Dynamic 13C flux analysis captures the reorganisation of adipocyte glucose metabolism in response to insulin

Summary Cellular metabolism is dynamic, but quantifying non-steady metabolic fluxes by stable isotope tracers presents unique computational challenges. Here, we developed an efficient 13C-tracer dynamic metabolic flux analysis (13C-DMFA) framework for modelling central carbon fluxes that vary over time. We used B-splines to generalise the flux parameterisation system and to improve the stability of the optimisation algorithm. As proof-of-concept, we investigated how 3T3-L1 cultured adipocytes acutely metabolise glucose in response to insulin. Insulin rapidly stimulates glucose uptake, but intracellular pathways responded with differing speeds and magnitudes. Fluxes in lower glycolysis increased faster than upper glycolysis. Glycolysis fluxes rose disproportionally larger and faster than the TCA cycle, with lactate a primary glucose end-product. The uncovered array of flux dynamics suggests glucose catabolism is additionally regulated beyond uptake to help shunt glucose into appropriate pathways. This work demonstrates the value of using dynamic intracellular fluxes to understand metabolic function and pathway regulation.