissolved in polyvinylpyrrolidone

The effects of a single injection of porcine FSH (pFSH) administered in long acting vehicle on the superovulatory response of milk (Sarda breed) sheep were determined during the anestrous season. The sheep (n=42), synchronized with intravaginal sponges (40 mg fluorogestone acetate -FGA- for 14 d) were submitted 24 h before sponge removal to three different superovulatory treatments. Group 1 (n=16) was treated with a single intramuscular (im) injection of 16 mg of pFHS dissolved in 30 % polyvinylpyrrolidone (PVP); Group 2 (n=12) was injected im with 6, 5, 3 and 2 mg of pFSH every 12 h over 2 d; Group 3 (n=14) was given 800 IU of PMSG and 12 mg of pFSH. All sheep were mated with a fertile ram. Embryos were recovered surgically at Day 7 of sponge removal and graded for the quality according to their morphology. The percentage of good quality embryos recovered was 84% in Group 1, 68% in Group 2 and 77% in Group 3. Data for the onset of estrus, number of corpora lutea (CL), number of unovulated follicles, embryo recovery rate, embryo quality and fertilization rate were recorded for the 3 groups. The onset of estrus, number of CL, number of unovulated follicles, fertilization rate and number of good quality embryos did not differ significantly among the 3 groups. The embryo recovery rate was significantly lower in the group treated with PMSG-FSH (Group 3) than in the 2 other groups. It is concluded that during the anestrous season a single injection im of pFSH results on average in a superovulatory response as good as the more traditional treatments like multiple injections of pFSH and PMSG-pFSH combined.     

Effects of duration and timing of progesterone priming on the incidence of corpora lutea with a normal life-span in Merino ewes induced to ovulate by the introduction of rams

Three experiments investigated the effects of progesterone on the incidence of corpora lutea with normal life-span in seasonally anovulatory Merino ewes induced to ovulate by the introduction of rams.Ewes in Experiments 1 and 2 received a single i.m. injection of 20 mg progesterone in oil at the time of, or up to 5 days before introduction of rams. Control ewes received oil alone. Progesterone priming had no effect on the proportion of ewes induced to ovulate, and the beneficial effect of progesterone on the proportion of corpora lutea with a normal life-span (P < 0.001, Experiment 1; P < 0.01, Experiment 2) was independent of its time of administration. Injection of progesterone delayed the time of the surge of LH compared to control ewes (P < 0.001), but only when given at the time of introduction of rams.Administration of progesterone by intravaginal devices for 6 h, 12 h or 24 h before introduction of rams (Experiment 3) increased the incidence of corpora lutea with normal luteal life-span compared to control ewes, but had no effect on the proportion of ewes ovulating. The incidence of corpora lutea with a normal life-span was significantly and positively correlated (R=0.977; P < 0.05) with duration of progesterone priming before introduction of rams.These results indicate that duration but not timing of progesterone priming is critical to its ability to eliminate short life-span corpora lutea. Such findings support the concept of a mode of action of progesterone at the ovarian level.     

FSH is superior to eCG for promoting ovarian response in Chinese Bamei gilts

The Bamei gilt is a Chinese native breed located in northwest China, which adapts to the extremely dry and cold environment and is distinguished for its excellent reproductive and maternal characters. To ensure sufficient numbers of embryos for transgenic and nuclear transfer research, hormonal induction of gilt estrus and superovulation may be necessary. The objective of this study was to compare the superovulation effects of equine chorionic gonadotropin (eCG, Group A) and FSH (Groups B-D) in Chinese Bamei gilts. The results show that though eCG could produce more corpora lutea (CL, 14.3) than the control (CL, 9.2), and the FSH treatments had significantly increased the number of CL compared with the eCG treatment. Within the different FSH protocols, the numbers of CL were significantly greater in Groups B (CL, 77.8) and C (CL, 66.8) than in Group D (CL, 42.7), however, ovarian cysts were observed in Groups B and C, but not in Group D. These data suggest that Group D (280 IU FSH) is a suitable protocol to facilitate the development of ovarian follicles and increase the number of useful embryos per gilt for embryos recovery. The optimal FSH protocol of superovulation in Bamei gilts appears to be: D13/100 IU, D14/80 IU, D15/60 IU, D16/40 IU plus prostaglandin (PG) 0.2mg, D17/hCG 1000 IU.     

Good quality sheep embryos produced by superovulation treatment without the use of progesterone devices

Multiple ovulation and embryo transfer (MOET) is a very important tool for the genetic improvement and preservation of endangered livestock. However, the success of a MOET programme highly depends on the number of transferable embryos in response to a superovulation treatment. Thus, the aim of this study was to compare the number and quality of embryos produced during natural oestrus under porcine FSH treatment without the use of progesterone devices to more traditional protocols. Forty Sarda sheep were divided into 2 groups: without sponges (WS) (n = 20) and with sponges (S) containing 40mg FGA for 12 d (n = 20) (control group); 350 I.U. of porcine FSH per sheep was administered in eight decreasing doses twice daily starting four days after estrus was detected (Day 0) in group WS and 48 h before sponge removal in group S. A single i.m. dose of 125 μg of cloprostenol was administered on Day 6 after estrus in group WS to induce luteolysis. Sheep were naturally mated 24 h after cloprostenol injection or sponge removal. Seven days after mating, an inguinal laparotomy was performed and the number of corpora lutea (CL) recorded. Embryos were recovered surgically by flushing each uterine horn. A total of 38 fresh and 22 vitrified embryos were transferred in pairs into 3 groups of recipients seven days after estrus detection: fresh embryos from group S (S-F) (n = 9), fresh embryos from group WS (WS-F) (n = 10) and vitrified embryos from group WS (WS-V) (n = 11). Data on the number of corpora lutea (CL), recovered ova and embryos (OER), and quality 1-2 and 3 embryos (EQ_1-2, EQ₃) per ewe were analyzed by ANOVA. Recovery (RR), fertility (FR) and quality 1-2 embryo (Q_1-2R) rates per treatment were analyzed by a Chi Square analysis. A Chi Square analysis was also applied to pregnancy rate (PR), lambing rate (LR) and twinning rate (TR) of fresh and vitrified embryos in order to analyze embryo transfer results. Among all superovulation variables analysed, results show statistically significant differences in mean number of CL/ ewe (9.3 ± 3.9 vs 7 ± 3.2), RR (67% vs 80 %) and FR (100% vs 80%) (P < 0.05) between WS and S groups respectively. There were no significant differences in PR (78%, 70% and 82%), LR (67%, 60% and 59%) and TR (71%, 71% and 44.4%) among S-F, WS-F and WS-V groups respectively. In conclusion, it is possible to produce a good number of transferable embryos during natural oestrus avoiding the use of sponges.     

Embryo production and progesterone profiles in ewes superovulated with different hormonal treatments

The superovulatory response and embryo yield following hormonal treatments of Merino ewes during late spring and their estrous cycle were evaluated. Ewes (n=17) were treated with progestagen-impregnated sponges and assigned to Group I (800 IU PMSG plus 11.5 mg FSH-p); Group II (1200 IU PMSG); Group III (1600 IU PMSG). Ewes were naturally mated and followed by laparotomy 6 days later. After laparotomy, ewes were injected with a prostaglandin analogue (PGF) and serum samples were obtained prior to surgery and then for 25 days to measure progesterone (P4) by radioimmunoassay. There were no differences among groups neither for estrous incidence (Group I: 83.3%; Group II: 83.3%; Group III: 100%), nor for the time interval to estrous onset (Group I: 26.4±2.4 h; Group II: 28.8±2.9 h; Group III: 24.0±3.8 h). Group I had more corpora lutea than Group II (14.2±1.2 and 6.2±0.8; P<0.05), and Group III was intermediate (11.0±3.0). There was a low incidence of persistent follicles in all treatments (Group I: 0.5±0.5; Group II: 0.6±0.4; Group III: 1.8±1.2). Number of collected ova were 9.0±2.6, 3.8±0.6 and 6.5±0.9 for Groups I, II and III, respectively. Significant differences in number of ova were detected between Groups I and II. Unfertilized ova did not differ among groups (Group I: 3.5±1.0; Group II: 2.8±0.8; Group III: 5.2±1.4; P>0.05). Embryos and high viability embryos were higher (P<0.05) in Group I (5.2±1.9 and 4.8±2.0) than in Group II (1.0±0.5 and 1.0±0.5) or Group III (1.2±0.6 and 1.0±0.5). Total plasma progesterone (P4) and P4 per corpus luteum before PGF administration did not vary (P>0.05) among groups (Group I: 71.0±14.7 and 4.9±0.7 nmol/l; Group II: 50.6±13.3 and 7.9±1.6 nmol/l; Group III: 90.4±42.6 and 6.8±1.8 nmol/l). There was a significant and positive correlation between P4 before PGF administration and number of corpora lutea (r=0.76). No significant differences were detected among groups for: interval PGF to P4 <3.18 nmol/l (Group I: 2.7±0.3 days; Group II: 1.8±0.6 days; Group III: 2.2±0.5 days), cycle length (Group I: 18.3±1.4 days; Group II: 17.9±0.5 days; Group III: 16.8±0.9 days), duration of P4 levels <3.18 nmol/l (Group I: 11.3±1.9 days; Group II: 7.1±1.0 days; Group III: 7.2±2.4 days), duration of P4 levels ≥3.18 nmol/l (Group I: 7.0±1.3 days; Group II: 10.8±0.8 days; Group III: 9.5±1.7 days) and peak of P4 (Group I: 7.4±0.4 nmol/l; Group II: 10.8±1.6 nmol/l; Group III: 9.2±1.9 nmol/l). It was concluded that PMSG–FSH-p treatment was more efficient than PMSG alone for superovulation and embryo production in ewes while P4 profiles were similar among groups.     

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).     

Doppler sonography of the uterine and ovarian arteries during a superovulatory program in horses

The aim of the present study was to investigate the effects of a gonadotropin treatment to induce superovulation on ovarian and uterine blood flow and its relationship with steroid hormone levels and ovarian response in mares, using color Doppler sonography. Each of six mares were examined sonographically in five cycles for 3 d (t1 to t3) during the follicular development phase (FDP) beginning at a follicle size of ≥ 22 mm, and for 4 d (D-4 to D-1; D0 = Ovulation) in the preovulatory phase (POP). After each examination, total estrogens (E_tot) and progesterone (P₄) levels were determined in peripheral plasma. Cycles 1, 3, and 5 (c₁, c3, c5) were unstimulated cycles (USC); in c2 and c4, the mares were stimulated (SC) with eFSH and inseminated when in estrus at 12 and 24 h after hCG administration. Embryo recovery was performed 6.5 d post ovulation. Cycle 5 c5 was an unstimulated cycle with hCG treatment, insemination, and embryo recovery. Ovarian and uterine blood flow was quantified by the blood flow volume (BFV) and the pulsatility index (PI) in ovarian and uterine arteries. The mean number of ovulations and developing CL was 1.3 ± 0.4 in USC and 4.4 ± 3.1 in stimulated cycles (SC) with no difference (P ≥ 0.05) between the ovaries within mares. No difference (P > 0.05) was observed in utBFV and utPI during FDP between USC and SC, but during POP, utPI was lower (P < 0.05) and utBFV higher (P < 0.001) in SC than USC. The ovBFV was higher (P < 0.01) and ovPI lower (P < 0.05) in SC compared to USC. All uterine and ovarian blood flow parameters were related to the number of developing follicles in SC. Parameters utPI (r = −0.67; P < 0.001) and ovPI (r = −0.53; P < 0.001) were negatively correlated with the number of ovulations on t3, and with the number of collected embryos on t3 (utPI: r = −0.81; P < 0.001), D-4 (utPI: r = −0.64; P < 0.0001), and D-1 (ovPI: r = −0.41; P < 0.01). P₄ levels were not positively correlated with utBFV (P > 0.05), but E_tot concentrations (D-4: r = 0.790; D3: r = 0.639; P < 0.001; D-1: r = 0.48; P < 0.001) and ovBFV from D-4 to D-1 (r = 0.64; P < 0.001) in SC were. The results of the present study show that in mares treatment with gonadotropins to induce superovulation is associated with a marked increase in uterine and ovarian perfusion, concurrent with the development of multiple follicles and an increase in E_tot levels. The increased blood flow seems to be related to the effectiveness of ovarian response to stimulation.     

Responses to n-3 fatty acid (LCPUFA) supplementation of gestating gilts,and lactating and weaned sows

Feeding n-3 long-chain polyunsaturated fatty acids (LCPUFA) to gilts or sows has shown different responses to litter growth, pre-weaning mortality and subsequent reproductive performance of the sow. Two hypotheses were tested: (1) that feeding a marine oil-based supplement rich in protected n-3 LCPUFAs to gilts in established gestation would improve the growth performance of their litters; and (2) that continued feeding of the supplement during lactation and after weaning would offset the negative effects of lactational catabolism induced, using an established experimental model involving feed restriction of lactating primiparous sows. A total of 117 primiparous sows were pair-matched at day 60 of gestation by weight, and when possible, litter of origin, and were allocated to be either control sows (CON) fed standard gestation and lactation diets, or treated sows (LCPUFA) fed the standard diets supplemented with 84 g/day of a n-3 LCPUFA rich supplement, from day 60 of first gestation, through a 21-day lactation, and until euthanasia at day 30 of their second gestation. All sows were feed restricted during the last 7 days of lactation to induce catabolism, providing a background challenge against which to determine beneficial effects of n-3 LCPUFA supplementation on subsequent reproduction. In the absence of an effect on litter size or birth weight, n-3 LCPUFA tended to improve piglet BW gain from birth until 34 days after weaning (P = 0.06), while increasing pre-weaning mortality (P = 0.05). It did not affect energy utilization by the sow during lactation, thus not improving the catabolic state of the sows. Supplementation from weaning until day 30 of second gestation did not have an effect on embryonic weight, ovulation rate or early embryonic survival, but did increase corpora lutea (CL) weight (P = 0.001). Eicosapentaenoic acid and docosahexaenoic acid (DHA) levels were increased in sow serum and CL (P < 0.001), whereas only DHA levels increased in embryos (P < 0.01). In conclusion, feeding n-3 LCPUFA to gilts tended to improve litter growth, but did not have an effect on overall subsequent reproductive performance.     

Animal and temporal effects on ovarian follicular dynamics in Brahman heifers

The aims of the current study were to determine if the pattern of ovarian follicular growth and development in Bos indicus heifers is different to that reported in Bos taurus breeds, and to examine the factors that determine which dominant follicle will ovulate. In addition, the extent to which variation in follicular dynamics is attributable to variation between animals and over time was evaluated. The ovaries of 17 Brahman heifers were examined daily by transrectal ultrasonography using a 7.5 MHz transducer for a total of 117 interovulatory intervals over a period of 10 months. Size and position of individual follicles ⪖5 mm in diameter, and size of corpora lutea (CL) were recorded. Circulating progesterone concentrations were determined from plasma samples obtained twice weekly. Although size of dominant follicles and CL within the ovaries of Bos indicus heifers were smaller than reported for Bos taurus breeds, the overall patterns of dominant follicle growth were similar. There were significant correlations between number of dominant follicles occurring prior to ovulation and time of appearance of the second dominant follicle, duration of detection of CL and size of the ovulatory follicle in the preceding oestrous cycle (P < 0.05). There were significant animal effects on a number of ovarian characteristics including number of dominant follicles per oestrous cycle (P < 0.001), with one heifer having four dominant follicles in more than a third of oestrous cycles observed. In addition, changes in daylength over the 10 month period were related to changes in duration of the interovulatory interval, persistence and maximum diameter of CL and size of ovulatory follicles. Liveweight change over the same period was related to changes in maximum diameter of the first dominant follicle.     

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.     

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).     

Embryo recovery from superovulated mouflons (Ovis gmelini musimon) and viability after transfer into domestic sheep

In this study multiple ovulation and embryo transfer (MOET) technology was tested as a method for increasing the number of offspring obtained from superovulated mouflons and then using Sardinian ewes as recipients. Two experiments were carried out over consecutive years. In Experiment 1, female mouflons received a standard superovulatory treatment during both breeding and anoestrous seasons. Sarda sheep, used as controls, received the same treatment. Mean superovulatory response (corpora lutea and large follicles) was higher in the domestic sheep than in the mouflons (4.8 vs. 10.1 and 4.2 vs. 8.8 in breeding and anoestrous seasons, respectively; P < 0.05). A high percentage of mouflons showed early luteal regression which negatively affected recovery rate (35% and 30% in mouflons vs. 69% and 71% in sheep) and the yield of embryos suitable for transfer (37% and 25% in mouflons vs 74% and 69% in sheep; P < 0.05). In Experiment 2, ten mouflons were treated by the same superovulatory protocol and divided into two groups. In the first (Group 5), embryos were recovered earlier by oviductal flushing and cultured in vitro with oviductal cells in CZB medium until the morula/blastocyst stage; in the second (Group 6), the usual embryo recovery time was followed. Recovery rate was higher in the former (89% vs. 31%; P < 0.01) than in the latter. After 4 days of culture, 53% of embryos reached compact morula or early blastocyst stage (16/30). Lambing rate was 57% for mouflon embryos transferred immediately and 56% for those cultured in vitro for 4 days; the lambing rate in the sheep control group was 71%. The length of gestation was longer in ewes carrying mouflons than in those carrying lambs (155 vs. 148 days).