Abstract
Cathinones (β-keto amphetamines), widely abused in recreational settings, have been shown similar or even worse toxicological profile than classical amphetamines. In the present study, the cytotoxicity of two β-keto amphetamines [3,4-dimethylmethcathinone (3,4-DMMC) and 4-methylmethcathinone (4-MMC)], was evaluated in differentiated dopaminergic SH-SY5Y cells in comparison to methamphetamine (METH). MTT reduction and NR uptake assays revealed that both cathinones and METH induced cytotoxicity in a concentration- and time-dependent manner. Pre-treatment with trolox (antioxidant) partially prevented the cytotoxicity induced by all tested drugs, while N-acetyl-l-cysteine (NAC; antioxidant and glutathione precursor) and GBR 12909 (dopamine transporter inhibitor) partially prevented the cytotoxicity induced by cathinones, as evaluated by the MTT reduction assay. Unlike METH, cathinones induced oxidative stress evidenced by the increase on intracellular levels of reactive oxygen species (ROS), and also by the decrease of intracellular glutathione levels. Trolox prevented, partially but significantly, the ROS generation elicited by cathinones, while NAC inhibited it completely. All tested drugs induced mitochondrial dysfunction, since they led to mitochondrial membrane depolarization and to intracellular ATP depletion. Activation of caspase-3, indicative of apoptosis, was seen both for cathinones and METH, and confirmed by annexin V and propidium iodide positive staining. Autophagy was also activated by all drugs tested. Pre-incubation with bafilomycin A1, an inhibitor of the vacuolar H+-ATPase, only protected against the cytotoxicity induced by METH, which indicates dissimilar toxicological pathways for the tested drugs. In conclusion, the mitochondrial impairment and oxidative stress observed for the tested cathinones may be key factors for their neurotoxicity, but different outcome pathways seem to be involved in the adverse effects, when compared to METH.
Graphic abstract
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Ajjimaporn A, Swinscoe J, Shavali S, Govitrapong P, Ebadi M (2005) Metallothionein provides zinc-mediated protective effects against methamphetamine toxicity in SK-N-SH cells. Brain Res Bull 67(6):466–475. https://doi.org/10.1016/j.brainresbull.2005.07.012
Andersen PH (1989) The dopamine inhibitor GBR 12909: selectivity and molecular mechanism of action. Eur J Pharmacol 166(3):493–504. https://doi.org/10.1016/0014-2999(89)90363-4
Anneken JH, Angoa-Perez M, Sati GC, Crich D, Kuhn DM (2018) Assessing the role of dopamine in the differential neurotoxicity patterns of methamphetamine, mephedrone, methcathinone and 4-methylmethamphetamine. Neuropharmacology 134(Pt A):46–56. https://doi.org/10.1016/j.neuropharm.2017.08.033
Arbo MD, Silva R, Barbosa DJ et al (2016) In vitro neurotoxicity evaluation of piperazine designer drugs in differentiated human neuroblastoma SH-SY5Y cells. J Appl Toxicol 36(1):121–130. https://doi.org/10.1002/jat.3153
Archer T, Kostrzewa RM (2018) Synthetic cathinones: neurotoxic health hazards and potential for abuse. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 1–10
Barbosa DJ, Capela JP, Silva R et al (2014) The mixture of “ecstasy” and its metabolites is toxic to human SH-SY5Y differentiated cells at in vivo relevant concentrations. Arch Toxicol 88(2):455–473. https://doi.org/10.1007/s00204-013-1120-7
Barbosa DJ, Capela JP, Feio-Azevedo R, Teixeira-Gomes A, Bastos Mde L, Carvalho F (2015) Mitochondria: key players in the neurotoxic effects of amphetamines. Arch Toxicol 89(10):1695–1725. https://doi.org/10.1007/s00204-015-1478-9
Cantarella G, Uberti D, Carsana T, Lombardo G, Bernardini R, Memo M (2003) Neutralization of TRAIL death pathway protects human neuronal cell line from beta-amyloid toxicity. Cell Death Differ 10(1):134–141. https://doi.org/10.1038/sj.cdd.4401143
Carvalho M, Carmo H, Costa VM et al (2012) Toxicity of amphetamines: an update. Arch Toxicol 86(8):1167–1231. https://doi.org/10.1007/s00204-012-0815-5
Chandramani Shivalingappa P, ** H, Anantharam V, Kanthasamy A, Kanthasamy A (2012) N-acetyl cysteine protects against methamphetamine-induced dopaminergic neurodegeneration via modulation of redox status and autophagy in dopaminergic cells. Parkinson’s Dis 2012:11. https://doi.org/10.1155/2012/424285
Chen L, Huang E, Wang H, Qiu P, Liu C (2013) RNA interference targeting alpha-synuclein attenuates methamphetamine-induced neurotoxicity in SH-SY5Y cells. Brain Res 1521:59–67. https://doi.org/10.1016/j.brainres.2013.05.016
Chen Q, Kang J, Fu C (2018) The independence of and associations among apoptosis, autophagy, and necrosis. Signal Transduct Target Ther 3:18. https://doi.org/10.1038/s41392-018-0018-5
Coccini T, Vecchio S, Crevani M, De Simone U (2019) Cytotoxic effects of 3,4-catechol-PV (one major MDPV metabolite) on human dopaminergic SH-SY5Y cells. Neurotox Res 35(1):49–62. https://doi.org/10.1007/s12640-018-9924-0
Coppola M, Mondola R (2012) Synthetic cathinones: chemistry, pharmacology and toxicology of a new class of designer drugs of abuse marketed as “bath salts” or “plant food”. Toxicol Lett 211(2):144–149. https://doi.org/10.1016/j.toxlet.2012.03.009
Corkery JM, Guirguis A, Papanti DG, Orsolini L, Schifano F (2018) Synthetic cathinones—prevalence and motivations for use. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 153–189
Cossarizza A, Baccarani-Contri M, Kalashnikova G, Franceschi C (1993) A new method for the cytofluorimetric analysis of mitochondrial membrane potential using the J-aggregate forming lipophilic cation 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1). Biochem Biophys Res Commun 197(1):40–45. https://doi.org/10.1006/bbrc.1993.2438
Costa VM, Carvalho F, Duarte JA, Bastos Mde L, Remiao F (2013) The heart as a target for xenobiotic toxicity: the cardiac susceptibility to oxidative stress. Chem Res Toxicol 26(9):1285–1311. https://doi.org/10.1021/tx400130v
den Hollander B, Sundstrom M, Pelander A et al (2014) Keto amphetamine toxicity-focus on the redox reactivity of the cathinone designer drug mephedrone. Toxicol Sci 141(1):120–131. https://doi.org/10.1093/toxsci/kfu108
den Hollander B, Sundstrom M, Pelander A et al (2015) Mitochondrial respiratory dysfunction due to the conversion of substituted cathinones to methylbenzamides in SH-SY5Y cells. Sci Rep 5:14924. https://doi.org/10.1038/srep14924
Escher BI, Hermens JL (2004) Internal exposure: linking bioavailability to effects. Environ Sci Technol 38(23):455A–462A. https://doi.org/10.1021/es0406740
Ferreira PS, Nogueira TB, Costa VM et al (2013) Neurotoxicity of “ecstasy” and its metabolites in human dopaminergic differentiated SH-SY5Y cells. Toxicol Lett 216(2–3):159–170. https://doi.org/10.1016/j.toxlet.2012.11.015
Ferreira C, Vaz AR, Florindo PR, Lopes A, Brites D, Quintas A (2019) Development of a high throughput methodology to screen cathinones’ toxicological impact. Forensic Sci Int 298:1–9. https://doi.org/10.1016/j.forsciint.2019.02.022
Fischer FC, Henneberger L, Konig M et al (2017) Modeling exposure in the Tox21 in vitro bioassays. Chem Res Toxicol 30(5):1197–1208. https://doi.org/10.1021/acs.chemrestox.7b00023
Fischer FC, Henneberger L, Schlichting R, Escher BI (2019) How to improve the dosing of chemicals in high-throughput in vitro mammalian cell assays. Chem Res Toxicol 32(8):1462–1468. https://doi.org/10.1021/acs.chemrestox.9b00167
Flippo KH, Strack S (2017) Mitochondrial dynamics in neuronal injury, development and plasticity. J Cell Sci 130(4):671–681. https://doi.org/10.1242/jcs.171017
Foroughi K, Khaksari M, Rahmati M, Bitaraf FS, Shayannia A (2019) Apelin-13 protects PC12 cells against methamphetamine-induced oxidative stress, autophagy and apoptosis. Neurochem Res 44(9):2103–2112. https://doi.org/10.1007/s11064-019-02847-9
Fricker M, Tolkovsky AM, Borutaite V, Coleman M, Brown GC (2018) Neuronal cell death. Physiol Rev 98(2):813–880. https://doi.org/10.1152/physrev.00011.2017
Gaspar H, Bronze S, Oliveira C et al (2018) Proactive response to tackle the threat of emerging drugs: Synthesis and toxicity evaluation of new cathinones. Forensic Sci Int 290:146–156. https://doi.org/10.1016/j.forsciint.2018.07.001
Gołembiowska K, Kamińska K (2018) Effects of synthetic cathinones on brain neurotransmitters. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 117–124
Graham DG (1978) Oxidative pathways for catecholamines in the genesis of neuromelanin and cytotoxic quinones. Mol Pharmacol 14(4):633–643
Heringa MB, Schreurs RH, Busser F, van der Saag PT, van der Burg B, Hermens JL (2004) Toward more useful in vitro toxicity data with measured free concentrations. Environ Sci Technol 38(23):6263–6270. https://doi.org/10.1021/es049285w
Huang YN, Wang JY, Lee CT, Lin CH, Lai CC, Wang JY (2012) l-ascorbate attenuates methamphetamine neurotoxicity through enhancing the induction of endogenous heme oxygenase-1. Toxicol Appl Pharmacol 265(2):241–252. https://doi.org/10.1016/j.taap.2012.08.036
Kabakov AE, Gabai VL (2018) Cell death and survival assays. In: Calderwood SK, Prince TL (eds) Chaperones: methods and protocols. Springer, New York, pp 107–127
Kanthasamy A, Anantharam V, Ali SF, Kanthasamy AG (2006) Methamphetamine induces autophagy and apoptosis in a mesencephalic dopaminergic neuronal culture model: role of cathepsin-D in methamphetamine-induced apoptotic cell death. Ann N Y Acad Sci 1074:234–244. https://doi.org/10.1196/annals.1369.022
Karila L, Benyamina A (2018) The effects and risks associated with synthetic cathinones use in humans. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 191–202
Kovalevich J, Langford D (2013) Considerations for the use of SH-SY5Y neuroblastoma cells in neurobiology. Methods Mol Biol 1078:9–21. https://doi.org/10.1007/978-1-62703-640-5_2
Larsen KE, Fon EA, Hastings TG, Edwards RH, Sulzer D (2002) Methamphetamine-induced degeneration of dopaminergic neurons involves autophagy and upregulation of dopamine synthesis. J Neurosci 22(20):8951–8960. https://doi.org/10.1523/JNEUROSCI.22-20-08951.2002
Leventelis C, Goutzourelas N, Kortsinidou A et al (2019) Buprenorphine and methadone as opioid maintenance treatments for heroin-addicted patients induce oxidative Stress in blood. Oxid Med Cell Longev 2019:9417048. https://doi.org/10.1155/2019/9417048
Maltese WA, Overmeyer JH (2014) Methuosis: nonapoptotic cell death associated with vacuolization of macropinosome and endosome compartments. Am J Pathol 184(6):1630–1642. https://doi.org/10.1016/j.ajpath.2014.02.028
Marino G, Niso-Santano M, Baehrecke EH, Kroemer G (2014) Self-consumption: the interplay of autophagy and apoptosis. Nat Rev Mol Cell Biol 15(2):81–94. https://doi.org/10.1038/nrm3735
Martinez-Clemente J, Lopez-Arnau R, Abad S, Pubill D, Escubedo E, Camarasa J (2014) Dose and time-dependent selective neurotoxicity induced by mephedrone in mice. PLoS One 9(6):e99002. https://doi.org/10.1371/journal.pone.0099002
Martins MJ, Roque Bravo R, Enea M et al (2018) Ethanol addictively enhances the in vitro cardiotoxicity of cocaine through oxidative damage, energetic deregulation, and apoptosis. Arch Toxicol 92(7):2311–2325. https://doi.org/10.1007/s00204-018-2227-7
Matsunaga T, Morikawa Y, Kamata K et al (2017) α-Pyrrolidinononanophenone provokes apoptosis of neuronal cells through alterations in antioxidant properties. Toxicology 386:93–102. https://doi.org/10.1016/j.tox.2017.05.017
Mbah NE, Overmeyer JH, Maltese WA (2017) Disruption of endolysosomal trafficking pathways in glioma cells by methuosis-inducing indole-based chalcones. Cell Biol Toxicol 33(3):263–282. https://doi.org/10.1007/s10565-016-9369-2
Nara A, Aki T, Funakoshi T, Uemura K (2010) Methamphetamine induces macropinocytosis in differentiated SH-SY5Y human neuroblastoma cells. Brain Res 1352:1–10. https://doi.org/10.1016/j.brainres.2010.07.043
Nara A, Aki T, Funakoshi T, Unuma K, Uemura K (2012) Hyperstimulation of macropinocytosis leads to lysosomal dysfunction during exposure to methamphetamine in SH-SY5Y cells. Brain Res 1466:1–14. https://doi.org/10.1016/j.brainres.2012.05.017
Naseri G, Fazel A, Golalipour MJ et al (2018) Exposure to mephedrone during gestation increases the risk of stillbirth and induces hippocampal neurotoxicity in mice offspring. Neurotoxicol Teratol 67:10–17. https://doi.org/10.1016/j.ntt.2018.03.001
Panel M, Ghaleh B, Morin D (2018) Mitochondria and aging: a role for the mitochondrial transition pore? Aging Cell 17(4):e12793. https://doi.org/10.1111/acel.12793
Papaseit E, Olesti E, de la Torre R, Torrens M, Farre M (2017) Mephedrone concentrations in cases of clinical intoxication. Curr Pharm Des 23(36):5511–5522. https://doi.org/10.2174/1381612823666170704130213
Pearson JM, Hargraves TL, Hair LS et al (2012) Three fatal intoxications due to methylone. J Anal Toxicol 36(6):444–451. https://doi.org/10.1093/jat/bks043
Pereira-Oliveira M, Reis-Mendes A, Carvalho F, Remiao F, Bastos ML, Costa VM (2019) Doxorubicin is key for the cardiotoxicity of FAC (5-fluorouracil + adriamycin + cyclophosphamide) combination in differentiated H9c2 cells. Biomolecules. https://doi.org/10.3390/biom9010021
Perry SW, Norman JP, Barbieri J, Brown EB, Gelbard HA (2011) Mitochondrial membrane potential probes and the proton gradient: a practical usage guide. Biotechniques 50(2):98–115. https://doi.org/10.2144/000113610
Pivtoraiko VN, Harrington AJ, Mader BJ et al (2010) Low-dose bafilomycin attenuates neuronal cell death associated with autophagy-lysosome pathway dysfunction. J Neurochem 114(4):1193–1204. https://doi.org/10.1111/j.1471-4159.2010.06838.x
PreADMET® (2020) https://preadmet.bmdrc.kr/adme/. Accessed 28 Feb 2020
Qi L, Gang L, Hang KW et al (2011) Programmed neuronal cell death induced by HIV-1 tat and methamphetamine. Microsc Res Tech 74(12):1139–1144. https://doi.org/10.1002/jemt.21006
Rahman MA, Bishayee K, Huh SO (2016) Angelica polymorpha maxim induces apoptosis of human SH-SY5Y neuroblastoma cells by regulating an intrinsic caspase pathway. Mol Cells 39(2):119–128. https://doi.org/10.14348/molcells.2016.2232
Romero A, Ramos E, Ares I et al (2017) Oxidative stress and gene expression profiling of cell death pathways in alpha-cypermethrin-treated SH-SY5Y cells. Arch Toxicol 91(5):2151–2164. https://doi.org/10.1007/s00204-016-1864-y
Rouxinol D, Carmo H, Carvalho F, Bastos MdL, Dias da Silva D (2019) Pharmacokinetics, pharmacodynamics, and toxicity of the new psychoactive substance 3,4-dimethylmethcathinone (3,4-DMMC). Forensic Toxicol. https://doi.org/10.1007/s11419-019-00494-x
Safhi MM, Alam MF, Hussain S et al (2014) Cathinone, an active principle of Cathaedulis, accelerates oxidative stress in the limbic area of swiss albino mice. J Ethnopharmacol 156:102–106. https://doi.org/10.1016/j.jep.2014.08.004
Schifano F, Orsolini L, Papanti D, Corkery J (2017) NPS: medical consequences associated with their intake. In: Baumann MH, Glennon RA, Wiley JL (eds) Neuropharmacology of new psychoactive substances (NPS): the science behind the headlines. Springer International Publishing, Cham, pp 351–380
Siedlecka-Kroplewska K, Wrońska A, Stasiłojć G, Kmieć Z (2018) The designer drug 3-fluoromethcathinone induces oxidative stress and activates autophagy in HT22 neuronal cells. Neurotox Res 34(3):388–400. https://doi.org/10.1007/s12640-018-9898-y
Simmler LD (2018) Monoamine transporter and receptor interaction profiles of synthetic cathinones. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 97–115
Simmons SJ, Kim E, Gentile TA, Murad A, Muschamp JW, Rawls SM (2018) Behavioral profiles and underlying transmitters/circuits of cathinone-derived psychostimulant drugs of abuse. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 125–152
Sivandzade F, Bhalerao A, Cucullo L (2019) Analysis of the mitochondrial membrane potential using the cationic JC-1 dye as a sensitive fluorescent probe. Bio Protoc 9(1):e3128. https://doi.org/10.21769/BioProtoc.3128
Soares J, Costa VM, Gaspar H et al (2019) Structure-cytotoxicity relationship profile of 13 synthetic cathinones in differentiated human SH-SY5Y neuronal cells. Neurotoxicology 75:158–173. https://doi.org/10.1016/j.neuro.2019.08.009
Sulzer D, Zecca L (2000) Intraneuronal dopamine-quinone synthesis: a review. Neurotox Res 1(3):181–195. https://doi.org/10.1007/bf03033289
Teixeira-Gomes A, Costa VM, Feio-Azevedo R, Bastos Mde L, Carvalho F, Capela JP (2015) The neurotoxicity of amphetamines during the adolescent period. Int J Dev Neurosci 41:44–62. https://doi.org/10.1016/j.ijdevneu.2014.12.001
Trabbic CJ, Dietsch HM, Alexander EM et al (2014) Differential induction of cytoplasmic vacuolization and methuosis by novel 2-indolyl-substituted pyridinylpropenones. ACS Med Chem Lett 5(1):73–77. https://doi.org/10.1021/ml4003925
Tsatsakis A, Docea AO, Calina D et al (2019) A mechanistic and pathophysiological approach for stroke associated with drugs of abuse. J Clin Med. https://doi.org/10.3390/jcm8091295
Turcant A, Deguigne M, Ferec S et al (2017) A 6-year review of new psychoactive substances at the Centre antipoison Grand-Ouest d’Angers: clinical and biological data. Toxicol Anal Clin 29(1):18–33. https://doi.org/10.1016/j.toxac.2016.12.001
UNODC (2017) World drug report 2017, vol. 4. Market analysis of synthetic drugs: Amphetamine-type stimulants, new psychoactive substances. United Nations Office on Drugs and Crime, Vienna
Usui K, Aramaki T, Hashiyada M, Hayashizaki Y, Funayama M (2014) Quantitative analysis of 3,4-dimethylmethcathinone in blood and urine by liquid chromatography-tandem mass spectrometry in a fatal case. Leg Med 16(4):222–226. https://doi.org/10.1016/j.legalmed.2014.03.008
Valente MJ, Amaral C, Correia-da-Silva G et al (2017a) Methylone and MDPV activate autophagy in human dopaminergic SH-SY5Y cells: a new insight into the context of β-keto amphetamines-related neurotoxicity. Arch Toxicol 91(11):3663–3676. https://doi.org/10.1007/s00204-017-1984-z
Valente MJ, Bastos MdL, Fernandes E, Carvalho F, Guedes de Pinho P, Carvalho M (2017b) Neurotoxicity of β-keto amphetamines: deathly mechanisms elicited by methylone and MDPV in human dopaminergic SH-SY5Y cells. ACS Chem Neurosci 8(4):850–859. https://doi.org/10.1021/acschemneuro.6b00421
Venkatesan A, Uzasci L, Chen Z et al (2011) Impairment of adult hippocampal neural progenitor proliferation by methamphetamine: role for nitrotyrosination. Mol Brain 4(1):28. https://doi.org/10.1186/1756-6606-4-28
Vermes I, Haanen C, Steffens-Nakken H, Reutelingsperger C (1995) A novel assay for apoptosis flow cytometric detection of phosphatidylserine expression on early apoptotic cells using fluorescein labelled annexin V. J Immunol Methods 184(1):39–51. https://doi.org/10.1016/0022-1759(95)00072-i
Wiergowski M, Aszyk J, Kaliszan M et al (2017) Identification of novel psychoactive substances 25B-NBOMe and 4-CMC in biological material using HPLC-Q-TOF-MS and their quantification in blood using UPLC-MS/MS in case of severe intoxications. J Chromatogr B Anal Technol Biomed Life Sci 1041–1042:1–10. https://doi.org/10.1016/j.jchromb.2016.12.018
Wojcieszak J, Andrzejczak D, Woldan-Tambor A, Zawilska JB (2016) Cytotoxic activity of pyrovalerone derivatives, an emerging group of pychostimulant designer cathinones. Neurotox Res 30(2):239–250. https://doi.org/10.1007/s12640-016-9640-6
Wojcieszak J, Andrzejczak D, Kedzierska M, Milowska K, Zawilska JB (2018) Cytotoxicity of alpha-pyrrolidinophenones: an impact of alpha-aliphatic side-chain length and changes in the plasma membrane fluidity. Neurotox Res 34(3):613–626. https://doi.org/10.1007/s12640-018-9923-1
Wu CW, ** YH, Yen JC et al (2007) Enhanced oxidative stress and aberrant mitochondrial biogenesis in human neuroblastoma SH-SY5Y cells during methamphetamine induced apoptosis. Toxicol Appl Pharmacol 220(3):243–251. https://doi.org/10.1016/j.taap.2007.01.011
Wurita A, Hasegawa K, Minakata K et al (2014) Postmortem distribution of alpha-pyrrolidinobutiophenone in body fluids and solid tissues of a human cadaver. Leg Med 16(5):241–246. https://doi.org/10.1016/j.legalmed.2014.05.001
**ong Q, Ru Q, Tian X et al (2018) Krill oil protects PC12 cells against methamphetamine-induced neurotoxicity by inhibiting apoptotic response and oxidative stress. Nutr Res 58:84–94. https://doi.org/10.1016/j.nutres.2018.07.006
Yamamoto BK, Moszczynska A, Gudelsky GA (2010) Amphetamine toxicities: classical and emerging mechanisms. Ann N Y Acad Sci 1187:101–121. https://doi.org/10.1111/j.1749-6632.2009.05141.x
Yamamuro A, Kishino T, Ohshima Y et al (2011) Caspase-4 directly activates caspase-9 in endoplasmic reticulum stress-induced apoptosis in SH-SY5Y cells. J Pharmacol Sci 115(2):239–243. https://doi.org/10.1254/jphs.10217sc
Zaitsu K (2018) Metabolism of synthetic cathinones. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 71–96
Zawilska JB, Wojcieszak J (2018) Novel psychoactive substances: classification and general information. In: Zawilska JB (ed) Synthetic cathinones: novel addictive and stimulatory psychoactive substances. Springer International Publishing, Cham, pp 11–24
Acknowledgements
Jorge Soares acknowledges University of Porto/Faculty of Medicine University of Porto through FSE—Fundo Social Europeu, NORTE2020—Programa Operacional Regional do Norte for his grant (NORTE-08-5369-FSE-000011). This work was supported by the Applied Molecular Biosciences Unit—UCIBIO through UID/MULTI/04378/2019 support with funding from FCT/MCTES through national funds. Centro de Química Estrutural, BioISI—Biosystems and Integrative Sciences Institute, and MARE—Marine and Environmental Sciences Centre, acknowledges the financial support of Fundação para a Ciência e Tecnologia—FCT (UIDB/00100/2020, UID/MULTI/04046/2019 and UIDB/04292/2020, respectively). Vera Marisa Costa acknowledges Fundação da Ciência e Tecnologia (FCT) for her grant (SFRH/BPD/110001/2015) that was funded by national funds through FCT—Fundação para a Ciência e a Tecnologia, I.P., under the Norma Transitória—DL57/2016/CP1334/CT0006. The authors wish to thank the Laboratório de Polícia Científica da Polícia Judiciária (LPC-PJ) for providing the smartshop product within the scope of the protocol established between LPC-PJ, FCUL, and FFUP.
Author information
Authors and Affiliations
Corresponding authors
Ethics declarations
Conflict of interest
The authors declare that there is no conflict of interest.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Electronic supplementary material
Below is the link to the electronic supplementary material.
204_2020_2761_MOESM1_ESM.tiff
Supplementary file1. Supplementary Table 1: Concentrations of 3,4-DMMC, 4-MMC and METH inducing 25, 50 and 75 % of toxicity (IC25, IC50 and IC75, respectively) in RA-TPA differentiated SH-SY5Y cells after a 24-h exposure. MTT and NR concentration-response curves were fitted using least squares as the fitting method and the comparisons between the curves concerning the two cytotoxicity assays performed (MTT reduction and NR uptake) were made using the extra sum-of-squares F test [*p < 0.05, ***p < 0.001 4-MMC (NR uptake) vs. 4-MMC (MTT reduction); ##p < 0.01 METH (NR uptake) vs. METH (MTT reduction)]. Data, as mean and 95 % confidence Intervals (95 % CI), are presented in mM. At the bottom of the table, the MTT and NR concentration-response curves can be observed (TIFF 1390 kb)
Rights and permissions
About this article
Cite this article
Soares, J., Costa, V.M., Gaspar, H. et al. Adverse outcome pathways induced by 3,4-dimethylmethcathinone and 4-methylmethcathinone in differentiated human SH-SY5Y neuronal cells. Arch Toxicol 94, 2481–2503 (2020). https://doi.org/10.1007/s00204-020-02761-y
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00204-020-02761-y