Determination of dopamine concentrations in brain extracellular fluid using microdialysis with short sampling intervals, analyzed by ultra high performance liquid chromatography tandem mass spectrometry (original) (raw)

2015, Journal of Pharmacological and Toxicological Methods

Dopamine Heroin Microdialysis Ultra high performance liquid chromatography tandem mass spectrometry UHPLC-MS/MS Introduction: An increase in striatal dopamine is considered essential for the rewarding and reinforcing effects of drugs of abuse. We have developed and validated an ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method for the analysis of dopamine in rat brain extracellular fluid (ECF) sampled with microdialysis. The method was applied to monitor changes in dopamine concentrations over time after an intravenous bolus injection of heroin. Methods: Dopamine and dopamine-d3 were analyzed using a 2.1 × 100 mm Aquity T3 column, 1.7 μm particle size, with a formic acid and methanol gradient. The run time of the method was 2.5 min including equilibration time. Results: The method had an LOQ of 0.15 ng/mL, which equals 0.55 pg on column. The calibration curves were linear in the tested area of 0.15 to 16 ng/mL. Inter-assay coefficients of variation varied between 5-17%, with an accuracy expressed as bias of − 10 to 5%. The intra-assay coefficients of variation varied between 9-15%, with an accuracy of −3-7%. Discussion: Heroin metabolism is very rapid. Sampling intervals of only 2 min were thus required to obtain an adequate number of samples of dopamine analysis accompanying the concentration-time profile of opioids in the brain. Applying a flow of 2 μL/min, 4 μL of dialysate were sampled at 2 min intervals, in 7 μL internal standard. The injection volume onto the UPLC column was 10 μL. Analyses of microdialysate samples from a rat given heroin i.v. showed that it was possible to measure baseline levels and rapid changes in dopamine concentrations with very short sampling periods.