Kinetic parameters of respiration in rat permeabilized hepatocytes upon Са2+ 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 K0.5 for succinate, at rotenone presence, are not changed in cont­rol animals after increase of Са2+ сoncentration from 0.1 to 1 µM. Maximal velocity Vmax 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, Vmax is 1.5 times lower, and K0.5 – 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, Vmax is 1.6 times less at low Са2+ and K0.5 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.

Keywords

• hepatocytes
• mitochondria
• respiration
• calcium
• taurine