In vivo monitoring of glutathione and cysteine in rat caudate nucleus using microdialysis on-line with capillary zone electrophoresis-laser induced fluorescence detection

Abstract

A fully-automated method for monitoring thiols in vivo using microdialysis coupled on-line with capillary zone electrophoresis with laser-induced fluorescence detection was developed. Dialysates were derivatized on-line with monobromobimane and automatically transferred to the separation capillary by a flow-gated interface. Analytes were detected on-column using the 2 mW, 354 nm line of a He-Cd laser for excitation. Dialysis probes were perfused at 79 nl/min resulting in relative recoveries of nearly 100%, which allowed quantitative monitoring. On-line detection limits for these analytes were in the 20–40 nM range and the response was linear up to 20 μM. The system was applied to the measurement of glutathione and cysteine in the extracellular space of the caudate nucleus of anesthetized rats. The measured basal concentrations of glutathione and cysteine were 2.0±0.1 μM and 2.3±0.3 μM, respectively which agree well with literature values. Increases in glutathione and cysteine were monitored with 180 s temporal resolution during stimulation by infusion of potassium. The average concentration of glutathione and cysteine during stimulation was 3.0±0.9 and 3.3±0.5 μM ( n=3), respectively. This system is the first to obtain high relative recoveries and high temporal resolution simultaneously for multiple thiols with microdialysis sampling in the brain.