РОЛЬ РІАНОДИНЧУТЛИВИХ Са2+-КАНАЛІВ У РЕГУЛЯЦІЇ ДИХАННЯ МІТОХОНДРІЙ ПЕЧІНКИ ЩУРІВ

Abstract

The role of mRyRs in regulation of rats hepatocytes mitochondrial respiration has been investigated at different submitochondrial concentrations of Ca 2+ and by oxidation of succinate, pyruvate and α-ketoglutarate. Isolated mitochondria were obtained by differential centrifugation method. The intactness of the received mitochondria was determined by electron microscopy method. As an indicator of the absence of plasma membrane residues and endoplasmic reticulum membranes, the ATPase activity of the mitochondria suspension has been defined based on changes in the content of inorganic phosphate in the medium after the incubation of mitochondria with eosin Y, oubain and tapsigargin. The rate of respiration has been determined by polarographic method with Clark’s electrode using. To identify the role of mRyRs in mitochondrial respiration regulation, the mitochondria suspension was preincubated with ryanodine at a concentration of 0.05 μmol/l or ryanodine was introduced at a concentration of 0.1 μmol/l directly into a polarographic chamber. Decreasing of mitochondrial oxygen consumption has been specified after the preincubation of the mitochondria suspension with ryanodine for 5 and 10 minutes by the succinate oxidation. Due to the direct application of ryanodine to mitochondria in a polarographic chamber with a Ca2+ concentration in the medium of 0.1 and 1 μmol/l by succinate oxidation, the rate of oxygen consumption decreased in the states S 4 , S 3 and S 4 ATP . The preincubation of mitochondria with ryanodine by pyruvate oxidation, caused a decreasing of oxygen consumption, as after the direct addition of ryanodine to a polarographic chamber with a concentration of Ca2+ in a medium of 0.1 μmol/l. At the concentration of Ca2+ in the medium of 1 μmol/l increasing of oxygen consumption has been detected under the influence of ryanodine in the states S 4 , S 3 by 12.2 and 17.2 % respectively. The mitochondrial oxygen consumption by the α-ketoglutarate oxidation after the addition of ryanodine to a polarographic chamber with a concentration of Ca2+ 0.1 μmol/l in the medium has increased in the state S 4 , and S 3 , while in the respiratory medium with a concentration of Ca2+ 1 μmol/l has increased in the state S 4 by 19.1 % and decreased by 14.6 % in state S 4 ATP . Data analysis suggests that mRyRs in the hepatocytes mitochondria is an important link in the adaptive regulatory bond between the value of membrane potential of mitochondria and the intensity of their respiration at low values of the non-mitochondrial Ca2+ concentration and pyruvate oxidation. If the oxidation of α-ketoglutarate predominates, or the sub-mitochondrial concentration of Ca2+ is high, its mRyRs yields inhibition of respiration.