VITRIFICATION OF BOVINE IVP EMBRYOS: AGE OF EMBRYOS AND EXPOSURE TIME TO CRYOPROTECTANT INFLUENCE VIABILITY

Avaliou-se diferentes tempos de exposicao e concentracoes de crioprotetores na vitrificacao de embrioes bovinos PIV. No primeiro experimento, foram utilizados blastocistos do dia 7 (Bx-D7). No tratamento 1 (T1), 82 embrioes foram expostos por 1 min. a solucao de equilibrio (SE1 = 10% EG + 10% dimetilsulfoxido (DMSO), seguido da exposicao por 20 segundos a solucao de vitrificacao (SV1 = 20% EG + 20% DMSO). No Tratamento 2 (T2) 84 embrioes foram expostos por 3 minutos a SE2 (8,25% EG + 8,25% DMSO), seguido de 45 segundos na SV2 (16,5% EG + 16,5% DMSO). No segundo experimento adotou-se os mesmos procedimentos do primeiro, porem com Bx D8. A remocao dos crioprotetores foi executado em duas etapas de cinco minutos, em 0,3 e 0,15M de sacarose. Os embrioes foram incubados por 72 horas, avaliando-se as taxas de re-expansao e eclosao (12 e 72 horas, respectivamente). No primeiro experimento, a taxa de re-expansao no T1 (91,6%) foi superior a do T2 (82,0%) (p 0,05). No segundo experimento, as taxas de re-expansao nao diferiram entre T1 e T2 (65,8 e 68,7% respectivamente), porem a taxa de eclosao do T1 (51,7%) foi superior a do T2 (33,2%) (p Abstract Different exposure times and cryoprotectant concentrations were evaluated in vitrification of D7 and D8 IVP bovine embryos. In the first experiment, D7 expanded blastocysts (Bx) were used. In Treatment 1 (T1) 82 embryos were exposed for 1 minute to an equilibrium solution (ES1 = 10% EG + 10% DMSO), followed by 20 seconds exposure to vitrification solution (VS1 = 16.5% EG + 16.5% DMSO). In Treatment 2 (T2) 84 embryos were exposed for 3 minutes to ES2 (8.25% EG + 8.25% DMSO), followed by 45 seconds exposure to VS2 (16.5% EG + 16.5% DMSO). Embryos were loaded in OPS with 5µL VS, and plunged into liquid nitrogen. The second experiment used D8 Bx as previously described. Cryoprotectant removal was performed in two steps, in 0.3 and 0.15M sucrose gradients for 5 minutes each. Embryos were then incubated for 72 hours, and re-expansion and hatching rates evaluated at 12 and 72 hours, respectively. In the first experiment, re-expansion rate in T1 (91.6%) was higher (P 0.05). In second experiment, re-expansion rates did not differ between T1 and T2 (65.8% and 68.7%) respectively, while hatching rate in T1 (51.7%) was higher (P