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Conselho Nacional de Desenvolvimento Científico e Tecnológico / The pathogenesis of Trypanosoma evansi (T. evansi) infection on central and peripheral nervous system (CNS) and immunologic system it is not completely understood. This study was conducted to investigate the effects of infection by T. evansi on the behavioral performance of rats, relating them to neurochemical alterations of the glutamatergic system and activity of important enzymes such as Ca2 + ATPase, Na + / K + ATPase and acetylcholinesterase (AChE) in the brain. Furthermore, we evaluated the activity of AChE in the T. evansi, using a biochemical protocol and discussed some possibilities for the interaction between the presence of this enzyme in the parasite and its possible relation with the host. Also, we investigated the in vivo effect of curcumin as an antitrypanosomal agent to contribute in the search for new therapies. To this end, we conducted a series of studies, as follow that are presented as scientific articles: in the Article I are presented data from rats experimentally infected with T. evansi and in which the activity of AChE in total brain, cerebellum and blood and plasma butyrylcholinesterase activity on days 3 (T3) and 5 (T5) post infection (pi) were evaluated. In the Article II trypanosomes were isolated by Dietil Amino Etil-celulose column to perform enzymatic assays of AChE activity in the parasite. Article III presents the results of daily oral administration of Curcumin (20 or 60 mg / kg) as a preventive treatment (for 30 or 15 days pre-infection) or as curative treatment (for 15 days pi) for T. evansi infection in rats. The animals were evaluated for parasitemia, proinflammatory cytokines (IFN-γ, TNF-α, IL-1, IL-6) and anti-inflammatory (IL-10) in plasma. Also blood AChE activity was studied. In the manuscript I were evaluated behavioral changes (memory and anxiety), glutamate uptake, activity of AChE and ATPases (Ca +2 and Na +, K +) in brain structures of T. evansi infected rats (5 and 30 dpi). Together these studies suggest that rats infected with T. evansi have cognitive impairment, probably caused by changes on energy metabolism (ATPases) and in the cholinergic and glutamatergic systems. That way, alterations observed during behavioral tests probably indicate the progression of clinical disease as a result of neurochemical dysfunction. It was also demonstrated that it is possible to biochemically detect the AChE activity in the T. evansi and this could cause the hydrolysis of acetylcholine in the parasite. It was also demonstrated that pretreatment with curcumin induces immunomodulatory effects, altering inflammatory cytokines and activity of AChE in rats infected with T. evansi. The treatments reduced parasitic load and mortality. Therefore, curcumin should be considered as a feeding supplement in trypanosomiasis endemic areas. / Com este trabalho, buscou-se compreender os mecanismos da patogenia do Trypanosoma evansi no sistema nervoso central (SNC), periférico e imunológico. Para isso, foram avaliados os efeitos da infecção por T. evansi sobre o desempenho comportamental de ratos, relacionando-os com alterações neuroquímicas do sistema glutamatérgico e com as atividades de importantes enzimas cerebrais como a Ca+2ATPase, a Na+/K+ATPase e a acetilcolinesterase (AChE). Avaliou-se também, a atividade da AChE no T. evansi usando um protocolo de bioquímica, buscando discutir algumas possibilidades de interação entre a presença desta enzima no parasita e suas possíveis relações com o hospedeiro. Também, foi investigado o efeito in vivo da curcumina como agente tripanocida, a fim de contribuir na busca de novas terapias. Com estes propósitos, foram realizadas varios estudos, sendo: no artigo I buscou-se avaliar se a infecção pelo protozoário altera o sistema colinérgico. Para isso, foram utilizados ratos experimentalmente infectados com T. evansi e avaliada a atividade da AChE no cérebro total, no cerebelo e no sangue e a atividade da butirilcolinesterase plasmática nos dias 3 e 5 pós infecção (pi); no artigo II tripanossomas foram isolados por coluna de dietilaminoetilcelulose para realizar ensaios enzimáticos da atividade da AChE no parasito; no artigo III doses diárias de Curcumina (20 ou 60 mg / kg) diluída em óleo de milho foram administradas diariamente por via oral (gavagem) como tratamento preventivo (30 ou 15 dias pré-infecção) e como tratamento durante quinze dias pi. Os animais foram avaliados quanto a parasitemia, as citocinas pró-inflamatórias (IFN-γ, TNF-α, IL-1, IL-6) e anti-inflamatória (IL-10) plasmáticas e a atividade sanguínea da AChE; no manuscrito I foram avaliadas as alterações comportamentais (memória e ansiedade), a captação de glutamato, atividade da AChE e das ATPases (Ca +2 e Na +, K +) nas estruturas encefálicas nos dias 5 e 30 de ratos infectados com T. evansi. Com estes estudos, pode-se concluir que ratos infectados com T. evansi apresentam déficit cognitivo, provavelmente causados por alterações nos parâmetros do metabolismo energético (ATPases) e nos sistemas colinérgico e glutamatérgico, sugerindo que as alterações observadas durante testes comportamentais provavelmente indicam uma disfunção neuroquímica. Também foi demonstrado que é possível detectar bioquimicamente a atividade da AChE no T. evansi e esta deve ser responsável pela hidrólise da acetilcolina no parasita. Quando os animais infectados foram tratados com curcumina demonstrou-se que o pré-tratamento com este composto induz efeitos imunomodulatórios, alterando os parâmetros inflamatórios tais como as citocinas e a atividade da AChE sanguínea. Desta forma, sugere-se que a curcumina pode ser utilizada como um suplemento alimentar em áreas endêmicas de tripanossomíase, uma vez que reduziu a carga parasitária e a mortalidade em ratos infectados com T. evansi, contribuindo assim com novas alternativas de tratamento.

Les hélicases sont des enzymes extrêmement répandues qui sont essentielles pour mener à bien l'ensemble des processus cellulaires impliquant des acides nucléiques. Ces protéines parviennent à annihiler les structures secondaires et tertiaires qui peuvent se former dans l'ADN ou l'ARN grâce à l'énergie puisée par l'hydrolyse de l'ATP. Les hélicases ne sont toutefois pas l'exclusivité des organismes eucaryotes et procaryotes, mais bon nombre de virus codent aussi pour des enzymes possédant cette fonction. La famille des Flaviviridae ne fait pas exception à ce groupe puisque l'ensemble des virus de cette famille code pour une hélicase nommée NS3. La famille des Flaviviridae est une famille virale d'une importance monumentale étant donné la présence de nombreux pathogènes humains parmi ses rangs. On y dénote le virus de l'hépatite C, le virus de la dengue et le virus du Nil occidental pour ne nommer que ceux-là. Ces virus ont aussi un autre point en commun, le manque criant d'une thérapie efficace pour traiter les patients atteints. La protéine NS3 et plus particulièrement son activité hélicase est une cible potentielle envisageable pour le traitement de ces virus étant donné l'aspect essentiel de cette activité dans le cycle de réplication de ceux-ci. L'étude présentée dans ce mémoire a pour dessein d'étudier le site d'hydrolyse de l'ATP de la protéine NS3 du virus du Nil occidental. Le but final est de découvrir les déterminants moléculaires qui mènent à la sélection du substrat à hydrolyser, tant au niveau du substrat qu'au niveau de la protéine. Du côté du substrat, une panoplie d'analogues de purines fut testée pour déterminer la capacité de ces molécules à compétitionner pour le site actif de la protéine ainsi que pour la faculté de la protéine à hydrolyser ces substrats. Du côté de la protéine, une gamme de mutants de NS3 fut exprimée et l'habileté de ces mutants à discriminer entre les différents substrats qui lui étaient présentés a été mesurée. Les acides aminés qui ont été sélectionnés pour être mutés l'ont été soit pour leur ressemblance à un motif de reconnaissance de l'ATP chez d'autres hélicases, soit pour leur proximité du substrat, déterminée grâce à un modèle de la protéine bâti sur la structure cristallisée de la protéine NS3 du virus de la dengue. L'ensemble des résultats obtenus a permis de dresser le portrait des atomes de l'ATP qui sont liés par la protéine en plus de mettre à jour plusieurs acides aminés qui sont impliqués directement ou indirectement dans la sélection du substrat par NS3. Il semble en effet que la présence d'un groupement donneur de liaison hydrogène soit nécessaire à la position six d'une purine pour permettre un bon niveau d'hydrolyse de la molécule, permettant ainsi à la protéine de faire la distinction entre l'ATP et le GTP. Aussi, l'apport insoupçonné dans le processus d'hydrolyse d'acides aminés tels que Arg202, Ans417 ou encore Arg185 a été démontré. Finalement, différentes expériences permettant d'élargir les connaissances sur la protéine NS3 ainsi que sur les hélicases en général sont proposées. Des recherches visant à déterminer les ressemblances et les différences au niveau de la discrimination entre sources d'énergie à utiliser entre les diverses protéines NS3 des Flaviviridae vont permettre d'amener à un autre niveau la compréhension du mécanisme d'hydrolyse de cette protéine. Des études s'appliquant à modifier différents acides aminés afin de façonner la protéine NS3 pour qu'elle hydrolyse des analogues particuliers ou qu'elle lie des acides nucléiques précis pourraient aussi permettre de mieux comprendre les bases de la reconnaissance des substrats de NS3 en plus de fournir éventuellement des outils moléculaires voués à des fonctions multiples.

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Les ATPases F1F0 sont présentes chez la majorité des bactéries, notamment les mycoplasmes qui sont caractérisés par un génome réduit et un mode de vie parasitaire. En plus de l’opéron codant l’ATPase F1F0, des clusters apparentés de sept gènes ont été identifiés dans le génome de nombreux mycoplasmes. Au cours de cette thèse, nous avons cherché à caractériser l’évolution et la fonction de ces clusters supplémentaires. Quatre des protéines codées par ces clusters présentent des similarités structurales avec les sous-unités α, β,  et ε de l’ATPase F1F0, résultant en une potentielle structure F1-like. Les trois autres protéines ne présentent aucune similarité avec des protéines connues. Une localisation transmembranaire est prédite pour deux d’entre elles. Deux types d’ATPase F1-like, Type 2 et Type 3, ont été identifiés. Les clusters de Type 2 et de Type 3 pourraient être originaires du groupe phylogénétique Hominis, les clusters de Type 3 ayant vraisemblablement été disséminés par des transferts horizontaux de gènes entre mycoplasmes colonisant le même hôte. Les gènes du cluster de Type 3 de Mycoplasma mycoides subsp. mycoides sont organisés en opéron et exprimés en milieu axénique. Des études de mutagénèse et de complémentation démontrent que le cluster de Type 3 est associé à une activité ATPase majeure des fractions membranaires. Des analyses biochimiques suggèrent que l’activité ATPase du cluster est sensible au ∆pH mais pas au ∆Ψ. Ces analyses suggèrent que le sodium et le potassium ne sont pas impliqués dans le fonctionnement de l’ATPase F1-likeX0. Les sous-unités des ATPases F1-likeX0 et F1F0 présentent un comportement différent en présence de détergents. L’ensemble de ces expériences suggèrent que l’ATPase F1-likeX0 est un complexe plus fragile que l’ATPase F1F0. Nos résultats montrent qu’en dépit d’une tendance à la réduction de génome, les mycoplasmes ont développé et échangé des ATPases sans équivalent chez d’autres bactéries. Nous proposons un modèle dans lequel une structure F1-like est associée avec un domaine hypothétique X0, enchâssé dans la membrane des mycoplasmes. / F1F0 ATPases have been found in most bacteria, including mycoplasmas that are characterized by drastically reduced genomes and a parasitic lifestyle. In addition to the typical operon of eight genes encoding genuine F1F0 ATPase, related clusters of seven genes were identified in many mycoplasmas. In this work, we investigated the evolution and the function of these supplementary clusters. Four proteins encoded by these clusters present structural similarities with subunits α, β,  and ε of F1F0 ATPases, resulting in potential F1-like structures. The three other encoded proteins did not show any similarity to known proteins. Transmembrane helices were predicted for two of them, suggesting a membrane localisation. Two types of F1-like ATPases, Type 2 and Type 3, were identified. Clusters encoding Type 2 and Type 3 ATPases were assumed to originate from the Hominis group of mycoplasmas. Further spreading of Type 3 ATPases towards other phylogenetic groups by horizontal gene transfers in between mycoplasmas sharing a same host was proposed on the basis of phylogenetic trees and genomic context. Functional analyses indicated that genes of Type 3 cluster in the ruminant pathogen Mycoplasma mycoides subsp. mycoides were organized as an operon. Proteomic analyses indicated that the seven encoded proteins were produced during growth in axenic media. Mutagenesis and complementation assays demonstrated that Type 3 cluster was associated with a major ATPase activity of membrane fractions. Biochemical analyses indicated that this ATPase activity was sensitive to ΔpH but not to ΔΨ. These analyses suggested that Na+ and K+ were not involved in the F1-likeX0 functioning. Our results indicated a behaviour of F1-likeX0 ATPase subunits that is different to that of F1F0 ATPase subunits in presence of detergents. Altogether, these analyses suggest that the F1-likeX0 complex could be more fragile than the F1F0 complex. Our results showed that despite their tendency to genome reduction, mycoplasmas have evolved and exchanged specific F1-like ATPases with no known equivalent in other bacteria. We propose a model in which the F1-like structure is associated with a hypothetical X0 sector embedded in the membrane of mycoplasmal cells.

Doctor of Philosophy / Entomology / Yoonseong Park / Tick salivary secretion is crucial for survival and for successful feeding. Tick saliva includes excretory water/ions and bioactive components for compromising the hosts' immune responses, and provides a direct route for pathogen transmission. Control of the tick salivation involves autocrine/paracrine dopamine, the most potent stimulator of tick salivation. Our research group reported the presence of two dopamine receptors in the salivary glands of the blacklegged tick (Ixodes scapularis): dopamine receptor (D1) and invertebrate specific D1-like dopamine receptor (InvD1L). Dopamine-induced salivary secretion was orchestrated by two distinct physiological roles via activation of the two dopamine receptors (Chapter 2). Low concentration of dopamine activated D1 receptor on epithelial cells of salivary gland acini leading inward fluid transport. High concentration of dopamine activated InvD1L receptors on axonal projections innervating myoepithelial cells modulating pumping/gating actions for emptying luminal saliva into the main duct. Thus, ticks coordinated salivary secretion with duo dopamine receptors. Dopamine-mediated saliva production involves an important downstream component, Na/K-ATPase (Chapter 3). Na/K-ATPase was found in the epithelial cells of all types of acini. However, Na/K-ATPase had two different functions in salivary secretion in different acini: 1) dopamine-mediated production of primary saliva in distally located salivary gland acini type-2/- 3, and 2) dopamine-independent resorption in proximally located salivary gland acini type-1. Type-1 acini were also found to function in direct water absorption of off-host ticks, which could be a potential route for delivery of acaricides. Chapter 4 investigated the comparative transcriptomics of the lone star tick underlying the processes of pathogen acquisition. Differential expression analyses in pathogen-exposed ticks revealed a number of transcripts that are important in the tick-pathogen interaction. These included genes for tick immunity against pathogen and for modulation of tick physiology facilitating a pathogen’s invasion and proliferation. My study expanded the understanding of physiological mechanisms controlling tick salivation. In addition, transcriptomics of ticks in interaction with pathogen identified several genes that are relevant in vector/pathogen interactions. The knowledge obtained in my study will facilitate to the development of novel methods for the disruption of tick feeding and pathogen transmission.

A procura por novos agentes com ação antifúngica está se intensificando nos últimos anos. A incidência de infecções fúngicas causadas por espécies do gênero Candida sp tem aumentado consideravelmente, principalmente em pacientes imunocomprometidos, nos quais podem apresentar gravidade. Por esse motivo, há uma intensa busca por novas substâncias com atividade antifúngica concomitante com pesquisa de novos alvos farmacológicos. A proteína H+-ATPase presente na membrana plasmática dos fungos é um novo alvo promissor, já que é exclusiva de células fúngicas. O Ebselen, mostrou ser um bom inibidor de H+-ATPase, sendo utilizado em alguns estudos como padrão de inibição dessa enzima. O eugenol, presente nos óleos essenciais de várias espécies vegetais é conhecido por apresentar diversas propriedades biológicas, incluindo ação antifúngica, a qual também parece estar relacionada à inibição desta ATPase microbiana. Soma-se a isso, o fato das 1,2-benzisoxazolin-3-onas, serem uma classe pouco explorada quanto ao seu perfil de ação biológica. Nesse contexto, foram planejados e sintetizados por hibridização molecular seis derivados benzisoxazolinônicos do eugenol e substâncias correlatas. Destes, quatro são inéditos, sendo que nenhum deles foi explorado quanto ao potencial antifúngico. Os produtos planejados foram sintetizados por meio de procedimentos sintéticos clássicos, tendo em comum as etapas finais de formação de intermediários do tipo ácido hidroxâmico seguido de ciclização às benzisoxazolinonas via reação de Mitsunobu. Os produtos finais foram caracterizados pelos métodos espectrométricos usuais. Na sequência, os produtos finais, bem como intermediários de síntese, foram avaliados quanto à sua atividade antifúngica in vitro frente a espécies de Candida spp. por meio de testes de microdiluição. Adicionalmente, os produtos mais promissores tiveram avaliados seus perfis de citotoxidade para determinação do índice de seletividade. Os testes revelaram que todos os compostos sintetizados apresentam atividade antifúngica superior à do eugenol. As N-fenilbenzisoxazolinonas (18, 19 e 20) foram as mais promissoras, apresentando, em alguns casos, valores de IC50 comparáveis ou até igual ao do fármaco fluconazol contra as espécies C. krusei e C. glabrata. O composto 19 apresentou um IC50 de 142 M frente a C. krusei, valor esse bem próximo ao alcançado pelo fluconazol (104 µM), sendo também o único híbrido planejado que se mostrou fungicida. O produto de hibridização 18, alvo do planejamento desse trabalho, foi o mais ativo contra C. glabrata com um CI50 de 53 µM. Ele obteve um índice seletividade bom (IS 3,4) e sua potência foi igual à do fluconazol (CI50 52 µM). Essa classe de heterociclos é ainda pouco explorada e mostrou ter atividade antifúngica, sendo passível de alterações estruturais para otimização dessa atividade biológica. / The incidence of fungal infections caused by Candida spp. has increased considerably, especially in immunocompromised patients, who may present gravity. The most widely used antifungal agents in the treatment of these infections are amphotericin B and azole. However, resistence to drugs and severe side effects limit the success of therapeutics. For these reasons, there is an intensive search for new substances with antifungal activity, mainly those acting in new molecular targets. Ebselen, a seleno-organic compound is a good inhibitor of H+-ATPase, an essential and unique yeast enzime. On the other hand, eugenol is a natural product present in the essential oils of several plant species, especially in clove, is and it known for its various biological properties, including antifungal action, which also seems to be related to inhibition of this microbial ATPase. Likewise, of benzisoxazolinones, are heterocycles chemically similar to ebselen, but relatively little explored as bioactive compounds. In this context, we describe here the design and synthesis of benzisoxazolinones derived from eugenol (our related substances) and ebselen, planned according to molecular hybridization strategy. They were synthesized by classical synthetic procedures and the final step consisted cyclization in hydroxamic acids into benzisoxazolinone via Mitsunobu reaction. The final products were obtained with yields ranging from 5 to 70% and were characterized by standard spectroscopic methods. Next, the final products, and synthetic intermediates were evaluated for their in vitro antifungal activity against Candida spp. by microdilution tests. In addition, the most promising products were evaluated for their cytotoxicity profiles for determining the selectivity indexes. Tests have shown that all synthesized compounds have antifungal activity higher than eugenol. The N-phenylbenzisoxazolinones (18, 19 and 20) were the most promising ones showing in some instances IC50 values comparable oe equal to those of fluconazole against C. krusei and C. glabrata. Compound 19 showed an IC50 of 142 µM against C. krusei, very close to that achieved by fluconazole (104 µM) and was the only hybrid with fungicidal activity. Product 18, eugenol-ebselen hybrid, was the most active one against C. glabrata with an IC50 of 53 µM, good selectivity index (3.4) and potency equal to that of fluconazole (IC50 52 µM). These results highlight these derivatives as good compounds for structural optimization and further studies, as investigations on its molecular mechanism of antifungal action. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES

In this work we present the first characterization of a Cu2+-transporting ATPase. The thermophilic bacteria Archaeoglobus fulgidus contains two genes, CopA and CopB, encoding for CPx-ATPases. CopB belongs to the subgroup IB-4 of the CPX-ATPases. These enzymes are characterized by a CPH motif in the 6th transmembrane domain and a His-rich N-terminus metal binding domain (MBD). CopB was heterologously expressed in E. coli. Membranes were prepared and used to measure activity. CopB was active at high temperature (75º C), high ionic strength and pH 5.7. The enzyme was activated by Cu2+, and in to a lesser extent by Ag+ and Cu+. CopB showed a Vmax = 5 µmol/mg/h and a high apparent affinity (K1/2 = 0.28 ± 0.09 μM) for Cu2+. Uptake of 64Cu2+ into everted vesicles was also measured in order to show that Cu2+ is not only activating the enzyme but being transported. Compared with CopB-WT, CopB-T (lacking the N-terminus MBD) did not show any difference in its activation by the different metal ions, demonstrating that the cytoplasmic MBD has no role in the metal selectivity. CopB-T also showed a 40 % decrease in the ATPase activity. CopB-WT and CopB-T presented similar levels of phosphorylation. However, CopB-T exhibited a reduced rate of dephosphorylation (slower transition from the E2P to the E2 conformation). These observations suggest a regulatory role for the cytoplasmic MBD.

Considerando que a deficiência hormonal causada pela ovariectomia promove alterações nas atividades das enzimas Na+,K+-ATPase, acetilcolinesterase (AChE) e ectonucleotidases e pode prejudicar a memória em ratas, no presente trabalho nós investigamos a influência do exercício físico sobre a ativação da Na+,K+-ATPase e acetilcolinesterase em hipocampo e córtex cerebral causada pela ovariectomia em ratas adultas, bem como sobre a hidrólise de nucleotídeos de adenosina no córtex cerebral e no soro. Também investigamos o efeito do exercício sobre a memória espacial e aversiva em ratas adultas ovariectomizadas. Ratas Wistar adultas foram divididas em quatro grupos: sham (submetidas à cirurgia sem a remoção dos ovários), exercício, ovariectomizadas (Ovx) e Ovx+exercício. Trinta dias após a cirurgia, os animais foram submetidos a um mês de exercício físico por 20 min, três vezes por semana. Logo após, as ratas foram decapitados, o soro coletado e o hipocampo e córtex cerebral dissecados, ou foram submetidas às tarefas de esquiva inibitória e labirinto aquático de Morris. Os dados demonstraram que o exercício físico reverte a ativação das atividades da Na+,K+-ATPase e AChE em hipocampo e córtex cerebral de ratas ovariectomizadas. A ovariectomia diminuiu a hidrólise de AMP no córtex cerebral e não alterou a hidrólise de nucleotídeos de adenosina sérica. Exercício per se diminuiu a hidrólise de ADP e AMP em córtex cerebral. Resultados também mostraram que ratas ovariectomizadas apresentaram prejuízo na memória aversiva e espacial (memória de referência e de trabalho), quando comparadas ao grupo controle (sham). Confirmando nossa hipótese, o déficit na memória foi revertido pelo exercício físico. Nossos achados mostram que a ovariectomia prejudica significativamente a memória/aprendizado aversiva e espacial e altera as enzimas Na+,K+-ATPase e AChE, e que o exercício físico preveniu tais efeitos. Esses dados sugerem que o exercício físico pode se mostrar uma estratégia importante para minimizar déficits cognitivos encontrados em mulheres pósmenopáusicas. / Hormone deficiency following ovariectomy causes activation of Na+,K+- ATPase and acetylcholinesterase (AChE), that has been related to cognitive deficits in experimental animals, and memory impairment. Considering that physical exercise presents neuroprotector effects, we decide to investigate whether exercise training would affect enzyme activation in hippocampus and cerebral cortex, as well as adenosine nucleotide hydrolysis in synaptosomes from cerebral cortex of ovariectomized rats, and ovariectomy-induced memory deficits in inhibitory avoidance and Morris water maze tasks. Female adult Wistar rats were assigned to one of the following groups: sham (submitted to surgery without removal of the ovaries), exercise, ovariectomized (Ovx) and Ovx plus exercise. Thirty days after surgery, animals were submitted to one month of exercise training for 20 min, three times per week. After, rats were euthanized, blood serum was collected and hippocampus and cerebral cortex were dissected, or rats were tested in inhibitory avoidance and Morris water maze tasks. Data demonstrated that exercise reversed the activation of Na+,K+-ATPase and AChE activities both in hippocampus and cerebral cortex of ovariectomized rats. Ovariectomy decreased AMP hydrolysis in cerebral cortex and did not alter adenosine nucleotides hydrolysis in blood serum. Exercise per se decreased ADP and AMP hydrolysis in cerebral cortex. On the other hand, AMP hydrolysis in blood serum was increased by exercise in ovariectomized adult rats. Results also show that ovariectomized rats presented impairment in aversive memory and spatial navigation, both in reference and working memory protocols, when compared to sham group (control). Confirming our hypothesis, ovariectomized rats submitted to exercise had those impairments prevented. We conclude that ovariectomy significantly impairs aversive and spatial reference learning/memory and related enzymes, and that physical exercise prevents such effects. These findings support that physical exercise might constitute an important strategy to minimize cognitive deficits found in postmenopausal women.