OBJECTIVE: Recent progress suggests that exenatide, a mimetic of glucagon-like peptide-1 (GLP-1), might lower glycemia independent of increased beta-cell response or reduced gastrointestinal motility. We aimed to investigate whether exenatide stimulates glucose turnover directly in insulin-responsive tissues dependent or independent of insulinemia. RESEARCH DESIGN AND METHODS: An intraportal glucose infusion clamp was used in dogs to measure glucose turnover to encompass potent activation of the putative glucose/GLP-1 sensor in the porto-hepatic circulation with exenatide. The modified glucose clamp was performed in the presence of postprandial hyperinsulinemia and hyperglycemia with exenatide (20 microg) or saline injected at 0 min. Furthermore, the role of hyperglycemia versus hyperinsulinemia in exenatide-mediated glucose disposal was studied. RESULTS: With hyperinsulinemia and hyperglycemia, exenatide produced a significant increase in total glucose turnover by approximately 30%, as indicated by portal glucose infusion rate (saline 15.9 +/- 1.6 vs. exenatide 20.4 +/- 2.1 mg x kg(-1) x min(-1), P < 0.001), resulting from increased whole-body glucose disposal (R(d), approximately 20%) and increased net hepatic uptake of exogenous glucose ( approximately 80%). Reducing systemic hyperglycemia to euglycemia, exenatide still increased total glucose turnover by approximately 20% (saline 13.2 +/- 1.9 vs. exenatide 15.6 +/- 2.1 mg x kg(-1) x min(-1), P < 0.05) in the presence of hyperinsulinemia, accompanied by smaller increments in R(d) (12%) and net hepatic uptake of exogenous glucose (45%). In contrast, reducing hyperinsulinemia to basal levels, exenatide-increased total glucose turnover was completely abolished despite hyperglycemia (saline 2.9 +/- 0.6 vs. exenatide 2.3 +/- 0.3 mg x kg(-1) x min(-1), P = 0.29). CONCLUSIONS: Exenatide directly stimulates glucose turnover by enhancing insulin-mediated whole-body glucose disposal and increasing hepatic uptake of exogenous glucose, contributing to its overall action to lower postprandial glucose excursions.