Cortisol has a well-defined circadian rhythm. The aim of the study was to examine the effect of the morning rise in cortisol concentration on lipolysis in adipose tissue. Ten healthy subjects were studied on two occasions, and six of these were studied on three occasions. During the first two occasions, either a control or cortisol suppression study was performed by using metyrapone, and on the third occasion exogenous cortisol replacement was given in addition to metyrapone. Lipolysis in the subcutaneous adipose tissue of the anterior abdominal wall was studied by measurement of arteriovenous differences. Reduction in the early morning rise in cortisol led to significantly decreased venoarterialized differences for nonesterified fatty acids (P < 0.05) and glycerol (P < 0.01), attributable in part to decreased hormone-sensitive lipase (EC 22.214.171.124) action (P < 0.05) in adipose tissue. At the same time the arterialized plasma triacylglycerol concentration increased (P < 0.005) with a significant reduction in the adipose lipoprotein lipase (EC 126.96.36.199) rate of action (P < 0.05). In the replacement study, values were identical to those of the control study, showing that metyrapone had no nonspecific effects on lipolysis. We conclude that the morning rise in plasma cortisol concentration plays an important role in the regulation of lipolysis in adipose tissue in normal healthy adults.