Efficacy of Deslorelin Acetate (SucroMate) on Induction of Ovulation in American Quarter Horse Mares

Equine clinicians rely on ovulation induction agents to provide a timed ovulation in mares for optimal breeding management. Numerous studies have been performed on the efficacy of human chorionic gonadotropin (hCG) to induce ovulation in the mare, but limited clinical data are available for the new deslorelin acetate product SucroMate. This study was designed to evaluate the efficacy of SucroMate (deslorelin) in comparison with hCG to induce ovulation. American Quarter horse mares (n = 256) presented to Colorado State University for breeding management were used in this study. Mares received either deslorelin or hCG when a follicle ≥35 mm was detected by transrectal ultrasound in the presence of uterine edema. Ultrasonographic examinations were subsequently performed once daily until ovulation was detected. Deslorelin was administered to 138 mares during168 estrous cycles, and hCG was given to 118 mares during 136 estrous cycles. Mares administered deslorelin had a similar (P < .05) higher ovulation rate (89.9%) within 48 hours following drug administration than mares administered hCG (82.8%). There are no effects of season or age on ovulation rates in either treatment group. Twenty-one mares administered deslorelin and 11 mares administered hCG were monitored by transrectal ultrasound every 6 hours to detect ovulation as part of a frozen semen management program. Average intervals from deslorelin or hCG administration to ovulation were 41.4 ± 9.4 and 44.4 ± 16.5 hours, respectively. Results of this study indicate that SucroMate is effective at inducing a timed ovulation in the mare.     

Comparison of Compounded Deslorelin and hCG for Induction of Ovulation in Mares

Ovulation-inducing agents are routinely used in broodmare practice. The objective of this study was to compare the efficacy of two compounded deslorelin products and human chorionic gonadotropin (hCG) in inducing ovulation in a clinical reproduction program. Breeding records of 203 mares administered an ovulation-inducing agent during the 2006 breeding season were reviewed. Estrous cycles were included for comparison if agents were administered when the largest follicle was 35 to 45 mm in diameter and endometrial edema was present. There was no significant difference (P > .05) in interval to ovulation for mares receiving deslorelin (1.9 ± 0.7 days) or hCG (2.0 ± 0.7 days). The percentage of mares that ovulated within 48 hours after treatment was also not significantly different between the agents (90.1% and 88.3%, respectively). In summary, clinical efficacy at inducing a timed ovulation in estrual mares with follicles 35 to 45 mm was similar between compounded deslorelin and hCG.     

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.     

Clinical experience with deslorelin (ovuplantTM) in a kentucky thoroughbred broodmare practice (1999)

Palpation records of 155 Throughbred broodmares maintained on one of seven farms (3–80 mares per farm) that were administered deslorelin on one or more estrous cycles (204 treated cycles) during the 1999 breeding season were retrospectively examined. Some deslorelin-treated mares were also treated with hCG (2500 units intravenously), or had no ovulation-inducing drugs administered, during different estrous cycles of the same season. Most mares were treated with an ovulation- inducing drug after returning to their resident farm following breeding and were subsequently examined by transrectal ultrasonography daily until ovulation was confirmed, and again 13–14 and 15–16 days after ovulation for determination of pregnancy status.Per-cycle pregnancy rate for all 155 mares bred was 53%, and for all deslorelin breeding was 57%. Per-cycle pregnancy rates for mares ovulating 0–1 days, 1–2 days, and 2–3 days after treatment with deslorelin did not differ (P>0.05). Forty-six mares received more than one treatment during the breeding season, yielding 115 breedings (estrous cycles) for comparison of pregnancy rates among treatment. Per-cycle pregnancy rates for these mares did not differ among treatments (P>0.10).No differences due to treatment were detected in mean interval to ovulation (P>0.10). Mean interovulatory interval was longer for deslorelin-treated mares than for untreated or hCG treated mares (P>0.01). Eighty percent (80%) of deslorelin-treated mares had interovulatory intervals of 18–25 days, and 19% had interovulatory intervals>25 days. Ninety-seven percent (97%) of untreated or hCG-treated mares had interovulatory interovulatory intervals>25 days. More deslorelin-treated mares had extended (>25 days) interovulatory intervals than hCG- or nontreated-mares (P>0.05). In this group of Thoroughbred mares, it appeared that season (month) and management (farm) factors had only minor effects on the incidence of extended interovulatory intervals following use of deslorelin.     

Strategies for Increasing Reproductive Efficiency in a Commercial Embryo Transfer Program With High Performance Donor Mares Under Training

Exercise stress has a negative impact on embryo transfer efficiency (ET). For example, a 34% embryo recovery rate, 43% incidence of poor quality embryos, and a 29% pregnancy rate after transfer have been reported. Administration of nonsteroidal anti-inflammatory drugs (NSAIDs) may reduce the inflammatory response produced after nonsurgical embryo transfer. In addition, progesterone supplementation is commonly administered to some recipient mares to improve uterine conditions before the transfer and to ensure adequate progestational support compatible with pregnancy. The aim of the study was to evaluate embryo recovery rates using BioRelease deslorelin versus hCG and to increase posttransfer pregnancy rates by jointly administering BioRelease progesterone and a NSAID (flunixin or meloxicam) to recipient mares. Seventeen upper-level showjumping mares stabled and in daily training were used as embryo donors. To induce ovulation, 1-mg IM BioRelease deslorelin (BioRelease Technologies, Lexington, KY) was injected in treated cycles (n = 66), or 2500-IU hCG IV (Ovusyn, Syntex, Buenos Aires, Argentina) was given in control cycles (n = 79) when a ≥35 mm follicle was present. Artificial insemination with extended fresh semen (at least 500 × 10⁶ progressively motile sperm) was carried out in both groups immediately after injecting the ovulation induction agent. Day 8 embryos were recovered and nonsurgically transferred using a speculum and a cervical traction forceps. Recipient mares (n = 73) were randomly assigned to one of three groups: Group A received a single injection of 1.5-g IM BioRelease progesterone (Progesterone LA 300, BioRelease Technologies) and 3 IV injections of 0.5 g of flunixin meglumine (Flunix Deltavet, Argentina), one injection administered the day of the transfer and one on each of the next two successive days. Group B received 1.5-g IM BioRelease progesterone and a single dose of 1.5-g IM BioRelease meloxicam (Meloxicam LA, BioRelease Technologies) at the moment of embryo transfer. Group C did not receive any treatment. Pregnancy diagnosis was carried out 7 days after transfer. Results were analyzed using comparisons of proportions. More embryos were recovered per cycle (13% increase) when donor mares in training were induced to ovulate with BioRelease deslorelin (60.6%; 40/66) than with hCG (46.8%; 37 of 79; P < .05). Although both recipient groups given NSAIDs in combination with BioRelease progesterone numerically had higher pregnancy rates (A: 70.8%; 17/24 and B: 75%; 15/20) compared with nontreated control recipients (47.1%; 33/70), pregnancy rates were significantly higher only in recipients given LA meloxicam treatment at the time of transfer (P < .05). The LA meloxicam is released over a 72-hour period making it more practical to use as it requires a single IM injection versus the 3 IV flunixin meglumine injections. Thus, to minimize the effects of exercise stress on ET efficiency, a combination of BioRelease deslorelin to induce ovulation in donors and BioRelease progesterone and LA meloxicam in recipients at the time of transfer may offer an interesting alternative for improving results in commercial ET programs.     

Does Clinical Treatment with Phenylbutazone and Meloxicam in the Pre‐ovulatory Period Influence the Ovulation Rate in Mares?

The presence of anovulatory haemorrhagic follicles during the oestrous cycle of mares causes financial impacts, slowing conception and increasing the number of services per pregnancy. Non‐steroidal anti‐inflammatory drugs (NSAIDs) such as meloxicam and phenylbutazone are used in the treatment of several disorders in mares, and these drugs can impair the formation of prostaglandins (PGs) and consequently interfere with reproductive activity. This study aimed to evaluate the effects of treatment with NSAIDs on the development of pre‐ovulatory follicles in mares. In total, 11 mares were studied over three consecutive oestrous cycles, and gynaecological and ultrasound examinations were performed every 12 h. When 32‐mm‐diameter follicles were detected, 1 mg of deslorelin was administered to induce ovulation. The first cycle was used as a control, and the mares received only a dose of deslorelin. In the subsequent cycles, in addition to receiving the same dose of deslorelin, each mare was treated with NSAIDs. In the second cycle, 4.4 mg/kg of phenylbutazone was administered, and in the third cycle, 0.6 mg/kg of meloxicam was administered once a day until ovulation or the beginning of follicular haemorrhage. All of the mares ovulated between 36 and 48 h after the induction in the control cycle. In the meloxicam cycle, 10 mares (92%) did not ovulate, while in the phenylbutazone cycle, nine mares (83%) did not ovulate. In both treatments, intrafollicular hyperechoic spots indicative of haemorrhagic follicles were observed on ultrasound. Thus, our results suggested that treatment with meloxicam and phenylbutazone at therapeutic doses induced intrafollicular haemorrhage and luteinization of anovulatory follicles.     

Superovulation in cycling mares using equine follicle stimulating hormone (eFSH)

Superovulation would potentially increase the efficiency and decrease the cost of embryo transfer by increasing embryo collection rates. Other potential clinical applications include improving pregnancy rates from frozen semen, treatment of subfertility in stallions and mares, and induction of ovulation in transitional mares. The objective of this study was to evaluate the efficacy of purified equine follicle stimulating hormone (eFSH; Bioniche Animal Health USA, Inc., Athens, GA) in inducing superovulation in cycling mares. In the first experiment, 49 normal, cycling mares were used in a study at Colorado State University. Mares were assigned to 1 of 3 groups: group 1, controls (n = 29) and groups 2 and 3, eFSH-treated (n = 10/group). Treated mares were administered 25 mg of eFSH twice daily beginning 5 or 6 days after ovulation (group 2). Mares received 250 (of cloprostenol on the second day of eFSH treatment. Administration of eFSH continued until the majority of follicles reached a diameter of 35 mm, at which time a deslorelin implant was administered. Group 3 mares (n = 10) received 12 mg of eFSH twice daily starting on day 5 or 6. The treatment regimen was identical to that of group 2. Mares in all 3 groups were bred with semen from 1 of 4 stallions. Pregnancy status was determined at 14 to 16 days after ovulation.In experiment 2, 16 light-horse mares were used during the physiologic breeding season in Brazil. On the first cycle, mares served as controls, and on the second cycle, mares were administered 12 mg of eFSH twice daily until a majority of follicles were 35 mm in diameter, at which time human chorionic gonadotropin (hCG) was administered. Mares were inseminated on both cycles, and embryo collection attempts were performed 7 or 8 days after ovulation.Mares treated with 25 mg of eFSH developed a greater number of follicles (35 mm) and ovulated a greater number of follicles than control mares. However, the number of pregnancies obtained per mare was not different between control mares and those receiving 25 mg of eFSH twice daily. Mares treated with 12 mg of eFSH and administered either hCG or deslorelin also developed more follicles than untreated controls. Mares receiving eFSH followed by hCG ovulated a greater number of follicles than control mares, whereas the number of ovulations from mares receiving eFSH followed by deslorelin was similar to that of control mares. Pregnancy rate for mares induced to ovulate with hCG was higher than that of control mares, whereas the pregnancy rate for eFSH-treated mares induced to ovulate with deslorelin did not differ from that of the controls. Overall, 80% of mares administered eFSH had multiple ovulations compared with 10.3% of the control mares.In experiment 2, the number of large follicles was greater in the eFSH-treated cycle than the previous untreated cycle. In addition, the number of ovulations during the cycle in which mares were treated with eFSH was greater (3.6) than for the control cycle (1.0). The average number of embryos recovered per mare for the eFSH cycle (1.9 ± 0.3) was greater than the embryo recovery rate for the control cycle (0.5 ± 0.3).In summary, the highest ovulation and the highest pregnancy and embryo recovery rates were obtained after administration of 12 mg of eFSH twice daily followed by 2500 IU of hCG. Superovulation with eFSH increased pregnancy rate and embryo recovery rate and, thus, the efficiency of the embryo transfer program.

Introduction

Induction of multiple ovulations or superovulation has been an elusive goal in the mare. Superovulation would potentially increase the efficiency and decrease the cost of embryo transfer by increasing embryo collection rates.[1 and 2] Superovulation also has been suggested as a critical requirement for other types of assisted reproductive technology in the horse, including oocyte transfer and gamete intrafallopian transfer. [2 and 3] Unfortunately, techniques used successfully to superovulate ruminants, such as administration of porcine follicle stimulating hormone and equine chorionic gonadotropin have little effect in the mare. [4 and 5]The most consistent therapy used to induce multiple ovulations in mares has been administration of purified equine pituitary gonadotropins. Equine pituitary extract (EPE) is a purified gonadotropin preparation containing approximately 6% to 10% LH and 2% to 4% FSH.[6] EPE has been used for many years to induce multiple ovulations in mares [7, 8 and 9] and increase the embryo recovery rate from embryo transfer donor mares. [10] Recently, a highly purified equine FSH product has become available commercially.The objectives of this study were to evaluate the efficacy of purified eFSH in inducing superovulation in cycling mares and to determine the relationship between ovulation rate and pregnancy rate or embryo collection rate in superovulated mares.

Materials and methods

Experiment 1

Forty-nine normally cycling mares, ranging in age from 3 to 12 years, were used in a study at Colorado State University. Group 1 (control) mares (n = 29) were examined daily when in estrus by transrectal ultrasonography. Mares were administered an implant containing 2.1 mg deslorelin (Ovuplant, Ft. Dodge Animal Health, Ft. Dodge, IA) subcutaneously in the vulva when a follicle 35 mm in diameter was detected. Mares were bred with frozen semen (800 million spermatozoa; minimum of 30% progressive motility) from 1 of 4 stallions 33 and 48 hours after deslorelin administration. The deslorelin implants were removed after detection of ovulation.[11] Pregnancy status was determined at 14 and 16 days after ovulation.Group 2 mares (n = 10) were administered 25 mg of eFSH (Bioniche Animal Health USA, Inc., Athens, GA) intramuscularly twice daily beginning 5 or 6 days after ovulation was detected. Mares received 250 g cloprostenol (Estrumate, Schering-Plough Animal Health, Omaha, NE) intramuscularly on the second day of eFSH treatment. Administration of eFSH continued until a majority of follicles reached a diameter of 35 mm, at which time a deslorelin implant was administered. Mares were subsequently bred with the same frozen semen used for control mares, and pregnancy examinations were performed as described above.Group 3 mares (n = 10) received 12 mg of eFSH twice daily starting 5 or 6 days after ovulation and were administered 250 μg cloprostenol on the second day of treatment. Mares were randomly selected to receive either a deslorelin implant (n = 5) or 2500 IU of human chorionic gonadotropin (hCG) intravenously (n = 5) to induce ovulation when a majority of follicles reached a diameter of 35 mm. Mares were bred with frozen semen and examined for pregnancy as described above.

Experiment 2

Sixteen cycling light-horse mares were used during the physiologic breeding season in Brazil. Reproductive activity was monitored by transrectal palpation and ultrasonography every 3 days during diestrus and daily during estrus. On the first cycle, mares were administered 2500 IU hCG intravenously once a follicle 35 mm was detected. Mares were subsequently inseminated with pooled fresh semen from 2 stallions (1 billion motile sperm) daily until ovulation was detected. An embryo collection procedure was performed 7 days after ovulation. Mares were subsequently administered cloprostenol, and eFSH treatment was initiated. Mares received 12 mg eFSH twice daily until a majority of follicles were 35 mm in diameter, at which time hCG was administered. Mares were inseminated and embryo collection attempts were performed as described previously.

Statistical analysis

In experiment 1, 1-way analysis of variance with F protected LSD was used to analyze quantitative data. Pregnancies per ovulation were analyzed by x² analysis. In experiment 2, number of large follicles, ovulation rate, and embryo recovery rate were compared by Student,'s t-test. Data are presented as the mean S.E.M. Differences were considered to be statistically significant at p < .05, unless otherwise indicated.

Results

In experiment 1, mares treated with 25 mg eFSH twice daily developed a greater number of follicles 35 mm in diameter (p = .001) and ovulated a greater number of follicles (p = .003) than control mares (Table 1). However, the number of pregnancies obtained per mare was not significantly different between the control group and the group receiving 25 mg eFSH (p = .9518). Mares treated with 12 mg eFSH and administered either hCG or deslorelin to induce ovulation also developed more follicles 35 mm (p = .0016 and .0003, respectively) than untreated controls. Mares receiving eFSH followed by hCG ovulated a greater number of follicles (p = .003) than control mares, whereas the number of ovulations for mares receiving eFSH followed by deslorelin was similar to that of control mares (p = .3463). Pregnancy rate for mares induced to ovulate with hCG was higher (p = .0119) than that of control mares, whereas the pregnancy rate for eFSH-treated mares induced to ovulate with deslorelin did not differ from that of controls (p = .692). Pregnancy rate per ovulation was not significantly different between control mares (54.5%) and mares treated with eFSH followed by hCG (52.9%). The lowest pregnancy rate per ovulation was for mares stimulated with 25 mg eFSH and induced to ovulate with deslorelin. The mean number of days mares were treated with 25 mg or 12 mg of eFSH was 7.8 ± 0.4 and 7.5 ± 0.5 days, respectively. Overall, 80.0% of mares administered eFSH had multiple ovulations compared with 10.3% of control mares.     

Treatment of transition phase mares with progesterone intravaginally and with deslorelin or hCG to assist ovulations

Over four years, four investigators in the Northern Hemisphere treated 413 privately owned transition phase mares between late February and early April, for the purpose of breeding such mares early in the season. Mares received an intravaginal device (CIDR-B) carrying 1.9 g progesterone, for about 12 days. Thereafter mares forming preovulatory follicles >30 mm were either treated with a short acting implant releasing the GnRH analog deslorelin (Ovuplant™) or with 1,500—2,500 IU hCG, or not. Follicle sizes were determined with ultrasonography at admission to the study (i.e. day of CIDR-B insertion), at intervals during treatment, at device removal and in 24 (to 48) hour intervals thereafter to determine the time for treatment to induce and accelerate ovulation and to ovulation, respectively. Pregnancies were determined by ultra-sonography between Days 14 to 18 after breeding, mostly 12 to 14 days after ovulation. Based on the size of the largest follicle at admission, mares were grouped into Classes with a ollicle diameter of 10 mm or less in Class I, and mares with follicles 11-20 mm, 21-30 mm and >30 mm in Classes II, III and IV, respectively. Overall, 80.2% of all mares responded to treatment with estrus and 80.7% ovulated. For mares in Classes I to IV, the rate of mares bred and becoming pregnant was 53.4% and 66.7%, 65.6% and 58.7%, 87.5% and 52.3%, and 75.0% and 52.0%, respectively. The overall pregnancy rate was 55.6% for the first breeding in response to treatment. Mares not assisted with Ovuplant or hCG were bred at a significantly lower rate (<0.0001) and the pregnancy rate was lower, 44.4% vs. 54.2% and 60.5%, respectively. Treatments with Ovuplant or hCG ensured ovulation rates of 96.0 and 84.9% versus 53.3% in unassisted mares overall. Follicle diameters increased significantly with CIDR-B in situ, and progressed after device removal to >30 mm within 4.0 days and to ovulation 5.3 days. Those mares in Class I responding to treatment (ca 60%) did not differ from Class II to IV mares in almost all the parameter evaluated. Significant differences were seen in the UK in response to treatment between years for the percentage of mares showing heat, ovulated, were bred and became pregnant.     

The Effect of Sulpiride Treatment During the Periovulatory Period on Prolactin Concentration and Ovulation in the Mare

Sixteen estrous cycles from 10 cyclic mares were randomly assigned to a control or sulpiride group (n = 8 each). All mares received 1,500 IU of human chorionic gonadotropin (hCG) (hour 0) during estrus with a follicular diameter ≥32 mm. Mares were scanned every 12 hours until ovulation. In the treatment group, beginning at hour 0, each mare received 1.5 mg/kg of sulpiride every 12 hours intra-muscularly until ovulation or formation of a luteinized unruptured follicle (LUF). Concentrations of luteinizing hormone (LH) and prolactin (PRL) were measured by radioimmunoassay. In each group, there were 10 preovulatory follicles for the eight cycles. The ovulation rate (9/10, 90%) was similar in the control and sulpiride groups. Two mares formed an LUF, which was first detected at hours 48 and 72 for the sulpiride and control mares, respectively. The interval from hCG to ovulation was 49.5 ± 11.1 and 43.5 ± 5.8 hours, for the control and sulpiride groups, respectively (P > .5). LH followed the typical preovulatory surge pattern, with no difference between groups (P > .5). Sulpiride administration increased PRL concentration in treated mares at 24 (P < .1), 36, and 48 hours (P < .05) after treatment. In conclusion, sulpiride administration every 12 hours increased PRL concentration in treated mares after 24 hours of the beginning of treatment. However, at this time window and concentration, PRL did not have any effect on ovulation. The control mare that developed an LUF had a PRL concentration similar to other ovulatory control mares (always ≤10 ng/mL).     

Comparison of Efficacy of Two Dose Rates of Histrelin to Human Chorionic Gonadotropin for Inducing Ovulation in Broodmares

Between February 15 and May 17, 2011, a total of 88 broodmares (10 maiden, 10 barren, and 68 foaling) maintained on pasture in southeast Texas were examined three times weekly (Tuesday, Thursday, Saturday) by transrectal palpation and ultrasonography. On Tuesday or Thursday, mares in estrus with uterine edema, a relaxed cervix, and a dominant follicle ≥34 mm in diameter were alternately assigned to treatment with the following: group (1) 2,500-unit human chorionic gonadotropin (hCG), intravenous; group (2) 1.0-mg BioRelease Histrelin (Biorelease Technologies, Lexington, KY), intramuscular; or group (3) 0.5-mg BioRelease Histrelin, intramuscular. Ovulation was confirmed by ultrasonographic examination. The percentage of mares ovulating within 2 days appeared to be similar between maiden, barren, and foaling mares, so responses for all mares were totaled for analysis. A nonsignificant trend for higher ovulation rates within 2 days was noted for both dose rates of histrelin compared with hCG treatment (31/37, 84%; 34/37, 92%; and 33/36, 92% for groups 1-3, respectively) (P = .45). Ovulatory responses appeared to improve for both products as the season progressed, yet no differences were detected between response rates to histrelin or hCG for any month (P ≥ .50). The use of 1.0- or 0.5-mg BioRelease Histrelin was found to be at least equally effective as hCG treatment for inducing ovulation within 2 days of treatment throughout the breeding season.     

Efficacy of Long-Acting Formulations of Estradiol or Progesterone Plus Estradiol on Estrous Synchronization in Broodmares

A preliminary trial was performed to evaluate the ability of sustained release preparations of estradiol-17β or progesterone plus estradiol-17β to synchronize estrus in cyclic mares. Group 1 mares were treated with a 50 mg intramuscular (IM) injection of sustained release estradiol-17β, while group 2 mares were treated with estradiol plus 1.5 g of sustained release progesterone. All mares received an IM injection of 10 mg of prostaglandin-F₂α (PGF₂α) 10 days after steroid treatment. Mares were examined by transrectal ultrasonography on Days 1 and 10 of treatment and then at ≤2 day intervals to monitor follicle size. Once a follicle ≥30 mm diameter and uterine edema were detected, 0.5 mg of the GnRH analog histrelin was administered IM. Mares were examined daily thereafter to detect ovulation. Group 1 mares did not exhibit ovulation synchrony (ovulations occurred 12-22 days after steroid treatment), whereas ovulation synchrony was satisfactory in group 2 mares (interval to ovulation being 20.4 ± 1.5 days, range 17-22 days). Using sustained release preparations of progesterone plus estradiol-17β, with PGF₂α administered on Day 10, could eliminate the need for daily injections of steroid preparations in oil when synchronizing estrus and ovulation.     

Pregnancy and twinning rates in Thoroughbred mares following the administration of human chorionic gonadotropin (hCG)

AIM: To determine the effect of hCG administration to cycling Thoroughbred mares, on pregnancy and twinning rates and the number of serves in the treated cycle. METHODS: A retrospective case control approach was conducted involving 2119 mare ovulatory cycles, on 1110 mares over a 7-year period. Data were collected by 1 of the authors during routine stud work at 3 commercial Thoroughbred farms in the Waikato region of New Zealand. The hCG (1500 IU) was administered by intravenous injection to selected mares 24 h before the expected time of breeding. Mares were scanned for pregnancy (singleton or twins) 14 days after the onset of dioestrus or detection of ovulation. Multilevel logistic regression analyses were used to identify the risk factors associated with the outcomes of interest while simultaneously controlling for possible confounding factors. RESULTS: Treatment with hCG tended to improve the odds of pregnancy (p=0.06), produced a 3-fold increase in the odds of twins (p<0.001), and increased the odds of a mare having a single serve in the treated ovulatory cycle (p=0.036). The first ovulatory cycle of a season in which a mare was bred was associated with a lower odds of pregnancy (p=0.02), and a lower odds of twins (p=0.003), when compared with subsequent cycles. Lactating mares were less likely to be diagnosed with twins (p=0.005), and were more likely to have a single serve (p<0.001), in any one ovulatory cycle than non-lactating mares. CONCLUSIONS: This report supports the role of hCG as an important therapeutic tool in veterinary management of broodmares for optimal reproductive performance. Mares treated with hCG must be managed in the knowledge that they have an increased likelihood of twins.     

Reproductive performance of commercial broodmares after induction of ovulation with HCG or Ovuplant™ (deslorelin)

Soon after Ovuplant™, the sustained-release implant containing the gonadotropin releasing hormone (GnRH) agonist deslorelin, was approved for commercial use in the United States for induction of ovulation in mares, anecdotal field observations were reported that some Ovuplant™—treated mares that did not become pregnant experienced a delayed return to estrus and prolonged inter-ovulatory interval. Although those observations have been subsequently confirmed, further data on how mares respond to Ovuplant™ compared to human chorionic gonadotropin (hCG) during the post-treatment period is needed. The objective of this study was to further evaluate the clinical use of Ovuplant™ by comparing the reproductive performance of commercial broodmares treated with hCG or Ovuplant™. This retrospective study was completed by examining the 1999 reproductive records of 106 mares treated with hCG during 134 estrous cycles and 117 mares treated with Ovuplant™ during 151 estrous cycles. There were no differences (P > 0.10) in follicle size at the time of treatment (39.4 ± 0.5 vs. 38.9 ± 0.5 mm), interval from treatment to ovulation (2.2 ± 0.1 vs. 2.2 ± 0.1 days), proportion of mares that failed to ovulate after treatment (3.0 vs. 4.6 %), or per-cycle pregnancy rate (47.7 vs. 51.4 %) between hCG-and Ovuplant™-treated mares, respectively. The interval from ovulation to return to estrus (25.8 ± 1.3 vs. 15.5 ± 0.6 days) and the inter-ovulatory interval (30.4 ± 1.5 vs. 20.8 ± 0.6 days) were longer (P<0.001) for Ovuplant™-compared to hCG-treated mares, and the proportion of non-pregnant mares that failed to return to estrus within 30 days after ovulation (31.4 vs. 1.5 %) was higher (P<0.001) for Ovuplant™-compared to hCG-treated mares, respectively. For Ovuplant™—treated mares, follicle size at the time of treatment tended (P<0.1) to be smaller for mares that failed to return to estrus within 30 days compared to mares that returned to estrus within 30 days (37.1 ± 1.1 vs. 40.1 ± 0.6 mm, respectively). Also, the average date of ovulation during the calendar year was later (P < 0.05) for Ovuplant™—treated mares that failed to return to estrus within 30 days compared to those that returned to estrus within 30 days (May 15 ± 4 vs. April 30 ± 4 days). The results of this study confirm previous reports that although the ovulatory response and fertility were not different for hCG- and Ovuplant™—treated mares, mares treated with Ovuplant™ that did not become pregnant had a significantly delayed return to estrus and prolonged inter-ovulatory interval. Based on recently published information, it appears this effect is due to Ovuplant™—induced down-regulation of the pituitary gland, which suppresses subsequent follicular growth and development. This study also demonstrated that follicle size and/or season may influence the probability that Ovuplant™—treated mares would experience a delayed return to estrus/ovulation; therefore, further work is needed to determine whether these or other factors are related to this specific outcome following Ovuplant™—treatment.     

The post-ovulatory rise in progesterone is lower and the persistence of oestrous behaviour longer during the first compared with the second cycle of the breeding season in mares

Mares are seasonally polyoestrous breeders. Therefore, the first ovulation of the season, following winter anoestrus, is the only cycle in which mares ovulate without the presence of an old CL from the previous cycle. The objective of this study was to compare the length of oestrous behaviour, and plasma progesterone concentrations during the early post-ovulatory period between mares after the first and second ovulation of the breeding season. Overall, 38 mares and 167 oestrous periods were used in the study. From those, 11 mares were used during the first and subsequent oestrous period to measure and compare the post-ovulatory rise in progesterone concentration, whereas all the mares were used to compare the length of the post-ovulatory oestrous behaviour between the first and subsequent cycles of the breeding season. The persistence of the post-ovulatory oestrus was longer (p < .001) following the first ovulation of the year (median of 52 h) compared with the subsequent ovulations (median of 36 h for second and later ovulations groups; n = 38 mares). The progesterone concentration at any of the four 8 h-intervals analysed (28, 36, 76 and 84 h post-ovulation) was lower (p < .01) following the first versus the second ovulation of the year. By 36 h post-ovulation the progesterone concentration of mares at the second ovulation of the year had passed the threshold of 2 ng/ml (2.1 ± 0.33 ng/ml), whereas in the first cycle it was 1.2 ± 0.13 ng/ml. In conclusion, mares had lower progesterone concentrations in their peripheral circulation and longer persistence of oestrous behaviour following the first ovulation of the year compared with the second and subsequent ovulatory periods of the breeding season.     

Comparison of HCG, buserelin and luprostiol for induction of ovulation in cycling mares

Sixty nonlactating light-horse mares were used to compare the efficacy of hCG, buserelin (a GnRH analog) and luprostiol (a PGF₂α analog) for induction of ovulation in cycling mares. Mares were assigned to 1 of 4 treatments: 1) controls; 2) 40 μg buserelin IM at 12 hr intervals during estrus until ovulation; 3) 7.5 mg IM luprostiol; and 4) 3,300 IU hCG. Treatments were given once a mare obtained a ≥35mm follicle and had been in estrus ≥2 days. Both buserelin and hCG shortened (p<0.05) the interval from treatment to ovulation compared to controls; whereas, luprostiol failed to hasten ovulation. Number of follicles ovulated was similar among all 4 groups. Although buserelin and hCG were equal in their ability to induce ovulation, an average of 3.8 injections of buserelin was required for hastening of ovulation.     

Comparison of Different Treatments for Oestrous Induction in Seasonally Anovulatory Mares

The aim of this study was to evaluate the effects of different treatments for induction and synchronization of oestrus and ovulation in seasonally anovulatory mares. Fifteen mares formed the control group (C), while 26 mares were randomly assigned to three treatment groups. Group T1 (n = 11) were treated with oral altrenogest (0.044 mg/kg; Regumate^®) during 11 days. Group T2 (n = 7) was intravaginally treated with 1.38 g of progesterone (CIDR^®) for 11 days. In group T3 (n = 8), mares were also treated with CIDR^®, but only for 8 days. All mares received PGF2α 1 day after finishing the treatment. Sonographic evaluation of follicles, pre‐ovulatory follicle size and ovulation time was recorded. Progesterone and leptin levels were analysed. Results show that pre‐ovulatory follicles were developed after the treatment in 88.5% of mares. However, the pre‐ovulatory follicle growth was dispersal, and sometimes it was detected when treatment was not finished. While in mares treated with intravaginal device, the follicle was soon detected (1.5 ± 1.2 days and 2.3 ± 2.0 days in T2 and T3 groups, respectively), in T1 group, the pre‐ovulatory follicle was detected slightly later (3.9 ± 1.6 days). The interval from the end of treatment to ovulation did not show significant differences between groups (T1 = 13.1 ± 2.5 days; T2 = 11.0 ± 3.6 days; T3 = 13.8 ± 4.3 days). The pregnancy rate was 47.4%, similar to the rate observed in group C (46.7%; p > 0.05). Initial leptin concentrations were significantly higher in mares, which restart their ovarian activity after treatments, suggesting a role in the reproduction mechanisms in mares. It could be concluded that the used treatments may be effective for oestrous induction in mares during the late phase of the seasonally anovulatory period. Furthermore, they cannot synchronize oestrus, and then, it is necessary to know the reproductive status of mares when these treatments are used for oestrous synchronization.     

Ovulation,Pregnancy Rate and Early Embryonic Development in Vernal Transitional Mares Treated with Equine‐ or Porcine‐FSH

The objective of this study was to compare the efficacy of purified equine‐ and porcine‐FSH treatment regimes in mares in early vernal transition. Mares (n = 22) kept under ambient light were examined ultrasonographically per‐rectum, starting January 30th. They were assigned to one of two treatment groups using a sequential alternating treatment design when a follicle ≥ 25 mm was detected. In the eFSH group, mares were treated twice daily with equine‐FSH, and in the pFSH group mares were treated twice daily with porcine‐FSH; treatments were continued until follicle(s) ≥ 35 mm, and 24 h later hCG was administered. Oestrous mares were inseminated with fresh semen and examined for pregnancy on days 11–20 post‐ovulation. In the eFSH group, 11/11 (100%) mares developed follicle(s) ≥ 35 mm, 8/11 (73%) ovulated and 6/8 (75%) conceived. In the pFSH group, 5/11 (45%) developed follicle(s) ≥ 35 mm, 4/11 (36%) ovulated and 3/4 (75%) conceived. Treatment with eFSH resulted in a greater ovarian stimulation; higher number of pre‐ovulatory‐sized follicles, higher number of ovulations and higher number of embryos (p < 0.05). Following ovulation, serum progesterone concentrations were correlated with the number of CLs and supported early embryonic development; maternal recognition of pregnancy occurred in all pregnant mares. We concluded that eFSH can be used to effectively induce follicular growth and ovulation in vernal transitional mares; however, if bred, diagnosis and management of twins’ pregnancies would be required prior to day 16 because of the increased risk of multiple embryos per pregnancy. Conversely, the current pFSH treatment regime cannot be recommended.     

Effects of repeated hCG injections on reproductive efficiency in mares

Thirty reproductively sound mares were divided into treatment and control groups. In the treatment group, consisting of 14 mares, 2500 I.U. of human chorionic gonadotropin (hCG) was administered intravenously during estrus, in the presence of a 35 mm follicle over five successive cycles in 1987, and at least two cycles in 1988. Beginning with the second cycle of treatment in 1988, these mares were bred to a fertile stallion. The control group, consisting of 16 mares, was followed for two to five cycles in either the 1987 or 1988 season and six of these mares were bred to fertile stallions. Throughout the study period, blood was collected from the mares in the treatment group for analysis of anti-hCG antibodies and cross reactivity of the antibody to purified equine lutenizing hormone (eLH) and equine chorionic gonadotropin (eCG).In 1987, after the first three injections of hCG, mean duration of estrus in treated mares tended to be shorter than in control mares (P<.10). After all five hCG injections in 1987, mean ovulation time for treated mares was shorter than in control mares (P<0.01). However, after two to five hCG injections in 1987, seven treated mares (50%) had some individual ovulation times that did not differ from the control mares.Initially, following the first three injections of hCG in 1988, mean duration of estrus tended to be shorter (P<0.1) in treated mares compared to control mares. A reduction in mean ovulation time was observed after the first two hCG injections of 1988 (P<0.01). However, after one to four hCG injections in 1988, eight treated mares (57.1%) had some individual ovulation times that did not differ from controls.In 1987, all 14 treated mares developed significant levels of antibodies to hCG after one to four injections, and again in 1988, were positive for anti-hCG antibodies after one to three injections. However, no correlation was observed between magnitude of the immune response and duration of ovulation time or pregnancy rate. In cross reactivity studies, no significant binding of plasma anti-hCG antibodies to either eLH or eCG was observed in vitro.Overall, pregnancy and foaling rates of treated (85.7%) and control mares (83.3%) did not differ. Additionally, no difference was observed in number of inseminations per estrus between treated and control mares. In this study, with successive injections of hCG, the expected shortened time to ovulation was not elicited consistently in all mares. However, mares continued to ovulate, conceive and foal in the presence of significant levels of anti-hCG antibodies.     

Timed artificial insemination in crossbred mares: Reproductive efficiency and costs

Timed artificial insemination (TAI) has boosted the use of conventional artificial insemination (CAI) by employing hormonal protocols to synchronize oestrus and ovulation. This study aimed to evaluate the efficiency of a hormonal protocol for TAI in mares, based on a combination of progesterone releasing intravaginal device (PRID), prostaglandin (PGF_2α) and human chorionic gonadotropin (hCG); and compare financial costs between CAI and TAI. Twenty-one mares were divided into two groups: CAI group (CAIG; n = 6 mares; 17 oestrous cycles) and TAI group (TAIG; n = 15 mares; 15 oestrous cycles). The CAIG was subjected to CAI, involving follicular dynamics and uterine oedema monitoring with ultrasound examinations (US), and administration of hCG (1,600 IU) when the dominant follicle (DF) diameter's ≥35 mm + uterine oedema + cervix opening. The AI was performed with fresh semen (500 × 10⁶ cells), and embryo was recovered on day 8 (D8) after ovulation. In TAI, mares received 1.9 g PRID on D0. On D10, PRID was removed and 6.71 mg dinoprost tromethamine was administered. Ovulation was induced on D14 (1,600 IU of hCG) regardless of the DF diameter's, and AI was performed with fresh semen (500 × 10⁶ cells). On D30 after AI, pregnancy was confirmed by US. The pregnancy rate was 80.0% in TAIG and 82.3% in CAIG (p > .05). The TAI protocol resulted in 65% reduction in professional transport costs, and 40% reduction in material costs. The TAI was as efficient as CAI, provided reduction in costs and handlings, and is recommended in mares.     

The Use of Equine Follicle Stimulating Hormone to Increase Equine Chorionic Gonadotropin in the Pregnant Mare

Equine chorionic gonadotropin (eCG), obtained from pregnant mares, is used for assisted reproductive technologies in laboratory rodents and livestock. The objective of the present study was to use equine follicle-stimulating hormone (eFSH) to increase the incidence of twin pregnancies, through multiple ovulations, and increase eCG. Nineteen light horse–type mares were enrolled in the study. The control group (n = 9) was bred with fresh or cooled semen and given human chorionic gonadotropin (hCG) at the time of breeding. The second group (n = 10) was given 12.5 mg of eFSH intramuscularly twice a day beginning 5–7 days after ovulation. Prostaglandin F2α was administered intramuscularly the second day of eFSH treatment. Treatment with eFSH continued until follicles were >35 mm in diameter, and mares were then given no treatment for 36 hours. The mares were then bred with fresh or cooled semen from the same stallion as the control group and given hCG. Blood samples were taken weekly from day 35 to day 105 after ovulation. Serum concentration of eCG was obtained, and data were analyzed with multivariate analysis using the mixed procedure. Significance was set at P < .05. Data were combined for all mares carrying twins and compared with those carrying singletons. The group of mares carrying twins had higher peak concentrations of eCG and higher values for area under the curve compared with mares carrying singletons (P < .05). These results suggest inducing twins could be a method used to increase eCG production.