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Perturbations in Mouse Embryo Development and Viability Caused by Ammonium Are More Severe after Exposure at the Cleavage Stages¹

Research Centre for Reproductive Health,³ Department of Obstetrics and Gynaecology, University of Adelaide, Woodville, South Australia 5011, Australia Repromed,⁴ Dulwich, South Australia 5065, Australia

ABSTRACT

The presence of ammonium in culture medium has a detrimental effect on embryo physiology and biochemistry; however, the stage at which the embryo is most sensitive to this effect is unknown. The aim of this study was to determine the exact stage at which the embryo is most vulnerable to ammonium by exposing the preimplantation embryo to 300 µM ammonium either at the precompaction stage (between the zygote and two-cell or the two-cell to eight-cell) or at the postcompaction stage (between the eight-cell and blastocyst). This study determined that exposure of embryos to ammonium at the precompaction stage from either the zygote to two-cell stage or from the two-cell to the eight-cell stage did not affect the rate of development to the blastocyst stage; however, the resultant blastocysts had decreased cell numbers and inner cell mass cells. Furthermore, these blastocysts had increased levels of cellular apoptosis and perturbed levels of Slc2a3 expression and glucose uptake. Transfer of these blastocysts revealed that, while implantation was not affected, the number of fetuses was reduced by culture with ammonium at the precompaction stage and fetal development was delayed, as observed by reduced crown-rump length and maturity. In contrast, the later stage embryo was more resistant to the negative effects of ammonium, with only Slc2a3 expression and fetal maturity affected. This raises the possibility that the later stage embryo is more able to protect itself from in vitro-derived stress and that the majority of in vitro-induced damage to mouse embryos is inflicted at the early stages of development.

assisted reproductive technology, early development, embryo, gene regulation, in vitro fertilization

¹ Supported by the Australian NHMRC project grant 299037. D.L.Z. is a recipient of a TQEH postgraduate scholarship, J.G.T. a NHMRC Senior Research Fellowship, and M.L. is a recipient of a R.D. Wright Fellowship from the NHMRC.

² Correspondence: Michelle Lane, Research Centre for Reproductive Health, University of Adelaide, Level 4, Maternity Building, The Queen Elizabeth Hospital, Woodville Road, Woodville, South Australia 5011, Australia. FAX: 61 8 8222 7521; michelle.lane{at}adelaide.edu.au

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