174. ATMOSPHERIC OXYGEN ALTERS THE EMBRYONIC METABOLOME AS QUANTIFIED BY CARBOHYDRATE UPTAKE AND AMINO ACID UTILISATION

Abstract

Oxygen regulates embryo development at both the cleavage and post compaction stages. In this study we investigated the effects of atmospheric oxygen during the preimplantation stages, on the post-compaction embryonic metabolome through quantification of glucose consumption and amino acid utilization. Zygotes obtained from F1 hybrid mice (C57BLxCBA) were randomly allocated to either 5 or 20% oxygen. In the first experiment, following the first 48 h of culture, embryos were cultured individually in 1 μL drops of modified G2 medium (0.5 mM glucose) and moved to fresh drops of medium every 24 h. The glucose concentration in the spent media samples, including controls containing no embryo, was determined by microfluorimetry. In the second experiment, embryos which had developed to the early blastocyst stage after 72 h were cultured for a further 24 h in groups of 10 in 2 μL drops of G2. Analysis of amino acid utilization was performed using liquid chromatography-triple quadrupole mass spectrometry. Glucose consumption by embryos cultured in 5% oxygen was significantly greater on day 4 and day 5 (4.89 ± 0.29 and 6.13 ± 0.41 pmol/embryo/h) compared to embryos cultured in 20% oxygen (2.59 ± 0.40 and 5.09 ± 0.28 pmol/embryo/h; P < 0.05). In contrast amino acid utilisation by embryos cultured in 5% oxygen was significantly less than embryos cultured in 20% oxygen (P < 0.05). The data generated will help to determine the aetiology of oxygen toxicity to the preimplantation embryo. Higher glucose utilisation by embryos in 5% oxygen is consistent with their improved development. Conversely, the increased utilisation of amino acids by blastocysts in 20% oxygen may reflect an adaptation to increased oxidative stress as a result of culture in a non-physiological oxygen concentration. This study demonstrates that atmospheric oxygen during the preimplantation period perturbs the embryonic metabolome which results in a compensatory increase in amino acid utilisation.