Maturation of the reconstructed oocytes by germinal vesicle transfer in rabbits and mice

The present study was designed to evaluate the feasibility of germinal vesi cle (GV) transfer in rabbits and mice. The GV oocytes were collected from o varies and cultured in 20 mug/mL 3-isobutyl-1-methylxanthin (IBMX) in TCM19 9 medium, which caused oocytes to shrink, enlarging the perivitelline space to facilitate the GV removal and transfer. Pairs of GV - cytoplast complex es were fused with electric pulses, and the fused, reconstructed oocytes we re cultured in TCM199 for 24 h. Results are as follows: 1) The exposure tim e of rabbit GV oocytes to IBMX medium affected the success of GV removal. F or oocytes cultured for 2 and 3 h in IBMX medium, removed rates were 56% an d 44, respectively, significantly higher (P < 0.05) than removal rates of G V oocytes cultured for 1 and 4 h (27% and 27%, respectively); 2) There was no significant difference (P > 0.1) in fusion and maturation rates of rabbi t reconstructed oocytes collected at 72 and 84 h after initiation of FSH in jection to donors; 3) eCG in the maturation media improved development of r abbit-to-rabbit GV transferred oocytes but had no positive effect on mouse- to-rabbit GV transferred oocytes; 4) When mouse GV-karyoplasts were injecte d into enucleated rabbit oocytes, fusion rates of GV-karyoplasts measuring 40- to 50-mum and 80- to 90-mum in diameters obtained were 84% and 93%, res pectively. The rates were significantly higher (P < 0.05) than fusion rates after transferring GV-karyoplasts measuring 30- to 35-<mu>m in diameter (6 3%). The maturation rate (89%) of reconstructed oocytes composed of 80- to 90-mum mouse GV-karyoplasts and rabbit GV-enucleated cytoplasts was higher than that seen for oocytes composed of 40- to 50-mum (77%, P<0.05) or 30- t o 35-<mu>m (59%, P<0.01) mouse karyoplasts. Thirty-five of the 63 (56%) mat ure mouse-to-rabbit reconstructed oocytes had the normal complement of 20 c hromosomes. (C) 2001 by Elsevier Science Inc.