The effect of exercise training on the antilipolytic action of insulin was studied in rat adipocytes. Exercise training enhanced lipolysis induced by norepinephrine. Insulin dose dependently inhibited norepinephrine- (1 microM) stimulated lipolysis in both groups. Its inhibition rate was significantly greater in the trained than in the control group. Thus, exercise training enhanced the antilipolytic action of insulin. In the control group, insulin (1,000 microU/ml) reduced the displacement rate of [3H]CGP-12177 binding to adipocytes by low concentrations of (-)-norepinephrine. The slope factor without insulin was 0.76, whereas that with insulin was 0.95. In the trained group, insulin did not affect the competition binding of (-)-norepinephrine for [3H]CGP-12177. The displacement rate of [3H]CGP-12177 binding from adipocytes by low concentrations of (-)-norepinephrine was significantly greater in the trained than in the control group. The number of surface beta-adrenergic receptors per adipocyte was smaller in the trained than in the control group. Cilostamide, which blocks the antilipolytic action of insulin, restored lipolysis in both groups. The recovery rate was significantly greater in the trained than in the control group. These findings suggest that the enhanced antilipolytic action by insulin in the trained group occurs at a site distal to the binding of norepinephrine to beta-adrenergic receptors and that it is due to the increased activity of particulate low-Michaelis constant phosphodiesterase.