When stimulated with 6 mmol/L glucose, a minimal, transient insulin secretory response was observed from perifused rat islets. The inclusion of 5 micromol/L nateglinide significantly amplified release. Elevating glucose to 8 or 10 mmol/L resulted in an increasing insulin secretory response that was again markedly potentiated by the further inclusion of nateglinide. The calcium channel antagonist, nitrendipine, abolished secretion to 8 mmol/L glucose plus nateglinide. Unlike nateglinide, rosiglitazone (5 micromol/L), troglitazone (1 to 10 micromol/L), or darglitazone (10 micromol/L), 3 peroxisome proliferator-activated receptor gamma (PPARgamma) agonists, were without any acute stimulatory effect on insulin release in the simultaneous presence of 6 to 10 mmol/L glucose. Glucose (8 to 10 mmol/L) significantly increased inositol phosphate accumulation. Nateglinide amplified this response. Nitrendipine reduced inositol phosphate (IP) accumulation in response to the combination of 8 mmol/L glucose plus 5 micromol/L nateglinide. Rosiglitazone had no effect on IP accumulation. These results confirm the efficacy of nateglinide as a potent glucose-dependent insulin secretagogue that exerts its stimulatory effect, at least in part, through the activation of phospholipase C (PLC). No acute potentiating effect of rosiglitazone on either insulin secretion or IP accumulation could be detected in isolated rat islets.