Study of regulation of mitochondrial respiration in vivo. An analysis of influence of ADP diffusion and possible role of cytoskeleton.

摘要: Abstract The purpose of this work was to investigate the mechanism regulation mitochondrial respiration in vivo different muscles normal rat and mice, transgenic mice deficient desmin. Skinned fiber technique used study cells heart, soleus white gastrocnemius skeletal these animals. Also, cardiomyocytes were isolated from permeabilized by saponin “ghost” (phantom) produced extraction myosin with 800 mM KCl. Use confocal immunofluorescent microscopy anti-desmin antibodies showed good preservation mitochondria cytoskeletal system phantom cells. Kinetics ADP also studied detail before after binding anti-desmine intermediate filaments. In skinned cardiac or muscle fibers but not fast twitch kinetics characterized very high apparent K m (low affinity) equal 300–400 μM, exceeding that for factor 25. m. soleus, partial inhibition NaN 3 did decrease significantly, excluding possible explanation low affinity its rapid consumption due oxidative activity intracellular diffusion problems. However, short treatment trypsin decreased constant value 40–70 confirming earlier proposition sensitivity is controlled some cytoplasmic protein. Phantom which contain mostly cytoskeleton retain shape, values ADP. Therefore, they are probably most suitable studies cellular factors control function vivo. change heart desmin-deficient changes observed: two populations observed, one usually second Morphological observations electron confirmed existence distinct mice. results conform conclusion reason observed phosphorylation slow permeability outer membrane porins problems into inside Most probably, there a protein associated cytoskeleton, controls porin pores (VDAC) Desmin itself does display type pores, absence appearance disorganised structure altered population lacking unknown VDAC controlling Thus, may be functional connection between mitochondria, structural organisation still unidentified factor(s).