Sucrose feeding reduces the ability of insulin to suppress glucose production and hepatic gluconeogenesis. The present study examined the effect of a high-sucrose diet on early insulin-signaling steps in the liver. Rats were provided a high-starch (STD, control diet) or high-sucrose diet (HSD) for 3 wk. On the day of study, overnight-fasted rats were anesthetized and injected with either saline (n = 5/diet group) or insulin (2 mU/kg, n = 5/diet group) via the portal vein. Portal venous blood and liver tissue were harvested 2 min after injections. Portal vein plasma glucose levels were not significantly different among groups, pooled average 147 +/- 12 mg/dl. Western blot analysis revealed no significant differences in the amount of insulin receptor (IR), insulin receptor substrates-1 and -2 (IRS-1, IRS-2), and the p85 subunit of phosphatidylinositol (PI) 3-kinase. In contrast, the amount of the p110beta subunit of PI 3-kinase was increased approximately 2-fold in HSD vs. STD (P < 0.05). After saline injection, tyrosine phosphorylation (pY) of IR, IRS-1, and IRS-2 was not significantly different between groups. However, PI 3-kinase activity associated with phosphorylated proteins was increased approximately 40% in HSD vs. STD (P < 0.05). After insulin injection, pY of the IR was not different between groups, whereas pY of IRS-1 and IRS-2 was reduced (P < 0.05) in HSD vs. STD. In addition, association of IRS-1 and IRS-2 with p85 was significantly reduced in HSD vs. STD. These data demonstrate that an HSD impairs insulin-stimulated early postreceptor signaling (pY of IRS proteins, IRS interaction with p85). Furthermore, the increased amount of p110beta and increased basal PI 3-kinase activity suggest a diet-induced compensatory response.