To investigate the cellular mechanism of insulinotropic effect of glutamate in pancreatic beta cells, we utilized patch-clamp technique to monitor directly the activities of ATP-sensitive potassium channels (K-ATP channels). Dimethylglutamate (5 mM), a membrane-permeable analog of glutamate, augmented the insulin release induced by the stimulatory concentrations of glucose (p < 0.05-0.01). In the cell-attached configurations, dimethylglutamate reversibly and significantly suppressed the K-ATP channel activities (P < 0.01). On the other hand, no significant effect was observed when glutamate itself was applied to the inside-out patches, whereas the prompt and reversible suppression was recorded in the case of ATP (p < 0.01). These results indicate that the insulinotropic action of glutamate in beta cells could be derived from the inhibition of K-ATP channel activities, probably due to generation of messengers via intracellular metabolism such as ATP. (C) 2003 Elsevier Inc. All rights reserved.