Endocrine Profile Comparisons of Fat Versus Moderately Conditioned Mares Following Parturition

The influence of progesterone, luteinizing hormone (LH), and estrogen on the mare's estrous cycle has been well researched and documented, but other endocrine profiles have not received as much attention. To evaluate endocrine concentrations in fat-conditioned (body condition score [BCS] of 7–8) versus moderately conditioned mares (BCS of 5–6), 24 mares were allotted to and maintained in respective groups from late gestation until pregnancy was confirmed after breeding on the second postpartum estrus. Serum concentrations of thyroxine (T₄), insulin-like growth factor-1 (IGF-1), and leptin were assayed to characterize circulating blood concentrations. Additionally, LH and progesterone serum concentrations were assayed to evaluate the estrous cycle status of the mares. Leptin and progesterone concentrations were not different (P > .05) between the groups. Nevertheless, serum concentrations of T₄ were higher (P < .01) and IGF-1 concentrations lower (P < .01) in moderately conditioned as compared with fat-conditioned mares during times of ovulation and the interovulatory period. Furthermore, serum concentrations of LH were found to be different between the groups only when the estrous cycle approached the second ovulation (P < .0001). Results of this study suggest that mares maintained in a BCS of 5 or greater are similar in terms of reproductive efficiency. Although the circulating serum concentrations of T₄ and IGF-1 are different after parturition, their influence does not affect reproductive capabilities of mares with a BCS of 5 or greater.     

Retrospective study on the incidence of postinsemination uterine fluid in mares inseminated with frozen/thawed semen

Uterine fluid accumulation has been reported after insemination or natural breeding of mares. This retrospective study examined the factors affecting the incidence of uterine fluid after insemination of frozen semen. Specifically, this study determined the association between mare age, reproductive status, fluid accumulation, and pregnancy rates in mares. Records were available from 283 warmblood mares throughout 496 cycles. Mares were divided into maiden, foaling, and barren and age groups of 3 to 9, 10 to 16, and more than 16 years. Mares were inseminated only once with frozen semen within 4 to 8 hours before or after ovulation. Ultrasound examinations were performed 12 to 18 hours after insemination. A depth of at least 20 mm of fluid was considered significant. Mares with less than 20 mm were treated with oxytocin, and those with more than 20mm of fluid were given oxytocin and uterine lavage. Pregnancy determination was performed at 14 to 16 and 30 to 50 days after ovulation. Fluid level of more than 20 mm was recorded in 25% of the cycles. Barren mares and aged mares (10-16 and > 16 years) had a higher incidence of uterine fluid accumulations. Per-cycle pregnancy rate was lower (45%) in mares with uterine fluid than in mares without uterine fluid (51%). This difference was primarily due to the reduction in fertility of mares who were older than 16 years and retained fluid after insemination. Apparently, oxytocin and lavage treatments provided acceptable fertility in the other groups of mares that had uterine fluid.

Introduction

Use of equine frozen semen is accepted by the majority of horse registries. According to several field studies,[1, 2, 3, 4 and 5] insemination of frozen semen has resulted in acceptable pregnancy rates. Postbreeding fluid accumulation is a physiologic inflammation that clears the uterus of foreign material such as excess spermatozoa, seminal plasma, bacteria, and extenders. [6, 7, 8, 9 and 10] Uterine fluid can be easily diagnosed with ultrasonography. [10, 11 and 12] Persistent postbreeding uterine fluid has been associated with a decrease in fertility after natural mating or artificial insemination (AI) of fresh semen. [11, 12 and 13] Predisposing factors to persistent fluid accumulations are reduced myometrial contractions, poor lymphatic drainage, large overstretched uterus, and cervical incompetence. [7, 14 and 15] Normal mares are able to expel uterine fluid quickly after inseminations, whereas susceptible mares accumulate fluid in their uterine lumen for more than 12 hours after breeding or insemination. [10]It is commonly stated that insemination with frozen semen leads to greater post-AI fluid accumulation than insemination with fresh or cooled semen or after natural mating. Apparently, there is only 1 controlled study on this comparison.[7] The authors reported that infusion of frozen semen resulted in a greater inflammatory response than natural breeding. In a field study, [16] 16% of mares naturally mated had persistent postbreeding fluid accumulations compared with a 30% rate reported for mares inseminated with frozen semen. [1 and 2] More recently, Watson et al. [17] reported a postbreeding fluid accumulation rate of 16%, which is identical to that reported for natural mating. [16] It is difficult to compare studies because details of mare selection and insemination or breeding frequencies are not always reported. Obviously, a higher proportion of barren and aged mares in a study would increase the incidence of postbreeding fluid accumulation. [1 and 2]The study presented herein was a retrospective study designed to determine the incidence of postbreeding fluid accumulation in a large number of mares inseminated with frozen semen. Associations were determined between mare age, reproductive status and fluid accumulation, and pregnancy rate in mares with and without uterine fluid accumulation.

Materials and methods

Mares

Records were available from 283 warmblood mares inseminated with frozen semen at the Cristella Veterinary Clinic in Italy during 1998 to 2001. Mares ranging in age from 3 to 20 years were inseminated with semen that was frozen in 10 centers and was from 34 stallions. The broodmare population was subdivided into 3 reproductive groups: 89 maiden mares (mean age, 7.2 years), 106 foaling mares (mean age, 9.4 years), and 87 barren mares (mean age, 11.9 years). Maiden mares older than 7 years were selected with biopsy scores of 1 or 2 only. Barren mares were open for no more than 2 consecutive seasons and had negative cytology and bacteriology scores. Age groups were divided as follows: 3 to 9 years (n = 132), 10 to 16 years (n = 137) and older than 16 years (n = 14). Data from 496 cycles were used. Distribution of the estrous cycles was 172, 157, and 167 in the maiden, foaling, and barren groups, respectively; and 224, 244, and 28 in the youngest, intermediate, and oldest groups, respectively.

Mare reproductive management and artificial insemination protocol

During estrus, all mares underwent a daily ultrasound examination with a 5-mHz transrectal probe (SA 600 Vet; Medison Inc., Seoul, South Korea) until 1 or more 35-mm ovarian follicles were detected. Ovulation was then induced by the intravenous administration of 2000 IU of human chorionic gonadotropin (hCG). Ultrasound examination was performed 12 hours after hCG treatment and then every 4 to 8 hours until ovulation occurred. Mares were inseminated only once within a period of 4 to 8 hours before or after ovulation. The semen used was thawed according to the distribution center's instructions and had the following minimum post-thaw quality requirements: not less than 200 × 10⁶ progressively motile spermatozoa per dose and a minimum of 30% progressive spermatozoal motility. Foaling mares were not inseminated at their first postpartum (“foal heat”) estrous period, because pregnancy rates are recognized to be lower than during the subsequent estrous periods.[18] During the first postpartum estrus, ovarian ultrasound scan examinations were performed every 2 to 3 days until an ovulation was detected. A prostaglandin F₂α injection was given 5 days later to short-cycle the mare.

Postinsemination monitoring

An ultrasound examination of the reproductive tract was performed 12 to 18 hours after insemination to detect any intrauterine fluid accumulation. The presence and depth of intrauterine fluid was recorded. Twenty millimeters or more of grade II or III intrauterine fluid[19] was recorded as a significant amount of fluid. Mares with less than 20 mm of fluid were treated with an intravenous injection of 20 IU oxytocin. For mares with more than 20 mm of fluid, oxytocin was administered, and the uterus was flushed daily with buffered saline solution: 1-L aliquots were infused and recovered until the recovered fluid was clear. In these mares, oxytocin treatment was repeated up to 3 times daily. Post insemination treatments were performed for no more than 4 days after ovulation had occurred.Pregnancy diagnosis was performed with ultrasound at 14 to 16 days after ovulation. Scans were then repeated at 30 and 50 days of gestation to confirm the presence in the uterus of an apparently healthy developing conceptus.

Statistical analysis

χ² Analysis was used to determine the effect of reproductive status and age on the incidence of fluid accumulation. In addition, the influence of persistent uterine fluid accumulation on pregnancy rates per cycle was determined for each reproductive class and age by using χ² analysis.

Results

The per-cycle pregnancy rate at 14-16 days after ovulation was 49.3% (245/496 cycles). By the end of the season, 245 of 283 mares (86.5%) were confirmed pregnant. Fluid level of at least 20 mm (grade II or III) was recorded in 126 of the 496 cycles (25.4%). Barren mares had a higher (P < .05) incidence of postbreeding fluid accumulation (64/167; 38.3%) than maiden (34/172; 19.7%) and foaling (28/157, 17.8%; Table 1) mares. The incidence of fluid accumulation was also higher in mares older than 16 years (19/28; 67.8%) than those aged 10 to 16 years (69/244; 28.2%) and 3 to 9 years (38/224; 17%). The incidence of uterine fluid was also higher (P < .05) for mares aged 10 to 16 years than those aged 3 to 9 years (Table 2). Overall, the per-cycle pregnancy rate was lower (P < .05) for mares with post-AI fluid accumulations than for those with no uterine fluid or only a small quantity of fluid (57/126, 41.9% vs 188/360, 56.2%). Pregnancy rates were similar (P > .05) for mares with or without uterine fluid when comparisons were made within maiden and barren mare groups. However, more foaling mares became pregnant when no fluid was detected after insemination. Pregnancy rate for this group (68.1%) was higher than that for maiden (44.2%) and barren (44.6%) mares (Table 3). Older mares with uterine fluid accumulations had a lower per-cycle pregnancy rate (36.8%) than mares in the same group but without fluid. Surprisingly, if no fluid was detected, the highest pregnancy rates were in mares older than 16 years ( Table 4).     

Twinning in mares: A review of recent studies

Recent studies on twinning are reviewed. Multiple ovulations were more frequent in thoroughbreds (19%) than in quarter horses (9%) and Appaloosas (8%). The multiple ovulation rate was reduced approximately 50% in foaling mares compared to barren and maiden mares. There was a high degree of repeatability of double ovulations and twin pregnancies within mares and within family lines.Only one embryo was found in each of 23 pituitary extract-treated mares with multiple, synchronous ovulations (<2 days apart) and in each of 39 brood-farm mares with double, synchronous ovulations. Pregnancy rates (number of mares pregnant, regardless of number of embryos/mare) were significantly higher for double, synchronous ovulations than for single ovulations in artificially stimulated mares (58% vs 38%) and in brood-farm mares (83% vs 54%). The results indicated that ova produced by synchronous, double ovulations are viable and fertilizable (indicated by the higher pregnancy rates), but that one of the resulting embryos is eliminated (indicated by the absence of twins).Synchronous, double ovulations were not recorded in association with any of 107 sets of natrually occurring twins. Most (76%) of the twin sets were associated with one detected ovulation. The remaining twins were associated either with one ovulation, but a large unovulated follicle was present at the time of the last examination (10%), or with asynchronous, double estrous ovulations (14%). Twins originated more frequently (P < .05) from asynchronous, double estrous ovulations (9/57) than from synchronous, double ovulations (0/39).Approximately 50% of the mares in which twin embryos were diagnosed rectally before day 31, had 1 foal. However, mares in which twins were recorded as present at day 32–36 and day 40–42 had a single foal in only 17% and 6% of the mares, respectively. The methods used for intervention when twins were diagnosed were unsatisfactory. Complete termination of pregnancy with a prostaglandin or intrauterine flushing resulted in failure to establish a singleton pregancy during the remaining breeding season in 10/11 mares. Attempts to eliminate one embryo resulted in loss of both embryos in 6/7 mares.     

Follicular Fluid Prolactin and the Periovulatory Prolactin Surge in the Mare

Prolactin may play multiple roles in equine reproduction. Prolactin appears to be associated with seasonal reproduction, and fluctuating prolactin levels during the estrous cycle suggest that it may play a role in estrous cyclicity as well. The purpose of this research was to investigate the activity of prolactin during the follicular phase of the estrous cycle. In experiment 1, prolactin concentrations were determined from plasma samples collected at least every other day throughout the estrous cycle. Periovulatory (ovulation ± 1 day) prolactin concentrations were compared with concentrations during early diestrus (days 2−10 postovulation). In experiment 2, prolactin concentrations were measured in follicular fluid collected from 74 follicles of various sizes. Follicles were grouped into small (≤20 mm), medium (21−35 mm), and large (>35 mm) size categories. Prolactin concentrations increased during the periovulatory period in cycling mares. This periovulatory surge was superimposed on baseline prolactin concentrations that varied with season. Prolactin was present in significant quantities in the follicular fluid. Follicular fluid prolactin concentrations were lowest in small follicles and increased in medium and large follicles. Concentrations did not differ between medium and large follicles. Follicular fluid prolactin concentrations were lower in autumnal follicles compared with summer follicles of comparable size. It is possible that the short-term surge in circulating prolactin around ovulation could be linked to the significant levels of prolactin in follicular fluid. Ovulation releases a relatively large volume of fluid into the peritoneum. The prolactin in this fluid could be a contributor to the periovulatory prolactin surge.     

Exogenous eFSH, follicle coasting, and hCG as a novel superovulation regimen in mares

The objective of this study was to evaluate various equine follicle-stimulating hormone (eFSH) treatment protocols and the effect of “follicle coasting” on ovulation and embryo recovery rates in mares. Cycling mares (n = 40) were randomly assigned to one of four groups 7 days after ovulation: (1) 12.5 mg eFSH twice daily until follicles were 35 mm or larger; (2) 12.5 mg eFSH twice daily until follicles were 32 mm or larger; (3) 12.5 mg eFSH twice daily for 3.5 days followed by 12.5 mg eFSH enriched with luteinizing hormone (LH) twice daily until follicles were 35 mm or larger; and (4) 25 mg eFSH once daily until follicles were 32 mm or larger. Mares in groups 1 and 3 were injected with human chorionic gonadotropin (hCG) (2500 IU intravenously) at the end of eFSH treatment, whereas mares in groups 2 and 4 were given hCG approximately 42 and 54 hours, respectively, after the last eFSH treatment (“follicle coasting”). Nonsurgical embryo collection was performed 6.5 to 7.5 days after ovulation. Each mare experienced a nontreated estrous cycle before being reassigned to a second treatment. Ovulation rates for mares in treatment groups 1 to 4 were 3.3 ± 0.4, 4.1 ± 0.4, 3.5 ± 0.4, and 2.8 ± 0.4 (mean ± SEM; P < .05), respectively. One or more embryos were recovered from more than 80% of mares in each treatment group, and embryo recovery rate per flush was similar among treatment groups (1.9 ± 0.3, 2.6 ± 0.3, 1.9 ± 0.3 and 1.9 ± 0.3, respectively; P > .05). The overall embryo recovery rate was 2.1 ± 1.5 embryos per flush. In summary, ovulation rate was higher for mares treated with eFSH (3.4 ± 0.4) compared with non-treated controls (1.1 ± 0.2). Ovulation rate in mares in which hCG was delayed (follicle coasting) was higher (P < .05) when treatments were given twice per day versus once per day. Administration of equine luteinizing hormone (eLH) in conjunction with eFSH did not have an advantage over mares treated only with eFSH.     

Use of a Mycobacterial Cell Wall Extract (MCWE) in Susceptible Mares to Clear Experimentally Induced Endometritis With Streptococcus zooepidemicus

The ability of an immunomodulator, mycobacterial cell wall extract (MCWE), to clear uterine infection in susceptible mares after an experimental challenge withStreptococcus zooepidemicus was evaluated. Thirty mares susceptible to endometritis, based on the presence of uterine fluid during both diestrus and estrus, were selected from a herd of 896 and inoculated with a live culture of 5 × 10⁶ CFU of S. zooepidemicus on day 1 of estrus. Twenty-four hours later, mares were evaluated by ultrasonography, bacteriology, exfoliative cytology, and uterine biopsy to confirm infection. Forty-eight hours after inoculation, and on confirmation of uterine infection, mares were randomly assigned to one of four unbalanced experimental treatments to receive 1500 μg MCWE IU (n = 10) or IV (n = 10), or placebo IU (n = 5) or IV (n = 5). Mares were examined at ovulation and 7 days post-ovulation for uterine fluid via transrectal ultrasonography and for bacteriology, exfoliative cytology, and uterine biopsy. Efficacy was based on the ability of the mare to clear endometritis as determined by negative bacteriology and reduced numbers of polymorphonuclear cells (PMNs) on uterine biopsy. Because no statistical difference was detected between routes of administration on day 7 post-ovulation, the data sets were combined and re-analyzed to evaluate overall efficacy. Endometritis was observed in all placebo-treated mares 7 days post-ovulation, whereas treatment with MCWE resulted in the elimination of endometritis in 35% of the mares by the time of ovulation, and 70% of the mares by 7 days post-ovulation. Treatment with MCWE, compared with the placebo group, resulted in a significant decrease in the number of mares positive for endometritis at ovulation based on exfoliative cytology and bacteriology (P < .01) and at 7 days post-ovulation based on biopsy, exfoliative cytology, and bacteriology (P < .001). Results indicate that MCWE was an effective treatment for the elimination of endometritis caused by S. zooepidemicus in mares.     

Uterine clearance mechanisms during the early postovulatory period in mares

Uterine response to inoculation with Streptococcus zooepidemicus organisms, 51Cr-labeled 15-microns microspheres, and charcoal was evaluated in 9 mares (4 resistant and 5 susceptible to endometritis) to determine mechanical and cellular clearance rates during the early postovulatory period. Mares were inoculated at estrus prior to ovulation during estrous cycles 1, 3, and 5. Uterine swab specimens for aerobic and anaerobic bacteriologic culture and serum for progesterone determination were obtained on postovulation day 3 during estrous cycle 1, on the day of ovulation during estrous cycle 3, and on postovulation day 5 during estrous cycle 5. Immediately thereafter, the uterus was irrigated with 50 ml of sterile physiologic saline solution containing tracer amounts of 125I-labeled human serum albumin. Streptococcus zooepidemicus was isolated from 10 of 15 (67%) uterine specimens collected from susceptible mares and incubated aerobically. Escherichia coli also was isolated from 2 of the 10 specimens incubated aerobically. Anaerobic bacteriologic culture of specimens from all mares yielded no growth. Chromium-labeled microspheres were recovered twice from 2 susceptible mares, on day 0 and day 5. Charcoal was retained in 5 specimens collected from 3 susceptible mares. Bacteriologic culture of specimens from resistant mares did not yield growth. On day 0, chromium-labeled microspheres and charcoal were recovered once from 1 resistant mare. Mares susceptible to endometritis accumulated more fluid within the uterine lumen after ovulation than did resistant mares (mean +/- SEM, 52.73 +/- 15.22 ml and 7.41 +/- 1.96 ml, respectively; P less than 0.01). From this study, it appeared that uterine cellular and bactericidal mechanisms are dysfunctional during the early postovulatory period. However, there appeared to be no disruption of the mechanisms responsible for mechanical clearance of materials inoculated in the uterus.     

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.     

Stallion-like Behavior in Mares: Review of Incidence, Characteristics, Ovarian Activity, and Role of Testosterone

Stallion-like sexual behavior in mares is rare, except in association with ovarian tumors or hormonal treatments. The rarity of the phenomenon was confirmed in a recent 3-year study. The mean number of mares with detected stallion-like behavior, including mounting with thrusts, during an entire ovulatory season was 5.7 (17/3 years) in a herd averaging 105 mares (5% incidence/mare/season). From a total of 17 mountings of an estrous mare by another mare, 15 occurred when the mounting mare was in the follicular phase and two when in the early luteal phase. Plasma testosterone concentration on the day of mounting was higher (P < 0.01) in the mounting mares (17.7 ± 2.3 pg/ml) than in the standing mares (10.9 ± 0.5 pg/ml). No other deviation in the endocrine, behavioral, or morphologic aspects of the estrous cycle was observed. In another study, testosterone was assayed daily from 7 days before to 4 days after ovulation in seven mares during estrous cycles with no detected mare-on-mare mountings. Concentrations during the follicular phase were highest on the days corresponding to when mare-on-mare mounting was detected in the previous study. It is concluded that the rare occurrence of stallion behavior by untreated mares with no detected ovarian tumors is a consequence of an unusually high, apparently transient fluctuation in circulating testosterone at the time of mounting.     

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.     

Efficacy of the gnrh agonist deslorelin acetate for inducing ovulation in mares relative to age of mare and season

Deslorelin acetate (Ovuplant™, Fort Dodge), a GnRH agonist, is commonly used to induce ovulation in cycling mares. Although its efficacy in hastening ovulation has been previously reported, the effects of age of mare and month of administration on percent of mares responding and interval to ovulation have not been studied.Data was gathered from reproduction records of 376 mares receiving deslorelin acetate at the Equine Reproduction Laboratory, Colorado State University, from 1995 to 1999. Age of mare, date of administration, size of largest follicle at treatment, and interval to ovulation were recorded. Age of mare was categorized into five groups: 2–4, 5–9, 10–14, 15–19, and greater than or equal to 20 years. Date of administration was divided into four groups: March and April, May and June, July and August, and September and October.A higher (p < 0.05) percentage of mares aged 10–14 (98.5%) ovulated in response to deslorelin acetate than mares aged 2–4 or 5–9 (90.2% or 91.0%, respectively) or mares aged 15–19 or ≥ 20 (87.9% or 83.8%, respectively). Mares ≥ 20 had the lowest ovulation rate (83.8%). However, mares ≥ 20 that responded to deslorelin acetate had a shorter (p < 0.05) interval from treatment to ovulation (1.7 ± 0.1 days) than mares 2–4 and 5–9 years of age (1.9 ± 0.1 and 1.9 ± 0.0 days, respectively).Deslorelin acetate was more effective in inducing ovulation in the July and August (95.4%) (p < 0.01) and September and October (95.7%) (p = 0. 04) than in the March and April (81.1%). Mares treated in May through October also experienced shorter (p < 0.05) intervals to ovulation than mares treated in March and April.     

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.     

Effect of Chronic Administration of Oxytocin on Corpus Luteum Function in Cycling Mares

The objective of this study was to determine if intramuscular administration of 60 units of oxytocin once daily for 29 days, regardless of when treatment was initiated during the estrous cycle (i.e., without monitoring estrous behavior and/or detecting ovulation), would induce prolonged corpus luteum (CL) function in cycling mares. Mares were randomly assigned to two groups: (1) saline-treated control (n = 7) and (2) oxytocin-treated (n = 9) subjects. Control mares received 3 cc of saline, and oxytocin-treated mares received 60 units (3 cc) of oxytocin intramuscularly for 29 consecutive days. Treatment was initiated in all mares on the same day (day 1), independent of the day of the cycle. Jugular blood samples for determination of progesterone concentration were collected three times weekly (M, W, and F) for 21 days before treatment was initiated to confirm that all mares had a luteal phase of normal duration immediately before treatment. Beginning on the first day of treatment, blood samples were collected daily for eight days and then three times weekly through day 80. Mares were considered to have prolonged CL function if serum progesterone remained >1.0 ng/mL continuously for at least 25 days after the end of the treatment period. The proportion of mares with prolonged CL function was higher in the oxytocin-treated group than in the saline-treated group (7/9 vs. 1/7, respectively; P < .05). Three of the seven oxytocin-treated mares that developed prolonged CL function initially underwent luteolysis within 4–7 days of the start of oxytocin treatment and then developed prolonged CL function after the subsequent ovulation during the treatment period. In the other four oxytocin-treated mares that developed prolonged CL function, progesterone remained >1.0 ng/mL throughout the treatment period and into the post-treatment period. All mares with prolonged CL function maintained elevated progesterone concentrations through at least day 55 of the study. In conclusion, intramuscular administration of 60 units of oxytocin for 29 consecutive days effectively prolonged CL function in mares, regardless of when treatment was initiated during the estrous cycle. Importantly, this represents a protocol for using oxytocin treatment to prolong CL function that does not require detection of estrous behavior or day of ovulation.     

A field study on early pregnancy loss in standardbred and thoroughbred mares

Early pregnancy loss in the mare is a major cause of infertility and economic loss. To study this important problem, sequential ultrasound examinations were completed on breeding farm mares (n = 404 pregnancies). The incidence of pregnancy loss between Week 2 and Week 8 post ovulation was 42 losses out of 404 pregnancies.(10.4%) More (p<0.05) pregnancies were lost at 2–4 weeks post ovulation than at 4–6 or 6–8 weeks post ovulation (23/42 vs 9/42 or 10/42). The number of days from detection of pregnancy loss until the subsequent ovulation was higher (p<0.01) for mares in which pregnancy loss was detected at 6–8 weeks post ovulation than for mares in which pregnancy loss was detected at 2–4or 4–6 weeks post ovulation (21.1 ± 3.90 days vs 12.7 ± 1.59 or 9.5 ± 1.05 days,respectively). Thirty-one of 45 mares which lost pregnancies were again bred. Sixty-five percent (20/31) of these mares again became detectably pregnant but forty percent (8/20) of thesepregnancies were subsequently lost.     

Ultrasonic evaluation of the reproductive tract of the mare: Ovaries

Ultrasonography is useful for monitoring the dynamic follicular and luteal changes of equine ovaries, since it permits rapid, visual, non-invasive access to the reproductive tract. A 5 MHz transducer has greater resolving power and is far more suitable for evaluation of ovaries than a 3–3.5 MHz transducer. Follicles as small as 2–3 mm can be seen and the corpus luteum can usually be identified throughout its functional life. In a study involving daily ovarian examinations, there was a pronounced change in shape of the preovulatory follicle from a roughly spherical to a pear-shaped or oblong form in 66% of the ovulatory periods, This change usually occurred on the day preceding ovulation. The occurrence of ovulation was detectable by the disappearance of a large follicle. In addition, the ovulation site on day 0 was characterized by an intense echogenic area in 88% of 32 ovulations. The developing corpus luteum retained the echogenicity for a mean of 2.4 days. In a blind study, the location of the corpus luteum, as determined by ultrasound, agreed with a previous independent determination of the side of ovulation by palpation in 88% of the 40 bred mares on days 0–14. In all of the 12 mares that were in estrus, the location of the corpus luteum could not be ascertained. In another study, the corpus luteum was identified for a mean of 16 days in 14 estrous cycles. One or more days before the corpus luteum became ultrasonically unidentifiable, it developed increased echogenicity in 36% of the mares, indicating greater tissue density. It is concluded that ultrasonic evaluation of the corpus luteum is superior to digital evaluation by rectal palpation. Some of the potential applications of ultrasonic examination of the ovaries include: 1) obtaining important, sometimes definitive, information by a single examination for judging whether a mare has entered the ovulatory season, 2) aiding in estimating the stage of the estrous cycle, 3) detecting double preovulatory-sized follicles which are in close apposition and difficult to discern by palpation, 4) detecting failure of ovulation or anovulatory estrus by the absence of a corpus luteum, 5) differentiating a persistent corpus luteum from anovulatory or anestrous conditions, 6) diagnosing certain pathological conditions such as peri-ovarian cysts and ovarian tumors, and 7) diagnosing anovulatory hemorrhagic follicles.     

Effects of a gonadotropin-releasing hormone antagonist and altrenogest on luteinizing hormone concentration and ovulation in the mare

The objective of Experiment 1 was to determine a dose and frequency of gonadotropin-releasing hormone (GnRH) antagonist administration to effectively suppress serum luteinizing hormone (LH) concentration and to delay ovulation when administered to mares. The objectives of Experiment 2 were 1) to determine the effects of subcutaneous or intravenous administration of a GnRH antagonist or oral altrenogest on serum LH concentration in the estrual mare; and 2) to determine the effectiveness of human chorionic gonadotropin (hCG) in inducing ovulation in mares with suppressed LH concentrations. In Experiment 1, mares (N = 20) were randomly assigned and treated with either 5% mannitol (control, single subcutaneous injection, 1 mL, at time 0; n = 5); low-dose GnRH antagonist (single subcutaneous injection, 0.01 mg/kg, at time 0; n = 5); frequent low-dose GnRH antagonist (subcutaneous injections, 0.01 mg/kg, at 0, 6, 18, and 24 hours; n = 5); or high-dose GnRH antagonist (single subcutaneous injection, 0.04 mg/kg, at time 0; n = 5). Both the frequent low-dose and high-dose GnRH antagonist treatments resulted in significantly lower LH concentrations compared with controls at 90, 102, and 114 hours after treatment (P < .05). In Experiment 2, mares (N = 38) were randomly assigned and treated with subcutaneous sterile saline (control), altrenogest (oral), subcutaneous GnRH antagonist, or intravenous GnRH antagonist. LH concentration for the altrenogest group was lower than the control group at 3, 4, 18, and 30 hours after treatment (P < .05). LH concentration for both the subcutaneous and intravenous GnRH antagonist groups were lower compared with the control group at several time points (P < .05). Based on these data, dose but not frequency of administration of a GnRH antagonist lowered LH concentration in the estrous mare but did not delay ovulation. In addition, serum LH concentrations can be lowered and ovulation effectively postponed in mares treated with altrenogest followed by administration of hCG. This indicates that serum LH concentrations can be lowered and ovulation effectively postponed in mares treated with altrenogest followed by administration of hCG.     

Relationship between estrous behavior in pregnant mares and the presence of a female conceptus

Unsolicited reports of estrous behavior in mares thought to be pregnant were received from owners or caretakers of Arabian mares. Estrous behavior was confirmed and mares were examined for pregnancy. Gender of the conceptus was determined at foaling in 11 mares in which estrous behavior was confirmed while an apparently viable, ultrasonically normal-appearing conceptus was present. In 9 mares in which the day of ovulation was known (Day 0), the estrous behavior occurred on Day 12, 13 or 14 (5 mares), Day 18 or 20 (2 mares), Day 40 (1 mare) and Day 60 (1 mare). In another study, 55 pony mares were observed for estrous behavior every 3 days for 20 minutes during Days 11 to 40. Estrous behavior was observed in 1 mare (2%) on Day 24. Combined for the 2 studies, the incidence of a female conceptus (12/12) was greater (P<0.01) than the incidence of a male conceptus (0/12) in mares that exhibited estrous behavior.     

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.     

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.     

Treatment of Persistent Mating‐Induced Endometritis in Mares with the Non‐Steroid Anti‐Inflammatory Drug Vedaprofen

Recently, successful treatment of mares with a history of persistent mating‐induced endometritis (PMIE) with dexamethasone has been reported. As systemic treatment of horses with glucocorticoids should be handled with caution, we tested the hypothesis that treatment with the non‐steroid anti‐inflammatory drug (NSAID) vedaprofen, an inhibitor of cyclooxygenase‐2, may have comparative, positive effects on fertility. Barren mares with a history of repeated PMIE were treated with vedaprofen (n = 8; initially 2 mg/kg bodyweight followed by 1 mg/kg orally twice daily) from 1 day before the first insemination to 1 day after ovulation or left untreated (n = 9). All mares received oxytocin (20 I.E. s.c.) thrice daily. Uterine swabs were collected for bacteriology and cytology. The day after ovulation, fluid accumulation was detected in three control mares and four treated mares (n.s.). The percentage of neutrophils in uterine cytology was significantly increased in comparison to the day before ovulation irrespective of treatment. Pregnancy was confirmed in two of nine mares in the control group and seven of eight mares in the treatment group (p < 0.05). NSAIDs may positively affect fertility in mares with a history of PMIE.