Insulin secretion from pancreatic β-cells plays a central role in glucose homeostasis and impaired insulin secretion contributes to the development of diabetes. Metabolic signals in β-cells are crucial in regulation of insulin secretion. For example, ATP generated by glucose metabolism is a key signal in glucose-induced insulin secretion (GIIS), which is the most important mechanism of insulin secretion. In addition to GIIS, potentiation of GIIS also is required for normal regulation of insulin secretion. cAMP is a well-known second messenger in β-cells that potentiates insulin secretion in a glucose-dependent manner. Utilizing this effect, incretin (gut hormones that increase cAMP in the β-cells)-related anti-diabetic drugs have been developed recently. However, the mechanism of the glucosedependency of cAMP action in insulin secretion remains unknown. Using metabolome-based analysis, we find that glutamate the malate-aspartate (MA) shuttle, an NADH shuttle linked to glycolysis, is essential for potentiation of insulin secretion by incretin/cAMP signaling. We are currently identifying a novel metabolic signal in cAMP-induced insulin secretion. Metabolomics is a powerful approach to identification of novel metabolic signals and potential therapeutic targets in insulin secretion.