DNA damage in brain cells and behavioral deficits in mice after treatment with high doses of amantadine Vanessa Kaefer, a,b Juliane Garcia Semedo, a Vivian Francília Silva Kahl, a Rafael Gomes Von Borowsky, b Janaína Gianesini, b Tarso Benigno Ledur Kist, c Patrícia Pereira b and Jaqueline Nascimento Picada a * ABSTRACT: Amantadine (AMA) is an uncompetitive antagonist of the N-methyl-D-aspartate receptor, with clinical application, acting on treatment of influenza A virus and Parkinson’s disease. It has been proposed that AMA can indirectly modulate dopaminergic transmission. In high doses, the central nervous system is its primary site of toxicity. To examine deleterious effects on CNS induced by AMA, this study evaluated possible neurobehavioral alterations induced by AMA such as stereotyped behavior, the effects on locomotion and memory and its possible genotoxic/mutagenic activities. Adult male CF-1 mice were treated with a systemic injection of AMA (15, 30 or 60 mg kg -1 ) 20 min before behavioral tasks on open field and inhibitory avoidance. Higher AMA doses increased the latency to step-down inhibitory avoidance test in the training session in the inhibitory avoidance task. At 60 mg kg -1 AMA induced impairing effects on locomotion and exploration and hence impaired habituation to a novel environment. Stereotyped behavior after each administration in a 3-day trial was observed, suggesting effects on dopaminergic system. Amantadine was not able to induce chromosomal mutagenesis or toxicity on bone marrow, as evaluated by the micronucleus assay. At the lowest dose tested, AMA did not induce DNA damage and it was unable to impair memory, locomotion, exploration or motivation in mice. However, higher AMA doses increased DNA damage in brain tissue, produced locomotor disturbances severe enough to preclude testing for learning and memory effects, and induced stereotypy, suggesting neurotoxicity. Copyright © 2010 John Wiley & Sons, Ltd. Keywords: amantadine; behavior; comet assay; memory; micronucleus assay; toxicity INTRODUCTION Amantadine is a drug used for the treatment of influenza A virus infections (Davies et al., 1964; Dolin et al., 1982; Scholtissek and Webster, 1998; Fleming, 2001), as well as for Parkinson’s disease (Pinter et al., 1999; Nishikawa et al., 2009), dementia, multiple sclerosis, chronic pain and in cocaine withdrawal (Fisher et al., 2000; Meythaler et al., 2002). It is usually proposed that AMA is an uncompetitive antagonist drug of the N-methyl-D-aspartate (NMDA) receptor and it increases the biosynthesis and release of dopamine in the striatum (Peeters et al., 2002; Thomas et al., 2005). Aminoadamantanes, like AMA, are well-tolerated drugs and have less psychotomimetic properties when compared with other uncompetitive NMDA receptor antagonists (Camarasa et al., 2008). Parkinson’s disease patients who have not developed psy- chiatric symptoms as adverse reactions to drug treatment may develop CNS symptoms when plasmatic AMA concentrations exceed 3.0 mg ml -1 (Nishikawa et al., 2009). The steady-state plasma level in patients receiving the recommended daily dose of 200 mg is 0.2–0.9 mg ml -1 (Pacifici et al., 1976; Nishikawa et al., 2009). Amantadine has a plasma half-life of about 12 h and its time to peak concentration is about 3.3 h (Verhagen Metman et al., 2002). The central nervous system is one of the primary sites of toxic- ity from chronic AMA overdose or small intentional ingestions (Snoey and Bessen 1989; Strong et al., 1991; Macchio et al., 1993). In large, acute overdoses, however, AMA toxicity primarily results in manifested cardiovascular toxicity. Death or cardiopulmonary arrest attributed to AMA has been reported in cases of ingested doses greater than 2 g and serum level equal to 39 mg ml -1 (Sartori et al., 1984; Pimentel and Hughes, 1991; Hartshorne et al., 1995; Schwartz et al., 2008). *Correspondence to: J. N. Picada, Laboratório de Genética Toxicológica, Programa de Pós-Graduação em Genética e Toxicologia Aplicada, Universidade Luterana do Brasil, ULBRA, Av. Farroupilha 8001, Bairro São José, Canoas, RS, CEP: 92425-900, Brazil. E-mail: jnpicada@cpovo.net a Laboratório de Genética Toxicológica, Programa de Pós-Graduação em Genética e Toxicologia Aplicada, Universidade Luterana do Brasil, ULBRA, Av. Farroupilha 8001, Bairro São José, Canoas, RS, CEP: 92425-900, Brazil b Laboratório de Farmacologia e Toxicologia, Programa de Pós-Graduação em Genética e Toxicologia Aplicada, ULBRA, Canoas, RS, Brazil c Laboratório de Métodos, Departamento de Biofísica, Instituto de Biociências, Uni- versidade Federal do Rio Grande do Sul,UFRGS, Porto Alegre, RS, Brazil Research Article Received: 10 February 2010, Revised: 26 April 2010, Accepted: 29 April 2010 Published online in Wiley Online Library: 23 June 2010 (wileyonlinelibrary.com) DOI 10.1002/jat.1550 745 J. Appl. Toxicol. 2010; 30: 745–753 Copyright © 2010 John Wiley & Sons, Ltd.