Development of in vitro matured/fertilized bovine embryos in a chemically defined medium: influence of oxygen concentration in the gas atmosphere.
The influence of oxygen concentrations in the gas atmosphere on the development of IVM/IVF bovine embryos was determined by culturing them in a microdrop of modified synthetic oviduct fluid medium supplemented with amino acids, insulin and PVA (mSOFai). After removing the cumulus cells at 18 hr post-insemination, presumptive zygotes were cultured in mSOFai for 104-106 hr under 5% CO2 with various O2 concentrations (2.5 to 20%). Reduced O2 (5-10%) improved the development to the morula stage, and 5% O2 gave the highest development. In the next experiment, morulae obtained after 102-104 hr of culture, were further cultured for 50 hr in mSOFai with 2mM glucose under 5 and 20% O2. An increase in the mean cell number in blastocysts, but not in the frequency of blastocysts, was observed under 5% O2. In the third experiment, zygotes were cultured for 152-154 hr in mSOFai under 5 and 20% O2, or cocultured with bovine oviduct epithelial cells in TCM199 + 10% FCS under 5% CO2 in air. Percentage of blastocysts for mSOFai in 5% O2 doubled to that for 20% O2, and was similar to that for coculture. Moreover, mean cell number in the blastocysts for mSOFai in 5% O2 was significantly higher than that for coculture. Results demonstrate that oxygen concentration critically affects embryonic development through zygotes to blastocysts, and suggest that around 5% 02 is optimal. It also indicates that bovine zygotes can be cultured up to the blastocyst stage using a chemically defined medium with rates similar to those of a conventional coculture system.
Department of Clinical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan.
Y
Y
Takahashi
M
M
Hishinuma
M
M
Matsui
H
H
Tanaka
H
H
Kanagawa
D000596:Amino Acids / Q000032:analysis / Q000494:pharmacology; D000818:Animals; D002417:Cattle / Q000196:embryology*; D003470:Culture Media / Q000032:analysis / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005314:Embryonic and Fetal Development / Q000187:drug effects* / Q000502:physiology; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods / Q000662:veterinary*; D007328:Insulin / Q000032:analysis / Q000494:pharmacology; D010100:Oxygen / Q000494:pharmacology*; D011247:Pregnancy; D015053:Zygote / Q000187:drug effects / Q000502:physiology
D000596:Amino Acids; D003470:Culture Media; D007328:Insulin; D010100:Oxygen