Excess glutamine in embryo culture media alters mitochondrial and autophagy-related dynamics associated with impaired postnatal development.

The preimplantation period is a critical nutrient-sensing window influencing metabolic disease risk, as established by the developmental origins of health and disease theory (DOHaD). In this study, we examined the postnatal consequences of increased glutamine concentration, an important energy source, by altering embryonic metabolic development. In vivo-fertilised ICR mouse embryos were cultured in modified Chatot-Ziomek-Bavister medium with 1 mM L-glutamine (Gln) and bovine serum albumin (BSA) as the control and compared with elevated Gln media (2 mM) with bovine serum albumin (+Gln) or without BSA (+Gln-BSA). Early embryonic developmental analysis showed that both +Gln and +Gln-BSA presented reduced blastocyst cell number, primarily in trophectoderm cells, associated with increased apoptosis. Embryos showed altered metabolic homeostasis, including reduced mitochondrial membrane potential and mitochondrial morphology, altered autophagy-related activity (indicated by discrepancies between the DAPGreen assay and LC3-positive autophagosomes), and reduced heterochromatin methylation signals (H3K27me3). Offspring derived from these embryos subsequently exhibited irregular body weight trajectories, reduced survival, and impaired glucose tolerance. These findings indicate that in vitro exposure to elevated glutamine during early development is associated with altered mitochondrial dynamics and autophagy-related responses, interrupting intracellular mechanisms with lifelong metabolic consequences.