Acute changes in blood glucose concentration have major effects on gastrointestinal motor function. Patients with diabetes mellitus have an increased prevalence of gastroesophageal reflux. Transient lower esophageal sphincter (LES) relaxation (TLESR) is the most common sphincter mechanism underlying reflux. The aim of this study was to investigate the effect of acute hyperglycemia on triggering TLESRs evoked by gastric distension in healthy volunteers. TLESRs were stimulated by pressure-controlled and volume-controlled (500 ml) gastric distension using an electronic barostat and performed on separate days. On each day, esophageal manometry was performed in the sitting position during gastric distension for 1 h under euglycemia (5 mM), and either marked hyperglycemia (15 mM) or physiological hyperglycemia (8 mM) in randomized order was maintained by a glucose clamp. Marked hyperglycemia doubled the rate of TLESRs in response to both pressure-controlled [5 (3-10.5, median or interquartile range) to 10 (9.5-14.5) per hour, P < 0.02] and volume-controlled [4 (2.5-7.5) to 10.5 (7-12.5) per hour, P < 0.02] gastric distension but had no effect on basal LES pressure. Physiological hyperglycemia had no effect on the triggering of TLESRs or basal LES pressure. In healthy human subjects, marked hyperglycemia increases the rate of TLESRs. Increase in the rate of TLESRs is independent of proximal gastric wall tension. Mechanisms underlying the effect remain to be determined. Hyperglycemia may be an important factor contributing to the increased esophageal acid exposure in patients with diabetes mellitus.