Diets enriched in monounsaturated fatty acids (MUFA)s have been shown to benefit glycemic control. Furthermore, MUFAs specifically stimulate secretion of the antidiabetic hormone, Glucagon-like peptide-1 (GLP-1) in vitro. To determine whether the MUFA-induced benefit in glycemic tolerance in vivo is due to increased GLP-1 release, lean Zucker rats were pair-fed a synthetic diet containing 5% fat derived from either olive oil (OO; 74% MUFA) or coconut oil (CO; 87% saturated fatty acids; SFA) for 2 weeks. Food intake and body weight gain were similar for both groups over the feeding period. The OO group had improved glycemic tolerance compared with the CO group in both oral and duodenal glucose tolerance tests [area under curve (AUC) 121 +/- 61 vs. 290 +/- 24 mM.120 min, P < 0.05; and 112 +/- 28 vs. 266 +/- 65 mM.120 min, P < 0.05, respectively]. This was accompanied by increased secretion of gut glucagon-like immunoreactivity (gGLI; an index of GLP-1 levels) in the OO rats compared with the CO rats (402 +/- 96 vs. 229 +/- 33 pg/ml at t = 10 min, P < 0.05). Tissue levels of GLP-1 and plasma insulin and glucagon levels were not different between the two groups. To determine the total contribution of GLP-1 to the enhanced glycemic tolerance in OO rats, the GLP-1 receptor antagonist exendin(9-39) (Ex(9-39)) was infused 3 min before a duodenal glucose tolerance test. Ex(9-39) abolished the benefit in glycemic tolerance conferred by OO feeding (OO+Ex(9-39) vs. CO+Ex(9-39), P = NS), and resulted in a deterioration of glycemic tolerance in the OO+Ex(9-39) group when compared with the OO controls (AUC 331 +/- 21 vs. 112 +/- 28 mM.120 min, P < 0.05). To probe the mechanism by which the OO diet enhanced GLP-1 secretion, a GLP-1-secreting L cell line was incubated for 24 h with either 100 microM oleic acid (MUFA) or 100 microM palmitic acid (SFA) and subsequently challenged with GIP, a known stimulator of the L cell. Preexposure to oleic acid but not to palmitic acid significantly increased GIP-induced GLP-1 secretion when compared with controls (55 +/- 12% vs. 34 +/- 9%, P < 0.01). These results demonstrate that the benefit in glycemic tolerance obtained with MUFA diets occurs in association with increased GLP-1 secretion, through a mechanism of enhanced L cell sensitivity. These results suggest that diet therapy with MUFAs may be useful for the treatment of patients with impaired glucose tolerance and/or type 2 diabetes through increased GLP-1 secretion.