Congenital hyperinsulinism is a disorder of pancreatic ␤-cell function characterized by failure to suppress insulin secretion in the setting of hypoglycemia, resulting in brain damage or death if untreated. Loss-of-function mutations in the K ATP channel (composed of two subunits: Kir6.2 and SUR-1) are responsible for the most common and severe form of congenital hyperinsulinism. Most patients are unresponsive to available medical therapy and require palliative pancreatectomy. Similar to the human condition, the SUR-1 ؊/؊ mouse is hypoglycemic when fasted and hyperglycemic when glucose-loaded. We have previously reported that the glucagon-like peptide-1 receptor antagonist exendin-(9 -39) raises fasting blood glucose in normal mice. Here we examine the effect of exendin-(9 -39) on fasting blood glucose in SUR-1 ؊/؊ mice. Mice were randomized to receive exendin-(9 -39) or vehicle. Fasting blood glucose levels in SUR-1 ؊/؊ mice treated with exendin-(9 -39) were significantly higher than in vehicle-treated mice and not different from wild-type littermates. Exendin-(9 -39) did not further worsen glucose tolerance and had no effect on body weight and insulin sensitivity. Isolated islet perifusion studies demonstrated that exendin-(9 -39) blocked amino acid-stimulated insulin secretion, which is abnormally increased in SUR-1 ؊/؊ islets. Furthermore, cAMP content in SUR-1 ؊/؊ islets was reduced by exendin-(9 -39) both basally and when stimulated by amino acids, whereas cytosolic calcium levels were not affected. These findings suggest that cAMP plays a key role in K ATP -independent insulin secretion and that the GLP-1 receptor is constitutively active in SUR-1 ؊/؊ ␤-cells. Our findings indicate that exendin-(9 -39) normalizes fasting hypoglycemia in SUR-1mice via a direct effect on insulin secretion, thereby raising exendin-(9 -39) as a potential therapeutic agent for K ATP hyperinsulinism.Congenital hyperinsulinism is a genetic disorder of pancreatic ␤-cell function characterized by failure to suppress insulin secretion in the setting of hypoglycemia, resulting in brain damage or death if inadequately treated. Loss-of-function mutations in the K ATP -sensitive channel (composed of two subunits: Kir6.2 and SUR-1) are responsible for the most common and severe form of hyperinsulinism (K ATP HI). K ATP -sensitive channels couple the metabolic state of the ␤-cell to membrane potential by sensing changes in intracellular ATP concentration. In pancreatic ␤-cells, closure of the channel in response to elevation of the ATP/ADP ratio following stimulation with glucose leads to depolarization of the membrane and activation of voltage-dependent calcium channels with resultant exocytosis of insulin-containing granules (1). Thus, K ATP -sensitive channels play a critical role in the triggering pathway of glucosestimulated insulin secretion (2). In contrast, the amplifying pathway of insulin release operates independently of K ATP -sensitive channels and functions to augment fuel-stimulated insulin secretion (3). Electrophysiologi...