We previously showed in vitro that glutamate stimulates insulin release via an alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor. Here we address a more physiological question concerning the in vivo effect of intravenously or orally administered glutamate on insulinemia and glycemia in fed and fasted rats. In anesthetized fed rats, the intravenous administration of glutamate at 9 and 30 mg/kg transiently increased insulinemia in a dose-dependent manner. The insulin-secretory effect of glutamate (9 mg/kg) was blocked by an antagonist of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors. In anesthetized fasted rats, glutamate at 9 mg/kg was ineffective, but during an intravenous glucose tolerance test (0.5 g/kg), glutamate markedly potentiated insulin release and increased the glucose disappearance rate. In conscious rats, the intragastric administration of glutamate at 200 mg/kg elicited a transient insulin response in fed animals and had no effect in fasted animals but, during an oral glucose tolerance test (1 g/kg), enhanced insulin secretion and reduced the hyperglycemia. Glutamate was effective at plasma concentrations of 200-300 microM. In conclusion, intravenously and orally administered glutamate stimulates insulin secretion in vivo via an excitatory amino acid receptor and improves glucose tolerance.