Background: Postbariatric hypoglycemia (PBH) is a potentially serious complication after Roux-en-Y gastric bypass (RYGB), and impaired counterregulatory hormone responses have been suggested to contribute to the condition. Objectives: We evaluated counterregulatory responses during postprandial hypoglycemia in individuals with PBH who underwent RYGB. Setting: University hospital. Methods: Eleven women with documented PBH who had RYGB underwent a baseline liquid mixed meal test (MMT) followed by 5 MMTs preceded by treatment with (1) acarbose 50 mg, (2) sitagliptin 100 mg, (3) verapamil 120 mg, (4) liraglutide 1.2 mg, and (5) pasireotide 300 μg. Blood was collected at fixed time intervals. Plasma and serum were analyzed for glucose, insulin, glucagon, epinephrine, norepinephrine, pancreatic polypeptide (PP), and cortisol. Results: During the baseline MMT, participants had nadir blood glucose concentrations of 3.3 ±.2 mmol/L. At the time of nadir glucose, there was a small but significant increase in plasma glucagon. Plasma epinephrine concentrations were not increased at nadir glucose but were significantly elevated by the end of the MMT. There were no changes in norepinephrine, PP, and cortisol concentrations in response to hypoglycemia. After treatment with sitagliptin, 8 individuals had glucose nadirs <3.2 mmol/L (versus 4 individuals at baseline), and significant increases in glucagon, PP, and cortisol responses were observed. Conclusions: In response to postprandial hypoglycemia, individuals with PBH who underwent RYGB only had minor increases in counterregulatory hormones, while larger hormone responses occurred when glucose levels were lowered during treatment with sitagliptin. The glycemic threshold for counterregulatory activation could be altered in individuals with PBH, possibly explained by recurrent hypoglycemia.