In schizophrenia, subtle aberrations in the brain cause functional disturbances like psychotic symptoms and social disability. There are, however, also disturbances outside the CNS indicating a systemic manifestation in the disease. The aim of the present thesis was to gain deeper understanding of the pathophysiological mechanisms underlying schizophrenia with a particular interest in peripheral and systemic manifestations with relevance for the increased risk of obesity and metabolic complications seen in the disease. Therefore, resting energy expenditure (REE), physical capacity, and relevant body composition variables were measured in patients with schizophrenia and in healthy controls. Also niacin skin flush response and electrodermal activity (EDA) were studied. Patients with schizophrenia exhibited significantly lower REE expressed as kJ/kg, and also lower values compared with predicted levels than the controls. The difference could not be attributed to medication or variations in body composition between the two groups. There was a gender difference with the lowest levels found in male patients. Male patients exhibited significantly lower physical capacity in terms of predicted maximal oxygen uptake capacity and faster increase in respiratory quotient than male controls. The oral niacin test revealed a significantly delayed skin flush reaction in patients compared with controls. The patients also exhibited lower EDA response. There was a significant association in response patterns for the niacin and the EDA tests in the patients, but not in controls. In a test-retest study in patients there was acceptable stability for EDA measures but low test-retest stability for niacin variables. The previously found association in responses for the two tests was, however, replicated. The results gain support for the concept of schizophrenia as a disease with systemic manifestations including metabolic dysregulation. The findings add to the understanding of the weight gain and the increased risk for cardiovascular morbidity seen in this condition.

Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Being the harvest time of apples concentrated in a short period, fruit storage is needed for market regulation. New storage technologies are being developed, such as dynamic controlled atmosphere with respiratory quotient (DCA-RQ), that needs to be evaluated under different temperatures and with the application of growth regulators such as aminoethoxyvinylglycine (AVG), naphthalene acetic acid (NAA) and 1-methylcyclopropene (1-MCP). This thesis is composed of four papers, which resulted from studies with the following objectives: to evaluate volatile compounds concentration in ‘Galaxy’ and ‘Royal Gala’ apples stored under DCA-RQ, DCA with chlorophyll fluorescence (DCA-CF), controlled atmosphere (CA) with and without 0,625 μL L-1 of 1-MCP application; to evaluate if DCA-RQ and DCA-CF allow employing higher temperatures during apple storage; to evaluate the effect of the application of AVG (0.83 kg ha-1) in combination with NAA (40g ha-1), as well as the effects of 1-MCP in the production of volatile compounds and physical-chemical quality of apples stored under DCA monitored with chlorophyll fluorescence and respiratory quotient; identify if the stress by low O2 partial pressure during RQ measurement has an effect on apple quality conservation. ‘Galaxy’ and ‘Royal Gala’ apples were stored for 9 months plus 7 days at 20 °C. It is worth noting that ‘Galaxy’ apples stored under DCA-RQ1.5 maintained better quality when compared to CA, ultralow oxygen (ULO – 0.4kPa) and DCA-CF, also increasing the production of esters responsible for the aroma. Besides, 1-MCP reduced volatile compounds concentration, even in apples stored under DCA-RQ1.5. Likewise, DCA-CF had a similar effect to ULO with 0.4 kPa of O2. The DCA-RQ1.3 and DCA-CF allowed storing ‘Galaxy’ apples under temperatures higher (2.0 and 2.5 °C) than recommended (1.5 °C). In addition, 1-MCP application did not have additional effects when fruit were stored under DCA and higher temperatures. Higher temperatures (2.0 and 2.5 °C) could be used without reducing the production of important volatile compounds. In DCA-RQ, the stress by low O2 employed during RQ measurement has presented a beneficial effects, with fruit maintaining higher flesh firmness, lower mealiness and decay incidence compared to fruit without RQ calculation. ‘Royal Gala’ apples stored under DCA-RQ1.3 and DCA-CF, submitted to the application of AVG and NAA during prehavest did show an increase in the concentration of volatile compounds. However, the combination of AVG plus NAA and the storage under DCA-RQ1.3 or DCA-CF presents an excellent alternative for storing apples, because maintains higher flesh firmness, reduce physiological disorders and maintain higher percentage of healthy fruit. / Devido à colheita de maçãs ser concentrada em curto período, faz-se necessário o armazenamento de uma grande parte dos frutos para regulação do mercado. Existem tecnologias de armazenamento em desenvolvimento, como a atmosfera controlada dinâmica com quociente respiratório (ACD-QR), a qual precisa ser avaliada em diferentes temperaturas e em frutos com aplicação de fitorreguladores como o 1-metilciclopropeno (1-MCP), a aminoetoxivinilglicina (AVG) e o ácido naftaleno acético (ANA). Esta tese é composta por quatro artigos científicos que resultaram de trabalhos com os seguintes objetivos: Avaliar a produção de compostos voláteis em maçãs ‘Galaxy’ e ‘Royal Gala’ armazenadas em ACD-QR, atmosfera controlada dinâmica com fluorescência de clorofilas (ACD-FC), atmosfera controlada (AC) convencional e com aplicação de 1-MCP; avaliar se a ACD-QR e a ACD-FC permitem utilizar temperaturas mais elevadas para o armazenamento das maçãs; avaliar o efeito da aplicação da AVG (0,83 kg ha-1) em combinação com o ANA (40g ha-1), bem como o efeito do 1-MCP na produção de compostos voláteis e qualidade físico-química dos frutos armazenados em ACD monitorada com fluorescência de clorofilas e com quociente respiratório; identificar se o estresse por baixa pressão parcial de O2 durante o período de cálculo do quociente respiratório possui efeito na conservação da qualidade de maçãs. Foram utilizadas maçãs ‘Galaxy’ e ‘Royal Gala’, armazenadas por nove meses mais sete dias de exposição à temperatura de 20 °C. Pode-se destacar que a maçã ‘Galaxy’ quando armazenada em ACD com QR 1,5 (ACD-QR1,5) manteve melhor qualidade, do que o armazenamento em AC convencional com pressões parciais de O2 ultrabaixas (ULO – 0,4 kPa CO2) e em ACD-FC, além de aumentar a produção de ésteres responsáveis pela formação do aroma. Além disso, a aplicação de 0,625 μL L-1 de 1-MCP reduziu a concentração de compostos voláteis, mesmo em frutos armazenados em ACD-QR1,5. A ACD-FC teve efeito similar ao armazenamento em ULO com 0,4 kPa de O2. A ACD-QR1,3 e a ACD-FC permitiram armazenar a maçã ‘Galaxy’ em temperatura mais elevada (2,0 e 2,5 °C) do que a normalmente recomendada (1,5 °C). Além disso, a aplicação de 1-MCP não teve efeito adicional em maçãs armazenadas com temperatura mais elevada quando em ACD. A alta temperatura (2,0 e 2,5 °C) pôde ser usada em ACD-QR1,3 sem perda na produção de compostos voláteis importantes. No armazenamento em ACD-QR, o estresse causado pelas baixas pressões parciais de O2 a que os frutos são submetidos durante o período de cálculo do QR (13 h), que pode atingir 0,0 kPa, possuiu efeito benéfico, pois manteve firmeza da polpa mais elevada, menor incidência de polpa farinácea e podridões, comparado aos frutos que não sofreram esse estresse por baixo O2. O armazenamento de maçãs ‘Royal Gala’ em ACD-QR1,3 e ACD-FC tratadas em pré-colheita com AVG e ANA não aumentaram a concentração de compostos voláteis. Entretanto, a aplicação associada de AVG e ANA e o armazenamento em ACD-QR1,3 ou ACD-FC foram excelentes alternativas para a conservação dos frutos, pois mantiveram a firmeza da polpa, reduziram os distúrbios fisiológicos, proporcionando maior volume de frutos sadios.

Le métabolisme est la somme des réactions chimiques dans un organisme. L’énergie leur permet d’effectuer la biosynthèse, le maintien des fonctions vitales et l’activité physique. Si l’énergie est transformée en chaleur nous pouvons déterminer le taux de transformation d’énergie chimique en taux métabolique (TM) et nous pouvons donc calculer les besoins énergétiques d’un animal. Le principal objectif de ce travail est d’évaluer le métabolisme énergétique chez la punaise hématophage Rhodnius prolixus. Nos résultats montrent, que le TM de l’alimentation chez R. prolixus atteint jusqu’à 17 fois le TM au repos. Le quotient respiratoire est de 0,83 pendant le repos et 0,52 pendant l’alimentation. De plus, la désactivation des protéines permettant gérer le choc thermique, diminuent le TM pendant la digestion. Le coût énergétique (CE) de la marche peut atteindre jusqu’à 1,7 fois le CE pendant le repos. Finalement, le CE de la production d’un oeuf chez R. prolixus est approximativement de 11,7 J. L’ensemble de nos résultats apportent des outils qui fournissent des éléments nécessaires pour mieux contrôler la transmission de maladies vectorielles. / Metabolism is the sum of all the chemical reactions in an organism. Energy uptake allows animals to perform biosynthesis, maintenance, and external work. If the energy produced is converted into heat, we can estimate the energy metabolism as the rate of conversion of chemical energy into metabolic rate (MR), which allows the calculation of energy requirements. The main objective of this work is to assess the metabolic cost of physiological work in the blood-sucking bug Rhodnius prolixus. Our results show that feeding is costly. The MR during feeding in R. prolixus reaches up to 17 times the MR during rest. The mean respiratory quotient is 0.83 during rest and 0.52 during feeding. We showed that the deactivation of Heat Shock Proteins on R. prolixus, causes a diminution of MR during digestion. Then, we showed that the energy cost (EC) during walking can reach up to 1.7 times the EC during rest. Finally, we found that the EC of production of one egg of R. prolixus was 11.7 J. All our results provide tools to a better understanding of biology and ecology of an hematophagous insect to provide the necessary elements to better control of transmission of vector-borne diseases.

Insufficient sleep may be associated with obesity via increased energy intake and/or decreased energy expenditure. The present study therefore aimed to investigate effects of sleep restriction on energy balance in healthy young adults. Participants (14 men, 13 women) aged 35.3 ± 1.0 y with 23.6 ± 0.2 kg/m2 BMI completed a randomized, crossover study exposed to short and habitual sleep with 4 wk washout. Controlled diets were provided during the first 4 d, followed by 2 d of ad libitum eating. Ad libitum energy intake, energy expenditure and physical activity level were determined as well as energy balance and body weight. Results showed that ad libitum energy intake (p = 0.031), as well as total fat (p = 0.018) increased after short compared with habitual sleep, but physical activity level, energy expenditure, energy balance, and body weight remained unaffected by sleep duration. In conclusion, sleep deprivation elevates energy intake, which may lead to positive energy balance over time and increase the risk of weight gain and/or obesity.

Die Belastung des Energiestoffwechsels der Hochleistungskuh ist in der peripartalen Phase am größten. Die Regulation der Futteraufnahme und des Energiestoffwechsels durch den Nucleus arcuatus (ARC) des Hypothalamus spielt eine entscheidende Rolle während dieser Phase. Zahlreiche Metabolite und Hormone, wie z.B. das Peptidhormon Ghrelin, beeinflussen die Expression des orexigenen (futteraufnahmesteigernden) Neuropeptids Agouti-related Protein (AgRP) im ARC des Hypothalamus. Das Ziel dieser Arbeit war es, den Zusammenhang zwischen Körperfettmobilisierung und orexigenen Signalen im peripartalen Zeitraum der Hochleistungskuh näher zu untersuchen. Hierfür wurden 20 multipare Hochleistungskühe der Rasse Deutsch-Holstein (2.-4. Laktation) 7 Wochen ante partum (ap) bis 6 Wochen post partum (pp) untersucht. Die Tiere wurden in Anbindehaltung aufgestallt und entsprechend der jeweiligen Produktionsperiode bedarfsgerecht energetisch versorgt. Die ad libitum Futteraufnahme und Milchleistung wurden täglich gemessen und die Milchzusammensetzung wöchentlich analysiert. Das Körpergewicht und die Rückenfettdicke (RFD) wurden ebenfalls wöchentlich bestimmt. Einmal wöchentlich wurden Blutproben genommen, um die Konzentration von nicht-veresterten Fettsäuren (NEFA), Triglyceriden (TG) und Aminosäuren zu bestimmen. Eine Leberbiopsie wurde am 34. Tag ap und am 3., 18. und 30. Tag pp entnommen. In der 5. Woche ap und in der 2. Woche pp wurden die Tiere in eine Respirationskammer eingestallt und darin jeweils am 1. Tag ad libitum versorgt, während ihnen am 2. Tag das Futter für 10 h entzogen wurde. Die Futteraufnahme bzw. die kompensatorische Futteraufnahme nach Futterentzug wurden ebenfalls gemessen. Mit Hilfe der indirekten Kalorimetrie wurden der Sauerstoffverbrauch, die Bildung von Kohlenstoffdioxid und Methan gemessen, und der Respiratorische Quotient (RQ), die Fettoxidation (FOX) und die Kohlenhydratoxidation (KOX) berechnet. An beiden Tagen des Aufenthaltes in der Respirationskammer wurden stündlich Blutproben entnommen und die Konzentration von Acyl- und Gesamtghrelin, NEFA und TG bestimmt. Am 40. Tag pp wurden die Tiere geschlachtet und der ARC entnommen. In Studie 1 wurden 16 Tiere, basierend auf ihren Leberfettgehalt (LFC) am 18. Tag pp in eine Gruppe mit hohem LFC (H, n=8) und eine mit niedrigem LFC (L, n=8) eingeteilt. Für die Studie 2 wurden 18 Kühe entsprechend ihrer NEFA-Blutplasmakonzentration am Schlachttag in eine Gruppe mit hoher NEFA- (H, n=9) und eine mit niedriger NEFA-Plasmakonzentration (L, n=9) eingeteilt. In Studie 1 konnte gezeigt werden, dass die Acyl- und Gesamtghrelin- Plasmakonzentrationen nicht mit der pp Futteraufnahmesteigerung von Hochleistungskühen korrelierten. H-Kühe, die im Vergleich zu L-Kühen einen höheren Leber- und Milchfettgehalt, eine größere RFD und einen geringeren RQ aufwiesen, zeigten während des 10-stündigen Futterentzuges den größeren Anstieg der Acylghrelinkonzentration sowie ein größeres Acyl- und Gesamtghrelin-Verhältnis. Signifikante Korrelationen zwischen dem präprandialen Acyl:Gesamtghrelin- Verhältnis und zahlreichen Parametern des Fettstoffwechsels, wie bspw. LFC, Milchfettgehalt, RQ und RFD, lassen einen Zusammenhang zwischen Ghrelin, dem Fettstoffwechsel und der Fettverteilung erkennen. In Studie 2 konnte nachgewiesen werden, dass die in der Frühlaktation auftretende unterschiedliche Aktivierung hypothalamischer AgRP-Neurone von H- und L-Kühen nicht mit deren Futteraufnahme assoziiert ist. Die höhere NEFA-Plasmakonzentration, die höhere RFD, die höhere FOX und der höhere Sauerstoffverbrauch der H-Kühe waren jeweils signifikant mit der geringeren Aktivierung hypothalamischer AgRP-Neurone korreliert. Diese Korrelationen belegen einen Zusammenhang zwischen dem prozentualen Anteil aktivierter AgRP-Neurone und dem Sauerstoffverbrauch sowie der Substratverstoffwechselung während der Frühlaktation. Zusammenfassend lässt sich schlussfolgern, dass die untersuchten Signale des orexigenen Systems im peripartalen Zeitraum der Hochleistungskuh nicht mit der Futteraufnahme, jedoch mit dem Fett- und Energiestoffwechsel assoziiert waren. Ferner lassen die Resultate den Schluss zu, dass die Futteraufnahme bereits vor der Kalbung durch den Körperfettgehalt determiniert ist, und dass die Fettmobilisierung per se kaum einen Einfluss auf die Futteraufnahmesteigerung in der Frühlaktation besitzt.

Insufficient sleep may be associated with obesity via increased energy intake and/or decreased energy expenditure. The present study therefore aimed to investigate effects of sleep restriction on energy balance in healthy young adults. Participants (14 men, 13 women) aged 35.3 ± 1.0 y with 23.6 ± 0.2 kg/m2 BMI completed a randomized, crossover study exposed to short and habitual sleep with 4 wk washout. Controlled diets were provided during the first 4 d, followed by 2 d of ad libitum eating. Ad libitum energy intake, energy expenditure and physical activity level were determined as well as energy balance and body weight. Results showed that ad libitum energy intake (p = 0.031), as well as total fat (p = 0.018) increased after short compared with habitual sleep, but physical activity level, energy expenditure, energy balance, and body weight remained unaffected by sleep duration. In conclusion, sleep deprivation elevates energy intake, which may lead to positive energy balance over time and increase the risk of weight gain and/or obesity.

La première partie de mon travail de thèse a été de définir le rôle joué par le récepteur nucléaire Rev-erb alpha dans les mécanismes de synchronisation par la nourriture d’une horloge circadienne putative, non encore localisée, appelée « horloge alimentaire ». La seconde partie de mon travail a consisté à étudier la participation de Rev-erb alpha dans les régulations des métabolismes glucidique et lipidique. L’ensemble de nos données indique que le répresseur transcriptionnel Rev-erb alpha joue un rôle charnière dans les fonctions circadiennes ainsi que dans le métabolisme. En effet, d’un point de vue circadien, l’absence de Rev-erb alpha altère la synchronisation à l’heure des repas – démontré par une réduction des sorties comportementales et physiologiques de l’horloge alimentaire, ainsi que par l’absence d’ajustement du rythme de la protéine d’horloge PER2 dans l’oscillateur cérébelleux. Sur le plan métabolique, la délétion de ce gène modifie notamment le métabolisme des lipides – démontré par une accumulation excessive de tissu adipeux, une utilisation préférentielle des acides gras, ainsi qu’une perte de contrôle de l’expression de la Lipoprotéine lipase. / The work performed during this PhD thesis aimed at investigating the role of the transcriptional silencer Rev-erbα in both the circadian clockwork of the food-entrainable oscillator and metabolic regulations. Firstly, by evaluating food-anticipatory components in animals fed once a day at the same time, we showed that mice lacking Rev-erbα display a reduction in locomotor activity prior to food access compared to littermate controls. Accordingly, the rises in body temperature and corticosterone that anticipate mealtime are also diminished. Interestingly, daily p-ERK expression in hypothalamic regions and daily PER2 expression in the cerebellum of Rev-erbα KO mice are not phase-adjusted to feeding time. These results indicate that Rev-erbα participates in the integration of feeding signals and in food-seeking behaviors. Secondly, by investigating energy balance in fasted, normal chow or high-fat fed animals, we revealed that Rev-erbα KO mice exhibit greater reliance on lipid fuels as energy substrates, contributing to a mild hyperglycemic state. We also found that Lipoprotein lipase (Lpl) expression, is strongly up-regulated in peripheral tissues of Rev-erbα KO mice, predisposing mice to obesity. In this regard, we uncovered a new molecular pathway that ties clock-driven Lpl expression to energy homeostasis. These findings highlight the significance of daily Rev-erbα oscillations to prevent the appearance of the metabolic syndrome.In conclusion, we provide evidence that REV-ERBα may be a part of the food-entrainable oscillator clockwork that triggers food-anticipatory components, and represents a pivotal player to link the core clock machinery to metabolic pathways.

Die Belastung des Energiestoffwechsels der Hochleistungskuh ist in der peripartalen Phase am größten. Die Regulation der Futteraufnahme und des Energiestoffwechsels durch den Nucleus arcuatus (ARC) des Hypothalamus spielt eine entscheidende Rolle während dieser Phase. Zahlreiche Metabolite und Hormone, wie z.B. das Peptidhormon Ghrelin, beeinflussen die Expression des orexigenen (futteraufnahmesteigernden) Neuropeptids Agouti-related Protein (AgRP) im ARC des Hypothalamus. Das Ziel dieser Arbeit war es, den Zusammenhang zwischen Körperfettmobilisierung und orexigenen Signalen im peripartalen Zeitraum der Hochleistungskuh näher zu untersuchen. Hierfür wurden 20 multipare Hochleistungskühe der Rasse Deutsch-Holstein (2.-4. Laktation) 7 Wochen ante partum (ap) bis 6 Wochen post partum (pp) untersucht. Die Tiere wurden in Anbindehaltung aufgestallt und entsprechend der jeweiligen Produktionsperiode bedarfsgerecht energetisch versorgt. Die ad libitum Futteraufnahme und Milchleistung wurden täglich gemessen und die Milchzusammensetzung wöchentlich analysiert. Das Körpergewicht und die Rückenfettdicke (RFD) wurden ebenfalls wöchentlich bestimmt. Einmal wöchentlich wurden Blutproben genommen, um die Konzentration von nicht-veresterten Fettsäuren (NEFA), Triglyceriden (TG) und Aminosäuren zu bestimmen. Eine Leberbiopsie wurde am 34. Tag ap und am 3., 18. und 30. Tag pp entnommen. In der 5. Woche ap und in der 2. Woche pp wurden die Tiere in eine Respirationskammer eingestallt und darin jeweils am 1. Tag ad libitum versorgt, während ihnen am 2. Tag das Futter für 10 h entzogen wurde. Die Futteraufnahme bzw. die kompensatorische Futteraufnahme nach Futterentzug wurden ebenfalls gemessen. Mit Hilfe der indirekten Kalorimetrie wurden der Sauerstoffverbrauch, die Bildung von Kohlenstoffdioxid und Methan gemessen, und der Respiratorische Quotient (RQ), die Fettoxidation (FOX) und die Kohlenhydratoxidation (KOX) berechnet. An beiden Tagen des Aufenthaltes in der Respirationskammer wurden stündlich Blutproben entnommen und die Konzentration von Acyl- und Gesamtghrelin, NEFA und TG bestimmt. Am 40. Tag pp wurden die Tiere geschlachtet und der ARC entnommen. In Studie 1 wurden 16 Tiere, basierend auf ihren Leberfettgehalt (LFC) am 18. Tag pp in eine Gruppe mit hohem LFC (H, n=8) und eine mit niedrigem LFC (L, n=8) eingeteilt. Für die Studie 2 wurden 18 Kühe entsprechend ihrer NEFA-Blutplasmakonzentration am Schlachttag in eine Gruppe mit hoher NEFA- (H, n=9) und eine mit niedriger NEFA-Plasmakonzentration (L, n=9) eingeteilt. In Studie 1 konnte gezeigt werden, dass die Acyl- und Gesamtghrelin- Plasmakonzentrationen nicht mit der pp Futteraufnahmesteigerung von Hochleistungskühen korrelierten. H-Kühe, die im Vergleich zu L-Kühen einen höheren Leber- und Milchfettgehalt, eine größere RFD und einen geringeren RQ aufwiesen, zeigten während des 10-stündigen Futterentzuges den größeren Anstieg der Acylghrelinkonzentration sowie ein größeres Acyl- und Gesamtghrelin-Verhältnis. Signifikante Korrelationen zwischen dem präprandialen Acyl:Gesamtghrelin- Verhältnis und zahlreichen Parametern des Fettstoffwechsels, wie bspw. LFC, Milchfettgehalt, RQ und RFD, lassen einen Zusammenhang zwischen Ghrelin, dem Fettstoffwechsel und der Fettverteilung erkennen. In Studie 2 konnte nachgewiesen werden, dass die in der Frühlaktation auftretende unterschiedliche Aktivierung hypothalamischer AgRP-Neurone von H- und L-Kühen nicht mit deren Futteraufnahme assoziiert ist. Die höhere NEFA-Plasmakonzentration, die höhere RFD, die höhere FOX und der höhere Sauerstoffverbrauch der H-Kühe waren jeweils signifikant mit der geringeren Aktivierung hypothalamischer AgRP-Neurone korreliert. Diese Korrelationen belegen einen Zusammenhang zwischen dem prozentualen Anteil aktivierter AgRP-Neurone und dem Sauerstoffverbrauch sowie der Substratverstoffwechselung während der Frühlaktation. Zusammenfassend lässt sich schlussfolgern, dass die untersuchten Signale des orexigenen Systems im peripartalen Zeitraum der Hochleistungskuh nicht mit der Futteraufnahme, jedoch mit dem Fett- und Energiestoffwechsel assoziiert waren. Ferner lassen die Resultate den Schluss zu, dass die Futteraufnahme bereits vor der Kalbung durch den Körperfettgehalt determiniert ist, und dass die Fettmobilisierung per se kaum einen Einfluss auf die Futteraufnahmesteigerung in der Frühlaktation besitzt.

Consistently, reports in the literature have identified that a sedentary lifestyle contributes to the progression of a range of chronic degenerative diseases. The measurement of energy expenditure and physical activity pattern in children is a challenge for all professionals interested in paediatric health and from a broader perspective, the public health fraternity charged with considering longer term health consequences of physical inactivity. The primary objective of this thesis was to identify a suitable indirect and objective measurement technique for the assessment of energy expenditure and physical activity pattern in children. The ideal characteristics of such a technique are that it should be reproducible and have been validated against a criterion reference method. To achieve this goal, a series of methodological studies were undertaken (Chapters II and III). This work was essential to increase accuracy during the individualised laboratory calibration process and further minimise prediction errors when analysing data from 7 days of monitoring under free-living conditions in the second part of the study (Chapters IV and V). In the first study to verify the combined effect of body position, apparatus and distraction on children's resting metabolic rate (RMR), experiments were carried out on 14 children aged 8-12 (mean age = 10.1 years ± 1.4). Each participant underwent 2 test sessions, one week apart under three different situations: a) using mouthpiece and nose-clip (MN) or facemask (FM); b) sitting (SEAT) or lying (LY) and c) TV viewing (TV) or no TV viewing. In the first session, following 20 min rest and watching TV, the following protocol was used: LY: 20 min - stabilisation; 10 min using MN and 10 min using FM. Body position was then changed to seated: 20 min stabilisation; 10 min using FM; 10 min using MN. In the second session, FM and MN order was changed and participants did not watch TV. Data were analysed according to the eight combinations among the three studied parameters. Repeated measures ANOVA indicated statistically significant differences for &VO2 (p=0.01) and RMR (p=0.02), with TVMNSEAT showing higher values than TVFMLY. Bland-Altman analysis showed a bias for &VO2, &VCO2, RQ and RMR between TVFMLY and TVMNSEAT of -17.8±14.5 ml.min-1, -8.8±14.5 ml. min-1, 0.03±0.05 and -115.2±101.9 kcal.d-1, respectively. There were no differences in RMR measurements due to body position and apparatus when each variable was isolated. Analyses of distraction in three of four combinations indicated no difference between TV and no TV. In summary, different parameter combinations can result in increased bias and variability and thereby reported differences among children's RMR measurement. The second study dealt with treadmill adaptation and determination of self-selected (SS) walking speed. Assessment of individual and group differences in metabolic energy expenditure using oxygen uptake requires that individuals are comfortable with, and can accommodate to, the equipment being utilised. In this study, a detailed proposal for an adaptation protocol based on the SS was developed. Experiments were carried out on 27 children aged 8-12 (mean age = 10.3±1.2 yr). Results from three treadmill tests following the adaptation protocol showed similar results for step length with no significant differences among tests and lower and no statistically significant variability within- and between-days. Additionally, no statistically significant differences between SS determined over-ground and on a treadmill were verified. These results suggest that SS speed determined over-ground is reproducible on a treadmill and the 10 min familiarisation protocol based on this speed provided sufficient exposure to achieve accommodation to the treadmill. The purpose of the third study was to verify within- and between-day repeatability and variability in children's oxygen uptake ( &VO2), gross economy (GE) [ &VO2 divided by speed] and heart rate (HR) during treadmill walking based on SS. 14 children (mean age = 10.2±1.4 yr) undertook 3 testing sessions over 2 days in which four walking speeds, including SS, were tested. Within- and between-day repeatability was assessed using the Bland and Altman method and coefficients of variability (CV) were determined for each child across exercise bouts and averaged to obtain a mean group CV value for &VO2, GE and HR per speed. Repeated measures ANOVA showed no statistically significant differences in within- or between-day CV for &VO2, GE or HR at any speed. Repeatability within and between-day for &VO2, GE and HR for all speeds was verified. These results suggest that submaximal &V O2 during treadmill walking is stable and reproducible at a range of speeds based on children's SS. In the fourth study, the objective was to establish the effect of walking speed on substrate oxidation during a treadmill protocol based on SS. Experiments were carried out on 12 girls aged 8-12 (mean age = 9.9±1.4 yr). Each participant underwent 2 test sessions, one week apart. Workloads on the treadmill included 2 speeds slower than SS (1.6 [V1] and 0.8 km.h-1 [V2] slower than SS), SS (V3), and a speed 0.8 km.h-1 faster than SS (V4). Indirect calorimetry from respired gas measurements enabled total fat (FO) and carbohydrate (CHO) oxidation rates to be calculated according to the non-protein respiratory quotient (Peronnet and Massicote, 1991) and percentage of CHO and FO calculations using equations from McGilvery and Goldstein (1983). Repeated measures ANOVA followed by a Tukey Post Hoc test (p< 0.05) was used to verify differences in CHO and FO rates among speeds. Paired T-test was used to verify differences in CHO and FO rates between tests per velocity. The reliability between-day was assessed using intraclass correlation coefficient (ICC). Results showed significant differences for CHO among all speeds, as well as significant differences for FO between V1 and V2 against V3 and V4 in both tests. Analyses between trials per velocity showed no significant substrate use differences as well as acceptable reliability. At the self-selected speed (V3) there was an accentuation in FO reduction as well as an increase in CHO oxidation. The purpose of the fifth study was to determine whether there were differences in substrate oxidation between girls (G) and women (W) during a treadmill protocol based on SS. Experiments were carried out on 12 G aged 8-12 (mean age = 9.9±1.4 yr) and 12 W aged 25-38 (mean age = 32.3±3.8 yr). The treadmill protocol included 6 min workloads followed by 5 min rest periods. Workloads included 2 speeds slower than SS (1.6 (V1) and 0.8 km.h-1 (V2) slower than SS), SS (V3), and a speed 0.8 km.h-1 faster than SS (V4). Total fat and carbohydrate (CHO) oxidation rates were calculated from indirect calorimetry according to the non-protein respiratory quotient. Repeated measures ANOVA followed by a Tukey Post Hoc test was used to verify intra-test differences in CHO and fat oxidation rates among speeds. Intergroup differences were analysed using paired T-test. Fat utilisation in W achieved a plateau at a relative velocity 0.8 km.h-1 slower than SS, but for G, fat utilisation increased until SS, and then stabilised upon reaching the higher velocity. CHO oxidation curves rose abruptly above V2 for W, while for G the acute increase occurred after SS (V3). Collectively, these results indicate that as walking intensity increases G are able to meet the energy demands of the work by increasing fat oxidation together with the increased CHO oxidation up to SS. In contrast for W, increasing CHO oxidation is associated with an early decrease in fat utilisation at a velocity slower than the self-selected speed. The sixth study dealt with validation of indirect techniques for the measurement of energy expenditure in free-living conditions against the DLW technique. Experiments were carried out on 19 children aged 8-12 (mean age = 10.3±1.0 yr). To indirectly predict energy expenditure 12 different procedures were used. Only one procedure, combining activity and heart rate (AHbranched), was based on a group equation, the others were based on individualised regression. Three of the individually-based techniques were able to accurately predict energy expenditure in free-living conditions. These tecniques were HRPAnetRMR using HRnet [HR exercise minus sleep HR (SHR)] against PAnet (measured PA exercise minus measured RMR) and upper and lower body equations corrected by RMR; HRPAnet4act using the same procedure but corrected by the mean resting &VO2 for 4 resting activities [(4act) = supine watching TV, sitting watching TV, sitting playing computer games and standing], and HRPALBnet4act using only lower body activities and corrected by 4act. HRPAnetRMR was only slightly more accurate than HRPAnet4act and HRPALBnet4act, but this technique is only adjusted by RMR whereas the other two are heavily dependent on more complex laboratory calibration. Bland and Altman (1986) analyses showed no significant differences between AHbranched predicted and measured TEE using the DLW technique. A SEE of 79 kcal.d-1 and a mean difference of 72 kcal.d-1, with a 95% CI ranging from -238 to 93.9 kcal.d-1 was found. In addition, no significant differences between predicted HRPAnetRMR and measured TEE using DLW were found, showing an SEE of 99 kcal.d-1 and a mean difference of -67 kcal.d-1, and a 95% CI ranging from -276.6 to 141.9 kcal.d-1. AHbranched and HRPAnetRMR were both valid and similarly suitable for the prediction of energy expenditure in children under free-living conditions. Significant associations between DLWAEE and the after-school time window indicated that this time window as an important discretionary period representative of children physical activity. However, the duration of the after-school time windows should be more carefully considered. Accelerometer data showed a better association between the largest after-school time window (3.5 hr) and measured TEE. The final study, completed with 19 children aged 8-12 (10.3±1.0 yr) highlighted, under laboratory conditions across a range of walking and running speeds, the inadequacy of the use of the standard MET in children. This traditional approach overestimates energy expenditure with an increased difference linearly related to speed increments. Minute-by-minute analyses of 7 days of free-living monitoring showed an average overestimation of 64 minutes per day for moderate-to-vigorousphysical- activity (MVPA) using the standard MET compared with the individually measured MET. For all intensities, these differences were statistically significant (p< 0.001). The second part of this study showed a variability of 20% in the average time spent at MVPA when comparing HR I 140 bpm and HR > 50%P &VO2 (P &VO2 = the highest &VO2 observed during an exercise test to exhaustion). Results of the current study compared to observations in the literature showed that HR I 140 bpm consistently estimates lower MVPA time than HR > 50%P &VO2. When these two PA indices were compared with individual and standard MET measured minute-byminute, statistically significant differences were verified among all of them at MPA, but no differences were verified at VPA, except between individual and standard METs. However, whether each one of the PA indices used are under- or overestimating time at MVPA is still debatable due to the lack of a gold standard. Finally, each index used in this study classified different numbers of participants as achieving the PA target of 60 min.d-1. The wide variability between indices when attempting to classify children who are achieving the recommended target is cause for great concern because habitually these indices are utilised as screening tools in paediatric and public health settings and used to guide behavioural interventions.