Características comportamentais de ratos SHR e LEWIS relacionadas à ansiedade e ao consumo de álcool após situações de estresse

Dissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas. Programa de Pós-Graduação em FarmacologiaO estresse provoca diversas alterações neurobiológicas, endócrinas e comportamentais que nem sempre são percebidas por todos os indivíduos da mesma intensidade, talvez por influência de fatores genéticos. Para estudarmos a inter-relação dos fatores genético e ambiental no comportamento, fizemos uso de duas linhagens isogênicas contrastantes para ansiedade e consumo de álcool (LEW e SHR). Estas linhagens foram submetidas a quatro protocolos de estresse com características distintas de estresse: imobilização única (IMOB-2h), imobilização repetida (IMOB-10d), variado severo (VARI-10d) e variado moderado e prolongado (VARIm-28d) e, posteriormente, testadas para verificar o nível de ansiedade (inata e condicionada) no modelo do labirinto em cruz elevado (LCE). Os mesmos estressores, exceto o IMOB-2h, foram utilizados para verificar o consumo de álcool destes roedores. Nossos resultados indicaram que o IMOB-10d e o VARI-10d promoveram alterações do tipo ansiogênico sendo que o IMOB-10d apareceu de modo mais pronunciado em ambas as linhagens e sexo. A re-exposição ao LCE não promoveu aumento da ansiedade nos animais previamente estressado pelo IMOB-10d e em SHR fêmeas previamente estressada nos demais protocolos. Em contraste com os dados obtidos no LCE, um aumento do consumo de álcool foi observado nos animais submetidos ao VARIm-28d em SHR machos sugerindo que o mesmo protocolo de estresse é capaz de promover alterações de comportamento distintas, dependendo do protocolo pós estresse. Com isso, ficou demonstrado que características relacionadas à ansiedade parecem não determinar o grau de consumo de álcool já que o uso de linhagens mais #ansiosas# e protocolos de estresse que promovem estados elevados de ansiedade não foram capazes de elevar o consumo de etanol. Stress situations provoke neurobiological, endocrine and behavioral changes that not always are perceived for all people perhaps in the same way maybe due to genetic influences. To study the relationship of the genetic factors and the environment (stress situation) in the behavior, we have used two contrasting inbred strains of rats for anxiety and in the alcohol consumption (Lewis and SHR rats). These strains had been submitted to four different stress protocols with distinct stress characteristics: acute immobilization (IMOB-2h), predictable immobilization (IMOB-10d), variable and severe stress (VARI-10d) and variable, moderate and chronic stress, and later tested to verify the level of anxiety (innate and condicionated) in the elevated plus maze (EPM). Our results indicate that the IMOB-10d and the VARI-10d promote anxiogenic-like alterations but the IMOB-10d produced a bigger alteration than VARI-10d in both male and females. The re-exposition to the EPM did not promote an increase of the anxiety level in animals when they were earlier stressed by the IMOB-10d and in the stressed females SHR by any protocol. In contrast with our results in the EPM, an increase of alcohol consumption were observed in the rodents submitted on VARIm-28d stress protocol but only in male SHR suggesting that the same stress protocol is capable of promote distinct alteration depending of the protocol after a stress situation. In conclusion, we have demonstrated that the anxiety-like characteristic apparently do not determine the degree of alcohol consumption once the use of more #anxious# rodent strain and several stress protocols that promote enhanced anxiety state did not increase of alcohol consumption

Envolvimento do neuropeptídeo Y na resiliência ao medo condicionado contextual em ratos

Tese (doutorado) - Universidade Federal de Santa Catarina, Centro de Ciências Biológicas, Programa de Pós-Graduação em Farmacologia, Florianópolis, 2013.O neuropeptídeo Y (NPY) é o peptídeo mais abundante no sistema nervoso central (SNC), sendo encontrado em regiões envolvidas com a regulação do estresse, memória, ansiedade e medo, como o hipocampo (HPC), amígdala e córtex pré-frontal. Embora alguns trabalhos suportem o envolvimento do NPY na mediação do comportamento emocional tanto em roedores como em humanos, ainda existe uma carência de dados mostrando o papel do NPY na resiliência ao estresse. Para isso, nosso estudo buscou verificar o envolvimento do NPY exógeno e endógeno, além de investigar a participação do receptor Y1 no teste do condicionamento aversivo contextual (CAC). Primeiramente, baseado no amplo espectro de atividades fisiológicas desempenhadas pelo sistema NPY, buscou-se observar os efeitos de baixas doses de NPY e do agonista Y1 Leu31Pro34-NPY (LP-NPY) e assim minimizar a incidência de efeitos adversos ao tratamento do medo condicionado, como prejuízos cognitivos, sedação e alterações da emocionalidade. Então, verificamos que a administração intracerebroventricular (i.c.v.) de NPY (3 pmol) e de LP-NPY (1 pmol) inibiram a aquisição e a consolidação (imediata e tardia) da memória de medo avaliada no teste do CAC. Por sua vez, somente o NPY facilitou a extinção, além de prejudicar a reconsolidação do medo condicionado. O pré-tratamento com o antagonista Y1, BIBO3304 (BIBO) bloqueou os efeitos do NPY, indicando a importante participação do receptor Y1, embora se sugira que a extinção do medo tenha também o envolvimento do receptor Y2. Para investigar a influência do NPY endógeno, submetemos os ratos ao ambiente enriquecido (AE) por 14 dias e após, verificamos o comportamento destes ratos na extinção do CAC, além de verificar a expressão do receptor Y1 nos ratos ?enriquecidos? no HPC, estrutura do SNC amplamente envolvida no condicionamento contextual. Como esperado, o AE facilitou a extinção do medo no teste do CAC, confirmando a participação do NPY. O envolvimento do NPY e do receptor Y1 foi confirmado com a detecção de níveis elevados da proteína do receptor Y1 no HPC em ratos enriquecidos. Interessantemente, esta elevação ocorreu somente nos ratos ?enriquecidos? que foram submetidos ao CAC, sugerindo que o AE favorece a resiliência ao aumentar a reatividade do receptor Y1, deixando este preparado para modular as respostas fisiológicas/comportamentais quando da ocorrência de situações traumáticas. Também procuramos verificar se a extinção do medo no teste do CAC era facilitada pelo bloqueio do auto-receptor Y2 com BIIE0246 (BIIE), já que este receptor regula a liberação de NPY. Nossos resultados mostraram que o tratamento com BIIE inibiu completamente a expressão do medo no teste do CAC, fazendo com que o comportamento dos ratos tratados comparáveis a ratos que não foram condicionados. Todavia, este efeito observado com o tratamento com BIIE permanece mesmo após a interrupção do tratamento, sugerindo que este efeito não é estado-dependente de uma atividade ansiolítica. Em adição, a participação de NPY endógeno é confirmada ao bloquear o receptor Y1 com BIBO, que reverteu os efeitos observados com o tratamento com o BIIE. De fato, mesmo quando não mais presente no organismo, o NPY continua a exercer seus efeitos, provavelmente através de mecanismo de neuroproteção, como observado em nosso experimento onde o NPY foi administrado 7 dias antes do teste do CAC, onde o NPY mas não o LP-NPY facilitou a extinção do medo. Com base no exposto acima, nossos dados mostram um efeito robusto do NPY inibindo o medo no teste do CAC, resposta essa que é mediada em parte pelo receptor Y1. Estes efeitos podem estar relacionados com o aumento da resiliência do SNC às situações traumáticas, sendo então uma abordagem de potencial terapêutico importante para o TEPT. 2013-12-06T00:41:21

The role of hippocampal NMDA receptors in long-term emotional responses following muscarinic receptor activation

Extensive evidence indicates the influence of the cholinergic system on emotional processing. Previous findings provided new insights into the underlying mechanisms of long-term anxiety, showing that rats injected with a single systemic dose of pilocarpine--a muscarinic receptor (mAChR) agonist--displayed persistent anxiogenic-like responses when evaluated in different behavioral tests and time-points (24 h up to 3 months later). Herein, we investigated whether the pilocarpine-induced long-term anxiogenesis modulates the HPA axis function and the putative involvement of NMDA receptors (NMDARs) following mAChRs activation. Accordingly, adult male Wistar rats presented anxiogenic-like behavior in the elevated plus-maze (EPM) after 24 h or 1 month of pilocarpine injection (150 mg/kg, i.p.). In these animals, mAChR activation disrupted HPA axis function inducing a long-term increase of corticosterone release associated with a reduced expression of hippocampal GRs, as well as consistently decreased NMDAR subunits expression. Furthermore, in another group of rats injected with memantine--an NMDARs antagonist (4 mg/kg, i.p.)--prior to pilocarpine, we found inhibition of anxiogenic-like behaviors in the EPM but no further alterations in the pilocarpine-induced NMDARs downregulation. Our data provide evidence that behavioral anxiogenesis induced by mAChR activation effectively yields short- and long-term alterations in hippocampal NMDARs expression associated with impairment of hippocampal inhibitory regulation of HPA axis activity. This is a novel mechanism associated with anxiety-like responses in rats, which comprise a putative target to future translational studies

Raptor-Mediated Proteasomal Degradation of Deamidated 4E-BP2 Regulates Postnatal Neuronal Translation and NF-κB Activity

The translation initiation repressor 4E-BP2 is deamidated in the brain on asparagines N99/N102 during early postnatal brain development. This post-translational modification enhances 4E-BP2 association with Raptor, a central component of mTORC1 and alters the kinetics of excitatory synaptic transmission. We show that 4E-BP2 deamidation is neuron specific, occurs in the human brain, and changes 4E-BP2 subcellular localization, but not its disordered structure state. We demonstrate that deamidated 4E-BP2 is ubiquitinated more and degrades faster than the unmodified protein. We find that enhanced deamidated 4E-BP2 degradation is dependent on Raptor binding, concomitant with increased association with a Raptor-CUL4B E3 ubiquitin ligase complex. Deamidated 4E-BP2 stability is promoted by inhibiting mTORC1 or glutamate receptors. We further demonstrate that deamidated 4E-BP2 regulates the translation of a distinct pool of mRNAs linked to cerebral development, mitochondria, and NF-κB activity, and thus may be crucial for postnatal brain development in neurodevelopmental disorders, such as ASD

Impact of host and environmental factors on β-glucuronidase enzymatic activity: implications for gastrointestinal serotonin

The gastrointestinal tract houses a reservoir of bacterial-derived enzymes which can directly catalyze the metabolism of drugs, dietary elements, and endogenous molecules. Both host and environmental factors may influence this enzymatic activity, with the potential to dictate the availability of the biologically-active form of endogenous molecules in the gut and influence inter-individual variation in drug metabolism. We aimed to investigate the influence of the microbiota, and the modulation of its composition, on fecal enzymatic activity. Intrinsic factors related to the host, including age, sex, and genetic background, were explored. Fecalase, a cell-free extract of feces, was prepared and used in a colorimetric-based assay to quantify enzymatic activity. To demonstrate the functional effects of fecal enzymatic activity, we examined β-glucuronidase-mediated cleavage of serotonin β-D-glucuronide (5-HT-GLU) and the resultant production of free 5-HT by HPLC. As expected, β-glucuronidase and β-glucosidase activity were absent in germ-free mice. Enzymatic activity was significantly influenced by mouse strain and animal species. Sex and age significantly altered metabolic activity with implications for free 5-HT. β-glucuronidase and β-glucosidase activity remained at reduced levels for nearly two weeks after cessation of antibiotic administration. This effect on fecalase corresponded to significantly lower 5-HT levels as compared to incubation with pre-antibiotic fecalase from the same mice. Dietary targeting of the microbiota using prebiotics did not alter β-glucuronidase or β-glucosidase activity. Our data demonstrate that multiple factors influence the activity of bacterial-derived enzymes which may have potential clinical implications for drug metabolism and the deconjugation of host-produced glucuronides in the gut

Enduring neurobehavioral effects induced by microbiota depletion during the adolescent period

The gut microbiota is an essential regulator of many aspects of host physiology. Disruption of gut microbial communities affects gut-brain communication which ultimately can manifest as changes in brain function and behaviour. Transient changes in gut microbial composition can be induced by various intrinsic and extrinsic factors, however, it is possible that enduring shifts in the microbiota composition can be achieved by perturbation at a timepoint when the gut microbiota has not fully matured or is generally unstable, such as during early life or ageing. In this study, we investigated the effects of 3-week microbiota depletion with antibiotic treatment during the adolescent period and in adulthood. Following a washout period to restore the gut microbiota, behavioural and molecular hallmarks of gut-brain communication were investigated. Our data revealed that transient microbiota depletion had long-lasting effects on microbiota composition and increased anxiety-like behaviour in mice exposed to antibiotic treatment during adolescence but not in adulthood. Similarly, gene expression in the amygdala was more severely affected in mice treated during adolescence. Taken together these data highlight the vulnerability of the gut microbiota during the critical adolescent period and the long-lasting impact manipulations of the microbiota can have on gene expression and behaviour in adulthood

Enduring neurobehavioral effects induced by microbiota depletion during the adolescent period

Peer ReviewedThe gut microbiota is an essential regulator of many aspects of host physiology. Disruption of gut microbial communities affects gut-brain communication which ultimately can manifest as changes in brain function and behaviour. Transient changes in gut microbial composition can be induced by various intrinsic and extrinsic factors, however, it is possible that enduring shifts in the microbiota composition can be achieved by perturbation at a timepoint when the gut microbiota has not fully matured or is generally unstable, such as during early life or ageing. In this study, we investigated the effects of 3-week microbiota depletion with antibiotic treatment during the adolescent period and in adulthood. Following a washout period to restore the gut microbiota, behavioural and molecular hallmarks of gut-brain communication were investigated. Our data revealed that transient microbiota depletion had long-lasting effects on microbiota composition and increased anxiety-like behaviour in mice exposed to antibiotic treatment during adolescence but not in adulthood. Similarly, gene expression in the amygdala was more severely affected in mice treated during adolescence. Taken together these data highlight the vulnerability of the gut microbiota during the critical adolescent period and the long-lasting impact manipulations of the microbiota can have on gene expression and behaviour in adulthood.Science Foundation Irelan

18 years of the European Journal of Wildlife Research: profile and prospects

Expanding on the foundation of Zeitschrift für Jagdwissenschaft in 1955, the European Journal of Wildlife Research (EJWR) continues to publish original research and reviews on all aspects of wildlife science regardless of the geographic region. Eighteen years after publication of the first issue of EJWR, we briefly reflect on the journal’s profile and prospects. Our target audience includes researchers, wildlife biologists, forestry and game management professionals, wildlife veterinarians, and other specialists, but we also aim at providing a resource of relevant information and scientific debate for practitioners and every person interested in wildlife science. With ecosystems being at the highest level of pressure due to land use change and other effects of the global crisis, the journal is in a key position to communicate relevant research to the scientific community around the world.Peer reviewe