The abnormal insulin secretion found in human diabetics and animal models of diabetes has been attributed to the deleterious effects of chronic hyperglycemia and/or elevated circulating levels of nonesterified fatty acids (NEFAs). In this study, abnormal glucose-induced insulin secretion (GIIS) was generated by a 48-hour infusion of glucose and assessed by the isolated perfused pancreas technique. In these hyperglycemic animals, abnormal GIIS is accompanied by a decrease in plasma NEFAs, while plasma and, more importantly, islet triglycerides remain at levels comparable to those in the controls. It is concluded that the abnormal insulin secretion in this glucose infusion model was likely caused by 48 hours of hyperglycemia and not by changes in circulating or islet lipids.