Prostaglandin F2α Treatment Research Articles

101 Treatment with gonadotrophin-releasing hormone on Day 7 or 21 does not reduce pregnancy loss in dairy heifers receiving invitro-produced embryos

Previous research has shown that induction of an accessory corpus luteum (CL) by administration of gonadotrophin-releasing hormone (GnRH) on Day 5 increases circulating progesterone and reduces pregnancy loss between Days 33 and 60 in heifers receiving invitro-produced (IVP) embryos. Therefore, the objective of the present study was to determine whether timing of induction of an accessory CL influenced pregnancy loss in IVP recipients. Holstein heifers (n=1,658) were synchronized using a modified 5-day CIDR CO-Synch protocol. Briefly, heifers received an intravaginal progesterone device (CIDR) on Day −8, Day −3: CIDR removed and first prostaglandin F2α (PGF2α) treatment, Day −2: second PGF2α, and Day 0: GnRH (G1, 100µg of gonadorelin acetate). Oestrus expression was evaluated on Day 0 with the use of an oestrus detection device. Transfer of fresh IVP embryos was performed on Day 7±1 after G1. At the time of transfer, heifers were randomly assigned, in a 2×2 factorial design, to receive GnRH (200µg of gonadorelin acetate) or remain as Control (untreated) at two different times: Day 7 (time of embryo transfer (ET)) or Day 21. Thus, the following groups were formed: Control-Control (n=410); Control-GnRH (n=409); GnRH-Control (n=419); and GnRH-GnRH (n=420). All heifers were evaluated by transrectal ultrasonography to determine number, size, and location of CL on Days 5, 14, 21, 28, 32, and 60 and pregnancy status on Days 28, 32, and 60. Data collected from each heifer included embryo stage and quality, oestrus expression, body condition score, number of transfers, and technician. Pregnancies per embryo transfer (P/ET) and pregnancy loss data were analysed by logistic regression (SAS 9.4). Ovulation to Day 7 treatment was greater (P<0.01) in GnRH treated heifers (70.0%; 585 out of 836) than in untreated Controls (2.7%; 22 of 819). Ovulation to Day 21 treatment was determined only in animals pregnant at Day 28 and was greater (P<0.01) in heifers treated with GnRH (37.6%; 153 of 407) than in untreated controls (1.0%; 4 of 390). There was no effect of Day 7 treatment (P>0.68) or Day 21 treatment (P>0.18), nor a Day 7×Day 21 treatment interaction (P>0.48) on P/ET at Day 32 or 60 (Table 1). Treatment with GnRH on Day 7 or 21 did not alter pregnancy loss between Days 32 and 60 (P>0.10). Heifers with an accessory CL present at Day 32 (11.7%; 41 of 350) had similar (P=0.55) pregnancy loss compared with heifers with no accessory CL (14.4%; 54 of 375). The number of CL present on Day 32 did not affect (P=0.23) pregnancy loss; however, heifers with 3 CL (4.9%; 3 of 61) tended (P<0.10) to have reduced pregnancy loss compared with heifers with 1 CL (14.4%; 54 of 375) or 2 CL (13.2%; 38 of 289). In conclusion, treatment with GnRH on Day 7 or 21 induced an accessory CL; however, it did not affect P/ET on Days 32 or 60 and pregnancy loss. Table 1.Pregnancies per embryo transfer (P/ET) and pregnancy loss in embryo recipients based on timing of treatment with gonadotrophin-releasing hormone (GnRH) Treatment P/ET Day 32% (n) P/ET Day 60% (n) Pregnancy loss% (n) Day 7 Day 21 Control Control 42.4 (410) 36.1 (410) 15.0 (174) GnRH 46.0 (409) 40.7 (409) 11.7 (188) GnRH Control 43.7 (419) 36.8 (419) 15.9 (183) P-value GnRH 43.1 (420) 38.8 (420) 10.0 (181) Day 7 treatment 0.68 0.72 0.96 Day 21 treatment 0.51 0.18 0.11 Interaction 0.48 0.66 0.71

Hormonal mechanisms regulating follicular wave dynamics II: Progesterone decreases diameter at follicle selection regardless of whether circulating FSH or LH are decreased or elevated

Selection of a single dominant follicle is morphologically manifested by diameter deviation between the future dominant follicle (F1) and the future largest subordinate follicle (F2). Conventional deviation is defined as F2≥7 mm when F1 reaches ∼8.5 mm whereas, undersized deviation is if F2<7 mm when F1 reaches ∼8.5 mm. Greater frequency of undersized deviation has been temporally associated with greater circulating progesterone (P4) and greater FSH but reduced LH in observational studies. Experiment 1 was conducted to directly test if elevating P4 increased the likelihood of undersized deviation and altered circulating concentrations of LH and FSH. Experiment 2 was conducted to test if increasing LH action by treatment with exogenous porcine LH or human chorionic gonadotropin (hCG) in the presence of elevated P4, would stimulate growth of F2 and increase the likelihood of conventional deviation. Ovaries were evaluated by ultrasound and blood samples collected every 12 h after development of a new wave following follicle ablation on D6 (D0 = ovulation). Data were normalized to F1≥7.5 mm and compared using SAS software. In experiment 1 (n = 20), the CL was regressed by prostaglandin F2α treatment and heifers were randomized on D6 into control (no P4 treatment) or P4 treatment (75 mg every 12 h for 5.5 d) beginning when F1 reached ∼3 mm (P4-3 mm group) or ∼6 mm (P4-6 mm group). The P4 treatment significantly increased the frequency of undersized deviation from 0% (controls) to 54%, decreased LH by 44%, and increased FSH by 32%. In experiment 2 (n = 27) heifers were randomized on D6 into control (saline) or treatment with the LH analogs - pLH (1.25 mg porcine LH/12 h) or hCG (160 IU initially and subsequently 96 IU/24 h).Treatment with LH analogs significantly increased P4 (control, 4.6 ± 0.3 ng/mL; pLH, 6.6 ± 0.4 ng/mL; and hCG, 8.9 ± 0.4 ng/mL) and decreased FSH (control, 0.46 ± 0.03 ng/mL; combined-pLH/hCG, 0.34 ± 0.02 ng/mL). However, F1 and F2 diameter and frequency of conventional (37%) and undersized (48%) deviations were similar between the control and combined-pLH/hCG groups. In conclusion, elevated P4 was directly linked to undersized deviation but the P4 effect on decreasing F2 diameter occurred independently of the P4 effects on FSH and LH concentrations.

A simple and practical short-term timed artificial insemination protocol using estradiol benzoate with prostaglandin F2α in lactating dairy cows

To minimize costs and labor for short-term ovulation synchronization protocol, we developed one wherein each treatment—drug administration and timed artificial insemination (TAI)—was performed 24 h apart. The objective of the present study was to evaluate this short-term ovulation synchronization protocol in lactating dairy cows. Data were derived from 133 inseminations performed in 120 cows (32 primiparous and 88 multiparous), and the ovaries of these cows were scanned using ultrasound. The cows detected to have a functional corpus luteum (CL) received prostaglandin F2α (PGF) as a luteolytic agent. The cows were randomly assigned to two treatment groups: (1) treatment with estradiol benzoate (EB) 24 h after PGF treatment, and TAI 24–28 h after EB treatment (EB group); and (2) treatment with gonadotropin-releasing hormone agonist (GnRH) 56 h after PGF treatment, and TAI 16–20 h after GnRH treatment (GnRH group). As a luteolytic agent, either dinoprost (DP; 25 mg) or D-cloprostenol (DCLP; 0.15 mg) was administered intramuscularly in each treatment group. Pregnancy per AI (P/AI) was significantly higher in the DP- or DCLP-treated cows in the EB group when compared with their counterparts in the GnRH group (64.5% vs. 33.3%, P = 0.03 in the DP-treated cows and 51.1% vs. 27.3%, P = 0.04 in the DCLP-treated cows, respectively). Regarding parity, multiparous cows had greater P/AI in the EB group than in the GnRH group (52.8% vs. 26.7%, P = 0.01), whereas primiparous cows showed no significant intergroup difference (65.2% vs. 41.7%, P = 0.28). To conclude, the use of a convenient synchronization protocol comprising the administration of PGF and EB 24 h apart, rather than PGF and GnRH 56 h apart, has greater potential to improve pregnancy rates after TAI in lactating dairy cows given that a functional CL was accurately detected. This beneficial effect of the protocol using EB was clearly demonstrated in multiparous cows.

118 A retrospective analysis of follicular dynamics, LH, estradiol-17β, and progesterone in prostaglandin F2α-induced estrus of Beetal goats

The objective of the study was to reveal the factors associated with variable interval to ovulation following a single dose of prostaglandin F2α (PGF2) in Beetal goats. In experiment 1, the extent of ovulation time was demonstrated after synchronizing Beetal goats (n=38) by using a single injection of PGF2 based on corpus luteum detection by B-mode transrectal ultrasonography. Ovulations were detected every 6h from PGF2 until ovulation. To explore the changes in follicular dynamics, endocrine profiles and behavioural events due to the variation in the interval to ovulation, another set of goats (n=13; experiment 2) were synchronized by a single dose of PGF2 given at random days of the luteal phase. Follicular dynamics were monitored every 12h from PGF2 until ovulation and oestrus was detected by apronized bucks every 6h from PGF2 treatment. Blood sampling (3mL) for oestradiol-17β and progesterone was done every 12h from PGF2 until ovulation and every 2h for LH assay by radioimmunoassay. Data were analysed by Student’s t-test, general linear models, and Pearson coefficient correlation (SPSS, version 20.0, SPSS Inc., Chicago, IL, USA). Results revealed that ovulation time varied from 60 to 96h after PGF2, and 57% of ovulations occurred by 72h after PGF2 (experiment 1). Based on this observation, goats (n=13) in second experiment were retrospectively divided into early and late ovulators (i.e. 72 and 84h following PGF2, respectively). Onset of oestrus, peak oestradiol-17β concentration, and LH surge after PGF2 were first observed in early than late ovulators (P&amp;lt;0.05). The goats having greater follicle diameter and smaller corpus luteum diameter at the time of PGF2 ovulated earlier than late ovulators (5.4±0.2v. 4.3±0.2mm and 10±0.6v. 11.8±0.3mm, respectively; P&amp;lt;0.05). Likewise, plasma progesterone concentration tended to be lower (P=0.087) in early than late ovulators. However, interval from onset of oestrus to peak LH and ovulation, duration of standing oestrus, interval from LH peak to ovulation, diameter and growth rate of preovulatory follicles, and number of ovulations were similar (P&amp;gt;0.05) between early and late ovulators. In conclusion, the size of dominant follicle and corpus luteum at the time of PGF2 determines the interval to ovulation following a single dose of PGF2 during the luteal phase.

49 Collection of Day 7 equine embryos in aluteal cycles in mares

Serial administration of prostaglandin F2α (PGF2) in early diestrus has been recently shown to prevent luteal function in mares and is associated with delayed embryo development in induced aluteal cycles (progesterone deprived environment; &amp;lt;1.0ng mL−1; Leisinger et al. 2018 Theriogenology 105, 178-183). We hypothesised that embryos collected on Day 7 during induced aluteal cycles would be developmentally delayed compared with those collected from control cycles (progesterone deprived environment but supplemented with exogenous progestogen). Mares were monitored until a preovulatory follicle=35mm and the presence of uterine oedema were detected by ultrasonography. Mares were treated IV once with 2000IU of human chorionic gonadotropin and artificially inseminated every other day with total motile spermatozoa from one stallion of known fertility until ovulation. Mares were examined twice daily to determine the occurrence of ovulation. After ovulation, mares were randomly assigned to the control group (serial PGF2 treatment+long-acting altrenogest) or AL group (serial PGF2 treatment only). Using a protocol to induce aluteal cycles (Leisinger et al. 2018 Theriogenology 105, 178-183), mares in the AL group (n=7) were treated twice daily with 10mg of PGF2 (Lutalyse, dinoprost tromethamine, Zoetis, Florham Park, NJ, USA) IM on Days 0, 1, 2, and then once daily on Days 3 and 4. Mares in the control group (n=4) were treated with serial PGF2 treatment as mentioned before, and treated with a single injection of 225mg altrenogest (BioRelease Altrenogest LA 150; BET Pharm, Lexington, KY, USA) IM at the time of ovulation. On Day 7 post-ovulation, embryo collection was performed by uterine flushing using lactated Ringer’s solution. The developmental stage of embryos, diameter, and quality were determined using a stereomicroscope and photographed. Embryos were washed 3 times in commercial embryo holding medium (EmCare™ Holding Solution, ICP Bio, Spring Valley, WI) and stained with 1µg mL−1 of 4’,6-diamidino-2-phenylindole at 38.5°C for 15min to determine the number of nonviable cells. Data were analysed using t-test or Mann-Whitney U test where appropriate. Statistical significance was set at P=0.05. Data are reported as mean±s.e.M. Overall, the developmental stage of control embryos differed from AL embryos (P&amp;lt;0.03). In the control group, all embryos (n=4) collected were classified as expanded blastocysts. In contrast, embryos (n=7) collected from the AL group were all early blastocysts. Furthermore, the mean embryo diameter differed in control v. AL groups, 401±8mm v. 166±2mm, respectively (P&amp;lt;0.001). The quality grades of control embryos (1.3±0.3) were not different from the AL group (1.3±0.2; P&amp;gt;0.05). All embryos of both groups had less than 10% of blastomeres stained with 4’,6-diamidino-2-phenylindole. This study showed that morphologically normal equine embryos of acceptable quality can be collected on Day 7 in aluteal cycles.

Second prostaglandin F2α treatment during Ovsynch protocol does not improve fertility outcomes in dairy cows.

The objective of this study was to evaluate the effect of a second prostaglandin F2α (PGF2α) treatment during Ovsynch on luteal regression and fertility in dairy cows, compared with standard Ovsynch. The study was conducted on 111 Holstein Friesian multiparous cows on commercial dairy farm. The cows in the experimental group (n=48) received two treatments of PGF2α 24 hours apart during Ovsynch. The cows in the control group (n=63) were synchronized with standard Ovsynch. To assess the progesterone (P4) concentration blood samples were collected at the day of PGF2α treatment and at the 2nd GnRH treatment. Pregnancy was evaluated by ultrasound examination 37-40 days after timed artificial insemination (TAI) by ultrasound. Cows diagnosed pregnant were re-examined between days 70-80 after TAI. The percentage of cows with complete corpus luteum (CL) regression (P4⟨0.5 ng/ml at the time of the 2nd GnRH treatment) was 89.6 % after two PGF2α treatments and 88.9 % after one PGF2α treatment. There were no statistically significant differences (p>0.05) in the pregnancies per artificial insemination (P/AI) between the experimental and control group (P/AI). However, the pregnancy loss rate was lower in cows receiving two PGF2α treatments than in the control animals (0.0 % vs. 6.4 %; p⟨0.05). In conclusion, the second PGF2α treatment during Ovsynch protocol had no significant effect on CL regression and P/AI in dairy cows. The pregnancy losses until days 75-80 after TAI were significantly lower after two PGF2α treatments than after one PGF2α treatment.

Effect of dose and timing of prostaglandin F2α treatments during a Resynch protocol on luteal regression and fertility to timed artificial insemination in lactating Holstein cows

Our objective was to evaluate the effect of a second PGF2α treatment (25 mg of dinoprost) or a double dose of PGF2α (50 mg of dinoprost) during a Resynch protocol on luteal regression and pregnancies per artificial insemination (P/AI) in lactating dairy cows. Lactating Holstein cows (n = 1,100) were randomly assigned at a nonpregnancy diagnosis to receive (1) Ovsynch (control: 100 µg of GnRH; 7 d, 25 mg of PGF2α; 56 h, 100 µg of GnRH), (2) Ovsynch with a second PGF2α treatment (GPPG: 100 µg of GnRH; 7 d, 25 mg of PGF2α; 24 h, 25 mg of PGF2α; 32 h, 100 µg of GnRH), or (3) Ovsynch with a double dose of PGF2α (GDDP: 100 µg of GnRH; 7 d, 50 mg of PGF2α; 56 h, 100 µg of GnRH). All cows received timed artificial insemination (TAI) approximately 16 h after the second GnRH treatment (G2). Pregnancy diagnosis was performed by transrectal palpation 39 ± 3 d after TAI, and pregnancy status was reconfirmed 66 d after TAI. Blood samples collected from a subset of cows in each treatment at the first PGF2α treatment (n = 394) and at G2 (n = 367) were assayed for progesterone (P4). Data were analyzed by logistic regression using the GLIMMIX procedure of SAS (SAS Institute Inc., Cary, NC). At 39 d after TAI, GPPG cows tended to have more P/AI than control cows [35% (137/387) vs. 31% (107/349)], whereas P/AI for GDDP cows [32% (118/364)] did not differ from that for control cows. Pregnancy loss from 38 to 66 d did not differ among treatments and was 8% (30/362). The percentage of cows with complete luteal regression (P4 <0.4 ng/mL at G2) tended to differ among treatments and was greater for GPPG cows than for GDDP and control cows (94% vs. 88% vs. 88%, respectively). Overall, cows with P4 <1 ng/mL at the first PGF2α treatment had fewer P/AI than cows with P4 ≥1 ng/mL (27% vs. 38%), whereas cows with P4 ≥0.4 ng/mL at G2 had fewer P/AI than cows with P4 <0.4 ng/mL (15% vs. 38%). We conclude that adding a second PGF2α treatment 24 h after the first within a Resynch protocol tended to increase the proportion of cows undergoing complete luteal regression and P/AI, whereas treatment with a double dose of PGF2α at a single time did not.

Prostaglandin F2α-induced luteolysis involves activation of Signal transducer and activator of transcription 3 and inhibition of AKT signaling in cattle

Prostaglandin F2α (PGF) induces the precipitous loss of steroidogenic capabilities and cellular death in the corpus luteum of many species, yet the molecular mechanisms underlying this event are not completely understood. Signal transducer and activator of transcription 3 (STAT3) was activated in granulosa cells during follicle atresia, whereas AKT is immediately down-regulated in the corpus luteum after PGF treatment in cattle; however, their involvement in both functional and morphological luteolysis in monovular species still need to be determined. Blood samples and corpus lutea were collected from cows before (0) and 2, 12, 24, and 48 hr after PGF treatment on Day 10 of the estrous cycle (4-5 cows per time point). Serum progesterone concentrations decreased by threefold (p < 0.05) within 2 hr, confirming functional luteolysis. The mRNA abundance of the pro-apoptotic gene BAX increased 12-48 hr post-PGF treatment (p < 0.05), while morphological luteolysis was observed 24 and 48 hr after PGF treatment, based on the loss of plasma membrane integrity, reduction of cytoplasmic volume, and pyknotic nuclei. Phosphorylated STAT3 increased, peaking at 12 hr, and remained elevated until 48 hr after PGF treatment. SOCS3 transcript abundance also increased (p < 0.05) starting at 2 hr post-PGF treatment. In contrast, AKT phosphorylation decreased by 12 hr after treatment. Thus, activation of STAT3 and inactivation of AKT signaling are involved in structural regression of the corpus luteum.

Purulent vaginal discharge in grazing dairy cows: Risk factors, reproductive performance, and prostaglandin F2α treatment

The objectives of this study were to assess the association of a 4-point scale of vaginal discharge score (VDS) with time to pregnancy to define criteria for a practical case of purulent vaginal discharge (PVD) in dairy cows, to test the risk factors for PVD, and, finally, the effect of a dose of PGF2α on cure and reproductive performance. In experiment 1, grazing Holstein cows (n = 2,414) had their vaginal discharge scored at ∼32 d in milk (DIM) on a 4-point scale, the effect of VDS on the hazard of pregnancy by 300 DIM was then assessed to derive a case definition of PVD. Risk factors for PVD and self-cure were also assessed. In experiment 2, grazing Holstein cows (n = 6,326) from 5 herds were checked for PVD at ∼30 DIM. Cows with PVD were assigned to receive one dose of 500 μg of PGF2α analog (Cloprostenol; Ciclase, Syntex SA, Buenos Aires, Argentina) per cow (odd ear tag number) or to remain untreated (even tag number). Cure was declared if cows presented clear normal vaginal discharge (VDS-0) at visit 2 (∼62 DIM). Data were analyzed with Cox's regression and mixed logistic models. In experiment 1, cows with VDS ≥1 had lower hazard of pregnancy and longer calving to pregnancy interval than cows with VDS-0. This finding was not affected by the time at which the diagnosis was performed. Therefore, a cow ≥21 DIM and having VDS ≥1 was used to define a case of PVD. The odds of PVD were greater in primiparous cows compared with multiparous, in cows with abnormal calving compared with those with normal calving, and in those losing BCS peripartum. In experiment 2, PGF2α treatment tended to slightly increase the hazard of pregnancy (adjusted hazard ratio = 1.13). Conversely, PGF2α had no effect on the odds of cure of PVD [adjusted odds ratio (AOR) = 1.19], pregnancy at first service (AOR = 1.03), or pregnancy by 100 DIM (AOR = 0.89) or 200 DIM (AOR = 1.27). In conclusion, cows with VDS ≥1 can be considered to have PVD because of their lower hazard of pregnancy and longer calving to pregnancy interval (up to 48 d). Important risk factors are parity, calving, and body condition score loss peripartum. Optimal time of diagnosis is ≥28 to 35 DIM because cows experience a high self-cure rate. Self-cure is also affected by parity, prepartum BCS, and VDS. Finally, as treatment with one dose of PGF2α had a small effect on the hazard of pregnancy and no effect on clinical cure, its therapeutic use in grazing dairy cows with PVD is not recommended.

CIDR and Ultrasound-Fetometry Scan; Comparative Study Between Sheep and Goat in Egypt

The present study aims to evaluate the impact of controlled internal drug release (CIDR) in combination with prostaglandin F2α (PGF2α) and pregnant mare serum gonadotropin (PMSG) treatments on reproductive performance of dairy ewes and does besides, evaluation the accuracy of pregnancy diagnosis during the breeding season. Twenty female animals divided as Ossimi ewes (n=10) and Baladi does (n=10) were subjected to ovsynch regimen. Ultrasonographic examinations were conducted weekly using real-time B-mode ultrasound scanner by transrectal linear and transabdominal convex probes starting from day 14 post service. This study showed that estrus response was 100% in both ewes and does. The conception rates were 90% in ewes and 80 % in does. The incidence of twining was higher in ewes when compared to that in does with incidence rate 67% and 50 in ewes and does, respectively. The accuracy of ultrasound examination for determining fetal number was 100% for single and twin pregnancy. The birth rate was 100 % for both species; and this was confirmed at lambing / kidding. In conclusion, our results indicate that using biotechnologies such as ovsynch regimen and pregnancy diagnosis by ultrasonography are useful to improve the management of reproduction in ewes and does.

101 DOSE AND TIMING OF ADMINISTRATION OF PROSTAGLANDIN F2α DURING FIXED-TIME EMBRYO TRANSFER IN AN IN VITRO-PRODUCTION PROGRAM

Synchronization protocols for fixed-time embryo transfer (ET) contribute significantly to the overall cost of an in vitro-produced-ET program, primarily through the cost of drugs and the labour required. Optimization of synchronization protocols to reduce cost, while providing high fertility, have the potential to improve overall efficiency and profitability. The objective of the present study was to evaluate the effect of dose and schedule of administration of prostaglandin F2α (PGF) during a synchronization protocol for fixed-time ET. Holstein and cross-bred Holstein heifers (n = 3766) were synchronized using a modified 5-day CIDR Synch as follows: Day 0: CIDR inserted; Day 5: CIDR removed, PGF2α treatment; Day 8: gonadotropin-releasing hormone (GnRH; 100 μg of gonadorelin). On Day 5, at the time of CIDR removal, heifers were randomly assigned to a 2 × 2 factorial design to receive either a full or half dose of PGF (Cloprostenol; 500 v. 250 μg) and 1 (Day 5) or 2 (Day 6) administrations resulting in the following treatments: full dose Day 5 + Day 6 (n = 938); full dose Day 5 (n = 938); half dose Day 5 + Day 6 (n = 946); and half dose Day 5 (n = 944). Heifers were evaluated by ultrasonography 5 days after GnRH to determine presence and size of the corpus luteum. Heifers with a corpus luteum received a fresh in vitro-produced embryo 7 ± 1 days after GnRH administration, and pregnancy was determined by ultrasonography 32 and 60 days after GnRH. Fertility data were analysed by logistic regression and included the fixed effects of dose, time, and their interaction. Fertility results are shown in Table 1. Utilisation rate (transferred/treated) was not affected by dose (P = 0.66), time (P = 0.19), or their interaction (P = 0.17). The percentage of heifers detected in oestrus was not affected by dose (P = 0.13), time (P = 0.72), or their interaction (P = 0.89). There were no significant differences between doses of PGF (P = 0.32), time (P = 0.71), or their interaction (P = 0.80) on pregnancies per ET on Day 32. Similarly, no differences were found on pregnancies per ET on Day 60 between doses (P = 0.35), time (P = 0.96), or their interaction (P = 0.89). In addition, pregnancy loss between Day 32 and 60 was not affected by dose (P = 0.76), time (P = 0.66), or their interaction (P = 0.54). In conclusion, the use of a half dose of PGF once on Day 5 resulted in comparable utilisation rate and fertility as the observed with 2 full dose applications 24 h apart. As a result, the overall cost of the fixed-time ET program can be reduced by eliminating the need for a second PGF treatment and by decreasing the dose without compromising fertility. Table 1. Utilisation rate, oestrus expression, pregnancies per ET (P/ET), and pregnancy loss in recipients receiving either a full or half dose of prostaglandin F2α on Days (D) 5 and 6 or once on D 5

The effect of equine chorionic gonadotrophin (ecg) injection combined with prostaglandin F2α (pGF2α) and gonadotrophin releasing hormone (GnRH) treatment on reproductive performance of Zandi ewes during non-breeding season

In this study, we aimed to investigate reproductive performance in estrus-induced Zandi ewes treated with equine chorionic gonadotrophin (eCG) injection in combination with prostaglandin F2α (PGF2α)and gonadotrophin releasing hormone (GnRH) during non-breeding season. The estrous cycle was synchronized using controlled internal drug release (CIDR) for 12 days. The ewes were randomly assigned to five groups (n=100 in each group). The first group (control) did not receive hormone injection, the second group (eCG) received 400 IU of eCG at the time of CIDR removal, the third group (eCG+PGF2α) received 400 IU of eCG and 1 ml of PGF2α at the time of CIDR removal, the fourth group (eCG+GnRH) received 400 IU of eCG at the time of CIDR removal and 1 ml of GnRH injection on the day of insemination, and the fifth group (eCG+PGF2α+GnRH) received 400 IU of eCG, 1 ml of PGF2α at the time of CIDR removal, and 1 ml of GnRH on the day of insemination. The results showed that the heat response rate was higher in all the groups receiving hormonal injections compared with the control group (P 0.05). In conclusion, use of eCG at the time of CIDR removal and GnRH on the day of insemination increased reproductive performance. Therefore, it can be applied as a strategy for increasing lambing rate in ewes.

1060 Effect of dose and timing of prostaglandin F2αtreatments during a Resynch protocol on luteal regression and fertility to timed artificial insemination in lactating Holstein cows

1060 Effect of dose and timing of prostaglandin F2αtreatments during a Resynch protocol on luteal regression and fertility to timed artificial insemination in lactating Holstein cows

Effect of the interval from Prostaglandin F2α treatment to ovulation on Embryo Recovery Rate and subsequent Pregnancy Rate in a commercial Equine Embryo Transfer program

Effect of the interval from Prostaglandin F2α treatment to ovulation on Embryo Recovery Rate and subsequent Pregnancy Rate in a commercial Equine Embryo Transfer program

Adding a second prostaglandin F2α treatment to but not reducing the duration of a PRID-Synch protocol increases fertility after resynchronization of ovulation in lactating Holstein cows

Our objective was to evaluate the effect of a second PGF2α treatment and duration of an Ovsynch protocol that included a progesterone-releasing intravaginal device (PRID) on progesterone (P4) concentrations and pregnancies per artificial insemination (P/AI) after resynchronization of ovulation and timed artificial insemination (TAI). Lactating Holstein cows (n=821) were assigned randomly at a nonpregnancy diagnosis (d 0) to 3 resynchronization protocols: (1) GnRH, d 0; PGF2α, d 7; GnRH, d 9.5 (7D1PGF); (2) GnRH, d 0; PGF2α, d 7; PGF2α, d 8; GnRH, d 9.5); (7D2PGF); or (3) GnRH, d 2; PGF2α, d 7; PGF2α, d 8; GnRH, d 9.5 (5D2PGF). All cows received a PRID at the first GnRH treatment of the resynchronization protocol, which was removed at the first PGF2α treatment, and all cows received TAI approximately 16h after the second GnRH treatment. Blood samples were collected from a subgroup of cows at each treatment of the resynchronization protocols. At 32 d after TAI, cows receiving a second PGF2α treatment (7D2PGF + 5D2PGF cows) had more P/AI (42.6 vs. 35.7%) than cows receiving a single PGF2α treatment (7D1PGF cows). For cows treated with a second PGF2α treatment, decreasing the duration of the protocol did not increase P/AI (41.4 vs. 43.8% for 7D2PGF vs. 5D2PGF cows). At 60 d after TAI, P/AI did not differ between cows treated with the 1 PGF2α (7D1PGF cows) or 2 PGF2α (7D2PGF + 5D1PGF cows) treatments (32.5 vs. 37.9%, respectively). In addition, reducing the duration of the protocol did not increase P/AI at 60 d after TAI (37.8 vs. 38.5% for 7D2PGF vs. 5D2PGF cows). Pregnancy loss from 32 to 60 d after TAI was not affected by the number of PGF2α treatments (8.5 vs. 10.6%, for 7D1PGF vs. 7D2PGF + 5D2PGF cows) or the duration of the protocol (9.1 vs. 12.1%, for 7D2PGF vs. 5D2PGF cows). The percentage of cows with incomplete luteal regression at the second GnRH treatment tended to differ among treatments and was lowest for 7D2PGF cows, intermediate for 5D2PGF cows, and greatest for 7D1PGF cows (1.9 vs. 6.9 vs. 11.0%, respectively). In conclusion, addition of a second PGF2α treatment tended to decrease the percentage of cows with incomplete luteal regression and increased P/AI 32 d after AI, whereas decreasing the duration of the Ovsynch protocol did not increase P/AI.

7 KISSPEPTIN SLOWLY ADMINISTERED DURING PROESTRUS IMPROVES FOLLICLE GROWTH AND OVULATION IN ANESTROUS COWS

This study evaluated whether kisspeptin (Kp) administered slowly from the moment of prostaglandin F2α (PGF) treatment for fixed-time AI protocol of anestrous cows provides suitable dominant follicle (DF) growth similarly to eCG. Postpartum, anestrous, primiparous Nelore cows (n = 50; &lt;8-mm follicles with no corpus luteum) and lean (body condition score 2.7 ± 0.1; 0–5 score scale) were enrolled. On Day 0, females received an intravaginal device with 1.0 g of progesterone (Sincrogest®, Ourofino, Brazil) and 2 mg of oestradiol benzoate IM (Sincrodiol®, Ourofino, Brazil). On Day 8, the device was removed, and cows were administered 500 μg of sodic cloprostenol IM (Sincrocio®, Ourofino, Brazil) and 1 mg of oestradiol cypionate IM (E.C.P.®, Zoetis, Brazil). Females were blocked by dominant follicle diameter and allotted in following groups: (1) control (CTRL), 1.5 mL of saline IM; (2) eCG, 300 IU IM of eCG (Novormon®, Intervet, Brazil); and (3) Kp SC with an osmotic minipump (9.65 μg kg–1 of Kp diluted in saline) over 48 h. Dominant follicle growth and ovulation evaluation were performed by ultrasound at 0, 24, 48, 60, 72, 84, and 96 h after PGF treatment. Dominant follicle growth rate was calculated by difference on DF from 0 to 48 h. Ovulation dispersion was analysed by proc GLIMMIX of SAS (SAS Institute Inc., Cary, NC, USA). Follicle variables, ovulation rate, and interval from PGF to ovulation were analysed by orthogonal contrast. Differences were considered when P &lt; 0.05, and tendencies were considered when P &lt; 0.1. There were no difference between CTRL and growth promoters (eCG and Kp) or between growth promoters on DF diameter at 0 h, larger follicle registered from 0 h to ovulation, ovulatory DF diameter, ovulation rate, and interval between PGF treatment and ovulation (Table 1). Dominant follicle diameter at 48 h was larger for growth promoters than for CTRL. Treatment with Kp proved as efficient as eCG in the DF growth rate, with tendency (P = 0.1) to be greater than CTRL. Data from our laboratory (Sales et al. 2011 Anim. Rep. Sci. 124, 12–18.) with same model and animal profile (n = 150 per group) presented positive effect of eCG treatment for growth and size of DF. Cows treated with Kp ovulated in only 3 specific times (60, 72, and 84 h), unlike eCG (48, 60, 72, and 84 h) and CTRL (24, 48, 60, 72, 84, and 96 h; P &gt; 0.05). Also, it is noted that Kp treatment dose and releasing does not cause negative effect on follicle growth and ovulation (down-regulation on LH releasing) or alteration on interval of PGF treatment to ovulation. Thus, synchronized anestrous cows treated with Kp delivered slowly during proestrus have similar results to eCG on follicle growth and ovulation. Table 1.Ovarian variables of anestrous Nelore cows treated with saline, eCG (300 IU), or kisspeptin (Kp; 9.65 μg kg–1). Data were analysed by orthogonal contrast, where C1 = growth promoter (eCG and Kp) v. control; C2 = eCG v. Kp

110 TREATMENT WITH GnRH ON DAY 5 REDUCES PREGNANCY LOSS IN HEIFERS RECEIVING IN VITRO-PRODUCED EXPANDED BLASTOCYSTS

The hypothesis was that GnRH on Day 5 of a synchronized cycle in embryo transfer recipients would increase progesterone (P4) concentrations, embryo size, and fertility. Holstein and cross-bred Holstein heifers (n = 1562) were synchronized using a modified 5-day CIDR Co-Synch as follows: Day –8 CIDR inserted; Day –3 CIDR removed; prostaglandin F2α treatment; Day –2 second prostaglandin F2α; Day 0 gonadotropin-releasing hormone (G1, 100 μg of gonadorelin acetate) to induce ovulation. On Day 5.5, heifers were assigned in a completely randomised design to 1 of 2 treatments: Control (untreated) or GnRH (200 μg of gonadorelin acetate). Transfer of fresh in vitro-produced embryos was performed between d 6 and 8 after G1. Data collected from each heifer included embryo stage and quality, body condition score, technician, interval from G1 to transfer, and number of previous transfers. All heifers were evaluated by transrectal ultrasonography on Day 5, 33, and 62 and a subset of heifers was scanned on Day 12 (n = 718; to determine ovulation to treatment) and another subset on Day 33 (n = 296; 16-s video to determine embryo and amniotic vesicle size). Serum P4 was determined from a subset of heifers on Day 12 (n = 467). Fertility data were analysed by logistic regression (LOGISTIC procedure, SAS 9.4), whereas continuous outcomes were analysed by ANOVA (MIXED procedure). Ovulation to Day 5.5 gonadotropin-releasing hormone was 83.9% (302/360) in GnRH-treated heifers v. 3.3% (12/358) in Control (P &lt; 0.001). Progesterone on Day 12 was greater in GnRH-treated heifers 7.2 ± 0.1 ng mL–1 v. Controls 6.0 ± 0.1 ng mL–1 (P &lt; 0.001). There was an effect of embryo stage at Day 33 and 60 of pregnancy, with Stage 7 having greater P/ET than Stage 6 embryos. Treatment with GnRH did not alter pregnancy per embryo transfer with either embryo stage but decreased pregnancy loss in Stage 7 embryos, as shown in Table 1. Embryo size measured as crown-rump length (CRL) did not differ, as shown in Table 1. Similarly, amniotic vesicle volume (AVV) was not different between GnRH (549.1 ± 16 mm3) and Control (543.5 ± 14 mm3; P = 0.86), nor was there an interaction between treatment and embryo stage (P = 0.71). In addition, neither AVV (P = 0.22) nor CRL (P = 0.41) were associated with pregnancy loss between Day 33 and 60. In conclusion, treatment with GnRH on Day 5 resulted in increased P4 and a reduction in pregnancy loss in heifers receiving a Stage 7 embryo without changing conceptus size. Table 1.Pregnancies per embryo transfer (P/ET), crown-rump length (CRL), and pregnancy loss in embryo recipients receiving gonadotropin-releasing hormone (GnRH) on Day 5.5 v. control

Effect of a second treatment with prostaglandin F2α during the Ovsynch protocol on luteolysis and pregnancy in dairy cows

The main objective of this study was to evaluate the effect of a second treatment with prostaglandin F2α (PGF) during Ovsynch on regression of the corpus luteum (CL) and on fertility to the timed artificial insemination. Two experiments were performed. In both experiments, cows were randomized to receive (1) no additional treatments with PGF=1 PGF, or (2) a second PGF treatment at 24h after the first PGF treatment=2 PGF. The first experiment (n=344 synchronized lactating dairy cows that received artificial insemination at 81±3d in milk) used the Double-Ovsynch protocol for synchronizing ovulation. Blood samples were collected at the PGF and final GnRH treatments (72 and 16h before timed artificial insemination) during the breeding Ovsynch protocol, to determine CL regression in response to the protocol. Treatment with 2 PGF increased CL regression from 83.0% with 1 PGF to 97.0% with 2 PGF. The effect of 2 PGF on CL regression was observed in both primiparous and multiparous cows. Cows with lower (2.0 to 4.8ng/mL) versus greater (4.9 to 12.0ng/mL) circulating progesterone at the time of PGF had lower percentage of cows with complete CL regression after 1 PGF (66.7 vs. 88.1%) but not after 2 PGF (95.1 vs. 97.6%). Experiment 2 used 2,148 lactating dairy cows on 11 dairy farms in 4 different regions of the United States. Cows were synchronized with Ovsynch and received timed artificial insemination at 60±3d in milk. Cows that received 2 PGF had a tendency for increased pregnancies per artificial insemination (P/AI) compared with cows with 1 PGF (36.1 vs. 33.3%). This tendency for improvement in P/AI was observed in multiparous but not in primiparous cows. Combining data from the 2 experiments indicated a 9.45% relative increase in P/AI for cows receiving 2 compared with 1 PGF (37.6 vs. 34.4%) with the increase in P/AI observed in multiparous but not in primiparous cows. Thus, a second PGF treatment in Ovsynch-type protocols can increase pregnancy success by about 10%, primarily due to enhanced fertility in multiparous cows.

Modifications to Ovsynch improve fertility during resynchronization: Evaluation of presynchronization with gonadotropin-releasing hormone 6 d before initiation of Ovsynch and addition of a second prostaglandin F2α treatment

Lactating Holstein cows (n=897) were randomly assigned to a 2×2 factorial arrangement of treatments to compare the main effects of presynchronization with GnRH 6 d before beginning an Ovsynch protocol (±GnRH) and a second PGF2α treatment 24h after the first (1 vs. 2 PGF2α) on pregnancies per artificial insemination (P/AI). This resulted in the following 4 treatments: (1) an Ovsynch protocol (GPG, control); (2) presynchronization with GnRH followed by an Ovsynch protocol (GGPG); (3) an Ovsynch protocol with a second PGF2α treatment (GPPG); and (4) presynchronization with GnRH followed by an Ovsynch protocol with a second PGF2α treatment (GGPPG). All cows were submitted for first timed artificial insemination (TAI) using a Presynch Ovsynch protocol, and cows detected in estrus after the second PGF2α treatment of the Presynch portion of the protocol were inseminated and removed from the experiment. Nonpregnant cows were resynchronized using an Ovsynch protocol initiated 32±3 d after artificial insemination. Blood samples were collected at the first GnRH treatment (G1), at the PGF2α treatment (PGF), and at the last GnRH treatment (G2) of the Ovsynch protocol and were assayed for progesterone (P4) concentrations. Overall, P/AI tended to be greater for cows receiving a second PGF2α treatment compared with cows not receiving the second PGF2α treatment (40 and 37% for GGPPG and GPPG treatments, respectively, vs. 33 and 32% for GGPG and GPG treatments, respectively). Interestingly, treatment effects on P/AI were detected only for resynchronized cows receiving second and greater TAI, but not for cows receiving first TAI. Fewer cows presynchronized with GnRH had low (<0.5ng/mL) P4 at G1 than cows not presynchronized (13 vs. 25%), and P4 was greater at PGF2α for cows presynchronized with GnRH (4.4 vs. 4.0ng/mL). Cows receiving 2 PGF2α treatments had lower P4 at G2 than cows receiving 1 PGF2α treatment (0.15 vs. 0.35ng/mL). Differences in P4 at PGF2α were detected only for resynchronized cows and not for cows submitted for first TAI. We conclude that presynchronization with GnRH 6 d before beginning an Ovsynch protocol tended to increase P4 at the first GnRH treatment of an Ovsynch protocol, and that a second PGF2α treatment 24h after the first decreased P4 at TAI, thereby increasing P/AI in cows resynchronized for second and greater TAI.

Effect of Cloprostenol Administration on Interval to Subsequent Ovulation and Anovulatory Follicle Formation in Quarter Horse Mares

Prostaglandin F2α (PGF) treatment is routinely used in the reproductive management of mares to induce luteolysis and allow a subsequent return to estrus. The objective of this retrospective study was to assess the effect of follicle size at the time of administration of cloprostenol on interval to subsequent ovulation. A secondary objective was to determine the incidence of hemorrhagic anovulatory follicle (HAF) formation after PGF administration. Reproductive records of 275 mares monitored over a total of 520 estrous cycles were evaluated. All mares received a single intramuscular dose of 250 μg of the synthetic PGF analog cloprostenol sodium between days 5 and 12 after ovulation. The average interval from PGF to ovulation was 8.4 ± 2.5 days. The interval from PGF administration to subsequent ovulation was inversely proportional to the diameter of the largest follicle at the time of treatment. Administration of cloprostenol to mares with a large (≥35 mm in diameter) diestrous follicle resulted in one of three outcomes—ovulation within 48 hours (13.4%) with variable uterine edema, ovulation after 48 hours usually accompanied by the presence of uterine edema (73.1%), or regression without ovulation followed by emergence and eventual ovulation of a new dominant follicle (13.4%). There was no effect of mare age or season on interval from PGF to ovulation. The overall incidence of HAF development after PGF administration in this study was low (2.5%).