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Daniel Hass, Celia Bisbach, Elizabeth Giering, Thomas Mundinger, James Hurley; In vivo 13C-succinate infusion reveals gluconeogenesis in the mouse retina and eyecup. Invest. Ophthalmol. Vis. Sci. 2021;62(8):2228.
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Gluconeogenesis is a process wherein metabolites are diverted from the Krebs cycle to become glycolytic intermediates. Gluconeogenesis is critical for blood glucose homeostasis, and is thought to be restricted to liver, kidney, intestine, and muscle tissue. Previous liturature suggests that amphibian retinas are capable of this process, and our goal is to determine whether gluconeogenesis occurs in the mammalian retina.
We infused male and female C57BL6/J mice with 100mg/kg 13C4-succinate through jugular catheters and euthanized mice 1, 2, 3, 5, and 12 minutes following the infusion, dissecting and snap-freezing retina and eyecup tissue (EC; a complex of choroid, sclera, and RPE; n=3-5/tissue/time point). Un-infused mice served as a ‘0 minute’ control. At 0 and 5 minutes post-infusion, we also collected liver tissue (n=3).Frozen samples were derivatized with methoxyamine and TBDMS, and analyzed by gas-chromatography mass spectrometry to determine the abundance of 13C-labeled metabolites in the Krebs cycle (succinate, fumarate, malate, and citrate) and glycolysis (pyruvate, phosphoenolpyruvate, and 3-phosphoglycerate).
Infused 13C4-succinate populated the bloodstream (>95% of blood succinate was labeled) and led to succinate infiltration in to retina (8% of pool) and EC (57% of pool). Surprisingly, 13C from infused succinate was incorporated into 13C3-pyruvate, 13C3-phosphoenolpyruvate, and 13C3-3-phosphoglycerate in both tissues. These metabolites respectively accumulated at a rate of 0.2 (0.96), 0.13 (0.96), and 0.14 (0.92) % pool/minute (mean slope; R2) in the retina, and 0.1 (0.59), 0.07 (0.68), 0.09 (0.97) % pool/minute in the EC. There was no observable accumulation of 13C on glycolytic intermediates in liver, thus our observations are not explained by gluconeogenesis in the liver.
Retina succinate pools are far less labeled by succinate than ECs, yet in both tissues the diversion of Krebs cycle intermediates (succinate) to 13C labeled glycolytic metabolites occurred at a similar rate. These data suggest that (a) both tissues are capable of gluconeogenesis and (b) the retina diverts a larger fraction of 13C from succinate to gluconeogenesis than EC tissue. These findings are not recapitulated in liver tissue or ex vivo retina or EC tissue supplied with 13C4-succinate, suggesting that the cellular environment may dictate the activity of this pathway.
This is a 2021 ARVO Annual Meeting abstract.
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