Impact of the feeding eycle upon the neuronal membrane properties of rat hippocampal neurones : the involvement of voltage-gated sodium and calcium currents and the maintenance of plasma membrane organization

Tese de doutoramento, Bioquímica (Biofísica Molecular), Universidade de Lisboa, Faculdade de Ciências, 2018Feeding behaviour and energy balance is regulated by the central nervous system, through a concerted endeavour of different brain areas. The hippocampus, historically regarded as a substrate for learning and memory processes, has also been implicated in such energy regulation. In recent years, researchers have established that hippocampal neurones form a memory of a meal and act to delay meal initiation during the postprandial period. However, more experiments are needed to identify the processes involved in such control. The present thesis starts to fill this gap, by identifying possible neuronal mechanisms by which the hippocampus processes satiety and meal termination. By assessing the functioning of ion currents/channels and the lipid composition and organization of the plasma membrane throughout the feeding cycle, this study furnishes a global perspective of the effect of post-prandial and fasting conditions upon intrinsic neuronal plasma membrane (PM) properties. The involvement of ion channels of rat hippocampal CA1 neurones in a feeding cycle context has already been studied. Indeed, the feeding cycle was found to impact the excitability of these neurones by modulating the activity of voltage-gated potassium currents. This finding has urged further investigation to evaluate the broadness of the effect of feeding cycle on the activity of other ion channels. Hence, it was critical to address the involvement of a) voltage-gated sodium (Na+) currents/channels, given their importance in the initiation and propagation of action potentials, and b) voltage-gated calcium (Ca2+) currents/channels, as they mediate the influx of this ubiquitous second messenger, with wide-ranging physiological roles, into the interior of the neurones. The influence of feeding cycle on the biophysics of Na+ and Ca2+ channels was undertaken in neurones acutely isolated from the CA1 subfield of the rat hippocampus. Two classes of neurones were used: those obtained from animals that fasted overnight (‘fasted neurones’) and those from animals that, after such period, were fed (‘fed neurones’). Voltage-gated Na+ currents were recorded by applying electrophysiological voltage clamp techniques - namely whole-cell (WC) and excised inside-out patches. Fed neurones, in comparison to fasted neurones, showed increased mean maximum macroscopic Na+ current density (1.5 ± 0.12mA.cm-2 vs. 1±0.10mA.cm-2) and a greater single-channel conductance (16.7 ± 0.76pS vs. 12.6 ± 1.30pS). Furthermore, the larger amplitude of the ‘window current’ obtained in fed neurones, derived from hyperpolarized activation curves and depolarized steady-state of inactivation curves (h∞), indicates a greater Na+ channel availability to respond to activation. Such variation is supported by a higher concentration of Nav1.2 isoform at the plasma membrane-enriched fractions of hippocampus of fed animals. Overall, the results indicate a variation in the biophysics and expression of voltage gated Na+ channels of rat hippocampal CA1 neurones, pointing out that feeding cycle changes the neuronal excitability. Voltage-gated Ca2+ currents were analysed with whole-cell recordings. It was observed heterogeneity in whole-cell Ca2+ currents, here sorted into three categories – ‘A’, ‘B’, and ‘C’ currents. The differential distribution of these currents between fed and fasted neurones determined significant alterations on the inactivation properties of Ca2+ currents. The increased values of the time-constant of inactivation - τh -, observed upon feeding, can be ascribed to a conspicuous slowly-inactivating current mainly assigned to fed neurones (current ‘A’), as oppose to the fastest kinetics of inactivation, solely seen in fasted neurones (current ‘C’). Furthermore, in fed neurones, a depolarizing shift of the most depolarized component (Vh2) of the voltage-dependence of h∞ was observed, which indicates that fasted neurones inactivate at more negative membrane potentials. Altogether, these observations point to a facilitated entry of Ca2+ into the soma of fed neurones, which, ultimately, potentiates the Ca2+-dependent intracellular events. The observed influence of feeding cycle on the biophysical and molecular expression of voltage-gated Na+ and Ca2+ channels did not have repercussions on the lipid environment of the PM. The plasma membrane-enriched fractions of rat hippocampus were labeled with molecular probes: 1,6-diphenyl 1,3,5-hexatriene (DPH) and trans-parinaric acid (t-PnA). By assessing the fluorescence properties of these probes, it was possible to study the molecular organization and lateral heterogeneity (in the membrane plane) of the lipid domains. Specifically, two types of fluorescence spectroscopy measurements were used, either in steady state (anisotropy measurements) and time-resolved domains (fluorescence intensity decay). The molecular biophysics analysis indicated that the order and rigidity of the acyl chains of the phospholipids constituents of the PM is not altered during the feeding cycle. Furthermore, the proportion of the different lipid domains at the surface of the neuronal PM is identical between conditions, which clearly indicates that the lateral heterogeneity of such domains is similar throughout the feeding cycle. This observation must be interpreted at a hydrophobic core level, where the t-Pna and DPH preferentially locate within the PM. The lipid content of the plasma membrane of rat hippocampus also did not endure any variation during the feeding cycle. The ratios calculated for the total lipid, phospholipid and cholesterol content were identical between the membranes of fed and fasted animals. The results concerning the molecular biophysics and biochemical characterization of the lipids imbedded in the neuronal plasma membrane indicate that neurones must have a shield mechanism to preserve their functional viability, regardless of the peripheral metabolic state. In summary, the greater levels of neuronal excitability and the promotion of Ca2+ entry into the neurones upon feeding may imply a subsequent increase on neuronal synaptic performance. A positive relationship between feeding and higher levels of synaptic plasticity-related phenomena (formation and consolidation of memories) is suggested, which could help to explain the role of hippocampus on the regulation of energy intake, mainly due to its role on meal-related episodic memories. This work gives new insights into the function of hippocampus on energy homeostasis, by adding new elements to the equation, namely, voltage-gated Na+ and Ca2+ channels.Fundação para a Ciência e a Tecnologia (FCT

Case study

This paper describes the numerical results of a CFD parametric studies of the influence on the thermal performance of a cold room by (1) the number and arrangement (n = 6, 9, 18, 30 and 40) of meat carcasses hanging in an airway; (2) the air temperature in the antechamber (T = 5ºC; 14ºC and 22ºC); and of (3) opening the door of the cold room. The case studies were tested in a real facility. A steady state 3D CFD model of the cold room is developed, allowing the detailed evaluation of the airflow and heat transfer for each case study. The thermal conduction within the meat carcasses is also predicted. The numerical predictions show that the average air temperature inside the cold room as well as the average carcass temperatures increase with the number of products and depend on the arrangement of the carcasses in the airway. A minimum space between carcasses is required to improve the refrigeration process. Similarly, the heat load of the cold room increases with the air temperature of the antechamber and when the door of the cold room is opened. The carcasses temperature profile becomes less uniform when the door is opened, being the temperature of carcasses located near the door strongly affected. These parametric studies allow to evaluate details of the air flow and heat transfer by convection and conduction inside goods that can contribute to the design of cold rooms in order to focus on the improvement of thermal performance and consequently of food safety and energy efficiency.info:eu-repo/semantics/publishedVersio

Estudo paramétrico por DFC da influência da capacidade e disposição do armazenamento de carcaças cárneas no desempenho térmico de câmaras de refrigeração

A necessidade de racionalizar energia, resultado do aumento do consumo energético, levou à procura de novas metodologias e soluções, a fim de otimizar processos industriais para promover a eficiência energética. É no contexto da otimização de sistemas de refrigeração, que surge o desenvolvimento deste trabalho. A aplicação da DFC na indústria agroalimentar é recente, pelo que o potencial em volta da otimização dos diferentes processos é elevado. A DFC permite de uma forma rápida e económica, simular processos e fenómenos, que experimentalmente consumiriam tempo e recursos. O presente estudo aborda a refrigeração de carcaças bovinas, tendo por base uma situação real. O trabalho desenvolvido assenta em 3 estudos base: a análise da influência da capacidade e disposição das carcaças, a influência da temperatura da antecâmara e avaliação do impacto da abertura das portas das câmaras de refrigeração. A análise efetuada, baseia-se em métodos computacionais com recurso à dinâmica de fluidos computacional, de modo a prever o escoamento de ar dentro da câmara e avaliar o comportamento térmico da mesma, consoante a alteração dos parâmetros. Os resultados obtidos, fruto das previsões numéricas, preveem que a refrigeração das carcaças depende do espaçamento entre elas e da temperatura do ar na antecâmara influencia o desempenho térmico da câmara. O estudo prevê um desperdício de energia, quando se abre a porta, permitindo que ar refrigerado saia, desaproveitando o sistema de frio. O estudo propõe pequenas melhorias, de modo a atenuar os efeitos dos diferentes parâmetros estudados.The current goal to save energy, due to energy consumption increase, lead us to find new strategies and solutions, in order to optimize industrial procedures and promote energy efficiency. The current work appears in this context, to optimize refrigeration systems. The application of CFD in food industry is recent, and its potential is powerful, in order to optimize different kinds of processes. CFD allows to simulate processes and phenomena in a quickly and economic way, which could take to much time and resources. The present study, focus on bovine carcasses refrigeration, taking in account a real situation. The developed work has 3 case studies: the capacity and disposal influence of the carcasses, the antechamber temperature influence and the impact on thermal performance of opening doors. The work is based in computational fluids dynamics methods, which goal is to provide air temperature and velocity field previsions inside the refrigeration chamber, in order to evaluate the temperature changes due to different parameters. The results obtained by means of CFD, predict a minimal space between carcasses, in order to reach effective refrigeration. The antechamber temperature influences the thermal performance. The study predicts a waste of energy, when the door is opened. The colder air leaves the chamber, from the bottom, and the warm air enters from the outside. The study proposes some changes to the actual facility with the main goal of increase the thermal performance

Experimental study and numerical modeling of the thermal performance of cold rooms for storage of meat products

This paper contributes to the energy efficiency and thermal performance improvement in cold rooms used for conservation and storage of meat products, commonly used in the agro industrial sector. An experimental analysis of the thermal performance of a cold room in the Covilhã region (Portugal) is developed, verifying its external conditions and construction parameters. A steady state 3D CFD model of the cold room is developed, allowing the detailed evaluation of the airflow and heat transfer. The CFD model setup required the definition of a physical-mathematical model based on the equations of continuity, momentum and energy. The comparison of numerical and experimental results allowed the validation of the CFD model. Thus, a parametric study is developed, which consists in changing the boundary condition values at the discharge grilles of evaporators. The parametric predictions show that the cold system, for product loading defined on the experimental analysis, is oversized, thus enabling the modification of the discharge air temperature and velocity. In this modification, the energy consumption is reduced maintaining similar thermal performance and ensuring the food safety. Thus, it is predicted an improvement of the energy efficiency and a consequent savings of environmental resources, energy and financial resources.info:eu-repo/semantics/publishedVersio

REFLEXOS DE UMA SOCIEDADE CONTEMPORÂNEA ACERCA DA EROTIZAÇÃO PRECOCE

O desenvolvimento infantil caracteriza-se enquanto artifício indissociável à vida de todo e qualquer sujeito, marcado por uma fase da sua existência. A sexualidade é um fenômeno complementar à vida do indivíduo, acompanhando-lhe em todas as fases do seu desenvolvimento. O referido trabalho tem como proposta principal, analisar a erotização precoce a partir de paradigmas de uma sociedade contemporânea, bem como, investigar o desenvolvimento infantil considerando as diferentes fases da vida do sujeito, discorrer o papel da família frente ao desenvolvimento, considerando as manifestações precoces dessa fase e, identificar o aparecimento da erotização precoce e a influência do meio frente a tal aparecimento. É possível perceber, a partir do estudo, que intensas discussões têm surgido frente ao desenvolvimento infantil, considerando inúmeras teorias que buscam refletir e discutir a temática. Sabe-se que a sexualidade é algo marcante em todas etapas da vida de cada um, de modo que a família assume papel primordial no sentido de contribuir ao sujeito melhores condições para lidar com essa questão. Percebe-se também, grande contribuição por parte da família e sociedade frente ao processo de estimulação precoce, onde cada vez mais crianças adentram o mundo adulto mesmo que de maneira indireta, de onde se tornam visíveis comportamentos incomuns à respectiva fase

Comparação entre Algoritmos de Aprendizado de Máquina para a Identificação de Floresta Tropical Sazonalmente Seca

O objetivo do estudo foi avaliar o potencial dos algoritmos de aprendizado de máquina k-Nearest Neighbor (kNN), Random Forest (RF), Support Vector Machine (SVM) e Artificial Neural Networks (ANN) na identificação das fitofisionomias da Caatinga a partir de imagens OLI/Landsat 8. Para tanto, foi elaborado um arquivo vetorial de treinamento com as amostras dos diferentes padrões dos usos e cobertura da terra, a fim de orientar os algoritmos no processo de classificação. A validação das classificações foi obtida por meio da validação cruzada, do tipo k-fold, com 30 repetições, sendo avaliada a qualidade da classificação a partir dos valores expressos pelo coeficiente Kappa. Para verificar a existência de diferenças significativas entre os algoritmos foi aplicado o teste estatístico de Friedman e Nemenyi. O algoritmo RF apresentou os maiores valores para o coeficiente Kappa, expressando um valor médio de 0,9841. Por outro lado, a ANN demonstrou desempenho inferior aos demais, englobando um valor médio de 0,7551, ocasionado pela confusão espectral na identificação da classe nuvem/sombra com a classe água. Apesar de todos os algoritmos testados apresentarem bons resultados, o algoritmo RF diferiu significativamente dos demais, expressando resultados superiores quando aplicado à identificação de padrões espaciais na Caatinga. Conclui-se que o uso de algoritmos de aprendizagem de máquina é eficiente na identificação de fitofisionomias da Caatinga, com destaque para o RF, o qual englobou melhor a variação dos padrões espectrais dos usos, podendo ser utilizado para estudos posteriores relacionados com a Caatinga.

Beira Interior Norte

O presente trabalho visa caracterizar a indústria agroalimentar utilizadora de sistemas de frio, na região da Beira Interior Norte. O estudo efetuado tem por base o levantamento de dados e características energéticas das empresas participantes num processo de inquirição e análise. As empresas do setor agroindustrial analisadas encontram-se distribuídas por cinco fileiras: carnes, lácteos, vinho, distribuição e hortofrutícola. O objetivo do artigo consiste na comparação de indicadores energéticos relativos ao consumo de energia eléctrica das empresas do mesmo setor e entre fileiras, permitindo analisar o desempenho energético das empresas e dos setores. Sendo o sector agroalimentar responsável por elevados padrões de consumo de energia, a utilização racional de energia parte da análise de trabalhos desta índole, a fim de promover medidas de eficiência energética no sector.info:eu-repo/semantics/publishedVersio

Collagen membrane from bovine pericardium for treatment of long bone defect

The treatment of bone regeneration failures has been constantly improved with the study of new biomaterials. Techgraft® is a collagen membrane derived from bovine pericardium, which has been shown to have biocompatibility and effectiveness in tissue repair. However, its use in orthopedics has not yet been evaluated. Therefore, the purpose of this study was to characterize a bovine pericardium collagen membrane and evaluate the effects of its use in the regeneration of a bone defect in rat tibia. Scanning electron microscopy, atomic force microscopy, weight lost and water uptake tests, and mechanical test were performed. Afterwards, the membrane was tested in an experimental study, using 12 male Sprague Dawley rats. A bone defect was surgically made in tibiae of animals, which were assigned to two groups (n = 6): bone defect treated with collagen membrane (TG) and bone defect without treatment (CONT). Then, tibiae were submitted to micro-CT. The membranes preserved their natural collagen characteristics, presenting great strength, high water absorption, hydrophilicity, and almost complete dissolution in 30 days. In the experimental study, the membrane enhanced the growth of bone tissue in contact with its surface. A higher bone volume and trabeculae number and less trabecular space was observed in bone defects of the membrane group compared to the control group at 21 days. In conclusion, the Techgraft membrane seems to have favorable characteristics for treatment of long bone repair.The authors gratefully acknowledge the financial support from the São Paulo Research Foundation (reference 2017/20051-0) and the scholarship from the Coordination for the Improvement of Higher Education Personnel

merlin, an improved framework for the reconstruction of high-quality genome-scale metabolic models

Genome-scale metabolic models have been recognised as useful tools for better understanding living organisms metabolism. merlin (https://www.merlin-sysbio.org/) is an open-source and user-friendly resource that hastens the models reconstruction process, conjugating manual and automatic procedures, while leveraging the user's expertise with a curation-oriented graphical interface. An updated and redesigned version of merlin is herein presented. Since 2015, several features have been implemented in merlin, along with deep changes in the software architecture, operational flow, and graphical interface. The current version (4.0) includes the implementation of novel algorithms and third-party tools for genome functional annotation, draft assembly, model refinement, and curation. Such updates increased the user base, resulting in multiple published works, including genome metabolic (re-)annotations and model reconstructions of multiple (lower and higher) eukaryotes and prokaryotes. merlin version 4.0 is the only tool able to perform template based and de novo draft reconstructions, while achieving competitive performance compared to state-of-the art tools both for well and less-studied organisms.Centre of Biological Engineering (CEB, UMinho); Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit; this work is a result of the project 22231/01/SAICT/2016: Biodata.pt Infraestrutura Portuguesa de Dados Biologicos, supported by the ´ PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF); FCT for providing PhD scholarships [DFA/BD/08789/2021 J.C, DFA/BD/8076/2020 E.C., SFRH/BD/139198/2018 to F.C., SFRH/BD/131916/2017 R. Rodrigues]; FCT for the Assistant Research contract of Oscar Dias obtained under CEEC Individual 2018.info:eu-repo/semantics/publishedVersio

Combined Impact of Sample Size and Modeling Approaches for Predicting Stem Volume in Eucalyptus spp. Forest Plantations Using Field and LiDAR Data

Light Detection and Ranging (LiDAR) remote sensing has been established as one of the most promising tools for large-scale forest monitoring and mapping. Continuous advances in computational techniques, such as machine learning algorithms, have been increasingly improving our capability to model forest attributes accurately and at high spatial and temporal resolution. While there have been previous studies exploring the use of LiDAR and machine learning algorithms for forest inventory modeling, as yet, no studies have demonstrated the combined impact of sample size and different modeling techniques for predicting and mapping stem total volume in industrial Eucalyptus spp. tree plantations. This study aimed to compare the combined effects of parametric and nonparametric modeling methods for estimating volume in Eucalyptus spp. tree plantation using airborne LiDAR data while varying the reference data (sample size). The modeling techniques were compared in terms of root mean square error (RMSE), bias, and R2 with 500 simulations. The best performance was verified for the ordinary least-squares (OLS) method, which was able to provide comparable results to the traditional forest inventory approaches using only 40% (n = 63; ~0.04 plots/ha) of the total field plots, followed by the random forest (RF) algorithm with identical sample size values. This study provides solutions for increasing the industry efficiency in monitoring and managing forest plantation stem volume for the paper and pulp supply chain