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Measurement of TNF induced oxidative stress in the paraventricular nucleus using electron paramagnetic resonance spectroscopy
Author(s) -
Mariappan Nithya,
Francis Joseph
Publication year - 2009
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.23.1_supplement.700.2
Subject(s) - apocynin , electron paramagnetic resonance , oxidative stress , chemistry , reactive oxygen species , radical , superoxide , nadph oxidase , endocrinology , medicine , tumor necrosis factor alpha , nucleus , biophysics , nuclear magnetic resonance , biochemistry , neuroscience , biology , enzyme , physics
Recent findings from our lab suggest that TNF plays an important role in sympathoexcitation in heart failure. We have demonstrated that TNF treatment induces oxidative stress and contributes to end organ damage. In this study, we used a novel EPR method to determine whether chronic TNF treatment induces oxidative stress in the paraventricular nucleus of the hypothalamus. Adult male Sprague‐Dawley rats were divided into four groups: control; TNF treatment; TNF + apocynin (APO; 200 μmol/kg); and TNF + TEMPOL (TEMP; 300 μmol/kg). TNF (40 μg/kg) was injected daily for five days. Rats were sacrificed in a CO 2 chamber. We used EPR, the most sensitive and definitive method for quantification of free radicals, to determine reactive oxygen species levels in the PVN. EPR is superior to other free radical detection methods in that it allows for the direct measurement of specific free radicals using specific spin probes. The PVN was micro punched and treated with the specific spin probes 1‐hydroxy‐3‐methoxycarbonyl‐2, 2, 5, 5‐tetramethylpyrrolidine (CMH), for the measurement of total reactive oxygen species and superoxide by EPR. Treatment with TNF increased total ROS and superoxide levels in the PVN (p<0.05). In contrast, treatment with the NADPH oxidase inhibitor apocynin or with tempol attenuated the TNF induced oxidative stress. These findings suggest that EPR is a novel and sensitive method for the measurement of free radicals in brain nuclei.