Use of deslorelin short-term implants to induce ovulation in cycling mares during three consecutive estrous cycles

ANIMAL REP$XK&IOY Animal ReproductionScience 39 (1995) 129-140

Use of deslorelin short-term implants to induce ovulation in cycling mares during three consecutive estrous cycles E.L. Mumford”, E.L. Squiresa7*, E. J6chleb, L.A. Harrisona, T.M. Nett”, T.E. Trigg” “Animal Reproduction and Biotechnology Laboratory, Department of Physiology, Colorado State University, Fort Collins, CO 80523, USA bWolfgang Jiichle Association, Inc.. Denville. NJ 07834, USA “Peptide Technology, Ltd., Dee Why. N.S. W. 2099, Australia

Accepted26 January 1995

Abstract

Alternatives to human chorionic gonadotropin (hCG) for inducing ovulation in cycling mares over several consecutive cycles were explored. Placebo, one, three or five short-term implants each containing 2.2 mg of gonadotropin-releasing hormone (GnRH) analogue (deslorelin) were administered to cycling mares after identification of a follicle over 30 mm. Mares were treated over three consecutive cycles, and artificially inseminated during the third cycle only. Serum was assayed for concentrations of luteinizing hormone (LH) , follicle-stimulating hormone (FSH), and progesterone during each cycle. All deslorelin doses decreased the interval to ovulation (4.0 days, 2.6 days, 2.4 days and 2.0 days), increased the proportion of mares ovulating within 48 h (20.0%, 83.3%, 73.3% and 85.7%)) and decreased the diameter of the largest follicle at ovulation (45.8 mm, 38.0 mm, 41.0 mm and 37.2 mm) for the placebo, 2.2 mg, 6.6 mg and 11.0 mg groups, respectively (P < 0.05). The interovulatory interval was lengthened in the 11.O mg group compared with all other groups (21.2 days, 21.8 days, 26.4 days and 32.7 days for the placebo, 2.2 mg, 6.6 mg and 11.Omg groups, respectively, P < 0.05). No differences (P > 0.05) were detected in serum concentrations of progesterone or pregnancy rate among the groups. Cycle number had no effect (P > 0.05) on the reproductive parameters or serum concentrations of progesterone. Concentrations of LH were higher in Cycle 3 in the 6.6 mg and 11.O mg groups, and FSH concentrations were decreased in Cycles 2 and 3 in treated mares. Deslorelin was efficacious for inducing ovulation in cycling mares with no diminished activity over three consecutive cycles. However, ovarian suppression and changes in serum concentrations of gonadotropins were noted at higher doses. Keywords:

GnRH analogue; Mares; Ovulation induction

* Correspondingauthor: Tel. 303-491-8373;Fax. 303-491-8419.

SSDIO378-4320(95)01383-O

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1. Introduction The duration of behavioral estrus and diameter of ovarian follicle(s) prior to ovulation vary considerably among normal, cycling mares. Therefore, predicting the time of ovulation is difficult, and mares often must be bred many times unless a hormonal treatment is given to induce ovulation. Human chorionic gonadotropin (hCG) is the only product commercially available for induction of ovulation in the mare. Numerous investigators have reported that ovulation occurs within 48 h of treatment with hCG (Sullivan et al., 1973; Voss et al., 1975; Duchamp et al., 1987; Squires et al., 1988). However, treatment with hCG over several consecutive cycles results in antibody formation (Roser et al., 1979; Duchamp et al., 1987; Wilson et al., 1990) and/or diminished effectiveness (Voss et al., 1975; Wilson et al., 1990). Recently, a short-term implant (STI) releasing 2.2 mg of the gonadotropin-releasing hormone ( GnRH) analogue deslorelin acetate was reported to reliably induce ovulation within 48 h after administration to cycling mares (Klug et al., 1992; Meinert et al., 1992; Meyers et al., 1992; McKinnon et al., 1993; Squires et al., 1994). Fitzgerald et al. ( 1993) demonstrated a temporary cessation of ovarian activity in cycling mares administered high doses of a GnRH analogue (goserelin) over 28 days. Apparently, the effects of multiple treatments and high doses of deslorelin ST1 on ovulatory response, estrus duration, pregnancy rate and serum hormone concentration have not been examined. Therefore, this study was designed to evaluate the effect of placebo, 2.2,6.6, and 11.0 mg of deslorelin STI on: ( 1) ovulatory response, follicular response, estrus duration and serum hormone concentration; (2) pregnancy rate; (3) the above reproductive parameters after deslorelin administration over three consecutive cycles.

2. Materials and methods Forty non-lactating, light-horse mares of mixed breeding were used in this study. Ages ranged from 3 to 20 years. Determination of whether mares were cycling was based on documentation of at least one ovulation during a 3 week acclimatization period. Mares were housed in paddocks, and each received 1.5 kg of a grain mixture and 8.5 kg alfalfa hay daily, and had constant access to water and salt/mineral blocks. Mares were exposed to a stallion daily for detection of estrous behavior, and their ovaries were palpated and scanned ultrasonographically per rectum every 3 days, or daily during behavioral or clinical estrus (presence of endometrial folds and a follicle at least 30 mm in diameter) throughout the study. Upon detection of an ovarian follicle over 30 mm in diameter, each mare was assigned at random to one of four treatment groups of ten mares each, and was injected subcutaneously in the neck with one ( 1 X ), three (3 X ) or five (5 X ) implants, each containing 2.2 mg of deslorelin acetate (Peptide Technology Ltd., Dee Why, N.S.W., Australia) or placebo implants. This protocol was repeated for each mare on three consecutive cycles; mares remained in the same dose group each cycle. During the third estrous cycle, mares were artificially

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inseminated every other day with 500 million progressively motile spermatozoa collected from a single fertile stallion, beginning on the day of treatment and continuing until ovulation (generally one or two inseminations). Mares were inseminated on the day of ovulation if they continued to exhibit behavioral estrus. Mares were evaluated ultrasonographically per rectum on 18 and 50 days after ovulation to determine pregnancy status. Mares not pregnant on day 18 were managed according to the above protocol for a fourth cycle, but no deslorelin was given. Five mares each from the control, 1 X and 3 X groups and six mares from the 5 X group were randomly selected for serum hormone analysis. Blood was collected from these mares via jugular venipuncture twice daily beginning at the onset of behavioral estrus, or upon detection of a follicle at least 30 mm in diameter, and continuing until 1 day after ovulation. Samples were then obtained once every other day from day 2 until day 16 after ovulation. Serum was removed and frozen until assayed for concentrations of luteinizing hormone (LH) (Nett et al., 1975) and follicle-stimulating hormone (FSH) (Nett et al., 1979). Serum collected on days 4,8 and 12 after ovulation was also assayed for concentrations of progesterone (Niswender, 1973). One-way analyses of variance (ANOVA) were used to compare mean interval from treatment to ovulation, day of estrus at treatment, diameter of the largest follicle present at treatment, diameter of the largest follicle present within 24 h before ovulation (follicle diameter at ovulation), duration of estrus, and interovulatory interval among treatment groups in each cycle. Repeated measures analysis of variance was used to compare these parameters among cycles within each treatment group. When ANOVA detected differences at P < 0.05, Tukey’s HSD multiple comparison test was used to compare groups. Two-way ANOVA was used to evaluate groupxcycle interactions. Chi-square analyses among groups were used to compare the proportion of mares ovulating within 48 h after treatment, and the proportion of mares pregnant 50 days after the first insemination, and by the end of the study. Repeated measures analysis of variance was used to compare serum concentrations of LH, FSH and progesterone over time, not only among treatment groups within each cycle but also among cycles within each treatment group. Hormonal concentrations were evaluated after being normalized by day of ovulation (from 2 days before until 16 days after ovulation; day 0 being the day of ovulation) and by day of treatment (from day of treatment until 3 days after treatment; day 0 being the day of treatment). Tukey’s HSD test was used to compare overall serum concentrations of LH and FSH among cycles and among groups. When cycle X time or group X time differences were detected at P < 0.05, LSD was used to compare groups or cycles on a given day.

3. Results There were no effects (P > 0.05) of cycle or cycle X group interactions for mean interval from treatment to ovulation, percentage of mares ovulating within 48 h after treatment, day of estrus at treatment, diameter of the largest follicle present at treatment, diameter of the largest follicle detected prior to ovulation, duration of estrus, or interovulatory interval; therefore, cycles were combined in further analyses. Data from mares ovulating follicles other than the follicle assumed to be dominant at the time of treatment (two, one, one and

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Table 1 Reproductive parameters following treatment with various doses of deslorelin: combined data for three cycles ParameteP

Dose of deslorelin (mg) 0 (placebo)

Interval to ovulation (days) Percent ovulating in48h Day of estrus at treatment Follicle diameter at treatment (mm) Follicle diameter at ovulation (mm) Duration of estrus (days) Inter-ovulatory interval (days)

2.2

6.6

4.0 f 1.7a

2.6 f 1.9b

2.4* 1.5b

20.0 (6/30)a

83.3 (25/30)b

73.3 (22/30)b

1.6f l.la

2.2 f 1.6ab

2.8 k2.7ab

11.0 2.0kOSb 85.7 (24/28)b 4.2 &4.4b

36.9k4.8

35.0*5.9

36.2 f4.7

33.1 k2.6

45.8 f5.0a

38.0f5.0bc

41.0&6.2b

37.2 f 3.8~

5.9f2.3

6.1 f2.4

5.9rt3.2

21.2f3.4a

21.8f4.6a

26.4 f 3.5a

7.6f5.5 32.7 f 10.4b

“Proportion, or mean f standard deviation. Values in a row followed by different letters differ at P < 0.05.

two mares from the placebo, 1 X ,3 X and 5 X groups, respectively) were not included in analysis of interval to ovulation, estrus duration, or interovulatory interval for the associated cycle. The interval to ovulation after deslorelin injection was shorter (P < 0.05) for all deslorelin-treated groups than for controls (Table 1) . In addition, a greater (P < 0.05) percentage of deslorelin-treated mares ovulated within 48 h compared with controls. The response of mares for these two parameters was similar (P > 0.05) among the three doses of deslorelin. The number of days in estrus prior to treatment (day of estrus at treatment) was greater (P < 0.05) for mares given five implants than for controls. Although the diameter of the largest follicle at treatment was similar (P > 0.05) among groups, its diameter at ovulation was less for treated mares than for controls, and mares in the 5 X group ovulated smaller follicles than mares in the 3 X group. Although not critically examined, mares given 6.6 or 11 mg of deslorelin were subjectively noted to have diminished follicular activity for 6-10 days after ovulation. Mares in the 5 X group had longer interovulatory intervals than mares in all other groups (P < 0.05). The number of interovulatory intervals at least 25 days in duration was 17/18, 16118, 6118 and 2118 for the placebo, 2.2 mg, 6.6 mg and 11 mg groups, respectively. Two mares in the 5 X group never met tbe criteria for a third injection, and one mare in the 3 X group did not ovulate after the third injection. There was no effect of treatment on pregnancy rate after tbe first (treated) or second (untreated) breeding cycle (Table 2). Serum concentration of progesterone did not vary by group or time X group for any cycle, nor did it vary by cycle or time X cycle for any treatment group (P > 0.05). The intra-assay coefficient of variation for progesterone was 11.7% and the radioimmunoassay (RIA) sensitivity was 23.4 pg ml- ‘.

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Table 2 Percentage of mares pregnant after treatment with deslorelin for three cycles Deslorelin dose (mg)

Percent pregnant at 50 days Cycle 3

Final

0 2.2 6.6 11.0

80.0 (8/10) 80.0 (8/10) 77.8 (719) 87.5 (7/8)

80.0 (8/10) 80.0 (8/10) 88.9 (8/9) 100.0 (8/8)

Serum concentrations of LH were elevated in all deslorelin-treated groups at 12 h compared with controls (Fig. 1; P < 0.05). Mares in the 3 X group had higher concentrations of LH (P < 0.05) than all other groups at 12 h after treatment in Cycle 2 (69.5 ng, 242.8 ng, 393.5 ng and 247.3 ng ml- ’ for the placebo, 1 X , 3 X and 5 X groups, respectively) and Cycle 3 (50.9 ng, 396.3 ng, 662.5 ng and 459.5 ng ml- ’for the placebo, 1 X , 3 X and 5 X groups, respectively). LH concentration continued to be elevated (P < 0.05) over control values in the 5 X group at 24 h (Cycle 1,43.9 ng, 167.7 ng, 148.2 ng and 208.9 ng ml- ’ for the placebo, 1 X , 3 X and 5 X groups, respectively), in the 3 X and 5 X groups at 24 h (Cycle 2, 132.3 ng, 110.0 ng, 157.7 ng and 184.1 ng ml- ’for the placebo, 1 x ,3 x and 5 X groups, respectively) and in the 3 X and 5 X groups (Cycle 3) at both 24 h (61.2 ng, 153.7 ng, 423.7 ng and 317.4 ng ml-’ for the placebo, 1 X, 3 X and 5 X groups, respectively) and 36 h (83.5 ng, 174.0 ng, 383.7 ng and 368.3 ng ml-’ for the placebo, 1 X ,3 X and 5 X groups, respectively). During Cycle 3, LH was also higher (P < 0.05) in

I

20

40 60 Hours

80

Fig. 1. Mean LH (ng ml- ’) among groups. Pooled standard deviation 5 1.O,64.0 and 104.8 for Cycles I,2 and 3, respectively. Hour 0, time of treatment. Asterisks indicate differences among groups (P< 0.05).

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20

40

60

80

Hours Fig. 2. Mean FSH (ng ml-‘) among groups. Pooled standard deviation 18.0, 12.8 and 6.5 for Cycles 1.2 and 3, respectively. Hour 0, time of treatment. Asterisks indicate differences among groups (P< 0.05).

3 X than 1 X mares at 24 h and in both 3 X and 5 X mares than 1 X mares at 36 h. The overall LH response was similar among treatment groups during Cycles 1 and 2, but overall concentrations of LH were higher (P < 0.05) in 3 X mares compared with all other groups during Cycle 3. The intra- and inter-assay coefficients of variation for LH were 8.4% and 7.4%, respectively. The RIA sensitivity was 4.6 ng ml-‘. In all cycles, serum concentrations of FSH were elevated (PO.O5)among Cycles 1, 2 and 3. In contrast, mares in the 3 X and 5 X groups had higher (P< 0.05) serum concentrations of LH after the third injection compared with the first or second injection. There was a significant (P< 0.05) time X cycle interaction for serum concentration of LH in the 5 X group (Fig. 5). The concentration of LH in the 5 X

E.L. Mumford et al. /Animal Reproduction Science 39 (1995) 129-140

-2

0

2

4

6

6

10

12

14

135

16

Days Fig. 3. Mean LH (ng rnt-‘) among groups during Cycle 2. All three cycles showed similar LH patterns relative to ovulation. Pooled standard deviation 72.4. Day 0, day of ovulation. Asterisks indicate differences among groups (P
'"-2

0

2

4

6

6

10

12

14

16

Days Fig. 4. Mean FSH (ng ml-‘) among groups during Cycle 2. All three cycles showedsimilarFSH patterns relative to ovulation. Pooled standard deviation 12.3. Day 0, day of ovulation. Asterisks indicate differences among groups (P ‘<0.05).

I

I

20

I

40

I

60

I

60

Hours Fig. 5. Mean LH (ng ml-‘) among cycles in 5 X mares. Hour 0, time of treatment. Pooled standard deviation 62.7. Asterisks indicate differences among cycles (P< 0.05).

serum concentration of LH in the 5 X group (Fig. 5). The concentration of LH in the 5 X group was higher (P < 0.05) in Cycle 3 than in Cycles 1 and 2 at 12 h (239.9 ng, 247.3 ng and 459.5 ng ml-’ for Cycles 1, 2 and 3, respectively) and 36 h (154.3 ng, 190.0 ng and 368.3 ng ml- ’ for Cycles 1, 2 and 3, respectively) after treatment, and was higher than in Cycle 1 at 48 h after treatment (123.3 ng, 184.1 ng and 263.0 ng ml-’ for Cycles 1,2 and 3, respectively).

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” E F ;

100 60

t? 20 I

1

20

,

,

40

I

60

I

80

Hours Fig. 6. Mean FSH (ng ml-‘) among cycles in 1 X mares. Hour 0, time of treatment. Pooled standard deviation 18.2. Asterisks indicate differences among cycles (P < 0.05).

Serum concentrations of FSH differed among cycles for the 1 X 3 X and 5 X groups (Fig. 6; P < 0.05). In each deslorelin-treated group, concentrations of FSH were higher during Cycle 1 than Cycles 2 and 3. There was a significant (P<0.05)time X cycle interaction for serum concentrations of FSH in the 1 X and 3 X groups. Concentrations of FSH in the 1 X and 3 X groups were higher in Cycle 1 than in Cycles 2 and 3 at 12 h after treatment ( 136.3 ng, 75.8 ng and 67.8 ng ml-’ for Cycles 1,2 and 3, respectively; P <0.05). ,

The interval to ovulation and percentage of mares ovulating within 48 h after injection of 2.2 mg deslorelin were similar to those reported previously (Klug et al., 1992; Meinert et al., 1992; Meyers et al., 1992; Squires et al., 1994; interval to ovulation ranging from 36 to 49 h, the percentage of mares ovulating within 48 h from 88 to 100%). In the present study, the proportion of mares ovulating within 48 h was not altered by administration of doses greater than 2.2 mg. In previous studies, the effect of repeated deslorelin treatment was not critically determined (McKinnon et al., 1993; Squires et al., 1994). We found no adverse effect of three consecutive treatments with a 2.2, 6.6, or 11 mg dose of deslorelin on the proportion of mares ovulating within 48 h or on interval to ovulation, since these parameters were similar among treatment cycles. The reduction in follicle diameter at ovulation for treated mares was similar to that reported previously for mares treated with deslorelin (McKinnon et al., 1993) or buserelin (Harrison et al., 1991)) and was not dose-dependent. Because ovulation could have occurred up to 24 h after follicle sizes were recorded, however, this parameter contains a large inherent error. The 11 mg dose of deslorelin resulted in treatment later in estrus after the first cycle (P = 0.050). During Cycle 1, the mean day of estrus at treatment was 1.8 compared with 4.3 and 7.0 during Cycles 2 and 3, respectively (data per cycle not shown). This could have resulted from suppressed follicular development and/or delayed recruitment of dominant follicles, initiated either by a delay in luteolysis or by a lack of circulating FSH, and is consistent with our subjective finding of diminished follicular activity for 6-10 days after

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ovulation in 3 X and 5 X mares. Additionally, mares given 11 mg of deslorelin had a longer mean interovulatory interval than other groups. A similar trend was noted in mares administered 6.6 mg of deslorelin; the longest interval in this dose group was 33 days. Two mares in the 11 mg group never met the criteria for treatment (follicle over 30 mm, and behavioral estrus or endometrial folds) after the second cycle, and were not treated a third time. The ovaries of one mare became inactive (follicles less than 20 mm), and the ovaries of the other had only limited follicular activity. One mare from the 3 X group did not ovulate after treatment in the third cycle; the dominant follicle appeared to have luteinized in situ. Unfortunately, none of these mares had been selected for analysis of serum hormone concentrations, and therefore, the mechanism for reduced follicular activity was not determined. Fitzgerald et al. ( 1993) reported temporary suppression of ovarian activity in most mares treated with implants delivering high doses of GnRH analogue over a 28 day period during the breeding season. This was often associated with elevated serum concentrations of progesterone suggesting a persistent corpus luteum. In the present study, administration of 2.2, 6.6, or 11 mg of deslorelin during es&us did not affect the function of the resulting corpus luteum. Both serum concentrations of progesterone 4,8 and 12 days after ovulation, and pregnancy rates were similar among groups. Others have reported no effect of treatment with GnRH or GnRH analogues on diestrous concentrations of progesterone (Johnson, 1986; Squires et al., 1988; Squires et al., 1994) and on pregnancy rates (Meyers et al., 1992; Squires et al., 1994). A normal pre-ovulatory pattern of endogenous LH was observed in mares treated with placebo, with the peak concentration occurring 1 day after ovulation. In contrast, LH in deslorelin-treated mares peaked 12-24 h after treatment (24-36 h prior to ovulation), as has been reported previously (Meinert et al., 1992; McKinnon et al., 1993; Squires et al., 1994). There was no additional LH peak associated with ovulation. No clear dose response was detected in serum concentrations of LH. At several time points the concentration of LH was higher in mares given 6.6 mg deslorelin than mares given the 11 mg dose. Despite the differences in LH patterns between placebo and GnRHtreated mares for all cycles, overall concentrations of LH during the 3 day post-treatment period only differed between the control and 3 X groups during Cycle 3. This suggested that the total pituitary release of LH was similar among all deslorelin-treated and control mares. Likewise, other studies (Johnson, 1986; Becker and Johnson, 1992) reported no overall differences in LH released between mares receiving pulsatile GnRH and untreated mares during the peri-ovulatory period. Concentrations of FSH peaked 12 h after GnRH treatment and returned to baseline within 36 h, as in previous reports (Meinert et al., 1992; Squires et al., 1994). The peak concentrations of FSH were inversely related to dose, with the 1 X mares having the highest

concentration of FSH at 12 h post-treatment in all three cycles. The elevated concentration of FSH noted in placebo-treated mares during early die&us was not detected in deslorelintreated mares. Concentrations of FSH were suppressed throughout diestrus in mares treated with 6.6 or 11 mg of deslorelin (Fig. 4), but increased late in diestrus in mares given 2.2 mg. This may explain the diminished ovarian activity and extended interovulatory intervals in mares receiving 6.6 or 11 mg of deslorelin. Continuous infusions of 30-50 pg day-’ of buserelin consistently suppressed FSH secretion in stallions (Boyle et al., 1991). Perhaps the higher doses of deslorelin in the current study promoted inhibin and/or estradiol secre-

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tion or suppressed activin secretion, thereby diminishing FSH synthesis and/or secretion. Increased circulating concentrations of estradiol have been shown to suppress concentrations of FSH (Silvia et al., 1987) as well as folliculogenesis (Burns and Douglas, 1981) in the mare. Serum concentrations of LH and FSH were affected by cycle. In mares administered 6.6 or 11 mg of deslorelin, post-treatment concentrations of LH increased in Cycle 3, suggesting a priming effect at the level of the pituitary of prior deslorelin treatment at the higher doses. Conversely, peak concentrations of FSH decreased after Cycle 1 in all deslorelin-treated groups. As with the decreased circulating FSH in early diestrus described above, decreases in secreted FSH over cycles could be due to effects of consecutive treatments with deslorelin on secretion of estradiol, inhibin and/or activin. Alternatively, deslorelin may have differential effects on LH and FSH secretion directly. In another study (E.L. Squires and E.L. Mumford, unpublished data), 10 cycling mares were implanted with ten deslorelin implants for a total dose of 22 mg deslorelin, or with placebo implants. All mares were killed and necropsied 15 days after treatment. In deslorelin-treated mares, six of the ten ovaries had become inactive and showed no visible or palpable structures. Of the four remaining ovaries, a single follicle was detected in one, a corpus albicans in another, and the third and fourth ovaries each had one corpus luteum. Placebo-treated mares showed only one inactive ovary and a total of 25 detectable follicles involving eight ovaries. These differences were significant (P < 0.05). At this dose, suppression of ovarian activity seems to have occurred quickly. Because GnRH and its analogues work primarily at the level of the pituitary, the potential for down-regulation of pituitary GnRH receptors, diminished serum concentrations of gonadotropins, and lack of ovarian activity exists if high doses of deslorelin are used. This response has been well-documented in other species (Nett et al., 1981; Mahler et al., 1991). Based on the results of this study, high doses of GnRH given over three consecutive cycles suppressed FSH but increased the circulating concentration of LH. It is not known if this was a direct effect of deslorelin on regulation of LH and FSH synthesis and/or secretion, or an effect of deslorelin-induced promotion or suppression of an intermediate product. Suppression of ovarian activity at higher doses of deslorelin, exemplified by the three aberrant mares in Cycle 3 and by extended interovulatory intervals, could be a result of low diestrous concentrations of FSH, high LH, or both. Despite these perturbances, interval from treatment to ovulation and cyclicity were not affected in most mares.

5. Conclusion

Administration of deslorelin acetate as a slow-release implant consistently induced cycling mares to ovulate within 48 h, and this effect was not diminished when mares were treated over three consecutive cycles. We found no added benefit of increased dose on induction of ovulation of pre-ovulatory follicles. We therefore conclude that use of deslorelin 2.2 mg implants is a safe, effective alternative to hCG, especially when administered over multiple cycles.

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Acknowledgments Funded in part by Peptide Technology, Australia.

Ltd., P.O. Box 444, Dee Why, N.S.W. 2099,

References Becker, S.E. and Johnson, A.L., 1992. Effects of gonadotropin-releasing hormone infused in a pulsatile or continuous fashion on serum gonadotropin concentrations and ovulation in the mare. J. Anim. Sci., 70: 120% 1215. Boyle, M.S., Skidmore, J., Zhang, J. and Cox, J.E., 1991. The effects of continuous treatment of stallions with high levels of a potent GnRH analogue. J. Reprod. Fettil. Suppl., 44: 169-182. Burns, P.J. and Douglas, R.H., 1981. Effects of daily administration of estradiol-17P on follicular growth, ovulation, and plasma hormones in mares. Biol. Reprod., 24: 10261031. Duchamp, G., Bour, B., Combamous, Y. and Palmer, E., 1987. Alternative solutions to hCG induction of ovulation in the mare. J. Reprod. Fertil. Suppl., 35: 221-228. Fitzgerald, B.P., Peterson, K.D. and Silvia, P.J., 1993. Effect of constant administration ofagonadotropin-releasing hormone agonist on reproductive activity in mares: Preliminary evidence on suppression of ovulation during the breeding season. Am. J. Vet. Res., 54: 1746-1751. Harrison, L.A., Squires, E.L. and McKinnon, A.O., 1991. Comparison of hCG, buserelin, and luprostiol for induction of ovulation in cycling mares. Equine Vet. Sci., 11: 163. Johnson, A.L., 1986. Pulsatile administration of gonadotropin-releasing hormone advances ovulation in cycling mares. Biol. Reprod., 35: 1123-l 130. Klug, E., Meinert, K., Lepel, J.V., Erbsloh, J., Biet, K., Lubekke, M., Bader, H., Me&, H. and Jochle, W., 1992. Acceleration of ovulation in the mare with a short-acting, subcutaneous implant of the GnRH analog deslotelin. Proc. 12th Int. Congress on Animal Reproduction and AI, The Hague, Vol. 4, pp. 1879-188 1. Mahler, C., Verhelst, J., Chaban, M. and Denis, L., 1991. Prolactin and pituitary gonadotropin values and responses to acute luteinizing hormone-releasing hormone (LHRH) challenge in patients having long-term treatment with depot LHRH analogue. Cancer, 67: 557-559. McKinnon, A.O., Nobelius, A.M., Tarrida de1 Marmol Figueroa, S., Skidmore, J., Vasey, J.R. and Trigg, T.E., 1993. Predictable ovulation in mares treated with an implant of the GnRH analogue deslorelin. Equine Vet. J., 25: 321-323. Meinert, C., Silvia, J.F.S., Kroetz, I., Klug, E., Trigg, T.E., Hoppen, H.O. and Jijchle, W., 1992. Advancing the time of ovulation in the mare with a short-term implant releasing the GnRH analogue deslorelin. Equine Vet. J., 25: 65-68. Meyers, P.J., JBchle, W., Samper, J.C., Sealey, L., Conboy, H.S., Squires, E., Reid, M.P., Bowman, R.T., Taylor, B.C., Gimenez, T., Schlager, R.L. and Dimmick, M.A., 1992. Efficacy of the GnRH analog, deslorelin, given as a short-acting subcutaneous implant, in accelerating ovulation in the mare. Proc. 12th Int. Congress on Animal Reproduction and AI, The Hague, Vol. 4, pp. 1897-1899. Nett, T.M., Holtan, D.W. and Estergreen, V.L., 1975. Levels of luteinizing hormone, prolactin and oestrogens in serum of postpartum mares. J. Reprod. Fertil. Suppl., 23: 201-206. Nett, T.M., Pickett, B.W. and Squires, E.L., 1979. Effects of Equimate (ICI-81008) on levels of luteinizing hormone, follicle stimulating hormone, and progesterone during the estrous cycle of the mare. J. Anim. Sci., 48: 69-75. Nett, T.M., Crowder, M.E., Moss, G.E. and Duello, T.M., 1981. GnRH-receptor interaction v. down-regulation of pituitary receptors for GnRH in ovariectomized ewes by infusion of homologous hormone. Biol. Reprod., 24: 1145-l 155. Niswender, G.D., 1973. Influence of the site of conjugation on the specificity of antibodies to progesterone. Steroids, 22: 413424. Roser, J.F., Kiefer, B.L., Evans, J.W., Neely, D.P. and Pacheco, C.A., 1979. The development of antibodies to human chorionic gonadotrophin following its repeated injection in the cyclic mare. J. Reprod. Fertil. Suppl., 27: 170-173.

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Sullivan, J.J., Parker, W.G. and Larson, L.L., 1973. Duration of estrus and ovulation time in nonlactating mares given human chorionic gonadotropin during three successiveestrous periods. J. Am. Vet. Med. Assoc., 162: 895-898. Voss, J.L., Sullivan, J.J., Pickett, B.W., Parker, W.G., Burwash, L.D. and Larson, L.L., 1975. The effect of hCG on duration of oestrus, ovulation time and fertility in mares. J. Reprod. Fertil. Suppl., 23: 297-301. Wilson, C.G., Downie, CR., Hughes, J.P. and Roser, J.F., 1990. Effects of repeated hCG injection on reproductive efficiency in mares. J. Equine Vet. Sci., 10: 301-308.