Este trabalho tem o objetivo avaliar o efeito da adrenalectomia bilateral no processo neurodegenerativo e cicatricial na via nigro estriatal dopaminérgica lesada pela 6- hidroxidopamina (6-OHDA) em ratos, e deste modo contribuir para a interpretação dos efeitos dos hormônios adreno esteróides nos processos neurodegenerativos e neurotróficos nas lesões do sistema nervoso central. Ratos Wistar adultos jovens foram submetidos à cirurgia de retirada bilateral das glândulas adrenais ou cirurgia simulada para a mesma. Após 2 dias, os animais foram submetidos à cirurgia estereotáxica para lesão do sistema nigro estriatal dopaminérgico através da injeção unilateral de 6-OHDA na substância negra. Os animais receberam diariamente doses de reposição hormonal de corticosterona (10mg/kg) ou solução veículo. Após um período de 72 horas, 1 semana e 3 semanas, os animais foram sacrificados por decapitação, e as regiões do mesencéfalo ventral e corpo estriado foram dissecados bilateralmente. O tecido foi processado para o método de Western blot onde foram analisados os seguintes marcadores: tirosina hidroxilase (TH enzima que participa da via de conversão da dopamina), proteína ácida fibrilar glial (GFAP proteína do filamento intermediário do citoesqueleto do astrócito), bem como as moléculas da matriz extracelular fosfacan, neurocan, sulfato de condroitina e NG2, a proteína Laminina e finalmente, o fator de crescimento fibroblástico-2 (FGF-2) na região do mesencéfalo ventral, onde observa-se ações tróficas nos neurônios dopaminérgicos nigras que podem exercer efeitos no processo inflamatório pelas suas ações gliogênica e angiogênica. Deste modo, a neurodegeneração dopaminérgica foi avaliada pelos níveis da TH, a ativação astrocitária pelos níveis da GFAP, o processo de cicatrização pela regulação das moléculas da matriz extracelular e as respostas tróficas pelo FGF-2. Pelos resultados obtidos é possível que hormônios glicocorticóides adrenais modulem os elementos envolvidos na neurodegeneração e reparo e cicatrização do sistema dopaminérgico. / This study has the objective to evaluate the effect of the bilateral adrenalectomy in the neurodegenerative process and cicatrization on the nigroestriatal pathway injury through 6-hydroxydopamine (6-OHDA) in rat, and in this way contribute for the interpretation of the adreno esteroides hormones effects in the neurodegenerative and neurotrophics process in the central nervous system.injury Adult rats wistar were submitted in a surgery to take the bilateral adrenal gland or a simulate surgery of the same. After 2 days the animals was involved into a stereotaxic surgery to nigroestriatal dopaminergic lesion with a unilateral injection of the 6-OHDA in the nigra. The animals received daily corticosterone hormonal (10mg/kg) or vehicle solution. After a period of 72 hours, 1 week and 3 weeks, the animals were decapitated, and the regions of the ventral midbrain and striatum it was bilaterally dissected. The tissue was processed by Western blot method and analyzed for the following markers: tyrosine hydroxylase (TH - enzyme that join of the dopamine conversion pathway), glial fibrilar acid protein (GFAP protein of the intermediary filament of the astrocyte citoesqueleto, the moleculars extracellular matrix phosphacan, neurocan, chondroitin sulfate and NG2, the Laminin protein and finally, the neurotrophic factor (FGF-2) in the region of ventral midbrain, where watched trophic actions in the dopaminergic neurons that could have effects in the inflammatory processes by your gliogenic and angiogenic actions. In this way, the dopaminergic neurodegeneration was evaluated by the TH levels, the astrocytary activation by the GFAP levels, the cicatrization processes by the molecules regulation of the extracellular matrix and the trophics FGF-2 answers. By the results it is possible that adrenal glucocorticoid hormones modulate the elements involved in neurodegeneration, repair and cicatrization of the dopaminergic system.

The vivid memory of an emotional event, as well as memory for incidental details associated with the arousing event, has been referred to collectively as a “flashbulb memory”. An important aspect of flashbulb memory in people is that an emotional event enhances memory of contextual details, such as the weather, or clothes one was wearing at the time of the event. Therefore, an emotional event not only produces a detailed memory of the event, itself, but also enhances memory for contextual details that would otherwise not be remembered. The first goal of this work is to describe the development of my animal model of flashbulb memory, including a discussion of the importance of the timing between an emotional event and incidental, contextual cues. The second goal is to address the time-dependent neuroendocrine processes involved in stress-induced memory enhancement in rats. The involvement of brain structures, namely the hippocampus and amygdala, and hormones, including corticosterone and epinephrine, that interact to produce a composite memory of the contextual cues occurring in close temporal proximity to an emotional event are discussed. The results of Experiment 1 validate the animal model of flashbulb memory whereby an emotional event (predator exposure) produced memory for context cues that, under control conditions, would be forgotten. This memory enhancement only occurred when the emotional event was close in temporal proximity to training in the task. Experiment 2 provided evidence that epinephrine administration close in time to training mimicked the context memory formation induced by brief predator exposure, while propranolol, a β-adrenergic antagonist, as well as CPP, an NMDA receptor antagonist, blocked this effect. The results of Experiment 3 revealed that propranolol, CPP, and dexamethasone also blocked the brief predator stress-induced context memory formation. The results of Experiment 4 revealed that cannulated animals infused with aCSF (control) did not show evidence of predator stress-induced memory, therefore methodological issues within this experiment are addressed. Finally, the results of Experiment 5 revealed that adrenalectomy eliminated the predator stress-induced context memory compared to sham operated animals, suggesting that endogenous stress hormones are required for stress-induced context memory formation. Further, adrenalectomized rats supplemented with epinephrine before training did show evidence of context memory enhancement suggesting that epinephrine eliminated the memory impairment produced by adrenalectomy, and was sufficient to enhance memory in the absence of corticosterone. Overall this approach has provided insight into the time-dependent neuroendocrine processes involved in the formation of flashbulb, and potentially traumatic, memories in people.

The mesocorticolimbic system is involved in fundamental processes that drive motivational behaviors essential for survival (feeding, reproduction and sexual behavior, etc.), as well as neurochemical activity involved in mood regulation. Stressful life events are an important cause of dysregulated psychological functioning, which in some leads to a pathophysiology of mood disorders. A source of such disorder could be, among other underlying factors, an impairment of synaptic plasticity induced by alterations in the levels of neurotrophins and/or aberrant glucocorticoid responses. The role of the brain derived neurotrophic factor (BDNF) and its high affinity receptor tyrosine-related kinase B (TrkB) in the mesocorticolimbic reward circuitry has been largely studied in adulthood, yet a possible role of this system in mediating memory and emotional responses induced by stress during the juvenile, adolescence period has not been elucidated. The proposed set of thesis studies are designed to investigate the roles of BDNF and TrkB signaling, via the selective and non-competitive TrkB antagonist, ANA-12 (N-[2-[[(Hexahydro-2-oxo-1H-azepin-3-yl)amino]carbonyl]phenyl] benzo[b]thiophene-2-carboxamide), in the expression of stress-induced changes in the brain stress circuitry (including the medial prefrontal cortex (mPFC), hypothalamic-pituitary-adrenal (HPA) axis, and hippocampus) and reward signaling systems of the brain (including the nucleus accumbens (NAc) and ventral tegmental area (VTA)). In addition, experiments aim to determine behavioral changes following stress exposure in male and female Wistar rats. Finally, the possible interplay between BDNF, dopamine, glutamate and orexins in response to repeated stress is examined. Articles 1 and 2, aimed to assess the biochemical and behavioral effects of direct ANA-12 infusion (0.25 µg/ 0.5µl) into the nucleus accumbens shell during exposure to a 10-day heterotypic stress paradigm in male rats. Specifically, Article 1 demonstrated a key role for BDNF/TrkB signaling to regulate stress-induced effects. Notably, the impact of ANA-12 to attenuate anxiety-like behavior in repeatedly stressed rats while increasing anxiety behavior in non-stress rats suggest an interesting behavioral and neurochemical state-dependent process induced by TrkB receptor signaling. Article 2 supports the key role for BDNF secretion in basal and stress-induced behaviors in rats suggesting an influence of TrkB in sociability, motivation and passive avoidance. Furthermore, this role of TrkB extended to increased expression of orexin A in the Perifornical area (PfA) and a decrease in the ventral CA1 of the hippocampus, and in stress-induced elevations in orexinergic projections to the VTA, of which reductions were observed in non-stress groups treated with ANA-12. Article 3 demonstrated gender-specific behavioral and biochemical responses in different developmental periods and the impact of TrkB activation, dependent on stress exposure, to affect the regulation of TrkB receptor isoforms (full length and truncated TrkB, TrkB.FL and TrkB.T1, respectively) in adulthood. Results revealed increased CORT responses in adolescent females relative to males and attenuated CORT secretions in both genders by TrkB inhibition. Elevated activity levels in young adult females and increased passive coping behavior in the forced swim in stress-naïve females were also noted, in addition to novel observations on brain region and sex differences in TrkB receptor isoforms. Taken together, thesis findings derived from applications of ANA-12, shall foster knowledge on the contribution of BDNF in regulation of mood upon stress exposure at times when the brain is undergoing important maturation and remodelling, as well as on the relationship of stress exposure during adolescence and lasting brain and behavioral disorders in adulthood.

The 3Rs of reduction, refinement and replacement are the guiding principles of animal research and embedded in national and international legislation regulating the use of animals in scientific procedures. Awake imaging by MRI of rodents can offer a reduction by increasing the quality of scientific data through longitudinal imaging using less animals by avoiding a serial sacrifice design and refinement through reducing the stressful effects animals are exposed to, in comparison to existing models. Before awake imaging can become an established biomarker it must be demonstrated that pathology is traceable with comparable or an improvement on results using existing biomarkers. To validate awake imaging of rodents three study types were conducted in two different rodent species: imaging of the progression of Aspergillus fumigatus infection in the mouse lung using anaesthetized animals; analysis of stress in rats during imaging and imaging restraint; imaging of the abdomen in awake rats and a prospective study into the utility of this method for imaging the progression of Candida albicans renal infection. The first study type used an established model of invasive pulmonary aspergillosis in mice to test the utility of MRI for tracking infection in the lung parenchyma. Images of collapsed parenchyma were obtained and shown to increase as the infection progressed. Further work is required to establish this as a clinically relevant biomarker. The study type used restraint, blood collection and imaging as stressors using corticosterone levels as a surrogate of stress. Analysis of levels in blood and faeces by RIA and ELISA allowed comparison of stress during anaethetised and awake imaging for the first time. There were no differences in rat corticosterone levels during anaethetised and awake imaging indicating that awake imaging was no more stressful than currently used procedures. The third study type employed restrainers and acclimatisation to MRI scanner noise to acclimatize rats for awake abdomen imaging. Both anaethetised and awake rats were imaged with FLASH and IntraGate™ sequences. These methods were utilized in an established model of disseminated candidiasis by imaging the kidney. Comparable image quality was obtained in awake animals, with the utility of the method validated by imaging differences in renal pathology between vehicle and low and high dose treated animals. In conclusion, the first steps have been taken towards establishing awake animal imaging by MRI. The imaging is no more stressful than using an anaesthetic and was a useful biomarker in the rat abdomen and capable of tracking disease development. Further work is required to make the technique fully quantitative and automated and hence become a useful tool for monitoring progression of fungal infection and other pathology.

Orientadores: Regina Celia Spadari-Bratfisch, Dora Maria Grassi-Kassisse / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-07T15:56:51Z (GMT). No. of bitstreams: 1 Hatore_EdgarTeruhiko_M.pdf: 10729043 bytes, checksum: a8e99d57d617133d73cabb8f544052b9 (MD5) Previous issue date: 2006 / Resumo: Os efeitos do estresse por natação sobre marcadores metabólicos diferem daqueles do estresse por choques nas patas. Estas diferenças poderiam ser conseqüências da natureza do estressor, que difere nos dois modelos, ou da intensidade de estresse, que parece ser menor quando os animais são submetidos à natação do que quando submetidos a choque nas patas. O objetivo deste trabalho é avaliar o efeito da intensidade do estressor sobre a mobilização de substratos metabólicos. Ratos Wistar adultos foram distribuídos em grupos: controle, não submetido à natação; e ratos submetidos à natação em água a 35°C, 24°C ou 18°C. As sessões de natação ocorreram em três dias consecutivos com 5, 15 e 15 min de duração, respectivamente. Os ratos foram pesados antes da primeira e da última sessão de natação e após a última sessão sua temperatura retal foi determinada por um tele-termômetro; foram anestesiados e uma amostra de sangue foi coletada por punção cardíaca. Foram retiradas amostras do fígado e dos músculos gastrocnêmio, sóleo e ventricular cardíaco. Adipócitos foram isolados do tecido adiposo epididimal e incubados com agonistas adrenérgicos. O glicerol e o lactato liberados no meio infranadante foram considerados como indicadores da sensibilidade das respostas lipolítica e glicolítica aos agonistas, respectivamente. Foram determinadas também as concentrações plasmáticas de glicose, lactato, ácidos graxos livres, glicerol e corticosterona, além da concentração muscular e hepática de glicogênio. O peso corporal dos ratos dos quatro grupos não apresentou diferença estatisticamente significativa antes da primeira ou da terceira sessão de natação. Após natação em temperaturas de 24°C e 18°C os ratos apresentaram hipotermia (30,7 +/- 0,7°C e 24,0 +/- 0,5°C, respectivamente) em relação os grupos controle (37,2 +/- O,12°C) e natação a 35°C(36,7 +/- 0,2°C), entre os quais não houve diferença estatisticamente significativa. Após a terceira sessão de natação, a concentração plasmática de corticosterona aumentou significativamente nos grupos submetidos à natação em 18°C (294,7 +/-17,9 ng/mL) e 24°C (289,5 +/- 27,1 ng/mL) do que 35°C (166,3 +/- 11,8 ng/mL), sendo os três significativamente diferentes do controle (57,2 +/- 10,0 ng/mL), enquanto que as concentrações sanguíneas de lactato e de ácidos graxos livres estavam significativamente elevadas em relação ao controle, mas não diferiram entre os grupos de ratos submetidos à natação. As concentrações de glicogênio das amostras hepáticas, cardíacas e do músculo sóleo de ratos submetidos à natação não diferiram significantemente do grupo controle. Porém, as concentrações de glicogênio nas porções branca e vermelha do músculo gastrocnêmio estavam reduzidas após natação. Em adipócitos isolados de ratos submetidos à natação houve aumento da liberação basal de glicerol e de lactato, mas as respostas ao d-butiril-AMPc, à noradrenalina e à adrenalina, assim como a sensibilidade às duas catecolaminas,não diferiu significantemente do controle. Concluímos que a intensidade do agente estressor pode ser modulada pela temperatura da água em protocolos de estresse por natação, de modo que as temperaturas mais baixas em relação à temperatura corporal do rato determinam respostas de estresse mais intensas, evidenciadas pela concentração plasmática de corticosterona. Entretanto, as alterações observadas nos marcadores metabólicos analisados são independentes da intensidade do estressor, no paradigma de estresse utilizado neste trabalho / Abstract: The effects of swimming stress on metabolic markers are different than those induced by foot-shock stress. These differences might be due either to stress intensity, which seems to be minor when the rats are submitted to swimming than foot-shock stress, or to the type of stressor agent. The aim of this work is to evaluate the effect of the stressor intensity on metabolic markers. Adult Wistar rats were distributed in groups: control, not submitted to swimming; or swimming stressed rats with the water temperature at 35°C, 24°C or 18°C. Swimming sessions occurred daily, for 3 days, with 5, 15 and 15 min duration, respectively. Before the first and last sessions the rats were weight and after the last swimming session the rat's rectal temperature was determined using a tele-thermometer; then the rats were anesthetized and a blood sample was collected through cardiac puncture. Samples of the liver, as well as the gastrocnemius, soleus and cardiac muscles were also collected. Adipocytes were isolated from the epididimal adipose tissue and were incubated with adrenergic agonists: The glycerol and lactate released in the infranatant were considered as indicative of in vitro lipolytic and glycolytic sensitivity to the agonists,respectively. The plasma concentrations of glucose, lactate, free fat acids, glycerol and corticosterone, as well as the muscular and hepatic glycogen concentrations were determined. The results have shown that the rat' s body weights were not altered by swimming. After swimming at 24 C and 18 C the rats showed severe hypothermia (30.7+/-0.7°C and 24.0+/- 0.5 C, respectively, p<0.05), whereas swimming at 35C did not alter the rats rectal temperature (36.7 +/-0.2°C) compared to control (37.2 +/- 0.12C). The plasma levels of corticosterone significantly enhanced in 18°C (294.7 +/- 17.9 ng/mL), 24°C (289.5+/- 27.1 ng/mL) and 35°C (166.3 +/-11.8 ng/mL) swimming groups, compared to control (57.2 +/- 10.0 ng/mL). The glucose and glycerol plasma levels were not altered, whereas the blood lactate and free fat acids levels significantly increased after swimming, with no difference between the stressed groups. Swimming did not alter the glycogen concentrations of the liver as well as cardiac and soleus muscle, but caused a decrease in the white and red gastrocnemius compared to control, with the decrease being independent of the water temperature. The in vitro basal glycerol and lactate release by adipocytes increased, similarly in all swimming groups. The lipolytic response to d-butiryl cAMP,norepinephrine and epinephrine were not different between groups. Catecholamines determined a biphasic effect on lactate release by adipocytes with the concentrations in the range up to 10 nM being stimulant and those higher than 10 nM being inhibitory of lactate release. No differences were observed between groups. We conclude that the stress intensity might be modulated by the water temperature in protocols of swimming stress, with lower temperatures being more stressful than those similar to the rats body temperature, as determined by the plasma levels of corticosterone. We also conclude that the swimming stress paradigm altered the analyzed metabolic markers in a manner that was independent of the stressor intensity (homotypic), in the stress paradigm used in this work / Mestrado / Fisiologia / Mestre em Biologia Funcional e Molecular

Stress cannot be fully avoided in an animal´s life, and the way that animals handle stress can be crucial for their health. Coping styles in animals are describing consistent sets of behaviour and physiological stress responses. Two forms of coping are typically described; a more proactive type that is more bold, active, and aggressive, and, in contrast, a less active, more shy reactive type. Coping styles can be used to learn how animals react to stress, and thus improve our handling of stress in captive animals. To investigate the existence of the reactive-proactive coping styles in the red junglefowl (Gallus gallus), both male and female red junglefowl were exposed to behavioural assays measuring boldness, activity, exploration, and aggression. These behavioural measures were compared to blood plasma concentrations of the stress hormone corticosterone, hence exploring the link between behaviour and physiological response to stress. I show positive correlations between boldness, activity, and exploration measured in a novel arena test, and boldness and activity measured in a novel object test. This somewhat confirms the inter-related behavioural aspect of reactive-proactive coping styles. Further, some of these behaviours correlated with post-stress concentrations of corticosterone, which somewhat links behaviour to the physiological aspect of coping styles. This study tentatively supports the reactive-proactive coping styles by showing a link between behaviours and the physiological stress response in the red junglefowl, with some similarities to what have been observed in other species across taxa.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第23467号 / 医博第4774号 / 新制||医||1053(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 森信 暁雄, 教授 上野 英樹, 教授 小柳 義夫 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

No description available.

The benefits of intravenous therapy have become more and more apparent over the years. Medications can be given rapidly with an expectant rapid onset of action. The response to the drugs or fluids can often be closely controlled by regulating the dose or rate of administration. Frequently, adequate blood and tissue levels needed to eradicate many serious infections can be reached only by this route. Intravenous therapy is an especially appropriate method when the use of the oral tract, for one reason or another, cannot be used. The development of intravenous therapy, however, did not proceed without its difficulties. Problems of allergic reactions, incompatible blood groups, bacterial contamination, particulate matter, thrombophlebitic syndromes, stability of solutions, and incompatibilities of admixtures soon became apparent. The purpose of this paper is to explore certain aspects of the latter problem, i.e., intravenous incompatibilities.

Sexual selection theory tries to explain evolution of apparently useless traits which mainly developed in males of numerous species. One such trait is also rich and vibrant coloration, typical for many of the bird species. These traits are difficult to be falsified, and therefore they honestly signal quality of the individual. Carotenoid coloration reflects the health condition and melanin coloration the social status, even though this traditional division might not be as strict according to the latest studies. Apart from these ways of maintaining honesty, recent studies are focusing more also on the effect of hormones, mainly two steroids: male sexual hormone - testosterone -, and the "stress" hormone - corticosterone. Both hormones could positively influence male's sexual traits such as ornamental coloration. On the other hand, elevated levels of these hormones possess risk to the organism (higher energetic expenditure, chronic stress), therefore also might potentially become costly. Using photographs of birds in standardized conditions as well as spectrophotometry I analyzed the plumage coloration of males of the Yellowhammer (Emberiza citrinella). Concentrations of testosterone and corticosterone deposited in feathers were analyzed using the LC-MS/MS. Also, for males in breeding season 2015,...