OBJECTIVE Based on the hypothesis that provision of glucose is good and fatty acids are bad for the ischaemic myocardium, the aims of this study were to determine i) the effects of different substrates on sarcolemmal permeability during normoxia, low-flow hypoxia (HLF) and reperfusion, ii) whether increased membrane permeability is associated with ultrastructural damage and increased influx of Ca2+ into cells and iii) whether changes in membrane permeability correlate with myocardial function and high energy phosphate metabolism. METHODS The isolated rat heart subjected to HLF was used as model of global ischaemia, and sarcolemmal permeability assessed by release of LDH from and influx of lanthanum and Ca2+ into myocardial tissue. Myocyte structural injury was also evaluated quantitatively, and mechanical activity was monitored throughout the experimental protocol. RESULTS Regardless of the substrate used, HLF caused a 80-90% and 20-40% reduction in myocardial oxygen uptake and coronary flow rate, respectively. Palmitate (0.5 mM conjugated to 0.1 mM albumin) or substrate-free perfusion caused ultrastructural damage and loss of normal sarcolemmal integrity during both normoxia and HLF. Although reperfusion reversed injury in some cells, in general, myocytes exhibited myofibrillar contracture, while membrane integrity recovered to some extent, as indicated by reduced lanthanum influx. Intracellular Ca2+ increased significantly upon reperfusion. Mechanical function as well as tissue high energy phosphates were significantly depressed during both HLF and reperfusion. Glucose, on the other hand, protected against ischaemia-induced structural damage and loss of sarcolemmal integrity. Reperfusion in these experiments resulted in almost complete recovery of normal morphology, ultrastructure and sarcolemmal integrity, while intracellular Ca2+ remained unchanged. Mechanical function and tissue high energy phosphates were significantly higher in glucose-perfused hearts than in palmitate-perfused or substrate-free hearts. Glucose was also able to attenuate the harmful effects of palmitate on myocardial ultrastructure, membrane integrity, mechanical function, energy metabolism and prevented Ca2+ overloading during reperfusion. CONCLUSIONS The results provide new evidence for the protective role of glucose during myocardial ischaemia and reperfusion. Although the exact mechanism of the beneficial actions of glucose remains to be established, the results suggest that glycolytic flux and thus glycolytically derived ATP protect against ischaemic damage via preservation of membrane integrity.