Download PDF
CC BY-NC-ND 4.0 · Planta Medica International Open 2021; 8(03): e88-e95
DOI: 10.1055/a-1516-4182
Original Papers

Neuroprotective Effects of Delta-9-Tetrahydrocannabinol against FeSO4- and H2O2-Induced Cell Damage on Dopaminergic Neurons in Primary Mesencephalic Cell Culture

Rudolf Moldzio‡
1   Institute of Medical Biochemistry, Department of Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria
,
Alexander Unterberger‡
1   Institute of Medical Biochemistry, Department of Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria
,
Christopher Krewenka
1   Institute of Medical Biochemistry, Department of Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria
,
Barbara Kranner
1   Institute of Medical Biochemistry, Department of Biomedical Sciences, University of Veterinary Medicine Vienna, Vienna, Austria
,
Khaled Radad
2   Department of Pathology, Faculty of Veterinary Medicine, Assiut University, Assiut, Egypt
3   Department of Pathology, College of Medicine, King Khalid University, Abha, Saudi Arabia
› Author Affiliations
› Further Information
   Permissions and Reprints

Abstract

Delta-9-Tetrahydrocannabinol and other phytocannabinoids have been previously demonstrated to possess neuroprotective effects in murine mesencephalic cell culture models of Parkinson’s disease, in which increased levels of superoxide radicals led to the loss of dopaminergic neurons. In these models, delta-9-tetrahydrocannabinol did not scavenge these radicals but displayed antioxidative capacity by increasing glutathione levels. Based on these findings, in the present study, we investigated whether the neuroprotective effect of delta-9-tetrahydrocannabinol can also be detected in FeSO4- and H2O2-stressed cells. Mesencephalic cultures were concomitantly treated with FeSO4 (350 μM) or H2O2 (150 μM) and delta-9-tetrahydrocannabinol (0.01, 0.1, 1, 10 μM) on the 12th days in vitro for 48 h. On the 14th DIV, dopaminergic neurons were stained immunocytochemically by tyrosine hydroxylase, and fluorescently using crystal violet, Hoechst 33342, and JC-1. FeSO4 and H2O2 significantly reduced the number of dopaminergic neurons by 33 and 36%, respectively, and adversely affected the morphology of surviving neurons. Moreover, FeSO4, but not H2O2, significantly decreased the fluorescence intensity of crystal violet and Hoechst 33342, and reduced the red/green ratio of JC-1. Co-treatment with delta-9-tetrahydrocannabinol at the concentrations 0.01 and 0.1 μM significantly rescued dopaminergic neurons in FeSO4 and H2O2-treated cultures by 16 and 30%, respectively. delta-9-Tetrahydrocannabinol treatment also led to a higher fluorescence intensity of crystal violet and Hoechst 33342, and increased the red/green fluorescence ratio of JC-1 when concomitantly administered with FeSO4 but not H2O2. To conclude, delta-9-tetrahydrocannabinol rescues dopaminergic neurons against FeSO4- and H2O2-induced neurotoxicity. Using fluorescence dyes, this effect seems to be mediated partially by restoring mitochondrial integrity and decreasing cell death, particularly in FeSO4-treated cultures.

Key words

delta-9-tetrahydrocannabinol - neuroprotection - Parkinson’s disease - oxidative stress - dopaminergic neurons

Supplementary Material



Publication History

Received: 31 January 2021
Received: 31 March 2021

Accepted: 04 May 2021

Article published online:
11 August 2021

© 2021. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution-NonDerivative-NonCommercial-License, permitting copying and reproduction so long as the original work is given appropriate credit. Contents may not be used for commercial purposes, or adapted, remixed, transformed or built upon. (https://creativecommons.org/licenses/by-nc-nd/4.0/).

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

 

  • References

  • 1 Russo EB. History of cannabis and its preparations in saga, science, and sobriquet. Chem Biodivers 2007; 4: 1614-1648
    CrossrefPubMedGoogle Scholar
  • 2 Mechoulam R. The pharmacohistory of Cannabis sativa. In: Mechoulam R. Cannabinoids as therapeutic agents. Boca Raton, FL: CRC Press; 1986: 1-19
    CrossrefGoogle Scholar
  • 3 Bielawiec P, Harasim SE, Chabowski A. Phytocannabinoids: Useful Drugs for the Treatment of Obesity? Special Focus on Cannabidiol. Front Endocrinol (Lausanne) 2020; 11: 114
    CrossrefPubMedGoogle Scholar
  • 4 Maroon J, Bost J. Review of the neurological benefits of phytocannabinoids. Surg Neurol Int 2018; 9: 91
    CrossrefPubMedGoogle Scholar
  • 5 Pertwee RG. The diverse CB1 and CB2 receptor pharmacology of three plant cannabinoids: delta9-tetrahydrocannabinol, cannabidiol and delta9-tetrahydrocannabivarin. Br J Pharmacol 2008; 153: 199-215
    CrossrefPubMedGoogle Scholar
  • 6 Pisanti S, Malfitano AM, Ciaglia E, Lamberti A, Ranieri R, Cuomo G, Abate M, Faggiana G, Proto MC, Fiore D, Laezza C, Bifulco M. Cannabidiol: State of the art and new challenges for therapeutic applications. Pharmacol Ther 2017; 175: 133-150
    CrossrefPubMedGoogle Scholar
  • 7 Athanasiou A, Clarke AB, Turner AE, Kumaran NM, Vakilpour S, Smith PA, Bagiokou D, Bradshaw TD, Westwell AD, Fang L, Lobo DN, Constantinescu CS, Calabrese V, Loesch A, Alexander SPH, Clothier RH, Kendall DA, Bates TE. Cannabinoid receptor agonists are mitochondrial inhibitors: A unified hypothesis of how cannabinoids modulate mitochondrial function and induce cell death. Biochem Biophys Res Commun 2007; 364: 131-137
    CrossrefPubMedGoogle Scholar
  • 8 Wolff V, Schlagowski AI, Rouyer O, Charles AL, Singh F, Auger C, Schini-Kerth V, Marescaux C, Raul JS, Zoll J, Geny B. Tetrahydrocannabinol induces brain mitochondrial respiratory chain dysfunction and increases oxidative stress: A potential mechanism involved in cannabis-related stroke. Biomed Res Int 2015; 2015: 323706
    CrossrefPubMedGoogle Scholar
  • 9 Moldzio R, Pacher T, Krewenka C, Kranner B, Novak J, Duvigneau JC, Rausch WD. Effects of cannabinoids Δ(9)-tetrahydrocannabinol, Δ(9)-tetrahydrocannabinolic acid and cannabidiol in MPP+affected murine mesencephalic cultures. Phytomedicine 2012; 15: 819-824
    CrossrefPubMedGoogle Scholar
  • 10 Pöhn B, Krewenka C, Kranner B, Duvigneau JC, Rausch WD, Moldzio R. Phytocannabinoids tetrahydrocannabinol and cannabidiol act against rotenone induced damages in murine cell cultures. Planta Med 2012; 78: PD171
    Article in Thieme ConnectPubMedGoogle Scholar
  • 11 Poewe W, Seppi K, Tanner C, Halliday G, Brundin P, Volkmann J, Schrag A-E, Lang AE. Parkinson disease. Nat Rev Dis Primers 2017; 3: 17013
    CrossrefPubMedGoogle Scholar
  • 12 Váradi C. Clinical Features of Parkinson’s Disease: The Evolution of Critical Symptoms. Biology (Basel) 2020; 9: 103
    PubMedGoogle Scholar
  • 13 Rokad D, Ghaisas S, Harischandra DS, Jin H, Anantharam V, Kanthasamy A, Kanthasamy AG. Role of neurotoxicants and traumatic brain injury in α-synuclein protein misfolding and aggregation. Brain Res Bull 2017; 133: 60-70
    CrossrefPubMedGoogle Scholar
  • 14 Kalinderi K, Bostantjopoulou S, Fidani L. The genetic background of Parkinson’s disease: current progress and future prospects. Acta Neurol Scand 2016; 134: 314-326
    CrossrefPubMedGoogle Scholar
  • 15 Jenner P, Olanow CW. The pathogenesis of cell death in Parkinson’s disease. Neurology 2006; 66: S24-S36
    CrossrefPubMedGoogle Scholar
  • 16 Farooqui T, Farooqui A. Lipid-mediated oxidative stress and inflammation in the pathogenesis of Parkinson’s disease. Parkinson’s Disease 2011; 2011: 247467
    PubMedGoogle Scholar
  • 17 Khan FH, Sen T, Maiti AK, Jana S, Chatterjee U, Chakrabarti S. Inhibition of rat brain mitochondrial electron transport chain activity by dopamine oxidation products during extended in vitro incubation: implications for Parkinson’s disease. Biochimica et Biophysica Acta 2005; 1741: 65-74
    CrossrefPubMedGoogle Scholar
  • 18 Dias V, Junn E, Mouradian MM. The role of oxidative stress in Parkinson’s disease. J Parkinsons Dis 2013; 3: 461-491
    CrossrefPubMedGoogle Scholar
  • 19 Gao G, Change YZ. Mitochondrial ferritin in the regulation of brain iron homeostasis and neurodegenerative diseases. Front Pharmacol 2014; 5: 19
    PubMedGoogle Scholar
  • 20 Ward RJ, Zucca FA, Duyn JH, Crichton RR, Zecca L. The role of iron in brain ageing and neurodegenerative disorders. Lancet Neurol 2014; 13: 1045-1060
    CrossrefPubMedGoogle Scholar
  • 21 Whittemore ER, Loo DT, Cotman CW. A detailed analysis of hydrogen-peroxide-Induced Cell Death in Primary Neuronal Culture. Neuroscience 1995; 61. Neuroscience 1995; 67: 921-932
    CrossrefPubMedGoogle Scholar
  • 22 Dusek P, Roos PM, Litwin T, Schneider SA, Flaten TP, Aaseth J. The neurotoxicity of iron, copper and manganese in Parkinson’s and Wilson’s diseases. J Trace Elem Med Biol 2015; 31: 193-203
    CrossrefPubMedGoogle Scholar
  • 23 Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J. Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol 2007; 39: 44-84
    CrossrefPubMedGoogle Scholar
  • 24 Krishnan CV, Garnett M, Chu B. Spatiotemporal oscillations in biological molecules: Hydrogen peroxide and Parkinson’s disease. Int J Electrochem Sci 2008; 3: 1364-1385
    CrossrefPubMedGoogle Scholar
  • 25 Michel PP, Vyas S, Agid Y. Toxic effects of iron for cultured mesencephalic dopaminergic neurons derived from rat embryonic brains. J Neurochem 1992; 59: 118-127
    CrossrefPubMedGoogle Scholar
  • 26 Reichelt D, Radad K, Moldzio R, Rausch WD, Reichmann H, Gille G. Comparable Neuroprotective Effects of Pergolide and Pramipexole on Ferrous Sulfate-Induced Dopaminergic Cell Death in Cell Culture. CNS Neurol Disord Drug Targets 2016; 15: 1325-1332
    CrossrefPubMedGoogle Scholar
  • 27 Du X, Xu H, Jiang H, Xie J. Akt/Nrf2 activated upregulation of heme oxygenase-1 involves in the role of Rg1 against ferrous iron-induced neurotoxicity in SK-N-SH cells. Neurotox Res 2013; 24: 71-79
    CrossrefPubMedGoogle Scholar
  • 28 Liu R, Liu W, Doctrow SR, Baudry M. Iron toxicity in organotypic cultures of hippocampal slices: role of reactive oxygen species. J Neurochem 2003; 85: 492-502
    CrossrefPubMedGoogle Scholar
  • 29 Ayton S, Lei P, Adlard PA, Volitakis I, Cherny RA, Bush AI, Finkelstein DI. Iron accumulation confers neurotoxicity to a vulnerable population of nigral neurons: implications for Parkinson’s disease. Mol Neurodegener 2014; 9: 27
    CrossrefPubMedGoogle Scholar
  • 30 Campos SS, Diez GR, Oresti GM, Salvador GA. Dopaminergic Neurons Respond to Iron-Induced Oxidative Stress by Modulating Lipid Acylation and Deacylation Cycles. PLoS One 2015; 10: e0130726
    CrossrefPubMedGoogle Scholar
  • 31 Ismail N, Ismail M, Fathy SF, Musa SNA, Imam MU, Foo JB, Iqbal S. Neuroprotective effects of germinated brown rice against hydrogen peroxide induced cell death in human SH-SY5Y cells. Int J Mol Sci 2012; 13: 9692-9708
    CrossrefPubMedGoogle Scholar
  • 32 Nguyen CH, Krewenka C, Radad K, Kranner B, Huber A, Duvigneau JC, Miller I, Moldzio R. THC (Δ9-Tetrahydrocannabinol) Exerts Neuroprotective Effect in Glutamate-affected Murine Primary Mesencephalic Cultures Through Restoring Mitochondrial Membrane Potential and Anti-apoptosis Involving CB1 Receptor-dependent Mechanism. Phytother Res 2016; 30: 2044-2052
    CrossrefPubMedGoogle Scholar
  • 33 Lastres-Becker I, Molina-Holgado F, Ramos JA, Mechoulam R, Fernández-Ruiz J. Cannabinoids provide neuroprotection against 6-hydroxydopamine toxicity in vivo and in vitro: relevance to Parkinson’s disease. Neurobiol Dis 2005; 19: 96-107
    CrossrefPubMedGoogle Scholar
  • 34 Hampson AJ, Grimaldi M, Axelrod J, Wink D. Cannabidiol and (-) Delta9-tetrahydrocannabinol are neuroprotective antioxidants. Proc Natl Acad Sci USA 1998; 95: 8268-8273
    CrossrefPubMedGoogle Scholar
  • 35 Chen J, Lee CT, Errico S, Deng X, Cadet JL, Freed WJ. Protective effects of Delta (9)-tetrahydrocannabinol against N-methyl-d-aspartate-induced AF5 cell death. Brain Res Mol Brain Res 2005; 134: 215-225
    CrossrefPubMedGoogle Scholar
  • 36 Rosales-Corral S, Hernández L, Gallegos M. Cannabinoids in neuroinflammation, oxidative stress and neuro excitotoxicity. Pharm Anal Acta 2015; 6: 3
    PubMedGoogle Scholar
  • 37 Zeissler M, Eastwood J, Mccorry K, Hanemann CO, Zajicek JP, Carroll CB. Delta-9-tetrahydrocannabinol protects against MPP+toxicity in SH-SY5Y cells by restoring proteins involved in mitochondrial biogenesis. Oncotarget 2016; 7: 46603-46614
    CrossrefPubMedGoogle Scholar
  • 38 Marsicano G, Moosmann B, Hermann H, Lutz B, Behl C. Neuroprotective properties of cannabinoids against oxidative stress: role of the cannabinoid receptor CB1. J Neurochem 2002; 80: 448-456
    CrossrefPubMedGoogle Scholar
  • 39 Radad K, Al-Shraim M, Al-Emam A, Moldzio R, Rausch WD. Neurotoxic effects of acrylamide on dopaminergic neurons in primary mesencephalic cell culture. Folia Neuropathol 2019; 57: 196-204
    CrossrefPubMedGoogle Scholar
  • 40 Feoktistova M, Geserick P, Leverkus M. Crystal violet assay for determining viability of cultured cells. Cold Spring Harb Protoc 2016; 2016: pdb.prot087379
    CrossrefPubMedGoogle Scholar
  • 41 Duvigneau JC, Trovato A, Müllebner A, Miller I, Krewenka C, Krenn K, Zich W, Moldzio R. Cannabidiol Protects Dopaminergic Neurons in Mesencephalic Cultures against the Complex I Inhibitor Rotenone Via Modulation of Heme Oxygenase Activity and Bilirubin. Antioxidants (Basel) 2020; 9: 135
    CrossrefPubMedGoogle Scholar
  • 42 Chazotte B. Labeling nuclear DNA with hoechst 33342. Cold Spring Harb Protoc 2011; 2011: pdb.prot5557
    CrossrefPubMedGoogle Scholar
  • 43 Crowley LC, Marfell BJ, Waterhouse NJ. Analyzing Cell Death by Nuclear Staining with Hoechst 33342. Cold Spring Harb Protoc 2016; 2016 DOI: 10.1101/pdb.prot087205.
    CrossrefPubMedGoogle Scholar
  • 44 White RJ, Reynolds IJ. Mitochondrial depolarization in glutamate-stimulated neurons: An early signal specific to excitotoxin exposure. J Neurosci 1996; 16: 5688-5697
    CrossrefPubMedGoogle Scholar