Control of glycolysis and fatty acid oxidation in ischemic myocardium was studied in isolated working rat hearts. Coronary flow was reduced to the whole heart. In ischemic tissue, oxygen consumption, glycolysis and fatty acid oxidation all decreased in proportion to the restriction in coronary flow. Inhibition of glycolysis developed at the level of glyceraldehyde-3-phosphate dehydrogenase. Restricted flux through this step appeared to result from accumulation of lactate, H+ and NADH. The rate of glycolysis was inversely related to accumulation of lactate. Additions of high levels of lactate to the perfusate inhibited glycolysis in aerobic, anoxic and ischemic hearts. The mechanism of this effect of lactate in anaerobic hearts is unknown, but does not appear to be related to pH changes. Oxidation of fatty acids was restricted at the level of beta-oxidation and high levels of both long-chain acyl CoA and carnitine derivatives accumulated.