Coixol amplifies glucose-stimulated insulin secretion via cAMP mediated signaling pathway

Recently, we reported the role of coixol (6-methoxy-2(3H)-benzoxazolone), an alkaloid from Scoparia dulcis, in glucose-dependent insulin secretion; however, its insulin secretory mechanism(s) remained unknown. Here, we explored the insulinotropic mechanism(s) of coixol in vitro and in vivo. Mice islets were batch incubated, perifused with coixol in the presence of agonists/antagonists, and insulin secretion was measured by ELISA. Intracellular cAMP levels were measured using enzyme immunoassay. K⁺- and Ca²⁺-currents were recorded in MIN6 cells using whole-cell patch-clamp technique. The in vivo glucose tolerance and the insulinogenic index were evaluated in diabetic rats treated with coixol at 25 and 50 mg/kg, respectively. Coixol, unlike sulfonylurea, enhanced insulin secretion in batch incubated and perifused islets at high glucose, with no effect at basal glucose concentrations. Coixol showed no pronounced effect on the inward rectifying K⁺- and Ca²⁺-currents in whole-cell patch recordings. Moreover, coixol-induced insulin secretion was further amplified in the depolarized islets. Coixol showed an additive effect with forskolin (10 μM)-induced cAMP level, and in insulin secretion; however, no additive effect was observed with isobutylmethylxanthine (IBMX, 100 μM)-induced cAMP level, nor in insulin secretion. The PKA inhibitor H-89 (50 μM), and Epac2 inhibitor MAY0132 (50 μM) significantly inhibited the coixol-induced insulin secretion (P < 0.01). Furthermore, insulin secretory kinetics revealed that coixol potentiates insulin secretion in both early and late phases of insulin secretion. In diabetic animals, coixol showed significant improvement in glucose tolerance and on fasting blood glucose levels. These data suggest that coixol amplifies glucose-stimulated insulin secretion by cAMP-mediated signaling pathways.