Identification of the signals for glucose-induced insulin secretion in INS1 (832/13) β-cells using metformin-induced metabolic deceleration as a model.

Metabolic deceleration in pancreatic β-cells is associated with inhibition of glucose-induced insulin secretion (GIIS), but only in the presence of intermediate/submaximal glucose concentrations. Here, we used acute metformin treatment as a tool to induce metabolic deceleration in INS1 (832/13) β-cells, with the goal of identifying key pathways and metabolites involved in GIIS. Metabolites and pathways previously implicated as signals for GIIS were measured in the cells at 2-25 mm glucose, with or without 5 mm metformin. We defined three criteria to identify candidate signals: 1) glucose-responsiveness, 2) sensitivity to metformin-induced inhibition of the glucose effect at intermediate glucose concentrations, and 3) alleviation of metformin inhibition by elevated glucose concentrations. Despite the lack of recovery from metformin-induced impairment of mitochondrial energy metabolism (glucose oxidation, O consumption, and ATP production), insulin secretion was almost completely restored at elevated glucose concentrations. Meeting the criteria for candidates involved in promoting GIIS were the following metabolic indicators and metabolites: cytosolic NAD/NADH ratio (inferred from the dihydroxyacetone phosphate:glycerol-3-phosphate ratio), mitochondrial membrane potential, ADP, Ca, 1-monoacylglycerol, diacylglycerol, malonyl-CoA, and HMG-CoA. On the contrary, most of the purine and nicotinamide nucleotides, acetoacetyl-CoA, HO, reduced glutathione, and 2-monoacylglycerol were not glucose-responsive. Overall these results underscore the significance of mitochondrial energy metabolism-independent signals in GIIS regulation; in particular, the candidate lipid signaling molecules 1-monoacylglycerol, diacylglycerol, and malonyl-CoA; the predominance of K/Ca signaling control by low ADP·Mg rather than by high ATP levels; and a role for a more oxidized state (NAD/NADH) in the cytosol during GIIS that favors high glycolysis rates.