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Succinate in hypoxia-reperfusion induced tissue damage.

A physiologically relevant substrate for in vitro assessment of mitochondrial ROS production and other functions in ischemia/hypoxia/reperfusion models.

 Starkov A.A.

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             It has long been established that succinate oxidation can support the highest rate of ROS production in mammalian mitochondria (reviewed in (Starkov 2008)). However, some researchers still dismiss the physiological significance of this fact. Their arguments are chiefly inspired by the widely used and as much widely misunderstood practice of using millimolar concentrations of succinate added together with Complex I inhibitor rotenone. The latter is added to prevent the inhibition of succinate dehydrogenase by intramitochondrially accumulated oxaloacetate. However, rotenone can be efficiently replaced with physiologically relevant glutamate which removes oxaloacetate trough the transamination reaction. Notably, such a replacement does not diminish the succinate-supported ROS production in mitochondria (Zoccarato et al. 2007). Regarding the succinate concentration, it is 0.20.5 mM in most tissue, much less than 25mM typically used in experiments with isolated mitochondria, indeed. However, tissue concentrations of succinate increase several folds to the millimolar range under ischemic and hypoxic conditions.

            It had been shown that in rat brain, 5 min of ischemia results in 8-10 folds decrease in concentrations of glycolytic intermediates and mitochondrial NAD-linked oxidative substrates pyruvate, citrate, alpha-ketoglutarate, oxaloacetate, fumarate, and malate. However, succinate concentration increased by ~300% to the millimolar range (Folbergrova et al. 1974; Benzi et al. 1979). To note, succinate remained moderately (~35%) elevated 15 min after the onset of the reperfusion, when all other metabolites recovered to the control levels (Benzi et al. 1979). Similar results were obtained in another study which also demonstrated that succinate in post-ischemic brain returns to the control levels only after 30 min reperfusion (Benzi et al. 1982). Another interesting finding is that hypoxia significantly (>60%) activated succinate and glutamate oxidation by isolated rat brain mitochondria (Khazanov et al. 1992; Poborskii 1997). Succinate has also been shown to inhibit the oxidation of pyruvate and other NAD-linked respiratory substrates and cause an over-reduction of mitochondrial pyridine nucleotides (Konig et al. 1969). Considering these findings an accumulation of succinate during hypoxia, its slow removal upon reperfusion, and its ability to inhibit the oxidation of NAD linked substrates it seems quite reasonable to suggest that succinate is the major substrate oxidized in vivo by mitochondria during first 30 min of reperfusion. Therefore, utilizing succinate in assays of mitochondrial functions in vitro is physiologically sound and in fact most relevant to studies on the role of mitochondria in hypoxia-reperfusion induced tissue damage, especially in experiments addressing mitochondrial ROS production.

         

References


Benzi, G., E. Arrigoni, F. Marzatico and R. F. Villa (1979). "Influence of some biological pyrimidines on the succinate cycle during and after cerebral ischemia." Biochem Pharmacol 28(17): 2545-50.


Benzi, G., O. Pastoris and M. Dossena (1982). "Relationships between gamma-aminobutyrate and succinate cycles during and after cerebral ischemia." J Neurosci Res 7(2): 193-201.


Folbergrova, J., B. Ljunggren, K. Norberg and B. K. Siesjo (1974). "Influence of complete ischemia on glycolytic metabolites, citric acid cycle intermediates, and associated amino acids in the rat cerebral cortex." Brain Res 80(2): 265-79.


Khazanov, V. A., A. N. Poborsky and M. N. Kondrashova (1992). "Air saturation of the medium reduces the rate of phosphorylating oxidation of succinate in isolated mitochondria." FEBS Lett 314(3): 264-6.

Konig, T., D. G. Nicholls and P. B. Garland (1969). "The inhibition of pyruvate and Ls(+)-isocitrate oxidation by succinate oxidation in rat liver mitochondria." Biochem J 114(3): 589-96.


Poborskii, A. N. (1997). "[Effect of research conditions on succinate oxidation in brain mitochondria in circulatory hypoxia]." Patol Fiziol Eksp Ter(1): 10-2.


Starkov, A. A. (2008). "The role of mitochondria in reactive oxygen species metabolism and signaling." Ann N Y Acad Sci 1147: 37-52.


Zoccarato, F., L. Cavallini, S. Bortolami and A. Alexandre (2007). "Succinate modulation of H2O2 release at NADH:ubiquinone oxidoreductase (Complex I) in brain mitochondria." Biochem J 406(1): 125-9.

Last Updated ( Wednesday, 16 September 2009 )
 

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