SPIN-TRAPPING OF NITRIC-OXIDE PRODUCED IN-VIVO IN SEPTIC-SHOCK MICE

A nitric oxide (.NO) spin-trapping technique combined with electron pa ramagnetic resonance (EPR) spectroscopy has been employed to measure t he in vivo production of .NO in lipopolysaccharide (LPS)-treated mice. The in vivo spin-trapping of .NO was performed by injecting into mice a metal-chelator complex, consisting of N-methyl-D-glucamine dithioca rbamate (MGD) and reduced iron (Fe2+), that binds to .NO and forms a s table, water-soluble [(MGD)(2)-Fe2+-NO] complex, and by monitoring con tinuously the in vivo formation of the latter complex using an S-band EPR spectrometer. At 6 h after intravenous injection of LPS, a three-l ine EPR spectrum of the [(MGD)(2)-Fe2+-NO] complex, was observed in th e blood circulation of the mouse tail; the [(MGD)(2)-Fe2+] complex was injected subcutaneously 2 h before EPR measurement. No signal was det ected in control groups. Administration of N-G-monomethyl-L-arginine, an .NO synthase inhibitor, caused a marked reduction in the in vivo EP R signal of the [(MGD)(2)-Fe2+-NO] complex, suggesting that the .NO de tected is synthesized via the arginine-nitric oxide synthase pathway. The results presented here demonstrated, for the first time, the in vi vo real time measurement of .NO in the blood circulation of conscious, LPS-treated animals.