In vitro fertilisation of Chinese hamster oocytes by spermatozoa that have undergone ionophore A23187-induced acrosome reaction, and their subsequent development into blastocysts.
To enhance potential use of the Chinese hamster, Cricetulus griseus, in developmental and cytogenetic studies of mammalian gametes and embryos, techniques for in vitro fertilisation and embryo culture were developed in the species. Spermatozoa were recovered from the vasa deferentia of mature males, and incubated in modified TYH medium for 1 h at 37 degrees C under 5% CO2 in air. They were then treated with ionophore A23187 (20 microM) for 10 min to induce the acrosome reaction. Following ionophore treatment, superovulated oocytes were collected from hormonally stimulated females and incubated with the acrosome-reacted spermatozoa for 2 h at 37 degrees C under 5% CO2 in air. In this study, 245 oocytes were used for insemination, and 198 (80.8%) were found to be penetrated by sperm; among them, 194 ova (98.0%) were determined to be monospermic. The monospermic ova were then cultured in TYH supplemented with 1 mM hypotaurine under the same gas phase. Within 30 h of fertilisation, 182 ova (93.8%) cleaved to the 2-cell stage, and subsequently 163 ova (84.0%) developed beyond the 2-cell stage. Thus, obstinate developmental arrest at the 2-cell stage ('2-cell block') was not observed in this species. Ultimately, 65.5% of monospermic ova reached morula to blastocyst stages.
Tateno
H
H
Department of Biological Sciences, Asahikawa Medical College, Japan.
Kamiguchi
Y
Y
D000177:Acrosome / Q000187:drug effects*; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001755:Blastocyst / Q000166:cytology / Q000502:physiology*; D000001:Calcimycin / Q000494:pharmacology*; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D004622:Embryo, Mammalian / Q000166:cytology / Q000502:physiology; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods* / Q000706:statistics & numerical data; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D009865:Oocytes / Q000502:physiology; D013094:Spermatozoa / Q000187:drug effects* / Q000502:physiology
D000900:Anti-Bacterial Agents; D007476:Ionophores; D000001:Calcimycin