PPAR¿ is obligatory for fat mass generation and is thought to determine the amount of TAG stored per fat cell. We investigated whether ligand availability for PPAR¿ is rate limiting in fat mass generation and substrate metabolism. Twenty healthy men (20¿29 years) were randomly assigned to receive the PPAR¿ ligand rosiglitazone (RSG) (8 mg/d) (n 10) or a placebo (n 10) during a stay of 7 d in a respiration chamber. Food intake was ad libitum, resulting in positive energy balances of 32·2 MJ (placebo) and 44·7 MJ (RSG). Fat cell size and expression of PPAR¿, adipocyte fatty acid-binding protein (aP2), adipsin, adiponectin and fasting-induced adipose factor (FIAF) were determined in subcutaneous abdominal fat biopsies. The total amount of fat stored and the amount of TAG per fat cell were not different between groups. For the entire group, fat cell size was decreased after overeating (P = 0·02). FIAF mRNA levels were decreased after overeating in the RSG group (P = 0·01), with a trend towards a decrease in the placebo group. Unexpectedly, RSG treatment did not influence the expression levels of PPAR¿ and of the PPAR¿ responsive genes aP2, adiponectin and adipsin. In addition, RSG resulted in a larger increase in plasma TAG during overeating than placebo treatment. These results suggest that in healthy, non-obese males the PPAR¿ ligand RSG influences TAG metabolism, independent of its PPAR¿ transcriptional activity in the subcutaneous adipose tissue.