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The development of preimplantation mouse embryos under different oxygen tensions in vitro
THERIOGENOLOGY
THE
DEVELOPMENT OF PREIMPLANTATION MOUSE EMBRYOS UNDER DIFFERENT OXYGEN TENSIONS IN VITRO P. Quinn and Gaye M. Harlow Department of Biological Sciences, University of Newcastle, New South Wales 2308.
The culture of preimplantation mammalian embryos in vitro has usually been performed using an atmosphere of 5% CO2 in air in accordance with the recommendations of Brinster (1). Under these conditions, however, the development of most mouse zygotes is blocked at the 2-cell stage, but if an atmosphere of 5% 02 : 5% CO : 90% N is used, zygotes develop into blastocysts during 4 days sf culturz in vitro (2). A study was made of the development of preimplantation mouse embryos to the blastocyst stage under various oxygen tensions in vitro. Embryos, at various stages of development from day 1 through to day 4, were collected from mated, superovulated Fl hybrid females and cultured in rubber-stoppered glass culture tubes in a simple, chemically defined culture medium (2). The optimal oxygen tension for development to the blastocyst stage was between 2.1/2% and 5%. One- and two-cell embryos, collected from females on days 1 and 2 of pregnancy respectively, had a more sharply defined range of oxygen tension capable of supporting development than embryos collected on days 3 ana 4. At all stages of development, however, more embryos developed to the blastocyst stage under 5% o2 compated to the numbers developing under higher oxygen tensions (20% and 40% 02). The numbers of blastomeres were counted in those embryos which developed to the blastocyst stage under 5% and 20% o . At each stage of development, those embryos developing to the blas s ocyst stage under 20% O2 had fewer blastomeres than the blastocysts which developed under 5% o*. As the time required for development to the blastocyst stage in vitro increased, there were fewer blastomeres present at the blastocyst stage. These results indicate that the cleaving mouse embryo has an optimal oxygen requirement in vitro of about 5%. As the oxygen tension is increased, fewer embryos develop to the blastocyst stage and in those which do develop, there are fewer cell divisions. It is recommended, therefore, that in studies of the fertilization and development of preimplantation mammalian embryos in vitro, consideration be given to the effects of oxygen on the embryos. An atmosphere containing 5% O2 would appear to be more beneficial to the embryo than one containinq 20% 02. REFERENCES 1.
Brinster, R.L. A method for in vitro cultivation of mouse ova fromtwo-cell to blastocyst. Expl. Cell Res.32: 205-208 (1963). -
2.
Whitten, W.K. Nutrient requirements for the culture of preimplantation embryos in vitro. Advances in the Bio Sciences -6: 129-141 (1971).
OCTOBER
1977
VOL. 8 NO. 4
161
THE
DEVELOPMENT OF PREIMPLANTATION MOUSE EMBRYOS UNDER DIFFERENT OXYGEN TENSIONS IN VITRO P. Quinn and Gaye M. Harlow Department of Biological Sciences, University of Newcastle, New South Wales 2308.
The culture of preimplantation mammalian embryos in vitro has usually been performed using an atmosphere of 5% CO2 in air in accordance with the recommendations of Brinster (1). Under these conditions, however, the development of most mouse zygotes is blocked at the 2-cell stage, but if an atmosphere of 5% 02 : 5% CO : 90% N is used, zygotes develop into blastocysts during 4 days sf culturz in vitro (2). A study was made of the development of preimplantation mouse embryos to the blastocyst stage under various oxygen tensions in vitro. Embryos, at various stages of development from day 1 through to day 4, were collected from mated, superovulated Fl hybrid females and cultured in rubber-stoppered glass culture tubes in a simple, chemically defined culture medium (2). The optimal oxygen tension for development to the blastocyst stage was between 2.1/2% and 5%. One- and two-cell embryos, collected from females on days 1 and 2 of pregnancy respectively, had a more sharply defined range of oxygen tension capable of supporting development than embryos collected on days 3 ana 4. At all stages of development, however, more embryos developed to the blastocyst stage under 5% o2 compated to the numbers developing under higher oxygen tensions (20% and 40% 02). The numbers of blastomeres were counted in those embryos which developed to the blastocyst stage under 5% and 20% o . At each stage of development, those embryos developing to the blas s ocyst stage under 20% O2 had fewer blastomeres than the blastocysts which developed under 5% o*. As the time required for development to the blastocyst stage in vitro increased, there were fewer blastomeres present at the blastocyst stage. These results indicate that the cleaving mouse embryo has an optimal oxygen requirement in vitro of about 5%. As the oxygen tension is increased, fewer embryos develop to the blastocyst stage and in those which do develop, there are fewer cell divisions. It is recommended, therefore, that in studies of the fertilization and development of preimplantation mammalian embryos in vitro, consideration be given to the effects of oxygen on the embryos. An atmosphere containing 5% O2 would appear to be more beneficial to the embryo than one containinq 20% 02. REFERENCES 1.
Brinster, R.L. A method for in vitro cultivation of mouse ova fromtwo-cell to blastocyst. Expl. Cell Res.32: 205-208 (1963). -
2.
Whitten, W.K. Nutrient requirements for the culture of preimplantation embryos in vitro. Advances in the Bio Sciences -6: 129-141 (1971).
OCTOBER
1977
VOL. 8 NO. 4
161