-The glucose-dependent secretion of the insulinotropic hormone glucagon-like peptide-1 (GLP-1) is a critical step in the regulation of glucose homeostasis. Two molecular mechanisms have separately been suggested as the primary mediator of intestinal glucose-stimulated GLP-1 secretion (GSGS): one is a metabotropic mechanism requiring the sweet taste receptor type 2 (T1R2) ϩ type 3 (T1R3) while the second is a metabolic mechanism requiring ATP-sensitive K ϩ (KATP) channels. By quantifying sugar-stimulated hormone secretion in receptor knockout mice and in rats receiving Roux-en-Y gastric bypass (RYGB), we found that both of these mechanisms contribute to GSGS; however, the mechanisms exhibit different selectivity, regulation, and localization. T1R3Ϫ/Ϫ mice showed impaired glucose and insulin homeostasis during an oral glucose challenge as well as slowed insulin granule exocytosis from isolated pancreatic islets. Glucose, fructose, and sucralose evoked GLP-1 secretion from T1R3 ϩ/ϩ , but not T1R3 Ϫ/Ϫ , ileum explants; this secretion was not mimicked by the K ATP channel blocker glibenclamide. T1R2 Ϫ/Ϫ mice showed normal glycemic control and partial small intestine GSGS, suggesting that T1R3 can mediate GSGS without T1R2. Robust GSGS that was K ATP channeldependent and glucose-specific emerged in the large intestine of T1R3 Ϫ/Ϫ mice and RYGB rats in association with elevated fecal carbohydrate throughout the distal gut. Our results demonstrate that the small and large intestines utilize distinct mechanisms for GSGS and suggest novel large intestine targets that could mimic the improved glycemic control seen after RYGB.glucagon-like peptide-1; insulin; T1R3; glucose-stimulated potassium ion channel; enteroendocrine l cells THE BODY TIGHTLY REGULATES blood glucose levels, and disruption of the homeostatic mechanisms that underlie normal glycemic control can have significant deleterious effects. For example, the prolonged hyperglycemia associated with type 2 diabetes mellitus (T2DM) increases the risk of cardiovascular disease, neuropathy, retinopathy, kidney disease, and death (66). Hormonal signals arising in the gastrointestinal tract are key components of the homeostatic mechanisms controlling blood glucose levels after a meal. Ingestion of carbohydrate and other nutrients promotes the secretion of insulinotropic hormones such as glucagon-like peptide-1 (GLP-1) from the gut, resulting in a surge of insulin production before blood glucose levels rise (11,32). This early response contributes to increased glucose disposal during absorption and helps to prevent hyperglycemia. GLP-1 mimetics and inhibitors of GLP-1 degradation help increase insulin biosynthesis and secretion from pancreatic -cells and are valuable additions to previous treatment regimens for T2DM patients (11,32).Despite the importance of intestinal glucose sensing and glucose-stimulated gut hormone secretion, the mechanisms underlying these processes have remained elusive. The distinct glucose-sensing mechanisms found in the pancreas and in the gustat...