The effects of insulin and leptin on fatty acid uptake in differentiated (adipocytes) and undifferentiated 3T3-L1 cells were investigated. It was demonstrated that in undifferentiated 3T3-L1 cells, insulin and leptin have no effect on fatty acid uptake. In differentiated 3T3-L1 adipocytes, insulin had a concentration-dependent stimulatory effect on fatty acid uptake, whereas leptin on its own had no effect. Leptin, when coincubated with 10 nmol/L insulin, resulted in a concentration-dependent inhibition of the insulin-stimulated fatty acid uptake in differentiated 3T3-L1 cells. These results indicate that leptin has a direct inhibitory effect on the stimulation of fatty acid uptake by insulin in differentiated murine adipocytes.