Progesterone-based fixed-time artificial insemination protocols for dairy cows: Gonadotropin-releasing hormone versus estradiol benzoate at initiation and estradiol cypionate versus estradiol benzoate at the end.

Our objectives were to evaluate ovarian dynamics and fertility comparing 2 treatments at the start of a progesterone (P4)-based fixed-time artificial insemination (FTAI) protocol and 2 treatments at the end of the protocol. Thus, 1,035 lactating Holstein cows were assigned in a random phase of the estrous cycle to 1 of 4 treatments using a completely randomized design with a 2×2 factorial arrangement. At the beginning of the protocol (d -10), cows received GnRH or estradiol benzoate (EB) and, at the end, EB (d -1) or estradiol cypionate (ECP; d -2), resulting in 4 treatments: GnRH-EB, GnRH-ECP, EB-EB, and EB-ECP. All cows received an intravaginal P4 device on d -10, which was removed on d -2. Cows also received PGF2α on d -3 and -2. The FTAI was performed on d 0. Ovaries were evaluated by ultrasound for corpus luteum (CL) presence and regression (d -10 and -3) and follicle measurements (d -10 and 0), as well as the uterus for percentage pregnant per AI (P/AI; d 32 and 60). Blood samples were collected (d -10 and -3) for P4 measurements. Treatment with GnRH rather than EB tended to increase P/AI on d 32 (38.2 vs. 33.7%) and on d 60 (32.9 vs. 28.9%). More cows treated with GnRH had CL on d -3 compared with EB-treated cows (77.3 vs. 58.3%), due to less CL regression between d -10 and -3 (24.7 vs. 43.8%) and more cows with a new CL on d -3 (35.9 vs. 25.0%). Cows treated with GnRH also had greater P4 concentrations on d -3 than EB cows (3.4 vs. 2.0 ng/mL). Increased circulating P4 at the start of the protocol (d -10) decreased the probability of ovulation to EB or GnRH at that time. Cows from GnRH group also ovulated a larger-diameter follicle at the end of the protocol (15.5 vs. 14.7mm). No difference between EB and ECP in P/AI on d 32 (34.8 vs. 37.0) and 60 (30.8 vs. 31.0%) or in pregnancy loss (11.1 vs. 15.4%) was detected and we found no interaction between treatments for P/AI. Independent of treatment, cows with CL on d -10 and -3 had the greatest P/AI on d 60 (36.9%). In conclusion, treatments at the end of the protocol were similar for ECP or EB and we found no additive effect or interactions on P/AI between treatments. However, cows treated with GnRH rather than EB on d -10 had less luteolysis and tended to have greater P/AI, probably because P4 concentrations were greater during the protocol. Finally, regardless of treatments, cows with CL at the beginning of the protocol as well as at the time of PGF2α had greater fertility.

Melo LF ，Monteiro PLJ Jr ，Surjus RS ，Drum JN ，Wiltbank MC ，Sartori R ... -  《-》

Increasing estradiol benzoate, pretreatment with gonadotropin-releasing hormone, and impediments for successful estradiol-based fixed-time artificial insemination protocols in dairy cattle.

With the objective to optimize fixed-time artificial insemination (FTAI) protocols based on estradiol benzoate (EB) and progesterone (P4), we performed 2 experiments (Exp.) in dairy cows. In Exp. 1 (n=44), we hypothesized that increased EB (EB3=3 mg vs. EB2=2 mg) on d 0 would improve synchronization of ovarian follicle wave emergence. Likewise, in Exp. 2 (n=82), we hypothesized that a GnRH treatment on d -3 (early in a follicular wave on d 0) versus d -7 (presence of a dominant follicle on d 0) would better synchronize wave emergence. Moreover, results from both experiments were combined to identify reasons for the lack of synchronization. All cows were treated with EB at the time of introduction of a P4 implant (d 0). On d 7, cows were given 25 mg of prostaglandin F2α; on d 8, the implant was removed and cows were given 1mg of estradiol cypionate. All cows received FTAI on d 10. In both experiments, daily ultrasound evaluations were performed and, in Exp. 2, circulating P4 was evaluated during the protocol. Pregnancy per artificial insemination (P/AI) was determined on d 31 and 59 after FTAI. In Exp. 1, EB dose did not change time to wave emergence, but EB3 compared with EB2 decreased the percentage of cows with a corpus luteum on d 7 (19.8 vs. 55.3%) and time to ovulation (10.4 vs. 10.9 d). In Exp. 2, although we detected a tendency for delayed follicle wave emergence after the start of the FTAI protocol in cows ovulating to GnRH given on d -7, there was no difference in percentage of cows with a synchronized wave emergence (~80%). Regardless of treatment, more cows with P4<0.1 ng/mL, compared with P4≥0.1 and <0.22 ng/mL at the time of AI, ovulated to the protocol (81.2 vs. 58.0%) and had increased P/AI (47.4 vs. 21.4%). An analysis of data from both experiments showed that only 73.8% (93/126) of cows had synchronized wave emergence, and only 77.8% (98/126) of cows ovulated at the end of the protocol. Fertility was much greater in cows that had emergence of a new wave synchronized and ovulated to end of the protocol [P/AI 61.3% (46/75)] compared with cows that failed to present one or both of the outcomes above [15.7% (8/51)]. Thus, although current FTAI protocols using EB and P4 produce P/AI between 30 and 40% for lactating dairy cows, there remains room for improvement because less than 60% (75/126) of the cows were correctly synchronized. Starting the FTAI protocol without the dominant follicle or increasing the dose of EB to 3mg was not effective in increasing synchronization rate.

Monteiro PL Jr ，Borsato M ，Silva FL ，Prata AB ，Wiltbank MC ，Sartori R ... -  《-》

Effect of adding a gonadotropin-releasing-hormone treatment at the beginning and a second prostaglandin F2α treatment at the end of an estradiol-based protocol for timed artificial insemination in lactating dairy cows during cool or hot seasons of the yea

Our hypothesis was that fertility could be increased in a timed artificial insemination (TAI) protocol based on estradiol (E2) and progesterone (P4) by combining GnRH with E2-benzoate at the start of the protocol to increase circulating P4 during preovulatory follicle development and by using 2 prostaglandin F2α (PGF) treatments at the end to decrease P4 near TAI. Lactating Holstein cows (n=1,808) were randomly assigned during the cool or hot season of the year to receive TAI (d 0) following 1 of 3 treatments: (1) control: controlled internal drug-release insert + 2mg of E2-benzoate on d -11, PGF on d -4, controlled internal drug-release insert withdrawal + 1.0mg of E2-cypionate on d -2, and TAI on d 0; (2) 2PGF: identical to control protocol with addition of a second PGF treatment on d -2; (3) GnRH: identical to 2PGF protocol with addition of a 100-μg GnRH treatment on d -11. Pregnancy diagnoses were performed on d 32 and 60 after TAI. Season had major effects on many reproductive measures, with cool season greater than hot season in percentage of cows with corpus luteum (CL) at PGF (62.9 vs. 56.2%), ovulatory follicle diameter (15.7 vs. 14.8mm), expression of estrus (86.7 vs. 79.9%), ovulation following the protocol (89.7 vs. 84.3%), and pregnancies per artificial insemination (P/AI; 45.4 vs. 21.4%). The GnRH protocol increased percentage of cows with CL (control=56.9%; 2PGF=55.8%; GnRH=70.5%) and P4 at PGF (control=3.28±0.22; 2PGF=3.35±0.22; GnRH=3.70±0.21ng/mL), compared with control and 2PGF protocols. The GnRH protocol increased P/AI at the pregnancy diagnosis at 32d [37.3% (219/595)] and 60d [31% (179/595)] after TAI, compared with control [30.0% (177/604); 25.1% (145/604)], with intermediate results with 2PGF protocol [33.2% (196/609); 28.0% (164/609)]. The positive effects of GnRH treatment on P/AI were only detected during the cool season (GnRH=50.9%; 2PGF=44.2%; control=41.0%) and not during the hot season. In addition, the effect of GnRH was only observed in cows with low P4 (<3ng/mL) at the start of the protocol and not in cows that began the protocol with high P4. Furthermore, presence of CL at PGF interacted with follicle diameter such that cows with a CL at PGF had greater P/AI if they ovulated larger rather than smaller follicles near TAI. Thus, fertility to TAI can be improved by inducing ovulation at the beginning of an E2/P4-based protocol using GnRH treatment, particularly during the cool season of the year and in cows with low P4 at the start of the protocol.

Pereira MH ，Wiltbank MC ，Barbosa LF ，Costa WM Jr ，Carvalho MA ，Vasconcelos JL ... -  《-》

Comparison of gonadotropin-releasing hormone and estradiol benzoate plus gonadotropin-releasing hormone to initiate a progesterone-based timed artificial insemination resynchronization protocol in lactating dairy cows.

The present study compared 2 strategies to initiate a progesterone (P4)-based timed artificial insemination (TAI) protocol for lactating dairy cows: only GnRH or estradiol benzoate (EB) plus GnRH (EB+GnRH). Lactating Holstein cows (n = 487; 184 primiparous and 303 multiparous) from 2 commercial dairy herds were used for their second or greater services postpartum. Each week, cows that were nonpregnant at the pregnancy diagnosis 32 d after a previous AI were randomly assigned to 1 of 2 experimental groups that differed only in the strategy to initiate (d 0) the TAI protocol. On d 0, every cow received a 2.0-g P4 implant; in the EB+GnRH group, cows were treated with 2.0 mg i.m. of EB and 16.8 µg i.m. of the GnRH analog buserelin acetate, whereas in the GnRH group, cows received only 16.8 µg i.m. of GnRH. On d 7 after the initial treatment, 0.530 mg i.m. of cloprostenol sodium (PGF) was administered in all cows, followed by a second dose on d 8, concomitant with 1.0 mg i.m. of estradiol cypionate and P4 implant withdrawal. The TAI was performed on d 10 (48 h after P4 device withdrawal) in both experimental groups. Only conventional Holstein semen was used throughout the study. The percentage of cows with corpus luteum (CL) on d 0 (73%) and overall ovulation rate after d 0 (54%) did not differ between groups. The CL regression between d 0 and the first PGF treatment was greater in the EB+GnRH group than the GnRH group (42% vs. 31%). Consequently, the proportion of cows with CL at PGF was greater when only GnRH was used on d 0 compared with EB+GnRH (86% vs. 82%), and the mean number of CL at PGF was greater (1.23 vs. 1.11). The expression of estrus near TAI was greater in GnRH group (84% vs. 77%), and cows showing estrus had greater (44% vs. 10%) pregnancy per AI (P/AI) on d 32 for both treatments. We found no effect of the presence of CL on d 0 or at PGF, nor of ovulation after d 0 or CL regression between d 0 and d 7 on fertility. However, fertility was critically impaired when cows did not have CL at both times, d 0 and at PGF treatment. We did not observe any interaction between treatment and other variables, and the P/AI was similar in cows receiving EB+GnRH or only GnRH on d 0 (37.8% vs. 36.6%). In summary, although there was no detectable difference in P/AI between treatments, this study demonstrated potential negative physiological outcomes caused by EB treatment on d 0 (greater incidence of luteolysis after d 0 and fewer cows with CL at PGF treatment). Overall, we found no benefit of adding EB at the initiation of a P4-based TAI protocol on fertility compared with using GnRH alone, despite differences in ovarian dynamics and expression of estrus.

Consentini CEC ，Melo LF ，Abadia T ，Gonzales B ，Motta JCL ，Alves RLOR ，E Silva LO ，Wiltbank MC ，Sartori R ... -  《-》

Timed artificial insemination programs during the summer in lactating dairy cows: comparison of the 5-d Cosynch protocol with an estrogen/progesterone-based protocol.

The objective of this study was to compare a GnRH-based to an estrogen/progesterone (E2/P4)-based protocol for estrous cycle synchronization and fixed timed artificial insemination (TAI), both designed for synchronization of ovulation and to reduce the period from follicular emergence until ovulation in cows with a synchronized follicular wave. A total of 1,190 lactating Holstein cows (primiparous: n=685 and multiparous: n=505) yielding 26.5 ± 0.30 kg of milk/d at 177 ± 5.02 d in milk were randomly assigned to receive one of the following programs: 5-d Cosynch protocol [d -8: controlled internal drug release (CIDR) + GnRH; d -3: CIDR removal + PGF2α; d -2: PGF2α; d 0: TAI + GnRH] or E2/P4 protocol (d -10: CIDR + estradiol benzoate; d -3: PGF2α; d -2: CIDR removal + estradiol cypionate; d 0: TAI). Rectal temperature and circulating progesterone (P4) were measured on d -3, -2, 0 (TAI), and 7. The estrous cycle was considered to be synchronized when P4 was ≥ 1.0 ng/mL on d 7 in cows that had luteolysis (P4 ≤ 0.4 ng/mL on d 0). To evaluate the effects of heat stress, cows were classified by number of heat stress events: 0, 1, and 2-or-more measurements of elevated body temperature (≥ 39.1 °C). Pregnancy success (pregnancy per artificial insemination, P/AI) was determined at d 32 and 60 after TAI. The cows in the 5-d Cosynch protocol had increased circulating P4 at the time of PGF2α injection (2.66 ± 0.13 vs. 1.66 ± 0.13 ng/mL). The cows in the E2/P4 protocol were more likely to be detected in estrus (62.8 vs. 43.4%) compared with the cows in the 5-d Cosynch protocol, and expression of estrus improved P/AI in both treatments. The cows in the 5-d Cosynch protocol had greater percentage of synchronized estrous cycle (78.2%), compared with cows in the E2/P4 protocol (70.7%). On d 60, the E2/P4 protocol tended to improve P/AI (20.7 vs. 16.7%) and reduced pregnancy loss from 32 to 60 d (11.0 vs. 19.6%), compared with the 5-d Cosynch protocol. In cows withtheir estrous cycle synchronized, the E2/P4 protocol had greater P/AI (25.6 vs. 17.7%) on d 60 and lower pregnancy loss from 32 to 60 d (6.7 vs. 21.7%) compared with cows in the 5-d Cosynch protocol. Follicle diameter affected pregnancy loss from 32 to 60d only in the cows in the 5-d Cosynch protocol, with smaller follicles resulting in greater pregnancy loss. Pregnancy per AI at d 60 was different between protocols in the cows with 2 or more measurements of heat stress (5-d Cosynch=12.2% vs. E2/P4=22.8%), but not in the cows without or with 1 heat stress measurement. In conclusion, the 5-d Cosynch protocol apparently produced better estrous cycle synchronization than the E2/P4 protocol but did not improve P/AI. The potential explanation for these results is that increased E2 concentrations during the periovulatory period can improve pregnancy success and pregnancy maintenance, and this effect appears to be greatest in heat-stressed cows when circulating E2 may be reduced.

Pereira MHC ，Rodrigues ADP ，Martins T ，Oliveira WVC ，Silveira PSA ，Wiltbank MC ，Vasconcelos JLM ... -  《-》