Timing of the first cleavage post-insemination affects cryosurvival of in vitro-produced bovine blastocysts

The time of the first cleavage of bovine zygotes during in vitro culture ca n affect the rate of development and cell number of the blastocysts. The ai m of this work was to study the effect of the timing of first cleavage on t he cryosurvival of the resulting blastocysts, Following standard IVM and IV F, zygotes were cultured in modified synthetic oviduct fluid (SOF), with 10 % fetal calf serum (FCS) added 48 hr post insemination, in a humidified atm osphere of 5% CO2, 5% O-2 and 90% N-2, Embryos which cleaved by 24, 27 30, 33, or 36 hr after insemination (IVF) were harvested and further cultured t o the blastocyst stage (day 7 or day 8 post IVF), All developing blastocyst s on days 7 and 8 were classified into three groups and were cryopreserved by vitrification. Group A consisted of blastocysts (<150 mu m, small blasto cysts); group B consisted of expanded or hatching blastocysts (>150 mu m, l arge blastocysts); and group C consisted of morphologically poor quality bl astocysts. The vitrification solution consisted of 6.5 M glycerol and 6% bo vine serum albumin in PBS (VS3a). Thawed embryos were cultured further and survival was defined as the re-expansion and maintenance of the blastocoel over 24, 48, and 72 hr, respectively. Overall survival and hatching at 72 h r post-thawing was higher in blastocysts formed by day 7 than those formed by day 8 (60% vs, 40% survival; 63% vs. 45% hatching). Large blastocysts fr om day-7 and day-8 groups survived significantly better than small or poor quality blastocysts (76% vs. 63% and 31%; 72% vs. 30% and 26%, respectively ; P < 0.05). Day-7 blastocysts from the 27- and 30-hr cleavage groups survi ved significantly better than those from the 36-hr group (63% and 66% vs. 2 5%, P < 0.05). Day-8 blastocysts from later cleaved (30 hr) zygotes had a h igher survival than the 27-hr cleavage groups (52% vs. 26%, P < 0.05). Thes e results indicate that the day of blastocyst appearance, developmental sta ge, and timing of the first cleavage post-insemination can influence the cr yosurvival of bovine blastocysts following vitrification. Mol. Reprod. Dev. 53:318-324, 1999. (C) 1999 Wiley-Liss, Inc.