Various Conditions for the Fertilization of Rat Eggs in Vitro

Eggs recovered from oviducts of superovulated immature rats were inseminated in vitro with epididymal sperm of mature rats. A higher proportion of eggs (77%) was penetrated during incubation with 5% CO2 in air than with air alone (24%), probably due to the maintenance of a relatively constant pH in the presence of 5% CO2 in air. When sperm and eggs were incubated in a small droplet of medium in the presence of 5% CO2 in air, penetration rates increased gradually from 10% in a droplet of 5 µl to 88–79% in droplets of 100–400 µl. The sperm motility and survival at the end of incubation were poor in a very small droplet especially in the absence of eggs. A high proportion of eggs was penetrated (94%) whether follicular cells were removed or not when the eggs were examined 7–8.25 h after insemination; when examined 3.5–4 h after insemination, the proportion of penetrated eggs was lower (6%) in the presence than in the absence (21%) of follicular cells, indicating the unimportance of follicular cells for the capacitation of rat sperm and fertilization of rat eggs. By insemination of epididymal sperm from CD or Long—Evans strain rats, the proportion of eggs penetrated or undergoing fertilization and that of polyspermic eggs were not significantly different whether the eggs were from CD or Long—Evans strain.