The effects of administering estradiol benzoate plus progesterone during the growth or static phases of the dominant follicle in CIDR-treated lactating dairy cows

We studied the effects of administering estradiol benzoate (EB) plus progesterone (P4) as part of a CIDR-based protocol during the growth or static phases of dominant follicle development on follicular wave emergence, follicular growth, synchrony of ovulation and pregnancy rate following CIDR withdrawal, treatment with PGF(2alpha) and GnRH, and fixed-time artificial insemination (TAI). Forty-one previously synchronized lactating Holstein dairy cows were randomly allocated to three treatment groups. The control group (n=14) received a CIDR on the third day after ovulation only (Day 0). The two treatment groups were administered CIDRs comprising 2 mg EB and 50 mg P4 either on the third (T1, n=14) or eighth day (T2, n=13) after ovulation (Day 0). All cows received PGF(2alpha) after CIDR removal on Day 7, GnRH on Day 9, and TAI 16 h after GnRH treatment. The proportion of cows with follicular wave emergence within 8 days of treatment differed (P<0.01) among the control (14.3%), T1 (85.7%), and T2 groups (92.9%). However, the mean intervals between treatment and wave emergence were not significantly different. There were significant differences in the diameters of the dominant follicles on Day 7 (P<0.01) and in preovulatory follicles on Day 9 (P<0.01), with the largest follicles observed in the control group and the smallest follicles observed in the T2 group. In contrast, the numbers of cows showing synchronous ovulation after GnRH treatment (92.9 to 100.0%) and pregnancy following TAI (46.2 to 50.0%) were similar between the treatment groups. The results showed that, irrespective of the phase (growth or static) of the dominant follicle, administration of 2 mg EB plus 50 mg P4 to CIDR-treated lactating dairy cows induced consistent follicular wave emergence and development, synchronous ovulation after GnRH administration, and similar pregnancy rates following TAI.     

Effect of short-term calf removal at three stages of a follicular wave on fate of a dominant follicle in postpartum beef cows.

The hypothesis that ovulation in response to short-term (48 h) calf removal (CR) is dependent on the developmental stage of the dominant follicle was tested in two studies. The objective of Exp. 1 was to characterize the fate of a dominant follicle following 48-h CR on d 2, 4, or 8 of a postpartum follicular wave. Ovaries of 61 beef cows were examined daily by transrectal ultrasonography starting at d 20 to 21 postpartum. Treatments were no CR (n = 14) and CR on d 2 (n = 12), 4 (n = 16), or 8 (n = 10) of first detected follicular wave. Percentage of cows that ovulated a dominant follicle following treatment was not different among groups (P = 0.62). Maximum size of dominant follicles was larger in cows that ovulated (P = 0.002) than in cows that did not ovulate. The objectives of Exp. 2 were 1) to determine whether a follicular wave could be synchronized in anestrous cows following injection of 1 mg of estradiol benzoate (EB) and 200 mg of progesterone (P4; EB + P4); 2) to characterize the fate of dominant follicles following 48-h CR at three stages of a synchronized follicular wave; and 3) to determine whether estrous cycles of normal length followed ovulation in cows pretreated with EB + P4. Ovaries of 50 anestrous beef cows were examined daily as in Exp. 1. Treatments were sesame oil (SO) injected (i.m.) on d 25 postpartum and no CR (n = 9); EB + P4 and no CR (n = 9); EB + P4 and CR on 6 (n = 12), 8 (n = 9), or 12 (n = 11) d after injection. The EB and P4 injections were given on d 25 postpartum. Variability in day of emergence of subsequent follicular waves was lower in cows receiving EB + P4 than in SO-injected cows (P < 0.05). The percentage of cows that ovulated was not different (P = 0.16), but CR increased the percentage of cows that ovulated when groups that received EB + P4 were compared to the EB + P4 group that did not have CR (53.1 vs 11.1%, respectively; P < 0.05). Maximum diameter of dominant follicles was larger (P = 0.05) in ovulatory follicles. The luteal phase was longer (P < 0.03) in cows receiving EB + P4 injection (10.6 +/- 1.2 d) than in cows receiving SO (4.4 +/- 2.2 d). In summary, the maximum size of ovulatory follicles was greater than that of nonovulatory follicles, the ovulatory response of postpartum anestrous cows was maintained through d 8 of a follicular wave, synchronization of follicular waves was accomplished in postpartum cows using EB + P4, and the subsequent luteal phase length was increased in animals that were administered EB + P4.     

Comparison of two types of CIDR-based timed artificial insemination protocols for repeat breeder dairy cows

This study compared two types of controlled internal drug release (CIDR)-based timed artificial insemination (TAI) protocol for treatment of repeat breeder dairy cows. In the first trial of the experiment, 55 repeat breeder cows were randomly assigned to the following two treatments. (1) In the EB group, a CIDR device was inserted into the cows, and then the cows were administered an injection of 1 mg estradiol benzoate (EB) plus 50 mg progesterone (P4; Day 0). On Day 7, they were given an injection of PGF(2alpha) and the CIDR device was removed. The cows were given an injection of 1 mg EB on Day 8 and were subjected to TAI 30 h later (n=27). (2) In the gonadotrophin releasing hormone (GnRH) group, a CIDR device was inserted into the cows, and then the cows were administered an injection of 250 microg gonadorelin (GnRH; Day 0). On Day 7, they were given an injection of PGF(2alpha) and the CIDR device was removed. The cows were given an injection of 250 microg GnRH on Day 9 and were subjected to TAI 17 h later (n=28). In the second trial, 41 repeat breeder cows that were confirmed as not pregnant in the first trial were randomly assigned to the same two treatments used in the first trial (an EB group of 20 cows and a GnRH group of 21 cows). The ovaries of 15 cows from each group were examined by transrectal ultrasonography in order to observe the changes in ovarian structures, and blood samples were collected for analysis of serum P4 concentrations. The pregnancy rates following TAI in the first (18.5 vs. 32.1%) and second (40.0 vs. 38.1%) trials and the combined rates (27.7 vs. 34.7%) did not differ between the EB and GnRH groups. The proportions of cows with follicular wave emergence within 7 days did not differ between the EB (12/15) and GnRH groups (13/15). The interval to wave emergence was shorter (P<0.01) in the GnRH group than in the EB group, but there was no difference in the mean diameters of dominant follicles on Day 7 between the groups. Moreover, the proportions of cows with synchronized ovulation following a second EB or GnRH treatment did not differ between the groups. In conclusion, treatment with either EB or GnRH in a CIDR-based TAI protocol results in synchronous follicular wave emergence, follicular development, synchronous ovulation, and similar pregnancy rates for TAI in repeat breeder cows.     

The effects of varying the interval from follicular wave emergence to progestin withdrawal on follicular dynamics and the synchrony of estrus in beef cattle

The objective of this experiment was to examine the effects of varying the interval from follicular wave emergence to progestin (controlled internal drug-releasing insert, CIDR) withdrawal on follicular dynamics and the synchrony of estrus. A secondary objective was to assess the effects of causing the dominant follicle (DF) to develop in the presence or absence of a corpus luteum (CL) on follicular dynamics and the synchrony of estrus and ovulation. The experiment was designed as a 2 x 2 x 2 factorial arrangement of treatments with injection of GnRH or estradiol-17 beta and progesterone (E2 + P4) at treatment initiation, duration of CIDR treatment, and injection of PG (prostaglandin F2 alpha) or saline at the time of CIDR insertion as main effects. Estrous cycles (n = 49) in Angus cows were synchronized, and treatments commenced on d 6 to 8 of the estrous cycle. Cows were randomly assigned to receive a CIDR containing 1.9 g of P4 for 7 or 9 d. Approximately half the cows from each CIDR group received either GnRH (100 micrograms) or E2 + P4 (1 mg of E2 + 100 mg of P4) at CIDR insertion. Cows in GnRH or E2 + P4 groups were divided into those that received PG (37.5 mg) or saline at CIDR insertion. All cows received PG (25 mg) 1 d before CIDR removal. Daily ovarian events were monitored via ultrasound. The intervals from GnRH or E2 + P4 treatment to follicular wave emergence were 1.4 and 3.3 d, respectively (P < 0.05). The interval from follicular wave emergence to CIDR removal was longer (P < 0.05) for cows treated with GnRH (6.6 d) than those treated with E2 + P4 (4.7 d) and longer (P < 0.05) for those fitted with a CIDR for 9 d (6.5 d) than those with a CIDR in place for 7 d (4.8 d). Cows treated with PG or GnRH at CIDR insertion had a larger (P < 0.05) DF at CIDR removal than those treated with saline or E2 + P4. Treatment with a CIDR for 9 d also resulted in a larger (P < 0.07) DF at CIDR removal compared with cows fitted with a CIDR for 7 d. The interval from CIDR removal to estrus was shorter (P < 0.05) in cows treated with PG than those treated with saline. The synchrony of estrus and ovulation was not affected by any of the treatments (P > 0.05). Altering the interval from follicular wave emergence to progestin removal or creating different luteal environments in which the DF developed caused differences in the size of the DF at CIDR removal and the timing of the onset of estrus, but it did not affect the synchrony of estrus or ovulation.     

Changes in follicular vascularity during the first follicular wave in lactating cows

Increase in the blood supply to individual follicles appears to be associated with follicular growth rates and the ability to become the dominant follicle, while reduced thecal vascularity appears to be closely associated with follicular atresia. Therefore, this study aimed to determine the real-time changes in the vascularity of the follicle wall during the first follicular wave in cycling Holstein cows. Normally cycling and lactating cows (n=5) were examined by transrectal color Doppler ultrasonography (the sensitivity for velocity: > 2 mm/sec) to determine the changes in the vasculature of the follicle wall (presence or absence of blood flow) and the diameter of follicles. A new follicular wave and ovulation were induced by GnRH injection at 48 h after an injection of PGF2alpha analogue. The ovaries were scanned daily for 7 days after GnRH injection. Follicles >2.5 mm were classified into 3 groups by the changes in diameter as follows: 1) largest follicle, 2) second largest follicle, and 3) small follicles, which included all other follicles >2.5 mm. Before the follicle selection, there was no significant difference in the percentage of follicles with detectable blood flow between the subsequently determined largest and second largest follicles. After the follicle selection, the percentage of follicles with detectable blood flow significantly decreased among the second largest follicles. In addition, small follicles with detectable blood flow kept larger diameters than those without detectable blood flow from one day before the occurrence of follicle selection. It is likely that maintenance of follicle vasculature and appropriate blood supply to the larger follicles is essential for follicle dominance. In small follicles, the presence of blood flow within the wall also appears to be required for recruitment. Consequently, the data suggest that the change of the blood supply to an individual follicle closely relates to the dynamics of follicular growth in the first follicular wave in the cow.     

Comparison of oestrus synchronisation programmes in dairy cattle using oestradiol benzoate, short-acting progesterone and cloprostenol, or buserelin and cloprostenol

AIM: To evaluate the efficacy of a programme using oestradiol benzoate, progesterone and the prostaglandin-F2 (PG) analogue, cloprostenol, to synchronise oestrus and ovulation in dairy cows, compared with a programme using a gonadotropinreleasing hormone (GnRH) agonist, buserelin, and cloprostenol. METHODS: Twenty non-lactating dairy cows, at random stages of the oestrus cycle, were randomly assigned to 1 of 2 treatments. In Treatment 1 ( OPPG; n=10), cows were injected with 2 mg oestradiol benzoate intramuscularly (IM) plus 200 mg progesterone subcutaneously (SC) on Day 0, followed by 500 microg cloprostenol IM on Day 9 and 1 mg oestradiol benzoate on Day 10. In Treatment 2 (GPG; n=10), cows were injected with 10 microg buserelin IM on Day 0, 500 microg cloprostenol IM on Day 7 and 10 microg buserelin on Day 9. The ovaries of all cows were examined by ultrasonography, using an 8 MHz probe, from 5 days before the initial treatment until ovulation. Cows were observed for oestrus 3 times daily for 7 days after cloprostenol treatment. Blood samples were collected daily for determination of progesterone, and 6-hourly for 36 h after the second oestradiol or buserelin injection for the determination of follicle stimulating hormone (FSH) and luteinising hormone (LH) concentrations. RESULTS: The percentage of cows observed in oestrus was higher in the OPPG group than in the GPG group (100% vs 55.6%, p=0.018). Treatment with either short-acting progesterone plus oestradiol benzoate or buserelin was followed by atresia or ovulation of the dominant follicle. Emergence of a new follicular wave occurred earlier (p>0.001) in the GPG group (2.2+/-0.2 days) than in the OPPG group (3.6+/-0.2 days). There was no significant difference between treatment groups in the variation of time of follicular wave emergence or size of the largest follicles at either the time of initial treatment (10.8+/-1.4 mm vs 11.1+/-0.8 mm), cloprostenol treatment (13.8+/-0.7 mm vs 14.0+/-1.3 mm) or of ovulation (15.4+/-0.7 mm vs 17.6+/-1.1 mm; p=0.10). The LH surge occurred sooner after the second injection of buserelin (4.0+/-1.0 h) than after the second injection of oestradiol benzoate (22.8+/-1.2 h; p>0.001). The interval between the second injection of oestradiol benzoate or buserelin and ovulation did not differ significantly between treatment groups (1.7+/-0.3 days vs 1.6+/-0.2 days; p=0.69). CONCLUSIONS: The use of short-term progesterone treatment, combined with oestradiol benzoate for follicular wave synchronisation, and cloprostenol to cause lysis of residual luteal tissue, is a promising alternative to established methods of oestrus synchronisation in cows.     

Effect of follicular aspiration at the onset of progesterone-based timed artificial insemination on the follicular dynamics and fertility of early postpartum Japanese black cows

The effect of gonadotropin-releasing hormone analogue (GnRH-A) or follicular aspiration at the onset of progesterone-based timed artificial insemination (TAI) on subsequent follicular growth and synchronization of ovulation was examined in early postpartum Japanese Black cows. A total of 40 (22 in Exp. 1 and 18 in Exp. 2) Japanese Black cows at 20-30 days postpartum were fitted with a progesterone releasing internal device (PRID) for 7 days, injected with a prostaglandin F2α analogue upon removal of the PRID and GnRH-A 48 h later, and inseminated 18 h after GnRH-A injection. In Exp. 1, the animals were divided into three groups (untreated control, GnRH-A injection or follicular aspiration) of different treatments on the first day of PRID insertion (day 0), and the synchronized ovulation rate in the follicular aspiration group (100%; 8/8) tended to be higher (P = 0.077) than that in the control group (42.9%; 3/7). In Exp. 2, follicular growth in the GnRH (n = 9) and follicular aspiration (n = 9) groups was monitored by ultrasonography. Four out of the nine animals in the GnRH group had a corpus luteum on either day 4 or day 7 (OV group), and the other five animals had no induced ovulation (NOV group). The diameter of the ovulatory follicle on day 9 in the OV group (1.44 ± 0.11 cm) tended to be greater (P = 0.078) than that in the NOV group (1.13 ± 0.07 cm). Follicular aspiration at the onset of PRID-based TAI of early postpartum Japanese Black cows, regardless of the resumption of ovarian cyclicity, tended to result in a higher rate of synchronization of ovulation than that of the untreated controls.     

The effects of 3 gonadorelin products on luteinizing hormone release,ovulation, and follicular wave emergence in cattle

The objective was to determine luteinizing hormone (LH) secretion and follicular dynamics in cattle following administration of 3 gonadorelin formulations that are commercially available in Canada. In experiment 1, nonlactating Holstein cows (n = 4 per group) were randomly assigned to receive 100 micrograms gonadorelin diacetate tetrahydrate, intramuscularly (C; Cystorelin, or FE; Fertagyl). Blood samples (for LH analysis) were collected 0, 1, 2, and 4 hours after treatment. In experiment 2, nonlactating Holstein cows (n = 10 per group) were randomly allocated to receive 100 micrograms gonadorelin, intramuscularly as follows: 2 mL of C; 1 mL of FE; or 2 mL of Factrel (FA, gonadorelin hydrochloride). Gonadorelin treatment was done on days 6 or 7 after ovulation and blood samples for LH analysis were collected at 0, 1, 2, 4, and 6 hours after treatment. Ovaries were examined by ultrasonography, twice daily, to detect ovulation. A replicate was conducted using only C (n = 10) or FE (n = 10); blood samples were collected at 0, 1, 2, 3, and 4 hours. In experiment 3, beef heifers (n = 10 per group) were randomly assigned to receive 1 of 3 GnRH gonadorelin treatments (as in the first phase of experiment 2) on days 6 or 7 after ovulation and blood samples were collected at 0, 0.5, 1, 1.5, 2, and 4 hours. In experiments 2 and 3, both mean and mean peak plasma LH concentrations were higher (P < 0.05) in cattle treated with C. The proportion of dominant follicles that ovulated was higher (P < 0.02) in Holstein cows treated with C than in those treated with FE or FA (18/19, 11/19, and 4/7, respectively), but there was no significant difference among the products in beef heifers (6/10, 6/10, and 4/10, respectively). No significant differences were found in the interval from treatment to the emergence of the next follicular wave. In summary, C induced a greater LH release and this resulted in a higher ovulatory rate in Holstein cows but not in beef heifers.     

Expression of Angiopoietin (ANPT)-1, ANPT-2 and their Receptors in Dominant Follicles during Periovulatory Period in GnRH-Treated Cow

Ovarian follicular vasculature is involved in follicular development and ovulation. Angiopoietin (ANPT)-Tie system is important for vascularization of the tissue surrounding the developing follicles and corpus luteum (CL). To determine how the expression of ANPT-1, ANPT-2 and their receptors in the follicles would be associated with the ovulatory process, the present study was conducted to examine mRNA expressions of ANPT-1, ANPT-2 and their receptors during the periovulatory phase in gonadotrophin-releasing hormone (GnRH)-treated cows. The ovaries were collected by transvaginal ovariectomy (n = 5, cows/group) and the follicles (n = 5, one follicle/cow) were classified into following groups: before GnRH administration [before luteinizing hormone (LH) surge]; 3-5 h after GnRH (during LH surge); 10 h after GnRH; 20 h after GnRH; 25 h after GnRH (peri-ovulation); and early CL (days 2-3). The mRNA expression was analysed by quantitative real-time PCR (rotor-gene 3000). Angiopoietin-1 expression rapidly decreased at 3-5 h and kept low level at 10 h after GnRH treatment compared with that before GnRH, and returned to the level before LH surge in the follicles >20 h after GnRH treatment. The levels of ANPT-2 mRNA decreased at 10 and 25 h after treatment compared with other periods. The ratio of ANPT-2/ANPT-1 (an index for destabilization of blood vessels) increased in the follicles at 3-5 h after GnRH treatment only. Both of Tie-1 and Tie-2 receptor expressions decreased in the follicles at 25 h after GnRH treatment. The results of the present study indicated that mRNA expressions of ANPT-1, ANPT-2 and their receptors changed in the bovine follicles during periovulatory period. These results suggest that angiopoietin-Tie system is associated with the initiation of vasculature of follicle that grows towards ovulation.     

GnRH treatment at CIDR insertion influences ovarian follicular dynamics in Japanese black cows

Ovarian follicular dynamics and estrous synchronization after Gonadotropin-releasing hormone (GnRH) treatment at Controlled Internal Drug Releasing device (CIDR) insertion were investigated in Japanese Black cows. CIDR was inserted for eight cows at 7 days after estrus. Cows were allocated to either Group A: 8-day CIDR insertion with GnRH treatment on d 0 (n=4, d 0=CIDR insertion) or Group B: 8-day CIDR insertion (n=4). Both groups were injected with prostaglandin F2alpha (PGF2alpha) on d 7. Ultrasonography and blood sampling were performed twice daily. Intensive sampling was performed every 15 min for 8 hr to determine the pulsatile release of LH on d -1, d 5 and d 10. Three of four cows showed intermediate ovulation within 2 days after GnRH treatment during CIDR insertion in Group A, whereas no ovulation was found in Group B. Three of four cows in Group A and all four cows in Group B ovulated after CIDR removal. Plasma progesterone concentrations from d 3 to d 7 in three intermediate ovulatory cows in Group A (8.4 +/- 1.6 ng/ml) was significantly higher than those in Group B (4.1 +/- 1.2 ng/ml; 4 cows) during CIDR insertion (P<0.01). Interval to estrus and ovulation after CIDR removal was observed at 60.0 +/- 12.0 hr and 76.0 +/- 6.9 hr in three cows in Group A, and 75.0 +/- 15.1 hr and 93.0 +/- 20.5 hr in Group B, respectively. There was a significant increase in LH pulse frequency on d 10 compared on d -1 or d 5 in both groups (P<0.05), in addition those on d 10 in Group A tended to be higher than in Group B. As a result, GnRH treatment at CIDR insertion at 7 days after estrus induced intermediate ovulation with formation of corpus luteum (CL) and rather synchronized emergence of ovulatory follicle during CIDR insertion. These induced CL increased plasma progesterone concentrations and contributed to precise synchronization.     

Strategies to optimize reproductive efficiency by regulation of ovarian function

Pregnancy rate to the Ovsynch protocol can be improved if cows are presynchronized (i.e., two PGF(2alpha) injections given 14 days apart and the second injection of PGF(2alpha) given 12 days prior to the first GnRH of the Ovsynch program) so that a greater proportion of cows during the Ovsynch protocol ovulate to the first GnRH injection and have a CL at PGF(2alpha) injection. Pregnancy rates were normal in anestrous cows (39.6%) if they ovulated to both injections of GnRH. Estradiol cypionate (ECP) can be used to replace GnRH to induce ovulation as a modification of the Presync-Ovsynch program (i.e., Presync-Heatsynch). Pregnancy rates after TI were 37.1+/-5.8% for Presync-Ovsynch compared to 35.1+5.0% for Presync-Heatsynch. Use of ECP to induce ovulation was an alternative to GnRH in which greater uterine tone, ease of insemination and occurrence of estrus, improved acceptance by inseminators. A GnRH agonist (Deslorelin; 750 microg) implant inserted at 48 h after injection of PGF(2alpha), as a component of the Ovsynch protocol, induced ovulation, development of a normal CL and delayed follicular growth until 24 d after implant insertion. Utilization of Deslorelin implants (450 microg and 750 microg) to induce ovulation compared to GnRH (100 microg) within the Ovsynch protocol resulted in 27 d pregnancy rates (GnRH 100 microg, 39%; Deslorelin implants 450 microg, 40% and 750 microg, 27.5%) with 12.7%, 5.0% and 9.5% embryonic losses by 41 d of pregnancy, respectively. Induction of an accessory CL with injection of hCG on day 5 after insemination improved conception rates by 7.1%. Bovine somatotrophin injected at first insemination following a Presync-Ovsynch program in cycling-lactating dairy cows increased 74 days pregnancy rates (57.1%>42.6%).     

Clinical experience with native GnRH therapy to hasten follicular development and first ovulation of the breeding season

Breeding records of 48 Thoroughbred and Standardbred mares treated with native GnRH (500μg im, bid) during February—April, 1999 or 2000, on 7 farms in central Kentucky were retrospectively examined. Treated mares were classified as being in anestrus or early transition (n=42; if no signs of estrus occurred within 31/2 weeks and the largest follicle remained ≤25 mm in diameter or the first larger follicle(s) of the season regressed without ovulating), or were classified as being in late transition (n=6; if follicular growth achieved 30-40 mm diameter but ovulation had not yet occurred during the breeding season). Thirty-eight mares (38/48; 79%) ovulated in 13.7 ± 7.4 days. Interval to ovulation was negatively associated with size of follicles at onset of native GnRH therapy (P < 0.01). Per cycle pregnancy rate was 53% (19/36 mares bred). Ovulation inducing drugs were administered to 32 of the native GnRH treated mares (2500 units hCG intravenously, n = 20; deslorelin implant [Ovuplant™] subcutaneously, n=12), while 6 mares were not administered any additional drugs to induce ovulation. Per cycle pregnancy rate did not differ among mares treated only with native GnRH (2/5 mares bred; 40% PR), mares treated with native GnRH plus hCG (12/19 mares bred; 63% PR), or mares treated with native GnRH plus Ovuplant™ (5/12 mares bred; 42% PR) (P > 0.10). Additional treatment with either hCG or Ovuplant™ did not alter mean follicle size at ovulation or interovulatory interval (P > 0.10). The proportion of interovulatory intervals > 25 days was not different between mares receiving no additional treatment to induce ovulation (0/4; 0%) compared to mares receiving hCG to induce ovulation (3/8; 38%) (P > 0.10), but the proportion of interovulatory intervals > 25 days was greater for mares receiving Ovuplant™ to induce ovulation (5/7; 71%) compared to mares receiving no additional treatment to induce ovulation (P < 0.05). The proportion of mares with extended interovulatory intervals (i.e., > 25 days) did not differ between mares with follicles < 15 mm diameter (4/8, 50%) and those with follicles > 15 mm diameter (3/11, 27%) at onset of native GnRH treatment (P > 0.10). While concurrent untreated controls were not used in this study, the 79% response rate to twice daily administration of native GnRH is in agreement with other reports using pulsatile or constant infusion as methods of administration, confirming therapy can hasten follicular development and first ovulation of the breeding season. As with previous reports, follicle size at onset of treatment is an important determinant of interval from onset of native GnRH therapy to ovulation. Use of hCG or Ovuplant™ did not enhance ovulatory response in native GnRH treated mares. Use of Ovuplant™ during native GnRH therapy may increase the incidence of post-treatment anestrus in mares not becoming pregnant.     

Changes in the ovary and the peripheral concentrations of sex hormones after the aspiration of follicular fluid from the spontaneous follicular cysts of dairy cows

The aim of the present study was to clarify the ovarian and hormonal dynamics after the aspiration of follicular fluid in cows with follicular cysts. Follicular fluid was aspirated from the follicular cysts and follicles that were fated to become cystic follicles and other coexisting normal follicles, respectively, in lactating cows (n = 3). After the aspiration procedure, new follicles developed and reached a diameter of 25 mm without ovulation within 13–19 days. The plasma concentrations of inhibin decreased and follicle-stimulating hormone increased rapidly after the aspiration procedure, and subsequently increased and decreased, respectively, as a new follicle grew. No luteal structures developed after the aspiration procedure, and the animals’ plasma progesterone levels remained low. The present study indicates that the cystic follicles are never luteinized by the aspiration of follicular fluid, and consequently, new follicular cysts are observed to repeatedly develop.     

Dynamics and histological observation of first follicular wave in Japanese black cows

AIMS: To determine the turnover of the first follicular wave in Japanese black cows and quantitative immunohistological characteristics of the previously in vivo identified dominant follicle (DF) and largest subordinate follicle (SF) derived from ovariectomy on Day 7 (3 cows) and Day 10 (3 cows) (Day 0=estrus). Six cases of first follicular wave in cows were observed twice daily by ultrasound scanning. The number of follicles, diameter of DF and SF, and prevalence of apoptotic granulosa cells (GC) and theca cells (TC) were studied by TUNEL methods. At follicular wave emergence, 13.5 +/- 9.5 Class I (2-5 mm in diameter follicles) were found 12 hr after ovulation, and increased its number until Day 1 pm. Future DF and SF observed retrospectively were 4.9 +/- 0.8 mm and 4.9 +/- 0.9 mm at wave emergence. Deviation of DF and SF occurred on Day 3 pm with mean diameters of 8.9 +/- 1.3 mm and 6.8 +/- 0.9 mm, respectively. DF developed until Day 8 am with a maximum diameter of 14.4 +/- 1.8 mm (n=3) and then regressed. The follicular wall of the DF had a characteristic image of a healthy follicle on Day 7 and slightly atretic DF on Day 10, whereas SF showed heavy atresia on both Day 7 and Day 10 under HE staining. In the prevalence of apoptotic cells, DF were 4.4 +/- 1.0% and 17.9 +/- 4.9% on Day 7 and on Day 10 in GC, respectively, and 2.4 +/- 0.7% and 8.0 +/- 1.4% on Day 7 and on Day 10 in TC, respectively. These results showed that, 1) the first follicular wave in cows is characterized by 24 hr recruitment of small follicles and a gradual divergence of growth rates in future DF and SF, and 2) early regression of DF on Day 10 was preceded by severe apoptosis.     

Effect of an orally active progestin on follicular dynamics in cycling and anestrous postpartum beef cows

Although treatment of cycling cows with low concentrations of melengesterol acetate (MGA) results in formation of persistent follicles, in the absence of corpora lutea, it is not known whether persistent follicles form in anestrous cows in response to a similar treatment. The objective of this experiment was to determine the effect of long-term MGA treatment (14 d) on follicular dynamics and the secretion of estradiol in anestrous postpartum beef cows. Treatment groups (replicated over 2 yr) included the following: anestrous control (AC; n = 11), anestrous MGA (AM; n = 16), and cycling MGA (CM; positive control; n = 16). Angus-crossbred cows were assigned to treatment by age, cow body condition, and days postpartum. Cows were fed carrier (AC group) or 0.5 mg MGA x animal(-1) x d(-1) (AM and CM groups) for 14 d beginning approximately 38 d postpartum. Cows allotted to the CM group were injected with PGF2alpha, on the first day of MGA treatment to induce luteolysis. The preceding treatment (CM) results in formation of persistent follicles and secretion of elevated concentrations of estradiol. Ovaries of each cow were examined daily by transrectal ultrasonography beginning 5 to 7 d preceding the initiation of feeding MGA or carrier and continued until ovulation or 7 d following MGA feeding. There was no difference among groups in the stage of follicular wave or diameter of the largest follicle at the start of carrier or MGA feeding. The length of the follicular wave present at the start of MGA feeding was greater (P < 0.01) for cows in the CM (14.5 d, yr 1; 18.3 d, yr 2) group compared to the AM (9.4 d, yr 1; 7.9 d, yr 2) or AC (9.7 d, yr 1; 10.7 d, yr 2) groups. Maximum follicular diameter over both years was greater (P < 0.01) for the CM (20.6 mm) group than the AM (15.1 mm) or AC (16.4 mm) groups. Circulating concentrations of estradiol were also increased (P < 0.05) in the CM group compared to the AM or AC groups. However, MGA appeared to have no effect (P > 0.05) on the number of follicles recruited, growth rate of the dominant follicle during the first 6 d oftreatment, or growth rate to the maximum follicular diameter. In summary, MGA treatment did not increase the duration ot the follicular wave, maximum follicular diameter, or secretion of estradiol in anestrous postpartum cows, nor did MGA affect the number of follicles recruited or growth rate of dominant follicles in cycling or anestrous animals.     

Induction of ovulation with GnRH and PGF(2 alpha) at two different stages during the early postpartum period in dairy cows: ovarian response and changes in hormone concentrations

The aims of this study were 1) to determine whether dairy cows can be induced to ovulate by the treatment with gonadotropin releasing hormone (GnRH) followed by prostaglandin F(2 alpha) (PGF(2 alpha)) during the early postpartum period and 2) to describe their ovarian and hormonal responses according to ovarian status. Cows were divided in two groups and received 10 microg of buserelin followed by 500 microg of cloprostenol 7 days apart starting from 21 (GnRH21, n=7) or around 37 days postpartum (GnRH37, n=7). The groups were further classified according to presence (-CL) or absence (-NCL) of functional corpora lutea (CL) on the day of GnRH treatment (d 0): GnRH21-NCL (n=4), GnRH21-CL (n=3) and GnRH37-CL (n=7). Ovarian morphology was monitored and the concentrations of P(4), E(2), FSH and insulin-like growth factor 1 (IGF-1) were measured. All cows ovulated after administration of GnRH. The P(4) levels of the GnRH21-NCL group from d 0 to d 5 were lower than those of the GnRH21-CL (P<0.05) and GnRH37-CL groups (P<0.01). In contrast, the E(2) levels of the GnRH21-NCL group within d 2 to d 6 were higher (P<0.05) than those of the other groups. Compared with the GnRH37-CL group, the GnRH21-NCL group had more small follicles on d 2 (P<0.05), d 3 (P<0.01) and d 4 (P<0.01) and more large follicles on d 5 (P<0.05). The induced CL and new ovulatory follicles were larger in the GnRH21-NCL group compared with the GnRH21-CL (P<0.001 and P<0.01) and GnRH37-CL groups (P<0.001 and P<0.05). IGF-1 did not differ among the groups. The GnRH21-NCL group had higher FSH levels than the GnRH21-CL (P<0.01) and GnRH37-CL groups (P<0.001) on d 0. Low P(4) and high FSH levels may suggest higher gonadotropin support on the enhanced ovarian morphology of the GnRH21-NCL group. PGF(2 alpha) treatment induced CL regression and subsequent ovulation in 3/4 (75%), 3/3 (100%) and 7/7 (100%) cows in the GnRH21-NCL, GnRH21-CL and GnRH37-CL groups, respectively. In conclusion, a 7-day GnRH-PGF(2 alpha) synchronization protocol can effectively induce dairy cows to ovulate as early as 21 days postpartum, regardless of ovarian status.     

Effects of low-dose follicle-stimulating hormone administration on follicular dynamics and preovulatory follicle characteristics in dairy cows during the summer

The well-documented phenomenon of reduced conception rate in dairy cows during the hot season involves impaired functioning of the ovarian follicles and their enclosed oocytes. Three experiments were performed to examine the administration of low doses of follicle-stimulating hormone (FSH) to induce turnover of follicles that are damaged upon summer thermal stress and to examine whether this FSH administration has beneficial effects on preovulatory follicles. In experiment 1, synchronized heifers were treated with 100 mg of Folltropin-V (n = 7) or 4.4 mg of Ovagen (n = 6) on day 3 of the estrous cycle. Treatment with both FSH sources resulted in greater (P < 0.05) numbers of follicles than in control animals (n = 12) on day 6 of the estrous cycle, indicating that low doses of FSH can increase the number of emerging follicles in a follicular wave. In experiment 2, milking cows were assigned to a control group (n = 4) or treated with 2.2 mg (FSH-2.2; n = 6) or 4.4 mg (FSH-4.4; n = 5) Ovagen. Follicle-stimulating hormone was administrated on day 3 or 4 and day 10 or 11 of the estrous cycle, coinciding with emergence of the first and second follicular waves, respectively. The number of follicles emerging during the first wave tended to be higher (P < 0.1) in FSH-4.4-treated cows than in controls. The second-wave dominant follicles emerged 2 d later in the treated cows and were smaller in diameter (P < 0.05) than controls, 2 d before aspiration. Despite being younger, the preovulatory follicles of FSH-4.4 cows expressed a steroidogenic capacity that was similar to controls with a tendency toward greater insulin concentrations (P < 0.09). In experiment 3, milking cows were assigned to a control group (n = 6) or treated with 4.4 mg Ovagen (FSH-4.4; n = 6). Follicle-stimulating hormone was administrated on day 3 and day 12 or 13 of the estrous cycle. The number of emerging follicles was higher (P < 0.05) in the treated vs control cows. However, the features of the preovulatory follicle developed in the subsequent cycle did not differ between groups. In summary, low doses of FSH can efficiently induce follicular turnover accompanied by a modest effect on the preovulatory follicle of the treated cycle. It appears that the administration of low doses of FSH, precisely timed to synchronize with the emergence of follicular waves, might have a beneficial effect on the preovulatory follicle and its enclosed oocyte.     

Follicular, hormonal, and pregnancy responses of early postpartum suckled beef cows to GnRH, norgestomet, and prostaglandin F2alpha.

ABSTRACT: Cycling (n = 16) and noncycling (n = 24), early postpartum, suckled beef cows of three breeds were assigned randomly to three treatments: 1) 100-microg injection of GnRH plus a 6-mg implant of norgestomet administered on d -7 before 25 mg of PGF2alpha and implant removal on d 0 (GnRH+NORG); 2) 100 microg of GnRH given on d -7 followed by 25 mg of PGF2alpha on d 0 (GnRH); or 3) 2 mL of saline plus a 6-mg implant of norgestomet administered on d -7 followed by 25 mg of PGF2, and implant removal on d 0 (NORG). All cows were given 100 microg of GnRH on d +2 (48 h after PGF2alpha). Blood sera collected daily from d -7 to d +4 were analyzed for progesterone and estradiol-17beta, and ovaries were monitored daily by transrectal ultrasonography to assess changes in ovarian structures. Luteal structures were induced in 75% of noncycling cows in both treatments after GnRH, resulting in elevated (P < .01) progesterone on d 0 for GnRH+NORG-treated cows. Concentrations of estradiol-17beta (P < .01) and LH (P < .05) were greater on d +2 after GnRH for cows previously receiving norgestomet implants. Pregnancy rates after one fixed-time AI at 16 h after GnRH (d +2) were greater (P < .05) in GnRH+NORG (71%) than in GnRH (31%) and NORG (15%) cows. Difference in pregnancy rate was due partly to normal luteal activity after AI in over 87% of GnRH+NORG cows and no incidence of short luteal phases. The GnRH+NORG treatment initially induced ovulation or turnover of the largest follicle, induction of a new follicular wave, followed later by increased concentrations of estradiol-17beta and progesterone. After PGF2alpha, greater GnRH-induced release of LH occurred in GnRH+NORG cows before ovulation, and pregnancy rates were greater after a fixed-time AI.     

The effect of GnRH or oestradiol injected at pro-oestrus on luteal function and follicular dynamics of the subsequent oestrous cycle in non-lactating cycling Holstein cows

Oestrous synchronization involves synchronization of ovarian follicular turnover, new wave emergence, and finally induction of ovulation. The final step can be synchronized by the parenteral administration of either GnRH or oestradiol benzoate. This study investigated corpus luteum and follicular emergence after ovulation had been induced by the administration of either GnRH or oestradiol benzoate. The injection of oestradiol benzoate may have delayed the emergence of the first follicular wave subsequent to the induced ovulation; administration of oestradiol benzoate or GnRH lowered the progesterone rise so that the maximum dioestrous concentration of progesterone on Day 9 was lower when cows were treated during pro-oestrus compared to the spontaneously ovulating controls. One implication of findings from the present study is that induction of ovulation with either oestradiol benzoate or GnRH, administered 24 or 36 h after withdrawal of the CIDR device, respectively, may lower fertility. Future studies must identify the timing of administration relative to the time of CIDR device withdrawal and the optimum concentration of oestradiol benzoate or GnRH that would not have untoward effects on the development of the corpus lutea, particularly within the first week of dioestrus.     

Plasma concentrations of inhibin A in cattle with follicular cysts: relationships with turnover of follicular waves and plasma levels of gonadotropins and steroid hormones

We investigated the profiles of circulating levels of inhibin A and total inhibin in beef cows with follicular cysts in relation to the patterns of follicular development and circulating gonadotropins and steroid hormones. Turnover of follicular waves was monitored in five cows every 2 days for 70 days from 10 days after detection of estrus without ovulation. The mean interwave intervals were 19.6 ± 1.0 days (n = 18 waves with cysts from the five cows). Circulating levels of inhibin A were approximately 170 pg/ml before emergence of follicular waves with cysts and increased (P < 0.05) concomitantly with follicle emergence. High concentrations of inhibin A (greater than 300 pg/ml) were noted for 7 days during the growth phase of cystic follicles, but inhibin A levels decreased gradually when development of the cysts reached a plateau. This profile of inhibin A was similar to those of total inhibin and estradiol, but was inversely related to the changes in plasma FSH concentrations. LH pulse frequency and mean concentrations of LH in cows with cysts were higher than those observed in the luteal phase of normal cyclic cows. These results indicate that the capacity to secrete inhibin, as well as estradiol, is maintained in cystic follicles, the growth of which is extended by LH secretion at levels greater than those seen in the normal luteal phase. Inhibin A plays an important role in the extension of interwave intervals by suppressing recruitment of a new cohort of follicles.