Fertilization in vitro of Rat Ovarian Oocytes after Freezing and Thawing

Rat ovarian oocytes recovered from immature females 48 h after an injection of PMSG and 3, 6.5 and 8.5 h after an injection of hCG were frozen at −196°C. After thawing they were cultured for 14, 11, 7.5 and 5.5 h, respectively, and then inseminated with epididymal spermatozoa. When dimethylsulphoxide (DMSO) was added at 0°C, 46–54% of the oocytes recovered without injection of hCG were normal after thawing and culture; 33–34% of the normal oocytes were penetrated following insemination in the samples in which DMSO was diluted at room temperature or 37°C after thawing. However, when DMSO was diluted at room temperature, comparatively higher proportions of normal oocytes (63%) and of penetrated oocytes (47%) were observed in the samples to which DMSO was added at room temperature and 37°C, respectively. With increasing time of oocyte collection after injection of hCG, the proportions of normal oocytes cultured after thawing and of penetrated oocytes after insemination were progressively increased when DMSO was added and diluted at room temperature.