Adiponectin is a fat-derived hormone with insulin-sensitizing properties. In patients with type 2 diabetes plasma adiponectin levels are decreased. Since these patients are characterized by high plasma insulin and glucose concentrations, hyperinsulinemia and hyperglycemia could be responsible for the down-regulation of adiponectin. Insulin decreases adiponectin levels in humans. The effect of hyperglycemia is unknown. To determine the selective effects of insulin, glucose or its combination on plasma adiponectin, clamps were performed in six healthy males on four occasions in a cross-over design: (1=reference clamp) lower insulinemic-euglycemic clamp (insulin 100 pmol/L, glucose 5 mmol/L); (2) hyperinsulinemic-euglycemic clamp (insulin 400 pmol/L, glucose 5 mmol/L); (3) lower insulinemic-hyperglycemic clamp (insulin 100 pmol/L, glucose 12 mmol/L); (4) hyperinsulinemic-hyperglycemic clamp (insulin 400 pmol/L, glucose 12 mmol/L). Adiponectin concentrations and HMW (high-molecular-weight) to total adiponectin ratio were measured at the start and end of the 6-hour clamps. After the 6-hour study period, total plasma adiponectin levels were significantly (p=0.045) decreased by 0.63 µg/ml in the lower insulinemic-euglycemic clamp (clamp 1). In both euglycemic groups (clamp 1+2), adiponectin concentrations significantly declined (p=0.016) over time by 0.56 µg/mL, whereas there was no change in both hyperglycemic groups (clamp 3+4) (p=0.420). In neither of the clamps, the ratio of HMW to total adiponectin changed. We conclude that insulin suppresses plasma adiponectin levels already at a plasma insulin concentration of 100 pmol/L. Hyperglycemia prevents the suppressive effect of insulin. This suggests that, in contrast to glucose, insulin could be involved in the down-regulation of plasma adiponectin in insulin resistant patients.