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.     

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.     

Prolonged interovulatory interval and hormonal changes in mares following the use of Ovuplant™ to hasten ovulation

In its first year of commercial availability in the United States, reports from the field indicated that Ovuplant™ (a deslorelin-containing slow-release implant for hastening ovulation in mares) was associated with a delayed return to estrus in mares not becoming pregnant. Supposedly this effect was particularly prevalent in mares subsequently administered PGF_2α to cause luteal regression after embryo collection. The present experiment was conducted 1) to determine if the field observations were repeatable under controlled experimental conditions, and 2) to gather endocrine data that might yield information on the underlying cause(s) of this observation. Twenty-five light horse mares were used. Ovaries of each mare were examined by transrectal ultrasonography daily during estrus until ovulation. Once a follicle >30 mm was detected, the mare received either Ovuplant (treated group; N = 13) at the recommended dosage or a sham injection (controls; N = 12); treatments were administered in a manner to ensure that they were unknown to personnel involved with data collection. On day 7 after ovulation, each mare received a luteolytic injection of PGF_2ð. Mares were examined every other day until return to estrus or development of a 30 mm follicle, at which time daily examination was performed until ovulation. Jugular blood samples were collected daily. Two mares receiving Ovuplant did not return to estrus within 30 days and their data were not included in the statistical analyses; in contrast, no control mare exhibited such an extended interovulatory interval. For all other mares receiving Ovuplant, the interval between the first and second ovulations was longer (P = .0001) than that of control mares by an average of 6.2 days. In addition, plasma LH concentrations were lower (P <.05) in the treated mares on days 0 through 4, 9, 11, 18, and 19 after the first ovulation. Plasma FSH concentrations were also lower (P = .017) in treated mares from days 4 to 11 and on days 6 and 5 prior to the second ovulation (P = .005). Differences in progesterone and estradiol were observed but were less consistent than for LH and FSH. Mares receiving Ovuplant had fewer small (P =.026), medium (P = .003) and large (P = .045) follicles prior to the second ovulation. In conclusion, Ovuplant treatment at the recommended dosage decreased follicular activity after ovulation and increased the interovulatory interval in mares short-cycled with PGF_2ð. These effects appear to be mediated by a hyposecretion of LH and(or) FSH.     

Endocrine changes and ultrasonography of ovaries in suckled beef cows during resumption of postpartum estrous cycles

Changes in follicular and luteal structures were assessed and concentrations of estradiol and progesterone were measured in 13 Hereford X Angus suckled beef cows during resumption of estrous cycles. Transrectal ultrasonography was used to monitor follicular size, ovulation, and formation and regression of the corpus luteum (CL). The interval from parturition to first postpartum ovulation (FO) was 82 +/- 4.7 d. Serum progesterone remained low before FO. One cow exhibited standing estrus, two cows showed other signs of estrus, and 10 displayed no signs of behavioral estrus preceding FO. All cows exhibited standing estrus before the second postpartum ovulation (SO). All cows had a short luteal phase after FO, with an average interval of 8.5 +/- .2 d between FO and SO. Concentrations of estradiol in serum during the 8 d preceding ovulation were similar before FO and SO. Maximal diameter of the preovulatory follicle was similar before FO and SO. However, the ovulatory follicle was larger in diameter at 2 d (P = .02) and 3 to 8 d (P less than .005) before FO than before SO. The time from detection until ovulation was less (P = .005) for the ovulatory follicle preceding SO than for the follicle associated with FO (8.5 vs 10.2 d, respectively, SE = .4). The second-largest follicle was larger (P less than .005) in diameter during the 8 d preceding the FO than before the SO. The difference in size between the ovulatory follicle and the second-largest follicle on the day before ovulation was greater (P less than .005) preceding SO than preceding FO (8.7 vs 6.6 mm, respectively, SE = .4).(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

Induction of short-term pseudopregnancy in gilts by the administration of estradiol benzoate or estradiol dipropionate to achieve ovulatory synchronization for embryo collection

A study was conducted to investigate whether ovulation in gilts could be synchronized for embryo collection by the administration of estradiol benzoate (EB) or estradiol dipropionate (EDP) to induce pseudopregnancy, followed by the treatment with prostaglandin F_2α (PGF_2α) on 10 days after. Ten gilts each received a total of 20 mg of EB or EDP on Day 10 or EB on Day 10 and 14 to induce pseudopregnancy (Day 0 = onset of estrus). Donors received PGF_2α 10 or 15 days (as a control) after the first administration of estrogens and subsequently eCG and hCG, and were then inseminated artificially. The embryos were collected 7 days after the administration of hCG, and assessed for embryo yield and their developmental stages. All protocols resulted in good embryo yield (9.8–13.2 embryos in average), and the embryos showed average ability to develop to the expanded blastocyst stage (3.29–4.03 as developmental scores) without any significant differences among the protocols. These results suggest that the administration of PGF_2α 10 days after the treatment of gilts with EB or EDP would allow synchronization of ovulation and embryo collection, as well as shortening the period from estrus detection to embryo collection, thus improving embryo collection efficiency.     

Estrus synchronization and pregnancy rates in beef cattle given CIDR-B, prostaglandin and estradiol, or GnRH

Two experiments were conducted to determine estrous response and pregnancy rate in beef cattle given a controlled internal drug release (CIDR-B) device plus prostaglandin F2 alpha (PGF) at CIDR-B removal, and estradiol or gonadotropin releasing hormone (GnRH). In Experiment I, crossbred beef heifers received a CIDR-B device and 1 mg estradiol benzoate (EB), plus 100 mg progesterone (E + P group; n = 41), 100 micrograms gonadotropin releasing hormone (GnRH group; n = 42), or no further treatment (Control group; n = 42), on Day 0. On Day 7, CIDR-B devices were removed and heifers were treated with PGF. Heifers in the E + P group were given 1 mg EB, 24 h after PGF, and then inseminated 30 h later. Heifers in the GnRH group were given 100 micrograms GnRH, 54 h after PGF, and concurrently inseminated. Control heifers were inseminated 12 h after onset of estrus. The estrous rate was lower (P < 0.01) in the GnRH group (55%) than in either the E + P (100%) or Control (83%) groups. The mean interval from CIDR-B removal to estrus was shorter (P < 0.01) and less variable (P < 0.01) in the E + P group than in the GnRH or Control groups. Pregnancy rate in the E + P group (76%) was higher (P < 0.01) than in the GnRH (48%) or Control (38%) groups. In Experiment II, 84 cows were treated similarly to the E + P group in Experiment I. Cows received 100 mg progesterone and either 1 mg EB or 5 mg estradiol-17 beta (E-17 beta) on Day 0 and either 1 mg of EB or 1 mg of E-17 beta on Day 8 (24 h after CIDR-B removal), in a 2 x 2 factorial design, and were inseminated 30 h later. There were no differences among groups for estrous rates or conception rates. The mean interval from CIDR-B removal to estrus was 44.2 h, s = 11.2. Conception rates were 67%, 62%, 52%, and 71% in Groups E-17 beta/E-17 beta, E-17 beta/EB, EB/E-17 beta, and EB/EB, respectively. In cattle given a CIDR-B device and estradiol plus progesterone, treatment with either EB or E-17 beta effectively synchronized estrus and resulted in acceptable conception rates to fixed-time artificial insemination.     

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 estradiol cypionate (ECP) on ovarian follicular development and ovulation in dairy cattle.

The efficacy of estradiol cypionate (ECP) for synchronizing ovarian follicular development was determined in lactating Holstein-Friesian cattle. In Experiment 1, 13 cattle were given simultaneous intramuscular (i.m.) injections of 100 mg progesterone and 0 (control), 0.5 or 1.0 mg ECP on Day 3, after a synchronized ovulation (Day 0). Maximum diameter of the dominant follicle of Wave 1 was significantly larger in control cattle than in those given 0.5 or 1.0 mg ECP (means: 15.7, 13.2, and 12.9 mm, respectively). Mean day of emergence of Wave 2 was significantly later in controls than in those given 1.0 mg ECP, with the 0.5 mg group intermediate (Days 10.2, 8.8 and 9.5, respectively). In Experiment 2, 14 cattle were given a CIDR-B and IM injections of 1 mg ECP and 50 mg progesterone without regard to stage of cycle (treatment = Day 0). On Day 8, the CIDR-B was removed and 500 micrograms cloprostenol injected, IM. Mean days of wave emergence (Day 3.4; range: -2 to 7) and ovulation (Day 12.1; range: 10 to 14) indicated that ECP had limited efficacy for synchronizing follicular development and ovulation in dairy cattle when given at random stages of the estrous cycle.     

Effect of eFSH on Ovarian Cyclicity and Embryo Production of Mares in Spring Transitional Phase

The use of equine FSH (eFSH) for inducing follicular development and ovulation in transitional mares was evaluated. Twenty-seven mares, from 3 to 15 years of age, were examined during the months of August and September 2004, in Brazil. Ultrasound evaluations were performed during 2 weeks before the start of the experiment to confirm transitional characteristics (no follicles larger than 25 mm and no corpus luteum [CL] present). After this period, as the mares obtained a follicle of at least 25 mm, they were assigned to one of two groups: (1) control group, untreated; (2) treated with 12.5 mg eFSH, 2 times per day, until at least half of all follicles larger than 30 mm had reached 35 mm. Follicular activity of all mares was monitored. When most of the follicles from treated mares and a single follicle from control mares acquired a preovulatory size (≥35 mm), 2,500 IU human chorionic gonadotropin (hCG) was administered IV to induce ovulation. After hCG administration, the mares were inseminated with fresh semen every other day until ovulation. Ultrasound examinations continued until detection of the last ovulation, and embryo recovery was performed 7 to 8 days after ovulation. The mares of the treated group reached the first preovulatory follicle (4.1 ± 1.0 vs 14.9 ± 10.8 days) and ovulated before untreated mares (6.6 ± 1.2 vs 18.0 ± 11.1 days; P < .05). All mares were treated with prostaglandin F_2α (PGF_2α), on the day of embryo flushing. Three superovulated mares did not cycle immediately after PGF_2α treatment, and consequently had a longer interovulatory interval (22.4 vs 10.9 days, P < 0.05). The mean period of treatment was 4.79 ± 1.07 days and 85.71% of mares had multiple ovulations. The number of ovulations (5.6 vs 1.0) and embryos (2.0 vs 0.7) per mare were higher (P < 0.05) for treated mares than control mares. In conclusion, treatment with eFSH was effective in hastening the onset of the breeding season, inducing multiple ovulations, and increasing embryo production in transitional mares. This is the first report showing the use of FSH treatment to recover embryos from the first cycle of the year.     

Estradiol and progesterone concentrations resulting from the administration of an exogenous hormone regimen in diestrous embryo transfer recipients

Estradiol and progesterone concentrations were evaluated from diestrous embryo transfer recipient mares (5 to 14 days post-ovulation) which were treated with an exogenous hormone regimen. Upon detection of the donor mare's ovulation (0 hours), 10 mg PGF_2α was given to the recipient mare; at 12, 24 and 36 hours 20 mg estradiol cypionate; at 48 hours, 500 mg progesterone in oil and then 22 mg altrenogest at 60, 72 and 96 hours. Altrenogest (22 mg/day) was continued until end of the trial (detection of a fetal heart beat). Embryos were transferred non-surgically 6 or 7 days after the start of treatment.Plasma samples were evaluated over three periods; period 1-between recipient mare ovulation and prior to PGF_2α period 2-between PGF and embryo transfer and period 3-post-transfer. During periods 2 and 3, estradiol was higher (P<.05) for mares which were 10 to 14 days post-ovulation (late diestrous) as compared to mares which were 5 to 9 days post ovulation (mid-diestrous) when treatment began. Progesterone concentrations were higher (P<.05) for the mid-diestrous mares in the same periods. The pregnancy rate was higher for the late diestrous mares than the mid-diestrous mares (58% (7/12) vs 10% (1/10)). However, no difference (P>.05) was detected in estradiol or progesterone in the late diestrous mares which were pregnant or open. During period 2, estradiol was higher (P<.05) in the pregnant than open mares. Whereas, during period 3, progesterone was higher (P<.05) in the open mares.These data suggest that estradiol is important for the establishment of pregnancy in the mare. Furthermore, hormone treatment developed in this study appears to have some potential in synchronization of diestrus mares to be used as embryo recipients.     

Evaluation of Injectable Sustained Release Progestin Formulations for Suppression of Estrus and Ovulation in Mares

Thirty-one mares were used in an experiment to evaluate the effectiveness of three sustained-release injectable formulations of altrenogest and one formulation of medroxyprogesterone acetate (MPA) for long-term suppression of estrus and ovulation. Luteolysis was induced by injection of prostaglandin-F_2α (Lutalyse) on day 0 (6th day after the previous ovulation) and was immediately followed by treatment with 1) no injection (controls; n = 7), 2) 1.5 mL of an altrenogest solution in sustained-release vehicle (LA 150, 1.5 mL; 225 mg altrenogest; n = 6), 3) 3 mL (450 mg altrenogest) of the same solution (n = 6), 4) 500 mg altrenogest in lactide-glycolide microparticles suspended in 7-mL vehicle (MP 500; n = 6), or 5) 1.0 g MPA as a 5-mL suspension. Mares were checked for estrus daily, and their ovaries scanned every other day until a 25-mm or greater follicle was detected, after which they were scanned daily. Control mares returned to estrus an average of 3.9 days after Lutalyse administration; all the single-injection altrenogest formulations increased (P < .05) the days to return to estrus, with the greatest increase occurring in mares receiving MP 500. Return to estrus was not affected by MPA treatment. Time of ovulation was determined by serial ultrasound scans and confirmed by daily plasma luteinizing hormone (LH) and progesterone concentrations. Control mares ovulated an average of 8.8 days after Lutalyse administration. Treatment with 1.5 or 3 mL of LA 150 increased (P < .05) the mean days to ovulation to 16.5 and 21.2 days, respectively; MP 500 increased (P < .05) the days to ovulation to 33.5 days. Administration of MPA did not affect (P > .1) days to ovulation relative to control mares. The MP 500 treatment provided long-term suppression of estrus and ovulation and could prove useful for that purpose. Treatment with the LA 150 solutions provided shorter-term suppression, and a relatively tight grouping of the individual mares around the mean days to ovulation; these one-shot formulations could be useful for synchronizing ovulation in cyclic mares and inducing normal estrous cyclicity in vernal transitional mares exhibiting erratic, anovulatory estrous periods.     

Effects of time of insemination relative to ovulation on pregnancy rate and embryonic-loss rate in mares

The effects of pre-ovulatory and post ovulatory insemination on pregnancy rate and embryonic-loss rate were studied in 268 mares in two experiments. Within each experiment mares were randomised within replicates as follows: to be inseminated on the day the pre-ovulatory follicle reached 35 mm (pre-ovulatory group), to be inseminated on the day of ovulation (Day 0 group), and to be inseminated on the day after ovulation (Day 1 group). Ultrasonic pregnancy diagnoses were performed on Days 11, 12, 13 and 14 (Experiment 1) and Days 11, 12, 13, 14, 15, 20 and 40 (Experiment 2). Combined for the two experiments, pregnancy rates were different (P less than 0.01) among the three groups. For Experiment 2, pregnancy rate within the pre-ovulatory group was lower (P less than 0.05) for insemination 4 days or more before ovulation than for up to 3 days before ovulation. Pregnancy rate was lower (P less than 0.05) for the Day 0 group than for the pre-ovulatory group inseminated up to 3 days before ovulation. In Experiment 2, ovulation was detected by examinations every 6 h; pregnancy rate was greater (P less than 0.05) for mares inseminated 0 to 6 h after ovulation than for mares inseminated at 18 to 24 h. No pregnancies occurred when mares were inseminated 30 h or more after ovulation. The mean day of first detection of the embryonic vesicle was different (P less than 0.0001) among the three groups. Diameter of embryonic vesicle averaged over Days 11 to 15 also differed (P less than 0.0001) among groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

Estradiol interactions with dopamine antagonists in mares: Prolactin secretion and reproductive traits

Two experiments studied the effects of pretreatment with estradiol benzoate before treatment with a dopamine antagonist on prolactin secretion and reproductive traits in mares during (1) the seasonal anovulatory period and (2) the normal breeding season. Experiment 1 was performed in winter with 17 mares selected for low follicular activity. Nine mares received estradiol benzoate injections every other day for a total of 10 injections; 8 mares received similar injections of vehicle. Ten days after onset of injections, all mares were placed on daily injections of sulpiride (250 mg) for 35 days or until ovulation. Plasma prolactin concentrations were higher (P < .001) in mares receiving estradiol than in controls for all assessments from days 12 through 36. Plasma luteinizing hormone (LH) concentrations were also increased (P < .05) by estradiol treatment from days 14 to 23. Mean day of first ovulation was 73.6 for control mares and 29.0 for estradiol-treated mares (P = .016). Estradiol treatment greatly enhanced prolactin secretion in response to sulpiride and increased LH secretion in seasonally anovulatory mares, which together hastened the date of first ovulation by an average of 45 days. Experiment 2 was designed to assess the efficacy of a long-acting, single-injection microparticle preparation of another dopamine antagonist, domperidone, for increasing prolactin secretion in cyclic mares in the summer. The experimental design and procedures used in experiment 1 were repeated, except that a single 3-g domperidone-microparticle injection was administered on day 11 rather than 45 days of sulpiride injections. Day 0 was the first day of estrus for each mare. Prolactin concentrations were higher (P < .05) in mares receiving estradiol than in control mares from days 12 through 25 and after a thyrotropin-releasing hormone injection on d 21. Estrous cycle traits (time to ovulation and time of luteal regression) were not affected (P > .1) by treatment. Estradiol enhanced the prolactin response to a single injection of 3 g domperidone in cyclic mares in the summer in a manner similar to the estradiol enhancement of prolactin secretion in response to daily sulpiride injections in anovulatory mares in winter. Thus, the single injection of domperidone could possibly replace the daily sulpiride injections used in experiment 1 to induce ovulation in seasonally anovulatory mares; this needs to be tested in future experiments.     

Timing of induction of ovulation in mares treated with Ovuplant or Chorulon

Reliable induction of timed ovulation is an important managerial tool in any horse-breeding operation. Not only does breeding close to ovulation increase pregnancy rates when using cooled, frozen, or poor-quality semen, but it also reduces the number of inseminations needed per cycle, resulting in a more efficient breeding program. To better predict ovulation time in the long estrus period of the mare, one could increase the frequency of transrectal palpations and ultrasounds and/or implement hormonal therapies to induce ovulations. However, previous studies have been unclear on the exact timing of ovulation of mares treated with human chorionic gonadotropin (Chorulon, Intervet Inc, Millsboro, DE) or deslorelin acetate (Ovuplant, Pharmacia and UpJohn Co, Kalamazoo, MI). This study was designed to determine the timing of ovulation after Ovuplant or Chorulon treatment in normal cycling mares presented to the veterinary clinic. In addition, the pregnancy rates were determined for mares bred when a single insemination, using frozen or chilled semen, was performed at a fixed time (36 hours) after Ovuplant or Chorulon treatment. Thirty-two mares were given a subcutaneous injection of 7.5 mg of prostaglandin F2α (Lutlyse, Ft Dodge Animal Health, Ft Dodge, IA) 5 days after the last ovulation and were examined every 48 hours until estrus was detected based on a dominant follicle and the presence of endometrial edema as determined by ultrasonographic examination. Group 1 (N = 12) was treated intravenously with 2,500 units of Chorulon, and group 2 (N = 20) was treated subcutaneously with Ovuplant as soon as mares were determined to be in estrus. Once treated all mares were examined by rectal palpation and ultrasound at 0, 12, 24, 28, 30, 32, 34, 36, 38, 40, 42, 44, 48, 60, 72, 84, 96, hours or until ovulation was detected. Ovulation rate in response to Chorulon was 83.3% at 48 hours, 91.6% at 72 hours, and 100% at 96 hours. All of the mares in the Ovuplant-treated group had ovulated by 48 hours. Chi-square analysis of the data showed a significant (P < .01) variation in the distribution of ovulation times between mares treated with Chorulon and mares treated with Ovuplant. This study provides enough evidence to support the hypothesis that timing of ovulation is a more reliable event in mares treated with Ovuplant compared with those treated with Chorulon.     

Efficacy of Acupuncture Sites for Delivery of Agents to Control Estrus and Ovulation in Mares

This study was designed to determine if prostaglandin F₂α (PGF₂α) when administered on d 6 post-ovulation in a low dose in the lumbosacral space (LSS) would induce luteolysis while minimizing side effects usually associated with intramuscular administration of this analogue in mares. A second objective was to determine if human chorionic gonadotropin (hCG) injected into the LSS would reduce time to ovulation in the mare. Ten normally cycling mares served as their own controls in a crossover design, receiving intramuscular injections of PGF₂α(10 mg), intravenous injections of hCG (3000 IU) and injections of PGF and hCG at the acupuncture site (2 mg and 3000 IU, respectively), as well as sham injections of saline. Beginning 12 h after injection, mean progesterone concentrations were less (P<0.05) in PGF₂α-treated mares than in mares receiving saline. Moreover, progesterone concentrations were similar (P<0.001) between both groups of mares receiving PGF₂α. In addition, there was no difference (P>0.1) between mares receiving the acupuncture injection of PGF₂α and the intramuscular injection in days to ovulation. However, duration and severity of side effects associated with PGF₂α administration were dramatically decreased (P<0.01) when PGF₂α was delivered to the acupuncture site compared to intramuscular delivery. The time to ovulation was similar (P>0.1) for mares receiving shams, or hCG. These data indicate that delivery of 2 mg of PGF₂α in the LSS induces luteolysis and reduces the sweating and muscle cramping associated with PGF₂α administration. There was no advantage to the delivery of hCG in the LSS.     

Efficacy of synovex-S implants in suppression of estrus in the mare

The objective of this study was to determine if administration of Synovex-S^® implants, approved for use in cattle to promote weight gain and feed efficiency, would suppress the expression of behavioral estrus and/or alter follicular development and ovulation in the mare. Twenty-four clinically normal adult horses were randomly assigned to one of four treatment groups (6 mares per group) which received a total of 0, 8, 32 or 80 Synovex-S^® pellets 5 days after ovulation. An implant dose of 8 Synovex-S^® pellets contained a total of 200 mg of progesterone and 20 mg of estradiol benzoate. Mares were monitored daily by teasing and ultrasonography of the reproductive tract per rectum for 45 days. A blood sample was collected daily for progesterone analysis. All mares receiving Synovex-S^® implants returned to estrus at the predicted time. No differences were noted in duration of estrus, interovulatory interval or ovulation rate. In conclusion, subcutaneous administration of 8, 32 or 80 Synovex-S^® pellets did not suppress the expression of behavioral estrus or block ovulation in mares.     

Follicular persistence induced by adrenocorticotropic hormone administration in goats

The objective of the present study was to examine the effect of adrenocorticotropic hormone (ACTH) administration on the induction of persistent cystic follicle in the goat in order to establish a method to experimentally induce cystic follicle. Four cross-bred goats were intramuscularly administered ACTH at 0.78 and 6.25 μg/10 kg twice a day from Days 15 to 21 (Day 0 was defined as the day of last estrus). Follicular status in the ovary was monitored by ultrasound examination. The plasma concentrations of estradiol, progesterone and cortisol were measured. Treatment with ACTH at the 0.78 and 6.25 μg/10 kg levels caused persistent follicles (> 10 days delay from the expected ovulation date) in 50% of the goats in both treatment groups. In those animals, ovulation occurred 17 and 27 days and 11 and 12 days after the expected days in the 0.78 and 6.25 μg/10 kg groups, respectively. The maximum follicle diameters were 10 and 9 mm in the 0.78 and 6.25 μg/10 kg ACTH groups, respectively. In the control group, the estradiol concentration increased on Day 18 and remained at a high level for a few days. However, such an increase was not seen in both ACTH groups. The estradiol concentration increased gradually from Days 21 to 27 in the 6.25 μg/10 kg ACTH group. These results suggest the possibility that ACTH induces persistent follicles in goats, which may be related to the delay of the onset of estradiol secretion followed by its maintenance at a high concentration.     

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.     

Quantitative Echotexture Analysis for Prediction of Ovulation in Mares

The aim of this study was to predict the ovulation in mares by quantitative analysis of the echotextural changes of preovulatory follicular walls. Four mares of breeding age with 32 preovulatory follicles and 11 anovulatory follicles were observed by ultrasonography. The slope of the regression line of the follicular wall and the echogenicity score of granulosa layer (GL) and anechoic layer (AL) were measured from the images on Days -3 (Day 0 = ovulation), -2, and -1, respectively. GL was scored from 1 (anechoic) to 3 (echoic), and prominence of AL was recorded from 1 (gray and thin) to 3 (black and thick). The results indicated that the regression line of the follicular wall for 81.3% (26/32) of preovulatory follicles had the slope value ≥19.0 on Day -1, in which 4 of the 26 preovulatory follicles were ≥19.0 on Day -2 already. Mean slope value on Day -1 (21.9 ± 1.5) was significantly greater (P < .01) than on Day -2 (15.0 ± 1.4) and Day -3 (14.0 ± 1.1). All of the slope values for the 11 anovulatory follicles were <19.0 on any given day. GL and AL scores of preovulatory follicles were significantly greater (P < .01) than in anovulatory follicles on Days -3, -2 and -1; nevertheless, only 28.1% (9/32) of preovulatory follicles scored 3 for both GL and AL simultaneously on Day -1. All anovulatory follicles scored <2 for both GL and AL on Day -1. It was concluded that the slope of the regression line of the follicular wall is useful in predicting preovulatory follicles within 48 hours of ovulation when the value is ≥19.0. Of these follicles (N = 26), 84.6% (22/26) were predicted to ovulate within 24 hours, and 15.4% (4/26) within 24 to 48 hours.

Introduction

Insemination in mares by accurately predicting the time of ovulation may obtain maximum fertility with minimum use of semen, and therefore would definitely be a profitable advantage in the horse farming business. The optimal time for insemination with frozen-thawed semen usually include a shorter interval than if fresh semen or natural breeding is used. To achieve the maximal pregnancy rates with frozen-thawed semen, it is necessary to inseminate mares during a period between 12 hours pre- and 6 hours post-ovulation.[1] Therefore, if the timing of ovulation could be predicted, it would be helpful for the veterinarian to inseminate a mare only once per cycle if performed very close to the time of ovulation. [2] In recent years, many indicators have been reported for predicting impending ovulation in mares, including measurement of electrical resistance of the vaginal mucus, [3] the distinguishable endometrial folding pattern of uterus in estrus, [4] changes in size and shape of the preovulatory follicles, [5, 6 and 7] and the echotexture changes in the preovulatory follicular wall. [8] The latter has been more efficient for predicting the imminence of ovulation; nevertheless, their assessment of criterions was scored subjectively. The hypothesis for this study was based on the published report from Gastal et al in 1998 [8]; they found that 2 echotexture changes of the preovulatory follicle-increasing echogenicity of the granulosa layer and increasing prominence of an anechoic layer beneath the granulosa, were detected in the follicular wall as ovulation approached in mares. Computer-assisted image analysis is an advanced technology for diagnostic ultrasonography to improve the reproductive management of patients. [9, 10 and 11] The purpose of this study is to quantify the echotextural changes in the preovulatory follicular wall as ovulation approaches using computer-assisted image analysis, so that the quantified echotexture changes could serve as an indicator for prediction of ovulation in mares.

Materials and Methods

Animals and Ultrasonography

Four non-lactating and nonpregnant mixed mares between 4 and 14 years of age and weighing between 450 and 550 kg were studied from January to December 2001. The geographic area of the mares in this study was in subtropical Taiwan of the northern hemisphere. All mares were maintained on alfalfa/grass hay and had access to water and mineralized salt. A teaser stallion was introduced to detect the estrus signs of mares about 2 weeks after the end of the last estrus. Follicular changes were monitored with a real-time B-mode linear assay ultrasound scanner, equipped with a 7.5-MHz transrectal probe (Model Scanner 200 Vet, Pie Medical, The Netherlands). Upon detection of a preovulatory follicle, ultrasound examination was performed daily and continued until ovulation. A total of 32 preovulatory follicles and 11 anovulatory follicles were identified from a retrospective determination.Ultrasonographic images were recorded on Hi-8 MP videotape with a Sony DCR-TRV 120 Digital-8 camera. The brightness and contrast controls of the monitor and the time-gain compensation of the scanner were standardized to constant settings throughout the observation period.

Image Analysis

Still images were subsequently captured and saved as TIF files by computer using a digital image analysis program (Image-Pro Express V4.0 for Windows, Media Cybernetics, L.P., USA) with a resolution of 640 × 480 pixels and 256 shades of gray. Echotexture of the regions of interest was defined in terms of pixel intensity ranging from 0 (black) to 255 (white). Three ultrasonographic images of each preovulatory follicle at its distinctly discernible cross section were subsequently selected. To avoid the enhancement of through-transmission, sampling regions were located within the 10 or 2 o'clock position for measurement of pixel values (Fig 1). The pixel values were measured with the “Line Profile” tool, which involved sampling pixel values along a line traversing the follicle wall from the peripheral antrum, GL, AL, to the stroma. A graph of the pixel intensities along the line was produced ( Fig 2). The GL was defined as the highest pixel after which there was a sequential fall in gray-scale values. The pixel values along the curve (P0, P1, P2) were obtained as an average of 9 measurements (3 images per follicle and 3 lines per image) and were used to measure the slope of a regression line of the fall segment ( Fig 2).