Objective We recently found that cAMP sensor Epac2A is a target of sulfonylurea (SU). To investigate the interaction of incretin and SU through Epac2A in insulin secretion, we examined the combinatorial effects of SU and incretin on insulin secretion and activation of Epac2A/Rap1 signaling. Methods We examined the combinatorial effects of SUs and GLP-1 on insulin secretion from perfused pancreata of wild-type and Epac2A-deficient mice. Activation of Rap1 by SUs and 8-pCPT-2’-O-Me-cAMP (8-pCPT), an Epac-selective cAMP analog, in insulin-secreting MIN6 cells was examined by GTP-Rap1 pull-down assay. Results In MIN6 cells, Epac2A was activated by glibenclamide (GLB) or glimepiride (GLM) but not gliclazide (GLC). At 4.4 mM glucose, GLB and GLM induced insulin secretion in a biphasic manner: a transient increase immediately after stimulation (early phase), followed by sustained release (late phase). In the presence of GLP-1, GLB-induced insulin secretion was markedly potentiated in both early and late phases in wild-type mice. In Epac2A-deficient mice, potentiation by GLP-1 of both phases in GLB-induced insulin secretion was markedly reduced. In contrast, there was no significant difference between wild-type and Epac2A-deficient mice in the potentiation by GLP-1 of GLC-induced insulin secretion. In MIN6 cells, Epac2A was synergistically activated by combination of GLB and 8-pCPT. Activation of Rap1 was enhanced by combination of 8-pCPT with GLB or GLM but not GLC. Conclusion Incretin and SU synergistically augment insulin secretion through Epac2A/Rap1 signaling. This synergistic effect depends upon structure of SU.