The mitochondrial K+ cycle consists of influx and efflux pathways for K+ and anions. Net movement of K+ salts across the inner membrane causes changes of matrix volume, so regulation of the cycle is vital for maintaining the structural integrity of the organelle. The mitochondrial K+ cycle also appears to play important roles in cellular pathophysiology in vivo. Opening the mitochondrial ATP-sensitive K+ channel (mitoK(ATP)) prior to ischemia protects the heart from ischemia-reperfusion injury. MitoK(ATP) is an important player in the cell signaling pathways for ischemic protection and also for gene transcription, roles that appear to depend on the ability of mitoK(ATP) opening to trigger increased mitochondrial production of reactive oxygen species. MitoK(ATP) opening during both ischemia and reperfusion and during the high work state is found to preserve the structure of the intermembrane space and thereby maintains the normally low outer membrane permeability to adenine nucleotides. This review discusses the properties of the mitochondrial K+ cycle that help to understand the basis of these effects.