KINETIC PARAMETERS OF RESPIRATION IN RAT PERMEABILIZED HEPATOCYTES UPON Ca2+ IN VARIOUS CONCENTRATIONS IN MEDIUM AND PROLONGED INFLUENCE OF TAURINE

Abstract

The dependence of permeabilized rat hepatocytes respiration rate on oxidation substrates concentration at presence 0.1 and 1 µM Са²⁺ in the medium and amino sulfonic acid – taurine – effect on this subordination was studied. Experimental animals were injected with taurine (40 mg/kg of weight) for 28 days and control rats were injected with equal volume of water. Liver cells were permeabilized with digitonin (20 µg per 1 million cells). Respiration rate was determined by polarographic method using Clark’s electrode, upon either succinate or pyruvate with malate oxidation. Hill equation parameters were calculated applying {v; v/[S]^h} coordinates. The kinetic dependence of respiration upon succinate and pyruvate oxidation is well described by Hill equation in the mediums with studied concentration of Са²⁺ in control and under taurine action. Hill coefficient h and semi activation constant K_0.5 for succinate, at rotenone presence, are not changed in cont­rol animals after increase of Са²⁺ сoncentration from 0.1 to 1 µM. Maximal velocity V_max was slightly increased under such conditions. Kinetic parameters of respiration upon succinate oxidation and both Са²⁺ concentration, are not changed significantly under prolonged taurine injection. There is more essential difference between kinetic parameters of pyruvate-stimulated respiration at various Ca²⁺ concentrations. Thus, in control, V_max is 1.5 times lower, and K_0.5 – 10.7 times major at 0.1 µM Са²⁺ in comparison with 1 µM Са²⁺. Hill coefficient becomes less than 1 at both Са²⁺concentration. As a result of taurine injection, V_max is 1.6 times less at low Са²⁺ and K_0.5 is higher, but only 6 times. Hill coefficient of this process increases at 0.1 µM Са²⁺ and is 1.12, upon taurine impact. Substrate inhibition, inherent to the dependence of respiration rate on pyruvate concentration within the medium with 1 µM Са²⁺, starts developing, under taurine effect, at higher concentration of this substrate in comparison with control (3 mM comparatively to 0.35 mM). Thus, Са²⁺ concentration rise from 0.1 to 1 µM stimulates the rat hepatocytes mitochondria respiration upon exogenous pyruvate oxidation, but not succinate. Taurine under the prolonged in vivo action does not influence succinate-stimulated rotenone-insensitive respiration and intensifies processes of pyruvate-stimulated oxygen consumption in hepatocytes.