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(15) BIOENERGETIC IMPORTANCE OF MITOCHONDRIAL OUTER MEMBRANE AS DIFFUSION BARRIERE UNDER NORMAL AND IMPAIRED CONDITIONS F.N. Gellerich, S. Trumbeckaite,
J.R. Opalka, H.N. Rasmussen and S. Zierz
Reconstituted systems were developed to study the effect of oncotic pressure on the exchange of metabolites across the porin pores between intermembrane space and cytosol. In reconstituted systems consisting of isolated mitochondria, dextrans (simulating cytosolic proteins) and pyruvate kinase (simulating glycolytic ADP depletion) we investigated the influence of oncotic pressure on rate dependent concentration gradients (up to 22 µM ADP) across the outer membrane (OM) and on ADP-transport into mitochondria by shuttles (mt-creatine kinase, mt-adenylate kinase) and by channels (mt-hexokinase). With increasing additions of dextrans (weight/volume) the direct diffusion of metabolites through OM decreases, but the ADP-transport via mitochondrial kinases increases. These dextran effects are mainly caused by increased formation of multiprotein complexes between the adenine nucleotide translocator, porin and mitochondrial kinases reinforcing dynamic compartmentation of ADP. If OM becomes leaky, dynamic compartmentation decreases resulting in diminished extramitochondrial phosphorylation potentials. In addition, apoptotic signal molecules as cytochrome c can be released. Therefore, the intactness of OM is an important indicator for acute impairments of mitochondria e.g. at ischemia/reperfusion or inflammation. By means of skinned fiber technique, leaks in OM as well as in inner membranes (IM) of mitochondria are detectable in heart and skeletal muscle. Loss of mitochondrial cytochrome c can be detected either by respiration measurements or spectrophotometrically in homogenates of skinned fibers. With these approaches the effect of endotoxin on mitochondrial function in rabbit hearts before and after ischemia/reperfusion was investigated. Endotoxin caused diminished state 3 respiration of heart mitochondria (- 40 %) but did not affect the intactness of OM and IM (permeability transition). However, increasing permeability transition and leaks in OM were detectable in skinned fibers after increasing time of ischemia. Endotoxin clearly reinforced the functional impairment of mitochondria caused by ischemia/reperfusion.
For further information contact...Carmen Mannella: carmen@wadsworth.org |
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