Research on timed AI in beef cattle: Past, present and future, a 27-year perspective.

This review aimed to (1) summarize the results from fixed-timed artificial insemination (TAI) fertility studies performed during the last 27 years; (2) compile and evaluate, as examples from the literature base, the direct comparisons made of specific manipulations to synchronization protocols; (3) evaluate the impact of the TAI programs on the reproductive performance during the breeding season, and (4) provide perspective on the future of TAI programs in beef cattle. A search of the literature published from 1995 to 2021 was conducted to identify experiments in which synchronization of ovulation and TAI in beef cattle was performed. The primary outcome of interest was fertility expressed as pregnancies per TAI. The literature included two search engines, the SIS Web of Science and the US National Library of Medicine Institutes of Health through PubMed. After the initial search and screening, a total of 228 manuscripts were selected containing a total of 272,668 TAI. A dramatic increase in the number of publications and TAIs occurred throughout the years. Most of them were from Brazil and United States, followed by Canada, Argentina, Uruguay, and Australia. Two main types of TAI programs were identified: GnRH-based and E2/P4-based protocols. In terms of GnRH-based programs, two variations were evaluated in the present manuscript. First, we evaluated the effect of the progesterone implant during the protocol. The progesterone implant increased pregnancy/TAI (P/TAI) from 44.3 to 54.3%. Second, the use of a second prostaglandin F2α treatment in 5-d CO-synch program increased the P/TAI from 53.2 to 60.9%. In E2/P4-based programs, use of GnRH at TAI increased P/TAI from 54.7 to 59.2% in cows. However, no increase was detected in heifers. Other research showed that use of TAI can increase the overall proportion of the cows pregnant at end of the breeding season and produce earlier calvings compared with bulls. In conclusion, there have been a large number of excellent research studies that have been performed during the last 27 years on TAI in beef cattle. This technology is being utilized successfully in the beef cattle industry. This success is largely because of the valid research that underlies the application of the technology and the economic value of the technology.

Dimethylformamide Preserves the Integrity of Cryopreserved Goat Semen in a Soybean Lecithin-Based Extender.

This study investigated the cryoprotectant effects of dimethylformamide (DMF), ethylene glycol (EG), and dimethyl sulfoxide (DMSO) as substitutes for glycerol (GLY) in a soybean lecithin (SL)-based extender in the cryopreservation of buck sperm. In this study, the semen of three Saanen bucks was individually extended in SL supplemented with 5% GLY (control), DMF, EG, or DMSO. After this, the extended semen was cryopreserved and two straws from each group were thawed (37°C for 30 seconds), pooled, and analyzed for sperm motion parameters, plasma membrane integrity (PMI), acrosomal integrity (ACI), and high mitochondrial membrane potential (HMMP). Samples were analyzed after 15 minutes (T0) and after 2 hours of incubation at 37°C (T2). The results revealed higher values of motility (total and progressive) and sperm motion parameters for DMF than the other cryoprotectants (p < 0.0001). PMI and HMMP did not differ (p > 0.05) between GLY and DMF, but ACI was higher (p < 0.01) for DMF compared with GLY. Based on these results, DMF and GLY samples were used in heterologous in vitro fertilization assays by using bovine oocytes (n = 337) obtained from a slaughterhouse. No differences (p > 0.05) were observed between GLY and DMF for unfertilized (GLY: 38.8%; DMF: 25.33%), pronucleus (GLY: 25.68%; DMF: 27.92%), and cleavage rates (GLY: 35.52%; DMF: 46.75%). Based on these results, it is concluded that DMF preserves sperm motion characteristics and ACI better than GLY, EG, and DMSO, and it is the penetrating cryoprotectant of choice for the cryopreservation of buck sperm in SL extender.

Accessory corpus luteum regression during pregnancy II: reproductive outcomes.

The objective of this study was to evaluate the effect of accessory corpus luteum (CL) induction on fertility in dairy cows. On day 5 after artificial insemination (AI), lactating Holstein cows were assigned unequally to receive gonadotrophin-releasing hormone treatment (GnRH) (n = 641) or no treatment (control; n = 289). Cows had their blood sampled for progesterone (P4), and ovaries were scanned by ultrasound on days 5, 12, 19, 26, 33, 47, and 61 after AI. Pregnancy diagnosis was performed on days 26, 33, 47, and 61. On day 12, cows treated with GnRH were allocated to ipsilateral (n = 239) or contralateral (n = 241) groups based on the side of accessory CL formation relative to previous ovulation. Accessory CL cows had greater P4 than controls. In total, 52.7% (78/148) of pregnant cows in contralateral group had accessory CL regression earlier (

Accessory corpus luteum regression during pregnancy I: timing, physiology, and P4 profiles.

Inappropriate corpus luteum (CL) regression can produce pregnancy loss. An experimental model was utilized to investigate regression of accessory CL during pregnancy in dairy cows. Cows were bred (day 0) and treated with gonadotrophin-releasing hormone 6 days later to form accessory CL. Transrectal ultrasound (every other days) and blood samples for progesterone (P4; daily) were performed until day 56 of pregnancy. On day 28, 13 cows were confirmed pregnant, and accessory CL were found contralateral (n = 9) or ipsilateral (n = 4) to previous ovulation. On day 18, CL biopsy was performed to analyze mRNA expression for interferon-stimulated genes (ISGs). Luteolysis occurred more frequently in cows that had contralateral accessory CL (88.9% (8/9)) than in cows with ipsilateral accessory CL (0% (0/4)). Luteolysis of contralateral accessory CL occurred either earlier (days 19-23; 2/8) or later (days 48-53; 6/8) in pregnancy and occurred rapidly (24 h), based on daily P4. After onset of earlier or later accessory CL regression, circulating P4 decreased by 41.2%. There was no difference in luteal tissue mRNA expression for ISGs on day 18 between accessory and original CL and between CL that subsequently regressed or did not regress. On day 56, an oxytocin challenge dramatically increased prostaglandin F2α metabolite (PGFM) in all cows but produced no pregnancy losses, although cows with previous accessory CL regression had greater PGFM. In summary, ipsilateral accessory CL did not regress during pregnancy, whereas most contralateral CL regressed by 63 days of pregnancy, providing evidence for local mechanisms in regression of accessory CL and protection of CL during pregnancy.

Oxytocin-induced prostaglandin F2-alpha release is low in early bovine pregnancy but increases during the second month of pregnancy†.

Circulating prostaglandin F2α metabolite (PGFM) after an oxytocin challenge was evaluated throughout the first 2 months of pregnancy in lactating Holstein cows. On day 11, 18, and 25 after artificial insemination (AI), and on days 32, 39, 46, 53, and 60 of pregnancy, cows were challenged with 50 IU oxytocin, i.m. Blood was collected before (0 min), 30, 60, 90, and 120 min after oxytocin for plasma PGFM concentrations. Ultrasound evaluations were performed for pregnancy diagnosis on day 32-60 post-AI. Nonpregnant (NP) cows on day 18 were designated by a lack of interferon-stimulated genes in peripheral blood leukocytes and Pregnant (P) based on day 32 ultrasound. On day 11, P and NP were similar with low PGFM and no effect of oxytocin on PGFM. On day 18, oxytocin increased PGFM (3-fold) in NP with little change in P cows. Comparing only P cows from day 11 to 60, basal circulating PGFM increased as pregnancy progressed, with day 11 and 18, lower than all days from day 25 to 60 of pregnancy. Oxytocin-induced PGFM in P cows on day 25 was greater than P cows on day 18 (2.9-fold). However, oxytocin-induced PGFM was lower on day 25 compared to day 53 and 60, with intermediate values on day 32, 39, and 46 of pregnancy. Thus, the corpus luteum (CL) of early pregnancy (day 11, 18) is maintained by suppression of PGF, as reflected by suppressed PGFM in this study. However, during the second month of pregnancy, uterine PGF secretion was not suppressed since basal PGFM and oxytocin-induced PGFM secretion were elevated. Apparently, mechanisms other than suppression of oxytocin receptors maintain CL after day 25 of pregnancy.

Effects of adiponectin during in vitro maturation of goat oocytes: MEK 1/2 pathway and gene expression pattern.

Recent studies have shown that adiponectin, an adipokine predominantly produced by adipose tissue, regulates several reproductive processes. However, the mechanisms of action of adiponectin on the maturation of goat oocytes remain to be determined. The aim of this study was to investigate whether (a) adiponectin influences the meiotic maturation of goat oocytes; (b) MAPK MEK 1/2 mediates the effects of adiponectin; and 3) adiponectin differentially affects mRNA relative abundance of genes relevant for adiponectin signal transduction in goat oocytes. The addition of adiponectin (5 µg/ml) during the maturation of goat oocytes resulted in a higher percentage of successful nuclear maturation compared to those of the group without adiponectin (p < 0.05). Adiponectin-stimulated nuclear oocyte maturation was significantly impaired by a mitogen-activated protein kinase MEK 1/2 inhibitor, U0126 (p < 0.05). There was no evidence of any adiponectin-induced difference in the relative transcript abundances of AdipoR1, AdipoR2, AMPKα1, AMPKα2, PPARα and PPARγ genes. In conclusion, these results indicate that adiponectin has a positive effect on the meiotic maturation of goat oocytes through the MAPK MEK 1/2 pathway. Furthermore, the adiponectin does not affect the relative abundance of genes relevant for adiponectin signal transduction in goat oocytes.

Expression of adiponectin and its receptors (AdipoR1 and AdipoR2) in goat ovary and its effect on oocyte nuclear maturation in vitro.

Adiponectin is an adipokine secreted primarily by adipocytes and is involved in the control of male and female reproductive functions. Circulating levels of adiponectin are inversely correlated with body fat mass, and its biological effects are predominantly mediated through two receptors, AdipoR1 and AdipoR2. The aim of the present study was to verify the expression of the adiponectin system (adiponectin and its receptors, AdipoR1 and AdipoR2) in goat ovary using qPCR and immunohistochemistry analyses and further investigate the in vitro effects of recombinant adiponectin (5 µg/mL and 10 µg/mL) on goat oocyte nuclear maturation. We demonstrated that the mRNA and proteins of the adiponectin system are present in goat ovary. Gene and protein expression of AdipoR1 and AdipoR2 was detected in follicular cells (oocyte, cumulus, granulosa and theca) of small and large antral follicles, while adiponectin mRNA was not detected in oocytes from small and large follicles or in large follicle cumulus cells. Finally, addition of various concentrations of adiponectin in maturation medium affected the number of oocytes that reached metaphase II. In conclusion, in the present study, we detected expression of adiponectin and its receptors AdipoR1 and AdipoR2 in goat ovarian follicles. Furthermore, we demonstrated that recombinant adiponectin increases nuclear maturation of goat oocytes in vitro.

Embryo production in heifers with low or high dry matter intake submitted to superovulation.

This study investigated the influence of feed intake on superovulatory response and embryo production of Nelore heifers. Pubertal heifers were kept in a feedlot and were submitted to the same diets, but with different levels of feed consumption: High (1.7 M; n = 20) or Low (0.7 M; n = 19) feed intake. Heifers in the 1.7 M treatment consumed 170% (2.6% of body weight [BW] in dry matter) and the 0.7 M heifers ate 70% (1.1% of BW in dry matter) of a maintenance diet. After 7 wk on these diets, heifers were treated with eight decreasing doses of follicle-stimulating hormone (FSH) given every 12 h, totaling 133 mg Folltropin (Folltropin-V; Bioniche Animal Health, Canada) per heifer. Seven d after AI, heifers had their uteri flushed and embryos were recovered and graded according to the International Embryo Technology Society standards. Data were analyzed using the GLIMMIX procedure of SAS and results are presented as least-squares means ± SEM (P < 0.05). At the onset of the FSH treatment (Day 0 of the protocol), 1.7 M heifers had greater body condition score (BCS), BW and serum insulin concentrations than 0.7 M heifers (4.1 ± 0.1 vs. 3.0 ± 0.1; 462.5 ± 10.1 vs. 382.7 ± 10.4 kg; and 14.3 ± 1.7 vs. 3.5 ± 0.8 µIU/mL, respectively). The 0.7 M heifers had more follicles ≥6 mm at the time of the last FSH (Day 7; 47.9 ± 6.4 vs. 23.5 ± 4.3 follicles), related to a better follicle superstimulatory response to FSH. Similarly, 0.7 M heifers had more corpora lutea at the time of embryo collection (33.6 ± 1.4 vs. 15.7 ± 0.9) than the 1.7 M heifers, which resulted in greater number of recovered embryos and ova (9.9 ± 0.7 vs. 6.7 ± 0.6) and viable embryos (5.3 ± 0.5 vs. 3.8 ± 0.4), despite having similar proportions of viable embryos (∼62%). A negative correlation between circulating insulin and follicle superstimulatory response to FSH was observed (r = -0.68). Therefore, we conclude that high feed intake, for a long period of time, compromised the superovulatory response and embryo production potential of Bos indicus heifers possibly related to the elevation in circulating insulin.

Progesterone supplementation after ovulation: effects on corpus luteum function and on fertility of dairy cows subjected to AI or ET.

Three experiments were done to evaluate the effects of progesterone (P4) supplementation starting during metestrus on formation of the CL and on fertility of lactating dairy cows subjected to fixed-time artificial insemination (FTAI) or embryo transfer (ET). In experiment 1, 42 Holstein cows were randomly allocated to untreated (Control) or to receive an intravaginal implant containing 1.9 g of P4 from Day 3 to 20 after FTAI (controlled internal drug release [CIDR]). Blood samples were collected on Days 3, 4, 7, 11, 14, 17, 20, and 21 after FTAI to evaluate the effect of CIDR supplementation on plasma concentration of P4 using radioimmunoassay. Ultrasound scans were performed at Days 4, 7, 11, 14, and 20 to evaluate CL volume. In experiment 2, the effect on CIDR supplementation on fertility was evaluated in 668 Holstein and crossbred dairy cows that were subjected to FTAI and allocated randomly to untreated (AI-Control) or to receive a CIDR from Day 3 to 17 (AI-CIDR) after FTAI. In experiment 3, 360 Holstein cows were treated with PGF and after heat detection (Day 0), they were allocated to untreated (ET-Control) or to receive a CIDR from Day 4 ± 1 to 8 ± 1 (ET-CIDR-4) or a CIDR from 4 ± 1 to 18 ± 1 (ET-CIDR-14). In vitro-produced embryos were transferred on Day 8 ± 1. Pregnancy diagnoses were performed by ultrasound. In experiment 1, there was interaction between treatment and day in relation to plasma P4 on Days 4 and 7 due to CIDR supplementation. Independent of treatment, pregnant cows had higher plasma P4 from Day 14 to 21 than nonpregnant cows (P ≤ 0.05). Supplementation with CIDR did not alter CL development. In experiment 2, there was no effect of supplementation of P4 on pregnancy per AI on Day 32 (32.0% vs. 31.8%, for AI-Control and AI-CIDR, respectively) or pregnancy loss (15.6% vs. 17.6%, for AI-Control and AI-CIDR, respectively). In experiment 3, P4 supplementation compromised pregnancy per ET (P/ET) on Day 32 in both supplemented groups (39.7% vs. 21.3% vs. 15.2%, for ET-Control, ET-CIDR-4, and ET-CIDR-14, respectively), with no effect on pregnancy loss. Therefore, although CIDR insertion on Day 3 after FTAI did not affect CL function and increased circulating P4, it did not increase pregnancy per AI in lactating dairy cows submitted to FTAI. Moreover, P4 supplementation decreased pregnancy per ET in lactating recipient cows.