Contact sites are created by fusion of the inner and outer mitochondrial membrane. They represent a dynamic microcompartment for creatine kinase activity. In this microenvironment the active sites of creatine kinase, oxidative phosphorylation, and ADP/ATP transport interact during basal and stimulated metabolism. Electron microscopic studies showed that the occurrence rate of contact sites was changing according to the energy state of the mitochondrion (Biermans et al., 1989). The aim of this study was to stimulate the metabolism of the rat heart by extracellular calcium and to investigate the formation of contact sites and creatine kinase activity. Isolated rat hearts were perfused with Krebs-Henseleit solution containing 1.6 mmol Ca. After 15 min stabilization the hearts were subjected to 0.6, 0.9, 1.8, 2.2, 2.6, or 3.6 mmol Ca for a period of 15 min. Stimulation of the isolated rat hearts with increasing calcium concentrations was physiologically manifested by enhanced dp/dt and LVP values. A higher frequency rate of mitochondrial contact sites was observed and these were associated with higher creatine kinase activity. We suppose that induction of contact site formation is related to the functional activity of the Ca stimulated rat heart and thus efficient production and transport of energy is provided. (Tab. 1, Fig. 6, Ref. 18.).