S-NITROSOGLUTATHIONE IN RAT CEREBELLUM - IDENTIFICATION AND QUANTIFICATION BY LIQUID-CHROMATOGRAPHY MASS-SPECTROMETRY

Given the extreme lability and the facile inactivation of the messenge r nitric oxide (NO) by many reactive biochemical species, it has been suggested that some intermediate compounds, for example, S-nitrosothio ls, may act to stabilize NO and at the same time to preserve its biolo gical activity. To test this hypothesis, we investigated if the S-nitr osothiol of glutathione, which is the predominant low molecular weight thiol in CNS, is present in the rat brain. The HPLC analysis of cereb ellar extract from [S-35] cysteine-prelabeled slices suggested that S- nitrosoglutathione (GSNO) was indeed present in rat brain, To detect e ndogenous GSNO, a methodology based on liquid chromatography-mass spec trometry was developed, Besides an unequivocal identification of the e ndogenous GSNO, this method also permitted its precise quantification using N-15-labeled GSNO ([N-15]GSNO) as internal standard. GSNO level in adult cerebellum amounts to 15.4 +/- 1.4 pmol/mg of protein. This i s the first direct demonstration of the presence of endogenous GSNO in CNS. The packaging of NO in the form of GSNO might serve to facilitat e its transport, prolong its life, and target its delivery to specific effecters.