Induced parthenogenetic activation of oocytes of the marsupial Sminthopsis macroura.
Maturation of marsupial oocytes in vitro, an important step in the analysis of early developmental events, has a low success rate and results from the artificial activation of oocytes, which may not include nuclear maturation. In Sminthopsis macroura, 24-h culture of advanced antral follicles in medium containing 10 microg mL(-1) porcine pituitary luteinising hormone (LH) yielded 60% of mature polarised oocytes with the first polar body; follicles cultured in medium without LH yielded only immature oocytes. Parthenogenetic activation of follicular, oviducal or uterine oocytes occurred when a two-step protocol was used. Sixty-one oocytes, exposed to 10 microm calcium ionophore A23187 for 10 min followed by 10 microg mL(-1) cycloheximide (protein synthesis inhibitor) for 5 h and then cultured for 20-24 h, were scored for signs of activation, namely extrusion of the second polar body and formation of the pronucleus. In each of 43 oocytes (70%), the extruded second polar body was present. Sixteen oocytes were analysed on slides after fixation and staining and, in 13 oocytes (81%) in this group, the female pronucleus was visible. No activation occurred following incubation of oocytes in medium containing Sr(2+) for 5 h (n = 14), 8% ethyl alcohol solution for 8 or 12 min (n = 13) or 10 microm calcium ionophore A23187 (n = 13) for 10-20 min followed by culture for 20-24 h.
Maleszewski
Marek
M
Department of Embryology, Institute of Zoology, Warsaw University, Miecznikowa 1, Warszawa, Poland. maleszewski@biol.uw.edu.pl
Selwood
Lynne
L
D000818:Animals; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology*; D005260:Female; D007476:Ionophores / Q000494:pharmacology*; D007986:Luteinizing Hormone / Q000494:pharmacology; D008394:Marsupialia / Q000502:physiology*; D009865:Oocytes / Q000187:drug effects* / Q000502:physiology / Q000648:ultrastructure; D006080:Ovarian Follicle / Q000166:cytology; D010060:Ovulation; D010312:Parthenogenesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology*
D007476:Ionophores; D011500:Protein Synthesis Inhibitors; D000001:Calcimycin; D007986:Luteinizing Hormone; D003513:Cycloheximide; D002118:Calcium