Activation of Equine Nuclear Transfer Oocytes: Methods and Timing of Treatment in Relation to Nuclear Remodeling1

Early development of embryos produced by transfer of equine nuclei to bovine cytoplasts is superior to that of intraspecies equine nuclear transfer embryos. This may be related to differ-ences in chromatin remodeling or efficiency of activation be-tween the two oocyte types. The pattern of donor nucleus re-modeling was examined in equine-equine and equine-bovine re-constructed oocytes. Chromosome condensation occurred in equine cytoplasts by 2 h but was not seen in bovine cytoplasts until 4 h. We investigated the effect of activation of equine-equine reconstructed oocytes at,30 min or at 2 h after recon-struction. Four activation treatments were evaluated at each time point: injection of sperm extract alone, or in combination with 6-dimethylaminopurine (6-DMAP), cytochalasin B, or 1% dimethylsulphoxide. There was no significant difference in nor-mal cleavage rate or average nucleus number of embryos be-tween equine oocytes activated,30 min or at 2 h after recon-struction. The combination of 6-DMAP with sperm extract sig-nificantly (P, 0.01) improved cleavage rate compared with the other three treatments. Activation with sperm extract and 6-DMAP 2 h after donor nucleus injection gave the highest cleav-age (79%) and the highest cleavage with normal nuclei (40%). Sperm extract and 6-DMAP also effectively activated oocytes parthenogenetically, yielding 83 % cleavage and 73 % cleavage with normal nuclei. These results indicate that although nuclear remodeling occurs rapidly in equine cytoplasts, early activation does not improve embryonic development after reconstruction. embryo, fertilization, gamete biology, in vitro fertilization, ovu