IN-VITRO VIABILITY AND ULTRASTRUCTURAL-CHANGES IN BOVINE OOCYTES TREATED WITH A VITRIFICATION SOLUTION

Abattoir-derived oocytes were exposed to a concentrated cryoprotectant solution (DAP213: 2 M DMSO, 1 M acetamide, 3 M propanediol, and 10% F CS in TCM199) for 1.5 or 5 min at the germinal vesicle (GV) stage or a fter maturation in vitro (IVM). Their viability was assessed by in vit ro fertilization (IVF) and culture (IVC) to blastocysts. To investigat e the effect of DAP213 on the ultrastructure, GV and IVM oocytes were processed for transmission electron microscopy ITEM) before (control) or after exposure to the cryoprotectant. DAP213 induced profound ultra structural modifications to the microvilli and mitochondria, resulted in large vesicle formation, and, most significantly, caused the premat ure release of the cortical granules (CG). In IVM oocytes exposed to t he cryoprotectant for 5 min, exocytosis of CG into the perivitelline s pace was common and the IVF rate was reduced (P <.05). After exposure for 5 min, GV oocytes displayed clusters of CG comparable to controls, but after IVM-IVF, polyspermy rate was increased (P <.05). Furthermor e, treated GV oocytes showed a reduced rate of cleavage and blastocyst formation and an increased percentage of oocytes exhibiting alteratio ns in organelles, whereas the viability and ultrastructure of IVM oocy tes treated for 1.5 min was not different from controls. These observa tions demonstrate that 1) cortical granule kinetics is one of the key elements controlling fertilizability of bovine oocytes treated with cr yoprotectant, and 2) GV oocytes are more sensitive to the cryoprotecta nt than those that have already been matured in vitro, (C) 1995 Wiley- Liss, Inc.