Cryopreservation of canine embryos.

The assisted reproductive techniques (ARTs) such as in vitro fertilization, embryo transfer, and cryopreservation of gametes have contributed considerably to the development of biomedical sciences in addition to improving infertility treatments in humans as well as the breeding of domestic animals. However, ARTs used in canine species have strictly limited utility when compared with other mammalian species, including humans. Although successful somatic cell cloning has been reported, artificial insemination by frozen semen to date is only available for the improved breeding and reproduction for companion and working dogs as well as guide dogs for the blind. We describe here the successful cryopreservation of embryos and subsequent embryo transfer in dogs. Canine embryos were collected from excised reproductive organs after artificial insemination and subsequently cryopreserved by a vitrification method. When the 4-cell to morula stage of cryopreserved embryos were nonsurgically transferred into the uteri of nine recipient bitches using a cystoscope, five recipients became pregnant and four of them delivered a total of seven pups. The cryopreservation of embryos in canine species will facilitate the transportation and storage of genetic materials and will aid in the elimination of vertically transmitted diseases in dogs. In addition, this technique will contribute to the improved breeding of companion and working dogs such as guide dogs, drug-detecting dogs, and quarantine dogs.

Individual fertility differences in the frozen-thawed spermatozoa among semen donors in the Labrador Retriever.

Purpose: We aimed to validate and determine the possible application of transcervical insemination of frozen semen for improved breeding in guide dogs for the blind in Japan. Methods: From February 2004 to March 2007, a total of 53 Labrador Retriever bitches, used for the breeding of guide dogs for the blind, were transcervically inseminated with frozen-thawed semen from 13 males by means of a cystoscope. Results: The overall whelping and pregnancy rate with the frozen semen was 42%. Pregnancy rates ranged widely from 0 to 100% depending on the semen donor male. Of 13 males, 6 males exhibited severely poor fertility (less than 20% pregnancy rate) and 3 males exhibited high fertility (over 70% pregnancy rate) on artificial insemination. However, the spermatozoa motility after thawing was not significantly different among these dogs. In addition, heterospermic insemination revealed the optimal timing for transcervical insemination with frozen-thawed semen to be by day 6 after the LH surge. Conclusions: Although transcervical insemination of frozen-thawed semen is effective for breeding of guide dogs for the blind, some modification of freeze-thawing procedures might be required to overcome individual fertility differences in the frozen-thawed spermatozoa among semen donor dogs,. In addition, the motility of spermatozoa after thawing might not be an appropriate indicator of the relative fertility of frozen-thawed spermatozoa in dogs.

Beneficial effect of erythropoietin administration on murine infection with Trypanosoma congolense.

The effect of erythropoietin treatment on Trypanosoma congolense infection in mice was studied. Survival rates of mice were dramatically improved by treatment with recombinant human erythropoietin (r-hu-EPO; 5,000 U/kg) when infected with 1,000 cells of T. congolense IL3000 (P < 0.05). All the untreated mice infected with T. congolense IL3000 died by day 9 of infection; however, 100%, 50%, and 25% of the mice treated with r-hu-EPO for 8 days survived to day 20, day 40, and day 60 of the parasitical infection, respectively. Anti-8-hydroxy-2'-deoxyguanosine antibody, a biomarker for oxidative damage of DNA, yielded positive reactions in the cytoplasm of the parasites recovered from the mice treated with r-hu-EPO. These results, taken together, indicate that erythropoietin administration is effective for the treatment of T. congolense infection.

Cryopreservation of canine ovaries by vitrification.

The cryopreservation of ovarian tissues is a technology with significant potential for the preservation of the genetic resource materials of working dogs, including guide dogs for the blind. However, no attempt has been reported on cryopreservation of the canine ovary. Thus, we evaluated a vitrification method for cryopreservation of canine ovaries and determined the potential functionality of vitrified-warmed canine ovaries by means of transplantation into non-obese diabetic-severe combined immunodeficiency (NOD-SCID) mice. All ovarian tissues cryopreserved by vitrification were morphologically normal in terms of histology. Cryopreserved ovaries were transplanted into the ovarian bursa of the NOD-SCID mice, and the xenografts were recovered from 23 of 23 mice (100%) 4 weeks after the operation. The transplanted canine tissue was tightly adhered to the mouse ovary. Although antral follicle formation did not occur after grafting, proliferating cell nuclear antigen immunoreactivity was detectable in many of the granulosa cells in the primary follicles of the grafts. These results indicate that cryopreservation of the canine ovary by vitrification appears to have the potential to restore endocrine function and ovulation potential.

Exposure of C57BL/6J male mice to an electric field improves copulation rates with superovulated females.

It is well-known that there are considerable strain differences in the relative copulation rates between male and superovulated female mice. In particular, the C57BL/6J strain of mice has a lower rate of successful copulation. We examined the effect of exposure to an electric field on sexual behavior in C57BL/6J male mice. When C57BL/6J males were exposed to a 50 Hz, 45 kV/m electric field for 30 min per day for 11 days and placed in a cage with a superovulated female of the same strain, the successful copulation rates of males was significantly improved compared with unexposed males (P<0.05). These results suggest that the exposure of C57BL/6J male mice to an electric field improves their sub-fertility activity in mating with superovulated females.