title: Simultaneous quantification of monoamine neurotransmitters and their biogenic metabolites intracellularly and extracellularly in primary neuronal cell cultures and in sub-regions of guinea pig brain.
abstract: In the present paper, we describe a validated chromatographic method for the simultaneous quantification of monoamine neurotransmitters and their biogenic metabolites intracellularly and extracellularly in primary neuronal cell culture and in sub-regions of the guinea pig brain. Electrochemical detection provided limits of quantifications (LOQs) between 3.6 and 12nM. Within the linear range, obtained recoveries were from 90.9±9.9 to 120±14% and intra-day and inter-day precisions found to be less than 5.5% and 12%, respectively. The analytical method was applicable for quantification of intracellular and extracellular amounts of monoamine neurotransmitters and their metabolites in guinea pig frontal cortex and hippocampal primary neuronal cell cultures. Noradrenaline, dopamine and serotonin were found to be in a range from 0.31 to 1.7pmol per 2 million cells intracellularly, but only the biogenic metabolites could be detected extracellularly. Distinct differences in monoamine concentrations were observed when comparing concentrations in guinea pig frontal cortex and cerebellum tissue with higher amounts of dopamine and its metabolites, 3,4-dihydroxyphenylacetic acid and homovanillic acid in frontal cortex, as compared to cerebellum. The chemical turnover in frontal cortex tissue of guinea pig was for serotonin successfully predicted from the turnover observed in the frontal cortex cell culture. In conclusion, the present analytical method shows high precision, accuracy and sensitivity and is broadly applicable to monoamine measurements in cell cultures as well as brain biopsies from animal models used in preclinical neurochemistry.
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Ridebanevej 9, 1870 Frederiksberg C, Denmark. Electronic address: am.schoupedersen@sund.ku.dk.
authors.forename: Anne Marie V
authors.initials: AMV
authors.lastname: Schou-Pedersen
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Ridebanevej 9, 1870 Frederiksberg C, Denmark.
authors.forename: Stine N
authors.initials: SN
authors.lastname: Hansen
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Ridebanevej 9, 1870 Frederiksberg C, Denmark.
authors.forename: Pernille
authors.initials: P
authors.lastname: Tveden-Nyborg
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Ridebanevej 9, 1870 Frederiksberg C, Denmark.
authors.forename: Jens
authors.initials: J
authors.lastname: Lykkesfeldt
mesh_terms: D000818:Animals; D015306:Biogenic Monoamines / Q000032:analysis* / Q000378:metabolism; D001921:Brain / Q000378:metabolism; D001923:Brain Chemistry; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D055664:Electrochemical Techniques / Q000379:methods; D006168:Guinea Pigs; D009474:Neurons / Q000737:chemistry* / Q000378:metabolism; D018377:Neurotransmitter Agents / Q000032:analysis* / Q000378:metabolism; D061251:Primary Cell Culture
chemical_list: D015306:Biogenic Monoamines; D018377:Neurotransmitter Agents
keywords: Cell culture; Electrochemical detection; Guinea pig; HPLC; Monoamines