Reappraisal of the e.p.r. signals in (post)-ischaemic cardiac tissue

Abstract

The present study was designed to measure directly, using e.p.r. spectroscopy, oxygen-derived free radicals in (post)-ischaemic or (post)-anoxic rat hearts. Rat hearts were rapidly freeze-clamped at 77 K under normoxic, anoxic, ischaemic or reperfusion conditions. The samples were measured at three different temperatures (13, 77 and 115 K) and at several microwave power levels, and were compared with isolated rat heart mitochondria. Samples were prepared both by grinding and as tissue cuts. The two preparation techniques gave identical e.p.r. results, which excludes the occurrence of grinding artifacts. No free radical signals linked to reperfusion injury were detected. Several electron transfer centres known in the mitochondrial respiratory chain were measured. The signals previously assigned to post-ischaemic reperfusion injury were found to originate from electron transfer centres of the respiratory chain, predominantly the iron-sulphur cluster S-1 in succinate dehydrogenase. The differences in signal intensity between normoxic, ischaemic and reperfused hearts were found to result from the different redox stages of these centres under the various conditions tested. These findings do not necessarily imply that oxygenderived free radicals are not formed in cardiac tissue during (post)-ischaemic reperfusion. The constitutive background of paramagnetism from the respiratory chain, however, seriously hampers the direct detection of comparatively low concentrations of free radicals in cardiac tissue. It is therefore expedient to focus future experiments in this field on the use of spin-trapping agents.

Caloric restriction improves thermotolerance and reduces hyperthermia‐induced cellular damage in old rats

“…The relatively narrow line at g ϭ 2.005 (⌬Hpp Ϸ10 -15 G) is highly responsive to microwave power; the intensity of this line decreases relative to all other spectral lines with increasing microwave power (data not shown). This response, coupled with the g value and line width, is consistent with the g ϭ 2.005 species being an organic radical, such as a semiquinone radical (30). The broad features at g ϭ 2.25 and 1.935 are indicative of iron(III), possibly from mitochondrial succinate dehydrogenase (31).…”

Hyperthermia stimulates nitric oxide formation: electron paramagnetic resonance detection of .NO-heme in blood

“…The features at g = 2.021 and 2.005 have more narrow line widths typical of free radicals. The deflection at g = 2.005 (peak-to-peak EPR spectral line width = 10 G) is characteristic of a semiquinone free radical (33). The intensity of these signals increased in PVB as T, rose, and new features became apparent at g = 2.012 and 2.0017 (Fig.…”

Physiological and pathophysiological reactive oxygen species as probed by EPR spectroscopy: the underutilized research window on muscle ageing

“…The resulting EPR spectrum reflects iron‐sulfur centres of the partially reduced mitochondrial respiratory chain complexes I–III (upper panel) and are assigned as described in van der Kraaij et al . (). EPR parameters: sweep rate, 1.55 mT s −1 ; modulation amplitude, 0.7 mT; MW attenuation, 10 dB; gain, 200.…”

“…The EPR tube was then slowly fitted in a glass Dewar flask filled with liquid nitrogen at equilibrium inside a Magnettech Benchtop EPR spectrometer before starting spectral acquisition. The resulting EPR spectrum reflects iron-sulfur centres of the partially reduced mitochondrial respiratory chain complexes I-III (upper panel) and are assigned as described invan der Kraaij et al (1989). EPR parameters: sweep rate, 1.55 mT s −1 ; modulation amplitude, 0.7 mT; MW attenuation, 10 dB; gain, 200.J Physiol 594.16…”