Abstract
Excessive levels of dopamine in the synaptic cleft, induced by cocaine for example, activates dopaminergic receptors, mainly D1R, D2R, and D3R subtypes, contributing to neurotoxic effects. New synthetic 1-[(2,3-dihydro-1-benzofuran-2-yl)methyl]piperazine derivatives (the LINS01 compounds), designed as histaminergic receptor (H3R) ligands, are also dopaminergic receptor ligands, mainly D2R and D3R. This study aims to evaluate the neurotoxicity of these new synthetic LINS01 compounds (LINS01003, LINS01004, LINS01011, and LINS01018), as well as to investigate their protective potential on a cocaine model of dopamine-induced neurotoxicity using SH-SY5Y cell line culture. Neurotoxicity was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), lactate dehydrogenase (LDH), and automated cell counting with fluorescent dyes (acridyl orange and propidium iodide) assays. Concentration-response curves (CRCs) were performed for all LINS compounds and cocaine using MTT assay. The results show that LINS series did not decrease cell viability after 48h of exposure—except for 100 µM LINS01018, which was discontinued from the study. Likewise, MTT, LDH, and fluorescent dyes staining showed no difference is cell viability for LINS compounds at 10 µM. When incubated with 2.5 mM cocaine (lethal concentration 50) for 48h, 10 µM of each LINS compound, metoclopramide (D2R antagonist) and haloperidol (D2R/D3R antagonist), ameliorated cocaine-induced neurotoxicity. However, only metoclopramide, haloperidol, and LINS01011 compound significantly decreased LDH released in the culture medium, suggesting that this new synthetic compound presents a more robust effect. This preliminary in vitro neurotoxicity study suggests that LINS01 compounds are not neurotoxic, and that they play a promising role in preventing cocaine-induced neurotoxicity.
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Acknowledgements
The authors gratefully acknowledge the support of students Maria C. R. Azevedo, Rafaela Y. S. Almeida, and Emidio P. L. Junior from UNIFESP; Gabriela O. Pereira from the University of São Paulo; Dr. Rodrigo P. Ureshino from UNIFESP; and the English specialist Janaina P. de Almeida from the University of São Paulo.
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This work was supported by the São Paulo Research Foundation - FAPESP (grants 2016/23139-2, 2017/21834-8, and 2019/24028-8) and the National Council for Scientific and Technological Development - CNPq (grants 307829/2021-9 and 156320/2019-3). TM and JPSF are research fellows of CNPq.
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All authors contributed to the study conception and design. Material preparation, data collection, and analysis were performed by Laísa Aliandro dos Santos, Gabriela Salles dos Santos, Gustavo Ariel Borges Fernandes, Michelle Fidelis Corrêa, Carolina Aparecida de Faria Almeida, and Raphael Caio Tamborelli Garcia. The first draft of the manuscript was written by Raphael Caio Tamborelli Garcia and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.
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dos Santos, L.A., dos Santos, G.S., Fernandes, G.A.B. et al. Neurotoxicity Assessment of 1-[(2,3-Dihydro-1-Benzofuran-2-yl)Methyl]Piperazine (LINS01 Series) Derivatives and their Protective Effect on Cocaine-Induced Neurotoxicity Model in SH-SY5Y Cell Culture. Neurotox Res 40, 1653–1663 (2022). https://doi.org/10.1007/s12640-022-00601-8
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DOI: https://doi.org/10.1007/s12640-022-00601-8