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Effects of superovulation on endogenous LH secretion in cattle, and consequences for embryo production.

Stimulation of follicular growth during superovulation is achieved by the injection of FSH or compounds with high FSH-bioactivities. However, some LH-activity is required for follicle maturation. It is of relevance to evaluate, therefore, the effect of superovulatory treatments on endogenous LH secretion. Luteinizing hormone is secreted in discrete pulses, and the pattern of pulsatile LH secretion during superovulation is reviewed. Four of five published studies have shown that LH pulse frequency is significantly reduced by injection of eCG or FSH preparations. This suppression appears within 8 h of treatment Effects of superovulation on LH pulse amplitude are less consistent. The reasons for the decrease in pulse frequency have been investigated, and although the answer is not definitive, it would seem that increased follicular estradiol, acting perhaps in synergism with progesterone, may play a role. Changes in plasma progesterone concentrations are not related to changes in LH pulse frequency. What is the significance of decreased LH pulse frequency? We attempted to investigate this by inducing LH pulses during superovulation, but the result was a major reduction in ovulation rate. More research is required to determine if modification of endogenous LH secretion can improve superovulatory responses.

Animals↗

Drugs used to treat reproductive disorders.

As with all aspects of medicine, it is necessary to fully understand disease processes before treatment modalities can be designed. Many disorders in small animal reproduction have not been completely evaluated, and further investigation is ongoing. Many reproductive hormone products are available but have limited clinical applicability. Other products have potential use but are not commercially available. A thorough diagnostic evaluation is always indicated before embarking on a treatment regimen.

Abortion, Induced↗

The reproductive pattern and efficiency of female buffaloes.

Buffaloes play a prominent role in rural livestock production, particularly in Asia. Reproductive efficiency is the primary factor affecting productivity and is hampered in female buffalo by (i) inherent late maturity, (ii) poor estrus expression in summer, (iii) distinct seasonal reproductive patterns, and (iv) prolonged intercalving intervals. Ovarian function is central to these issues; hence, the focal point of this review is ovarian function in Bubalus bubalis, particularly, in relation to seasonal changes. Ovarian anatomy, follicular and luteal development development, and hormonal profiles during the estrous cycle are discussed. Review of the literature revealed a paucity of critically derived information on follicular and ovulatory patterns in buffalo, particularly, in relation to seasonal estrus/birthing. Efforts may be directed at understanding the process (recruitment, development, atresia) and temporal pattern (follicle selection, dominance, subordinate follicle suppression, follicle numbers, and, preovulatory changes) of follicular dynamics using techniques which permit serial assessment of changes occurring over time. Emphasis may be directed towards investigating follicular "waves" as a functional unit, rather than the estrous cycle, in the context of whole animal endocrinology. The data obtained from such basic studies may then be used to develop and test models for enhancing reproductive efficiency.

Animals↗

Progestagen synchronisation in the absence of a corpus luteum results in the ovulation of a persistent follicle in cyclic ewe lambs.

Progestagens are widely used to synchronise oestrous in sheep but the effects on follicular dynamics are not clear. We tested the hypothesis that when luteolysis occurs early during progestagen synchronisation prolonged growth of the ovulatory follicle will occur. Cyclic ewe lambs (40.0+/-0.3 kg) were divided into three groups: eight ewes (Long group) received a progestagen sponge (60 mg medroxyprogesterone acetate) from Days 5 to 19 after oestrous and eight ewes (Short group) received a progestagen sponge on Day 5 which was replaced on Day 10 and again on Day 15, and removed on Day 19 after oestrous. On Days 6 and 7, ewes in both groups received prostaglandin. A third group (n=5, Control) did not receive any treatment. The growth and development of follicles > or =2 mm in diameter were characterised using daily transrectal ultrasonography. On Day 18, blood samples were collected every 12 min for 8 h from five ewes in the Long and Short groups. Data were analysed by ANOVA. The maximum diameter and age (emergence to ovulation) of the ovulatory follicle was greater (P<0.01) in ewes in the Long group (7. 4+/-0.2 mm and 12.1+/-0.6 days) than in ewes in the Short group (6. 3+/-0.2 mm and 5.1+/-0.5 days) and Control group (6.3+/-0.4 mm and 6. 8+/-0.6 days). On Day 18 of the cycle, LH pulse frequency and oestradiol concentrations were greater (P<0.05) in ewes in the Long group (3.2+/-1.1 pulse per 8 h and 1.15+/-0.09 pg ml(-1)) than the Short group (0.8+/-0.4 pulses per 8 h and 0.54+/-0.08 pg ml(-1)). We suggest that the negative feedback efficacy of a long-term progestagen sponge decreased with time and led to an increase in LH pulse frequency and prolonged growth of the ovulatory follicle. We conclude that, in the absence of luteal progesterone, synchronisation with a single progestagen sponge for 14 days resulted in higher LH pulse frequency and ovulation of a persistent follicle with a larger maximum diameter, compared with controls.

Animals↗

Reproduction in Old World camels.

The article reviews aspects of the basic reproductive biology of the camel, such as puberty, breeding season, ovarian dynamics, synchronisation of ovarian activity and artificial insemination, and superovulation and embryo transfer. Pregnancy and parturition are also discussed.

Animals↗

In vivo oocyte recovery and in vitro embryo production from bovine oocyte donors treated with progestagen, oestradiol and FSH.

The effect of treatment of donor cattle with progestagen and oestradiol or FSH on in vivo oocyte recovery and in vitro embryo production was studied. Forty-eight beefxFriesian cows formed eight replicates of six treatments in a 2 (no steroid versus steroid)x3 (none, single or multiple dose(s) of FSH) factorial design in which follicles were aspirated once weekly for 3 weeks. Oocytes were graded, washed, matured for 20-24h and then inseminated with frozen/thawed semen from a single sire followed by coculture on granulosa cell monolayers. Treatment with steroid had no significant effect on any follicular, oocyte or embryo production variate other than to reduce the number (P<0.05) and the diameter of large follicles>10mm (P<0.01) present at aspiration. FSH increased numbers of medium (6-10mm) and large follicles (P<0.01) and there was a corresponding decrease in the number of small follicles (2-5mm; P<0. 01). The total number of follicles at aspiration increased from 17. 7+/-1.60 for animals not treated with FSH to 23.6+/-1.97 following multiple dose treatment with FSH (P<0.05). Significantly, more follicles were aspirated following FSH treatment (no FSH 9.7+/-1.09, single dose FSH 13.6+/-1.30, multiple dose FSH 17.3+/-1.52; P<0.05) and numbers of oocytes recovered per cow per week increased (no FSH 4.1+/-0.76, single dose FSH 5.3+/-0.87, multiple dose FSH 5.9+/-0. 94) but the differences were not significant. Significantly, more good oocytes (Category 1) were recovered from animals treated with FSH (P<0.05). There was no overall significant effect of FSH on embryo production rate or the total number of transferable embryos produced but the number of transferable embryos was highest following administration of multiple doses of FSH. In conclusion, progestagen plus oestradiol 17beta treatment did not affect follicle, oocyte and embryo production of oocyte donors aspirated once per week. FSH treatment, however, significantly increased the number of follicles aspirated and Category 1 oocytes recovered. Multiple dose administration of FSH resulted in the production of the highest number of transferable embryos but this effect was not significant.

Animals↗

The role of LH pulse frequency in ACTH-induced ovarian follicular cysts in heifers.

The aim of the present study was to induce ovarian cysts experimentally in cattle using ACTH and to closely examine the role of LH pulse frequency in ovarian cyst formation. Five regularly cycling Holstein-Friesian heifers (15-18-month-old) were used. Ovaries were scanned daily using an ultrasound scanner with a 7.5 MHz rectal transducer. Daily blood samples were obtained via tail venepuncture for hormone analyses. Additional blood samples (for FSH and LH pulses) were obtained through an indwelling jugular vein catheters every 15 min for 8 h on Days 2 (early luteal phase; ELP), 12 (mid-luteal phase; MLP) and 19 (follicular phase; FP) of control estrous cycle and on alternate days during follicular cyst (FC) formation and persistence. Cysts were induced using subcutaneous injections of ACTH (Cortrosyn) Z; 1 mg) every 12 h for 7 days beginning on Day 15 of the subsequent estrous cycle. Plasma concentrations of progesterone (P4), estradiol-17beta, FSH and LH were determined by double antibody radioimmunoassay while cortisol concentration was determined by enzyme immunoassay (EIA). Ovarian follicular and endocrine dynamics were normal during the control estrous cycles. Ovarian follicular cysts were induced in four of the five heifers. Mean maximum size of cysts was larger (P<0.05) than that of ovulatory follicles (26.78+/-3.65 versus 14.1+/-0.90 mm), respectively. Cortisol levels were increased during ACTH treatment. High concentrations of estradiol and low progesterone were observed after cyst formation. LH pulse frequency was significantly reduced (P<0.05) during cyst formation and persistence compared to ELP (7.5+/-0.75) and FP (6.5+/-0.58), but was not significantly (P=0.23) different from MLP (2.8+/-0.29) pulses. Mean LH pulse amplitude and concentrations were not different. Similarly, the mean pulse frequency, amplitude and concentration of FSH were not different between control study and cystic heifers. These results suggest that the LH pulse frequency observed following ACTH treatment may interact with high estradiol concentration to induce ovarian cyst formation in heifers.

Adrenocorticotropic Hormone↗

Preovulatory follicular status and diet affect the insulin and glucose content of follicles in high-yielding dairy cows.

Insulin and glucose may be limiting factors for ovarian function in dairy cows genetically selected for high milk yield. The effects of nutrition on the intrafollicular content of insulin and glucose were investigated in Israeli Holstein dairy cattle fed a basic total mixed ration and producing 34-39kg of milk daily. In experiment 1, carried out in 11 oestrus-synchronised cows, little variation in insulin concentration was found in plasma sampled during the luteal phase, but high variation was found in plasma sampled during the follicular phase. Therefore, in order to prevent confounding the effects of diet and of phase in cycle in the following experiments, experimental diets were fed during the luteal phase of synchronised oestrus cycles. In experiment 2, designed as Latin-Square, six cows received sequentially diets containing 17.1 (control) or 19.7% of crude protein, using two sources of supplementary protein, i.e. soyabean meal (SBM) and corn gluten meal (CGM), differing in ruminal degradability and leucine content. When dry matter intake was used as covariant, plasma insulin on day 16 was 29.5 and 26.4% higher in cows fed diets containing SBM and CGM than in the control (P<0.05). In experiment 3, 17 cows were individually fed the basic diet and then switched to isoenergetic diets containing SBM (n=5), CGM (n=6) or corn grain (CG, n=6) given from day 10 to 16 of the synchronised oestrus cycle. On the eve of day 16, and in the morning of day 17, they were administered PGF(2alpha) and the content of 26 largest follicles was aspirated by using the transvaginal ovum pick-up technique. Follicles were sorted into two classes (preovulatory and subordinate) according to oestradiol concentration and the progesterone:oestradiol ratio in follicular fluid (FF). Higher concentrations of insulin (0.282 versus 0.127ng/ml, P<0.0001) and of glucose (0.614 versus 0.386g/l, P<0.002), were found in FF from preovulatory follicles. The insulin concentration in the FF of cows fed the CG diet was 26% higher than in their counterparts fed CGM (P<0.04), SBM being intermediate. Dietary effects did not reach significance in subordinate follicles. The finding that preovulatory follicular status is associated with increased intrafollicular insulin and glucose suggests that insulin is involved in follicular maturation. The nutritional effect on intrafollicular glucose and insulin may have practical implications to optimise feeding in dairy cows during phases of the oestrus cycle.

Animal Feed↗

Superovulation in ewes by a single injection of pFSH dissolved in polyvinylpyrrolidone (PVP): effects of PVP molecular weight, concentration and schedule of treatment.

Three experiments were carried out to evaluate induction in ewes of superovulation and embryo production by a single injection of a porcine pituitary extract (pFSH) dissolved in polyvinylpyrrolidone (PVP), investigating the effects of PVP molecular weight and its concentration (Experiment I), time and method of treatment (Experiments II and III). All ewes were synchronized for estrus by vaginal sponges impregnated with fluorogestone acetate (FGA; 30 mg, 9 days) plus PGF(2alpha) (Cloprostenol, 50 microg, 48h before sponge removal - s.r.), and superovulated by 250 IU pFSH. In Experiment I, 60 Gentile di Puglia ewes were subdivided into five experimental groups (n = 12): Group A, the control, received six decreasing intramuscular (i.m.) doses of pFSH, 12 h apart, beginning 48h before s.r.; Groups B and C were given 48 h before s.r. a single i.m. injection of pFSH dissolved in PVP with MW = 10,000, respectively, at concentrations of 15 and 30% w/v; Groups D and E received the same treatments as for B and C using PVP with MW = 40,000. None of the pFSH-PVP treatments were effective in inducing superovulation. In Experiment II, 22 Leccese ewes were subdivided into two groups (n = 11): Group A, control received i.m. four decreasing doses of pFSH, beginning 24 h before s.r., 12h apart; Group B was given a single i.m. injection of pFSH dissolved in PVP (MW = 40,000 at 30% w/v), 24 h before s.r. The pFSH-PVP treatment provided an ovulation rate similar to the control and tended to enhance embryo yield (4.4 versus 2.4, P>0.05). In Experiment III, 60 Leccese ewes were subdivided into six treatment groups (n = 10). Groups A and D served as controls and received i.m. 12 h apart, six doses (from 48 h before s.r.) and four doses (from 24h before s.r.) of pFSH, respectively. Groups B and C were treated by a single injection of pFSH in PVP (MW = 10,000; 30% w/v) 48 h before s.r., respectively by i.m. or subcutaneous (s.c.) administration. Groups E and F received the same treatments as for B and C 24 h before s.r. Intramuscular pFSH-PVP administration 24 h before s.r. provided an ovulation rate (8.1), mean numbers of ova recovered (5.6) and fertilized (4.2) comparable to the six or four dose treatments and significantly higher (P <0.01) compared to the pFSH-PVP treatment carried out i.m. 48 h before s.r. These results show that a single injection of pFSH dissolved in PVP at 30% w/v, performed i.m. 24 h before s.r., is able to induce a superovulatory response comparable to that following multiple injection treatment, regardless of PVP molecular weight.

Animals↗

Alteration of gonadotrophin and steroid hormone release, and of ovarian function by a GnRH antagonist in gilts.

This study examined the impact of the gonadotrophin-releasing hormone (GnRH) antagonist Antarelix on LH, FSH, ovarian steroid hormone secretion, follicular development and pituitary response to LHRH in cycling gilts. Oestrous cycle of 24 Landrace gilts was synchronised with Regumate (for 15 days) followed by 800 IU PMSG 24h later. In experiment 1, Antarelix (n=6 gilts) was injected i.v. (0.5mg per injection) twice daily on four consecutive days from day 3 to 6 (day 0=last day of Regumate feeding). Control gilts (n=6) received saline. Blood was sampled daily, and every 20 min for 6h on days 2, 4, 6, 8 and 10. In experiment 2, gilts (n=12) were assigned to the following treatments: Antarelix; Antarelix + 50 microg LHRH on day 4; Antarelix + 150 microg LHRH on day 4 or control, 50 microg LHRH only on day 4. Blood samples were collected daily and every 20 min for 6h on days 2, 4 and 6 to assess LH pulsatility. Ovarian follicular development was evaluated at slaughter. Antarelix suppressed (P<0.05) serum LH concentrations. The amount of LH released on days 4-9 (experiment 1) was 8.80 versus 36.54 ngml(-1) (S.E.M.=6.54). The pattern of FSH, and the preovulatory oestradiol rise was not affected by GnRH antagonist. Suppression of LH resulted in a failure (P<0.05) of postovulatory progesterone secretion. Exogenous LHRH (experiment 2) induced a preovulatory-like LH peak, however in Antarelix treated gilts the LH surge started earlier and its duration was less compared to controls (P<0.01). Furthermore, the amount of LH released from day 4 to 5 was lower (P<0.01) in Antarelix, Antarelix + 50 and Antarelix + 150 treated animals compared to controls. No differences were estimated in the number of LH pulses between days and treatment. Pulsatile FSH was not affected by treatment. Mean basal LH levels were lower (P<0.05) after antagonist treatment compared to controls. Antarelix blocked the preovulatory LH surge and ovulation, but the effects of Antarelix were reduced by exogenous LHRH treatment. The development of follicles larger than 4mm was suppressed (P<0.05) by antagonist treatment. In conclusion, Antarelix treatment during the follicular phase blocked preovulatory LH surge, while FSH and oestradiol secretion were not affected. Antarelix failed to alter pulsatile LH and FSH secretor or pituitary responsiveness to LHRH during the preovulatory period.

Animals↗

Calf survival from embryo transfer-induced twinning in dairy-beef cows and the effects of synchronised calving.

In each of 4 years, 94-116 mature cows had two 6-7-day-old embryos, produced by the in vitro fertilisation of oocytes, inserted non-surgically into one uterine horn of each cow. Starting 5 days before the expected date of calving, the cows were continuously observed and assistance at calving was provided when required. In year 1, perinatal calf survival was similar in twin-calving (TC) and single-calving (SC) cows (98.1 versus 100% for calves born to TC and SC, respectively). There was a higher incidence of assistance at birth for TC (52%) than for SC (21%). In years 2 and 3, the calving of 30 SC and 33 TC was synchronised using an injection of Opticortinol (OP) 6-9 days before the injection of Estrumate and Dexol-5 (E+D). A further 34 SC calved naturally. Synchronised calving reduced the spread of calving from 16-25 to 8-9 days without reducing perinatal calf survival and had no significant effect on the incidence of assistance at birth in SC. The TC in years 2 and 3 had a high incidence of retained placenta at 48 h (70%) and a high incidence of assistance at birth (85%). In year 4, calving was synchronised in 16 SC and 21 TC with E+D and no pre-treatment with OP, while 15 SC were treated with both OP and E+D. There were no effects of the hormone treatment on perinatal calf survival and only small effects on the incidence of assisted births for SC. The incidence of retained placenta at 48 h was lower for SC pre-treated with OP (40%) than for SC (88%) and TC (76%) not pre-treated with OP. Continuous supervision over calving produced perinatal calf survival rates for TC that were similar to SC, despite the higher incidence of assistance of TC at parturition. Hormonal synchronisation of calving can halve the time required for continuous supervision of calving, but the hormone treatments exacerbate the already high incidence of retained placenta in TC.

Adrenal Cortex Hormones↗

Restoration of LH output and 17beta-oestradiol responsiveness in acutely ovariectomised holstein dairy cows pre-treated with a GnRH agonist (deslorelin) for 10 days.

The objectives of the study were firstly to identify the role of the ovary in maintaining plasma luteinising hormone (LH) concentrations in cows treated with an implant of a potent GnRH agonist (deslorelin), and secondly to characterise the changes in LH following ovariectomy (OVX) in the same animals. Oestrus was synchronised in mature Holstein dairy cows and deslorelin implants were inserted 17 days later into two-third of the cows. A further 10 days later (day 0) all cows had bilateral OVX performed. A control group (CON; n=4) received no treatment and had blood samples collected at 15-min intervals for 8h on the day prior to OVX (day -1) and similarly on days 4 and 10. One group (DES_IN; n=4) had implants in place for the duration of the study while another group had implants removed (DES_OUT; n=4) at the time of OVX. DES_IN cows were sampled hourly at each sampling session (days -1, +4 and +10), whereas DES_OUT cows were sampled similarly to CON except on day -1 when hourly samples were collected. Predictable post-operative increases in mean LH (0.61 ng/ml versus 1.79 ng/ml; P<0.01) and LH pulse amplitude (0.66 ng/ml versus 1.56 ng/ml; day -1 versus day +10; P<0.01) occurred after CON cows were ovariectomised. Smoothed LH means showed a delayed effect of time compared to arithmetic means. Pulse frequency was unchanged following OVX in CON cows. A comparison of all cows that had been treated with deslorelin from day -1 showed a significant elevation of smoothed mean LH compared to untreated cows (0.80 ng/ml versus 0.34 ng/ml; DES_IN and DES_OUT versus CON; P<0.05). DES_IN cows had a 54% reduction in mean LH from day -1 to +4 following OVX (1.05 ng/ml versus 0.48 ng/ml; P<0.01) indicating the probable involvement of the ovary in the maintenance of elevated basal LH. No further reduction was detected by day +10. The LH response to an intramuscular (IM) injection of 500 microg 17beta-oestradiol (E2) on day +11 varied significantly between treatment groups (P<0.01). CON cows showed a typical LH surge, reaching maximum concentrations (10.3 ng/ml) at 17.3h post-injection. Even though low amplitude LH pulsatility had been restored in DES_OUT cows by day +4, there was an inconsistent response to E2 on day +12; one cow had an apparently normal surge yet, others showed only attenuated responses. Pulse amplitude in DES_OUT cows was lower at days +4 and +10 compared to CON (P<0.05). DES_IN cows did not produce any surge after E2. Mean LH prior to OVX (day -1) remained unchanged following the 500 microg oestradiol injection (0.38 ng/ml versus 0.45 ng/ml pre-E2 versus post-E2 compared to 1.05 ng/ml pre-OVX). The results of this experiment implicated ovarian involvement in maintaining elevated basal LH output in cows that were chronically treated with a GnRH agonist. Individual cows varied in their LH surge response to exogenous E2 given 12 days after implant removal, even though LH pulse amplitude and frequency had been restored.

Animals↗

Effect of climate on the response to three oestrous synchronisation techniques in lactating dairy cows.

The reproductive efficiency of Friesian dairy cows was investigated in a three (oestrous synchronisation technique) x two (seasons of the year) factorial design. The 90 primiparous and multiparous cows (winter, n=42; summer, n=48) were allocated at random to three synchronisation treatments (n=30 cows per treatment). In treatment 1 (GPG), the cows were administered 15 mg PGF(2alpha) i.m. at 30 +/- 3 days postpartum, 100 microg GnRH i.m. at 51 +/- 3 days and 15 mg PGF(2alpha) 7 days later. A second 100 microg dose of GnRH was given after, further 2 days and fixed time AI occurred 16-20 h later. In treatment 2 (PG-PG), 15 mg PGF(2alpha) was administered i.m. to each cow on three occasions at successive 14 days interval starting at 30 +/- 3 days postpartum and the cows were inseminated at observed oestrus following the third dose of PGF(2alpha). Cows in treatment 3 (PG) had a single administration of 15 mg PGF(2alpha) i.m. at 57+/-3days postpartum and were inseminated as in treatment 2. Mean daily ambient temperature was 10.9 degrees C in winter (November-March) and 20.2 degrees C in summer (June-October). The cows were confined in an open-fronted shed and had ad libitum access to a complete diet with a 37:63 forage to concentrate ratio. Body condition score was assessed at 57 +/- 3 days postpartum. Cow rectal temperature at insemination, milk yield, reproductive data and climatic variables were recorded. Blood samples were collected for progesterone assay on days 4, 11, 18, 25, 32, 39 and 46 post-AI from 54 of the cows (19 GPG; 17 PG-PG; 18 PG). Pregnancy rate to first AI was 36.7% (11/30) for GPG and 16.7% (5/30) for both PG-PG and PG treatments. The difference was not significant. The cumulative pregnancy rate after third AI were GPG 83.3% (25/30), PG-PG 60.0% (18/30) and PG 60.0% (18/30; P<0.057). The cumulative pregnancy rate for cows inseminated in the winter (81.0%; 34/42) was higher (P<0.01) than for those inseminated in the summer (56.3%; 27/48). The interval from calving to first service was shorter (P<0.05) in treatment PG-PG (65.4+/-1.3 days) than in PG (69.2+/-1.3 days). Mean plasma progesterone concentrations post-AI of pregnant cows were higher (P<0.001) for GPG cows than those for PG-PG and PG cows. Plasma progesterone levels of pregnant cows tended to be higher (P=0.087) in winter than in summer. In conclusion, although the cumulative pregnancy rate was higher for GPG cows, it may be appropriate to correct the nutrition and management of the herd before resorting to synchronisation techniques to improve animal reproductive performances.

Animals↗

Embryo production using defined oocyte maturation and zygote culture media following repeated ovum pick-up (OPU) from FSH-stimulated Simmental heifers.

To determine whether differences in ovarian follicle populations and endocrine status at ovum pick-up (OPU) influenced the quality and developmental competence of oocyte-cumulus complexes (OCC's) collected from follicle stimulating hormone (FSH)-stimulated donors, 24 Simmental heifers had their ovarian follicles aspirated via transvaginal ultrasound-guided OPU at both 15 (OPU1) and 21 (OPU2) days following a synchronised oestrus, on four consecutive occasions at 15-week intervals. More OCC's were collected during OPU1 than OPU2 (means +/- S.E.M. = 7.2 +/- 0.47 versus 5.7 +/- 0.44; P = 0.01), but the respective percentages that were of good quality (categories 1 and 2) did not differ significantly (55 +/- 3% versus 47 +/- 3%). The incidence of zygote cleavage following OCC maturation (Medium 199; protein-free), in vitro fertilization (mTALP; including 0.6% (w/v) albumin) and culture (modified SOF; protein-free) was not significantly different (mean +/- S.E.M. = 81 +/- 2% and 71 +/- 7% for OPU1 and OPU2, respectively). Corresponding blastocyst yields from good quality OCC's (24 +/- 3% and 26 +/- 4%) also did not differ. Although the same 3-day FSH regimen was used immediately prior to each OPU session, plasma FSH concentrations were consistently lower at OPU1 than OPU2 (1.3 +/- 0.28 ng/ml versus 2.5 +/- 0.45 ng/ml; P < 0.05). In contrast, plasma progesterone concentrations were higher at OPU1 (6.6 +/- 0.48 ng/ml versus 3.9 +/- 0.53 ng/ml; P < 0.001), with concentrations at OPU2 being consistent with the presence of luteal tissues, including both persistent corpora lutea and luteinised follicle remnants following OPU1. Failure of the significant differences in follicular and endocrine status between OPU1 and OPU2 to alter the developmental competence of OCC's suggests that, probably as a result of its stabilising influence on nutritionally-sensitive intraovarian regulators of oocyte competence, the constant feeding regimen had a more profound effect on oocyte quality than observed shifts in the peripheral concentrations of some reproductive hormones. Finally, the study demonstrates that it is possible to generate acceptable numbers of in vitro blastocyst-stage embryos from high genetic merit heifers using strategies which restrict reliance on protein to the in vitro fertilization stage of the production process.

Animals↗

Embryo survival and recipient pregnancy rates after transfer of fresh or vitrified, in vivo or in vitro produced ovine blastocysts.

The aim of this study was to assess the effect of production system and of cryopreservation of ovine embryos on their viability when transferred to recipients. The experimental design was an unbalanced 2 x 2 factorial design of two embryo production systems (in vivo versus in vitro) and two embryo preservation conditions prior to transfer (transferred fresh versus transferred after vitrification/warming). For the production of blastocysts in vivo, crossbred donor ewes (n=30) were synchronised using a 13-day intravaginal progestagen pessary. Ewes received 1500 IU equine chorionic gonadotropin (eCG) 2 days before pessary withdrawal, and were mated 2 days after pessary withdrawal and embryos were recovered surgically (6 days after mating). Blastocysts were produced in vitro (IVP) using standard techniques. Recipients (n=95) were synchronised using a progestagen pessary and received 500 IU eCG at pessary removal and were randomly assigned to receive (two per recipient) in vivo fresh (n=10), in vivo vitrified (n=10), in vitro fresh (n=35) or in vitro vitrified (n=40) blastocysts. Recipients were slaughtered at day 42 of gestation and foetuses recovered. Pregnancy and embryo survival rates were recorded and analysed using CATMOD procedures. Foetal weights and crown-rump lengths were recorded and analysed using generalised linear model (GLM) procedures. There were no statistically significant interactions between the effects of embryo production system and preservation status at transfer on pregnancy rate and embryo survival. The pregnancy rate following transfer of fresh IVP blastocysts was lower (P<0.07) than that of in vivo embryos (54.3% versus 90.0%, respectively). Vitrification resulted in a decrease in pregnancy rate, the effect being more pronounced in the case of IVP embryos (54.3-5.0%, P<0.001) compared with in vivo embryos (90.0-50.0%), although the absolute change was similar (49.3% versus 40.0%). Transfer of fresh IVP blastocysts resulted in a higher proportion of single (78.9% versus 33.3%) and lower proportion of twin (21.1% versus 66.7%) pregnancies than those produced in vivo. This was reflected in a significant difference in embryo survival rate (fresh: 32.8% versus 75.0%, P<0.01; vitrified: 2.5% versus 35.0%, P<0.001, for IVP and in vivo blastocysts, respectively). Similarly, all pregnancies resulting from the transfer of vitrified/warmed IVP blastocysts were single pregnancies, while 40% of those from vitrified/warmed in vivo blastocysts were twin pregnancies; this was reflected in an embryo survival rate of 35.0% versus 75.0%, respectively. There was a significant effect (P=0.0184) of litter size on foetal weight but not on foetal length (P=0.3304). Foetuses derived from the fresh transfer of IVP blastocysts were heavier (6.4+/-0.2g versus 5.8+/-0.2g, respectively, P<0.05) and longer (5.2+/-0.1cm versus 4.8+/-0.1cm, respectively, P<0.01) than those derived from fresh in vivo blastocysts. There was no difference in these parameters as a consequence of vitrification of IVP embryos. However, in vivo blastocysts subjected to vitrification resulted in heavier (6.6+/-0.3g versus 5.8+/-0.2g, respectively, P=0.055) and longer (5.2+/-0.1cm versus 4.8+/-0.1cm, respectively, P<0.05) foetuses than their counterparts transferred fresh.

Animals↗

Reproductive responses following royal jelly treatment administered orally or intramuscularly into progesterone-treated Awassi ewes.

An experiment was conducted to determine whether natural royal jelly (RJ) paste administered orally or intramuscularly (i.m.) in conjunction with exogenous progesterone is associated with improved reproductive responses in ewes. Thirty 3-6-year-old Awassi ewes were randomly allocated into three (RJ-capsule, RJC; RJ-injection, RJI and control, CON) groups of 10 ewes each. All ewes were treated with intravaginal progesterone sponges for 12 days. Ewes in the RJC and RJI were administered orally or i.m. with a total of 3g of RJ given in 12 equal doses of 250 mg per ewe per day starting at the time of sponge insertion. At the time of sponge withdrawal (day 0, 0 h), ewes were exposed to three rams and checked for breeding marks at 6-h intervals for 3 days. Blood samples were collected from all ewes for analysis of progesterone concentrations. Pretreatment progesterone levels were <0.5 ng x ml(-1) in 16/30 and >1.3 ng x ml(-1) in the remaining ewes indicating luteal function and cyclicity. Similar reproductive responses and progesterone levels occurred in ewes of the RJC and RJI; therefore, data of the two groups were pooled. Following sponge insertion, progesterone levels increased rapidly and reached maximum values of 5.8+/-0.2 ng x ml(-1) within 2 days among ewes of the three groups, and then declined gradually to day 0 values of 1.6+/-0.1 and 1.9+/-0.1 ng x ml(-1) for the RJ-treated and CON ewes, respectively. The rate of progesterone decline was greater (P<0.001) in RJ-treated than in CON. Mean progesterone levels during the 12-day period were lower (P<0.001) in RJ-treated than in CON (2.8+/-0.2 ng x ml(-1) versus 3.3+/-0.2 ng x ml(-1)). Treatment with RJ resulted in greater (P<0.05) incidence of oestrus and shorter (P<0.05) intervals to onset of oestrus than CON. Based upon progesterone levels, ovulation occurred following day 0 in all ewes. Progesterone increased on day 3 in RJ-treated and on day 4 in CON ewes. Progesterone remained elevated through day 18 in 8/20 RJ-treated and 1/10 CON ewes (P=0.09). All pregnant ewes exhibited oestrus 14 h earlier (P<0.02), ovulated approximately 1 day earlier and had higher (P<0.001) luteal phase progesterone levels than non-pregnant ewes. Non-pregnant had higher (P<0.04) body weights than pregnant ewes. In conclusion, results demonstrate that both RJ treatments in conjunction with exogenous progesterone were equally capable of improving oestrus response and pregnancy rate.

Administration, Oral↗