To detect nitric oxide (NO) in the rat brain during lipopolysaccharide (LPS)-induced sepsis, electron paramagnetic resonance (EPR) was employed with the NO trapping technique, using an iron and N,N-diethyldithiocarbamate (DETC) complex. An X-band (about 9.5 GHz) EPR system detected a triplet signal (g = 2.038) derived from an NO-Fe-DETC complex being superimposed on the g(perpendicular) signal of Cu-DETC complex at liquid nitrogen temperature. The height of the triplet signal peaked seven hours after injection of 40 mg/kg of LPS, and over 25 x 10(4) U/kg of IFN-gamma enhanced the LPS-induced NO formation. Pretreatment with N(G)-monomethyl-L-arginine (NMMA), an NO synthase inhibitor, deleted only the triplet signal. A triplet signal (g(iso) = 2.040, aN = 1.28 mT) derived from the NO-Fe-DETC complex was also observed at ambient temperature. Then, a home-built 700 MHz EPR system was used to detect an NO signal in the septic rat brain in vivo. We successfully monitored the NO-Fe-DETC signal in the head region of a living rat under the condition that provided maximum height of the NO-Fe-DETC signal in the X-band EPR study. Pretreatment with NMMA again deleted the NO-Fe-DETC signal. This is the first EPR observation of endogenous NO in the brain of living rats.