title: Highly sensitive isotope-dilution liquid-chromatography-electrospray ionization-tandem-mass spectrometry approach to study the drug-mediated modulation of dopamine and serotonin levels in Caenorhabditis elegans.
abstract: Dopamine (DA) and serotonin (SRT) are monoamine neurotransmitters that play a key role in regulating the central and peripheral nervous system. Their impaired metabolism has been implicated in several neurological disorders, such as Parkinson's disease and depression. Consequently, it is imperative to monitor changes in levels of these low-abundant neurotransmitters and their role in mediating disease. For the first time, a rapid, specific and sensitive isotope-dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantification of DA and SRT in the nematode Caenorhabditis elegans (C. elegans). This model organism offers a unique approach for studying the effect of various drugs and environmental conditions on neurotransmitter levels, given by the conserved DA and SRT biology, including synaptic release, trafficking and formation. We introduce a novel sample preparation protocol incorporating the usage of sodium thiosulfate in perchloric acid as extraction medium that assures high recovery of the relatively unstable neurotransmitters monitored. Moreover, the use of both deuterated internal standards and the multiple reaction monitoring (MRM) technique allows for unequivocal quantification. Thereby, to the best of our knowledge, we achieve a detection sensitivity that clearly exceeds those of published DA and SRT quantification methods in various matrices. We are the first to show that exposure of C. elegans to the monoamine oxidase B (MAO-B) inhibitor selegiline or the catechol-O-methyltransferase (COMT) inhibitor tolcapone, in order to block DA and SRT degradation, resulted in accumulation of the respective neurotransmitter. Assessment of a behavioral output of the dopaminergic system (basal slowing response) corroborated the analytical LC-MS/MS data. Thus, utilization of the C. elegans model system in conjunction with our analytical method is well-suited to investigate drug-mediated modulation of the DA and SRT system in order to identify compounds with neuroprotective or regenerative properties.
authors.affiliation: Department of Toxicology, Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany; Department of Molecular Biology, University of Duisburg-Essen, Hufelandstraße 55, 45147 Essen, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Schumacher
authors.affiliation: Neuroscience Graduate Program, Vanderbilt University Medical Center, Nashville, TN, USA; Department of Molecular Pharmacology, Neuroscience, and Pediatrics, Albert Einstein College of Medicine, Bronx, NY, USA.
authors.forename: Sudipta
authors.initials: S
authors.lastname: Chakraborty
authors.affiliation: Department of Toxicology, Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Kleuser
authors.affiliation: Department of Molecular Biology, University of Duisburg-Essen, Hufelandstraße 55, 45147 Essen, Germany.
authors.forename: Erich
authors.initials: E
authors.lastname: Gulbins
authors.affiliation: Department of Food Chemistry, Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany.
authors.forename: Tanja
authors.initials: T
authors.lastname: Schwerdtle
authors.affiliation: Department of Molecular Pharmacology, Neuroscience, and Pediatrics, Albert Einstein College of Medicine, Bronx, NY, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Aschner
authors.affiliation: Department of Food Chemistry, Institute of Nutritional Science, University of Potsdam, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany. Electronic address: julia.bornhorst@uni-potsdam.de.
authors.forename: Julia
authors.initials: J
authors.lastname: Bornhorst
mesh_terms: D000818:Animals; D001577:Benzophenones / Q000494:pharmacology; D017173:Caenorhabditis elegans / Q000187:drug effects* / Q000378:metabolism*; D065098:Catechol O-Methyltransferase Inhibitors / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003903:Deuterium / Q000737:chemistry; D004298:Dopamine / Q000378:metabolism*; D057230:Limit of Detection; D008996:Monoamine Oxidase Inhibitors / Q000494:pharmacology; D009596:Nitrophenols / Q000494:pharmacology; D012642:Selegiline / Q000494:pharmacology; D012701:Serotonin / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013997:Time Factors; D000077867:Tolcapone
chemical_list: D001577:Benzophenones; D065098:Catechol O-Methyltransferase Inhibitors; D008996:Monoamine Oxidase Inhibitors; D009596:Nitrophenols; D012642:Selegiline; D012701:Serotonin; D003903:Deuterium; D000077867:Tolcapone; D004298:Dopamine
keywords: Caenorhabditis elegans; Dopamine; Isotope-dilution analysis; Liquid chromatography-tandem mass spectrometry; Serotonin