IN-DEPTH: REPRODUCTIVE ENDOCRINOLOGY Hormones and Breeding Carlos R.F. Pinto, MedVet, PhD, Diplomate ACT Author’s address: Theriogenology and Reproductive Medicine, Department of Veterinary Clinical Sciences, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210; e-mail: [email protected]. © 2013 AAEP. 1. Introduction affected by PGF treatment to induce estrus. In The administration of hormones to mares during other words, once luteolysis takes place, whether breeding management is an essential tool for equine induced by PGF treatment or occurring naturally, practitioners. Proper and timely administration of the events that follow (estrus behavior, ovulation specific hormones to broodmares may be targeted to and fertility) are essentially similar or minimally prevent reproductive disorders, to serve as an aid to affected (eg, decreased signs of behavioral estrus). treating reproductive disorders or hormonal imbal- Duration of diestrus and interovulatory intervals ances, and to optimize reproductive efficiency, for are shortened after PGF administration.1 The example, through induction of estrus or ovulation. equine corpus luteum (CL) is responsive to PGF These hormones, when administered exogenously, luteolytic effects any day after ovulation; however, act to control the duration and onset of the different only CL Ͼ5 days are responsive to one bolus injec- stages of the estrous cycle, specifically by affecting tion of PGF.2,3 Luteolysis or antiluteogenesis can duration of luteal function, hastening ovulation es- be reliably achieved in CL Ͻ5 days only if multiple pecially for timed artificial insemination and stimu- PGF treatments are administered. For that rea- lating myometrial activity in mares susceptible to or son, it became a widespread practice to administer showing delayed uterine clearance. In this discus- PGF as a single bolus injection (subcutaneous or sion, we address the effects and potential indications intramuscular) only after 5 to 6 days after ovulation. for the different hormones available to the equine This common practice perhaps contributed to the practitioner working with broodmare reproduction. prevailing assumption that PGF treatments have only significant effects in mares with CL Ͼ5 days. 2. Prostaglandin F2␣ Mares with a CL Ͼ5 days undergo luteolysis when The administration of natural or synthetic prosta- treated with one single injection (subcutaneous or glandin F2␣ (PGF) analogues causes interruption intramuscular) of 5 to 10 mg of dinoprost trometh- of luteal function by luteolysis. The diestrus stage amine or 250 to 500 ␮g of cloprostenol (synthetic of the estrous cycle is then shortened leading to PGF analogue). In the United States, dinoprost onset of estrus. Characteristics of estrus and ovu- tromethaminea is the only PGF drug approved by lation are not different from that occurring in natu- the Food and Drug Administration (FDA) for use in ral cycles. The inherent fertility of mares is not horses, although the use of the racemic formulation NOTES AAEP PROCEEDINGS ր Vol. 59 ր 2013 331 IN-DEPTH: REPRODUCTIVE ENDOCRINOLOGY of d,l-cloprostenolb is very common among practi- thor’s clinical experience that mares that repeatedly tioners working with horse breeding, especially be- ovulate two or three follicles respond to a single cause of its longer half-life than dinoprost hCG treatment similarly to mares ovulating only tromethamine salt preparations—hours versus one follicle during estrus. Because hCG is avail- minutes, respectively. Conversely, in other coun- able in 10,000-IU vials, it is common to have equine tries (Europe, South America, etc), formulations of practitioners administering bolus doses of 2000 or the synthetic analogue cloprostenol are approved 2500 IU to optimize the use of each vial. Although and available for use in horses. In those countries, the author has successfully used vials that have the racemic d,l-cloprostenol and the more potent been reconstituted for several weeks, it is recom- formulation containing only the d-enantiomer clo- mended that reconstituted vials be used within 2 prostenolc are available; only 25 to 37.5 ␮g per mare weeks and kept refrigerated during that period. of d-cloprostenol is needed to induce luteolysis in Intramuscular or intravenous routes can be used to mares at least 5 days after ovulation.4 administer hCG in mares, but it seems, anecdotally, that fewer ovulation failures are seen in mares re- 3. Induction of Ovulation ceiving hCG intravenously than in mares receiving The ability to induce ovulation is one of the most it intramuscularly. For example, intramuscular important strategies used in breeding management. doses of 5000 to 10,000 IU should be avoided. The goal of inducing ovulation in breeding manage- Mares do have development of antibodies to the ment of mares is to induce ovulation within 48 human hormone, but the presence of antibodies does hours, preferably between 24 to 48 hours after treat- not appear to interfere with ovulation.7 Neverthe- ment. In mares monitored closely by palpation less, some authors do recommend not treating mares per rectum and ultrasonography, ovulation occurs with hCG more than once or twice during the same within 36 to 48 hours. This fact is particularly breeding season, whereas others (including this au- important for mares being managed for artificial thor) have used it successfully (ovulation within 48 insemination with frozen-thawed semen. For opti- hours from administration) in mares treated five to mal chances to predict outcome and increase efficacy 10 times during the same breeding season. of the treatment, regardless of the ovulation-induc- ing agent being used, induction of ovulation should 5. Gonadotropin-Releasing Hormones be attempted only in mares found unquestionably to be in estrus and with a growing follicle at least Ն30 Administration of simple gonadotropin-releasing mm in diameter. The most common hormones used hormone (GnRH) analogues (equivalent to the nat- to induce ovulation include human chorionic gonad- ural decapeptide GnRH) to cycling mares has failed otropin (hCG) and the gonadotropin-releasing hor- to consistently induce ovulation. Whereas doses of ␮ mone agonists (GnRH), which are discussed below. 50 to 100 g of GnRH are sufficient to induce ovu- lation in cattle, doses as high as 1 to 4 mg have failed 4. Human Chorionic Gonadotropin to reliably induce ovulation in mares when admin- Human chorionic gonadotropind has been widely istered once or twice daily during estrus. Further- and extensively used as an ovulation-inducing more, only pulsatile administration (every hour) of agent because of its luteinizing hormone (LH)-like GnRH has been found to be effective in inducing activity. The hormone, hCG, is produced after be- ovulation in the mare; this method is, however, not ing extracted from urine of pregnant women. feasible for widespread clinical application. Since Chemically, hCG is a glycoprotein composed of an ␣- the chemical structure and peptide identification and a ␤-subunit; the ␣-subunit is similar to that of was reported in 1971, more than 2000 GnRH ana- human follicle-stimulating hormone (FSH) and LH; logues were synthesized in the following 15 years however, the ␤-subunit, despite containing the same after the 1971 report. Scientists working in re- amino acids present in the LH ␤-subunit, does have search and private pharmaceutical institutions an additional group of 23 amino acids and a high shared two main aims: (1) to create an analogue degree of sialylation that confers hCG a longer half- with high affinity for GnRH receptor binding and (2) life (several hours) than that of LH (20 to 30 min- to avoid or minimize enzymatic degradation by pro- utes). The hCG hormone is then lyophilized and teolysis. The GnRH molecule is susceptible to available in the United States in vials containing cleavage and inactivation by hypophyseal endopep- 10,000 international units (IU). Despite reports of tidases, especially between amino acids in position 5 mares being treated with hCG since 1939,5 hCG is and 6 and between 6 and 7. Another enzyme, car- not approved by the FDA for use in horses. Never- boxypeptidase, targets the bond between amino ac- theless, a single bolus dose of 1500 to 3300 IU is ids in positions 9 and 10. For these reasons, several typically sufficient to induced ovulation in mares GnRH synthetic analogues used in human and vet- during estrus, with distinct estrous uterine edema erinary medicine have (1) D-amino acid substitu- and with a viable, growing follicle Ͼ30 mm.6 tions in position 6 (glycine) and (2) the amino acid in Mares with more than one presumptive preovula- position 10 (another glycine) and its amide terminal tory follicle do not need to receive a double dose or a replaced by an ethylamide residue that is linked to second injection during that estrus. It is the au- the amino acid 9 proline.8 332 2013 ր Vol. 59 ր AAEP PROCEEDINGS IN-DEPTH: REPRODUCTIVE ENDOCRINOLOGY Deslorelin Table 1. Comparative Rank Order Potency of Synthetic GnRH Ana- logues in Relation to Natural GnRH In contrast with the poor or limited efficacy of simple GnRH analogues, more potent GnRH synthetic an- GnRH Buserelin Deslorelin Histrelin alogues such as deslorelin have a predictable effect Relative potency 1 20 144 210 in inducing ovulation within 48 hours when admin- istered as intramuscular injections or by means of subcutaneous implants. Deslorelin differs from the natural decapeptide GnRH in two amino acid sub- stitutions: in amino acid position 6,