In vivo monitoring of neurotransmitter release by microdialysis and on-line CE-LIF.

Abstract

An analytical method based on microdialysis sampling and micellar electrokinetic chromatography (MEKC) was developed to monitor neurotransmitter release in live animal brains. In vivo amino acids were sampled by microdialysis probe and derivatized on-line by the addition of naphthalene-2,3-dicarboxaldehyde (NDA) and cyanide to form fluorescent derivatives. Analytes were continuously delivered into a flow-gated interface and monitored by a home-built capillary electrophoresis laser-induced fluorescence (CE-LIF) system. The optimized condition allows 18 amino acid derivatives to be resolved within 30 s, which can be further reduced to 20 s by overlapped injections. Limits of detection for amino acids range from 30--50 nM. The integrity of this method was demonstrated by using freely-moving rats as a model to study the effect of acute alcohol injections and pharmacological administrations. The on-line sampling and separation scheme was further improved to detect low abundance endogenous compounds, such as dopamine and norepinephrine. The optimized in vitro detection limit for dopamine is &sim;2 nM. An MEKC condition was developed to allow dopamine to be resolved from compounds in dialysate. Local administrations of nomifensine and alpha-methyl-<italic>p</italic>-tyrosine (AMPT) were applied to characterize in vivo dopamine peak identity. The total in vivo electropherogram exhibits over 60 peaks and the temporal resolution can be as low as 1 min. Besides monitoring dopamine in vivo, the separation scheme also allows amphetamine to be derivatized and detected simultaneously. The dopamine uptake inhibition induced by intravenous cocaine infusions in freely-moving rat striatum was monitored every 90 s. The same cocaine dosage was administered at different infusion rates (within 5 or 100 s). Both 5 and 100 s infusions caused a similar magnitude of increase in basal dopamine level. However, the 5 s infusion caused a more rapid (&sim;3 min faster) dopamine concentration change than 100 s infusion did. Overlaid curves of simulated cocaine concentration, dopamine level and psychomotor changes exhibited a good correspondence. The experimental data suggest the neuronal chemical information can be correlated to behavior changes.