Activation regimens to prepare bovine oocytes for intracytoplasmic sperm injection

• Published in: Molecular reproduction and development 1998-08, Vol.50 (4), p.485-492
• Main Authors: Rho, G.J, Wu, B, Kawarsky, S, Leibo, S.P, Betteridge, K.J
• Format: Article
• Language: English
• Subjects: 6-dimethylaminopurine; activation; Adenine - analogs & derivatives; Adenine - pharmacology; Animals; Biological and medical sciences; bovine oocytes; Cattle; cell culture; cell number; chromosome number; Cleavage Stage, Ovum; Cytoplasm; embryogenesis; enzyme inhibitors; Female; fertilization (reproduction); Fertilization in Vitro - methods; Fundamental and applied biological sciences. Psychology; in vitro; injection; intracytoplasmic sperm injection; ionomycin; Ionomycin - pharmacology; ionophores; Ionophores - pharmacology; Male; Mammalian reproduction. General aspects; Microinjections; oocyte activation; oocyte culture; oocytes; Oocytes - drug effects; Oocytes - physiology; parthenogenesis; parthenote; parthenotes; polar bodies; pronucleus; Sperm-Ovum Interactions - drug effects; Sperm-Ovum Interactions - physiology; spermatozoa; Vertebrates: reproduction
• ISSN: 1040-452X; 1098-2795
• DOI: 10.1002/(SICI)1098-2795(199808)50:4<485::AID-MRD12>3.0.CO;2-1

Activation of bovine oocytes to produce a single haploid pronucleus in preparation for intracytoplasmic sperm injection (ICSI) has been investigated with various combinations of ionomycin and 6‐dimethylaminopurine (DMAP). Effects were evaluated by immunocytochemical staining, chromosomal analysis and assessment of development in vitro. Oocytes matured in vitro were exposed to: ionomycin alone (single or repeated treatments, Groups 1 and 2 respectively), ionomycin followed by DMAP (immediately or after a 3‐h delay, Groups 3 and 4), or no treatment (control, Group 5). They were then co‐cultured in M199 with bovine oviductal epithelial cells. Activation rates were not significantly different among groups but significantly fewer oocytes in Group 3 extruded a second 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 mixoploid or polyploid. Most of the parthenotes (88%) in Group 4 formed a single pronucleus besides extruding the second polar body and were therefore more suitable for ICSI than those of Groups 1 and 2 in which condensed chromosomes predominated. The respective rates of oocyte cleavage in Groups 1 to 4 were 24%, 36%, 70%, and 75%; corresponding blastocyst rates were 1%, 5%, 17%, and 8%. There were significantly fewer cells in the parthenotes of Groups 1, 2, and 4 than of Group 3, or of embryos produced by in vitro fertilization. Thus, delaying the addition of DMAP after ionomycin decreases chromosomal abnormalities and produces a high proportion of activated oocytes suitable for ICSI. Mol. Reprod. Dev. 50:485–492, 1998. © 1998 Wiley‐Liss, Inc.