Abstract
Acetylcholinesterase (AChE) hydrolyzes acetylcholine (ACh), a vital neurotransmitter that regulates muscle movement and brain function, including memory, attention, and learning. Inhibition of AChE activity can cause a variety of adverse health effects and toxicity. Identifying AChE inhibitors quickly and efficiently warrants developing AChE inhibition assays in a quantitative, high-throughput screening (qHTS) platform. In this chapter, protocols for multiple homogenous AChE inhibition assays used in a qHTS system are provided. These AChE inhibition assays include a (1) human neuroblastoma (SH-SY5Y) cell-based assay with fluorescence or colorimetric detection; (2) human recombinant AChE with fluorescence or colorimetric detection; and (3) combination of human recombinant AChE and liver microsomes with colorimetric detection, which enables detection of test compounds requiring metabolic activation to become AChE inhibitors. Together, these AChE assays can help identify, prioritize, and predict chemical hazards in large compound libraries using qHTS systems.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Soreq H, Seidman S (2001) Acetylcholinesterase - new roles for an old actor. Nat Rev Neurosci 2(4):294–302. https://doi.org/10.1038/35067589
Imbimbo BP (2001) Pharmacodynamic-tolerability relationships of cholinesterase inhibitors for Alzheimer’s disease. CNS Drugs 15(5):375–390. https://doi.org/10.2165/00023210-200115050-00004
Mirjana BC, Danijela ZK, Tamara DL-P, Aleksandra MB, Vesna MV (2013) Acetylcholinesterase inhibitors: pharmacology and toxicology. Curr Neuropharmacol 11(3):315–335. https://doi.org/10.2174/1570159X11311030006
Ali TB, Schleret TR, Reilly BM, Chen WY, Abagyan R (2015) Adverse effects of cholinesterase inhibitors in dementia, according to the pharmacovigilance databases of the United-States and Canada. PLoS One 10(12):e0144337. https://doi.org/10.1371/journal.pone.0144337
Inglese J, Auld DS, Jadhav A, Johnson RL, Simeonov A, Yasgar A, Zheng W, Austin CP (2006) Quantitative high-throughput screening: a titration-based approach that efficiently identifies biological activities in large chemical libraries. Proc Natl Acad Sci U S A 103(31):11473–11478. https://doi.org/10.1073/pnas.0604348103
Attene-Ramos MS, Miller N, Huang R, Michael S, Itkin M, Kavlock RJ, Austin CP, Shinn P, Simeonov A, Tice RR, Xia M (2013) The Tox21 robotic platform for the assessment of environmental chemicals--from vision to reality. Drug Discov Today 18(15–16):716–723. https://doi.org/10.1016/j.drudis.2013.05.015
Thomas RS, Paules RS, Simeonov A, Fitzpatrick SC, Crofton KM, Casey WM, Mendrick DL (2018) The US federal Tox21 program: a strategic and operational plan for continued leadership. ALTEX 35(2):163–168. https://doi.org/10.14573/altex.1803011
Li S, Huang R, Solomon S, Liu Y, Zhao B, Santillo MF, Xia M (2017) Identification of acetylcholinesterase inhibitors using homogenous cell-based assays in quantitative high-throughput screening platforms. Biotechnol J 12(5). https://doi.org/10.1002/biot.201600715
Li S, Zhao J, Huang R, Santillo MF, Houck KA, Xia M (2019) Use of high-throughput enzyme-based assay with xenobiotic metabolic capability to evaluate the inhibition of acetylcholinesterase activity by organophosphorous pesticides. Toxicol In Vitro 56:93–100. https://doi.org/10.1016/j.tiv.2019.01.002
Li S, Zhao J, Huang R, Travers J, Klumpp-Thomas C, Yu W, MacKerell AD, Sakamuru S, Ooka M, Xue F, Sipes NS, Hsieh J-H, Ryan K, Simeonov A, Santillo MF, Xia M (2021) Profiling the Tox21 chemical collection for acetylcholinesterase inhibition. Environ Health Perspect 129(4):047008. https://doi.org/10.1289/EHP6993
Inglese J, Johnson RL, Simeonov A, Xia M, Zheng W, Austin CP, Auld DS (2007) High-throughput screening assays for the identification of chemical probes. Nat Chem Biol 3(8):466–479. https://doi.org/10.1038/nchembio.2007.17
Pereira DA, Williams JA (2007) Origin and evolution of high throughput screening. Brit J Pharmacol 152(1):53–61. https://doi.org/10.1038/sj.bjp.0707373
Sipes NS, Martin MT, Kothiya P, Reif DM, Judson RS, Richard AM, Houck KA, Dix DJ, Kavlock RJ, Knudsen TB (2013) Profiling 976 ToxCast chemicals across 331 enzymatic and receptor signaling assays. Chem Res Toxicol 26(6):878–895. https://doi.org/10.1021/tx400021
Santillo MF, Liu Y (2015) A fluorescence assay for measuring acetylcholinesterase activity in rat blood and a human neuroblastoma cell line (SH-SY5Y). J Pharmacol Toxicol Methods 76:15–22. https://doi.org/10.1016/j.vascn.2015.07.002
Acknowledgments
This work was supported in part by the Intramural research program of the NCATS, NIH.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2022 This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply
About this protocol
Cite this protocol
Li, S., Li, A.J., Zhao, J., Santillo, M.F., Xia, M. (2022). Acetylcholinesterase Inhibition Assays for High-Throughput Screening. In: Zhu, H., Xia, M. (eds) High-Throughput Screening Assays in Toxicology. Methods in Molecular Biology, vol 2474. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2213-1_6
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2213-1_6
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2212-4
Online ISBN: 978-1-0716-2213-1
eBook Packages: Springer Protocols