November 2000

New Treatments for Infertility in Old Mares
by Heather Smith Thomas

Uterine infection is a common cause of infertility in older mares. Some mares are more susceptible to infection than others. Just the act of breeding introduces bacteria that can interfere with conception and pregnancy, but normal mares with a strong, healthy reproductive tract can resist this type of infection, quickly clearing the contaminants out of the uterus before the embryo emerges from the Fallopian tube to attach to the uterine lining. "All mares experience an inflammation in the uterus, known as endometritis, in response to a foreign substance such as semen," says Dr. Michelle LeBlanc, professor of large animal clinical sciences at University of Florida's College of Veterinary Medicine.

"In the early 1990's we still did not know why some mares were infertile," she says. Beginning in 1989 she led a 6-year research study on endometritis in mares and its effects on pregnancy. The problem of persistant endometritis, which is very common in older mares, costs horse breeders millions of dollars annually when mares fail to carry a pregnancy and incur related veterinary and management expenses.

Most mares are able to clear breeding-caused infection from the uterus within 24 hours, says LeBlanc. This leaves plenty of time for the arrival of the embryo, which is still travelling down the Fallopian tube where the sperm and egg united. "However, if inflammation remains in the uterus, this can lead to a fertility problem, since lingering infection can cause the death of the embryo when it enters the uterus about five days after ovulation."

Some mares, especially older mares, can't seem to defend themselves against the infections introduced at breeding. Even if they were free of infection at the time of breeding, these mares cannot throw off the effects of the inflammation in time to make the uterus healthy enough to nourish and sustain the embryo. It dies and the mare returns to heat.

LeBlanc and her researchers used scintigraphy (the process of putting a radioactive liquid, which emits gamma rays, into the uterus and then using a scintillation counter with a special camera to get an image, similar to an x-ray, of what was happening in the organ) to evaluate the changes after breeding. Problem mares were found to take longer than normal to clear the fluids and contaminants out of the uterus after breeding. "Normal mares pushed 60 percent of the debris out within 2 hours," says LeBlanc, "whereas problem mares didn't push any of it out."

In the normal mare, microscopic contaminants and bacteria are ingested by specialized white blood cells, she says. Larger particles and debris introduced by breeding are physically expelled from the uterus through secretions and contractions that send this drainage out through the cervix. "A normal mare clears out debris in 12 hours or less, whereas an abnormal mare starts to clear it out but the bacteria multiply rapidly. In a normal mare the bacteria are all gone by nine hours; in an infertile mare they attach to the uterine wall by nine hours and begin to multiply."

She says the white blood cells reach peak numbers at eight hours after breeding, then start dropping in a normal mare, and by 12 hours are all gone. "When white blood cells eat the bacteria, they grow and explode, creating an acid when they explode; the acid is part of the inside of the white cell. This acid is the signal or attractant for more white cells to keep coming into the uterus to fight the infection, but it also irritates the uterine lining and makes it more susceptible to infection. You have to stop the cycle of more white cells coming in," says LeBlanc.

The researchers found that older mares don't clear out the post-breeding debris very well because the uterus is saggy and lower in the abdomen. "The broad ligaments holding the uterus in place have stretched, especially after a mare has had a lot of foals. In the younger mare the uterus is right at the brim of the pelvis and when she walks around everything drains out. But in the old mare the uterus is like a hammock stretched between the two ovaries, sagging down. For her to drain out fluid, she'd have to stand on her hind legs. The problem is not an inability to fight off infection-her immune system is fine-her problem is that she just can't get rid of the debris in the uterus," says LeBlanc. Problem mares may take more than four days to flush contaminants from the uterus because of the combination of the sag, weaker activity of uterine muscles, and less consistent/more sporadic contractions. In some mares this leads to a chronic infection.

Researchers at the University of California (Davis) found that flushing the uterus with buffered saline solution helped clean it, increasing a mare's chances for successful pregnancy. According to their recommendations, the flushing (lavage-putting the fluid in and draining it back out) should not be done sooner than six hours or later than 20 hours after breeding, so as to give the sperm time to get through the uterus and into the Fallopian tube. The longer the debris and products of breeding are left sitting in the uterus, the more inflammation they can cause and the lower the mare's chances of pregnancy. The buffered saline solution is pumped into the uterus through a catheter, then drawn back out, and this process is repeated several times until the solution comes back clear and free of debris. This is often very effective but is time consuming, especially for some mares who may take as much as nine liters of fluid.

At University of Florida, LeBlanc began experimenting with the use of oxytocin in 1993, to see if this hormone could help the uterus flush itself. Oxytocin is produced naturally by the mare. It triggers labor, aids uterine contractions during both the birth and shedding of the placenta, helps the uterus shrink back to normal size, and stimulates milk let-down. In older mares, however, the production of this hormone is sometimes inadequate. This hormone has been used by veterinarians for many years in conjuction with birth problems, but LeBlanc's research was the first to use it as a treatment for something else. "Although oxytocin has been on the market for many years, most people didn't think it would cause the uterus to contract other than during the process of giving birth," says LeBlanc, "but when given after breeding, mares can clear all liquids from the uterus within 30 minutes."

She and her colleagues used scintigraphy to identify mares with persistent endometritis and found that the mares who retained infections after breeding could not clear the material out of the uterus. Oxytocin was the first of several drugs LeBlanc tried, and it gave the best results. During the first year of her experiments with oxytocin, a number of problem mares were successfully treated and able to continue raising foals.

Many veterinarians are now using oxytocin for problem mares. LeBlanc recommends first flushing with saline, six to eight hours after breeding, and then giving 1 ml. of oxytocin intravenously right after the flushing. "The following day we check the mare with ultrasound to see if she actually ovulated and to see if there is any fluid still in the uterus. If there's fluid, we lavage again and administer another injection of oxytocin. We don't recommend continuing these treatments much further than that-continual flushing may keep the uterus too irritated. It either works or it doesn't."

She also says you cannot treat all mares the same. Oxytocin is not the answer for every problem. "About 65 percent of Thoroughbred mares that have come to us for scintigraphy do have a delay in clearance of fluid from the uterus, and oxytocin helps, but the others don't. It is then the veterinarian's job to try to figure out what their problem is and how to deal with it.

Heidi Smith, DVM at Terrebonne Oregon, who specializes in infertility problems in older mares, says injections of oxytocin seem to help when a mare's own production is inadequate. She says it can be given right after breeding, or right after uterine infusion following breeding, and that two or three small multiple doses may have more effect than a single dose. She prefers infusing older mares after breeding, rather than flushing them. She uses saline mixed with an antibiotic such as liquid furacin and a little DMSO, and feels this type of infusion might trigger some oxytocin release in the mare. Depending on the mare, she may just infuse after breeding, or give oxytocin in addition to the infusion.

With lavage (flushing), most veterinarians wait at least six hours after breeding (to give the sperm time to be safely in the Fallopian tubes) but an infusion can be given immediately after breeding, she says, because it triggers more uterine activity and better sperm transport. She generally infuses a problem mare within four to eight hours after breeding, sometimes even as late as the next day, normally with good results. "Infusion can be followed with injection of oxytocin," says Smith. The oxytocin can be given eight to 12 hours after the breeding.

With some older mares she infuses before breeding, when they first come into heat, if the follicle is not yet ready to ovulate. To be safe, an infusion before breeding should be given at least 24 hours prior. "It may seem illogical with a big fluid-filled uterus to fill it more, but the infusion stimulates the uterus to contract and clean itself out," she says. If a mare has uterine cysts (common in older mares) and is not settling, cysts may be part of the problem. In these cases Smith infuses the mare two or three times during a heat cycle with a mix of 40 cc. saline and 20 cc. DMSO, then waits and breeds on the next cycle after checking the mare with ultrasound. In most cases this treatment will eliminate or shrink the cysts, she says, and thus the mare will be more apt to conceive.

At University of Florida, LeBlanc and her research team are now looking at ways to help diagnose problems in older mares. Not every veterinarian has access to scintigraphy. LeBlanc is working on diagnostic tests that can be used in the field, trying to develop quick and inexpensive tests that can determine if a mare has a delay in clearance of uterine fluids. They are also looking at ways to diagnose other problems that may be causing the infertility.

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