<p dir="ltr">High exposure to manganese (Mn) through the inhalation of welding fumes has been shown to have a toxic effect to the human brain, leading to parkinsonian-like symptoms such as changes in mood, cognition, and motor function. Oxidative stress and GABAergic dysfunction, two proposed mechanisms implicated in Mn neurotoxicity, can be measured by edited MR spectroscopy (MRS). Previous animal studies have found depleted levels of glutathione, the brain’s antioxidant, and GABA in response to exposure to Mn. Past welder studies have shown altered GABA levels in highly exposed welders. However, GSH has not yet been measured in welders. Recent advances in edited MRS allow for the simultaneous measurement of GABA and GSH, however, one sequence does not ‘fit all’. Thus, there is a need to ensure accurate and reproducible measurements of these metabolites in the study of neurological disorders, such as Mn neurotoxicity.</p><p dir="ltr">The overall goal of this dissertation is to establish a reproducible edited MRS protocol and ensure accurate measurement of metabolites in the context of Mn neurotoxicity. This work has been accomplished in three steps. First, we developed an optimized and reproducible HERMES sequence that allows for the consistently reliable measurement of GABA and GSH at 3T. Second, we investigated whether toenail concentrations of manganese (Mn) and iron (Fe) serve as biomarkers for levels of GABA, GSH, and Glx in the brains of welders exposed to these metals. This aim explores the potential for toenails to be used as a risk assessment tool by evaluating correlations between toenail metal levels and brain metals and metabolites. Lastly, we examined whether excessive accumulation of metals in the brain has an impact on the relaxation times of metabolites. Due to its paramagnetic properties, brain accumulation of two major components of welding fumes, Mn and iron (Fe), may be measured noninvasively through increased magnetic resonance imaging (MRI) relaxation rates, R1 and R2*, respectively. This aim delves into the potential effects of metal exposure on the physical properties of brain metabolites, which could shed light on the accuracy of quantification.</p><p dir="ltr">Overall, the dissertation is a successful step towards establishing reproducible edited MRS acquisitions, and the accurate quantification in the application of Mn neurotoxicity. This work focused on developing methodologies and assessing physical properties for accurate GABA and GSH measurements and investigating risk-assessment methods for metal-induced neurotoxicity.</p>

This thesis examines the developmental neurotoxic effects of exposure to low-dose ionizing radiation (IR), alone or together with xenobiotics, during a critical period of neonatal brain development in mice. During mammalian brain development there is a period called the brain growth spurt (BGS), which involves extensive growth and maturation of the brain. It is known that neonatal exposure during the BGS to xenobiotics can have a negative impact on neonatal brain development, resulting in impaired cognitive function in the adult mouse. In humans, the BGS starts during the third trimester of pregnancy and continues for approximately 2 years in the child.   The present thesis has identified a defined critical period, during the BGS, when IR can induce developmental neurotoxicity in mice. The observed neurotoxicity was not dependent on sex or strain and manifested as altered neurobehaviour in the adult mouse. Furthermore, fractionated dose exposures appear to be as potent as a higher acute dose. The cholinergic system can be a target system for developmental neurotoxicity of IR, since alterations in adult mouse cholinergic system susceptibility were observed. Co-exposure to IR and nicotine exacerbated the behavioural disturbances and cholinergic system dysfunction. Furthermore, co-exposure with the environmental agent paraquat has indicated that the dopaminergic system can be a potential target.   In this thesis, clinically relevant doses of IR and a sedative/anesthetic agent (ketamine) were shown to interact and exacerbate defects in adult mouse neurobehaviour, learning and memory, following neonatal exposure, at doses where the single agents did not have any impact on the measured variables. This indicates a shift in the dose-response curve for IR, towards lower doses, if exposure occurs during the neonatal brain development. In addition, co-exposed mice, showing cognitive defects, expressed elevated levels of tau protein in the cerebral cortex. Furthermore, exacerbation of neurochemical deviations were observed following co-exposure compared to irradiation alone. Further investigations of neurotoxic effects following fractionated or acute low-dose IR, modelling the clinical situation during repeated CT scans or levels of radiation deposited in non-target tissue during radiotherapy, and possible interaction effects with xenobiotics, is of great importance in the field of radioprotection.

The causes of Amyotrophic Lateral Sclerosis (ALS), a fatal neurodegenerative disease that results in skeletal muscle paralysis, remain unclear. However, a nuclear DNA and RNA binding protein called TAR DNA binding protein 43 (TDP-43) has emerged as a critical marker of ALS pathology. A previous drug screen conducted in the Zarnescu laboratory showed that anti-diabetic drugs can rescue lethality in a fruit fly model of ALS based on TDP-43. These results suggested that the insulin signaling pathway might be altered in motor neurons in a TDP-43 dependent manner. Therefore, we hypothesized that the insulin pathway is interacting with TDP-43 in vivo and may be contributing to TDP-43neurotoxicity. Using genetic interaction approaches in flies we found that TDP-43dependent locomotor defects are sensitive to the levels of insulin receptor activity. In addition, genetic interaction data suggest that Akt is hyperactivated in motor neurons expressing TDP-43, possibly as a compensatory mechanism to enable survival. Finally, upregulating protein synthesis through S6K and 4EBP appears to have beneficial effects. These findings support our hypothesis and provide insights into potential therapeutic strategies to help treat this devastating disease.

Murine neuronal networks, derived from embryonic frontal cortex (FC) tissue grown on microelectrode arrays, were used to investigate zinc toxicity at concentrations ranging from 20 to 2000 mM total zinc acetate added to the culture medium. Continual multi-channel recording of spontaneous action potential generation allowed a quantitative analysis of the temporal evolution of network spike activity generation at specific zinc acetate concentrations. Cultures responded with immediate concentration-dependent excitation lasting from 5 to 50 min, consisting of increased spiking and enhanced, coordinated bursting. This was followed by irreversible activity decay. The time to 50% and 90% activity loss was concentration dependent, highly reproducible, and formed linear functions in log-log plots. Network activity loss generally preceded morphological changes. 20% cell swelling was correlated with 50% activity loss. Cultures pretreated with the GABAA receptor antagonists bicuculline (40 mM) and picrotoxin (1 mM) lacked the initial excitation phase. This suggests that zinc-induced excitation may be mediated by interfering with GABA inhibition. Partial network protection was achieved by stopping spontaneous activity with either tetrodotoxin (200 nM) or lidocaine (250 mM). However, recovery was not complete and slow deterioration of network activity continued over 6 hrs. Removal of zinc by early medium changes showed irreversible, catastrophic network failure to develop in a concentration-dependent time window between 50% and 90% activity loss. Investigation of entry routes suggested the L-type but not N-type calcium channels to be the main entry pathway for zinc. Data are presented implicating the chloride channel to be an additional entry route.

Dans l’arsenal thérapeutique anticancéreux, les agents anti-microtubules (MTA) occupent une place essentielle dans le traitement de tumeurs solides et d’hémopathies malignes. Néanmoins, leur utilisation est limitée par l’induction d’une toxicité neurologique qui affecte la qualité de vie des patients et dont les mécanismes d’action demeurent peu compris. L’absence de solutions préventives ou curatives réellement efficaces, reflète la complexité des mécanismes d’action des MTA. Dans le cadre du projet « Mitotarget » (7ème PCRD) porté par le partenaire industriel Trophos, notre objectif était de préciser le mécanisme à l’origine de la neurotoxicité des MTA et d’évaluer le potentiel neuroprotecteur de l’olesoxime, composé ayant fait la preuve de son efficacité neuroprotectrice dans différents modèles de pathologies neurodégénératives. Nous montrons ici que les réseaux microtubulaire (dynamique des microtubules, localisation de la protéine EB1) et mitochondrial (motilité des mitochondries), cibles des MTA dans les cellules cancéreuses, sont aussi affectés dans les cellules de type neuronal. Leur préservation par l’olesoxime est nécessaire à l’établissement d’une neuroprotection. Ce travail met en évidence l’originalité du mécanisme d’action de l’olesoxime, premier neuroprotecteur capable d’agir tout à la fois sur les microtubules et les mitochondries, et souligne l’importance des liens étroits existant entre ces deux compartiments. Deux axes d’étude ont été initiés à la suite de ce projet afin de (i) déchiffrer les interconnexions microtubules-mitochondries dans la réponse des cellules cancéreuses aux MTA; (ii) préciser l’importance et la régulation post-traductionnelle de la protéine EB1 dans l’efficacité anti-migratoire des MTA. L’ensemble des données obtenues appelle à poursuivre la caractérisation des mécanismes de réponse aux agents anti-microtubules afin d’optimiser les stratégies thérapeutiques existantes. / Nowadays, the so-called Microtubule Targeting Agents (MTAs) remain benchmark clinical treatments displaying high efficiency and are still widely used against a broad spectrum of tumors and hemopathies. The new compounds in clinical development and the discovery of their anti-angiogenic properties make them a family booming. However, MTAs treatment is limited by the occurrence of neurological toxicities that greatly impair patients quality of life and which mechanisms of action are still poorly understood. The current absence of really efficient curative of preventive strategies underline the complexity of MTA mechanisms of action. In the framework of the “MitoTarget” project from the 7th PCRD,lead by the industrial partner Trophos, we aimed to precise MTA neurotoxic mechanisms and to evaluate neuroprotective potential of olesoxime, a compound that already showed to be efficient in various models of neurodegenerative diseases. Our data show that microtubular (microtubule dynamics parameters, EB1 protein localization) and mitochondria (mitochondria) networks, MTA targeted compartments in cancer cells, are damaged in neuronal-like cells. Interestingly, olesoxime neuroprotective activity implies preservation of both microtubule and mitochondria from MTA-induced damages. This work highlights the original mechanism of action of olesoxime as the first neuroprotective agent able to act on both microtubule and mitochondria and underlines the strengthened link existing between these compartments. It thus gave rise to two side projects with the aim to (i) decipher microtubule-mitochondria interconnections in response to MTA treatment; (ii) precise the importance and regulation of EB1 in the anti-migratory efficacy of MTA by looking at EB1 post-translational modifications. Altogether, the data obtained incite to keep on characterizing mechanisms involved in response to MTA in order to optimize the existing therapeutic strategies.

As doenças neurodegenerativas (DN), incluindo doença de Alzheimer e doença de Parkinson, estão entre as principais causas de mortalidade e morbidade nos países ocidentais. Não há ainda um tratamento definitivo para estas neuropatias, mas os estudos têm apontado para mecanismos comuns de toxicidade, que incluem disfunção mitocondrial, estresse oxidativo e apoptose. Assim, as mitocôndrias constituem alvos importantes para futuras estratégias de neuroproteção visando tratar, prevenir ou retardar a neurodegeneração. A biodiversidade da fauna brasileira representa uma fonte promissora e ainda pouco explorada de novas moléculas com (i) atividade neuroprotetora e, portanto, potencial para originar novos fármacos para o tratamento destas doenças; ou ainda com (ii) atividade neurotóxica, podendo ser utilizadas como ferramentas de estudo de mecanismos de neurotoxicidade. O presente estudo teve por objetivo investigar o possível potencial neuroprotetor e/ou neurotóxico de peptídeos de baixo peso molecular, obtidos a partir do veneno de diferentes espécies de serpentes do gênero Bothrops por meio da investigação dos efeitos destes compostos na função mitocondrial de cérebro de rato. Duas das frações estudadas (obtidas a partir do veneno da B. atrox e da B. jararaca) apresentaram um interessante potencial protetor contra o intumescimento osmótico mitocondrial e a produção de espécies reativas de oxigênio (ERO), eventos associados à transição de permeabilidade mitocondrial e à morte celular. Por outro lado, outra fração, obtida a partir do veneno da B. pirajai, apresentou potencial neurotóxico. Estes achados podem ser úteis para estudos mecanísticos e também no estabelecimento de futuras estratégias de tratamento das doenças neurodegenerativas, tendo as mitocôndrias como alvos terapêuticos (terapia alvo). / The neurodegenerative diseases, including Alzheimer\'s and Parkinson\'s diseases, are among the main causes of morbidity and mortality in Western countries. A definitive treatment for these neuropathies has not yet been found, but studies have indicated common mechanisms of toxicity, including mitochondrial dysfunction, oxidative stress and apoptosis. Therefore, mitochondria represent important targets for the future neuroprotective strategies aimed to treat, prevent or delay the neurodegeneration. The Brazilian fauna biodiversity represents a promising and under explored source of new molecules with (i) neuroprotective activity and potential to originate new drugs for the treatment of these diseases; or yet, with (ii) neurotoxic activity, representing tools to study neurotoxicity mechanisms. The present study aimed to investigate the possible neuroprotective and/or neurotoxic potential of low-molecular-mass peptides extracted from the venom of different species of Bothrops genus snakes by investigating their effects on rat brain mitochondrial function. Two of the studied fractions (from B. atrox and B. jararaca venoms) presented an interesting protective potential against both the mitochondrial swelling and reactive oxygen species (ROS) production, events associated with mitochondrial permeability transition and cell death. On the other hand, other fraction (from B. pirajai venom) presented a neurotoxic potential. These findings might be useful for mechanistic studies and also for the establishment of future strategies of neurodegenerative diseases treatment, using mitochondria as therapeutic targets (targeted therapy).

Pode-se dizer que o câncer, ao lado da AIDS, é a doença que atinge, a níveis mundiais, maior clamor da sociedade por desenvolvimento de cura e melhor qualidade de vida ao paciente. Várias terapias são utilizadas para o tratamento do câncer, podendo-se destacar a importância da quimioterapia em inúmeros protocolos de tratamento. Um dos fármacos mais antigos e mais utilizados na quimioterapia é a cisplatina, que ao longo dos seus mais de 30 anos na prática clínica demonstrou sua eficácia e foi incorporada no protocolo de tratamento de uma variedade de tumores. Entretanto, ao lado se sua comprovada eficácia, é inerente o desenvolvimento de vários efeitos colaterais resultante de sua toxicidade à células sadias. Neste trabalho, destaque é dado a neurotoxicidade, que surge em um grande número de pacientes que recebem tratamento com a cisplatina. Inúmeros esforços têm sido realizados na tentativa de prevenir ou controlar os efeitos tóxicos induzidos pela cisplatina, com atenção especial à suplementação do tratamento quimioterápico com compostos antioxidantes. Nesse sentido, este estudo visa avaliar a associação da curcumina, um composto polifenólico com notável atividade antioxidante e neuroprotetora, ao tratamento com cisplatina, em um modelo in vitro com culturas de células PC12. A curcumina é reconhecidamente um composto polifenólico com um amplo espectro de atividades biológicas, com suas propriedades neuroprotetoras demonstradas em vários modelos estudados estando principalmente relacionadas com seu potencial antioxidante. Os múltiplos mecanismos de ação dos compostos fenólicos e suas propriedades neuroprotetoras atribuídas a capacidade de inibição das espécies reativas de oxigênio indicam um potencial de inibição da neurotoxicidade que surgem durante o tratamento quimioterápico com a cisplatina, visto a reconhecida capacidade de geração de espécies reativas de oxigênio da cisplatina. Como a geração de espécies reativas de oxigênio intracelular pode resultar em danos ao DNA pela ação direta das espécies reativas ou indiretamente via produtos de degradação da peroxidação lipídica, nosso estudo avaliou o potencial protetor da curcumina contra a genotoxicidade induzida pela cisplatina em culturas de células PC12, analisando a indução de micronúcleos e a análise de quebras de cadeia simples, cadeia dupla do DNA e sítios álcali-labeis pelo ensaio do cometa. Antes da avaliação antigenotóxica da curcumina, foi realizada uma avaliação citotóxica e genotóxica, em que pode-se observar que em concentrações elevadas a curcumina é citotóxica e genotóxica às células PC12. O efeito protetor da curcumina foi avaliado em pré-tratamento das culturas de células PC12 com concentrações não tóxicas. Nas três concentrações de curcumina estudadas não houve indícios de interferência com a citotoxicidade ou o efeito citostático da cisplatina. Embora o efeito protetor da curcumina contra os danos induzidos ao DNA de células PC12 não tenha sido evidente nos resultados obtidos pelo ensaio do cometa, a curcumina foi eficaz na redução de micronúcleos induzidos pela cisplatina, de maneira semelhante nas três concentrações avaliadas. Os resultados positivos obtidos nesse estudo juntamente com os dados pré-existentes a respeito de efeitos protetores da curcumina incentivam novas pesquisas em relação aos possíveis benefícios da utilização da curcumina em terapia combinada aos quimioterápicos. / Several therapies are used for treating cancer and the importance of chemotherapy can be emphasized in many protocols of treatment. One of the oldest and most used drugs in chemotherapy is the cisplatin which, with its more than thirty years of clinical practice, has demonstrated its effectiveness being incorporated into the protocol of a variety of tumor treatments. On the other hand, it is intrinsic the development of various side effects resulting from its toxicity to healthy cells. This research gives emphasis to the neurotoxicity which appears in a large number of patients receiving the cisplatin treatment. Numerous efforts are being made in attempt to prevent or even control the toxic effects induced by cisplatin with special attention being given to the supplementation of chemotherapy treatment with antioxidant compounds. In that sense, this study aims to assess the association of curcumin, a polifenolic compound with remarkable antioxidant and neuroprotective activity, to the cisplatin treatment in an in vitro neuronal model with PC12 cell cultures. The curcumin is admittedly a polifenolic compound with a broad spectrum of biological activities and their neuroprotective properties, demonstrated in several models, are strongly related to its antioxidant potential. The multiple mechanisms of action of the phenolic compounds and their neuroprotective properties credited to the ability of inhibition of the reactive oxygen species indicate a potential inhibition of neurotoxicity that takes place during the chemotherapy treatment with cisplatin, given its well known ability to generate reactive oxygen species. Since the generation of reactive oxygen species intracellular can result in DNA damage by direct action of reactive species or indirect, through degradation products of the lipid peroxidation, our studies evaluated the potential protector of curcumin against the genotoxicity induced by cisplatin in PC12 cell cultures by examining the induction of micronuclei and analyzing DNA single strand breaks, double strand breaks, and alkali-labeis sites through the comet test. Before the antigenotoxic evaluation of the curcumin, a cytotoxic and genotoxic test was conducted where it was observed that when in high concentrations the curcumin is cytotoxic and genotoxic to the PC12 cells. The protective effect of curcumin was evaluated at the pre-treatment of PC12 cell cultures with non-toxic levels of concentration. In the three studied curcumin concentrations there was no evidence of interference with the cytotoxicity or cytostatic effect of the cisplatin. Although the protective effect of the curcumin put against the damage caused to the DNA of cells PC12 was not evident in the results obtained by the comet test, the curcumin was equally effective in the reduction of micronuclei induced by the cisplatin in all three concentrations evaluated. The positive results obtained in this study combined with the pre-existing data about the protective effects of the curcumin encourage some more new research on possible benefits of using curcumin in combination to chemotherapy.

Nos últimos anos, a prescrição e o consumo de medicamentos psicotrópicos vêm aumentando ao redor do mundo, crescendo assim a presença destes compostos em efluentes. Embora os esgotos domésticos e hospitalares em geral sejam encaminhados para as Estações de Tratamento de Esgotos (ETEs), o tratamento convencional aplicado não é eficiente na remoção da maioria dos fármacos e, consequentemente, estes poluentes alcançam os corpos hídricos. Frente a essas informações, é importante entender os possíveis danos causados tanto para seres humanos, como para o ecossistema como um todo, após a exposição a esses fármacos, por meio da água contaminada. Desta maneira, nossa hipótese é que os fármacos psicotrópicos presentes em amostras de água possam induzir a danos neuronais e no desenvolvimento de organismos não alvo, como peixes. Para responder a esta hipótese, avaliamos os efeitos teratogêncios e neurotóxicos de fármacos psicotrópicos como venlafaxina, haloperidol, carbamazepina e fluoxetina, por meio da análise de parâmetros de letalidade e sub-letalidade nos em Danio rerio, visando a determinação dos possíveis efeitos teratogênicos. Adicionalmente, utilizamos a determinação da atividade da enzima acetilcolinesterase como biomarcador de neurotoxicidade em larvas, após a exposição dos embriões e a avaliação do padrão de movimentação. Os fármacos foram testados individualmente e em misturas, para avaliar os efeitos da co-exposição, simulando um cenário mais real de contaminação aquática. Os resultados mostraram que os fármacos carbamazepina e venlafaxina não apresentaram nenhum dos efeitos analizados de forma estatisticamente significativa. Já os fármacos haloperidol e fluoxetina, mesmo não apresentando efeito sobre a teratogenicidade e movimentação, provocaram um aumento na atividade enzimática, em doses consideradas bastante baixas. Considerando que esta estimulação inicial voltou a níveis próximos ao controle negativo e, no caso da fluoxetina, um estudo inibição da atividade em doses mais altas, sugerimos a manifestação de hormesis. A mistura dos fármacos não apresentou efeito significativo nas avaliações realizadas, mostrando que o aumento da atividade enzimática dfoi antagonizado na quando em mistura. Nosso trabalho mostra a importância do estudo de toxicidade em doses baixas, para que uma correta avaliação do risco seja possível. / In the last years, the prescription and use of psychotropic drugs have being increasing around the world, consequently the discharge of these drugs and their metabolites have also increased in wastewater. Considering the presence in the environment and the toxic effects to target and non target species, these compounds were classified as emerging water contaminants. The main sources of these compounds are the domestic and hospital treated effluents, because the conventional treatment applied by effluent treatment plants is not efficient to remove most of drugs, and consequently these pollutants reach the water bodies. In this context, it is important to understand the possible damages to humans and ecosystems after exposure to these drugs through contaminated water. In that way, the hypothesis of this research is that psychotropic drugs present in water samples could induce neurologic and development damages in non-target organisms, such as fish. Therefore, we propose the evaluation of teratogenic and neurotoxic effects of the psychotropic drugs Fluoxetine, Carbamazepine, Venlafaxine and Haloperidol using the determination of the activity of cholinesterase enzyme as biomarker of neurotoxicity in larvae, after the exposure of Danio rerio eggs. Lethality and sub-lethality parameters will be also analyzed in the same organisms to establish the potential of teratogenic effects, the movement analysis was also performed. These drugs will be tested individually and in mixtures to evaluate the effects of coexposure simulating a real scenario of water pollution. Carbamazepine and venlafaxine don\'t responde with significant results in any of these evaluations. Haloperidol and Fluoxetine presented negative results in teratogenic and moviment effects, but they incresed the activity of acetylcholinesterase enzime, this result was different from another study, that these drugs in highest concentrations decrease this activity, so it can indicate the presence of hormesis effect. However, the mixtures of drugs don\'t present any significant effect on all evaluations, this result can suggest that these effects caused by haloperidol and fluoxetine on the enzime activity were canceled, it can be a interaction of antagonist effects on the mixtures.

O uso recreacional e os efeitos deletérios de drogas lícitas e/ou ilícitas é considerado um problema de saúde pública. Dentre estas drogas destaca-se a cetamina, um anestésico empregado na veterinária e o etanol, a droga lícita mais utilizada. Além dos efeitos centrais, estudos revelam que a cetamina e o etanol possuem propriedades imunomodulatórias. No entanto, poucos estudos são realizados com a associação destas drogas; assim, o objetivo deste estudo foi o de avaliar os efeitos tóxicos, imunotóxicos, o potencial oxidativo e os possíveis efeitos neurotóxicos resultantes do consumo associado ou não das mesmas. Para isso, ratos Wistar foram tratados uma vez ao dia, por até 28 dias, com injeções intraperitoneais (15 ou 30mg/kg/PV) de cetamina (K) ou com etanol a 10% (E) via oral (gavagem ou adicionado a água de bebida), ou ainda em associação (KE). O grupo controle recebeu apenas os veículos dos tratamentos. Ao final do experimento, os animais foram submetidos à eutanásia para realização de avaliações toxicológicas, imunológicas, oxidativas e antioxidativas, e de neurotransmissores centrais. Os resultados revelaram redução no ganho de peso dos animais tratados com KE e aumento de ácido ascórbico urinário. A redução de pH e glicose urinária foi observada nos grupos E e KE. Na bioquímica, todos os grupos apresentaram aumento de HDL, porém a associação das drogas levou a um aumento do colesterol, enquanto no grupo K, observou-se diminuição dos triglicérides e da fosfatase alcalina. Ainda, somente o grupo KE apresentou alterações na função renal. Todos os grupos experimentais exibiram alterações histopatológicas hepáticas e/ou vesicais. Os grupos E e KE apresentaram alterações no número de células mieloides e linfoides concomitantes ao aumento na celularidade de medula óssea. Apenas animais do grupo KE apresentaram alteração na resposta do tipo Th2. Foi observado o aumento da peroxidação lipídica nos animais tratados com K, bem como aumento na atividade de GPx e CAT e do conteúdo de GSSG dos animais tratados com E e/ou KE nos órgãos analisados. Ao nível central, observou-se elevação dos níveis de DA e NOR no hipocampo dos animais do grupo E, e 5HT nos animais do grupo K; além de aumento de DA e 5HT no córtex pré-frontal dos animais dos grupos E e K, respectivamente. Assim, concluímos que a associação KE promove redução do ganho de peso não relacionado ao consumo de ração. O etanol, em associação ou não, promove alterações nos parâmetros urinários; a cetamina promove diminuição nos níveis de FA e as drogas quando em associação alteram o perfil lipídico e renal na dependência da dose e do tempo de administração. Efeitos pró-oxidantes entraram em equilíbrio devido à ação de antioxidantes. O etanol, em associação ou não, promove alterações em células do sistema imune, no entanto, sem promover imunomodulação sobre suas respostas. O etanol e/ou a cetamina também promovem alterações histopatológicas hepáticas, enquanto que a associação ainda promove lesões vesicais. Além disso, ambas as drogas promovem alterações no perfil neuroquímico central; porém, quando associadas não promoveram efeitos sinérgicos sobre os parâmetros avaliados. / The recreational use and the deleterious effects of licit and/or illicit drugs are considered a public health issue. Among these drugs are ketamine, used in veterinary anaesthesia, and ethanol the most commonly licit drug. Besides their central effects, studies have shown that ketamine and ethanol have immunomodulatory properties. However, few studies are conducted with the association of both drugs; thus, we aimed to evaluate the toxic and immunotoxic effects, as well the oxidative potential and the possible neurotoxic effects resulted from the consumption of these drugs associated or not. For this, Wistar rats were treated once daily for up to 28 days with intraperitoneal injections (15 or 30mg/kg/BW) of ketamine (K) or orally with 10% ethanol (E) (gavage or drinking water) or the association of both treatments (KE). Control group received only vehicles. At the end of the experimental period, the animals were euthanized for toxicological, immunotoxicological, oxidative stress and antioxidant status evaluation, and central levels of neurotransmitters. KE animals showed reduced body weight gain and increased levels of urinary ascorbic acid. Reduction of urinary pH and glucose was observed in E and KE groups. Biochemistry analysis showed an increase in HDL levels in all experimental groups; although, KE showed an increase in cholesterol levels, while K group exhibited reduction in triglycerides and alkaline phosphatase (ALP) levels. Only KE group showed renal function alterations. All experimental groups showed hepatic and/or bladder histopathology alterations. E and KE groups exhibited alterations in myeloid and lymphoid cells number with concomitant increased in bone marrow cellularity. Only animals of KE group presented changes in Th2 response. Increased in lipid peroxidation was observed in K-treated animals, as well as GPx and CAT activities and GSSG content of animals treated with E and/or KE. We observed elevation in DA and NOR in hippocampus of E group and 5HT in K group, in addition to an increase in DA and 5HT of the prefrontal cortex of E and K groups, respectively. Thus, we conclude that KE association promotes reduction in body weight gain not related to food consumption. Ethanol, in combination or not, promotes urinary alterations; ketamine reduces ALP levels and the drugs, when in combination, alter lipid and renal profile depending on dose and time of administration. Pro-oxidant effects were balanced due to the action of antioxidants. Ethanol, associated or not, promotes alterations on immune cells without immunomodulatoty responses. Ethanol and/or ketamine also promotes liver histopathological alterations, while this association promotes bladder lesions. In addition, both drugs promote changes in central neurochemical levels; however, when associated do not promoted synergistic effects in evaluated parameters.

Submitted by PPG Zoologia (zoologia-pg@pucrs.br) on 2018-06-29T17:28:48Z No. of bitstreams: 1 Dissertac?a?o_Mestrado_Raphaela_ (003).pdf: 1487803 bytes, checksum: 85e14999295df94278920f21e8362c4d (MD5) / Approved for entry into archive by Sheila Dias (sheila.dias@pucrs.br) on 2018-07-09T11:35:44Z (GMT) No. of bitstreams: 1 Dissertac?a?o_Mestrado_Raphaela_ (003).pdf: 1487803 bytes, checksum: 85e14999295df94278920f21e8362c4d (MD5) / Made available in DSpace on 2018-07-09T11:42:43Z (GMT). No. of bitstreams: 1 Dissertac?a?o_Mestrado_Raphaela_ (003).pdf: 1487803 bytes, checksum: 85e14999295df94278920f21e8362c4d (MD5) Previous issue date: 2018-02-26 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES / The increase of environmental pollutants sources directly affects the ecosystem and human health, mainly due to loss of soil and water quality. Among these toxic agents are included endocrine disruptors, which can cause changes in biochemical pathways and physiological changes in organisms. Studies with different forms of bisphenols, compounds present in many products of human consumption and the environment, had already demonstrated extremely adverse effects on behavior and gene expression of targets related to neural and endocrine systems during development. The zebrafish, Danio rerio, is an excellent model organism for studying neurotoxicity, due to its advantages with respect to specific aspects of their development and a low maintenance cost when compared to other vertebrate model organisms. This work proposes the investigation of the effects of bisphenol A (BPA), its chlorinated derivatives and its substitutes (BPS and BPF), toxicological and behavioral parameters combined with characterization of its potential underlying cellular and molecular mechanisms such as cell death and proteins and gene expression in the nervous and endocrine systems. After the embryo and larvae were exposed to different concentrations of BPA and its derivatives, morphological analyzes and embryo survival were carried out, as well as behavioral tests of locomotion and cognitive evaluation of the larvae. In the concentrantion curve of the different bisphenolic compounds, no significant difference was observed in the survival, teratogenesis and hatching of the animals, however, a significant increase was detected in the exploratory parameters of the BPA and BPF groups. When comparing the aversive behavior of the BPA with the DMSO, there was no statistical difference between the groups, whereas in the BPS an improvement in the escape response of the larvae occurred. Additionally, behavioral and locomotor activity analyzes will be performed in adult animals previously treated with the compounds studied during the initial development. In this way, it is expected to contribute to the current knowledge of neurotoxicity mechanisms related to endocrine disruption caused by contaminants of the bisphenol family. / O crescimento das fontes de poluentes ambientais afeta diretamente o ecossistema e a sa?de humana, principalmente devido ? perda da qualidade do solo e da ?gua. Entre estes agentes t?xicos, est?o inclu?dos os disruptores end?crinos, os quais podem acarretar diversas mudan?as em vias bioqu?micas e altera??es fisiol?gicas em organismos. Estudos com diferentes formas de bisfen?is, compostos presentes em diversos produtos de consumo humano e inclusive no ambiente devido ? contamina??o por efluentes industriais, j? apresentaram efeitos extremamente nocivos na regula??o da express?o de genes relacionados ao sistema neural e end?crino durante o desenvolvimento, al?m de altera??es comportamentais severas. O zebrafish, Danio rerio, ? um excelente organismo modelo para estudar neurotoxicidade, uma vez que possui vantagens em rela??o a aspectos espec?ficos do seu desenvolvimento e um custo reduzido de manuten??o quando comparado a outros vertebrados modelos. Este projeto prop?s o estudo dos efeitos do bisfenol A (BPA), seus derivados clorados e substitutivos (BPS e BPF), sobre par?metros de embriotoxicidade e comportamento, buscando tamb?m caracterizar os mecanismos celulares e moleculares dos potenciais efeitos observados avaliando morte celular e n?veis de prote?nas e express?o de genes alvos em embri?es e larvas de zebrafish. Para tanto, ap?s a exposi??o dos embri?es e larvas a diferentes concentra??es de BPA e seus derivados, foram realizadas an?lises morfol?gicas e sobreviv?ncia dos embri?es; ensaios comportamentais de locomo??o e avalia??o cognitiva das larvas. Ao realizar a curva de concentra??o dos diferentes compostos bisfen?licos, n?o foi observada diferen?a significativa na sobreviv?ncia, teratog?nese e eclos?o dos animais, por?m, ao analisar o comportamento explorat?rio dos animais tratados com BPA e BPF foi detectado um aumento significativo nos par?metros explorat?rios das larvas. Ao compararmos o comportamento aversivo do BPA com o controle tratado com o ve?culo DMSO, n?o existiu diferen?a estat?stica entre os grupos, enquanto no BPS ocorreu uma melhora na resposta de escape das larvas. Adicionalmente, ser?o realizadas an?lises comportamentais e de atividade locomotora em animais adultos previamente tratados com os compostos estudados durante o desenvolvimento inicial. Desta forma, espera-se contribuir para o conhecimento atual dos mecanismos de neurotoxicidade relacionados ? disrup??o end?crina causada por contaminantes da fam?lia dos bisfen?is.