Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 87-99).

Orientador: Fabio Papes / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-27T05:41:39Z (GMT). No. of bitstreams: 1 Souza_MateusAugustodeAndrade_M.pdf: 6121419 bytes, checksum: a603ea19d560e8cfebddccca9b7d824a (MD5) Previous issue date: 2015 / Resumo: Em mamíferos, os neurônios sensoriais do Sistema Olfatório (OSNs) se encontram no interior da cavidade nasal, mas estão diretamente expostos ao ambiente externo. Por um lado, tal localização permite a esses neurônios o acesso imediato aos estímulos químicos ambientais, tomando vantagem do fluxo respiratório. Por outro lado, esses neurônios estão constantemente sujeitos a injúrias por agentes nocivos, como toxinas e patógenos, capazes de destruir essas células sensoriais. Sua perda constante, contudo, é contrabalanceada pela geração de novos OSNs durante toda a vida do indivíduo, fato que torna o Sistema Olfatório um dos poucos locais do organismo com neurogênese contínua na idade adulta. A regeneração dos OSNs tem atraído a atenção da comunidade científica tanto pelo seu potencial uso como modelo de estudo do Sistema Nervoso quanto pela sua potencial aplicação para o tratamento de doenças neurodegenerativas. Nesse sentido, muito conhecimento já foi produzido sobre a dinâmica de fatores de transcrição que acompanha a diferenciação dos progenitores neuronais olfatórios em OSNs. Porém, uma grande lacuna no conhecimento diz respeito a outros elementos capazes de coordenar esse processo, como os fatores moduladores da cromatina. Diante desse cenário, escolhemos como objeto de estudo as proteínas do Grupo Polycomb (PcG), que constituem uma maquinaria de controle transcricional relacionada a modificações na organização da cromatina. Neste trabalho, genes PcG selecionados foram caracterizados molecular e funcionalmente no epitélio olfatório principal de camundongos (MOE). Através de ensaios de hibridação in situ, cinco dos seis genes avaliados apresentaram expressão ubíqua por todo o epitélio (Cbx2, Cbx4, Phc2, Ezh1, Bcl6), enquanto um (Ezh2) mostrou-se expresso somente nos estratos basais do MOE. Em ensaios de colocalização, provamos que Ezh2 é expresso exclusivamente nos progenitores olfatórios, onde o processo de diferenciação se inicia, e em parte dos OSNs recém-diferenciados, ainda não funcionais. Esta foi a primeira vez que a expressão de um gene PcG foi analisada detalhadamente no Sistema Olfatório. O interessante perfil de expressão de Ezh2 foi sugestivo de um possível papel funcional relacionado à diferenciação dos progenitores olfatórios. Para investigar essa hipótese, utilizamos como ferramenta experimental a habilidade do MOE em se regenerar após a indução de injúrias específicas. Para isso, o MOE de camundongos foi lesionado quimicamente com o composto diclobenil, que leva à perda abrupta de OSNs, estimulando a proliferação e a diferenciação dos progenitores olfatórios para repovoar as regiões lesionadas. Os animais assim tratados receberam, por via intranasal, o fármaco GSK126, uma molécula inibidora específica da atividade da proteína EZH2. Acompanhando a regeneração subsequente do MOE, observamos que a inibição da atividade de EZH2 levou ao incremento de OSNs no epitélio, favorecendo a sua regeneração. Interessantemente, esse incremento também foi observado em MOEs não lesionados, mostrando que o efeito de GSK126 não é dependente da indução de injúrias prévias. Através dessa investigação molecular e funcional, buscamos contribuir para o melhor entendimento da diferenciação neuronal do MOE, e apontamos as proteína PcG como elementos importantes para esse processo / Abstract: In mammals, the olfactory sensory neurons (OSNs) are located inside the nasal cavity, but they are directly exposed to the external environment. Taking advantage of the respiratory flux, this location favors the access to the chemical stimuli presented by the environment. On the other hand, it leads OSNs to be continually damaged by pathogens and toxic substances carried by the inhaled air. However, the persistence of neuronal progenitors in the olfactory epithelium makes the constant reposition of the OSNs possible. This unique ability of regeneration makes the Olfactory System one of the few sites of neurogenesis throughout the adult life. Olfactory regeneration has attracted the attention scientific community because of its potential as a model of study of the Nervous System and application in the treatment of neurodegenerative diseases. A great amount of knowledge has been accumulated about the transcription factor dynamics that follows the differentiation of neuronal progenitors into OSNs. However, there is a great gap about other elements that could coordinate this process, such as chromatin modulator factors. In this scenario, we decided to study the Polycomb Group (PcG) proteins, a transcription control machinery involved in chromatin structure organization. In the present study, selected PcG genes were molecular and functionally analyzed in the mouse main olfactory epithelium (MOE). Using in situ hybridization assays, we characterized the expression of six PcG genes. Five of them were shown to be expressed throughout the MOE (Cbx2, Cbx4, Phc2, Ezh1, Bcl6), while one (Ezh2) was found only in the basal layers of this epithelium. Using colocalization strategies, we proved that Ezh2 gene is expressed exclusively in the olfactory progenitor cells, where the differentiation process begins, and in part of the newly differentiated OSNs that are still not functional. It was the first time that a PcG gene expression profile was finely analyzed in the Olfactory System. This interesting expression profile presented by Ezh2 suggested a possible involvement with the MOE neuronal progenitor differentiation. For this functional investigation, we used MOE¿s neuronal regeneration after specific injuries as an experimental tool. For this purpose, the MOE was chemically damaged by the compound dichlobenil, which causes a great loss of OSNs, stimulating proliferation and differentiation of neuronal progenitor cells, leading to the repopulation of the damaged tissue. Next, mice received by intranasal route the pharmacological inhibitor GSK126, which blocks EZH2 protein activity. The observation of the MOE regeneration that followed showed us that GSK126 application resulted in an increased number of OSNs, improving MOE regeneration. Interestingly, this increase was also found in intact MOEs, pointing that GSK126¿s effects do not depend on previous olfactory injuries. By this molecular and functional investigation, we aimed at a better understanding of olfactory neuronal differentiation, and we targeted the PcG proteins as relevant elements to this process / Mestrado / Genetica Animal e Evolução / Mestre em Genética e Biologia Molecular

Objective: Decrease of olfactory function in Parkinson's disease (PD) is a well-investigated fact. Studies indicate that pharmacological treatment of PD fails to restore olfactory function in PD patients. The aim of this investigation was whether patients with PD would benefit from “training” with odors in terms of an improvement of their general olfactory function. It has been hypothesized that olfactory training should produce both an improved sensitivity towards the odors used in the training process and an overall increase of olfactory function. Methods: We recruited 70 subjects with PD and olfactory loss into this single-center, prospective, controlled non-blinded study. Thirty-five patients were assigned to the olfactory training group and 35 subjects to the control group (no training). Olfactory training was performed over a period of 12 weeks while patients exposed themselves twice daily to four odors (phenyl ethyl alcohol: rose, eucalyptol: eucalyptus, citronellal: lemon, and eugenol: cloves). Olfactory testing was performed before and after training using the “Sniffin' Sticks” (thresholds for phenyl ethyl alcohol, tests for odor discrimination, and odor identification) in addition to threshold tests for the odors used in the training process. Results: Compared to baseline, trained PD patients experienced a significant increase in their olfactory function, which was observed for the Sniffin' Sticks test score and for thresholds for the odors used in the training process. Olfactory function was unchanged in PD patients who did not perform olfactory training. Conclusion: The present results indicate that olfactory training may increase olfactory sensitivity in PD patients.

Diss. (sammanfattning) Umeå : Univ., 2007. / Härtill 4 uppsatser.

Orientador: Fabio Papes / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-24T14:16:00Z (GMT). No. of bitstreams: 1 Tolentino_FelipeThadeu_M.pdf: 9244448 bytes, checksum: deea9f7963e05d8a997d9b5a554f9708 (MD5) Previous issue date: 2014 / Resumo: O Sistema Sensorial Olfativo de mamíferos é composto por vários subsistemas na cavidade nasal. Dentre estes, destacam-se o sistema olfativo principal e o sistema olfativo acessório ou vomeronasal. O primeiro realiza a detecção geral de odores e parece participar também da detecção de algumas substâncias que levam a respostas comportamentais instintivas (feromônios), enquanto o último é especializado na detecção desta classe de semioquímicos. A detecção dos estímulos sensoriais olfativos resulta em informações importantes que dependem de vias complexas para sua interpretação e para a geração de respostas apropriadas por parte do sistema nervoso central. Existem vários pontos ainda desconhecidos sobre o funcionamento do sistema olfativo, tanto no que diz respeito aos mecanismos moleculares subjacentes à escolha dos receptores a serem expressos por um dado neurônio sensorial ¿ sendo que cada neurônio olfativo expressa apenas um receptor dentro de uma grande família multi-gênica ¿ quanto em relação ao processamento da informação sensorial em centros cerebrais superiores. Neurônios sensoriais olfativos cultivados eficientemente in vitro seriam extremamente úteis, pois poderiam ser utilizados como ferramenta para o estudo destes problemas, como a investigação da atividade das células sensoriais olfativas, possibilitando, por exemplo, uma melhor compreensão dos mecanismos genéticos e moleculares por trás da expressão dos receptores olfativos e de suas propriedades de detecção. Neste trabalho foram desenvolvidas ferramentas baseadas em vetores retrovirais com o objetivo de induzir a diferenciação celular de neurônios olfativos in vitro, utilizando uma combinação de fatores de transcrição, por meio de transdução viral em células-alvo (fibroblastos murinos). Os retrovírus produzidos foram testados e algumas combinações de fatores de transcrição foram preliminarmente testadas, sendo capazes de induzir mudanças moleculares em fibroblastos acompanhadas da expressão de marcadores de neurônios sensoriais olfativos / Abstract: The mammalian Olfactory System enables the vast majority of animal species to identify the presence and quality of food, predators, competitors, conspecifics and potential mates in the environment. Olfactory stimuli detected by sensory neurons are interpreted by brain processing pathways to generate appropriate behavioral and endocrine responses. Despite its central importance in mammalian physiology, several aspects about the biology of this sensory system remain uncharacterized. For example, it is known that each olfactory sensory neuron (OSN) in the nasal cavity expresses only one gene out of a large multi-gene family coding for receptors involved in odorant and pheromone detection. However, the molecular mechanisms behind this process of olfactory receptor gene choice are not fully understood. The study of this and many other aspects of olfaction has been made difficult by the lack of appropriate in vitro cellular models. An efficient way to obtain cultured OSNs would thus be extremely useful, enabling researchers to investigate the sensory neuron¿s activity in a controllable environment, avoiding obstacles imposed by the cellular heterogeneity found in sensory organs in vivo. In this study, we aimed at obtaining OSNs directly differentiated from mouse embryonic fibroblasts (MEF) using the forced expression of specific transcription factors via retroviral vectors. We therefore developed tools based on retroviral vectors with the objective of differentiating olfactory sensory neurons in vitro, using viral transduction in target cells (murine fibroblasts) with combinations of select transcription factors. Retroviruses were tested and some combinations of transcription factors were tested on a preliminary basis, which were capable of inducing molecular alterations on fibroblasts followed by the expression of olfactory sensory neuron markers / Mestrado / Genetica Animal e Evolução / Mestre em Genética e Biologia Molecular

Chez les abeilles domestiques, les neurones à récepteurs olfactifs hébergés dans les antennes sont des neurones sensoriels primaires responsables de la détection des odeurs et des phéromones. L'information olfactive est ensuite acheminée par les nerfs antennaires jusqu'aux lobes antennaires qui constituent le premier étage d'intégration de l'information olfactive. Les abeilles butineuses sont exposées aux insecticides, notamment ceux de la classe des pyréthrinoïdes, qui sont utilisés pour la protection des plantes et la lutte contre les insectes considérés comme étant nuisibles.Nous avons caractérisé l'effet des pyréthrinoïdes sur les canaux sodiques activés par le potentiel (responsables des potentiels d'action) dans les deux premiers étages du système olfactif de l'abeille. Nos enregistrements électrophysiologiques en mode potentiel imposé dans les neurones à récepteurs olfactifs mis en culture révèlent que l'effet des pyréthrinoïdes de type I et II (notamment la tétraméthrine et la deltaméthrine) est amplifié par une intensification de l'activité électrique neuronale. Cette amplification survient notamment via le démasquage de canaux sodiques silencieux que nous avons également mis en évidence avec la toxine d'anémone de mer ATX-II. Le niveau maximal de canaux sodiques modifiés est atteint en quelques centaines de millisecondes. Dans les neurones centraux des lobes antennaires, cette amplification apparait très limitée voire absente avec les pyréthrinoïdes mais elle peut toutefois survenir en présence de l'alcaloïde végétal vératridine. Par ailleurs, dans ces neurones centraux, les pyréthrinoïdes semblent être à l’origine d’une accélération de l'inactivation lente des canaux sodiques auparavant décrite en présence de certains anesthésiques locaux. Les modifications différentielles observées dans les neurones périphériques et centraux pourraient être responsables des effets délétères des pyréthrinoïdes sur les capacités de perception, d'orientation et d'apprentissage de l'abeille domestique. / In domestic honeybees, the olfactory receptor neurons localized in the antennae are primary sensory neurons responsible for the detection of odor and pheromone compounds. The olfactory information is further conveyed to the antennal lobes by the antennal nerves. The antennal lobes are the first stage of integration of the olfactory information. Forager bees are exposed to insecticides, especially pyrethroids that are used for plant protection and eradication of pests.In the honeybee olfactory pathway, we investigated the effects of pyrethroids on the voltage-gated sodium channels (which underlie action potentials). Our patch-clamp recordings in the antennal olfactory receptor neurons maintained in cell culture reveal that the effects of type I and type II pyrethroids (e.g. tetramethrin and deltamethrin) are increased by an augmentation of neuronal electrical activity. The amplification of the effects of pyrethroids occurs as a result of the unmasking of silent sodium channels that we have also shown evidence for, with sea anemone toxin ATX-II. The maximal sodium channels modification takes place within few hundreds of milliseconds. In the central antennal lobe neurons, that amplification is rather limited or absent with pyrethroids but the plant alkaloid veratridine is able to induce such an amplification. Furthermore, in the latter cell type, pyrethroids cause an acceleration of the sodium channels slow inactivation. Such an effect has been previously reported for some local anesthetics. The differential actions of pyrethroids that we have observed in the peripheral and central neurons may be responsible for the impairment of learning performance, perception and disorientation exhibited by pyrethroid-exposed honeybees.

No description available.

Background: Patients with Panic Disorder (PD) direct their attention towards potential threat, followed by panic attacks, and increased sweat production. Onés own anxiety sweat odor influences the attentional focus, and discrimination of threat or non-threat. Since olfactory projection areas overlap with neuronal areas of a panic-specific fear network, the present study investigated the neuronal processing of odors in general and of stress-related sweat odors in particular in patients with PD. Methods: A sample of 13 patients with PD with/ without agoraphobia and 13 age- and gender-matched healthy controls underwent an fMRI investigation during olfactory stimulation with their stress-related sweat odors (TSST, ergometry) as well as artificial odors (peach, artificial sweat) as non-fearful non-body odors. Principal Findings: The two groups did not differ with respect to their olfactory identification ability. Independent of the kind of odor, the patients with PD showed activations in fronto-cortical areas in contrast to the healthy controls who showed activations in olfaction-related areas such as the amygdalae and the hippocampus. For artificial odors, the patients with PD showed a decreased neuronal activation of the thalamus, the posterior cingulate cortex and the anterior cingulate cortex. Under the presentation of sweat odor caused by ergometric exercise, the patients with PD showed an increased activation in the superior temporal gyrus, the supramarginal gyrus, and the cingulate cortex which was positively correlated with the severity of the psychopathology. For the sweat odor from the anxiety condition, the patients with PD showed an increased activation in the gyrus frontalis inferior, which was positively correlated with the severity of the psychopathology. Conclusions: The results suggest altered neuronal processing of olfactory stimuli in PD. Both artificial odors and stress-related body odors activate specific parts of a fear-network which is associated with an increased severity of the psychopathology.

Objective: Decrease of olfactory function in Parkinson's disease (PD) is a well-investigated fact. Studies indicate that pharmacological treatment of PD fails to restore olfactory function in PD patients. The aim of this investigation was whether patients with PD would benefit from “training” with odors in terms of an improvement of their general olfactory function. It has been hypothesized that olfactory training should produce both an improved sensitivity towards the odors used in the training process and an overall increase of olfactory function. Methods: We recruited 70 subjects with PD and olfactory loss into this single-center, prospective, controlled non-blinded study. Thirty-five patients were assigned to the olfactory training group and 35 subjects to the control group (no training). Olfactory training was performed over a period of 12 weeks while patients exposed themselves twice daily to four odors (phenyl ethyl alcohol: rose, eucalyptol: eucalyptus, citronellal: lemon, and eugenol: cloves). Olfactory testing was performed before and after training using the “Sniffin' Sticks” (thresholds for phenyl ethyl alcohol, tests for odor discrimination, and odor identification) in addition to threshold tests for the odors used in the training process. Results: Compared to baseline, trained PD patients experienced a significant increase in their olfactory function, which was observed for the Sniffin' Sticks test score and for thresholds for the odors used in the training process. Olfactory function was unchanged in PD patients who did not perform olfactory training. Conclusion: The present results indicate that olfactory training may increase olfactory sensitivity in PD patients.

Background: Patients with Panic Disorder (PD) direct their attention towards potential threat, followed by panic attacks, and increased sweat production. Onés own anxiety sweat odor influences the attentional focus, and discrimination of threat or non-threat. Since olfactory projection areas overlap with neuronal areas of a panic-specific fear network, the present study investigated the neuronal processing of odors in general and of stress-related sweat odors in particular in patients with PD. Methods: A sample of 13 patients with PD with/ without agoraphobia and 13 age- and gender-matched healthy controls underwent an fMRI investigation during olfactory stimulation with their stress-related sweat odors (TSST, ergometry) as well as artificial odors (peach, artificial sweat) as non-fearful non-body odors. Principal Findings: The two groups did not differ with respect to their olfactory identification ability. Independent of the kind of odor, the patients with PD showed activations in fronto-cortical areas in contrast to the healthy controls who showed activations in olfaction-related areas such as the amygdalae and the hippocampus. For artificial odors, the patients with PD showed a decreased neuronal activation of the thalamus, the posterior cingulate cortex and the anterior cingulate cortex. Under the presentation of sweat odor caused by ergometric exercise, the patients with PD showed an increased activation in the superior temporal gyrus, the supramarginal gyrus, and the cingulate cortex which was positively correlated with the severity of the psychopathology. For the sweat odor from the anxiety condition, the patients with PD showed an increased activation in the gyrus frontalis inferior, which was positively correlated with the severity of the psychopathology. Conclusions: The results suggest altered neuronal processing of olfactory stimuli in PD. Both artificial odors and stress-related body odors activate specific parts of a fear-network which is associated with an increased severity of the psychopathology.