Kinetics of increased generation of (.)NO in endotoxaemic rats as measured by EPR.

Abstract

Ferrous-diethyldithiocarbamate (Fe(DETC)(2)) chelate is a lipophilic spin trap developed for (.)NO detection by electron paramagnetic resonance (EPR) spectroscopy. Using this spin trap we investigated the kinetics of (.)NO production in endotoxaemia in rats induced by lipopolysaccharide (Escherichia coli, 10 mg/kg). The NO-Fe(DETC)(2) complex was found to give a characteristic EPR signal, and the amplitude of the 3rd (high-field) component of its hyperfine splitting was used to monitor the level of (.)NO. We found that in blood, kidney, liver, heart and lung (.)NO production starts to increase as early as 2 h after LPS injection, reaches the maximum 6 h after LPS injection and then returns to basal level within further 12-18 h. Interestingly, in the eye bulb the maximum of (.)NO production was detected 12 h after LPS, and the signal was still pronounced 24 h after LPS. In brief, the highly lipophilic exogenous spin trap, Fe(DETC)(2) is well suited for assessment of (.)NO production in endotoxaemia. We demonstrated that the kinetics of increased production of (.)NO in endotoxaemic organs, with the notable exception of the eye, do not follow the known pattern of NOS-2 induction under those conditions. Accordingly, only in early endotoxaemia a high level of (.)NO is detected, while in late endotoxaemia (.)NO detectability is diminished most probably due to concomitant oxidant stress.