A new variant of large-volume sample stacking injection (LVSS) was used in the capillary electrophoresis with capacitively coupled contactless conductivity detection (CE/C(4)D) determination of the neurotransmitters γ-aminobutyric acid (GABA), glycine (Gly) and glutamate (Glu) in microdialysates of periaqueductal gray matter (PAG). The separation capillary was filled to 98% from the injection side with a sample of microdialysate in acetonitrile. Simultaneously with turning on the separation voltage, the sample zone was forced out by the background electrolyte by increasing the pressure in the terminal capillary outlet vessel. As a consequence of the stacking effect, the analyte was concentrated from the large sample volume into a narrow zone at the sample/background electrolyte boundary close to the injection end of the capillary. Under these conditions, LOD values of 9, 10 and 15nM were determined in the model samples for GABA, Gly and Glu, respectively; RSD equalled 0.5% for the migration times and 1.0-1.9% for the peak areas, respectively. In analysis of microdialysates of PAG, LOD values of 29, 29 and 37nM were determined for GABA, Gly and Glu, respectively; RSD equalled 0.5-0.7% for the migration times and 2.6-8.2% for the peak areas, respectively. The determined basal levels of the neurotransmitters in PAG microdialysates are 0.08, 4.7 and 0.8μM for GABA, Gly and Glu, respectively. Carrageenan-induced hyperalgesia increases the Gly and Glu levels and reduces GABA in PAG microdialysate. Peroral administration of paracetamol in hyperalgesia effectively reduces the Gly value and has no effect on Glu and GABA.

Institute of Biochemistry Cell and Molecular Biology 3rd Faculty of Medicine Charles University Ruská 87 100 00 Prague 10 Czech Republic

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