(All Photos By Jessie Cohen, NZP Staff Photographer)
Smithsonian News Service - February 1989
Artificial insemination, test-tube fertilization, frozen storage of embryos and surrogate parenting. For many people in the 1980s, these new and often controversial procedures of artificial reproduction hold out a last hope for starting a family.
But today's reproductive technology is not limited to helping create human families. Reproductive specialists at the nation's zoos and research centers have begun to investigate ways to save endangered animals with these techniques.
"For some endangered species, the new procedures for reproduction--often designed to solve human fertility problems--offer a last hope for survival," Dr. David Wildt, a reproductive physiologist at the Smithsonian's National Zoological Park in Washington, D.C., says. Wildt often works in collaboration with researchers from other zoos around the country, such as the San Diego Zoo and the Henry Doorley Zoo in Omaha, Neb.
Artificial reproduction can dramatically increase the "breeding potential" of healthy individuals of an endangered species, Wildt continues. Sperm from a genetically fit male, for instance, might be removed, preserved and shipped around the world to impregnate many females of a particular species. And transporting an animal's cells is often much easier than shipping an entire animal, Wildt adds.
The new procedures can also help physically or behaviorally handicapped animals reproduce. For example, Wildt says, a female animal that is unable to mate naturally could be artificially inseminated. A male might have its sperm collected for use sometime later in an artificial reproduction attempt.
Animal rights activists often object to such biomedical research. The Smithsonian's Wildt, however, says, "people in the animal rights movement are concerned about the exploitation of animals for the benefit of mankind. We are reversing that philosophy. We are taking techniques of artificial reproduction developed to improve the fertility of humans and applying them to endangered animals."
Perhaps the most familiar procedure is artificial insemination--introducing semen into the uterus of a receptive female to fertilize her ova or eggs. But this procedure is not as straightforward as it sounds. Before artificially inseminating any animal, researchers must have detailed knowledge about its reproductive cycle, including such important facts as when females of a species ovulate--that is, produce viable eggs. For animals used in agriculture--such as cattle and sheep--an economic incentive has existed to develop this knowledge. Unfortunately, for exotic animals, this information is often lacking.
As an example, Wildt cites his work with a small number of black-footed ferrets, discovered in Wyoming. These animals were thought to be extinct. For protection, the ferrets were brought into a managed environment at a breeding facility set up by the Wyoming Department of Fish and Game. Scientists felt it was urgent to breed as many of these animals as possible and that artificial reproduction could increase their breeding potential. Because scientists knew very little about the black-footed ferret's reproductive biology, however, National Zoo researchers first tried artificial insemination with domestic ferrets, which are not endangered. "Domestic ferrets are good models for black-footed ferrets," Wildt explains. "Since the population of black-footed ferrets is so small, it would be dangerous to use them for developing new procedures."
A litter of 31 domestic ferrets produced by using artificial insemination showed how this work could be done safely with the 18 surviving black-footed ferrets. If the population of the black-footed ferrets can be increased significantly, scientists may be able to reintroduce the species to its wild habitat, Wildt says.
An important aspect of Wildt's work with the ferrets was the use of frozen ferret semen. Again guidelines for collecting and storing the semen were established by research with semen collected from domestic ferrets. Wildt; Dr. Mitchell Bush, then the National Zoo's chief veterinarian; and Dr. JoGayle Howard, another Zoo researcher, evaluated several different methods of freezing and storing the semen.
Essentially, the process they developed consists of freezing the semen--mixed with egg yolk as a protein-rich extender that provides nutrients and bulk to the small amounts of fluid--in pellets on blocks of dry ice. These pellets are then stored in canisters of liquid nitrogen at a bone-chilling -196 degrees Celsius. Semen from the domestic ferrets was frozen for a year-and-a-half before it was thawed and used for artificial insemination.
A cup filled with frozen pellets of ferret semen is placed into
a "cryogenic" storage tank. Liquid nitrogen in the tank
prevents damage to animal cells, allowing long-term storage.
The ability to freeze sperm and other reproductive cells is vital for endangered species, Wildt says. When only a few individuals of a species remain, the "gene pool," or number of different genes existing for that species, is greatly diminished. New offspring may be extremely vulnerable to genetic diseases. The males of the nearly extinct Florida panther, for example, often are genetically disposed to low sperm counts and abnormal sperm. The females of this species often suffer from dysfunction of their reproductive cycles, which is also a hereditary problem. Such problems, probably resulting from inbreeding, can rapidly lead to a species' extinction.
To produce the healthiest offspring possible, then, researchers attempt to breed animals in a species that are as distantly related as possible. With the ability to freeze and store reproductive cells, Wildt says, biologists can save this genetic material for one or more generations, thus artificially increasing the gene pool of the animals used in reproduction. "If scientists are to save this species from extinction," Wildt says of the black-footed ferrets, "they must be able to preserve semen for long periods of time. After several generations have passed, semen can be thawed and used to impregnate females that are genetically distant from the donor male."
Other species threatened with extinction include large cats, such as the clouded leopard, cheetahs and some tiger sub-species. In captivity, males and females of these species are frequently incompatible, maiming or even killing one another when paired for mating. Wildt's recent experiments with test-tube or in-vitro fertilization--another procedure developed for human beings--might offer these animals some hope.
Wildt and his colleagues began their research with domestic cats, again to determine how the procedure might be applied to the cats' endangered cousins. First, hormones were used to stimulate and produce eggs in the female cats' ovaries. Next, the eggs were removed from the cats with a very small needle and suction device. The eggs were then placed in a glass vessel where sperm from male cats was introduced.
At this stage, researchers watched the new embryos for proper development. Healthy embryos were placed back into the reproductive tracts of females that produced the eggs. In the spring of 1987, Wildt and his colleagues proudly announced the births of three litters of adorable kittens, proof that test-tube fertilization is possible with carnivorous species. More recently, Wildt and his colleagues have produced viable, in-vitro-conceived puma embryos. They have also begun similar research on the Florida panther, whose numbers have dwindled to fewer than 25.
"In-vitro fertilization requires an amazing amount of time in the laboratory and a lot of expensive and specialized equipment," Wildt says. "Working together, our graduate students at the National Zoo were able to solve most of the problems for the domestic cat effectively. Still, the work required two years of research, which is a long time when trying to produce a technique applicable to endangered species."
The World's first test-tube kittens.
The work with in-vitro fertilization holds other possibilities, Wildt says. For example, embryos from one species might be implanted in a female of a related species. This "surrogate-parenting" would be valuable when the females of an endangered species are somehow unfit for reproduction, but nonetheless able to produce viable ova.
Zoos might also keep embryos of animal species threatened in the wild in frozen storage, Wildt says. While saving the natural habitat of endangered species is the first priority of wildlife researchers, such techniques might provide another safeguard against extinction.
"Zoos are typically places for families to have fun and learn about wildlife from all over the world," Wildt says. "With this research into artificial reproduction, we also hope zoos can play an important role in saving the same wildlife for many generations to come. "
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