Abstract
A growing body of evidence indicates that creatine (Cr) exerts beneficial effects on a variety of pathologies where energy metabolism and oxidative stress play an etiological role. However, the benefits of Cr treatment for epileptics are still shrouded in controversy. In the present study, we found that acute Cr treatment (300 mg/kg, p.o.) prevented the increase in electroencephalographic wave amplitude typically elicited by PTZ (30, 45 or 60 mg/kg, i.p.). Cr treatment also increased the latency periods of first myoclonic jerks, lengthened the latency periods of the generalized tonic–clonic seizures and reduced the time spent in the generalized tonic–clonic seizures induced by PTZ (60 mg/kg). Administration of PTZ (all doses) decreased Na+, K+-ATPase activity as well as adenosine triphosphate (ATP) and adenosine diphosphate levels in the cerebral cortex, but Cr treatment prevented these effects. Cr administration also prevented increases in xanthine oxidase activity, adenosine monophosphate levels, adenosine levels, inosine levels and uric acid levels that normally occur after PTZ treatment (60 mg/kg, i.p.). We also showed that Cr treatment increased the total Cr (Cr + PCr) content, creatine kinase activity and the mitochondrial membrane potential (ΔΨ) in the cerebral cortex. In addition, Cr prevented PTZ-induced mitochondrial dysfunction characterized by decreasing ΔΨ, increasing thiobarbituric acid-reactive substance levels and increasing protein carbonylation. These experimental findings reinforce the idea that mitochondrial dysfunction plays a critical role in models of epileptic seizures and suggest that buffering brain energy levels through Cr treatment may be a promising therapeutic approach for the treatment of this neurological disease.
Similar content being viewed by others
References
Adhihetty PJ, Beal MF (2008) Creatine and its potential therapeutic value for targeting cellular energy impairment in neurodegenerative diseases. Neuromol Med 10:275–290
Adhihetty PJ, Hood DA (2003) Mechanisms of apoptosis in skeletal muscle. Basic Appl myol 13:171–179
Akerman KE, Wikstrom MK (1976) Safranine as a probe of the mitochondrial membrane potential. FEBS Lett 68:191–197
Andres RH, Ducray AD, Schlattner U, Wallimann T, Widmer HR (2008) Functions and effects of creatine in the central nervous system. Brain Res Bull 76:329–343
Beal MF (2011) Neuroprotective effects of creatine. Amino Acids 40:1305–1313
Bindokas VP, Lee CC, Colmers WF, Miller RJ (1998) Changes in mitochondrial function resulting from synaptic activity in the rat hippocampal slice. J Neurosci 18:4570–4587
Bonan CD, Amaral OB, Rockenbach IC, Walz R, Battastini AM, Izquierdo I, Sarkis JJ (2000) Altered ATP hydrolysis induced by pentylenetetrazol kindling in rat brain synaptosomes. Neurochem Res 25:775–779
Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
Brini M, Pinton P, King MP, Davidson M, Schon EA, Rizzuto R (1999) A calcium signaling defect in the pathogenesis of a mitochondrial DNA inherited oxidative phosphorylation deficiency. Nat Med 5:951–954
Bruce AJ, Baudry M (1995) Oxygen free radicals in rat limbic structures after kainate-induced seizures. Free Radic Biol Med 18:993–1002
Bruno AN, Oses JP, Amaral O, Coitinho A, Bonan CD, Battastini AM, Sarkis JJ (2003) Changes in nucleotide hydrolysis in rat blood serum induced by pentylenetetrazol-kindling. Brain Res Mol Brain Res 114:140–145
Canafoglia L, Franceschetti S, Antozzi C, Carrara F, Farina L, Granata T, Lamantea E, Savoiardo M, Uziel G, Villani F, Zeviani M, Avanzini G (2001) Epileptic phenotypes associated with mitochondrial disorders. Neurology 56:1340–1346
Carmody S, Brennan L (2010) Effects of pentylenetetrazole-induced seizures on metabolomic profiles of rat brain. Neurochem Int 56:340–344
Clapcote SJ, Duffy S, Xie G, Kirshenbaum G, Bechard AR, Rodacker Schack V, Petersen J, Sinai L, Saab BJ, Lerch JP, Minassian BA, Ackerley CA, Sled JG, Cortez MA, Henderson JT, Vilsen B, Roder JC (2009) Mutation I810N in the alpha3 isoform of Na+, K+-ATPase causes impairments in the sodium pump and hyperexcitability in the CNS. Proc Natl Acad Sci USA 106:14085–14090
Clausen T, Van Hardeveld C, Everts ME (1991) Significance of cation transport in control of energy metabolism and thermogenesis. Physiol Rev 71:733–774
Fiske CH, Subbarow Y (1925) The colorimetric determination of phosphorus. J Biol Chem 66:375–400
Folbergrova J, Kunz WS (2011) Mitochondrial dysfunction in epilepsy. Mitochondrion 12:35–40
Fox PT, Raichle ME, Mintun MA, Dence C (1988) Nonoxidative glucose consumption during focal physiologic neural activity. Science 241:462–464
Frantseva MV, Velazquez JL, Hwang PA, Carlen PL (2000) Free radical production correlates with cell death in an in vitro model of epilepsy. Eur J Neurosci 12:1431–1439
Fujisawa H, Kajikawa K, Ohi Y, Hashimot Y, Yoshida H (1965) Movement of radioactive calcium in brain slices and influences on it of protoveratrine ouabain potassium chloride and cocaine. Jpn J Pharmacol 15:327–334
Gallanti A, Tonelli A, Cardin V, Bussone G, Bresolin N, Bassi MT (2008) A novel de novo nonsense mutation in ATP1A2 associated with sporadic hemiplegic migraine and epileptic seizures. J Neurol Sci 273:123–126
Gualano B, Artioli GG, Poortmans JR, Lancha Junior AH (2009) Exploring the therapeutic role of creatine supplementation. Amino Acids 38:31–44
Gualano B, Roschel H, Lancha-Jr AH, Brightbill CE, Rawson ES (2011) In sickness and in health: the widespread application of creatine supplementation. Amino Acids 43:519–529
Gupta YK, Veerendra Kumar MH, Srivastava AK (2003) Effect of Centella asiatica on pentylenetetrazole-induced kindling, cognition and oxidative stress in rats. Pharmacol Biochem Behav 74:579–585
Holtzman D, Togliatti A, Khait I, Jensen F (1998) Creatine increases survival and suppresses seizures in the hypoxic immature rat. Pediatr Res 44:410–414
Ipsiroglu OS, Stromberger C, Ilas J, Hoger H, Muhl A, Stockler-Ipsiroglu S (2001) Changes of tissue creatine concentrations upon oral supplementation of creatine-monohydrate in various animal species. Life Sci 69:1805–1815
Jamme I, Petit E, Divoux D, Gerbi A, Maixent JM, Nouvelot A (1995) Modulation of mouse cerebral Na+, K(+)-ATPase activity by oxygen free radicals. Neuroreport 7:333–337
Jost CR, Van Der Zee CE, In ‘t Zandt HJ, Oerlemans F, Verheij M, Streijger F, Fransen J, Heerschap A, Cools AR, Wieringa B (2002) Creatine kinase B-driven energy transfer in the brain is important for habituation and spatial learning behaviour, mossy fibre field size and determination of seizure susceptibility. Eur J Neurosci 15:1692–1706
Kanemitsu H, Tamura A, Kirino T, Oka H, Sano K, Iwamoto T, Yoshiura M, Iriyama K (1989) Allopurinol inhibits uric acid accumulation in the rat brain following focal cerebral ischemia. Brain Res 499:367–370
Kilbride SM, Telford JE, Tipton KF, Davey GP (2008) Partial inhibition of complex I activity increases Ca-independent glutamate release rates from depolarized synaptosomes. J Neurochem 106:826–834
Klein AM, Ferrante RJ (2007) The neuroprotective role of creatine. Subcell Biochem 46:205–243
Karatzaferi C, De Haan A, Offringa C, Sargeant AJ (1999) Improved high-performance liquid chromatographic assay for the determination of "high-energy" phosphates in mammalian skeletal muscle. Application to a single-fibre study in man. J Chromatogr B Biomed Sci Appl 730:183–191
Klopstock T, Elstner M, Bender A (2011) Creatine in mouse models of neurodegeneration and aging. Amino Acids 40(5):1297–1303
Kraemer WJ, Volek JS (1999) Creatine supplementation. Its role in human performance. Clin Sports Med 18:651–666 ix
Lees GJ, Lehmann A, Sandberg M, Hamberger A (1990) The neurotoxicity of ouabain, a sodium-potassium ATPase inhibitor, in the rat hippocampus. Neurosci Lett 120:159–162
Li S, Stys PK (2001) Na+-K+-ATPase inhibition and depolarization induce glutamate release via reverse Na+-dependent transport in spinal cord white matter. Neuroscience 107:675–683
Magni DV, Oliveira MS, Furian AF, Fiorenza NG, Fighera MR, Ferreira J, Mello CF, Royes LF (2007) Creatine decreases convulsions and neurochemical alterations induced by glutaric acid in rats. Brain Res 1185:336–345
Masino SA, Geiger JD (2008) Are purines mediators of the anticonvulsant/neuroprotective effects of ketogenic diets? Trends Neurosci 31:273–278
McColl CD, Horne MK, Finkelstein DI, Wong JY, Berkovic SF, Drago J (2003) Electroencephalographic characterisation of pentylenetetrazole-induced seizures in mice lacking the alpha 4 subunit of the neuronal nicotinic receptor. Neuropharmacology 44:234–243
Mikati MA, Kurdit RM, Rahmeh AA, Farhat F, Abu Rialy S, Lteif L, Francis E, Geha G, Maraashli W (2004) Effects of creatine and cyclocreatine supplementation on kainate induced injury in pre-pubescent rats. Brain Inj 18:1229–1241
Mills PC, Smith NC, Harris RC, Harris P (1997) Effect of allopurinol on the formation of reactive oxygen species during intense exercise in the horse. Res Vet Sci 62:11–16
Oliveira MS, Furian AF, Rambo LM, Ribeiro LR, Royes LF, Ferreira J, Calixto JB, Otalora LF, Garrido-Sanabria ER, Mello CF (2009) Prostaglandin E2 modulates Na+, K+-ATPase activity in rat hippocampus: implications for neurological diseases. J Neurochem 109:416–426
Oses JP, Viola GG, de Paula Cognato G, Junior VH, Hansel G, Bohmer AE, Leke R, Bruno AN, Bonan CD, Bogo MR, Portela LV, Souza DO, Sarkis JJ (2007) Pentylenetetrazol kindling alters adenine and guanine nucleotide catabolism in rat hippocampal slices and cerebrospinal fluid. Epilepsy Res 75:104–111
Özogul F, Taylor AKD, Quantick P, Özogul Y (2000) A rapid HPLC-determination of ATP-related compounds and its applications to herring stored under modified atmosphere. Int J Food Sci Technol 35:549–554
Pan JW, Kim JH, Cohen-Gadol A, Pan C, Spencer DD, Hetherington HP (2005) Regional energetic dysfunction in hippocampal epilepsy. Acta Neurol Scand 111:218–224
Patel M (2004) Mitochondrial dysfunction and oxidative stress: cause and consequence of epileptic seizures. Free Radic Biol Med 37:1951–1962
Patel M, Li QY (2003) Age dependence of seizure-induced oxidative stress. Neuroscience 118:431–437
Patsoukis N, Zervoudakis G, Georgiou CD, Angelatou F, Matsokis NA, Panagopoulos NT (2005) Thiol redox state and lipid and protein oxidation in the mouse striatum after pentylenetetrazol-induced epileptic seizure. Epilepsia 46:1205–1211
Paxinos G, Watson C (1986) The rat brain in stereotaxic coordinates. Academic Press, San Diego
Poderoso JJ, Carreras MC, Lisdero C, Riobo N, Schopfer F, Boveris A (1996) Nitric oxide inhibits electron transfer and increases superoxide radical production in rat heart mitochondria and submitochondrial particles. Arch Biochem Biophys 328:85–92
Prajda N, Weber G (1975) Malignant transformation-linked imbalance: decreased xanthine oxidase activity in hepatomas. FEBS Lett 59:245–249
Rambo LM, Ribeiro LR, Oliveira MS, Furian AF, Lima FD, Souza MA, Silva LF, Retamoso LT, Corte CL, Puntel GO, de Avila DS, Soares FA, Fighera MR, Mello CF, Royes LF (2009) Additive anticonvulsant effects of creatine supplementation and physical exercise against pentylenetetrazol-induced seizures. Neurochem Int 55:333–340
Ribeiro MC, de Avila DS, Schneider CY, Hermes FS, Furian AF, Oliveira MS, Rubin MA, Lehmann M, Krieglstein J, Mello CF (2005) alpha-Tocopherol protects against pentylenetetrazol- and methylmalonate-induced convulsions. Epilepsy Res 66:185–194
Royes LF, Fighera MR, Furian AF, Oliveira MS, da Silva LG, Malfatti CR, Schneider PH, Braga AL, Wajner M, Mello CF (2003) Creatine protects against the convulsive behavior and lactate production elicited by the intrastriatal injection of methylmalonate. Neuroscience 118:1079–1090
Royes LF, Fighera MR, Furian AF, Oliveira MS, Myskiw Jde C, Fiorenza NG, Petry JC, Coelho RC, Mello CF (2006) Effectiveness of creatine monohydrate on seizures and oxidative damage induced by methylmalonate. Pharmacol Biochem Behav 83:136–144
Schneider Oliveira M, Flavia Furian A, Freire Royes LF, Rechia Fighera M, de Carvalho Myskiw J, Gindri Fiorenza N, Mello CF (2004) Ascorbate modulates pentylenetetrazol-induced convulsions biphasically. Neuroscience 128:721–728
Shin EJ, Jeong JH, Chung YH, Kim WK, Ko KH, Bach JH, Hong JS, Yoneda Y, Kim HC (2011) Role of oxidative stress in epileptic seizures. Neurochem Int 59:122–137
Souza MA, Oliveira MS, Furian AF, Rambo LM, Ribeiro LR, Lima FD, Dalla Corte LC, Silva LF, Retamoso LT, Dalla Corte CL, Puntel GO, de Avila DS, Soares FA, Fighera MR, de Mello CF, Royes LF (2009) Swimming training prevents pentylenetetrazol-induced inhibition of Na+, K+-ATPase activity, seizures, and oxidative stress. Epilepsia 50:811–823
Streijger F, Scheenen WJ, van Luijtelaar G, Oerlemans F, Wieringa B, Van der Zee CE (2010) Complete brain-type creatine kinase deficiency in mice blocks seizure activity and affects intracellular calcium kinetics. Epilepsia 51:79–88
Sullivan PG, Dube C, Dorenbos K, Steward O, Baram TZ (2003) Mitochondrial uncoupling protein-2 protects the immature brain from excitotoxic neuronal death. Ann Neurol 53:711–717
Tada H, Morooka K, Arimoto K, Matsuo T (1991) Clinical effects of allopurinol on intractable epilepsy. Epilepsia 32:279–283
Tonkonogi M, Sahlin K (1997) Rate of oxidative phosphorylation in isolated mitochondria from human skeletal muscle: effect of training status. Acta Physiol Scand 161:345–353
Vielhaber S, Von Oertzen JH, Kudin AF, Schoenfeld A, Menzel C, Biersack HJ, Kral T, Elger CE, Kunz WS (2003) Correlation of hippocampal glucose oxidation capacity and interictal FDG-PET in temporal lobe epilepsy. Epilepsia 44:193–199
Waldbaum S, Patel M (2010) Mitochondrial dysfunction and oxidative stress: a contributing link to acquired epilepsy? J Bioenergy Biomembr 42:449–455
Wyss M, Kaddurah-Daouk R (2000) Creatine and creatinine metabolism. Physiol Rev 80:1107–1213
Zagnoni PG, Bianchi A, Zolo P, Canger R, Cornaggia C, D’Alessandro P, DeMarco P, Pisani F, Gianelli M, Verze L et al (1994) Allopurinol as add-on therapy in refractory epilepsy: a double-blind placebo-controlled randomized study. Epilepsia 35:107–112
Acknowledgments
The authors thank Dr. Guilherme Bresciani for critical reading of the manuscript. This work was supported by FAPERGS/CNPq (Grant: #11/2082-4). L.F.F. Royes, M.R. Fighera and L.M. Rambo are the recipients of CNPq fellowships (Grant: #141164/2010-7). I. Della-Pace is the recipient of CAPES fellowships. We confirm that we have read the journal’s position on issues involved in ethical publication and affirm that this report is consistent with those guidelines. In addition, we would like to state that all authors have observed and approved the study and that no part of the submitted work has been published or is under consideration for publication elsewhere. Moreover, the present work was supported by government funding and has no financial or other relationships that might lead to a conflict of interest. We also would like to declare that all experiments were carried out according to the National Institute of Health Guide for the Care and Use of Laboratory Animals (NIH Publications No. 80-23) revised 1996 and that the University Ethics Committee approved the respective protocols.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Rambo, L.M., Ribeiro, L.R., Della-Pace, I.D. et al. Acute creatine administration improves mitochondrial membrane potential and protects against pentylenetetrazol-induced seizures. Amino Acids 44, 857–868 (2013). https://doi.org/10.1007/s00726-012-1408-6
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00726-012-1408-6