A high fructose diet induces insulin resistance in rats, although the exact molecular mechanism involved is unknown. In this study, we used immunoprecipitation and immunoblotting to examine the levels and phosphorylation status of the insulin receptor (IR) and insulin receptor substrate-1 (IRS-1), as well as the association of the IRS-I with phosphatidylinositol 3-kinase (PI 3-kinase), and phosphotyrosine phosphatase (SHP2) in the liver and muscle of rats fed a control or high fructose diet for 28 d. There were no differences in IR and the IRS-I protein levels in the liver and muscle of rats fed the control and high fructose diets. However, tyrosine-phosphorylation of the insulin receptor after insulin stimulation was reduced to 71 +/- 2% (P < 0.05) of control in the liver of the fructose-fed rats. In samples previously immunoprecipitated with anti-IRS-l antibody and blotted with antiphosphotyrosine antibody, the insulin-stimulated IRS-1 phosphorylation levels in the liver and muscle of the fructose-fed group were only 70 +/- 6% (P < 0.05) and 76 +/- 5% (P < 0.05) of those of control rats, respectively. The insulin-stimulated IRS-1 association with PI 3-kinase was reduced to 84 +/- 3% (P < 0.05) in the liver and to 84 +/- 4% (P < 0.05) in the muscle of the fructose-fed group compared with control rats. Insulin-stimulated IRS-1 association with SHP2 was reduced to 79 +/- 5% (P < 0.05) in liver of the fructose-fed rats. These data suggest that changes in the early steps of insulin signal transduction may have an important role in the insulin resistance observed in these rats.130615311535