Quantification of acetylcholine, an essential neurotransmitter, in brain microdialysis samples by liquid chromatography mass spectrometry

Abstract

Chemical neurotransmission has been the subject of intensive investigations in recent years. Acetylcholine is an essential neurotransmitter in the central nervous system as it has an effect on alertness, memory and learning. Enzymatic hydrolysis of acetylcholine in the synaptic cleft is fast and quickly metabolizes to choline and acetate by acetylcholinesterase. Hence the concentration in the extracellular fluid of the brain is low (0.1-6 nm). Techniques such as microdialysis are routinely employed to measure acetylcholine levels in living brain systems and the microdialysis sample volumes are usually less than 50 microL. In order to develop medicine for the diseases associated with cognitive dysfunction like mild cognitive impairment, Alzheimer's disease, schizophrenia and Parkinson's disease, or to study the mechanism of the illness, it is important to measure the concentration of acetylcholine in the extracellular fluid of the brain. Recently considerable attention has been focused on the development of chromatographic-mass spectrometric techniques to provide more sensitive and accurate quantification of acetylcholine collected from in-vivo brain microdialysis experiments. This review will provide a brief overview of acetylcholine biosynthesis, microdialysis technique and liquid chromatography mass spectrometry, which is being used to quantitate extracellular levels of acetylcholine.

Tonic and phasic release of glutamate and acetylcholine neurotransmission in sub-regions of the rat prefrontal cortex using enzyme-based microelectrode arrays

“…Microdialysis, with its collection times on the scale of several seconds to minutes, may allow for more ACh hydrolysis by acetylcholinesterase, resulting in lower reported concentrations. Thus, many scientists who use microdialysis for measuring ACh levels also perfuse an acetylcholinesterase inhibitor, such as neostigmine, to prevent ACh breakdown (Nirogi et al, 2010; Ragozzino et al, 1996). However, this technique may artificially affect the measured ACh levels in the extracellular space.…”

Analysis of acetylcholine from extracellular fluid in brain by in vivo microdialysis and LC–ESI-MS/MS with the stable isotope-labeled internal standard

“…The ACh peak had a bad shape and width in a simultaneous determination of multiple neurotransmitters [23]. Furthermore, the researchers analyzing ACh with MS detector have been listed in a review [24]. There were various columns and mobile phases for detection of ACh and the best LLOQ was 1 fmol on column.…”

Development and validation of a sample stabilization strategy and a UPLC–MS/MS method for the simultaneous quantitation of acetylcholine (ACh), histamine (HA), and its metabolites in rat cerebrospinal fluid (CSF)

“…ESI-MS/MS has been extensively used for the determination of endogenous biomarkers due to its high sensitivity and selectivity [16,27,28]. The ACh, HA, t-mHA, and t-MIAA molecules studies here all contain basic amine functionalities that allows for positive charging under acidic conditions and results in sufficient sensitivity using the ESI positive ionization mode without derivatization.…”

“…While analytical methodologies on the quantitative analysis of neurotransmitters in rat brain microdialysates have been extensively reported [16,18,27,37], only a few studies have analyzed their concentrations in the CSF [8]. Although in vivo microdialysis is used in preclinical studies to assess the release of neurotransmitters, the translation of these findings from preclinical to clinical studies has yet to be demonstrated.…”