Abstract
Studies have indicated that epilepsy, an important neurological disease, can generate oxidative stress and mitochondrial dysfunction, among other damages to the brain. In this context, the use of antioxidant compounds could provide neuroprotection and help to reduce the damage caused by epileptic seizures and thereby the use of anticonvulsant drugs. Rosmarinic acid (RA) is an ester of caffeic acid and 3,4-dihydroxyphenylactic acid that prevents cell damage caused by free radicals, acting as an antioxidant. It also presents anti-inflammatory, antimutagenic, and antiapoptotic properties. In this work, we used two models of acute seizure, 4-aminopyridine (4-AP) and picrotoxin (PTX)-induced seizures in mice, to investigate the anticonvulsant, antioxidant, and neuroprotective profile of RA. Diazepam and valproic acid, antiepileptic drugs already used in the treatment of epilepsy, were used as positive controls. Although RA could not prevent seizures in the models used in this study, neither enhance the latency time to first seizure at the tested doses, it exhibited an antioxidant and neuroprotective effect. RA (8 and 16 mg/kg) decreased reactive oxygen species production, superoxide dismutase activity, and DNA damage, measured in hippocampus, after seizures induced by PTX and 4-AP. Catalase activity was decreased by RA only after seizures induced by 4-AP. The activity of the mitochondrial complex II was increased by RA in hippocampus samples after both seizure models. The results obtained in this study suggest that RA is able to reduce cell damage generated by the 4-AP and PTX seizures and therefore could represent a potential candidate in reducing pathophysiological processes involved in epilepsy.
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References
Aebi H (1984) Catalase in Vitro. Methods Enzymol 105:121–126. https://doi.org/10.1016/S0076-6879(84)05016-3
Avoli M, de Curtis M (2011) GABAergic synchronization in the limbic system and its role in the generation of epileptiform activity. Prog Neurobiol 95:104–132. https://doi.org/10.1016/j.pneurobio.2011.07.003
Awad R, Muhammad A, Durst T, Trudeau VL, Arnason JT (2009) Bioassay-guided fractionation of lemon balm (Melissa officinalis L .) using an In Vitro measure of GABA transaminase activity. Phytother Res 23:1075–1081. https://doi.org/10.1002/ptr
Bhat AH, Dar KB, Anees S, Zargar MA, Masood A, Sofi MA, Ganie SA (2015) Oxidative stress, mitochondrial dysfunction and neurodegenerative diseases; a mechanistic insight. Biomed Pharmacother 74:101–110. https://doi.org/10.1016/j.biopha.2015.07.025
Coelho VR, Vieira CG, de Souza LP, Moysés F, Basso C, Papke DKM, Pires TR, Siqueira IR, Picada JN, Pereira P (2015) Antiepileptogenic, antioxidant and genotoxic evaluation of rosmarinic acid and its metabolite caffeic acid in mice. Life Sci 122:65–71. https://doi.org/10.1016/j.lfs.2014.11.009
Coelho VR, Vieira CG, de Souza LP, da Silva LL, Pfluger P, Regner GG, Papke DKM, Picada JN, Pereira P (2016) Behavioral and genotoxic evaluation of rosmarinic and caffeic acid in acute seizure models induced by pentylenetetrazole and pilocarpine in mice. Naunyn Schmiedeberg's Arch Pharmacol 389:1195–1203. https://doi.org/10.1007/s00210-016-1281-z
Coelho VR, Viau CM, Staub RB, de Souza MS, Pfluger P, Regner GG, Pereira P, Saffi J (2017) Rosmarinic acid attenuates the activation of murine microglial N9 cells through the downregulation of inflammatory cytokines and cleaved Caspase-3. Neuroimmunomodulation 24:171–181. https://doi.org/10.1159/000481095
da Silva CG, Ribeiro CAJ, Leipnitz G, Dutra CS, Wyse ATS, Wannmacher CMD, Sarkis JJF, Jakobs C, Wajner M (2002) Inhibition of cytochrome c oxidase activity in rat cerebral cortex and human skeletal muscle by D-2-hydroxyglutaric acid in vitro. Biochim Biophys Acta 1586:81–91. https://doi.org/10.1016/S09254439(01)00088-6
De Oliveira NCD, Sarmento MS, Nunes EA, Porto CM, Rosa DP, Bona SR, Rodrigues G, Marroni NP, Pereira P, Picada JN, Ferraz ABF, Thiesen FV, da Silva J (2012) Rosmarinic acid as a protective agent against genotoxicity of ethanol in mice. Food Chem Toxicol 50:1208–1214. https://doi.org/10.1016/j.fct.2012.01.028
Devinsky O, Vezzani A, De Lanerolle NC, Rogawski MA (2013) Glia and epilepsy: excitability and inflammation. Trends Neurosci 36:174–184. https://doi.org/10.1016/j.tins.2012.11.008
Dóczi J, Banczerowski-Pelyhe I, Barna B, Világi I (1999) Effect of a glutamate receptor antagonist (GYKI 52466) on 4-aminopyridine-induced seizure activity developed in rat cortical slices. Brain Res Bull 49(6):435–440. https://doi.org/10.1016/S0361-9230(99)00079-9
Fachel FNS, Schuh RS, Veras KS, Bassani VL, Koester LS, Henriques AT, Braganhol E, Teixeira HF (2019) An overview of the neuroprotective potential of rosmarinic acid and its association with nanotechnology-based delivery systems: a novel approach to treating neurodegenerative disorders. Neurochem Int 122:47–58. https://doi.org/10.1016/j.neuint.2018.11.003
Fallarini S, Miglio G, Paoletti T, Minassi A, Amoruso A, Bardelli C, Brunelleschi S, Lombardi G (2009) Clovamide and rosmarinic acid induce neuroprotective effects in in vitro models of neuronal death. Br J Pharmacol 157:1072–1084. https://doi.org/10.1111/j.1476-5381.2009.00213.x
Fischer JC, Ruitenbeek W, Berden JA, Trijbels JMF, Veerkamp JH, Stadhouders AM, Sengers RCA, Janssen AJM (1985) Differential investigation of the capacity of succinate oxidation in human skeletal muscle. Clin Chim Acta 153:23–36. https://doi.org/10.1016/0009-8981(85)90135-4
Fisher RS, Boas WE, Blume W, Elger C, Genton P, Lee P, Engel J (2005) Epileptic seizures and epilepsy: definitions proposed by the international league against epilepsy (ILAE) and the International Bureau for Epilepsy (IBE). Epilepsia 46(4):470–472. https://doi.org/10.1111/j.0013-9580.2005.66104.x
Fisher RS, Acevedo C, Arzimanoglou A, Bogacz A, Cross JH, Elger CE, Engel J, Forsgren L, French JA, Glynn M, Hesdorffer DC, Lee BI, Mathern GW, Moshé SL, Perucca E, Scheffer IE, Tomson T, Watanabe M, Wiebe S (2014) A practical clinical definition of epilepsy. Epilepsia 55(4):475–482. https://doi.org/10.1111/epi.12550
Freitas RM, Nascimento VS, Vasconcelos SMM, Sousa FCF, Viana GSB, Fonteles MMF (2004) Catalase activity in cerebellum, hippocampus, frontal cortex and striatum after status epilepticus induced by pilocarpine in Wistar rats. Neurosci Lett 365:102–105. https://doi.org/10.1016/j.neulet.2004.04.060
Gao LP, Wei HL, Zhao HS, Xiao SY, Zheng RL (2005) Antiapoptotic and antioxidant effects of rosmarinic acid in astrocytes. Pharmazie 60:62–65
Gok DK, Hidisoglu E, Ocak GA, Er H, Acun AD, Yargıcoglu P (2018) Protective role of rosmarinic acid on amyloid beta 42-induced echoic memory decline: implication of oxidative stress and cholinergic impairment. Neurochem Int 118:1–13. https://doi.org/10.1016/j.neuint.2018.04.008
Grigoletto J, de Oliveira CV, Grauncke ACB, de Sousa TL, Souto NS, de Freitas ML, Furian AF, Santos ARS, Oliveira MS (2016) Rosmarinic acid is anticonvulsant against seizures induced by pentylenetetrazol and pilocarpine in mice. Epilepsy Behav 62:27–34. https://doi.org/10.1016/j.yebeh.2016.06.037
Grings M, Moura AP, Parmeggiani B, Pletsch JT, Cardoso GMF, August PM, Matté C, Wyse ATS, Wajner M, Leipnitz G (2017) Bezafibrate prevents mitochondrial dysfunction, antioxidant system disturbance, glial reactivity and neuronal damage induced by sulfite administration in striatum of rats: implications for a possible therapeutic strategy for sulfite oxidase deficiency. Biochim Biophys Acta Mol basis Dis 1863(9):2135–2148. https://doi.org/10.1016/j.bbadis.2017.05.019
Guzy RD, Sharma B, Bell E, Chandel NS, Schumacker PT (2008) Loss of the SdhB, but not the SdhA, subunit of complex II triggers reactive oxygen species-dependent hypoxia-inducible factor activation and tumorigenesis. Mol Cell Biol 28(2):718–731. https://doi.org/10.1128/MCB.01338-07
Hartmann A, Speit G (1997) The contribution of cytotoxicity to DNA-effects in the single cell gel test (comet assay). Toxicol Lett 90:183–188. https://doi.org/10.1016/S0378-4274(96)03847-7
Hartmann A, Agurell E, Beevers C, Brendler-Schwaab S, Burlinson B, Clay P, Collins A, Smith A, Speit G, Thybaud V, Tice RR (2003) Recommendations for conducting the in vivo alkaline comet assay. Mutagenesis 18(1):45–51. https://doi.org/10.1093/mutage/18.1.45
Hasan ZA, Razzak RLA, Alzoubi KH (2014) Comparison between the effect of propofol and midazolam on picrotoxin-induced convulsions in rat. Physiol Behav 128:114–118. https://doi.org/10.1016/j.physbeh.2014.01.027
Jacob S, Nair AB (2016) An updated overview on therapeutic drug monitoring of recent antiepileptic drugs. Drugs RD 16:303–316. https://doi.org/10.1007/s40268-016-0148-6
Kalyanaraman B, Darley-Usmar V, Davies KJA, Dennery PA, Forman HJ, Grisham MB, Mann GE, Moore K, Roberts LJ, Ischiropoulos H (2012) Measuring reactive oxygen and nitrogen species with fluorescent probes: challenges and limitations. Free Radic Biol Med 52:1–6. https://doi.org/10.1016/j.freeradbiomed.2011.09.030
Kang M, Yun S, Won J (2003) Rosmarinic acid inhibits Ca2+-dependent pathways of T-cell antigen receptor-mediated signaling by inhibiting the PLC-Gamma1 and Itk activity. Blood 101(9):3534–3542. https://doi.org/10.1182/blood-2002-07-1992
Khamse S, Sadr SS, Roghani M, Hasanzadeh G, Mohammadian M (2015) Rosmarinic acid exerts a neuroprotective effect in the kainate rat model of temporal lobe epilepsy: underlying mechanisms. Pharm Biol 53(12):1818–1825. https://doi.org/10.3109/13880209.2015.1010738
Kudin AP, Kudina TA, Seyfried J, Vielhaber S, Beck H, Elger CE, Kunz WS (2002) Seizure-dependent modulation of mitochondrial oxidative phosphorylation in rat hippocampus. Eur J Neurosci 15:1105–1114. https://doi.org/10.1152/jn.01053.2003
Kwon YO, Hong JT, Oh KW (2017) Rosmarinic acid potentiates pentobarbital-induced sleep behaviors and non-rapid eye movement (NREM) sleep through the activation of GABAa-ergic systems. Biomol Ther 25(2):105–111. https://doi.org/10.4062/biomolther.2016.035
Li X, Fang P, Mai J, Choi ET, Wang H, Yang X (2013) Targeting mitochondrial reactive oxygen species as novel therapy for inflammatory diseases and cancers. J Hematol Oncol 6(1):19. https://doi.org/10.1186/1756-8722-6-19
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275
Martinc B, Grabnar I, Vovk T (2014) Antioxidants as a preventive treatment for epileptic process: a review of the current status. Curr Neuropharmacol 12:527–550. https://doi.org/10.2174/1570159X12666140923205715
Michiels C, Raes M, Toussaint O, Remacle J (1994) Importance of SE-glutathione peroxidase, catalase, and CU/ZN-SOD for cell survival against oxidative stress. Free Radic Biol Med 17(3):235–248. https://doi.org/10.1016/0891-5849(94)90079-5
Misra HP, Fridovich I (1972) The role of superoxide anion in the epinephrine and a simple assay for superoxide dismutase. J Biol Chem 247(10):3170–3175
Murphy MP (2009) How mitochondria produce reactive oxygen species. Biochem J 417:1–13. https://doi.org/10.1042/BJ20081386
Najjar S, Pearlman D, Miller DC, Devinsky O (2011) Refractory epilepsy associated with microglial activation. Neurologist 17(5):249–254. https://doi.org/10.1097/NRL.0b013e31822aad04
Ngugi AK, Bottomley C, Kleinschmidt I, Sander J, Newton CR (2010) Estimation of the burden of active and life-time epilepsy: A meta-analytic approach. Epilepsia 51:883–889. https://doi.org/10.1111/j.1528-1167.2009.02481.x
Orr AL, Quinlan CL, Perevoshchikova IV, Brand MD (2012) A refined analysis of superoxide production by mitochondrial sn-glycerol 3-phosphate dehydrogenase. J Biol Chem 287(51):42921–42935. https://doi.org/10.1074/jbc.M112.397828
Paranagama MP, Sakamoto K, Amino H, Awano M, Miyoshi H, Kita K (2010) Contribution of the FAD and quinone binding sites to the production of reactive oxygen species from Ascaris suum mitochondrial complex II. Mitochondrion 10:158–165. https://doi.org/10.1016/j.mito.2009.12.145
Patel M (2004) Mitochondrial dysfunction and oxidative stress: cause and consequence of epileptic seizures. Free Radic Biol Med 37:1951–1962. https://doi.org/10.1016/j.freeradbiomed.2004.08.021
Peña F, Tapia R (2000) Seizures and neurodegeneration induced by 4-aminopyridine in rat hippocampus in vivo: role of glutamate- and GAMA-mediated neurotransmission and of ion channels. Neuroscience 101(3):547–561
Pereira P, De Oliveira PA, Ardenghi P, Rotta L, Henriques JAP, Picada JN (2006) Neuropharmacological analysis of Caffeic acid in rats. Basic Clin Pharmacol Toxicol 99:374–378. https://doi.org/10.1016/j.phrs.2005.03.003
Pflüger P, Regner GG, Coelho VR, da Silva LL, Nascimento L, Viau CM, Zanette RA, Hoffmann C, Picada JN, Saffi J, Pereira P (2018) Gamma-Decanolactone improves biochemical parameters associated with pilocarpine-induced seizures in male mice. Curr Mol Pharmacol 11(2):162–169. https://doi.org/10.2174/1874467210666171002114954
Quinlan CL, Orr AL, Perevoshchikova IV, Treberg JR, Ackrell BA, Brand MD (2012) Mitochondrial complex II can generate reactive oxygen species at high rates in both the forward and reverse. J Biol Chem 287(32):27255–27264. https://doi.org/10.1074/jbc.M112.374629
Rahbardar MG, Amin B, Mehri S, Mirnajafi-Zadeh SJ, Hosseinzadeh H (2018) Rosmarinic acid attenuates development and existing pain in a rat model of neuropathic pain: an evidence of anti-oxidative and anti-inflammatory effects. Phytomedicine 40:59–67. https://doi.org/10.1016/j.phymed.2018.01.001
Rizk HA, Masoud MA, Maher OW (2017) Prophylactic effects of ellagic acid and rosmarinic acid on doxorubicin-induced neurotoxicity in rats. J Biochem Mol Toxicol 31(12). https://doi.org/10.1002/jbt.21977
Rocha J, Eduardo-Figueira M, Barateiro A, Fernandes A, Brites D, Bronze R, Duarte CMM, Serra AT, Pinto R, Freitas M, Fernandes E, Silva-Lima B, Mota-Filipe H, Sepodes B (2015) Anti-inflammatory effect of Rosmarinic acid and an extract of Rosmarinus officinalis in rat models of local and systemic inflammation. Basic Clin Pharmacol Toxicol 116:398–413. https://doi.org/10.1111/bcpt.12335
Rowles J, Olsen M (2012) Perspectives on the Development of Antioxidant Antiepileptogenic Agents. Mini-Reviews Med Chem 12:1015–1027. https://doi.org/10.2753/RES1060-939322036
Rowley S, Patel M (2013) Mitochondrial involvement and oxidative stress in temporal lobe epilepsy. Free Radic Biol Med 62:121–131. https://doi.org/10.1016/j.freeradbiomed.2013.02.002
Salami P, Lévesque M, Gotman J, Avoli M (2015) Distinct EEG seizure patterns reflect different seizure generation mechanisms. J Neurophysiol 113:2840–2844. https://doi.org/10.1152/jn.00031.2015
Salgo MG, Pryor WA (1996) Trolox inhibits Peroxynitrite-mediated oxidative stress and apoptosis in rat Thymocytes. Arch Biochem Biophys 333(2):482–488. https://doi.org/10.1006/abbi.1996.0418
Schapira AHV, Mann VM, Cooper JM, Dexter D, Daniel SE, Jenner P, Clark JB, Marsden CD (1990) Anatomic and disease specificity of NADH CoQ1 reductase (complex I) deficiency in Parkinson’s disease. J Neurochem 55:2142–2145. https://doi.org/10.1111/j.1471-4159.1990.tb05809.x
Selivanov VA, Votyakova TV, Pivtoraiko VN, Zeak J, Sukhomlin T, Trucco M, Roca J, Cascante M (2011) Reactive oxygen species production by forward and reverse electron fluxes in the mitochondrial respiratory chain. PLoS Comput Biol 7(3):1–17. https://doi.org/10.1371/journal.pcbi.1001115
Shiha AA, de la Rosa RF, Delgado M, Pozo MA, García-García L (2017) Subacute fluoxetine reduces signs of hippocampal damage induced by a single Convulsant dose of 4-Aminopyridine in rats. CNS Neurol Disord Drug Targets 16(6):694–704. https://doi.org/10.2174/1871527315666160720121723
Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175:184–191
Singh A, Kukreti R, Saso L, Kukreti S (2019) Oxidative stress: a key modulator in neurodegenerative diseases. Molecules 24(8):1–20. https://doi.org/10.3390/molecules24081583
Turrens JF (2003) Mitochondrial formation of reactive oxygen species. J Physiol 552:335–344. https://doi.org/10.1113/jphysiol.2003.049478
Van Houten B, Woshner V, Santos JH (2006) Role of mitochondrial DNA in toxic responses to oxidative stress. DNA Repair (Amst) 5:145–152. https://doi.org/10.1016/j.dnarep.2005.03.002
Venditti P, Di Stefano L, Di Meo S (2013) Mitochondrial metabolism of reactive oxygen species. Mitochondrion 13:71–82. https://doi.org/10.1016/j.mito.2013.01.008
Wahab A, Albus K, Gabriel S, Heinemann U (2010) In search of models of pharmacoresistant epilepsy. Epilepsia 51:154–159. https://doi.org/10.1111/j.1528-1167.2010.02632.x
Wang J, Xu H, Jiang H, Du X, Sun P, Xie J (2012) Neurorescue effect of rosmarinic acid on 6-hydroxydopamine-lesioned nigral domapine neurons in rat model of Parkinson´s disease. J Mol Neurosci 47:113–119. https://doi.org/10.1007/s12031-011-9693-1
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The authors were supported by the Brazilian’s agencies: National Council for Scientific and Technological Development (CNPq) (Dr. Guilhian Leipnitz and Dr. P Pereira), and Federal University of Rio Grande do Sul (UFRGS).
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JGL, LS, AMM, MSR, and DG conducted the behavioral and biochemical experiments. GRG and PFP analyzed data. GL, DJM, JGL, and PP developed the experimental design, analyzed the data, and wrote the paper. All authors read and approved the manuscript.
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Luft, J.G., Steffens, L., Morás, A.M. et al. Rosmarinic acid improves oxidative stress parameters and mitochondrial respiratory chain activity following 4-aminopyridine and picrotoxin-induced seizure in mice. Naunyn-Schmiedeberg's Arch Pharmacol 392, 1347–1358 (2019). https://doi.org/10.1007/s00210-019-01675-6
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DOI: https://doi.org/10.1007/s00210-019-01675-6