Abstract A comparative study of the transport of pyruvate in heart mitochondria from normal and triiodothyronine-treated rats has been carried out. It has been found that the rate of carrier-mediated (α-cyanocinnamate-sensitive) pyruvate uptake is significantly enhanced in mitochondria from triiodothyronine-treated rats as compared with mitochondria from control rats. The kinetic parameters of the pyruvate uptake indicate that only the V max of this process is enhanced whilst there is no change in the K m value. The enhanced rate of pyruvate uptake is not dependent on the increase of the transmembrane ΔpH value (both mitochondria from normal and triiodothyronine-treated rats exhibit the same ΔpH value) neither does it depend on the increase of the pyruvate carrier molecules (titration of these last with α-cyanocinnamate gives the same total number of binding sites). The pyruvate-dependent oxygen uptake is stimulated by 35–40% in mitochondria from hyperthyroid rats when compared with mitochondria from control rats. There is, however, no difference in either the respiratory control ratios or in the ADP O ratios between these two types of mitochondria. The heart mitochondrial phospholipid composition is altered significantly in hyperthyroid rats; in particular, negatively charged phospholipid such as cardiolipin and phosphatidylserine were found to increase by more than 50%. Minor alterations were found in the pattern of fatty acids with an increase of 20:4 18:2 ratio. It is suggested that the changes in the kinetic parameters of pyruvate transport in mitochondria from hyperthyroid rats involve hormone-mediated changes in the lipid composition of the mitochondrial membranes which in turn modulate the activity of the pyruvate carrier.