Fri, 05/24/2024 - 7:21pm

Ice, Ice, Baby

Using frozen semen

How successful are breedings done with frozen semen?


The most important thing to keep in mind when considering a frozen-semen breeding is that the lifespan of frozen semen is short, somewhere between 12 to 24 hours. Accurate ovulation timing for the breeding is crucial in order for the breeding to be successful. You want to ensure that the bitch has ovulated and that the eggs are ready to be fertilized when the semen arrives.

The fertility of the dog and the bitch are also important factors when planning frozen-semen breedings. It can be risky to use frozen semen on a virgin bitch or one that has a poor reproductive history. It may take more than the usual breeding unit if you are using a dog’s frozen semen that has a low motility percentage when thawed.

When making your arrangements, you want to deal with an experienced veterinary team that is familiar with ovulation timing and the proper thawing, evaluating and inseminating techniques of the frozen semen.


What is the best way to do ovulation timing?


Monitoring your bitch’s heat cycle to determine the optimal time to inseminate involves carrying out one or a combination of the following procedures.

Slides, or smears, of the cells of the vaginal lining can be stained and evaluated under the microscope to determine the progression of the heat cycle. Once the changes in the vaginal cells reach 50 percent cornification, blood testing should begin.

Vaginoscopy involves the use of a scope to examine changes in the wall of the vagina, which correlate to the stage of the heat cycle. Vaginoscopy can also identify anatomic abnormalities, such as strictures and persistent hymen, before breeding.

Blood samples can be taken to measure the hormones, progesterone and luteinizing hormone (LH). The number of blood samples that need to be taken to determine when to inseminate varies from bitch to bitch. I recommend starting testing five days after the onset of heat and then every second or third day until the LH surge or the progesterone level reaches 2.0 ng/dL. If you are doing LH testing, blood samples need to be drawn daily.

Progesterone is a hormone that rises during the estrous cycle preceding the release of LH. Progesterone is necessary to prepare the uterus for implantation of the fertilized eggs and for maintaining pregnancy. LH is the hormone that causes the release of the eggs from the ovary. The rise of the level of LH is the actual metabolic signal to the ovaries to release the eggs.

The day of the LH surge, or when the progesterone level reaches 2.0 ng/dL and keeps rising, is designated as day 1 of the bitch’s fertile period. The day or days recommended for insemination using frozen semen are day 5 when one insemination is being performed, and days 4 and 6 or days 5 and 6 if the bitch is being inseminated twice.


How is the semen frozen?


Semen freezing begins with collection of the semen by manual stimulation of the dog. A dog ejaculates in three sections, or “fractions.” The first is the pre-sperm faction, followed by the second, which is the sperm-rich fraction, and the third fraction, which contains prostatic fluid.

Although the third fraction is important during a natural breeding to provide volume to flush the semen into the bitch’s reproductive tract, it is actually harmful to the sperm during long-term storage. When collecting semen for freezing, care must be taken to separate the fractions, since only the second, sperm-rich fraction is used for freezing.

Once the semen is collected, it is evaluated under a microscope to record information about its color, density, volume, motility, quality of motion, and number of normal and abnormal sperm cells. The minimum values for normal semen are at least 70 percent progressively motile and at least 70 percent structurally normal. Ideally, these values should be higher in semen that is to be frozen, since the values will inevitably decrease through the freezing process.

Semen extender is added to the sample, and the semen is gradually cooled. Extenders contain ingredients to provide nutrients for the semen as well as ingredients to protect the sperm’s membranes from the freezing process.

Frozen semen is stored in either a pellet form or a straw form. To store in pellets, the cooled semen-extender mixture is dripped through a pipette onto a block of dry ice that has had shallow, uniform holes bored into the surface. The semen instantly freezes into spheres. The pellets can be transferred into vials, which are then stored in liquid nitrogen at the extreme temperature of -198oC, or -324oF.

To store the semen in straws, the semen-extender mixture is injected into straws, and the ends of the straws are sealed. The sealed straws are then placed in progressively colder steps, until finally they are plunged into liquid nitrogen.

After the semen-freezing process is complete, some pellets or a straw should be thawed and evaluated. It is normal for the motility of the semen to be decreased as compared to the motility before being frozen, but it should still ideally be at least 50 percent motile. If there is a big change in the quality after being frozen, it may be that the semen needs to be frozen with a different extender. In some cases, it takes multiple attempts at freezing before the best extender is identified for your dog. The post-thaw evaluation allows you to determine how many pellets or straws will be needed for each breeding unit.


What are the insemination procedures?


When thawed, the lifespan of frozen semen is 12 to 24 hours. For best results, procedures that place the thawed semen directly into the uterus maximize the chances for conception. The two techniques used are surgical insemination (SI) and transcervical insemination (TCI). The process of surgical insemination involves placing the bitch under general anesthesia, making a surgical incision into her abdomen, and injecting the semen directly into her uterus through the uterine wall.

The process of TCI involves passing a metal scope with a camera on its end into the vagina. The camera is advanced until the operator can see the cervix, and a catheter is passed through the cervical opening into the uterus. The semen is then flushed into the uterus through the catheter. Because most bitches in standing heat tolerate the process of TCI without sedation or anesthesia, it has fewer risks than SI.

Several studies have indicated that the success rate is the same for both procedures if performed by an experienced clinician. The cost of the equipment required for TCI and the difficulty of the procedure can make it challenging to find a veterinary practice that offers this service.


Should I have my dog collected and freeze the semen?


If you think your dog will be used in a breeding program now or at some time in the future, you should consider having him collected and the semen frozen. Once the semen is properly stored, it can be thawed and used at any point in the coming years. If you have a dog who is genetically valuable, you could have his semen frozen now and still take advantage of his genetics in two years, 15 years, 30 years or beyond.

Working with frozen semen can decrease the stress of long-distance breedings. The frozen semen can be shipped well before it is needed so it is on hand to be thawed and inseminated when the bitch is ready. Even for local breedings, if the dog has a busy show schedule or multiple breeding commitments, having frozen semen on hand can eliminate the problem of last-minute availability.

The best time to have your dog’s semen collected and frozen is when he is young, healthy and in the prime of his fertility. This will provide you with the best samples for freezing. It also safeguards his genetics in the event he was to pass away unexpectedly. It is possible in some situations to retrieve a semen sample postmortem by doing an epididymal collection. These samples are often less than ideal quality but can provide an opportunity to preserve specific genetics if his semen was not frozen earlier.

An important factor to consider is what will happen to your dog’s frozen semen if you were to pass away unexpectedly. Not all semen-freezing banks collect this information at the time of collection. Frozen semen is an asset that should be considered when making out your will. You may consider leaving money for the payment of storage fees or leaving the stored semen to a trusted friend or family member. By making arrangements ahead of time, you will avoid the potential destruction of a valuable contribution to the breed’s genetic potential.



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