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1 Medicine, University of Liverpool, Liverpool, United Kingdom
2 Institute of Gerontology, University of Michigan, Ann Arbor, Michigan, USA
3 Health Sciences Centre, University of Texas, San Antonio, Texas, USA
4 Paediatrics, University of California, San Francisco, California, USA
* To whom correspondence should be addressed. E-mail: mjj{at}liv.ac.uk.
Contractions of skeletal muscles produce increases in the concentrations of superoxide anions and activity of hydroxyl radicals in the extracellular space. The sources of these reactive oxygen species are not clear. We tested the hypothesis that following a demanding isometric contraction protocol, the major source of superoxide and hydroxyl radical activity in the extracellular space of muscles is the mitochondrial generation of superoxide anions and that with a reduction in MnSOD activity, the concentration of superoxide anions in the extracellular space is unchanged, but the concentration of hydroxyl radicals is decreased. For gastrocnemius muscles from 6-8 month old, adult wild type (Sod2+/+) and knockout mice heterozygous for the MnSOD gene (Sod2+/-), the concentrations of superoxide anions and hydroxyl radical activity were measured in the extracellular space by a microdialysis technique. A 15 min protocol of 180 isometric contractions induced a rapid and equivalent increase in the reduction of cytochrome c as an index of superoxide anion concentrations in the extracellular space of Sod2+/+ and Sod2+/- mice, whereas the hydroxyl radical activity measured by formation of 2,3 dihydroxybenzoate from salicylate increased only in the extracellular space of muscles of Sod2+/+ mice. The lack of a difference in the increase in superoxide anion concentration in the extracellular space of Sod2+/+ and Sod2+/- mice following the contraction protocol supported the hypothesis that the superoxide anions were not directly derived from the mitochondria. In contrast, the data obtained suggest that the increase in the hydroxyl radical concentration in the extracellular space of muscles from wild type mice following the contraction protocol, most likely results from degradation of hydrogen peroxide generated by MnSOD activity.
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