CHAPTER 45Insemination with Frozen Semen
Artificial insemination (AI) with frozen semen involves the timely introduction of an adequate number of sperm into the mare’s uterus. To ensure success in an AI program with frozen semen, knowledge of the reproductive tract of the mare is very important. Mare selection, as well as close monitoring of the cycle, are key factors that will determine the success of this kind of program (see Suggested Readings at the end of this chapter).
In the equine industry (especially in some breeds), artificial insemination with frozen semen is not well understood, and that makes many owners and breeders approach the subject with skepticism. Cryopreservation of spermatozoa remained an enigma until the serendipitous discovery of glycerol in 1948 by scientists in Cambridge, England.1,2 Regretfully, there is still much progress needed in processes of successful cryopreservation of equine spermatozoa (see Suggested Readings).
Since 1957, when the first foal born from frozen semen was reported,3 many attempts have been made to maximize this reproductive technique. In spite of these efforts, the technique has not developed as quickly as it has in bovine semen. Reasons for this include a lack of research, a lack of funds available, the relatively small number of mares bred with frozen semen,4 and little selection pressure for equine fertility because performance, not fertility, is the prime determinate for breeding.5 Successful pregnancies can be difficult to achieve with frozen stallion semen. Important factors that affect pregnancy rates with frozen semen are (1) the stallion, (2) semen processing and handling, (3) mare’s status, and (4) experience of the inseminator.6
ADVANTAGES
DISADVANTAGES
SELECTING THE PROPER MARE FOR ARTIFICIAL INSEMINATION WITH FROZEN SEMEN
Although all mares theoretically could be bred with frozen semen, the recognized lower pregnancy rate per cycle suggests only mares that are reproductively normal would be good candidates. To be considered reproductively normal, the mare should have a uterine biopsy of either I or IIA. Mares with biopsies ranked IIB or III have a very low expected foaling rate.7 Mares with known delayed uterine clearance should be avoided, as well as breeding during the foal heat. Other poor choices include mares not in good body condition and, in general, old mares or mares that have not been able to get pregnant under good management conditions.
The mare should be examined manually and with ultrasonography to identify any anatomic malformations, fluid accumulation, or any other abnormalities. Uterine cysts should be recorded in order to avoid confusion when evaluating for pregnancy. All mares (except maiden mares) should be cultured, and cytologic evaluation is strongly recommended, especially in older mares or mares that have not had foals for long time. When culturing a mare, it should be ensured that the swab enters the uterus and collects bacteria only from the body of the uterus.8 If required, Caslick’s procedure should be performed to avoid contamination of the reproductive tract.
TIMING OF THE INSEMINATION WITH FROZEN SEMEN
It is critical when inseminating with frozen semen to know when the mare is going to ovulate in order to deposit the semen as close as possible to the time of ovulation, preferably before ovulation. Synchronization of ovulation obviously has many advantages in a breeding program. Because the follicular status of the mares can be predetermined, mares can be mated at a more precise time, increasing the chances of pregnancy and decreasing embryonic loss.9 Currently there are two available products that help the clinician induce a reliable ovulation time and thus increase the success of insemination with frozen semen. Human chorionic gonadotrophin (hCG) (Chorulon, Intervet Inc., Holland) is a glycoprotein hormone with luteinizing hormone (LH) activity. More recently a biodegradable short-term implant of a gonadotropin-releasing hormone (GnRH) analogue, deslorelin (Ovuplant, Fort Dodge), has been given to mares to hasten ovulation.10 Although both products have been reported to control the time of ovulation in the mare, it has been suggested that Ovuplant has fewer complications such as failure to respond.
hCG reliably induces ovulation at 36 plus or minus 4 hours and Ovuplant at 41 plus or minus 3 hours.11 Among the disadvantages that synchronization with hCG has are inconsistencies in response between and within mares and antibody formation, which may or may not cause refractoriness and impair fertility.12 Ovulation induction with deslorelin has been associated with increased interovulatory intervals in some mares.13 However, a recent study confirmed that removing the deslorelin implant 48 hours after administration prevented the suppression of follicle-stimulating hormone (FSH) secretion and subsequent follicular development.14
There are two approaches currently used for insemination of frozen semen. Firstly and most commonly mares are inseminated immediately before, during, or after ovulation. This approach requires considerable effort and multiple examinations using palpation/ultrasonography. The principal advantages of this regimen are firstly, it decreases to only one the number of times a mare is bred during an individual estrous cycle, and secondly, it should avoid breeding mares with abnormal ovulations such as anovulatory hemorrhagic follicles.5
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