Development of a fast liquid chromatography-tandem mass spectrometry method for simultaneous quantification of neurotran
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RESEARCH PAPER
Development of a fast liquid chromatography-tandem mass spectrometry method for simultaneous quantification of neurotransmitters in murine microdialysate Christin Helmschrodt 1 & Susen Becker 1 & Stefanie Perl 1 & Anja Schulz 1 & Angelika Richter 1 Received: 11 June 2020 / Revised: 9 August 2020 / Accepted: 20 August 2020 # The Author(s) 2020
Abstract The continuous measurement of multiple neurotransmitters in microdialysate of freely moving mice to study neurochemical changes in specific brain regions requires a rapid and very sensitive quantitative analytical method. The quantitative analysis of 11 neurotransmitters and metabolites, including serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), melatonin (ME), dopamine (DA), levodopa (L-DOPA), 3-methoxytyramine (3-MT), norepinephrine (NE), epinephrine (EP), acetylcholine (ACh), choline (Ch), and γ-aminobutyric acid (GABA), was performed using a biphenyl column coupled to an API-QTrap 3200 (AB SCIEX) mass spectrometer in positive electrospray ionization mode. To the microdialysate samples, 0.5 ng of isotopically labeled standard was added for analyte quantification. A rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous analysis of monoamines, their precursor, and metabolites, as well as ACh, Ch, and GABA in murine microdialysate within 7.0 min. The limit of detection in artificial CSF ranged from 0.005 ng/mL (ME) to 0.75 ng/mL (NE and GABA). A comprehensive pre-analytical protocol was validated. Recovery was between 87 and 117% for neurotransmitter concentrations from 0.6 to 45 ng/mL with an inter-day accuracy of below 20%. Basal neurotransmitter values were determined in the striatum of mice over a time period of 3 h. This LC-MS/MS method, including a short and gentle sample preparation, is suitable for simultaneous measurements of neurotransmitters in murine cerebral microdialysate and enables the determination of basal neurotransmitter levels in specific brain regions to detect disease-related and drug-induced neurochemical changes.
Keywords Microdialysis . Neurotransmitter . Neurochemistry . LC-MS/MS . Basal values . Murine microdialysate Abbreviations ACh aCSF Ch DA EC ECD ESI EP GABA
Acetylcholine Artificial cerebrospinal fluid Choline Dopamine Electrochemical Electrochemical detection Electrospray ionization Epinephrine γ-Aminobutyric acid
* Christin Helmschrodt [email protected] 1
Institute of Pharmacology, Pharmacy and Toxicology, Faculty of Veterinary Medicine, University of Leipzig, An den Tierkliniken 15, 04103 Leipzig, Germany
5-HIAA 5-HT LC-MS/MS L-DOPA MD ME 3-MT NE NT
5-Hydroxyindoleacetic acid Serotonin Liquid chromatography tandem mass spectrometry Levodopa Microdialysis Melatonin 3-Methoxytyramine Norepinephrine Neurotransmitter
Introduction Neurochemistry provides insights into the pathophysiology of neurological diseases and mechanisms of drugs. In addition to anatomical and behavioral methods, this field complements the study of the
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