Pharmacological control of reproduction in mares requires the use of equine gonadotrophins to avoid induced immunological resistance. Crude equine gonadotrophins (CEG) have been used but the presence of equine luteinizing hormone (eLH) and follicle-stimulating hormone (eFSH) in CEG has led to disappointing results in superovulation studies. Separation of eLH and eFSH activities from CEG is necessary to overcome this problem. The hydrophobic properties of the two hormones were sufficiently different to permit their separation by hydrophobic interaction chromatography (HIC) on a phenyl Sepharose matrix. Good yields of separate FSH and LH fractions were readily obtained by stepwise elution and the method was adapted for large scale preparations of enriched fractions of eLH and eFSH. Two experiments were performed in vivo to evaluate the biological activity of the HIC fractions. Experiment 1 showed that biological activity of the LH fraction in inducing ovulation of preovulatory follicles was similar to that obtained with CEG, indicating that LH bioactivity was not altered by HIC. Experiment 2 demonstrated that biological activity of the FSH fraction was identical (as far as rate of ovulation was concerned) to that of CEG in superovulating mares, indicating that FSH activity was also not altered by HIC. Although we have not obtained better results with the separate equine gonadotrophins than with CEG, it is potentially advantageous to use preparations with single activity to obtain a controlled balance of FSH and LH activity. The HIC technique was chosen because it could easily be scaled up to provide the large amounts of the separate hormones needed for the treatment of a large number of mares.

The pharmacological control of reproduction in the mare requires the use of equine gonadotrophins to avoid induced immunological resistance. Crude equine gonadotrophins (CEG) have been used for several years but the presence of both equine luteinizing hormone (eLH) and follicle-stimulating hormone (eFSH) in CEG has so far led to disappointing results in superovulation. In order to overcome this problem, the separation of eLH and eFSH activities from CEG appeared necessary. The hydrophobic properties of the 2 hormones were different enough to permit their efficient separation by hydrophobic interaction chromatography (HIC) on a phenyl sepharose column. Separate FSH and LH fractions were readily obtained in good yield by stepwise elution and the method can be easily adapted for large scale preparations of enriched fractions of eLH and eFSH. Three in vivo experiments were performed to evaluate the biological activity of fractions obtained by HIC. Experiment 1 showed that the biological activity of the LH fraction in inducing ovulation of preovulatory follicles was similar to that obtained with crude CEG on the basis of their respective immunological LH activities. This indicates that the LH bioactivity was not altered by HIC. Similarly, experiment 2 demonstrated that the biological activity of the FSH fraction was identical to that of CEG to superovulate mares, indicating that FSH activity was not altered by HIC. Finally, experiment 3 revealed that ovarian responses with FSH were more homogeneous among animals and ovulations more synchronous than with CEG.

Cyclic mares were assigned to 1 of 3 treatments (n=15 per group): Group 1 received equine pituitary extract (EPE; 25 mg, i.m.) on Day 5 after ovulation; Group 2 received EPE on Day 12 after ovulation; while Group 3 received 3.3 mg of GnRH analogue (buserelin implant) on the day of ovulation and 25 mg, i.m. EPE on Day 12. Mares in each group were given 10 mg PGF2alpha on the first and second day of EPE treatment. The EPE treatment was continued daily until the first spontaneous ovulation, at which time 3,300 IU of human chorionic gonadotropin (hCG) were given to induce further ovulations. Mares in estrus with a >or=35 mm follicle were inseminated every other day with pooled semen from 2 stallions. Embryo recovery was attempted 7 days after the last ovulation. Follicular changes and embryo recovery during 15 estrous cycles prior to treatment were used as control data. During treatment, the number of follicles>or=25 mm was higher (P<0.05) for Day 5 than for Day 12 or control mares, but the number for Day-5 mares was similar (P>0.05) to that of mares treated with buserelin implants (Group 3). Initiation of EPE treatment on Day 5 resulted in a greater (P<0.05) number of ovulation (2.9) than on Day 12 (1.1) or in the control mares (1.3) but not in the buserelin-treated mares (1.8). The number of embryos recovered from mares in the Day 5 (1.2), Day 12 (1.0), buserelin (0.9) and control (0.9) groups was similar (P>0.05). The conclusions were 1) EPE initiated in early diestrus increased follicular development and ovulation and 2) treatment with GnRH analogue marginally improved response to EPE treatment.

Twelve anoestrous mares were treated with an intravaginal sponge containing 0.5 g allyl trembolone (Regumate; Roussel UCLAF, Paris) and 50 mg oestradiol benzoate for 7 days, followed by daily intramuscular (i.m.) injections of 25 mg crude equine pituitary extract (CEG), with (n = 6) or without (n = 6) 0.25 mg porcine growth hormone (pGH). No difference in ovarian response to this superovulation treatment was observed between the 2 groups (2.2 +/- 0.4 vs 2.3 +/- 0.4 ovulations per mare, respectively). CEG treatment was then combined with allyl trembolone (40 mg per os per day) and prolonged in the 12 mares, with (n = 6) or without (n = 6) a 7-day interruption of CEG treatment 7 days after the mares first ovulated. Ovarian responses showed that an interruption in gonadotrophin treatment was not necessary to allow the occurrence of a new follicular wave, but the interruption did seem to generate the growth of a more synchronous cohort of follicles. Ovulation rate in the second wave was higher than in the first although there was no effect of treatment interruption on ovulation rate in the second wave, which occurred during progestagen treatment and gave rise to persistent corpora lutea (36.5 +/- 5.9 days). The 6 mares that did not experience an interruption in treatment continued to receive daily CEG injections and oral progestagen. Two periods (Days 0-47 and Days 48-94) were compared for analysis of follicular growth.(ABSTRACT TRUNCATED AT 250 WORDS)