Vitro Fertilization In Rhesus Monkeys

In order to set up an effective route of in vitro fertilization and embryo transfer in rhesus monkeys, 5 male rhesus monkeys' semen were electroejaculated and evaluated. The superovulation protocols were made using gonadotropin for animals made in China, 15 female rhesus monkeys were stimulated and oocytes were recovered using invasive surgical techniques. In vivo or in vitro maturated oocytes were inseminated with liquefied, separated and capacitated motile sperm. Zygocytes were cultured in vitro, and 8-cell embryos were cryopreserved, three of which were thawed and transferred to a recipient on the fifteenth day of its menstrual cycle. The whole results were as followings.1. More semen was obtained using penile electroejaculation. Sperm concentration and motility showed no difference between penile electroejaculation and rectal probe electroejaculation after being clystered. The quality and quantity of semen reduced significantly using rectal probe electroejaculation without clystering. 1 percent a -chymotrypsin liquefied over 40 percent semen clot of rhesus monkeys.Dilution made semen quality declined.2. Among the four stimulating protocols, more than 20 oocytes were obtained in C and D. When the dose of HCG was added to 1500IU, 53.6 percent of oocytes were in metaphase II (MII). As the interval extended to 34h after HCG administration, the percentage of MII oocytes was 46.3. The percentage of normal morpha (68±10 to 82±7 and 84±6) and motility of sperm(72±l 1 to 85±6 and 83±9) could be improved by using the two motile sperm separation methods, but there was no difference in fertility rate. 136 oocytes were used in the experiment of IVF, 30 of which were damaged to death during capacitation; the rest were inseminated and 56 of which fertilizated. 6 of fertilized oocytes were fixed, and 28 cleavaged. 11 of the 28 fertilizated oocytes developed to 8-cell embryos. Although the cryosurvival rate of three embryos was quite high after thawed (100%, 3/3), pregnancy rate was zero after being transferred.The conclusions resulted from above are as followings:1.Clyster made rectal probe electroejaculation more effective, and penile electroejaculation was more effective than rectal probe electroejaculation. Proper anaesthesia was more suitable for semen collection for rhesus monkeys. The ejaculates can be partly liquefied by physical or chemical means.2. Scheme C and D showed better superovulating results. HCG is very important for oocytes maturation, and the longer interval after using HCG can improve the degree of maturation. January and February seems to be the best seasons for superovulation in rhesus monkeys.3. There was no significant difference in fertilization rate between Percoll and swim up groups both methods were effective motile sperm separation methods. Both caffeine and dbcAMP are necessary for inducing sperm to capacitation, they have different effects on the ability of sperm to bind to the zona and to undergo the acrosome reaction. The optimal sperm concentration for IVF is 2x105sperm/ml or so.Serum and hormone can improve the maturation rate of oocytes. And serum and amino acid are necessary for embryo development.4. The failure of pregnancy maybe resulted from multiple factors.