Ex vivo EPR detection of nitric oxide in brain tissue.

Abstract

The concentration of nitric oxide (NO) was measured in the brain of septic-shock animals by electron paramagnetic resonance spectrometry (EPR). NO was spin trapped and quantitated in several regions of the brain (cortex, hippocampus, hypothalamus, cerebellum, and olfactory bulb) as well as other organs (liver, kidney, and heart) of rats induced with lipopolysaccharide (LPS) using Fe(II)/dithiocarbamate complexes containing diethyldithiocarbamate (DETC) or N-methyl-D-glucamine (MGD). The spin trap, (DETC)(2)-Fe(II), complexed NO generated in all tissues examined, but (MGD)(2)-Fe(II) complex was ineffective in detecting NO in the brain of septic-shock rats, although identical amounts of NO were detected in the liver with either spin trap. A triplet EPR spectrum of (DETC)(2)-Fe(II)-NO with a(N) = 12.8 gauss and g = 2.04 was observed in the cortex, hippocampus, hypothalamus, cerebellum, but not the olfactory bulb. The amount of NO in the brain was about 20% of that found in the liver. The (DETC)(2)-Fe(II)-NO signal in all the tissues of septic-shock rats was markedly suppressed by preadministration of the nitric oxide synthase (NOS) inhibitors, N(G)-monomethyl-L-arginine (L-NMMA) or 3-bromo-7-nitroindazole, suggesting that the NO detected from brain tissue was produced enzymatically by NOS. In contrast to previous studies on the liver and other organs, phenyl-N-tert-butyl nitrone (PBN), did not suppress iNOS expression in brain tissue of LPS-treated rats. This could be due to a totally different regulation system for iNOS in liver versus brain tissue. Magn Reson Med 42:599-602, 1999.