Obesity induced by long-term consumption of a fat-rich diet causes marked endothelial dysfunction. In this study we aimed to determine whether endothelial impairment is due to obesity or the diet per se. Wistar rats were fed either standard laboratory chow throughout (controls), or given a highly palatable diet (diet-fed) for 3 days, or fed the diet for 3 days and then returned to chow for 3 further days before sacrifice (diet-to-chow). Body weight, fat and gastrocnemius muscle mass, and plasma levels of glucose, insulin and leptin were all comparable between the three groups. Diet-fed rats had significantly raised plasma non-esterified fatty acids (NEFA; P=0.0005) and triglyceride levels (P=0.00001). The diet-to-chow group had intermediate plasma NEFA and triglyceride levels (significantly higher than in controls, P=0.019 and P=0.0035 for NEFA and triglycerides, respectively). There were no changes in noradrenaline and KCl responses in mesenteric arteries, whereas vasorelaxation to both carbamylcholine and sodium nitroprusside were significantly attenuated in the diet-fed group (by up to 18%; P=0.00001). Both these responses remained largely impaired in the diet-to-chow group. By contrast, histamine-induced vasorelaxation was comparable between all three groups. Thus, short-term feeding with a palatable diet induces marked endothelium-dependent and -independent arterial dysfunction. These effects occurred in the absence of obesity and largely persisted after removal of the palatable diet. Diet per se can have important detrimental effects on arterial function, which may be mediated by raised NEFA and/or triglyceride levels.