The kinetics of the permeation of cardiac muscle cells by glucose and the effect of insulin on this process were examined in the isolated perfused rat heart. Previous attempts to determine the parameters of permeation by glucose are considered to be imprecise because endogenous insulin influenced experi¬ mental results, intracellular glucose was inaccurately measured and estimates of glucose utilisation were not associated with defined concentrations of extracellular or intracellular glucose. In principle the parameters of permeation can be deter¬ mined from a steady state relationship between glucose uptake and the intracellular and extracellular concentration of glucose. Alternatively they can be determined from the relationship between the uptake and the extracellular concentration of glucose. if the kinetics of glucose metabolism are taken into account. A novel apparatus for cardiac perfusion was therefore developed in which hearts were brought to a steady state of glucose permeation and utilisation. In the apparatus hearts were perfused with a small volume of perfusate under well controlled conditions. Glucose concentration in perfusates was estimated by rapid and accurate automated methods which met the requirements of the procedure. The utilisation of glucose was determined not only in the steady state but also throughout the approach to that state. The first method for the determination of the parameters of permeation was impracticable because accurate estimates of intracellular glucose could not be made with methods currentlyavailable. Consequently the kinetics of glucose utilisation were compared with the predictions of a mathematical model in which permeation was assumed to be a simple carrier mechanism and the phosphorylation of glucose to be an irreversible enzymecatalysed reaction. Agreement was found. Estimates made of the parameters of permeation in the presence and absence of insulin were in qualitative accord with previous work. The hormone increased the half saturation constant at least 3-fold and the maximum rate of permeation 5-fold. The estimates of the parameters of permeation in the presence of insulin may be inaccurate because in their determination the possibility of a concentration gradient in the extracellular glucose was ignored. However, the estimates provide a reason¬ able explanation of a phenomenon which was observed when the time-course of glucose utilisation was studied in the absence of exogenous insulin. A stimulation of utilisation, presumably by endogenous insulin, occurred consistently only at concentra¬ tions of glucose greater than ImM. It was concluded that the estimates are an improvement on previous values and that the apparatus is a useful addition to the methods available for the study of the metabolism of isolated hearts.