The aim of this thesis was to determine content of - a -bitter acids in samples of hop obtained from different types of growing regions. Object of research were cultivars of Saaz late, Sládek and Premiant cultivated in three growing regions (Žatecko, Úštěcko, Tršicko) in year 2015. Nine hop samples from conventional agriculture and one from bio production were examined. For all growing areas, from which hop cones were obtained, was determined altitude, rainfall and average temperatures during vegetation period. Quantity of bitter acids were measured by HPLC. Our calculated correlation coefficients showed that the higher altitude and average temperature the lower content of bitter acids was measured and more precipitation has positive impact on content of bitter acids.

A mandioca (Manihot esculenta Crantz) é uma das principais culturas do mundo, havendo grande variabilidade genética. As variedades são classificadas com base na palatabilidade e toxicidade das raízes, em mansas ou doces e bravas ou amargas. Apesar da importância, o potencial da mandioca é pouco explorado, não sendo conhecidos, em nível molecular, os elementos determinantes para as suas características. Assim, pretendeu-se identificar, empregando a 2D-PAGE, proteínas que possam estar associadas com as diferenças físico-químicas das raízes tuberosas de variedades de mesa (IAC 576-70 e IAC 06-01), indústria (Cigana Preta, IAC 12 e IAC 90) e de uso misto (Vassourinha Paulista). Após extração de proteínas e separação por 2D-PAGE, as imagens dos géis foram analisadas no programa Delta2D (DECODON), sendo realizada análise estatística utilizando-se ANOVA (p<0,01), Heat Map e Análises de Componentes Principais (ACP) e de Agrupamentos. Os 146 spots de interesse foram removidos dos géis e suas proteínas digeridas e sequenciadas por espectrometria de massas. Algumas proteínas refletiram as características fenotípicas das variedades em estudo, especialmente entre as de mesa e indústria. Pela ACP, foram explicados 54,54% da variabilidade entre as amostras. A primeira componente separou as variedades exclusivamente de mesa de todas as demais, enquanto a segunda separou a IAC 90 de todas as outras, sendo esta caracterizada por um perfil proteico diferente das demais amostras de uso industrial. A IAC 576-70 e a IAC 12 apresentaram alta correlação positiva, assim como, a Vassourinha e a Cigana. A Análise de Agrupamentos corroborou as informações da ACP, revelando que o proteoma das raízes tuberosas refletiu diferenças fenotípicas entre as variedades. / Cassava (Manihot esculenta Crantz) is a main crop with large genetic variability. The varieties are classified according palatability and toxicity of the roots as sweet or bitter cassavas. Despite its importance, little is known about the molecular basis of phenotypic characteristics. Therefore, this study aimed to identify proteins associated to the differences between the sweet (\'IAC 576-70\' e \'IAC 06-01\'), bitter (\'Cigana Preta\', \'IAC 12\' e \'IAC 90\') and the mixed-use (\'Vassourinha Paulista\') varieties by 2D-PAGE. After the protein extraction and separation by 2D-PAGE, the gel images were analyzed through the software Delta 2D (DECODON), and the statistical analysis were performed with ANOVA (p<0,01), Heat Map, Principal Component Analysis (PCA) and Cluster Analysis. The 146 significant spots were removed from the gels, digested and sequenced by mass spectrometry. Some proteins were related to the physico-chemical characteristics of the varieties, especially between the sweet and the bitter. Variability of the samples was explained at the level of 54,54% by the PCA. The first component separated the sweet varieties from all others while the second one separated the \'IAC 90\' from all others. This variety was characterized by a different protein profile among the bitter cassavas. The \'IAC 576-70\' and the \'IAC 12\' were positively correlated, as well as, \'Vassourinha\' and the \'Cigana\'. Cluster Analysis agreed the PCA information, revealing that the proteomes of the tuberous roots reflected phenotypic differences among the varieties.

Sensory and consumer sciences aim to understand the influences of product acceptability and purchase decisions. The food industry measures product acceptability through hedonic testing but often does not assess implicit or qualitative response. Incorporation of qualitative research and automated facial expression analysis (AFEA) may supplement hedonic acceptability testing to provide product insights. The purpose of this research was to assess the application of AFEA and qualitative analysis to understand consumer experience and response. In two studies, AFEA was applied to elucidate consumers emotional response to dairy (n=42) and water (n=46) beverages. For dairy, unflavored milk (x=6.6±1.8) and vanilla syrup flavored milk (x=5.9±2.2) (p>0.05) were acceptably rated (1=dislike extremely; 9=like extremely) while salty flavored milk (x=2.3±1.3) was least acceptable (p<0.05). Vanilla syrup flavored milk generated emotions with surprised intermittently present over time (10 sec) (p<0.025) compared to unflavored milk. Salty flavored milk created an intense disgust response among other emotions compared to unflavored milk (p<0.025). Using a bitter solutions model in water, an inverse relationship existed with acceptability as bitter intensity increased (rs=-0.90; p<0.0001). Facial expressions characterized as disgust and happy emotion increased in duration as bitter intensity increased while neutral remained similar across bitter intensities compared to the control (p<0.025). In a mixed methods analysis to enumerate microbial populations, assess water quality, and qualitatively gain consumer insights regarding water fountains and water filling stations, results inferred that water quality differences did not exist between water fountains and water filling stations (metals, pH, chlorine, and microbial) (p>0.05). However, the exterior of water fountains were microbially (8.8 CFU/cm^2) and visually cleaner than filling stations (10.4x10^3 CFU/cm^2) (p<0.05). Qualitative analysis contradicted quantitative findings as participants preferred water filling stations because they felt they were cleaner and delivered higher quality water. Lastly, The Theory of Planned Behavior was able to assist in understanding undergraduates' reusable water bottle behavior and revealed 11 categories (attitudes n=6; subjective norms n=2; perceived behavioral control n=2; intentions n=1). Collectively, the use of AFEA and qualitative analysis provided additional insight to consumer-product interaction and acceptability; however, additional research should include improving the sensitivity of AFEA to consumer product evaluation. / Ph. D.

Intensive Forschung hat in den vergangenen Jahrzehnten zu einer sehr detaillierten Charakterisierung des Geschmackssystems der Säugetiere geführt. Dennoch sind mit den bislang eingesetzten Methoden wichtige Fragestellungen unbeantwortet geblieben. Eine dieser Fragen gilt der Unterscheidung von Bitterstoffen. Die Zahl der Substanzen, die für den Menschen bitter schmecken und in Tieren angeborenes Aversionsverhalten auslösen, geht in die Tausende. Diese Substanzen sind sowohl von der chemischen Struktur als auch von ihrer Wirkung auf den Organismus sehr verschieden. Während viele Bitterstoffe potente Gifte darstellen, sind andere in den Mengen, die mit der Nahrung aufgenommen werden, harmlos oder haben sogar positive Effekte auf den Körper. Zwischen diesen Gruppen unterscheiden zu können, wäre für ein Tier von Vorteil. Ein solcher Mechanismus ist jedoch bei Säugetieren nicht bekannt. Das Ziel dieser Arbeit war die Untersuchung der Verarbeitung von Geschmacksinformation in der ersten Station der Geschmacksbahn im Mausgehirn, dem Nucleus tractus solitarii (NTS), mit besonderem Augenmerk auf der Frage nach der Diskriminierung verschiedener Bitterstoffe. Zu diesem Zweck wurde eine neue Untersuchungsmethode für das Geschmackssystem etabliert, die die Nachteile bereits verfügbarer Methoden umgeht und ihre Vorteile kombiniert. Die Arc-catFISH-Methode (cellular compartment analysis of temporal activity by fluorescent in situ hybridization), die die Charakterisierung der Antwort großer Neuronengruppen auf zwei Stimuli erlaubt, wurde zur Untersuchung geschmacksverarbeitender Zellen im NTS angewandt. Im Zuge dieses Projekts wurde erstmals eine stimulusinduzierte Arc-Expression im NTS gezeigt. Die ersten Ergebnisse offenbarten, dass die Arc-Expression im NTS spezifisch nach Stimulation mit Bitterstoffen auftritt und sich die Arc exprimierenden Neurone vornehmlich im gustatorischen Teil des NTS befinden. Dies weist darauf hin, dass Arc-Expression ein Marker für bitterverarbeitende gustatorische Neurone im NTS ist. Nach zweimaliger Stimulation mit Bittersubstanzen konnten überlappende, aber verschiedene Populationen von Neuronen beobachtet werden, die unterschiedlich auf die drei verwendeten Bittersubstanzen Cycloheximid, Chininhydrochlorid und Cucurbitacin I reagierten. Diese Neurone sind vermutlich an der Steuerung von Abwehrreflexen beteiligt und könnten so die Grundlage für divergentes Verhalten gegenüber verschiedenen Bitterstoffen bilden. / Intense research in the past decades has led to a detailed understanding of the mammalian taste system. Some important issues, however, have remained unanswered with the established methods that have been applied so far. One of these questions is whether different bitter substances can be distinguished. There are thousands of compounds which taste bitter to humans and elicit innate aversive behavior in animals. Moreover, these bitter substances are very heterogeneous regarding their structure as well as their effect on the organism. While many bitter tastants are potent poisons, others are harmless or even have beneficial effects in the amounts that are typically ingested. The ability to discriminate between those groups of bitter tastants could be an evolutionary advantage. Such a mechanism, however, is not known for mammals. The aim of this thesis was to study the processing of taste information in the first station of gustatory processing in the mouse brain, the nucleus of the solitary tract (NTS). Of particular interest was the question concerning discrimination of bitter tastants. To this end a new method was established for the taste system combining the advantages of methods used before while circumventing their disadvantages. The Arc catFISH method (cellular compartment analysis of temporal activity by fluorescent in situ hybridization), which allows the characterization of responses of large neuron populations to two stimuli, was used to analyze taste-processing cells in the NTS. In the course of this project a stimulus-induced Arc expression in the NTS was shown for the first time. The results demonstrated that Arc expression in the NTS appears specifically after stimulation with bitter tastants and that the Arc expressing neurons are located primarily in the gustatory part of the NTS. This indicates that Arc expression is a marker for bitter-processing gustatory neurons in the NTS. Upon stimulating twice with bitter compounds, distinct, yet overlapping neuron populations were identified, that reacted differently to the three bitter substances cycloheximide, quinine hydrochloride, and cucurbitacin I. Presumably these neurons are involved in the regulation of aversive reflexes and could form a basis for divergent behavior towards different bitter substances.

Die Wahrnehmung von Geschmacksempfindungen beruht auf dem Zusammenspiel verschiedener Sinneseindrücke wie Schmecken, Riechen und Tasten. Diese Komplexität der gustatorischen Wahrnehmung erschwert die Beantwortung der Frage wie Geschmacksinformationen vom Mund ins Gehirn weitergeleitet, prozessiert und kodiert werden. Die Analysen zur neuronalen Prozessierung von Geschmacksinformationen erfolgten zumeist mit Bitterstimuli am Mausmodell. Zwar ist bekannt, dass das Genom der Maus für 35 funktionelle Bitterrezeptoren kodiert, jedoch war nur für zwei unter ihnen ein Ligand ermittelt worden. Um eine bessere Grundlage für tierexperimentelle Arbeiten zu schaffen, wurden 16 der 35 Bitterrezeptoren der Maus heterolog in HEK293T-Zellen exprimiert und in Calcium-Imaging-Experimenten funktionell charakterisiert. Die Daten belegen, dass das Funktionsspektrum der Bitterrezeptoren der Maus im Vergleich zum Menschen enger ist und widerlegen damit die Aussage, dass humane und murine orthologe Rezeptoren durch das gleiche Ligandenspektrum angesprochen werden. Die Interpretation von tierexperimentellen Daten und die Übertragbarkeit auf den Menschen werden folglich nicht nur durch die Komplexität des Geschmacks, sondern auch durch Speziesunterschiede verkompliziert. Die Komplexität des Geschmacks beruht u. a. auf der Tatsache, dass Geschmacksstoffe selten isoliert auftreten und daher eine Vielzahl an Informationen kodiert werden muss. Um solche geschmacksstoffassoziierten Stimuli in der Analyse der gustatorischen Kommunikationsbahnen auszuschließen, sollten Opsine, die durch Licht spezifischer Wellenlänge angeregt werden können, für die selektive Ersetzung von Geschmacksrezeptoren genutzt werden. Um die Funktionalität dieser angestrebten Knockout-Knockin-Modelle zu evaluieren, die eine Kopplung von Opsinen mit dem geschmacksspezifischen G-Protein Gustducin voraussetzte, wurden Oozyten vom Krallenfrosch Xenopus laevis mit dem Zwei-Elektroden-Spannungsklemm-Verfahren hinsichtlich dieser Interaktion analysiert. Der positiven Bewertung dieser Kopplung folgte die Erzeugung von drei Mauslinien, die in der kodierenden Region eines spezifischen Geschmacksrezeptors (Tas1r1, Tas1r2, Tas2r114) Photorezeptoren exprimierten. Durch RT-PCR-, In-situ-Hybridisierungs- und immunhistochemische Experimente konnte der erfolgreiche Knockout der Rezeptorgene und der Knockin der Opsine belegt werden. Der Nachweis der Funktionalität der Opsine im gustatorischen System wird Gegenstand zukünftiger Analysen sein. Bei erfolgreichem Beleg der Lichtempfindlichkeit von Geschmacksrezeptorzellen dieser Mausmodelle wäre ein System geschaffen, dass es ermöglichen würde, gustatorische neuronale Netzwerke und Hirnareale zu identifizieren, die auf einen reinen geschmacks- und qualitätsspezifischen Stimulus zurückzuführen wären. / Taste impression is based on the interaction of taste, smell and touch. To evaluate the nutritious content of food mammals possess five distinct taste qualities: sweet, bitter, umami (taste of amino acids), sour and salty. For bitter, sweet, and umami compounds taste signaling is initiated by binding of tastants to G protein-coupled receptors. The interactions of taste stimuli, usually watersoluble chemicals, with their cognate receptors lead to the activation of the G protein gustducin, which, in turn, initiates a signal resulting in the activation of gustatory afferents. However, details of gustatory signal transmission and processing as well as neural coding are only incompletely understood. This is partly due to the property of some tastants to elicit several sensations simultaneously, unspecific effects caused by the temperature, viscosity, osmolarity, and pH of the solvents, as well as by mechanical stimulation of the tongue during stimulus application. The analysis of gustatory processing of taste information are mainly based on mouse models after stimulation with bitter taste stimuli. Even though it is known that the mouse genome codes for 35 bitter taste receptor genes only few of them had been analysed so far. For better understanding and interpretation of animal experiments 16 mouse bitter receptors had been analysed by Calcium Imaging experiments with HEK293T cells. The data reveal that mouse bitter taste receptors are more narrow tuned than human bitter taste receptors, proving that the ligand spectra of murine and human orthologous receptors are not complient. In order to avoid the disturbing effects of solvents and stimulus application on the analysis of gustatory information transfer and processing, I employ an optogenetical approach to address this problem. For this purpose I generated three strains of gene-targeted mice in which the coding regions of the genes for the umami receptor subunit Tas1r1, the sweet receptor subunit Tas1r2 or the bitter taste receptor Tas2r114 have been replaced by the coding sequences of different opsins (photoreceptors of visual transduction) that are sensitive to light of various wavelengths. In these animals I should be able to activate sweet, bitter, or umami signalling by light avoiding any solvent effects. In initial experiments of this project I demonstrated that the various visual opsins indeed functionally couple to taste signal transduction pathway in oocyte expression system, generating basic knowledge and foundation for the generation of the gene-targeted animals. The knockout-knockin strategies have been successfully realized in the case of all three mouse models, revealed by RT-PCR, in situ hybridization and immunohistochemical analysis of taste papillae. All data confirm that the particular taste receptors have been replaced by the different opsins in taste cells. Further analysis concerning the functional consequences of opsin knockin and taste receptor knockout are part of prospective work.

Orientador: Marli de Fátima Stradioto Papa / Banca: Marineide Rosa Vieira / Banca: César Júnior Bueno / Resumo: Plantas apresentam grande diversidade de compostos orgânicos que podem ter ação antifúngica. O objetivo deste estudo foi determinar a atividade antifúngica in vitro e in vivo de extratos de pacari e melão-de-são-caetano sobre o crescimento micelial e a germinação de esporos de Colletotrichum musae. Avaliou-se, também o efeito dos extratos no desenvolvimento de lesões de antracnose em bananas, da variedade Nanicão, sob condições de laboratório e a campo. Os extratos foram obtidos a partir de plantas secas ao sol e em estufa com circulação forçada de ar, moídas, utilizando-se como agente extrator a água ou álcool etílico. Somente o fungicida proporcionaram 100% de inibição do crescimento micelial e da germinação de esporos e o extrato de melão-de-são-caetano hidroetanólico, seco so sol, incorporado a 50% em BDA, proporcionaram 100% de inibição da germinação de esporos. Os extratos de pacari aquoso não diluído proporcionaram as menores percentagens de lesões de antracnose nos frutos. Os extratos hidroetanólicos de pacari retardaram a maturação dos frutos em quatro-cinco dias em relação a testemunha. As propriedades antifúngicas dos extratos de pacari e melão-de-são-caetano detectadas em nosso estudo evidenciaram o uso potencial dos mesmos como uma alternativa aos métodos adotados para o controle da antracnose em banana. / Abstract: Plants present a high diversity of organic compounds that can have antifungal activity. The objective of this study was to determine the in vitro and in vivo antifungal activity of pacari and bitter melon extracts on the mycelial growth and spores germination of Colletotrichum musae. We also aimed to evaluate the effect of these two extracts in the development of anthracnose lesions on bananas variety Nanicão, under laboratory and field conditions. The extracts were obtained by grinding sun- or stews with forced circulation of air plant tissues, using water or ethylic alcohol as extractors. Only the fungicide check inhibited 100% of the mycelial growth and the spores germination, and of bitter melon hydro-ethanolic extracts incorporated at 50% in PDA inhibited 100% of the spores germination. The non-diluted aqueous extracts of pacari provided the lowest percentages of anthracnose lesions on banana fruits. The hydro-ethanolic extract of pacari delayed the maturation of the fruits in at least four to five days in relation to the non-treated checks. The antifungal properties of the pacari and bitter melon extracts detected in our study evidenced their potential use as alternative to the methods commonly adopted for controlling banana anthracnose. / Mestre

Made available in DSpace on 2014-06-11T19:29:45Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-12-02Bitstream added on 2014-06-13T21:00:09Z : No. of bitstreams: 1 silva_mf_me_ilha.pdf: 503783 bytes, checksum: bbb9bd24eb36123d7fea9c4f63053199 (MD5) / Plantas apresentam grande diversidade de compostos orgânicos que podem ter ação antifúngica. O objetivo deste estudo foi determinar a atividade antifúngica in vitro e in vivo de extratos de pacari e melão-de-são-caetano sobre o crescimento micelial e a germinação de esporos de Colletotrichum musae. Avaliou-se, também o efeito dos extratos no desenvolvimento de lesões de antracnose em bananas, da variedade Nanicão, sob condições de laboratório e a campo. Os extratos foram obtidos a partir de plantas secas ao sol e em estufa com circulação forçada de ar, moídas, utilizando-se como agente extrator a água ou álcool etílico. Somente o fungicida proporcionaram 100% de inibição do crescimento micelial e da germinação de esporos e o extrato de melão-de-são-caetano hidroetanólico, seco so sol, incorporado a 50% em BDA, proporcionaram 100% de inibição da germinação de esporos. Os extratos de pacari aquoso não diluído proporcionaram as menores percentagens de lesões de antracnose nos frutos. Os extratos hidroetanólicos de pacari retardaram a maturação dos frutos em quatro-cinco dias em relação a testemunha. As propriedades antifúngicas dos extratos de pacari e melão-de-são-caetano detectadas em nosso estudo evidenciaram o uso potencial dos mesmos como uma alternativa aos métodos adotados para o controle da antracnose em banana. / Plants present a high diversity of organic compounds that can have antifungal activity. The objective of this study was to determine the in vitro and in vivo antifungal activity of pacari and bitter melon extracts on the mycelial growth and spores germination of Colletotrichum musae. We also aimed to evaluate the effect of these two extracts in the development of anthracnose lesions on bananas variety Nanicão, under laboratory and field conditions. The extracts were obtained by grinding sun- or stews with forced circulation of air plant tissues, using water or ethylic alcohol as extractors. Only the fungicide check inhibited 100% of the mycelial growth and the spores germination, and of bitter melon hydro-ethanolic extracts incorporated at 50% in PDA inhibited 100% of the spores germination. The non-diluted aqueous extracts of pacari provided the lowest percentages of anthracnose lesions on banana fruits. The hydro-ethanolic extract of pacari delayed the maturation of the fruits in at least four to five days in relation to the non-treated checks. The antifungal properties of the pacari and bitter melon extracts detected in our study evidenced their potential use as alternative to the methods commonly adopted for controlling banana anthracnose.

Chez les animaux et en particulier les insectes, l’alimentation comprend une phase d’examen sensoriel qui précède l’ingestion, afin notamment d’éviter d’ingérer des substances toxiques. Cette détection fait intervenir des cellules spécialisées dans la détection de telles molécules, cellules qui sont généralement qualifiées de sensibles aux goûts « amers ». A l’aide d’observations électrophysiologiques et comportementales, nous avons abordé comment un insecte modèle, la drosophile, était capable de détecter des substances potentiellement toxiques mélangées à des sucres à l’aide de ses neurones gustatifs. Dans une première partie, nous avons étudié la détection de la L-canavanine, qui est un acide aminé non protéique. Cette molécule est toxique pour l’homme comme pour les animaux car elle est confondue par le métabolisme avec un acide aminé, la L-arginine, et intégrée à sa place dans les protéines. En utilisant des constructions génétiques et en particulier le système UAS-Gal4, nous avons montré que la Lcanavanine est détectée par des cellules gustatives qui expriment une protéine réceptrice GR66a, qui est impliquée dans la détection de nombreuses substances amères. Nous avons également montré que, contrairement à la caféine, la détection de L-canavanine nécessite des protéines Gαo fonctionnelles. Nous avons ensuite étudié les interactions sucré-amer. Dans un premier travail, nous avons montré que l’addition de Lcanavanine une solution sucrée n’altérait pas la détection des sucres, contrairement à la strychnine qui peut complètement supprimer la détection du sucre dans les cellules gustatives. Grâce à des ablations spécifiques des cellules détectant l’amer, nous avons pu montrer que cette inhibition était une propriété intrinsèque des cellules sensibles aux. sucres. Les cellules sensibles aux sucres auraient donc des sites récepteurs non identifiés, sensibles à certains ligands amers. Nous avons également abordé des interactions inverses, à savoir l’inhibition de la détection de substances amères par des sucres, en confrontant 4 substances amères (denatonium, berberine, caféine, umbelliferone) à 12 sucres. Les observations que nous avons réalisées montrent que certains sucres exercent un effet inhibiteur sur la détection des molécules amères testées. En utilisant des outils génétiques permettant l’ablation des cellules sensibles aux sucres, nous avons montré que cette inhibition est une propriété intrinsèque des cellules sensibles à l’amer. Cependant, cet effet inhibiteur est loin d’être aussi efficace que l’inhibition des substances amères sur la détection des sucres. Dans une dernière partie, nous avons évalué la modulation de la détection gustative à l’aide d’analogues d’une neuro-hormone, la leucokinine, connue pour ses effets sur la diurèse. Lorsqu’elle est mélangée à une solution sucrée, ces analogues inhibent la détection des sucres par les sensilles gustatives, à la fois chez le moustique Aedes aegypti et chez la drosophile. La détection de substances « amères » par les cellules gustatives de drosophiles implique donc deux voies de codage : l’une, spécifique, concerne des cellules dédiées à la détection des substances amères ; l’autre, moins spécifique, affecte les cellules dédiées à la détection des sucres. De manière réciproque, ces cellules dédiées à la détection des molécules sont affectées par la présence de ligands sucrés. Le codage des informations gustatives à la périphérie est donc un phénomène plus complexe qui nécessite d’étudier plus précisément la détection de composés en mélanges. / In most animals including insects, ingestion is preceded by a close examination of the food, for example in order to detect the presence of potentially noxious chemicals. This detection involves specialized gustatory cells, which are generally described as sensitive to “bitter” tastes. Using electrophysiology and behavioral observations, we studied how a model insect, Drosophila melanogaster, can detect potentially toxic substances (described here as “bitter”) when mixed with sugar molecules, with their gustatory neurons. In a first part, we studied how L-canavanine is detected. Lcanavanine is a pseudo amino acid, which is confounded with L-arginine by the metabolism. Proteins which include Lcanavanine are non-functional and this compound is toxic for animals including insects. Using genetic constructions based on the UAS-Gal4 expression system, we showed that Lcanavanine is detected by gustatory cells expressing a receptor protein, GR66a, which is specific to most cells capable of detecting bitter substances. We also showed that, contrary to caffeine, the detection of L-canavanine requires functional Gαo proteins. Then, we studied some aspects of the detection of mixtures of sweet and bitter molecules. In a first approach, we contributed to establish that L-canavanine does not impact sugar detection, while other chemicals like strychnine completely inhibit sugar detection. By using the UAS-Gal4 system to ablate bitter-sensitive cells, we could demonstrate that such inhibition is a specific property of sugar- sensitive cells. These cells should have thus receptors for bitter substances which have not been identified yet. We also examined the reverse interaction, which is a possible role of sweet molecules to inhibit the detection of bitter substances. We examined the detection of denatonium, berberine, caffeine and umbelliferone in the presence of 12 different sugars, using behavioral and electrophysiology observations. By using genetic construction to ablate sugar-sensitive cells, we found that the sugar inhibitory action is not due to the presence of sugar-sensitive cells. It should be noted, however that in our experimental conditions, this inhibitory action is less efficient than the inhibition of bitter upon sugar detection. In a last part, we examined the modulation of gustatory perception by analogs of leucokinine, which is a neuropeptide involved in the diuresis of insects. We show that these analogs, when mixed with sugars in solution, can inhibit sugar detection by gustatory sensilla, both in Aedes aegypti mosquitoes and in Drosophila. The detection of bitter molecules by gustatory neurons in Drosophila thus involves two main coding channels: one is specific, and involves gustatory cells dedicated to the detection of bitter molecules; the second one, less specific, is affecting cells which are dedicated to the detection of sugar molecules. Gustatory coding is thus a more complex phenomenon than previously thought on the basis of examining responses to single molecules, thus urging to study the responses of gustatory receptors to more complex and natural mixtures.

Taste is one of the crucial factors that contributes to shaping eating behaviors and is also one of the leading reasons that affects our preferences to like or dislike some foods that mainly have a bitter taste. Variation in bitter phenotype (tasters and non-tasters) could influence diet quality and in turn body weight, which overall influenced health outcomes. The objective of this study was to identify whether bitter taste phenotype status influences anthropometric measurements, body fat percentage, and eating behaviors (liking and intake) in female college students. In this cross-sectional study (n = 86), female college students aged 18 to 22 from Mississippi State University were classified into one of two groups (taster or non-taster) by means of a taste test of filter paper saturated with the bitter compound 6-n-propylthiouracil (PROP). Adiposity was measured using anthropometric measurements and body composition was measured using bioelectrical impedance analysis. A food liking survey was administrated to identify how much participants liked or disliked various foods and beverages. Dietary intake of total energy intake, macronutrients, fruits, and vegetables were evaluated using the NIH Diet History Questionnaire. Bitter phenotype status was not significantly associated with adiposity indicators; however, it was associated with food liking scores for foods that have bitter and umami tastes such as kale and mushrooms. Additionally, bitter phenotype was associated with dietary intake for total fruits and vitamin C intakes. Ethnic background was the strongest independent variable that was significantly correlated with adiposity indicators and food liking. These results suggested that while bitter taste phenotype may influence eating behaviors in certain foods, it does not affect adiposity indicators and body fat percentage.

L'obésité constitue l'un des principaux problèmes de santé publique en ce début du 21ème siècle. Sa prévalence augmente régulièrement, en particulier chez les enfants. Ce constat n'est pas anodin car l'obésité est généralement associée à diverses pathologies graves (diabète de type 2, hypertension et cancer,…). Ainsi, des investigations sur les mécanismes impliqués dans la perception gustative des lipides alimentaires pourraient éclairer leurs rôles dans l’incidence de l’obésité.Plusieurs études ont démontré le rôle des endocannabinoïdes et des aliments amers dans l’obésité. Ainsi, nous avons étudié l’interaction (cross-talk) des récepteurs cannabinoïdes et du goût amer avec le goût lipidique. Cette thèse comporte ainsi deux volets : les récepteurs cannabinoïdes (CB1R), le goût amer et leurs interactions avec les récepteurs lipidiques.Dans la première partie, nous avons étudié le rôle régulateur de CB1R. Dans la présente étude, des tests comportementaux sur des souris CB1R-/- et des souris de type sauvage (WT) ont montré que l'invalidation du gène Cb1r était associée à une faible préférence pour les solutions contenant de l'huile de colza ou un acide gras à longue chaîne (AGLC) tel que l’acide linoléique (LA). L'administration de rimonabant, un agoniste-inverse de CB1R, chez la souris a également entraîné une faible préférence pour les acides gras alimentaires. Aucune différence dans l'expression des protéines CD36 et GPR120 n'a été observée dans les cellules des papilles gustatives des souris WT et CB1R-/-. La signalisation calcique via CD36 dans les cellules des papilles gustatives des souris CB1R-/- diminue de façon significative par rapport à celle observée dans les cellules gustatives des souris WT. Les cellules des papilles gustatives des souris CB1R-/- présentent également une diminution significative de l'ARNm de Pro-glucagon et de Glp-1r et un faible niveau basal de GLP-1. Nous rapportons que CB1R est impliqué dans la perception du goût du gras via la signalisation calcique et la sécrétion de GLP-1.Dans la seconde partie, nous avons d’abord caractérisé le phénotype de cellules fongiformes humaines (HTC-8). En effet, le projet de ma thèse comprend la caractérisation à l’échelle moléculaire des récepteurs amers et lipidiques et leur cross-talk dans ces cellules (collaboration BRAIN, Allemagne). Nous avons démontré que les cellules HTC-8 expriment PLCβ2 et l’α-gustducin à l’échelle des ARNm et des protéines. Elles expriment également TAS2R16 et TAS2R38 et ces mêmes cellules co-expriment CD36 et GPR120. Puis, nous avons étudié la signalisation via ces récepteurs en utilisant l’acide linoléique, un agoniste de CD36 et GPR120, la sinigrin, agoniste de TAS2R16 et TAS2R38, la salicin, agoniste du récepteur TAS2R16 et le phénylthiocarbamide, agoniste du récepteur TAS2R38. De plus, les études du signal calcique ont démontré que la signalisation en aval du goût gras partage une voie commune avec la signalisation en aval du goût amer, mettant en évidence un cross-talk entre ces deux modalités gustatives.Bien que nous ayons montré le cross-talk entre les modalités gustatives amère et lipidique, il nous reste à étudier ces phénomènes à l’échelle de l’organisme. Ces résultats, d’ores et déjà, montrent que le goût amer et le récepteur cannabinoïde-1 sont liés à la sensibilité au goût du gras et doivent être pris en compte pour la gestion de l'obésité. / Obesity is one of the major public health problems at the beginning of the 21st century. Its prevalence is increasing steadily, especially among children. This observation is not insignificant because obesity is generally associated with various serious pathologies (type 2 diabetes, hypertension and cancer, etc.). Thus, investigations into the mechanisms involved in the taste perception of dietary lipids could shed light on their roles in the incidence of obesity.Several studies have demonstrated the role of endocannabinoids and bitter foods in obesity. Thus, we studied the cross-talk of cannabinoid receptors and bitter taste with lipid taste. This thesis has two components: cannabinoid receptors (CB1R), bitter taste and their interactions with lipid receptors.In the first part, we studied the regulatory role of CB1R. In the present study, behavioral tests on CB1R-/- mice and wild-type (WT) mice showed that the invalidation of the Cb1r gene was associated with a low preference for solutions containing rapeseed oil or a long chain fatty acid (LCFA) such as linoleic acid (LA). Administration of rimonabant, a CB1R inverse agonist, in mice also resulted in a low preference for dietary fatty acids. No differences in the expression of CD36 and GPR120 proteins were observed in the taste buds cells of the WT and CB1R-/- mice. Calcium signaling via CD36 in the taste bud cells of CB1R-/- mice decreased significantly compared with those observed in the taste cells of WT mice. The taste bud cells of CB1R-/- mice also show a significant decrease in Pro-glucagon and Glp-1r mRNA and a low basal level of GLP-1. We report that CB1R is involved in the perception of fat taste via calcium signaling and secretion of GLP-1.In the second part, we first characterized the phenotype of human fungiform cells (HTC-8). Indeed, the project of my thesis includes the characterization on the molecular scale of bitter and lipid receptors and their cross-talk in these cells (collaboration BRAIN, Germany). We have demonstrated that HTC-8 cells express PLCβ2 and α-gustducin at the mRNA and protein level. They also express TAS2R16 and TAS2R38 and these same cells co-express CD36 and GPR120. Then, we studied signaling via these receptors using linoleic acid, a CD36 and GPR120 agonist, sinigrin, TAS2R16 agonist and TAS2R38, salicin, TAS2R16 receptor agonist, and phenylthiocarbamide, TAS2R38 receptor agonist. In addition, calcium signal studies have shown that downstream fatty signaling shares a common path with downstream bitter taste signaling, highlighting a cross-talk between these two taste modalities.Although we have shown the cross-talk between bitter and lipid taste modalities, we still have to study these phenomena at the level of the organism. These results, already, show that the bitter taste and the cannabinoid-1 receptor are related to the taste sensitivity of fat and must be taken into account for the management of obesity