Determination of extracellular glutathione in rat brain by microdialysis and high-performance liquid chromatography with fluorescence detection.

A method for the continuous monitoring of extracellular glutathione (GSH) concentrations in rat brain has been developed. This method involved the in vivo sampling of brain extracellular fluid by microdialysis perfusion and the subsequent analysis by high-performance liquid chromatography (HPLC) with fluorescence detection. Perfusates from the microdialysis probes were directly derivatized with methanolic monobromobimane which acted as the fluorescence tag. Separation of the derivatized perfusate was achieved on narrow-bore reversed-phase C18 columns. Recoveries of GSH from the microdialysis probes ranged from 1.5% to 4%. The basal extracellular GSH concentration in rat (Sprague-Dawley) brain cortex was found to be 2.10 +/- 1.78 microM (mean +/- S.D.) (results of 18 rats). Fluorescence detection and separation on narrow-bore columns provided adequate sensitivity for accurate determination of brain extracellular GSH concentrations in rats. With this method, the extracellular GSH concentrations in the cerebral cortex were found to be significantly elevated upon the onset of cerebral ischemia induced by the ligation of bilateral common carotid arteries.