Ca~(2+) Accumulation in Isolated Rat Heart Mitochondria under Maintenance of Mitochondrial Membrane Potential

摘要

It is known that mitochondria can accumulate calcium which regulates energy metabolism and cell death. In cardiomyocytes, about 90% of energy is generated just in the mitochondria. The heart cells are also affected by the rapid changes in intracellular Ca2+ concentration. Therefore, mitochondrial Ca2+-accumulation ability is crucial. The work was aimed at studying Ca2+ accumulation in isolated rat heart mitochondria under maintaining mitochondrial membrane potential and using different extramitochondrial Ca2+ concentrations. Isolated organelles were loaded with fluorescent dye Fluo-4 AM (2.5 umol/1) at a temperature of 26°C for 30 min. It has been revealed that in these conditions sufficiently high mitochondrial membrane potential is maintained, which is necessary for the functioning of calcium-transporting system in organelles. We have found that mitochondria are characterized by a limited ability to store ionized calcium, since adding of Ca2+ in concentrations of 10, 20, 50 umol/1 ensures a certain level of its accumulation in organelles with further cessation of fluorescent signal growth. Adding of 100 umol/1 Ca2+ to isolated mitochondria results in a significant increase in the fluorescence intensity (46% on the fifth minute, compared to thefluorescence induced by 20 umol/1 Ca2+ ) and presumably in the activation of cation release. It has been shown that ruthenium red (105 mol/1), an inhibitor of Ca2+-uniporter, prevents Ca2+ accumulation in organelles by 89% in the presence of 100 umol/1 Ca.2+ Heart mitochondria require Mg~(2+)-ATP complex (3mmol/l) to accumulate Ca,2+ to maintain the inner membrane potential, the activity of Ca2+ uniporter and energy processes in organelles. Thus, the process of Ca2+ accumulation in rat heart mitochondria requires the maintenance of mitochondrial membrane potential, the activity of Ca~(2+)-uniporter, it also depends on extramitochondrial Ca2+ concentration and the presence of Mg~(2+) ATP complex.