Exposure to progesterone before an ovulation synchronization protocol increases the follicular diameter and fertility of multiparous suckled Bos taurus cows.

The objective of this study was to evaluate the effect of administering injectable progesterone (P4i) before a timed artificial insemination (TAI) protocol on the follicular growth, ovulation, and pregnancy rate of Bos taurus suckled cows. The effect of P4i administration before the TAI on the pregnancy rate (P/AI) was evaluated in 576 suckled Bos taurus cows at 30-90 days postpartum. In addition, the effect of P4i administration before TAI on follicular dynamics was evaluated in subgroup of 401 suckled Bos taurus cows. On Day -10 (D-10), cows were divided into two experimental groups (Control and P4i). In this moment, P4i cows received i.m. 150 mg of injectable long-action progesterone. After that, both experimental groups received a synchronization protocol (Day 0; D0) that consisted of administration i.m. of 2 mg of estradiol benzoate and a progesterone intravaginal insert on D0. On Day 8 (D8), the progesterone insert was removed, and the cows received 500 µg of cloprostenol, 400 IU of eCG, and 1 mg of estradiol cypionate. TAI was performed 48 h after the removal of the progesterone insert. The ultrasound exams were performed in a subgroup of cows on Days 0, 8, 10 and 12 to evaluate the diameter of the largest follicle, rate of follicular growth and risks of single and double ovulation. The pregnancy diagnosis was performed 30 days after TAI in all cows to determine the pregnancy rate. The diameter of the largest follicle, on D10 (P = 0.84), rate of follicular growth (P = 0.14), ovulation rate (P = 0.40) and double ovulation rates (P = 0.23) did not differ between experimental groups. The pregnancy rate was greater in the P4i group [Control 46.2 % (133/288) vs. P4i 55.6 % (160/288); P = 0.03]. The diameter of the largest follicles (LF) on D0 (Control 11.6 ± 0.2 vs. P4i 13.3 ± 0.3) was greater (P = 0.01) in the P4i group. In conclusion, injectable progesterone before the ovulation synchronization protocol increased the diameter of the largest follicle on the D0 and the pregnancy rate in multiparous Bos taurus suckled beef cows.

Use of injectable progesterone at the beginning of the TAI protocol is not necessary in super-early resynchronization started 14 days after artificial insemination in Bos indicus beef heifers.

In a protocol for the resynchronization of ovulation starting 14 days (D14) after timed artificial insemination, the effects of short-action injectable progesterone (P4i) administration and the length of treatment with an intravaginal progesterone (P4) device on follicular dynamics, the conception rate (P/AI) and the percentage of false positives were evaluated in 1065 Nelore heifers previously timed-inseminated. On D14, P4 devices were inserted, and heifers allocated, based on a 2 × 2 factorial design, to receive (P4i) or not 50 mg of P4i (NoP4i), and to remove the P4 device after 6 (6Day) or 8 days (8Day). At the time of P4 device removal (D20 and D22), non-pregnancy diagnosis was performed using vascularization of the corpus luteum (VCL) evaluated. At this time, non-pregnant heifers received 150 µg of D-cloprostenol, 0.6 mg of estradiol cypionate and 300IU of eCG. TAI was performed 48 h after P4 device removal. For these heifers, ultrasound examinations were performed at P4 device removal and at TAI to evaluate follicular dynamics and at 30 days after TAI to evaluate the P/AI. Pregnant heifers based on VCL were evaluated using B-mode ultrasonography 10 days after Doppler ultrasound to evaluate the percentage of false positives. Statistical analyses were performed using the GLIMMIX procedure of SAS. There were no interaction effects between P4i and duration of treatment with a P4 device. The P/AI was diagnosed by Doppler, 1st TAI, 2nd TAI, total and percentage of false positives did not differ between heifers receiving or not P4i. Similarly, duration of treatment with a P4 device did not influence the P/AI by Doppler, 1st TAI, 2nd TAI and total. However, the percentage of false positives diagnoses was higher in 6Day heifers (P = 0.01). The diameter of largest follicle at P4 device removal was greater in the 8Day heifers (P = 0.001), and at TAI was higher in the P4i-treated heifers (P = 0.03). Additionally, the percentage of false positives diagnoses was lower in heifers that ovulated to the 1st TAI protocol (P = 0.001). In conclusion, for resynchronization 14 days after TAI, it is not necessary to inject P4i at the beginning of the protocol. In addition, P4 device removal after 6 instead of 8 days increases the percentage of false positives because of the earlier diagnosis (20 days after TAI), but does not interfere in P/AI of resynchronization protocol. Furthermore, the percentage of false positives is higher in heifers that did not ovulate to 1st TAI protocol.

Effects of eCG and FSH in timed artificial insemination treatment regimens on estrous expression and pregnancy rates in primiparous and multiparous Bos indicus cows.

Effects were evaluated in Bos indicus cows of eCG and FSH on follicular growth, estrous expression, and pregnancy per artificial insemination (P/AI) as a result of fixed-time artificial insemination (TAI). In Experiment 1, extent of timing-of-ovulation synchronization among cows was evaluated after imposing an estrogen/progesterone-based treatment regimen. At progesterone device removal (D8), cows were administered: eCG, or FSH or served as untreated Controls. In Experiment 2, percentage of cows P/AI was evaluated when the Experiment 1-treatment regimen was imposed. On D10, all cows were artificially inseminated. In Experiment 3, cows were assigned to two treatment groups (Control and eCG) on D8 to evaluate percentage of cows P/AI and estrous expression. In Experiment 1, follicular dynamics were similar among treatment groups. In Experiment 2, follicular growth was greater (P = 0.0001) with the eCG treatment. There was an interaction of treatment × parity (P = 0.007) on percentage of cows P/AI. There was a greater percentage of primiparous cows P/AI in the eCG-treated than Control and FSH-treated cows. There was a greater percentage of eCG-treated multiparous cows pregnant as a result of TAI than Control cows. There was an interaction of treatment × parity (P = 0.005) on P/AI in Experiment 3, in which the eCG effect was more pronounced in primiparous cows. Treatment with FSH, therefore, was not as effective as eCG in stimulation of follicular growth or enhancing percentage of cows pregnant as a result of TAI. Physiological effects of eCG, however, were also more evident in primiparous cows.

Nuclear maturation kinetics and in vitro fertilization of immature bovine oocytes injected into pre-ovulatory follicles.

The maturation kinetics and in vitro fertilization of immature bovine oocytes injected by the intra-follicular oocyte injection (IFOT) technique into pre-ovulatory follicles of previously synchronized cows were evaluated. In Experiment 1, grade I, II and III cumulus-oocyte complexes (COCs) were randomly distributed to one of three Groups: Matvitro22 (COCs matured in vitro for 22 h), MatFol20 and MatFol28 (COCs matured in vivo after being injected into a pre-ovulatory follicle of previously synchronized cows for 19.8 ± 0.1 h and 28.3 ± 0.1 h, respectively). Cows received 12.5 mg of LH (Lutropin, Bioniche, Canada) at the time of IFOT in the MatFol20 Group or 10 h after IFOT in the MatFol28 Group. MatFol20 and MatFol28 COCs were aspirated approximately 20 h after the LH injection for nuclear maturation kinetics and recovery rate assessment. In Experiment 2, grade I, II, and III COCs were randomly distributed into two Groups: Matvitro22 Group, COCs were matured and fertilized in vitro, and MatFol20 Group, COCs were matured as in the MatFol20 Group in Experiment 1, but COCs were fertilized in vitro. Putative zygotes were classified as fertilized, unfertilized or polyspermic. In Experiment 1, the recovery rate was lower (P < 0.001) in the MatFol20 Group (52.9%, 91/172) compared with MatFol28 (72.9%, 113/155). Rate of oocytes in germinal vesicle stage, metaphase I, anaphase I and telophase I were similar among Groups. However, oocytes matured in vivo for 28.3 h had lower rate of metaphase II (P = 0.001) and greater rates of degenerated (P = 0.001) and parthenogenetically activated (P = 0.001) oocytes. In experiment 2, the rates of polyspermy and degenerated were similar between Groups. However, the rate of fertilized oocytes was greater (P = 0.05) in oocytes in the MatFol20 Group. It is concluded that oocyte in vivo maturation for 19.8 h after IFOT does not compromise the nuclear maturation kinetics and increases in vitro fertilization rates. However, the extra 10 h of intra-follicular incubation time decreased oocyte viability.

Subclinical mastitis interferes with ovulation, oocyte and granulosa cell quality in dairy cows.

The objective was to evaluate the effect of mastitis by somatic cell count (SCC) on follicular growth, ovulation, oocytes and cumulus cells quality and on the concentration and size of exosomes in follicular fluid of dairy cows. In the study, crossbred cows (Bos taurus - Holstein x Bos indicus - Gir) were classified for analysis as Control (SCC<200.000 cells/mL) and Mastitis (SCC>400.000 cells/mL) groups. In experiment 1 (follicular dynamics), cows (n = 57) were submitted to ultrasound evaluations every 24 h, from progesterone-releasing-intravaginal-device (PRID) removal (D8) until 48 h later (D10). Thereafter, evaluations were performed every 12 h, until ovulation or up to 96 h after PRID removal. In experiment 2 (oocyte, cumulus complexes, and follicular fluid evaluation), cows (n = 26) were submitted to follicular aspiration (OPU) for oocyte quality and cumulus cells transcript evaluation. The amount of cumulus complexes transcripts (BCL2, BAX, PI3K, PTEN, FOXO3) was determined by Real-Time Polymerase Chain Reaction. Moreover, seven days after the OPU session, the dominant follicle was aspirated. Exosomes were isolated from the follicular fluid for evaluation of particle size and concentration. Ovulation rate [Control 77.4% (24/31) and Mastitis 57.7% (15/26); P = 0.09] and viable oocytes rate [Control 59.1% (130/220) and Mastitis 41.9% (125/298); P = 0.01] were higher in Control animals. Additionally, there was a greater number of degenerate oocytes [Control 6.7 ±â€¯1.2 and Mastitis 13.3 ±â€¯5.5; (P = 0.001)] in subclinical mastitis cows. There was greater abundance (P = 0.003) of BAX cumulus cell transcripts and exosome mean (P = 0.03) and mode (P = 0.02) was smaller in subclinical mastitis cows. In conclusion, ovulation rate, oocyte quality, and exosome diameter were smaller in cows with SCC>400.000 cells/mL.

Presynchronization by induction of a largest follicle using a progesterone device in GnRH-based-ovulation synchronization protocol in crossbred dairy cows.

The objective was to compare the fertility of dairy cows using a presynchronization protocol by induction of a largest follicle using a progesterone intravaginal device prior to an Ovsynch protocol (P4synch) with the Double-Ovsynch in lactating dairy cows. Lactating Bos indicus x Bos taurus crossbred cows (n = 440) were randomly allocated to one of two treatments: (I) Double-Ov (n = 228), GnRH (D-17), PGF2α 7 days later (D-10) and GnRH 3 days later (D-7) followed by an Ovsynch protocol 7 days later (GnRH on D0, PGF on D7, GnRH on D9); (II) P4synch (n = 212), insertion of a sustained release progesterone intravaginal device (D-10), 10 days later (D0) an Ovsynch protocol was initiated (GnRH on D0, PGF on D7, GnRH on D9) with progesterone device withdrawal on Day 7. All cows were artificially inseminated (TAI) 16 h after the second GnRH of the Ovsynch protocol and pregnancy diagnosis was performed by transrectal ultrasonography 30 and 60 days after TAI. A subset of cows (n = 52 for Double-Ov and n = 50 for P4synch) ultrasonography was performed on days 0, 7, 9 and 24 of the experimental period. There were no differences among treatments on presynchronization rate [presence of a follicle>12 mm on D0, Double-Ov 94.2% (49/52) and P4synch 92.0% (46/50); P = 0.66], follicular diameter on the 1st GnRH (Double-Ov 17.2 ±â€¯0.7 mm e P4synch 18.6 ±â€¯0.9 mm; P = 0.28), ovulation rate to the 1st GnRH [Double-Ov 86.3% (44/51) and P4synch 81.2% (39/48); P = 0.50], synchronization rate [presence of a follicle>12 mm on D9; Double-Ov 84.6% (44/52) and P4synch 86.0% (43/50); P = 0.84], follicular diameter on the 2nd GnRH (Double-Ov 17.5 ±â€¯0.6 mm and P4synch 18.0 ±â€¯0.5 mm; P = 0.48), ovulation rate to the 2nd GnRH [Double-Ov 90.9% (40/44) and P4synch 86.0% (37/43); P = 0.48] and CL diameter on D24 (Double-Ov 27.9 ±â€¯0.7 mm and P4synch 29.4 ±â€¯0.9 mm; P = 0.19). Corpus luteum presence on D0 was different (P = 0.03) among treatments [Double-Ov 57.7% (30/52) and P4synch 36.0% (18/50)]. There was no difference (P = 0.85) among the pregnancy per AI on day 30 [Double-Ov 39.0% (89/228) and P4synch 40.1% (85/212)], on day 60 [Double-Ov 34.8% (79/227) and P4synch 38.7% (82/212); P = 0.41] and pregnancy loss [Double-Ov 10.2% (9/88) and P4synch 3.5% (3/85); P = 0.08]. The presynchronization by induction of a largest follicle using a sustained release progesterone device prior to Ovsynch yielded similar results compared with the Double Ovsynch protocol on follicular development patterns and on the fertility of lactating dairy cows.

Exposure to progesterone previous to the protocol of ovulation synchronization increases the follicular diameter and the fertility of suckled Bos indicus cows.

The objective was to evaluate the effect of injectable progesterone previous to the timed artificial insemination (TAI) protocol on follicular growth, ovulation and pregnancy rate of suckled Bos indicus cows. In experiment 1, 10 days before the beginning to TAI protocol (D-10), 431 suckled-anestrus Nelore cows (249 multiparous and 182 primiparous), were allocated to one of three treatments groups (control, P4i and P4iGnRH). At this moment, cows in the P4i and P4iGnRH group received 150 mg of injectable progesterone intramuscularly (Sincrogest injetável®, Ouro Fino, Brazil). On Day 0 (D0), all cows were synchronized using an estradiol/progesterone-based TAI protocol. Simultaneously, in the P4iGnRH group, cows received 10 µg of Busereline intramuscularly (Sincroforte®, Ouro Fino, Brazil). Ultrasound exams were performed to evaluate the diameter of the largest follicle (D0, D8 and D10), ovulation rate and diameter of the corpus luteum (D24). In experiment 2, 681 suckled Nelore cows (356 multiparous and 325 primiparous) were synchronized using an estradiol/progesterone-based TAI protocol and received treatments similar to experiment 1. TAI was performed 48 h after removal of the progesterone (P4) device. Pregnancy diagnosis was 30 d after TAI. In experiment 3, blood samples were collected to evaluated the progesterone concentration for 168 h after administration of injectable progesterone intramuscularly. Statistical analyses were performed by GLIMMIX procedure of SAS. In experiment 1, the diameter of the largest follicle (LF) on D10 (P = 0.21), follicular growth rate (P = 0.34) and ovulation rate (P = 0.62) were similar among experimental groups. However, there was difference among groups for the LF on D0 [Control (10.9 ±â€¯0.2 mm)b, P4i (12.7 ±â€¯0.3 mm)a and P4iGnRH (12.6 ±â€¯0.3 mm)a; P = 0.001], LF on D8 [Control (9.7 ±â€¯0.2 mm)b, P4i (10.4 ±â€¯0.2 mm)a and P4iGnRH (9.9 ±â€¯0.2 mm)ab; P = 0.05], presence of the CL on D8 [Control 0% (0/136)b, P4i 0% (0/140)band P4iGnRH 26.4% (38/144)a; P = 0.001], diameter of the CL on D24 [Control (19.7 ±â€¯0.4 mm)ab, P4i (20.1 ±â€¯0.4 mm)a and P4iGnRH (18.5 ±â€¯0.4 mm)b; P = 0.001] and pregnancy rate [Control 35.0% (78/223)b, P4i 45.9% (105/229)a and P4iGnRH 40.6% (93/229)ab; P = 0.01]. The circulating concentration of P4 remained above 1.5 ng/mL until 168 h after the P4i treatment. In conclusion, the injectable progesterone previous to the TAI protocol increased diameter of the LF on D0 and D8 without interfering on the ovulation rate. Furthermore, such exposure increases the pregnancy rate in suckled Nelore cows.

Molecular and endocrine factors involved in future dominant follicle dynamics during the induction of luteolysis in Bos indicus cows.

The growth profiles of the future dominant follicle (DF) and subordinate follicle (SF) and the gene expression of the granulosa cells during luteolysis induction in Bos indicus cows were evaluated. Forty cows were synchronized with a progesterone and estradiol based protocol. After synchronization, cows with a corpus luteum (CL) were evaluated by ultrasonography every 12 h, beginning at eight days post ovulation. Cows identified with a follicle of at least 6.0 mm in diameter in the second wave were split into two groups (BD-before follicular deviation and AD-after follicular deviation. In the BD group cows received 500 µg of cloprostenol (a synthetic analogue of prostaglandin F2α) when the DF reached a mean diameter of 7.0 mm (6.5-7.5 mm). In the AD group, cows received 500 µg of cloprostenol when the DF reached a mean diameter of 8.0 mm (7.5-8.5 mm). Cows in both groups were submitted to aspiration of the DF at 96 and 72 h after prostaglandin was given. Follicular aspirations were performed to quantify IGF1R, LHR and PAPPA transcripts in the granulosa cells. The diameter of the DF at the moment of prostaglandin administration (P = 0.001) and the growth rate of the SF (P = 0.05) were greater in the AD group. There was greater abundance of LHR transcripts in BD cows (P = 0.04). The remaining variables tested were similar between the experimental groups (P > 0.05). In conclusion, the induction of luteolysis before follicular deviation does not interfere with dominant follicle dynamics. However, it causes granulosa cell LHR down regulation.

Although it induces synchronized ovulation, hCG reduces the fertility of Santa Ines ewes submitted to TAI / Embora induza a ovulação sincronizada, hCG reduz a fertilidade de ovelhas Santa Inês submetidas à IATF

The aim of this study was to evaluate hCG treatment on ovarian response and on pregnancy rate using a 9-day oestrus synchronization protocol in Santa Ines ewes. On a random oestrus cycle day, ewes received an intravaginal progesterone device (Primer-PR®, Tecnopec, Brazil). Nine days later (Day 9), 30µg of d-cloprostenol (Prolise®, Syntex, Argentina) and 250IU of eCG (Folligon®, Intervet, Brazil) were administered and the progesterone device was removed. This moment, the ewes were randomly assigned on two groups: Control Group and hCG Group. In the hCG Group, the ewes received 500IU of hCG (Vetecor®, Hertape-Calier, Spain) 24h after device removal. In the Control Group, the ewes did not receive any ovulation inductor. Control and hCG Groups ewes were inseminated 60h and 48h after device removal, respectively. There was no difference between the groups regarding the first ovulatory follicle diameter and the second ovulatory follicle. hCG Group ewes had shorter interval between device removal and ovulation (Control: 79.9±15.4h and hCG: 54.7±4.9h; P=0.001) and more synchronized ovulations. However, the treatment with hCG decreased the pregnancy rate after TAI (P=0,009). In conclusion, hCG administration improves ovulatory synchronisation, but causes a decrease in the pregnancy rate.(AU)

Avaliou-se o tratamento com hCG na resposta ovariana e na taxa de prenhez utilizando protocolo de sincronização do estro de nove dias em ovelhas Santa Inês. As ovelhas receberam um dispositivo intravaginal de progesterona em fase aleatória do ciclo (dia zero= D0). No momento da remoção do dispositivo (D9), as fêmeas receberam 30µg de d-cloprostenol (Prolise®, Syntex, Argentina) e 250UI de eCG (Folligon®, Intervet, Brasil). Nesse momento, as ovelhas foram aleatoriamente distribuídas em dois grupos de tratamento: controle sem indução de ovulação e tratamento com 500UI hCG para indução de ovulação. As ovelhas dos grupos controle e hCG foram inseminadas 60h e 48h após a remoção do dispositivo, respectivamente. Não houve diferença entre os grupos para o diâmetro do primeiro e do segundo folículo pré-ovulatório. As avelhas do grupo hCG apresentaram menor intervalo entre a remoção do dispositivo e a ovulação (grupo controle: 79.9±15.4h e grupo hCG: 54.7±4.9h; P=0.001) e maior sincronização das ovulações. No entanto, o tratamento com hCG diminuiu a taxa de prenhez após a IATF (P=0,009). Conclui-se que, apesar de a administração de hCG aumentar a sincronização da ovulação, reduz a taxa de prenhez.(AU)