The effect of tolbutamide on lipolysis and the intracellular concentration of cyclic AMP in vitro was examined in white fat cells isolated from epididymal adipose tissue of fasted rats. Incubations were performed in the presence and absence of glucose. Lipolysis was stimulated by theophylline and DL-arterenol plus theophylline. After eight minutes of incubation tolbutamide inhibited lipolysis induced by DL-arterenol plus theophylline but, in contrast to insulin, it increased the intracellular concentration of cyclic AMP. Tolbutamide also inhibits basal and theophylline- induced lipolysis but tolbutamine alone or in combination with theophylline had no appreciable effect on the intracellular concentration of cyclic AMP. The mechanism by which tolbutamide is antilipolytic and yet is able to increase the concentration of cyclic AMP is unknown. The fact that tolbutamide can inhibit lipolysis under conditions where cyclic AMP concentration is either increased or unchanged suggests that the antilipolytic effect...