Fertilization of Rat Eggs in Vitro by Epididymal Spermatozoa and the Development of Eggs Following Transfer

Summary. Newly ovulated eggs from immature rats treated with PMSG and HCG were `inseminated' in vitro with spermatozoa recovered from the cauda epididymidis of mature males. A modified Krebs-Ringer bicarbonate solution, supplemented with glucose, sodium pyruvate, sodium lactate, bovine serum albumin and antibiotics, was used. Eggs and spermatozoa were incubated in 0·4 ml medium under mineral oil at 37° C in a gas phase of 5% CO2 in air. Sperm penetration through the zona pellucida started 5 to 5½ hr after insemination and the transformation of sperm head(s) into the male pronucleus occurred about 2 hr later. Most of the penetrated eggs were at the two-cell stage 30 to 35 hr after insemination. The percentage of eggs undergoing fertilization was 88·7% (316/356) 7 to 20 hr after insemination and that of fertilized eggs at 30 to 46 hr after insemination was 75·3% (174/231). Only a few eggs cleaved to the four-cell stage after further incubation. A total of 203 two-celled eggs fertilized in vitro were transferred to the oviducts of fourteen pseudopregnant rats on the first day of dioestrus. Nine recipients became pregnant and forty-three fetuses and newborn young were obtained (21% of development).