ACTIVATION REGIMENS TO PREPARE BOVINE OOCYTES FOR INTRACYTOPLASMIC SPERM INJECTION

Activation of bovine oocytes to produce a single haploid pronucleus in preparation for intracytoplasmic sperm injection (ICSI) has been inve stigated with Various combinations of ionomycin and 6-dimethylaminopur ine (DMAP). Effects were evaluated by immunocytochemical staining, chr omosomal analysis and assessment of development in vitro. Oocytes matu red in vitro were exposed to: ionomycin alone (single or repeated trea tments, Groups 1 and 2 respectively), ionomycin followed by DMAP (imme diately or after a 3-h delay, Groups 3 and 4), or no treatment (contro l, Group 5). They were then co-cultured in M199 with bovine oviductal epithelial cells. Activation rates were not significantly different am ong groups but significantly fewer oocytes in Group 3 extruded a secon d polar body than in Groups 1, 2, and 4. Most parthenotes (60% to 80%) in Groups 1, 2, and 4 were haploid, whereas 82% in Group 3 were mixop loid or polyploid. Most of the parthenotes (88%) in Group 4 formed a s ingle pronucleus besides extruding the second polar body and were ther efore more suitable for ICSI than those of Groups 1 and 2 in which con densed chromosomes predominated. The respective rates of oocyte cleava ge in Groups 1 to 4 were 24%, 36%, 70%, and 75%; corresponding blastoc yst rates were 1%, 5%, 17%, and 8%. There were significantly fewer cel ls in the parthenotes of Groups 1, 2, and 4 than of Group 3, or of emb ryos produced by in vitro fertilization. Thus, delaying the addition o f DMAP after ionomycin decreases chromosomal abnormalities and produce s a high proportion of activated oocytes suitable for ICSI. Mol. Repro d. Dev. 50:485-492, 1998. (C) 1998 Wiley-Liss, Inc.