Intracellular oxygen metabolism during bovine oocyte and preimplantation embryo development

• Published in: Scientific reports Vol. 11; no. 1; pp. 21245 - 13
• Main Authors: McKeegan, Paul J., Boardman, Selina F., Wanless, Amy A., Boyd, Grace, Warwick, Laura J., Lu, Jianping, Gnanaprabha, Keerthi, Picton, Helen M.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 28.10.2021; Nature Publishing Group; Nature Portfolio
• Subjects: 631/136/1455; 631/80/2373; 631/80/642/333/1465; Abattoirs; Animals; Biomarkers; Blastocyst - metabolism; Blastomeres - metabolism; Carbonyl compounds; Cattle; Copy number; Cyanides; Embryonic Development - genetics; Embryos; Energy Metabolism; Female; Humanities and Social Sciences; Intracellular; Mammals; Mitochondria - genetics; Mitochondria - metabolism; Mitochondrial DNA; multidisciplinary; Oocytes; Oocytes - cytology; Oocytes - metabolism; Oxidative Phosphorylation; Oxygen; Oxygen - metabolism; Oxygen consumption; Pregnancy; Science; Science (multidisciplinary); Trophectoderm
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-021-99512-5

We report a novel method to profile intrcellular oxygen concentration (icO 2 ) during in vitro mammalian oocyte and preimplantation embryo development using a commercially available multimodal phosphorescent nanosensor (MM2). Abattoir-derived bovine oocytes and embryos were incubated with MM2 in vitro. A series of inhibitors were applied during live-cell multiphoton imaging to record changes in icO 2 associated with mitochondrial processes. The uncoupler carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) uncouples mitochondrial oxygen consumption to its maximum, while antimycin inhibits complex III to ablate mitochondrial oxygen consumption. Increasing oxygen consumption was expected to reduce icO 2 and decreasing oxygen consumption to increase icO 2 . Use of these inhibitors quantifies how much oxygen is consumed at basal in comparison to the upper and lower limits of mitochondrial function. icO 2 measurements were compared to mitochondrial DNA copy number analysed by qPCR. Antimycin treatment increased icO 2 for all stages tested, suggesting significant mitochondrial oxygen consumption at basal. icO 2 of oocytes and preimplantation embryos were unaffected by FCCP treatment. Inner cell mass icO 2 was lower than trophectoderm, perhaps reflecting limitations of diffusion. Mitochondrial DNA copy numbers were similar between stages in the range 0.9–4 × 10 6 copies and did not correlate with icO 2 . These results validate the MM2 probe as a sensitive, non-toxic probe of intracellular oxygen concentration in mammalian oocytes and preimplantation embryos.