Insights into the role of anaplerosis in insulin secretion: A 13C NMR study

Defining mechanisms and enzymatic paths critical to fuel-regulated insulin secretion are key goals of diabetes research. In this study, 13C-nuclear magnetic resonance spectroscopy and isotopomer analysis were used to investigate the link between insulin secretion and metabolic pathways associated with the tricarboxylic acid (TCA) cycle.

To this end, four insulinoma cell lines (betaTC3, betaTC-tet, INS-1 [832/13], R7T1) and porcine islets were examined under a variety of culture conditions (i.e. presence vs absence of amino acids and sera, and low vs high glucose).

Glucose consumption, insulin release, and glutamate isotopomeric patterns were influenced by media complexity (e.g. PBS, plain culture media, fully supplemented culture media). The 13C-labelled metabolites increased with media complexity and increasing glucose concentration, with the notable exception of aspartate, which was always higher under low-glucose conditions. The 13C-glutamate isotopomeric fractions were fitted to metabolic models to estimate the relative metabolic fluxes to the TCA cycle through key enzymatic processes. These indices of metabolism were compared with insulin secretion to determine correlative links. A model containing a single pool of pyruvate, an entrance to the TCA cycle via the pyruvate dehydrogenase complex, and two anaplerotic entrances, one through pyruvate carboxylase and another through an undefined (by the modelling program) source, provided the best fit to the data under all conditions tested, for all cell lines.

On the basis of our findings, a strong correlation may exist between stimulated insulin secretion and non-pyruvate carboxylase anaplerosis for the four cell lines examined in this study.