1 2 RESEARCH PAPER 3 Cerebral Malaria Causes Enduring Behavioral and Molecular Changes 4 in Mice Brain Without Causing Gross Histopathological Damage 5 Thaı´ze Lopes de Souza, a Ana Claudia Beck Grauncke, a Leandro Rodrigo Ribeiro, a * Fernanda Kulinski Mello, a 6 Sara Marchesan Oliveira, b Fa´tima Brant, c Fabiana S. Machado c and Mauro Schneider Oliveira a 7 a Graduate Program in Pharmacology and Department of Physiology and Pharmacology, Federal University of Santa Maria, Santa Maria, 8 Rio Grande do Sul, Brazil 9 b Graduate Program in Biological Sciences: Toxicological Biochemistry and Department of Biochemistry and Molecular Biology, Federal 10 University of Santa Maria, Santa Maria, Rio Grande do Sul, Brazil 11 c Graduate Program in Health Sciences: Infectious Diseases and Tropical Medicine and Department of Biochemistry and Immunology, Institute 12 of Biological Science, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil 13 14 Abstract—Malaria, parasitic disease considered a major health public problem, is caused by Plasmodium proto- zoan genus and transmitted by the bite of infected female Anopheles mosquito genus. Cerebral malaria (CM) is the most severe presentation of malaria, caused by P. falciparum and responsible for high mortality and enduring development of cognitive deficits which may persist even after cure and cessation of therapy. In the present study we evaluated selected behavioral, neurochemical and neuropathologic parameters after rescue from experimental cerebral malaria caused by P. berghei ANKA in C57BL/6 mice. Behavioral tests showed impaired nest building activity as well as increased marble burying, indicating that natural behavior of mice remains altered even after cure of infection. Regarding the neurochemical data, we found decreased a2/a3 Na + ,K + -ATPase activity and increased immunoreactivity of phosphorylated Na + ,K + -ATPase at Ser 943 in cerebral cortex after CM. In addition, [ 3 H]-Flunitrazepam binding assays revealed a decrease of benzodiazepine/GABA A receptor binding sites in infected animals. Moreover, in hippocampus, dot blot analysis revealed increased levels of protein carbonyls, suggesting occurrence of oxidative damage to proteins. Interestingly, no changes in the neuropathological mark- ers Fluoro-Jade C, Timm staining or IBA-1 were detected. Altogether, present data indicate that behavioral and neurochemical alterations persist even after parasitemia clearance and CM recovery, which agrees with available clinical findings. Some of the molecular mechanisms reported in the present study may underlie the behavioral changes and increased seizure susceptibility that persist after recovery from CM and may help in the future devel- opment of therapeutic strategies for CM sequelae. Ó 2017 Published by Elsevier Ltd on behalf of IBRO. Key words: Plasmodium berghei ANKA, chloroquine, cognitive deficits, GABA A receptor, Na + ,K + -ATPase, oxidative stress. 15 INTRODUCTION 16 Malaria is a serious tropical disease caused by different 17 species of protozoan Plasmodium (Phillips et al., 2017). 18 The World Health Organization (WHO) estimates that 19 212 million cases and 429,000 malaria-related deaths 20 were reported in 2015, affecting mainly people in the 21 sub-Saharan Africa region (WHO, 2016). Cerebral 22 malaria (CM) is a neurological complication which is asso- 23 ciated with high rates of morbidity and mortality, even 24 after recovery, and it has been considered the most sev- 25 ere clinical presentation of infection with P. falciparum 26 (Hunt et al., 2006; Opoka et al., 2009). In fact, high rates 27 of mortality are observed in patients diagnosed with CM, 28 and about one quarter of survivors presents long-term 29 neurological complications (John et al., 2008; Idro et al., 30 2010a, 2010b). 31 Experimental models are useful to investigate 32 mechanisms involved in disease and possible 33 development of sequelae, since clinical studies present 34 limitations including but not limited to ethical issues and 35 the need of invasive techniques (Lacerda-Queiroz et al., 36 2010). P. berghei ANKA (PbA) strain is considered a 37 well-characterized experimental model of CM (ECM) 38 (Hunt et al., 2010). In this context, results obtained with https://doi.org/10.1016/j.neuroscience.2017.10.043 0306-4522/Ó 2017 Published by Elsevier Ltd on behalf of IBRO. * Corresponding author. Address: Department of Physiology and Pharmacology, Federal University of Santa Maria, Av. Roraima, n° 1000, CEP 97105-900, Santa Maria, Rio Grande do Sul, Brazil. E-mail address: leandrorodrigoribeiro@yahoo.com.br (L. Rodrigo Ribeiro). NEUROSCIENCE RESEARCH ARTICLE T. Lopes de Souza et al. / Neuroscience xxx (2017) xxx–xxx Please cite this article in press as: Lopes de Souza T et al. Cerebral Malaria Causes Enduring Behavioral and Molecular Changes in Mice Brain Without Causing Gross Histopathological Damage. Neuroscience (2017), https://doi.org/10.1016/j.neuroscience.2017.10.043 1 NSC 18108 No. of Pages 10 11 November 2017