Contribution of estrogen and GluN2B subunit to the HtraKO mouse phenotype

Htr1a is one of the most widespread serotonin receptor across the brain, strongly expressed in CAI region of hippocampus. Our laboratory studies the phenotypic alteration in 5HTla- deficient mice (Htr1aK0), characterized an abnormal anxious-like behavior. Our aim is to evaluate the regulation of this cognitive process by understanding the circuitry involved. This phenotype sets up early during development and has durable effect in adulthood. Our laboratory showed that adult Htr1aK0 male mice displaying exuberant dendritic growth of oblique dendrites in a specific layer of a CAI pyramidal neurons, the stratum radiatum. Application of drugs in organotypic cultures and by in vivo injections revealed that GluN2B, a subunit of NMDA receptor highly expressed during development, is responsible for this dendritic exuberance. Immunohistochemistry highlighted in particular a synaptic enrichment of GluN2B in stratum radiatum of Htr1aK0 CAI pyramidal neurons at puberty. Finally, original analysis of Htr1aK0 mouse behavior showed a different response to anxiety between male and female. Htr1a activation down-regulates the CaMKII activity in the CAI pyramidal neurons. CaMKII directly favors the membrane conductance and stability of GluN2B at the synapse. In the context of the Htr1aK0 mouse, GluN2B is the final common pathway of our phenotype. This subunit is well known to regulate the threshold of LTD/LTP and the dendritogenesis during development. In my thesis, I establish a link between the gender differences in the morphology and the physiology in the Htr1aK0 mice during development to understand how these characteristics shape the circuit with prominent cognitive impacts in adulthood. My study highlighted that during development, Htr1aK0 male mice show a constant increase of the dendritic growth of oblique dendrites from early ages until adulthood associated with an increased physiological impact of altered GluN2A/GluN2B ratio. Whereas during puberty, synaptic contribution of GluN2B to NMDA response is higher in Htr1aK0 compared to WT male mice, this ratio comes back to normal values towards adulthood. However, this recovery of the ratio of GluN2A/GluN2B located at the synaptic level is concomitant with the lateral diffusion of excess GluN2B subunits, leading to extrasynaptic enrichment. The main impact was a lowering of the LTP threshold characterized by strong increased potentiation of synaptic strength after 5 Hz low frequency stimulation. Moreover, the extrasynaptic GluN2B overexpression leads to a shift of the maturation phase switch explaining the exuberant morphology. However, Htr1aK0 females characterized during the 3 first weeks of development by an increase of the dendritic growth of oblique dendrites showed starting at puberty that the dendrite arborization returns progressively to WT values. The physiological impact of GluN2B was investigated and directly linked to this morphology, since Htr1aK0 female mice does not show alteration of the synaptic strength during development. These observations show a compensation occurring in Htr1aK0 female, responsible for a rescue of the phenotype morphologically, physiologically and to be tested behaviorally. We highlighted then the biological processes underlying this compensation. During development, sexual hormones such as testosterone and estrogen are responsible to induce sexual differentiation of specific brain regions. I demonstrated that estrogen, but not testosterone, was able to reduce both in vitro and in vivo the dendritic arborization early during development, through activation of GPER-1, a G-coupled protein estrogen receptor, which phenocopy the activation of Htr1a by reducing GluN2B conductance and stability. I then identified a pathway, parallel to Htr1a, able to regulate GluN2B and responsible for the morphological and physiological phenotype in Htr1aK0 female mice. The specific rise of estrogen occurring at puberty in female is responsible for the compensation observed and induces a late rescue of the Htr1aK0 phenotype by activation GPER-1. -- Htr1a est un des récepteurs à la sérotonine les plus répandus dans le cerveau, fortement exprimé dans la région CAI de l'hippocampe. Notre laboratoire étudie les altérations phénotypiques de souris déficientes pour ce récepteur (Htr1aK0), caractérisées par un comportement avec des traits anxieux. Notre objectif est d'évaluer la régulation de ces processus cognitifs en comprenant les connexions nerveuses impliquées. Ce phénotype se met en place tôt au cours du développement et présente un effet durable à l'âge adulte. Notre laboratoire a montré que les souris Htr1aK0 mâles adultes se caractérisent par une croissance exubérante des dendrites obliques dans une couche spécifique des neurones pyramidaux du CAI, le stratum radiatum. L'application de drogues sur cultures organotypiques et par injections in vivo ont révélé que GluN2B, une sous-unité du récepteur NMDA fortement exprimée au cours du développement, est responsable de cette exubérance dendritique. Des expériences d'immunohistochimie ont notamment mis en évidence un enrichissement synaptique de GluN2B durant la puberté dans le stratum radiatum des neurones de la région CAI des souris Htr1aK0. Finalement, l'analyse originale du comportement des souris Htr1aK0 a montré une différence de réponse à l'anxiété entre mâles et femelles. L'activation de Htr1a diminue l'activité de la CaMKII dans les neurones pyramidaux du CAI. La CaMKII favorise directement la conductance et la stabilité de la sous-unité GluN2B à la synapse. Dans le contexte de la souris Htr1aK0, GluN2B est le « médiateur » de notre phénotype. Cette sous-unité est particulièrement connue pour réguler le seuil de LTD-LTP ainsi que la dendritogénèse durant le développement. Dans ma thèse, j'ai établi le lien entre les différences dépendant du genre dans la morphologie et physiologie des souris Htr1aK0 au cours du développement pour comprendre comment ces caractéristiques modulent le circuit accompagnés d'impacts cognitifs visibles à l'âge adulte. Mon étude a mis en évidence que durant le développement, les souris mâles Htr1aK0 montrent une constante augmentation de la croissance des dendrites obliques entre les premières semaines et l'âge adulte associée à une augmentation de l'impact physiologique du ratio GluN2A/GluN2B altéré. Alors que durant la puberté, la contribution synaptique de GluN2B à la réponse NMDA est plus haute chez la souris mâle Htr1aK0 que le WT, ce ratio revient à des valeurs normales à l'âge adulte. Cependant, cette récupération de l'expression du récepteur au niveau synaptique est concomitante avec la diffusion des sous-unités GluN2B excédantes, amenant alors à un enrichissement extrasynaptique. Le principal impact est une diminution du seuil de la LTP caractérisée par une forte potentiation de la plasticité après une stimulation basse fréquence à 5 Hz. De plus, la surexpression des GluN2B extrasynaptiques conduit à un décalage de la bascule à la phase de maturation, expliquant la morphologie dendritique exubérante. Cependant, les femelles Htr1aK0 initialement caractérisées pendant les 3 premières semaines du développement par une augmentation de la croissance des dendrites obliques montrent à partir de la puberté que cette arborisation dendritique retourne à des valeurs WT. L'impact physiologique de GLuN2B a été investigué et mis en lien avec cette morphologie, étant donné que les femelles Htr1aK0 ne montrent pas d'altération de la plasticité durant le développement. Ces observations montrent une compensation se produisant chez la femelle Htr1aK0, responsable d'une récupération du phénotype morphologique, physiologique et peut-être comportemental. Nous avons souligné les processus biologiques sous-jacent à cette compensation. Au cours du développement, les hormones sexuelles telles que la testostérone et l'estrogène sont responsables de la différentiation sexuelle de régions du cerveau spécifiques. J'ai démontré que l'estrogène, mais pas la testostérone, était capable de réduire in vitro et in vivo l'arborisation dendritique tôt dans le développement au travers de l'activation du récepteur GPER-1, un récepteur aux estrogènes couplés à un protéine G, qui phénocopie l'activation de Htr1a en réduisant la conductance et la stabilité de GluN2B à la membrane. J'ai identifié une voie de signalisation parallèle à celle de Htr1a, capable de réguler GluN2B et responsable du phénotype morphologique et physiologique de la souris femelle Htr1aK0. La montée spécifique d'estrogène se déroulant à la puberté chez la femelle est responsable de cette compensation et implique une récupération tardive du phénotype Htr1aK0 par l'activation de GPER-1.

Análise bioclimática como ferramenta para implementação do plano diretor do campus central da UFRN

This research investigates the microclimate and the morphology features of the central campus of the UFRN, in Natal-RN, through the use of bioclimatic tools of analysis in order to assist the implementation of the campus´ Master Plan. It develops a diagnosis of the evolution and growth of the urban space surveyed by analyzing its initial plan and the basic urban conception behind it, as well as the morphology and typologies utilized. The study makes a qualitative analysis of the local microclimate by using Katzschner (1997) methodology, with land-use and topography maps, building heights, vegetation and soil covering. It also makes use of the methodology proposed by Oliveira (1993), which examines, from the bioclimatic standpoint, the human environment as related to the urban form (site and built mass). It identifies zones whose climatic characteristics are representative of the local microclimate and classifies them into areas to be strictly preserved, areas to be protected and areas to be improved. By means of the methodology for spatial and environmental assessment developed by Bustos Romero (2001), the survey selects characteristic points of each area in order to register the environmental data relative to the two basic seasons found in the region where the campus is located, that is, the dry and the rainy season, so that it can evaluate changes in the environment which might have been caused by urban density growth, by arborization or by the influence of the urban form. It then proceeds to a quantitative and statistical survey of the collected data with the purpose of evaluating the degree of influence of the identified features over the environmental variables along the different scales of approach. The study shows the existence of different microclimates and emphasizes the relevance of the bioclimatic analysis of the built environment as a tool for the decision-making process along the development of the Master Plan for UFRN Central Campus

Visitantes florais de Erythrina speciosa Andr. (Leguminosae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Morfologia de frutos, sementes e plântulas de castanheira (Terminalia catappa L. - COMBRETACEAE)

O trabalho foi realizado com o objetivo de descrever morfologicamente os frutos, sementes e plântulas de castanheira. Foi feita a biometria dos frutos e das sementes e sua caracterização quanto à forma, por meio de mensurações com paquímetro e observações realizadas em estereomicroscópio com câmara clara. Os frutos de castanheira são carnosos, indeiscentes, do tipo nucóide, glabros, de coloração verde a vinácea, projeção das nervuras carpelares externamente evidentes, com epicarpo delgado, mesocarpo carnoso e esponjoso de coloração vinácea, com feixes vasculares conspícuos em corte transversal. Geralmente, cada fruto contém apenas uma semente. As sementes são exalbuminosas, de formas alongadas e cilíndricas, recobertas por endocarpo rígido de coloração marrom; possuem cerca de 2,5cm, 0,7cm e 0,7cm, de comprimento, largura e espessura, respectivamente. A germinação das sementes de castanheira é epígea, e a plantula é fanerocotiledonar.

Análise quali-quantitativa da arborização na praça XV de novembro em Ribeirão Preto - SP, Brasil

A Praça XV de Novembro, implantada em meados do século XIX, tem grande valor histórico-cultural, além de se constituir em uma das principais áreas verdes do centro da cidade de Ribeirão Preto. Visando ao conhecimento detalhado da vegetação para fins de orientação do manejo e conservação dessa área, foi feito um levantamento quali-quantitativo e fitossociológico das árvores e palmeiras da praça. Foram medidas altura e Diâmetro à Altura do Peito (DAP) e identificados todos os indivíduos de porte arbóreo (árvores e palmeiras) presentes na Praça, em nível de espécie. A praça ocupa uma área de 15.456,00 m², onde foram amostradas 42 espécies distribuídas por 19 famílias, num total de 161 indivíduos. Apesar de o local apresentar arborização com alto índice de diversidade de espécies (Shannon-Weaver de 3,14), os exemplares necessitam de maior atenção quanto a problemas ligados à fitossanidade e podas adequadas, fazendo que resulte em espaço seguro para os frequentadores e em boa qualidade paisagística.

Germinação de sementes de Mimosa caesalpiniaefolia Benth. em diferentes substratos e temperaturas

A Mimosa caesalpiniaefolia Benth. é uma espécie nativa da região Nordeste, que vem sendo progressivamente cultivada do Maranhão ao Rio de Janeiro. A planta apresenta grande potencial para arborização, cerca viva e produção de madeira. O presente trabalho teve como objetivo definir o tipo de substrato, a temperatura, o tempo médio e a frequência relativa de germinação, para auxiliar a condução de testes de germinação e vigor em sementes de Mimosa caesalpiniaefolia. Para tanto, realizou-se um experimento no Laboratório de Análise de Sementes do CCA-UFPB, em Areia-PB, em delineamento inteiramente casualizado com os tratamentos distribuídos em esquema fatorial 4x4, com os fatores temperaturas (20, 25, 30ºC constantes e 20-30ºC alternadas) e substratos (entre papel, sobre papel, entre areia e entre vermiculita), em quatro repetições de 25 sementes. Foram analisados os seguintes parâmetros: porcentagem de germinação e de plântulas anormais, primeira contagem da germinação e velocidade de germinação, tempo médio e frequência relativa de germinação, massa fresca e massa seca de plântulas. A temperatura de 25ºC mostrou-se mais adequada para a condução dos testes de germinação e vigor em sementes de Mimosa caesalpiniaefolia, independentemente do substrato utilizado. O substrato entre papel foi o mais apropriado para avaliação da qualidade fisiológica das sementes e a sincronização do processo germinativo foi maior no substrato entre papel, independentemente da temperatura empregada.

Caracterização de subpopulações de interneurônios imunorreativos para proteínas ligantes de cálcio no córtex pré-frontal do Sagui (Callithrix jacchus): distribuição e morfologia

Cortical interneurons are characterized by their distinct morphological, physiological and biochemical properties, acting as modulators of the excitatory activity by pyramidal neurons, for example. Various studies have revealed differences in both distribution and density of this cell group throughout distinct cortical areas in several species. A particular class of interneuron closely related to cortical modulation is revealed by the immunohistochemistry for calcium binding proteins calbindin (CB), calretinina (CR) and parvalbumin (PV). Despite the growing amount of studies focusing on calcium binding proteins, the prefrontal cortex of primates remains relatively little explored, particularly in what concerns a better understanding of the organization of the inhibitory circuitry across its subdivisions. In the present study we characterized the morphology and distribution of neurons rich in calcium-binding proteins in the medial, orbital and dorsolateral areas of the prefrontal cortex of the marmoset (Callithrix jacchus). Using both morphometric and stereological techniques, we found that CR-reactive neurons (mainly double bouquet and bipolar cells) have a more complex dendritic arborization than CB-reactive (bitufted and basket cells) and PV-reactive neurons (chandelier cells). The neuronal densities of CR- and CB-reactive cells are higher in the supragranular layers (II/III) whilst PV-reactive neurons, conversely, are more concentrated in the infragranular layers (V/VI). CR-reactive neurons were the predominant group in the three regions evaluated, being most prevalent in dorsomedial region. Our findings point out to fundamental differences in the inhibitory circuitry of the different areas of the prefrontal cortex in marmoset

Lymphatic drainage on healthy and neoplasic mammary glands in female dogs: Can it really be altered?

The purpose of this research was to study the mammary lymphatic drainage under a macroscopic and mesoscopic view, comparing the vascular pattern of healthy and neoplasic mammary glands injected with drawing ink alcoholic and fluorescein solutions, in 46 mongrel female dogs. The results pointed out that the thoracic gland is drained by the axillary lymph centre, but in mammary neoplasia either superficial cervical or ventral thoracic lymph centres can be involved. Cranial and caudal abdominal glands may be drained by the axillary, inguinofemoral and popliteal lymph centres. However, the popliteal drainage is specific for the healthy caudal abdominal mammary gland. The inguinal gland can be drained by both inguinofemoral and popliteal lymph centres in both neoplasic and healthy conditions. Regarding the mammary lymphatic communications, this research demonstrated that neoplasic glands present more types of anastomosis (40.9%), than healthy glands (33.33%), and an increase in contralateral anastomosis (50%) compared with healthy ones (33%). Given the data, the mammary neoplasia can change the lymphatic drainage pattern in terms of lymph centres and vascular arborization, thus forming new drainage channels and recruiting a larger number of lymph nodes. Lastly, some comments were made about the severity of a specific neoplasic mammary gland and conditions to be considered before making a decision in terms of the most adequate operative procedure, and suggestions for further investigations.

The Heart of the South American Rattlesnake, Crotalus durissus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Composto e vermicomposto de lixo urbano na produção de mudas de oiti (Licania tomentosa (Benth)) para arborização

Compost and vermicompost, produced from organic fraction of municipal solid wastes were studied as components of potting media for Licania tomentosa (oiti) seedlings production. The objective was to achieve data to amplify the utilization of these products, in order to utilize them to ornamental and urban arborization species. Increasing rates of organic fertilizers were used until the total soil substitution from the potting media. There were no fitotoxic effects. Compost or vermicompost caused higher germination index of seeds and higher growth of plants. The study demonstrated that it is possible to utilize these products in other activities beyond aplication in agricultural soil, and it can generate new markets to this kind of organic fertilizer.

Maturidade fisiológica das sementes do ipê amarelo, Tabebuia chrysotricha (Mart. Ex DC.) Standl

Ipê amarelo Tabebuia chrysotricha (Mart. Ex DC.) Standl. is a native forest species that presents heavy and durable wood: it is common to be used for urban arborization. The species is latte secondary and heliophyta. Occurs from Espirito Santo State until Santa Catarina in the Pluvial Atlantic Forest (Mata Atlântica). The seedling production is hard because their seeds have short longevity after dispersion, and so due to the difficulty to harvest winged seeds that are quickly dispersed by the wind. The present study was carried out to evaluate the physiological maturity of Ipê Amarelo seeds, to establish the best moment to harvest them. The study was to accompany the maturation of 100 fruits in just beginning development, from eleven different trees. Morphological characteristic measures were made weekly, with fruits still on trees, starting from the second week development when fruits presented the following averages: 6.3 cm length, 0.71 cm thickness, and 0.82 cm width. From the sixth week of fruit development we harvest them to proceed the germination test, water content rate, and electric conductivity test. The germination has begun by the seventh week development with 28% of germinated seedlings. The highest germination rate happened by the eighth week development, with 74.5% of germinated seedlings and on the ninth week the rate decrease to 65.5%, when the seeds were already in dispersion. Through the electric conductivity test the highest value obtained was 378.06 μS.g -1.cm -1 on the seventh week development and the lowest one was 183.28 μpS.g -1.cm -1 on eighth week. These results support those obtained in the germination test, because as higher is the electric conductivity of seeds, as higher is their deterioration level. Finally, the study allows concluding that the physiological maturation of ipê amarelo seeds has occurred before dispersion close to the eighth week development when fruits presented the following averages: 22.2 cm length, 1.37 cm width, and 1.05 cm thickness; water content rate was 61.8% and electric conductivity was 183.28 μS.g -1.cm -1. In that moment, the fruits presented greened brown coloration and started to show fissures.

Efeitos da luminosidade no crescimento de mudas de Caesalpinia ferrea Mart. ex Tul. (Leguminosae, Caesalpinoideae)

Caesalpinia ferrea is a species used a lot as a medicinal plant, for urban arborization and landscape design in the state of Amapa. Yet there is not much ecophysiological information available on it. Light is an important environment factor that controls processes associates with the accumulation of dry matter, contributing thus for plant growth. The object of this research was to study the effect of different luminosity levels on the growth of seedlings of this species. The seedlings were transplanted to plastics bags containing a mixture of soil and sand (2:1), which were maintained in the full sun, under artificial shade with reduction of 50% and 70% of the luminosity and under natural shade of a closed canopy of forest. The experimental design was completely randomized with five replicates. Seedlings submitted to natural shade showed strong growth inhibition. In the full sun, the seedlings presented higher net assimilatory rate (NAR), lower shoot/root ratio (SRR) and lower leaf area ratio (LAR). The results showed little difference in the growth and biomass allocation between seedlings kept in 50 and 70% shade; the seedlings submitted to this treatment presented higher values of SRR and LAR. This indicates plasticity which influences a possible increase in light capture and is important, therefore, to keep the seedling growth and survival under low light levels. The results as a whole showed morphological and physiological adjustment to different light levels in Caesalpinia ferrea.

Influence of hydric stress in the production of toxic principles in Nerium oleander L.

Many plants utilized in the urban center shows substances considered toxic whose production could be influenced by some factors, like hydric stress, including ornamental Nerium oleander L., widely used in gardens in various parts of the world, which presents production of cardioative glucosides, considered toxic. This study had the objective to evaluate the effect of field capacity in the biomass and the level of cardioative glucosides in seedlings of Nerium oleander. The experiment was carried at UNIDERP, in Campo Grande City, Mato Grosso do Sul State, Brazil, at the University for Development of State and Pantanal Region, using the experimental delineation in randomized blocks. There were 4 treatments (25%; 50%; 75% and 100% of the field capacity), 5 replications and 4 plants by parcel, totalling 80 plants. The evaluations were realized 60 days after the seedlings were planted. The quantitative analysis of the cardioative glucosides was realized by gravimetric test, after selective extraction of the glucosides. Were conclude that increase of the quantity of water in the soil raised the biomass production until 75% of the field capacity and increased the level of cardioative glucosides, showing that water management is very important and should be provided only the necessary to development of the plant.

Mapeamento de fragmentos de mata no município de Maringá, PR: uma abordagem da ecologia da paisagem

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Análise histórico-cultural, paisagística e quali-quantitativa dos elementos arquitetônicos da Praça Sete de Setembro, Ribeirão Preto, SP

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)