Effects of oestradiol and some of its esters on gonadotrophin release and ovarian follicular dynamics in CIDR-treated beef cattle

Three experiments were conducted to: (1) compare the effect of three oestradiol formulations on gonadotrophin release in ovariectomised cows; (2) compare the effects of either oestradiol-17beta (E-17beta) or oestradiol benzoate (EB), given at two doses, on the synchrony of ovarian follicular wave emergence in CIDR-treated beef cattle; and (3) determine the timing of ovulation of the dominant follicle of a synchronised follicular wave following administration of E-17beta or EB 24h after progesterone withdrawal. In Experiment 1, ovariectomised cows (n = 16) received a once-used CIDR on Day 0 (beginning of the experiment) and were allocated randomly to receive 5mg of E-17beta, EB or oestradiol valerate (EV) plus 100mg progesterone i.m. The CIDR inserts were removed on Day 7. There were effects of time, and a treatment-by-time interaction (P < 0.0001) for plasma concentrations of both oestradiol and FSH. Plasma oestradiol concentrations peaked 12h after treatment, with highest (P < 0.01) peak concentrations in cows given E-17beta; estradiol concentrations subsequently returned to baseline by 36 h in E-17beta-treated cows and by 96 h in EB- and EV-treated cows. Plasma FSH concentrations decreased by 12h after oestradiol treatment in all groups (P < 0.0001), reached a nadir at 24h, and increased by 60 h in all groups; plasma FSH reached higher (P < 0.02) concentrations in E-17beta-treated than in EB- or EV-treated cows. In Experiment 2, non-lactating Hereford cows (n = 29) received a new CIDR on Day 0 (beginning of the experiment), and were assigned randomly to receive 1 or 5mg of E-17beta or EB i.m. on Day 1. On Day 8, CIDR were removed and PGF was given. Transrectal ultrasonography was done once daily from 2 days before CIDR insertion to 2 days after CIDR removal, and then twice-daily to ovulation. Although there was no difference among groups in the interval from oestradiol treatment to follicular wave emergence (4.2 +/- 0.3 days; P = 0.5), 5mg of E-17beta resulted in the least variable interval to wave emergence (P < 0.005), compared with the other treatment groups which were not different (P = 0.1). For the interval from CIDR removal to ovulation, there were no differences among groups for either means (P = 0.5) or variances (P = 0.1). In Experiment 3, beef heifers (n = 32) received a once-used CIDR on Day 0 (beginning of the experiment) plus 100mg progesterone i.m. and were assigned randomly to receive 5mg E-17beta or 1mg EB i.m. On Day 7, CIDR were removed and all heifers received PGF. On Day 8 (24h after CIDR removal), each group was subdivided randomly to receive 1mg of either E-17beta or EB i.m. There was no effect of oestradiol formulation on interval from treatment to follicular wave emergence (4.1 +/- 0.2 days; P = 0.7) or on the median interval (76.6h; P = 0.7) or range (72-120 h; P = 0.08) from CIDR removal to ovulation. In summary, oestradiol treatments suppressed FSH in ovariectomised cows, with the duration of suppression dependent on the oestradiol formulation. Both E-17beta and EB effectively synchronised ovarian follicular wave emergence and ovulation in CIDR-treated cattle, and the interval from CIDR removal to ovulation did not differ in heifers given either E-17beta or EB 24h after CIDR removal.     

Effects of estradiol cypionate (ECP) on ovarian follicular dynamics,synchrony of ovulation,and fertility in CIDR-based,fixed-time AI programs in beef heifers

Estradiol cypionate (ECP) was used in beef heifers receiving a controlled internal drug release (CIDR; insertion = Day 0) device for fixed-time AI (FTAI) in four experiments. In Experiment 1, heifers (n = 24) received 1mg ECP or 1mg ECP plus 50mg commercial progesterone (CP) preparation i.m. on Day 0. Eight or 9 days later, CIDR were removed, PGF was administered and heifers were allocated to receive 0.5mg ECP i.m. concurrently (ECP0) or 24h later (ECP24). There was no effect of treatment (P = 0.6) on mean (+/-S.E.M.) day of follicular wave emergence (3.9+/-0.4 days). Interval from CIDR removal to ovulation was affected (P<0.05) only by duration of CIDR treatment (88.3+/-3.8h versus 76.4+/-4.1h; 8 days versus 9 days, respectively). In Experiment 2, 58 heifers received 100mg progesterone and either 5mg estradiol-17beta or 1mg ECP i.m. (E-17beta and ECP groups, respectively) on Day 0. Seven (E-17beta group) or 9 days (ECP group) later, CIDR were removed, PGF was administered and heifers received ECP (as in Experiment 1) or 1mg EB 24h after CIDR removal, with FTAI 58-60h after CIDR removal. Follicular wave emergence was later (P<0.02) and more variable (P<0.002) in heifers given ECP than in those given E-17beta (4.1+/-0.4 days versus 3.3+/-0.1 days), but pregnancy rate was unaffected (overall, 69%; P = 0.2). In Experiment 3, 30 heifers received a CIDR device and 5mg E-17beta, with or without 100mg progesterone (P) i.m. on Day 0. On Day 7, CIDR were removed and heifers received ECP as described in Experiment 1 or no estradiol (Control). Intervals from CIDR removal to ovulation were shorter (P<0.05) in ECP0 (81.6+/-5.0h) and ECP24 (86.4+/-3.5h) groups than in the Control group (98.4+/-5.6h). In Experiment 4, heifers (n = 300) received a CIDR device, E-17beta, P, and PGF (as in Experiment 3) and after CIDR removal were allocated to three groups (as in Experiment 2), with FTAI 54-56h (ECP0) or 56-58h (ECP24 and EB24) after CIDR removal. Pregnancy rate did not differ among groups (overall, 63.6%, P = 0.96). In summary, although 1mg ECP (with or without progesterone) was less efficacious than 5mg E-17beta plus 100mg progesterone for synchronizing follicular wave emergence, 0.5mg ECP (at CIDR removal or 24h later) induced a synchronous ovulation with an acceptable pregnancy rate to fixed-time AI.     

Effect of progestogen plus estradiol-17beta treatment on superovulatory response in beef cattle

A series of 3 experiments were conducted to evaluate superovulatory response following exogenously controlled follicular wave emergence in cattle. In Experiment 1 the hypothesis was tested that treatments with progestogen plus estradiol-17beta (E-17beta) would result in the emergence of a wave of ovarian follicles that are as responsive to exogenous gonadotropins as those of a spontaneous follicular wave. Beef cows and heifers either received a progestogen ear implant on Day 0 (ovulation) plus 5 mg im E-17beta on Day 1 and were superstimulated on Day 5, or did not receive implants but were superstimulated on Day 8 (expected day of emergence of the second follicular wave). The cattle received 400 mg NIH-FSH-P1 of Folltropin-V, given in a single subcutaneous injection or twice daily as intramuscular injections over 4 d. No significant differences were detected between the 2 groups in the number of corpora lutea (CL), ova/embryos collected, fertilized ova and transferable embryos. In Experiment 2 superstimulatory responses to a single subcutaneous injection of Folltropin-V were compared between heifers in which follicle wave emergence was synchronized with progestogen plus E-17beta at unknown stages of the estrous cycle with those treated following a conventional method of superstimulation at middiestrus. Superstimulation 4 d after E-17beta treatment in heifers with progestogen implants resulted in a similar superovulatory response and higher fertilization rates than those initiated 8 to 12 d after estrus. In Experiment 3 the ovarian response to a single- versus multiple-injection superstimulatory treatment protocol was compared in heifers given progestogen plus E-17beta to induce synchronous wave emergence. The number of CL, ova/embryos collected, fertilized ova and viable embryos were not different between groups. Superstimulatory treatments initiated 4 d after E-17beta treatment of cattle with progestogen implants resulted in comparable ovulatory responses to treatments initiated at the time of spontaneous wave emergence or during middiestrus. Synchronizing wave emergence in a group of randomly cycling cattle obviated the need of estrus detection and synchronization prior to superstimulation.     

The use of estradiol and/or GnRH in a two-dose PGF protocol for breeding management of beef heifers

The objective was to determine reproductive performance following AI in beef heifers given estradiol to synchronize ovarian follicular wave emergence and estradiol or GnRH to synchronize ovulation in a two-dose PGF-based protocol. In Experiment 1, 561 cycling (confirmed by ultrasonography), Angus heifers received 500 microg cloprostenol, i.m. (PGF) twice, 14 days apart (days 0 and 14) and were equally allocated to four groups in a 2 x 2 factorial design. On Day 7, heifers received either 2 mg estradiol benzoate (EB) and 50 mg progesterone (P), i.m. in oil (EBP group) or no treatment (NT group). Half the heifers in each group received 1mg EB, i.m. in oil on Day 15 (24h after the second PGF treatment) with TAI 28 h later (52 h after PGF), and the other half received 100 microg GnRH, i.m. on Day 17 (72 h after PGF) concurrent with TAI. All heifers were observed for estrus twice daily from days 13 to 17; those detected in estrus more than 16 h before scheduled TAI were inseminated 4-16 h later and considered nonpregnant to TAI. Overall pregnancy rate (approximately 35 days after AI) was higher in heifers that received EBP than those that did not (61.6% versus 48.2%, respectively; P < 0.002); but was lower in heifers that received EB after PGF than those that received GnRH (50.0% versus 59.8%; P < 0.02). Although estrus was detected prior to TAI in 77 of 279 heifers (27.6%) treated with EBP (presumably due to induced luteolysis), they were inseminated and 53.2% became pregnant. Overall pregnancy rates were 51.4, 68.3, 45.0, and 55.0% in the NT/GnRH, EBP/GnRH, NT/EB, and EBP/EB groups, respectively (P < 0.05). In Experiment 2, 401 cycling, Angus heifers were used. The design was identical to Experiment 1, except that 1.5mg estradiol-17beta (E-17beta) plus 50mg progesterone (E-17betaP) and 1mg E-17beta were used in lieu of EBP and EB, respectively. All heifers receiving E-17beta 24h after the second injection of PGF (NT/E-17beta and E-17betaP/E-17beta) were TAI 28 h later without estrus detection, i.e. 52 h after PGF. Heifers in the other two groups received 100 microg GnRH, i.m. 72 h after PGF and were concurrently TAI; heifers in these two groups that were detected in estrus prior to this time were inseminated 4-12h later and considered nonpregnant to TAI. Estrus rate during the first 72 h after the second PGF treatment was higher (P < 0.05) in the E-17betaP/GnRH group (45.0%; n = 100) than in the NT/GnRH group (16.0%; n = 100), but conception rate following estrus detection and AI was not different (mean, 57.4%; P = 0.50). Overall pregnancy rate was not significantly different among groups (mean, 46.9%; P = 0.32). In summary, the use of EB or E-17beta to synchronize follicular wave emergence and estradiol or GnRH to synchronize ovulation in a two-dose, PGF-based protocol resulted in acceptable fertility to TAI. However, when 2mg EB was used to synchronize follicular wave emergence, early estrus occurred in approximately 28% of heifers, necessitating additional estrus detection. A combination of estrus detection and timed-AI in a two-dose PGF protocol resulted in highly acceptable pregnancy rates.     

Neither plasma progesterone concentrations nor exogenous eCG affects rates of ovulation or pregnancy in fixed-time artificial insemination (FTAI) protocols for puberal Nellore heifers

The objective was to evaluate the effects of plasma progesterone (P4) concentrations and exogenous eCG on ovulation and pregnancy rates of pubertal Nellore heifers in fixed-time artificial insemination (FTAI) protocols. In Experiment 1 (Exp. 1), on Day 0 (7 d after ovulation), heifers (n = 15) were given 2 mg of estradiol benzoate (EB) im and randomly allocated to receive: an intravaginal progesterone-releasing device containing 0.558 g of P4 (group 0.5G, n = 4); an intravaginal device containing 1 g of P4 (group 1G, n = 4); 0.558 g of P4 and PGF_2α (PGF; 150 μg d-cloprostenol, group 0.5G/PGF, n = 4); or 1 g of P4 and PGF (group 1G/PGF, n = 3). On Day 8, PGF was given to all heifers and intravaginal devices removed; 24 h later (Day 9), all heifers were given 1 mg EB im. In Exp. 2, pubertal Nellore heifers (n = 292) were treated as in Exp. 1, with FTAI on Day 10 (30 to 36 h after EB). In Exp. 3, pubertal heifers (n = 459) received the treatments described for groups 0.5G/PGF and 1G/PGF and were also given 300 IU of eCG im (groups 0.5G/PGF/eCG and 1G/PGF/eCG) at device removal (Day 8). In Exp. 1, plasma P4 concentrations were significantly higher in heifers that received 1.0 vs 0.588 g P4, and were significantly lower in heifers that received PGF on Day 0. In Exp. 2 and 3, there were no significant differences among groups in rates of ovulation (65-77%) or pregnancy (Exp. 2: 26-33%; Exp. 3: 39-43%). In Exp. 3, diameter of the dominant ovarian follicle on Day 9 was larger in heifers given 0.558 g vs 1.0 g P4 (10.3 ± 0.2 vs 9.3 ± 0.2 mm; P < 0.01). In conclusion, lesser amounts of P4 in the intravaginal device or PGF on Day 0 decreased plasma P4 from Days 1 to 8 and increased diameter of the dominant follicle on Day 9. However, neither of these nor 300 IU of eCG on Day 8 significantly increased rates of ovulation or pregnancy.     

Ovarian follicular wave synchronization and induction of ovulation in postpartum beef cows

An experiment was designed to evaluate a) the effect of a progesterone-estradiol combined treatment on ovarian follicular dynamics in postpartum beef cows, and b) ovulation and the subsequent luteal activity after short-term calf removal and GnRH agonist treatment. Multiparous Angus cows (25 to 40 d after calving) were assigned to the following treatments: untreated (Control, n = 9); short term calf removal (CR, n = 8); progesterone (CIDR, n = 9) and progesterone plus estradiol-17 beta (CIDR + E-17 beta, n = 9). Progesterone treatment (CIDR) lasted 8 d and the day of device insertion was considered as Day 0. Cows in the CIDR + E-17 beta group also received an i.m. injection of 5 mg of E-17 beta on Day 1. On Day 8, calves were removed for 48 h (CR, CIDR and CIDR + E-17 beta groups) and 6 h before the end of calf removal these cows also received an i.m. injection of 8 micrograms of Busereline (GnRH). Anestrus was confirmed in all cows by the absence of luteal tissue and progesterone concentrations below 1 ng ml-1 at the beginning of the experiment. Although mean (+/- SEM) interval from the beginning of the experiment (Day 0) to wave emergence did not differ (P > 0.05) among treatment groups (Control, 1.9 +/- 1.0, range -2 to 7 d; CR, 3.9 +/- 0.7, range 0 to 6 d; CIDR, 2.8 +/- 0.5, range 0 to 4 d and CIDR + E-17 beta, 4.1 +/- 0.2, range 3 to 5), the variability was less (P < 0.05) in the CIDR + E-17 beta group. The proportion of cows ovulating 24 to 48 h after GnRH administration tended (P = 0.08) to be higher in cows from CIDR + E-17 beta group (8/9) than in those of CR (5/8) or CIDR (6/9) groups, respectively and was associated with a higher proportion (P < 0.05) of CIDR + E-17 beta treated cows (9/9) that had a dominant follicle in the growing/early static phase at the time of GnRH treatment compared to the other GnRH treated groups (5/8, and 4/9 for CR and CIDR groups, respectively). Two CR cows ovulated 0-24 h after GnRH and only one Control cow ovulated the day before the time of GnRH administration. Cows pretreated with progesterone had longer (P < 0.05) luteal lifespan (CIDR, 14.5 +/- 0.7, CIDR + E-17 beta, 13.9 +/- 0.6 d) than those not treated with CIDR (Control, 5, CR, 4.0 +/- 0.4). We conclude that progesterone plus estradiol treatment results in tightly synchronized wave emergence and high GnRH-induced ovulation rate with normal luteal activity in postpartum beef cattle.     

Induction of follicular wave emergence for estrus synchronization and artificial insemination in heifers

The objective was to synchronize follicular wave emergence among cattle for synchronization of estrus and ovulation, and to determine pregnancy rate after AI at observed estrus. At random stages of the estrous cycle, a controlled internal drug release device (CIDR-B) was inserted intravaginally (Day 0) in 67 cross-bred beef heifers, and they were randomly allocated to receive either no further treatment (Control; n = 18); 5 mg of estradiol-17beta and 100 mg of progesterone im (E/P; n = 16); 100 microg im of GnRH (GnRH; n = 16); or transvaginal ultrasound-guided follicular ablation of all follicles > or = 5 mm (FA; n = 17). All heifers received a luteolytic dose of PGF (repeated 12 h later), and CIDR-B were removed on Days 9, 8, 6 or 5, in Control, E/P, GnRH or FA groups, respectively, so the dominant follicle of the induced wave was exposed to exogenous progesterone for a similar period of time in each group. Mean (+/- SEM) intervals (and range, in days) from treatment to follicular wave emergence in these groups were 3.5 +/- 0.6 (-2 to 8), 3.4 +/- 0.1 (3 to 4), 1.5 +/- 0.3 (-1 to 4), and 1.0 +/- 0.1 (0 to 2), respectively. Although the interval was longest (P<0.01) in the E/P and Control groups, it was least variable (P<0.01) in the E/P and FA groups. Intervals (and range, in days) from CIDR-B removal (and first PGF treatment) to estrus were 2.3 +/- 0.2 (1.5 to 4.5), 2.2 +/- 0.2 (1.5 to 3.0), 2.1 +/- 0.1,(1.5 to 3.5), and 2.5 +/- 0.1 (2.0 to 3.5), and to ovulation were 3.5 +/- 0.2 (2.5 to 5.5), 3.4 +/- 0.1 (3.0 to 4.5), 3.5 +/- 0.1 (2.5 to 4.5), and 3.8 +/- 0.1 (3.0 to 4.5), for Control, E/P, GnRH and FA groups, respectively (ns). The proportion of heifers displaying estrus was higher in the Control than in the FA group (94% versus 65%, P<0.05) and intermediate in EP and GnRH groups (87% and 75%). Heifers were inseminated approximately 12 h prior to ovulation (based on estrous behavior and ultrasound examinations). Pregnancy rates were 78%, 80%, 69% and 65% for Control, E/P, GnRH and FA groups, respectively (P=0.73). Results support the hypothesis that synchronous follicular wave emergence results in synchronous follicle development and, following progesterone removal, synchronous estrus and ovulation with high pregnancy rates to AI. The synchrony of estrus and ovulation in the E/P, GnRH and FA groups suggest that these treatments, in combination with CIDR-B, could be adapted to fixed-time insemination programs.     

The effects of dose and duration of administration of pFSH during the first follicular wave on the ovulation rate of beef heifers

Four experiments were carried out to examine the effects of administration of pFSH (Vetrepharm) from Day 3 of the estrous cycle in conjunction with PG on Day 5 on follicular populations and ovulation rate in heifers. In Experiment 1, 47 heifers were allocated to 1 of 4 treatment groups (n = 11 to 12 per group): a) control, b) 1.5 mg pFSH, c) 2.0 mg pFSH or d) 2.5 mg pFSH until estrus. Heifers assigned to the 3 treatments had a higher ovulation rate than the controls (P < 0.05). In Experiment 2, 45 heifers were allocated to 1 of 5 treatment groups (n = 8 to 10 per group): a) control, b) 1.0 mg pFSH until PG, c) 1.0 mg pFSH until estrus, d) 1.5 mg pFSH until PG or e) 1.5 mg pFSH until estrus. From Day 5, heifers assigned to pFSH treatments had more large follicles than the controls (P < 0.05). There was no effect of treatment on the incidence of twin ovulations. In Experiment 3, 43 heifers were assigned to 1 of 3 groups (n = 11 to 16 per group): a) control, b) 1.0 mg pFSH until estrus or c) 1.5 mg pFSH until estrus. At slaughter, 14 d after administration of PG, the incidence of twin ovulations was 0/11, 7/16 and 8/16 for Groups a, b and c, respectively (P = 0.011). In Experiment 4, pFSH (1.5 mg) was administered to 3 groups during the development of the first dominant follicle: a) growth phase (n = 19); b) static phase (n = 17); and c) decline phase (n = 17). All pFSH-treated heifers had a higher ovulation rate than the controls (P < 0.05); heifers assigned to Group c had a higher ovulation rate than those in Groups a or b (P < 0.05). More heifers assigned to Group c (7/17) superovulated than in the other 2 groups (P < 0.05). In conclusion, administration of 1.0 or 1.5 mg pFSH twice daily beginning at Day 3 of the estrous cycle in association with the induction of luteolysis increased the ovulation rate significantly when pFSH treatment was continued to onset of estrus. The ovulation rate and the occurrence of multiple ovulations were significantly higher when pFSH was administered at the time that the first dominant follicle was in decline.     

Ovarian function in Nelore (Bos taurus indicus) cows after post-ovulation hormonal treatments

Maternal recognition of pregnancy in the cow requires successful signaling by the conceptus to block luteolysis. Conceptus growth and function depend on an optimal uterine environment, regulated by luteal progesterone. The objective of this study was to test strategies to optimize luteal function, as well as prevent a dominant follicle from initiating luteolysis. Nelore (Bos taurus indicus) beef cows (n=40) were submitted to a GnRH/PGF(2alpha)/GnRH protocol. Cows that ovulated from a dominant ovarian follicle (ovulation=Day 0) were allocated to receive: no additional treatment (G(C); n=7); 3000IU of hCG on Day 5 (G(hCG); n=5); 5mg of estradiol-17beta on Day 12 (G(E2); n=6); or 3000IU of hCG on Day 5 and 5mg of estradiol-17beta on Day 12 (G(hCG/E2); n=5). Ultrasonographic imaging of the ovaries, assessment of plasma progesterone concentration, and detection of estrus were done daily from Day 5 to the day of subsequent ovulation. Treatment with hCG induced an accessory CL, increased CL volume, and plasma progesterone concentration throughout the luteal phase (P<0.01). Estradiol-17beta induced atresia and recruitment of a new wave of follicular growth; it eliminated a potentially estrogen-active, growing ovarian follicle within the critical period for maternal recognition of pregnancy, but it also hastened luteolysis (Days 16 or 17 vs. Days 18 or 19 in non-treated cows). In conclusion, the approaches tested enhanced luteal function (hCG) and altered ovarian follicular dynamics (estradiol-17beta), but were unable to extend the life-span of the CL in Nelore cows.     

Resynchronization of previously timed-inseminated beef heifers with progestins

The objective was to determine the efficacy of a previously used CIDR or melengestrol acetate (MGA; 0.5mg/head/day) for resynchronization of estrus in beef heifers not pregnant to timed-AI (TAI). In three experiments and a field trial, heifers were reinseminated 6-12 h after first detection of estrus. Pregnancy diagnosis was done from approximately 25-43 days after either TAI or reinsemination. In Experiment 1, 79 heifers received a once-used CIDR from 13 to 20 days after TAI and 80 heifers were untreated controls. For these two groups, there were 34 and 35 heifers, respectively, not pregnant to TAI; median +/- S.E. intervals from TAI to onset of estrus were 22 +/- 0.2 days versus 20 +/- 0.6 days (P < 0.001); estrus rates were 70.6% versus 85.7% (P = 0.1); conception rates were 62.5% versus 76.7% (P < 0.3); and pregnancy rates were 44.1% versus 65.7% (P = 0.07), for CIDR and untreated (control) groups, respectively. In Experiment 2, heifers (n = 651) were TAI (Day 0) and 13 days later randomly assigned to one of seven groups (n = 93 per group) to receive a once-used CIDR (three groups; Days 13-20), MGA (three groups; Days 13-19), or no treatment (control group). Groups given a CIDR or MGA also received: no further treatment (CIDR or MGA alone); 1.5mg estradiol-17beta (E-17beta) and 50 mg progesterone (P4) in 2 mL canola oil on Day 13; or E-17beta and P4 on Day 13 and 0.5 mg E-17beta on Day 21 (24 h after CIDR removal or 48 h after the last feeding of MGA). Pregnancy rate to TAI was lowest (P < 0.05) for the group given a CIDR plus E-17beta and P4 on Day 13 and E-17beta on Day 21. Variability in return to estrus was greater (P < 0.001) in the control and MGA groups than in CIDR groups. Conception and pregnancy rates in heifers given a CIDR (65.1 and 61.4%) were higher (P<0.01) than those fed MGA (49.6 and 40.4%), but not different from controls (62.2 and 54.9%, respectively). In Experiment 3, 616 heifers received a once- or twice-used CIDR for 7 days, beginning 13+/-1 days after TAI, with or without a concurrent injection of 150 mg of P4 (2 x 2 factorial design). Pregnancy rate to TAI was 47.2%. In heifers that returned to estrus, there was no significant difference between a once- or twice-used CIDR for rates of estrus (68.8%, P < 0.3), conception (65.9%, P < 0.6) and pregnancy (45.3%, P < 0.8). Injecting progesterone at CIDR insertion increased the median interval from CIDR removal to onset of estrus (P < 0.05) and reduced rates of estrus (63.8% versus 73.8%, P<0.05), conception (60.5% versus 70.6%, P = 0.1) and pregnancy (38.6% versus 52.2%, P < 0.02). In a field trial, 983 heifers received a once-used CIDR for 7 days, beginning 13 +/- 1 days after TAI. Pregnancy rate to TAI was 55.2%. The median (and mode) of the interval from CIDR removal to estrus was 2.5 days. Estrus, conception and pregnancy rates were 78.2, 70.3 and 55.0% (overall pregnancy rate to TAI and rebreeding, 78.7%). In summary, a once- or twice-used CIDR for 7 days, starting 13 +/- 1 days after TAI resulted in the majority of nonpregnant heifers detected in estrus over a 4-day interval, with acceptable conception rates; however, injecting progesterone at CIDR insertion significantly reduced both estrus and pregnancy rates, and estradiol treatment after CIDR removal was associated with a decreased pregnancy rate to TAI. Fertility was higher in heifers resynchronized with a once-used CIDR than with MGA.     

Effect of purified llama ovulation-inducing factor (OIF) on ovarian function in cattle

Two experiments were designed to determine the effect of purified ovulation inducing factor (OIF) on ovarian function in cattle. In Experiment 1, prepubertal heifers (n = 11 per group) were treated on Day 5 (Day 0 = day of follicular wave emergence) of the follicular wave with an intramuscular dose of saline (1 mL), GnRH (100 μg), or purified OIF (1 mg/100 kg body weight). Ovulation occurred in 9/11 heifers treated with GnRH, and 1/11 heifers in each of the OIF- and saline-treated groups (P < 0.05). Compared to saline-treated controls, OIF treatment was associated with a smaller dominant follicle diameter (P < 0.01), a rise in plasma FSH concentration (P < 0.1), and earlier emergence of the next follicular wave (P < 0.05). In Experiment 2, sexually mature heifers were given either GnRH or purified OIF on Days 3, 6 or 9 of the first follicular wave (i.e., early growing, early static, or late static phase of the dominant follicle; n = 5 per group per day), or were untreated (n = 10). In heifers treated with OIF on Day 6, the dominant follicle diameter profile tended to be smaller than in controls, and was associated with a rise (P < 0.05) in plasma FSH concentrations. A similar rise in FSH was detected after OIF treatment on Day 9. Compared to untreated controls, treatment with OIF and GnRH was associated with a larger CL diameter (Days 3 and 6 groups; P < 0.05) and a greater concentration of plasma progesterone (Days 6 and 9 groups; P < 0.05). Treatment with purified OIF did not induce ovulation in heifers, but hastened new follicular wave emergence in prepubertal heifers, influenced follicular dynamics in a phase-specific manner in mature heifers, and was luteotrophic.     

Effect of estradiol valerate on ovarian follicle dynamics and superovulatory response in progestin-treated cattle

Three experiments evaluated the effects of estradiol valerate (EV) on ovarian follicular and CL dynamics, intervals to estrus and ovulation, and superovulatory response in cattle. Experiment 1 compared the efficacy of two norgestomet ear implants (Crestar and Syncro-Mate B; SMB) for 9 d (with PGF at implant removal), combined with either 5 mg estradiol-17beta and 100 mg progesterone (EP) or 5 mg EV and 3mg norgestomet (EN) im at the time of implant insertion on CL diameter and follicular wave dynamics. Ovaries were monitored by ultrasonography. There was no effect of norgestomet implant. Diameter of the CL decreased following EN treatment (P < 0.01). Mean (+/- S.D.) day of follicular wave emergence (FWE) was earlier (P < 0.0001) and less variable (P < 0.0001) in EP- (3.6 +/- 0.5 d) than in EN- (5.7 +/- 1.5 d) treated heifers. Intervals from implant removal to estrus (P < 0.001) and ovulation (P < 0.01) were shorter in EN- (45.7 +/- 11.7 and 74.3 +/- 12.6 h, respectively) than in EP- (56.4 +/- 14.1 and 83.3 +/- 17.0 h, respectively) treated heifers. Experiment 2 compared the efficacy of EP versus EN in synchronizing FWE for superovulation in SMB-implanted cows. At random stages of the estrous cycle, Holstein cows (n = 78) received two SMB implants (Day 0) and were randomly assigned to receive EN on Day 0 or EP on Day 1. Folltropin-V treatments were initiated on the evening of Day 5, with PGF in the morning and evening of Day 8, when SMB were removed. Cows were inseminated after the onset of estrus and embryos were recovered 7 d later. Non-lactating cows had more CL (16.7 +/- 11.3 versus 8.3 +/- 4.9) and total ova/embryos (14.7 +/- 9.5 versus 7.9 +/- 4.6) than lactating cows (P < 0.05). EP-treated cows tended (P = 0.09) to yield more transferable embryos (5.6 +/- 5.2) than EN-treated cows (4.0 +/- 3.7). Experiment 3 compared the effect of dose of EV on ovarian follicle and CL growth profiles and synchrony of estrus and ovulation in CIDR-treated beef cows (n = 43). At random stages of the estrous cycle (Day 0), cows received a CIDR and no further treatment (Control), or an injection of 1, 2, or 5 mg im of EV. On Day 7, CIDR were removed and cows received PGF. Follicular wave emergence occurred within 7 d in 7/10 Control cows and 31/32 EV-treated cows (P < 0.05). In responding cows, interval from treatment to FWE was longer (P < 0.05) in those treated with 5 mg EV (4.8 +/- 1.2 d) than in those treated with 1 mg (3.2 +/- 0.9 d) or 2 mg (3.4 +/- 0.8 d) EV, while Control cows were intermediate (3.8 +/- 2.0 d). Diameter of the dominant follicle was smaller (P < 0.05) at CIDR removal and tended (P = 0.08) to be smaller just prior to ovulation in the 5 mg EV group (8.5 +/- 2.2 and 13.2 +/- 0.6 mm, respectively) than in the Control (11.8 +/- 4.6 and 15.5 +/- 2.9 mm, respectively) or 1mg EV (11.7 +/- 2.5 and 15.1 +/- 2.2 mm, respectively) groups, with the 2mg EV group (10.7 +/- 1.5 and 14.3 +/- 1.7 mm, respectively) intermediate. Diameter of the dominant follicle at CIDR removal was less variable (P < 0.01) in the 2 and 5mg EV groups than in the Control group, and intermediate in the 1mg EV group. In summary, treatment with 5mg EV resulted in a longer and more variable interval to follicular wave emergence than treatment with 5mg estradiol-17beta, which affected preovulatory dominant follicle size following progestin removal, and may have also affected superstimulatory response in Holstein cows. Additionally, 5 mg EV appeared to induce luteolysis in heifers, reducing the interval to ovulation following norgestomet removal. Conversely, intervals to, and synchrony of, follicular wave emergence, estrus and ovulation following treatment with 1 or 2 mg EV suggested that reduced doses of EV may be more useful for the synchronization of follicular wave emergence in progestogen-treated cattle.     

Failure of the LH-releasing hormone agonist, deslorelin, to prevent development of a persistent follicle in heifers synchronized with norgestomet

The use of exogenous progestagens for estrus synchronization in cattle can result in a persistent dominant follicle which is associated with reduced fertility. We examined whether the LHRH agonist, deslorelin, would prevent the formation of a persistent follicle in heifers synchronized with norgestomet. The estrous cycles of heifers were synchronized with cloprostenol, and on Day 7 of the ensuing cycle the heifers received one of the following treatments for 10 d: Group C (n = 5), untreated control; Group N (n = 6), injection of a luteolytic dose of cloprostenol on Days 7 and 8 and implant of norgestomet from Day 7 to Day 17 (i.e. typical 10-day norgestomet implant period); Group D (n = 6), injection of cloprostenol on Days 7 and 8 and implants of deslorelin from Day 7 to Day 17; Group ND (n = 6), injections of cloprostenol and both norgestomet and deslorelin implants as above. Follicle growth was monitored using ultrasonography. Group-N heifers showed continued follicle growth and had larger follicles on Day 17 of the cycle than Group-C heifers (16.8 +/- 1.6 and 10.4 +/- 1.6 mm). Follicle growth for Group-D and ND heifers was similar and variable, and seemed to depend on follicle status at the initiation of treatment. Heifers with follicles of 5 to 10 mm (n = 9) in diameter either showed no follicle growth (2 9 ) or developed large follicles (7 9 ), while heifers with follicles approximately 12 mm (n = 3) in diameter showed follicle atresia with no further significant growth. On Day 17, size of the largest follicle was similar for Group-ND (14.3 +/- 2.9) and Group-D (16.8 +/- 1.6) heifers. Heifers in Group N showed estrous behavior 1.8 +/- 0.2 d after treatment, whereas heifers in Groups D and ND did not show estrus for 2 to 4 wk. The results show that combined treatment with progestagen and an LHRH agonist does not consistently prevent the development of a persistent dominant follicle and that return to estrus can be delayed after treatment with an LHRH agonist.     

Estrus synchronization in cattle using estradiol, melengestrol acetate and PGF

In Experiment 1, all cattle were fed MGA (0.5 mg/head/d) for 7 d (designated Days 0 to 6) and given PGF on Day 6. One-half were administered estradiol valerate (EV; 5 mg, im) on Day 0. At Location 1, a higher proportion (P < 0.005) of EV-treated heifers were detected in estrus and bred by AI between Days 7 and 13 than control heifers not receiving EV (27 of 33 versus 15 of 32), but the number of pregnancies (12 vs 10) was not significantly different. Eighty-three of 104 EV-treated and 89 of 106 control cows were inseminated, resulting in 50 and 45 pregnancies, respectively (not significant). At Location 2, cattle were similarly treated and exposed to bulls on Days 7 to 49. Fall pregnancy rate was higher (P < 0.015) for EV-treated than control heifers (44 of 48 vs 33 of 46), but was not significantly different for cows (22 of 26 vs 19 of 23). In Experiment 2, estradiol 17beta (E17beta; 5 mg, im) and progesterone (100 mg, im) were administered on Day 0 (instead of EV). In a third group (designated the PGF group), cattle were bred on Days 0 to 6, and PGF was administered on Day 6 to those not yet bred. For 213 cows, the percentage pregnant to a synchronized estrus was greater in the PGF group (72%) than in either the control group treated with MGA (49%; P = 0.005) or the group receiving MGA and E17beta (54%; P < 0.025). Fall pregnancy rates were 91, 89, and 96% for the 213 cows (not significant) and 89, 93, and 98% for 131 heifers (not significant) in the PGF, MGA and E17beta groups, respectively. In cattle without a functional CL, the average diameter of the largest follicle at Day 6 was 1 to 2 mm smaller in the E17beta + MGA group than in the MGA group (difference significant only in cows at Location 1). Combined for both locations, the synchronized pregnancy rate in heifers without a functional CL on Day 6 was higher (P < 0.05) in the E17beta + MGA group than in the MGA group (11 of 21, 52% versus 4 of 20, 20%). Estrogen treatment caused regression of ovarian follicles with emergence of a new follicular wave. Including estrogen in an estrus synchronization program utilizing MGA and PGF significantly increased fall pregnancy rate in heifers (at 1 location) and the synchronized pregnancy rate of heifers without a functional CL at the time of PGF treatment (combined for both locations).     

Effect of the metabolic environment at key stages of follicle development in cattle: focus on steroid biosynthesis

Cellular mechanisms that contribute to low estradiol concentrations produced by the preovulatory ovarian follicle in cattle with a compromised metabolic status are largely unknown. To gain insight into the main metabolic mechanisms affecting preovulatory follicle function, two different animal models were used. Experiment 1 compared Holstein-Friesian nonlactating heifers (n = 17) and lactating cows (n = 16) at three stages of preovulatory follicle development: 1) newly selected dominant follicle in the luteal phase (Selection), 2) follicular phase before the LH surge (Differentiation), and 3) preovulatory phase after the LH surge (Luteinization). Experiment 2 compared newly selected dominant follicles in the luteal phase in beef heifers fed a diet of 1.2 times maintenance (M, n = 8) or 0.4 M (n = 11). Lactating cows and 0.4 M beef heifers had higher concentrations of β-hydroxybutyrate, and lower concentrations of glucose, insulin, and IGF-I compared with dairy heifers and 1.2 M beef heifers, respectively. In lactating cows this altered metabolic environment was associated with reduced dominant follicle estradiol and progesterone synthesis during Differentiation and Luteinization, respectively, and in 0.4 M beef heifers with reduced dominant follicle estradiol synthesis. Using a combination of RNA sequencing, Ingenuity Pathway Analysis, and qRT-PCR validation, we identified several important molecular markers involved in steroid biosynthesis, such as the expression of steroidogenic acute regulatory protein (STAR) within developing dominant follicles, to be downregulated by the catabolic state. Based on this, we propose that the adverse metabolic environment caused by lactation or nutritional restriction decreases preovulatory follicle function mainly by affecting cholesterol transport into the mitochondria to initiate steroidogenesis.     

The dominant follicle exerts an interovarian inhibition on FSH-induced follicular development

An experiment was conducted to evaluate the role of the dominant follicle (DF) of the first wave in regulating follicular and ovulatory responses and embryonic yield to a superovulation regime with FSH-P. Twenty normally cycling Holstein-Freisian heifers (n = 20) were synchronized with GnRH and pgf(2alpha) and randomly assigned to a control or a treated group (n = 10 each). Treated heifers had the first wave dominant follicle removed via transvaginal, ultrasound-guided aspiration on Day 6 after a synchronized estrus. All heifers received a total of 32 mg FSH-P given in decreasing doses at 12 h intervals from Day 8 to Day 11 plus two injections of pgf(2alpha) (35 mg and 20 mg, respectively) on Day 10. Heifers were inseminated at 6 h and 16 h after onset of estrus. Follicular dynamics were examined daily by transrectal ultrasonography from Day 4 to estrus, once following ovulation, and at the time of embryo collection on Day 7. Blood samples were collected daily during the superovulatory treatment and at embryo collection. Follicles were classified as: small, /= 10 mm. Aspiration of the dominant follicle was associated with an immediate decrease in large follicles, and a linear rate increase in small follicles from Day 4 to Day 8 just prior to the FSH-P injections, (treatment > control: +0.33 vs. -0.22, number of small follicles per day; P < 0.10). During FSH-P injections, the increase in number of medium follicles was greater (P < 0.01) for treatment on Day 9-11 (treatment > control: Day 9, 3.2 > 1.8; Day 10, 9.2 > 4.7; Day 11, 13.1 > 8.3; +/- 0.56). Number of large follicles was greater in treatment at Day 11 (5.12 > 1.4 +/-0.21; P < 0.01). Mean number of induced ovulatory follicles (difference between number of follicles at estrus and Day 2 after estrus) was greater in treatment (13.4 > 6.3 +/- 1.82; P < 0.01). Plasma estradiol at Day 11 during FSH-P treatment was greater in treatment (32.5 > 15.8 +/- 2.6; P < 0.01). Plasma progesterone at embryo flushing (Day 7 after ovulation) was greater in treatment (7.4 > 4.9; P < 0.02); technical difficulties at embryo recovery reduced sensitivity of embryonic measurements. No changes in the distribution of unfertilized oocytes and embryo developmental stages were detected between control and treatment groups. Presence of dominant follicle of the first wave inhibited intraovarian follicular responses to exogenous FSH.     

Comparison of three methods of acute administration of progesterone on ovarian follicular development and the timing and synchrony of ovulation in Bos indicus heifers

The aim of this study was to induce the formation of a persistent dominant ovarian follicle and to compare the effects of 3 methods of acute administration of P4 on ovarian follicular development and on the timing and synchrony of ovulation. Stage of the estrous cycle was initially synchronized in Bos indicus heifers with a norgestomet implants (3 mg) for 10 d and with an analogue of PGF2 alpha (15 mg) on the first and last day of norgestomet treatment. Eight days after removal of the implants, heifers were randomly assigned to 4 groups. All heifers received a norgestomet implant (Day 0), which was removed 17 d later (Day 17); PGF2 alpha was administered on Days 0 and 4. Heifers in the control group (n = 5) received no other treatment. On Day 10 heifers in Group P4C (n = 5) were treated with a CIDR for 24 h; heifers in Group P4O (n = 5) were administered 100 mg i.m. of P4 in oil, while heifers in Group P4S (n = 5) were administered 100 mg i.m. of P4 in saline/alcohol. Data were analyzed using bootstrap estimates of location (mean) and spread (standard deviation; SD). Compared with the control heifers, day of emergence of the ovulatory follicle was delayed, and age and duration of dominance of the ovulatory follicle were reduced in the P4C and P4O heifers (P < 0.05) but not in the P4S heifers (P > 0.05). In all groups treated with P4 both the mean and variability (SD) in the timing of ovulation did not differ with that of the control group (P > 0.05) but there was less variability in the day of emergence, age, duration of dominance and diameter of the ovulatory follicle than in the control group (P < 0.05). Delayed timing and reduced synchrony (SD) of ovulation and greater age of the ovulatory follicle (P < 0.05) occurred in P4S heifers than in P4C heifers. We conclude that administration of 100 mg of P4 in oil is as effective as treatment with a CIDR for synchronizing emergence and ovulation of a newly recruited dominant follicle. However, reduced synchrony of ovulation, greater age of the ovulatory follicle and delayed timing of ovulation occurred following administration 100 mg of P4 in saline/alcohol compared with the CIDR device.     

Will the first dominant follicle of the estrous cycle of heifers ovulate following luteolysis on day 7?

The pattern of turnover of dominant follicles involves the sequential growth and regression of two to three dominant follicles during the estrous cycle. The dominant follicle that ovulates is the one that develops concomitantly with the regression of the corpus luteum. The aim of this paper was to determine if the first dominant follicle would ovulate following induction of luteolysis with prostaglandin F2 alpha analogues (PGF) on Day 7 of the cycle. Heifers (n = 43) were checked for estrus (Day 0); their ovaries were scanned daily from Day 6 of the cycle for one week, and the fate of the first dominant follicle was determined. Luteolysis was induced on Day 7 with PGF analogues, and blood samples were taken daily for progesterone and estradiol measurement and at 3-h intervals for 33 h for luteinizing hormone (LH) measurement. Of the 43 heifers given PGF, complete luteolysis occurred in 40 animals. Of these, the first dominant follicle ovulated in 37 heifers; the dominant follicle was not the ovulatory follicle in 2 heifers and the dominant follicle became cystic in one heifer.     

Endocrine and ovarian responses associated with the first-wave dominant follicle in cattle.

To examine endocrine and biochemical differences between dominant and subordinate follicles and how the dominant follicle affects the hypothalamic-pituitary-ovarian axis in Holstein cows, the ovary bearing the dominant follicle was unilaterally removed on Day 5 (n = 8), 8 (n = 8), or 12 (n = 8) of synchronized estrous cycles. Follicular development was followed daily by ultrasonography from the day of detected estrus (Day 0) until 5 days after ovariectomy. Aromatase activity and steroid concentrations in first-wave dominant and subordinate follicles were measured. Intact dominant and subordinate follicles were cultured in 4 ml Minimum Essential Medium supplemented with 100 microCi 3H-leucine to evaluate de novo protein synthesis. Five days after unilateral ovariectomy, cows were resynchronized and the experiment was repeated. Follicular growth was characterized by the development of single large dominant follicles, which was associated with suppression of other follicles. Concentrations of estradiol-17 beta (E2) in follicular fluid and aromatase activity of follicular walls were higher in dominant follicles (438.9 +/- 45.5 ng/ml; 875.4 +/- 68.2 pg E2/follicle) compared to subordinate follicles (40.6 +/- 69.4 ng/ml; 99.4 +/- 104.2 pg E2/follicle). Aromatase activity in first-wave dominant follicles was higher at Days 5 (1147.1 +/- 118.1 pg E2/follicle) and 8 (1028.2 +/- 118.1 pg E2/follicle) compared to Day 12 (450.7 +/- 118.1 pg E2/follicle). Concentrations of E2 and androstenedione in first-wave dominant follicles were higher at Day 5 (983.2 +/- 78.2 and 89.5 +/- 15.7 ng/ml) compared to Days 8 (225.1 +/- 78.6 and 5.9 +/- 14.8 ng/ml) and 12 (108.5 +/- 78.6 and 13.0 +/- 14.8 ng/ml). Concentrations of progesterone in subordinate follicles increased linearly between Days 5 and 12 of the estrous cycle. Plasma concentrations of FSH increased from 17.9 +/- 1.4 to 32.5 +/- 1.4 ng/ml between 0 and 32 h following unilateral removal of the ovary with the first-wave dominant follicle. Increases in plasma FSH were associated with increased numbers of class 1 (3-4 mm) follicles in cows that were ovariectomized at Day 5 or 8 of the cycle. Unilateral ovariectomy had no effects on plasma concentrations of LH when a CL was present on the remaining ovary. First-wave dominant follicles incorporated more 3H-leucine into macromolecules and secreted high (90,000-120,000) and low (20,000-23,000) molecular weight proteins that were not as evident for subordinate follicles at Days 8 and 12.(ABSTRACT TRUNCATED AT 400 WORDS)     

Alterations in follicular estradiol and gonadotropin receptors during development of bovine antral follicles

It was hypothesized that growth divergence of dominant and subordinate follicles during Wave 1 and growth termination of the dominant follicle would be associated with changes in the number of gonadotropin receptors on granulosa cells and estradiol in follicular fluid. To test this hypothesis, follicular development of 16 Holstein heifers was monitored by ultrasound, and follicles were collected on Days 2,4,6 and 10 (Day 0 = ovulation). Dominant follicles were compared across days, whereas dominant and largest subordinate follicles were compared on Days 2 and 4 only. The numbers of LH and FSH receptors on the granulosa cells of dominant follicles did not differ significantly over Days 2, 4, 6 and 10. In contrast, concentrations of estradiol in follicular fluid decreased (P < 0.05) from Days 2 to 10 (373 +/- 150 to 42 +/- 12 ng/ml) and concentrations of progesterone in follicular fluid increased (P < 0.05) from Days 2 to 10 (12.2 +/- 2.3 to 24.4 +/- 4.8 ng/ml). Correspondingly, the ratio of estradiol:progesterone in the dominant follicles decreased (P < 0.05) from Days 2 to 10. Comparisons between dominant and subordinate follicles indicated greater (P < 0.05) estradiol concentrations in the dominant follicle on Day 2, but the number of gonadotropin receptors was not different until Day 4. Thus, differences in concentrations of follicular fluid estradiol, but not numbers of granulosa cell gonadotropin receptors, were associated with the early growth divergence of dominant and subordinate follicles (Day 2) and the eventual growth termination of the dominant follicle (Day 10). Late divergence (Day 4) was associated with higher gonadotropin receptor numbers and follicular estradiol concentrations in the dominant than in the subordinate follicles. These results indicate that an increase in estradiol productivity of the selected dominant follicle occurred before an increase in the number of gonadotropin receptors.