Breeding management in the mare (Proceedings)

Introduction

With some minor breed differences, most mares have similar reproductive physiology and, from that respect, may be expected to respond similarly to a canned breeding management protocol. When designing breeding management protocols for a particular mare or stud farm, you must take into account many external factors. These should include the type of semen or breeding used (live cover, fresh AI, chilled/shipped AI, or frozen), when the stallion (or semen) is available, if the mare is to hold the pregnancy or be an embryo donor, if the mare is to be bred on foal heat, your availability to that farm (Do you visit every day? Do you work weekends?), and perhaps other factors unique to that particular farm. All of these variables will influence your decisions about when to schedule follow-up examinations, when to give certain drugs, when to inseminate, etc. This lecture will focus mostly on the normal physiological changes in the mare reproductive cycle and how we can manipulate the cycle. It is up to you to figure out how to effectively use those tools to manipulate the cycle around whatever unique parameters surround your particular farms. No matter what protocols you develop, the end goal is the same: foals on the ground the following year.

Estrous behavior

The earliest form of breeding management in horses was simply to tease the mare to the stallion and allow them to breed periodically while the mare showed signs of being in heat. Even with the technical advances of our day, including ultrasound and hormonal manipulation, the mare's natural reproductive behaviors still play an important role in determining if and when to breed her. Ultrasound allows us to monitor uterine edema, a very useful tool in monitoring the mare cycle, but inflammation can also cause edema. Follicles generally reach a certain size before they ovulate, but diestral follicles can reach those large sizes and then regress instead of ovulating. The cervix generally softens during estrus, but maiden mares will often have a tight cervix throughout their cycle. With all of these variables, it is often useful to be able to take a mare sending mixed signals through our routine reproductive examination and expose her to a stallion. Whether she is showing behavioral signs of estrus ("teasing in") or not ("teasing out") can be the deciding factor as to how you should proceed with her protocol.

Behavioral signs of estrus in the mare classically include raising the tail, squatting, everting the clitoris ("winking"), and urinating. The ears are alert and generally forward. The mare may squeal, playfully kick out (not to strike), or turn to push her hind end towards the stallion. A mare in diestrus will generally clamp her tail down tight over her vulva, keep her ears pinned back, squeal, and kick out to strike. The mare will be reluctant to approach the stallion.

The most important hormone regulating estrous behavior in the mare is progesterone. Progesterone is produced by the corpus luteum and exerts a strong inhibitory effect on estrous behavior, making diestrous mares strongly averse to the advances of the stallion. Estrogens (high during estrus, baseline during diestrus) do enhance estrous behavior, but are often not necessary for the mare to show signs of estrus. When the corpus luteum regresses at the end of diestrus and becomes inactive, no longer secreting progesterone, the mere absence of the inhibitory effects of that hormone are often enough to cause the mare to become receptive to the stallion. This is evidenced by the behavior of some ovariectomized mares, who will continually show signs of estrus, despite the absence of both progesterone and estrogens. Not all mares are this sensitive to the effects of progesterone, and some do require supplementary estrogens to show signs of being in heat. Another clinical application of these hormonal interactions is that mares will usually tease to the stallion for at least 24 hours after ovulation. Estrogen drops precipitously up to 24 hours prior to ovulation, but progesterone does not climb to levels significant enough to suppress estrous behavior for 1-2 days post ovulation. Just because a mare is still teasing to the stallion does not, therefore, mean that she still has a dominant follicle, and since the oocyte is only fertile for about 6-8 hours post-ovulation, it does not mean the mare is still in the fertile part of her cycle.

In a program where the mares are teased regularly to a stallion, the number of days that a mare has been teasing can be helpful information. Early in the breeding season, estrus may last up to 7 days. At the height of the breeding season, the estrus period shortens considerably and may only last a few days.

Transrectal palpation

The most basic and core reproductive veterinary skill in large animals is transrectal palpation. Noting the tone of the cervix will give valuable information that is not assessable via ultrasound. The cervix of the mare changes dramatically between diestrus and estrus due to the effects of progesterone and estrogen. In diestrus and pregnancy, the cervix is long, narrow, and has good tone. In estrus, the cervix is short, wide, and flaccid. Exceptions to these changes may be in maiden mares, who tend to maintain higher tone in the cervix even during estrus, and mares experiencing inflammation, who tend to have lower tone even during diestrus or pregnancy. In later stages of pregnancy, the tone of the cervix usually relaxes a little, in comparison to early pregnancy. Mares that have experienced a significant degree of chronic inflammation or cervical trauma may develop adhesions that will interfere with the usual cervical dynamic shifts in tone and give a false impression of perpetual tone.

Opposite all other domestic species, the mare's ovary has an external medulla and internal cortex. In the mare, ovulation happens at the ovulation fossa on the inner curvature of the ovary. Because ovulation happens internally, corpora lutea are not palpable in the mare. Instead, a misshapen ovary is usually indicative of a growing follicle. Follicles can grow quite large prior to ovulating (40-60 mm in diameter) and greatly distort the shape of the ovary. Not only does the size of the ovary change with a growing follicle, but the texture will change, as well. As a follicle approaches the ovulatory stage, it will soften considerably.

Transrectal Ultrasonography

Ultrasonography has revolutionized the reproductive examination of the mare. It enables us to more accurately follow the reproductive cycle, diagnose pathological changes, and detect twins at a stage we can reduce to a singleton pregnancy with great success. When evaluating the uterus of the mare, it is important to methodically scan the entire tract. This consistent, methodical approach will ensure that you do not miss any pathological changes, nor a pregnancy, nor a twin.

During estrus, the uterus develops a marked amount of edema due to the vasoconstrictive effects of estrogen on the uterine vasculature. This edema is easily assessed via transrectal ultrasound. Grading scales may differ between institutions. A common method is to grade the edema on a scale of 0 to 3, with 3 being maximal edema noted at the height of estrus. If this is the scale used, recognize that greater than a grade of 3 may exist with inflammatory changes. Monitoring uterine edema is a very useful tool in breeding management. A growing, dominant follicle will secrete estrogen enough to cause edema to grow over the course of a couple days from no edema to a grade 3. During the LH surge, and about 24 hours prior to actual ovulation, estrogen production from the follicle drops precipitously and the accompanying edema will also drop. Noting that a particular mare had a large amount of edema during a previous examination, but currently has much less, with the continued presence of a large follicle, is a strong clue that she is very likely to ovulate within the next 24 hours. It would be unwise, therefore, with such a mare, to schedule an insemination for the following day if it could be done on the day of examination instead.

Uterine cysts are a common finding during transrectal ultrasonographic examination of the uterus. Drawing a map of the cysts in the medical record of that mare will serve as a reference when performing the 14 day pregnancy examination. If any doubt exists as to whether a structure is a cyst or a vesicle, simply recheck the mare 1-2 days later. A cyst will not have changed in size, shape, or position.

Ovaries change dramatically through the estrous cycle. Follicular waves are constant throughout the cycle, and may consist of multiple small follicles (<10 mm in diameter), multiple medium follicles (10-30 mm), or a dominant follicle (usually >30 mm) with some combination of smaller accompanying follicles. During diestrus, these larger follicles will usually regress. During estrus, they continue to grow until somewhere between 35 and 60 mm when they will ovulate. After ovulation, the follicle will often fill temporarily with blood before luteinization. This structure is called a corpus hemorrhagicum (CH) and is typified by a hyperechoic border with a mottled center of mixed echogenicity. The CH is not always noted, nor does it always form. Often the only change noted after ovulation is the absence of the large follicle that had been present during the previous examination. Follow-up examination in either case (a CH or absence of a large follicle) a couple days later should reveal a corpus luteum (CL). Sometimes the CL is quick to form and is visible within a day after ovulation. The CL has a uniform echogenic appearance, which is slightly hyperechoic. During formation and then again during regression, the CL may take on a more marked hyperechoic appearance.

Predicting Ovulation

All of these tools are used in breeding management with the initial goal of predicting both when a mare might naturally ovulate, and when we might intervene to coordinate the ovulation appropriately with insemination. The effective veterinarian uses all of these tools together to understand accurately where the mare is currently in her cycle. Using one parameter alone (e.g. follicle size) will lead to mistakes.

Mares typically grow follicles to a size between 40 and 50 mm before they ovulate. Some mares may ovulate smaller follicles, and others (particularly draft breeds) will typically grow much larger follicles (50-60 mm) before ovulation. The texture of the follicle, as noted above, will soften as ovulation gets closer. A follicle approaching ovulation will often develop a thicker, hyperechoic border. And, finally, the shape of the follicle changes as it migrates towards the ovulation fossa. This change in shape is not always noted, because the distance from the ovulation fossa is variable, and, more importantly, the duration of the change in shape is probably only hours, and so is easily missed when only checking the mare every couple of days. The important point here is that if the change in shape is correctly identified, you should suspect ovulation within hours.

The dynamics of uterine edema have already been noted above. I will just remind you here that when predicting ovulation, a marked drop in uterine edema from one day to the next is a strong clue that ovulation is likely to happen within 24 hours.

Sometimes even after a complete examination of the uterus, ovaries, and cervix, you will be unsure as to whether the mare is in estrus or not. Do not forget the importance of teasing the mare to a stallion. A mare that is teasing in strongly will almost assuredly be in estrus. Sometimes a stallion is not readily available. One final diagnostic tool you may use is to perform a manual vaginal examination. After cleaning the mare, simply put on a sterile sleeve and insert your hand just past the vestibule. The vagina and cervix of a mare in estrus will typically be moist, whereas the vagina and cervix of a mare in diestrus will be drier and tacky.

Inducing Ovulation

It is not necessary to just be able to predict natural ovulation. We have the ability to pharmacologically induce ovulation. Two common drugs are used in equine practice to this effect. Deslorelin acetate is a gonadotropin releasing hormone (GnRH) analog and human chorionic gonadotropin (hCG) acts as a luteinizing hormone (LH) analog. Historically, the first formulation of deslorelin was as a subcutaneous implant. It was administered under the skin of the neck when a follicle in an estrous mare was at least 30 mm in diameter. In most mares, this resulted in ovulation between 40 and 48 hours later. Problems were noted, however, in that mares who failed to become pregnant did not return to estrus at the normally expected 3 week interval. Instead, many of these mares had prolonged interestrous intervals that ranged from days to even weeks. The remedy for this problem was to instead inject the implant under the skin of the vulva, which was much thinner and therefore it was easier to locate the implant later. After ovulation was documented, the implant was removed. These mares, if they did not become pregnant, cycled in the normal 21-day intervals. The implant is no longer available in the United States, though it is available in other countries. Currently, there is an injectable form of deslorelin that is available through certain compounding pharmacies. While no published data have yet become available (to my knowledge), anecdotally this form seems to work as well as the original implant with no reports of prolonged interestrous intervals.

The other drug, hCG, requires a slightly larger follicle to respond (>35 mm) and will cause ovulation between 24 and 48 hours (average 36 hours) in estrous mares. Because it is a human protein, there is concern about antibody formation. There are no reports of any anaphylactic reactions, but there are some data to support antibody formation correlated with inactivation of the hCG protein, nullifying the effects of the drug. These data show that repeated exposure to hCG in a breeding season may result in mares becoming resistant to the drug, and not ovulating during the anticipated interval. It is therefore recommended by some practitioners that hCG not be used more than 2 or 3 times in a season in any given mare. These data are challenged by other studies, and it remains a controversial topic.

Short-cycling Mares

If a mare is detected to be in diestrus (CL present, absence of uterine edema, tight cervix, teasing out, dry or tacky vagina), we often will give a drug to lyse the CL, removing the source of progesterone, and induce the mare to come into estrus sooner than she would naturally do so. The most common drug used for this purpose is PGF2α. The CL of the mare takes 5 or 6 days to completely form and mature. When PGF2α is administered to a mare who has a mature CL, the CL will rapidly regress and, in the absence of progesterone, the follicles present on the follicle will progress until one or two attain dominance and proceed to ovulation. While most mares will respond by 5 days post ovulation, some mares still have an immature CL at this stage. Waiting 6-7 days post ovulation before administering PGF2α to a mare will ensure that these slower mares also resond.

How rapidly a mare comes into heat after PGF2α administration depends entirely on the size of the follicles present on the ovary at the time of exposure to the drug. A mare with no readily detectable follicles present (<5 mm) may take as long as 7-9 days before showing signs of estrus. On the other hand, a mare with a large follicle (>40 mm) may respond by ovulating the large follicle by the next day, necessitating waiting another week before you can attempt to short cycle the mare again. It is therefore advisable to not short cycle mares with very large follicle, but rather to wait until that follicle begins to regress and then administer the PGF2α. Another protocol that has been tried with mixed success is to administer very low doses of PGF2α daily to mares with large follicles in order to mimic the natural release of prostaglandins from the endometrium and slowly bring the mare into estrus.

Post-breeding Evaluation and Treatment

Documenting ovulation is important in order to ensure that semen was delivered to the oocyte within an acceptable time frame (e.g. within 48 hours prior to ovulation in the case of fresh or chilled semen) and to accurately count when to perform the 14-16 day pregnancy examination. The post-breeding evaluation will also allow the practitioner to assess the uterus for unresolved inflammation, evidenced by persistent uterine edema or intralumenal fluid. Dependant upon the echogenic character and amount of fluid present, appropriate treatments consist of a drug to induce uterine smooth muscle contractions (e.g. oxytocin or cloprostenol), intrauterine antibiotic therapy, and uterine lavage. Details into specific treatment regimes are beyond the scope of this lecture. Whatever treatments are pursued, they should usually be completed by 2 days post-ovulation in order to allow the cervix time to close in anticipation of the embryo entering the uterus around 5.5 days post-ovulation.