Non-lethal sampling for assessment of mitochondrial function does not affect metabolic rate and swimming performance.

A fundamental issue in the metabolic field is whether it is possible to understand underlying mechanisms that characterize individual variation. Whole-animal performance relies on mitochondrial function as it produces energy for cellular processes. However, our lack of longitudinal measures to evaluate how mitochondrial function can change within and among individuals and with environmental context makes it difficult to assess individual variation in mitochondrial traits. The aims of this study were to test the repeatability of muscle mitochondrial metabolism by performing two biopsies of red muscle, and to evaluate the effects of biopsies on whole-animal performance in goldfish Carassius auratus. Our results show that basal mitochondrial respiration and net phosphorylation efficiency are repeatable at 14-day intervals. We also show that swimming performance (optimal cost of transport and critical swimming speed) was repeatable in biopsied fish, whereas the repeatability of individual oxygen consumption (standard and maximal metabolic rates) seemed unstable over time. However, we noted that the means of individual and mitochondrial traits did not change over time in biopsied fish. This study shows that muscle biopsies allow the measurement of mitochondrial metabolism without sacrificing animals and that two muscle biopsies 14 days apart affect the intraspecific variation in fish performance without affecting average performance of individuals. This article is part of the theme issue 'The evolutionary significance of variation in metabolic rates'.

Lactate signaling and fuel selection in rainbow trout: mobilization of energy reserves.

Lactate is now recognized as a regulator of fuel selection in mammals because it inhibits lipolysis by binding to the hydroxycarboxylic acid receptor 1 (HCAR1). The goals of this study were to quantify the effects of exogenous lactate on: 1) lipolytic rate or rate of appearance of glycerol in the circulation (Ra glycerol) and hepatic glucose production (Ra glucose), and 2) key tissue proteins involved in lactate signaling, glucose transport, glycolysis, gluconeogenesis, lipolysis, and ß-oxidation in rainbow trout. Measurements of fuel mobilization kinetics show that lactate does not affect lipolysis as it does in mammals (Ra glycerol remains at 7.3 ± 0.5 µmol·kg-1·min-1), but strongly reduces hepatic glucose production (16.4 ± 2.0 to 8.9 ± 1.2 µmol·kg-1·min-1). This reduction is likely induced by decreasing gluconeogenic flux through the inhibition of cytosolic phosphoenolpyruvate carboxykinase (Pck1, alternatively called Pepck1; 60% and 24% declines in gene expression and protein level, respectively). It is also caused by lactate substituting for glucose as a fuel in all tissues except white muscle that increases glut4a expression and has limited capacity for monocarboxylate transporter (Mct)-mediated lactate import. We conclude that lipolysis is not affected by hyperlactatemia because trout show no activation of autocrine Hcar1 signaling (gene expression of the receptor is unchanged or even repressed in red muscle). Lactate regulates fuel mobilization via Pck1-mediated suppression of gluconeogenesis and by replacing glucose as a fuel. This study highlights important functional differences in the Hcar1 signaling system between fish and mammals for the regulation of fuel selection.

Succinate oxidation rescues mitochondrial ATP synthesis at high temperature in Drosophila melanogaster.

Decreased NADH-induced and increased reduced FADH2 -induced respiration rates at high temperatures are associated with thermal tolerance in Drosophila. Here, we determined whether this change was associated with adjustments of adenosine triphosphate (ATP) production rate and coupling efficiency (ATP/O) in Drosophila melanogaster. We show that decreased pyruvate + malate oxidation at 35°C is associated with a collapse of ATP synthesis and a drop in ATP/O ratio. However, adding succinate triggered a full compensation of both oxygen consumption and ATP synthesis rates at this high temperature. Addition of glycerol-3-phosphate (G3P) led to a huge increase in respiration with no further advantage in terms of ATP production. We conclude that succinate is the only alternative substrate able to compensate both oxygen consumption and ATP production rates during oxidative phosphorylation at high temperature, which has important implications for thermal adaptation.

Temperature-dependent metabolic consequences of food deprivation in the European sardine.

Aquatic ecosystems can exhibit seasonal variation in resource availability and animals have evolved to cope with the associated caloric restriction. During winter in the NW Mediterranean Sea, the European sardine Sardina pilchardus naturally experiences caloric restriction owing to a decrease in the diversity and quantity of plankton. However, ongoing global warming has had deleterious effects on plankton communities such that food shortages may occur throughout the year, especially under warm conditions in the summer. We investigated the interactive effects of temperature and food availability on sardine metabolism by continuously monitoring whole-animal respiration of groups of control (fed) and food-deprived sardines over a 60-day experiment in winter (12°C) or summer (20°C) conditions under natural photoperiod. In addition, we measured mitochondrial respiration of red muscle fibres, biometric variables and energy reserves of individuals sampled at 30 and 60 days. This revealed that winter food deprivation elicits energy saving mechanisms at whole animal and cellular levels by maintaining a low metabolism to preserve energy reserves, allowing high levels of survival. By contrast, despite energy saving mechanisms at the mitochondrial level, whole animal metabolic rate was high during food deprivation in summer, causing increased consumption of energy reserves at the muscular level and high mortality after 60 days. Furthermore, a 5-day re-feeding did not improve survival, and mortalities continued, suggesting that long-term food deprivation at high temperatures causes profound stress in sardines that potentially impairs nutrient absorption.

Absence of mitochondrial responses in muscles of zebrafish exposed to several heat waves.

Heat waves are extreme thermal events whose frequency and intensity will increase with global warming. As metabolic responses to temperature are time-dependent, we explored the effects of an exposure to several heat waves on the mitochondrial metabolism of zebrafish Danio rerio. For this purpose, zebrafish were acclimated at 26 °C or 31 °C for 4 weeks and some fish acclimated at 26 °C underwent 2 types of heat waves: 2 periods of 5 days at 31 °C or 10 days at 31 °C. After this acclimation period, mitochondrial respiration of red muscle fibres was measured at 26 °C and 31 °C for each fish, with the phosphorylation (OXPHOS) and basal (LEAK) respirations obtained with activation of complex I, complex II or complexes I and II. The respiratory control ratio (RCR) and the mitochondrial aerobic scope (CAS) were also calculated at both temperatures after the activation of complexes I and II. Under our conditions, heat waves did not result in variations in any mitochondrial parameters, suggesting a high tolerance of zebrafish to environmental temperature fluctuations. However, an acute in vitro warming led to an increase in the LEAK respiration together with a higher temperature effect on complex II than complex I, inducing a decrease of mitochondrial efficiency to produce energy at high temperatures. Increased interindividual variability for some parameters at 26 °C or 31 °C also suggests that each individual has its own ability to cope with temperature fluctuations.

Different patterns of chronic hypoxia lead to hierarchical adaptive mechanisms in goldfish metabolism.

Some hypoxia-tolerant species, such as goldfish, experience intermittent and severe hypoxia in their natural habitat, causing them to develop multiple physiological adaptations. However, in fish, the metabolic impact of regular hypoxic exposure on swimming performance in normoxia is less well understood. Therefore, we experimentally tested whether chronic exposure to constant (30 days at 10% air saturation) or intermittent hypoxia (3 h in normoxia and 21 h in hypoxia, 5 days a week) would result in similar metabolic and swimming performance benefits after reoxygenation. Moreover, half of the normoxic and intermittent hypoxic fish were put on a 20-day normoxic training regime. After these treatments, metabolic rate (standard and maximum metabolic rates: SMR and MMR) and swimming performance [critical swimming speed (Ucrit) and cost of transport (COT)] were assessed. In addition, enzyme activities [citrate synthase (CS), cytochrome c oxidase (COX) and lactate dehydrogenase (LDH)] and mitochondrial respiration were examined in red muscle fibres. We found that acclimation to constant hypoxia resulted in (1) metabolic suppression (-45% SMR and -27% MMR), (2) increased anaerobic capacity (+117% LDH), (3) improved swimming performance (+80% Ucrit, -71% COT) and (4) no changes at the mitochondrial level. Conversely, the enhancement of swimming performance was reduced following acclimation to intermittent hypoxia (+45% Ucrit, -41% COT), with a 55% decrease in aerobic scope, despite a significant increase in oxidative metabolism (+201% COX, +49% CS). This study demonstrates that constant hypoxia leads to the greatest benefit in swimming performance and that mitochondrial metabolic adjustments only provide minor help in coping with hypoxia.

Changes in foraging mode caused by a decline in prey size have major bioenergetic consequences for a small pelagic fish.

Global warming is causing profound modifications of aquatic ecosystems and one major outcome appears to be a decline in adult size of many fish species. Over the last decade, sardine populations in the Gulf of Lions (NW Mediterranean Sea) have shown severe declines in body size and condition as well as disappearance of the oldest individuals, which could not be related to overfishing, predation pressure or epizootic diseases. In this study, we investigated whether this situation reflects a bottom-up phenomenon caused by reduced size and availability of prey that could lead to energetic constraints. We fed captive sardines with food items of two different sizes eliciting a change in feeding mode (filter-feeding on small items and directly capturing larger ones) at two different rations for several months, and then assessed their muscle bioenergetics to test for changes in cellular function. Feeding on smaller items was associated with a decline in body condition, even at high ration, and almost completely inhibited growth by comparison to sardines fed large items at high ration. Sardines fed on small items presented specific mitochondrial adjustments for energy sparing, indicating a major bioenergetic challenge. Moreover, mitochondria from sardines in poor condition had low basal oxidative activity but high efficiency of ATP production. Notably, when body condition was below a threshold value of 1.07, close to the mean observed in the wild, it was directly correlated with basal mitochondrial activity in muscle. The results show a link between whole-animal condition and cellular bioenergetics in the sardine, and reveal physiological consequences of a shift in feeding mode. They demonstrate that filter-feeding on small prey leads to poor growth, even under abundant food and an increase in the efficiency of ATP production. These findings may partially explain the declines in sardine size and condition observed in the wild.

Le changement global entraîne de profondes modifications des écosystèmes aquatiques, l'une des principales étant le déclin de la taille des adultes chez de nombreuses espèces de poissons. Au cours de la dernière décennie, les populations de sardines du Golfe du Lion (Nord-Ouest de la Méditerranée) ont montré une importante diminution de leur taille et de leur condition corporelle ainsi qu'une disparition des individus les plus âgés, qui n'ont pas pu être liées à la surpêche, à la pression de prédation ou aux épizooties. Dans cette étude, nous avons cherché à savoir si cette situation reflète un phénomène ascendant causé par la réduction de la taille et de la disponibilité des proies qui pourrait entraîner des contraintes énergétiques chez la sardine. Nous avons ainsi nourri des sardines captives avec des granulés de deux tailles différentes provoquant un changement de mode d'alimentation (filtration des petits granulés et capture directe des plus gros) et à deux rations différentes pendant plusieurs mois, puis nous avons évalué leur bioénergétique musculaire pour tester les changements au niveau de leur fonction cellulaire. L'alimentation à base de petits granulés a été associée à un déclin de la condition corporelle, même à une ration élevée, et à une croissance quasiment inhibée par rapport aux sardines nourries avec des plus gros granulés à une ration élevée. Les sardines nourries avec des petits granulés ont également présenté des ajustements mitochondriaux spécifiques pour économiser de l'énergie, indiquant un défi bioénergétique majeur. De plus, les mitochondries des sardines en mauvaise condition présentaient une faible activité oxydative basale, mais une efficacité élevée de production d'ATP. Notamment, lorsque la condition corporelle était inférieure à une valeur seuil de 1,07, proche de la moyenne observée dans la nature, elle était directement corrélée à l'activité mitochondriale basale dans le muscle. Ces résultats montrent un lien entre la condition de l'animal entier et la bioénergétique cellulaire chez la sardine, et révèlent les conséquences physiologiques d'un changement de mode d'alimentation. Ils démontrent que le nourrissage via la filtration de petites proies entraîne une faible croissance, même en cas de nourriture abondante, et une augmentation de l'efficacité de la production d'ATP. Ces résultats peuvent expliquer en partie le déclin de la taille et de la condition des sardines observé dans la nature.

Low oxygen levels can help to prevent the detrimental effect of acute warming on mitochondrial efficiency in fish.

Aerobic metabolism of aquatic ectotherms is highly sensitive to fluctuating climates. Many mitochondrial traits exhibit phenotypic plasticity in response to acute variations in temperature and oxygen availability. These responses are critical for understanding the effects of environmental variations on aquatic ectotherms' performance. Using the European seabass, Dicentrarchus labrax, we determined the effects of acute warming and deoxygenation in vitro on mitochondrial respiratory capacities and mitochondrial efficiency to produce ATP (ATP/O ratio). We show that acute warming reduced ATP/O ratio but deoxygenation marginally raised ATP/O ratio, leading to a compensatory effect of low oxygen availability on mitochondrial ATP/O ratio at high temperature. The acute effect of warming and deoxygenation on mitochondrial efficiency might be related to the leak of protons across the mitochondrial inner membrane, as the mitochondrial respiration required to counteract the proton leak increased with warming and decreased with deoxygenation. Our study underlines the importance of integrating the combined effects of temperature and oxygen availability on mitochondrial metabolism. Predictions on decline in performance of aquatic ectotherms owing to climate change may not be accurate, since these predictions typically look at respiratory capacity and ignore efficiency of ATP production.

Muscle bioenergetics of two emblematic Mediterranean fish species: Sardina pilchardus and Sparus aurata.

We investigated links between swimming behavior and muscle bioenergetics in two emblematic Mediterranean fish species that have very different ecologies and activity levels. European sardines Sardina pilchardus are pelagic, they swim aerobically, school constantly and have high muscle fat content. Gilthead seabream Sparus aurata are bentho-pelagic, they show discontinuous spontaneous swimming patterns and store less fat in their muscle. Estimating the proportion of red and white muscle phenotypes, sardine exhibited a larger proportion of red muscle (~10% of the body mass) compared to gilthead seabream (~5% of the body mass). We firstly studied red and white muscle fiber bioenergetics, using high-resolution respirometers, showing a 4-fold higher oxidation capacity for red compared to white muscle. Secondly, we aimed to compare the red muscle ability to oxidize either lipids or carbohydrates. Sardine red muscle had a 3-fold higher oxidative capacity than gilthead seabream and a greater capacity to oxidize lipids. This study provides novel insights into physiological mechanisms underlying the different lifestyles of these highly-prized species.

Artificial light at night disturbs the activity and energy allocation of the common toad during the breeding period.

The presence of artificial light at night (ALAN) is currently a global phenomenon. By altering the photoperiod, ALAN may directly affect the physiology and behaviour of many organisms, such as the timing of daily rhythms, hormonal regulation, food intake, metabolism, migration and reproduction. Surprisingly while it is known that ALAN exposure strongly influences health of humans and laboratory animals, studies on wildlife remain scarce. Amphibians are one of the most nocturnal groups of vertebrates and exhibit an unfavourable conservation status in most parts of the world. In order to gain insight into the consequences of ALAN, we experimentally exposed 36 adult breeding male common toads, Bufo bufo, to a light intensity of 0.1, 5 or 20 lux for 20 days, to investigate the activity using infrared cameras and the whole-body oxygen consumption by respirometry, as well as body mass and food intake. ALAN reduced toad activity over 24 h by 56% at 5 lux and by 73% at 20 lux. It did not affect the total energy expenditure but altered energy allocation. Indeed, standard energy expenditure increased by 28% at 5 lux and by 58% at 20 lux, while activity energy expenditure decreased by 18% at 5 lux and 38% at 20 lux. Finally, body mass and food intake were not affected. This study suggests that ALAN plays a large role in the activity and energy metabolism of common toads, which may have a long-term negative effect on the fitness of common toad populations. Generalizing these results to other taxa is crucial for conservation of biodiversity in an increasingly light world.

Mitochondrial oxidative phosphorylation efficiency is upregulated during fasting in two major oxidative tissues of ducklings.

Fasted endothermic vertebrates must develop physiological responses to maximize energy conservation and survival. The aim of this study was to determine the effect of 1-wk. fasting in 5-wk. old ducklings (Cairina moschata) from whole-body resting metabolic rate and body temperature to metabolic phenotype of tissues and mitochondrial coupling efficiency. At the level of whole organism, the mass-specific metabolic rate of ducklings was decreased by 40% after 1-wk. of fasting, which was associated with nocturnal Tb declines and shallow diurnal hypothermia during fasting. At the cellular level, fasting induced a large reduction in liver, gastrocnemius (oxidative) and pectoralis (glycolytic) muscle masses together with a fuel selection towards lipid oxidation and ketone body production in liver and a lower glycolytic phenotype in skeletal muscles. At the level of mitochondria, fasting induced a reduction of oxidative phosphorylation activities and an up-regulation of coupling efficiency (+30% on average) in liver and skeletal muscles. The present integrative study shows that energy conservation in fasted ducklings is mainly achieved by an overall reduction in mitochondrial activity and an increase in mitochondrial coupling efficiency, which would, in association with shallow hypothermia, increase the conservation of endogenous fuel stores during fasting.

Doping for sex: Bad for mitochondrial performances? Case of testosterone supplemented Hyla arborea during the courtship period.

Sexual selection has been widely explored from numerous perspectives, including behavior, ecology, and to a lesser extent, energetics. Hormones, and specifically androgens such as testosterone, are known to trigger sexual behaviors. Their effects are therefore of interest during the breeding period. Our work investigates the effect of testosterone on the relationship between cellular bioenergetics and contractile properties of two skeletal muscles involved in sexual selection in tree frogs. Calling and locomotor abilities are considered evidence of good condition in Hyla males, and thus server as proxies for male quality and attractiveness. Therefore, how these behaviors are powered efficiently remains of both physiological and behavioral interest. Most previous research, however, has focused primarily on biomechanics, contractile properties or mitochondrial enzyme activities. Some have tried to establish a relationship between those parameters but to our knowledge, there is no study examining muscle fiber bioenergetics in Hyla arborea. Using chronic testosterone supplementation and through an integrative study combining fiber bioenergetics and contractile properties, we compared sexually dimorphic trunk muscles directly linked to chronic sound production to a hindlimb muscle (i.e. gastrocnemius) that is particularly adapted for explosive movement. As expected, trunk muscle bioenergetics were more affected by testosterone than gastrocnemius muscle. Our study also underlines contrasted energetic capacities between muscles, in line with contractile properties of these two different muscle phenotypes. The discrepancy of both substrate utilization and contractile properties is consistent with the specific role of each muscle and our results are elucidating another integrative example of a muscle force-endurance trade-off.

Novel energy-saving strategies to multiple stressors in birds: the ultradian regulation of body temperature.

This study aimed to examine thermoregulatory responses in birds facing two commonly experienced stressors, cold and fasting. Logging devices allowing long-term and precise access to internal body temperature were placed within the gizzards of ducklings acclimated to cold (CA) (5°C) or thermoneutrality (TN) (25°C). The animals were then examined under three equal 4-day periods: ad libitum feeding, fasting and re-feeding. Through the analysis of daily as well as short-term, or ultradian, variations of body temperature, we showed that while ducklings at TN show only a modest decline in daily thermoregulatory parameters when fasted, they exhibit reduced surface temperatures from key sites of vascular heat exchange during fasting. The CA birds, on the other hand, significantly reduced their short-term variations of body temperature while increasing long-term variability when fasting. This phenomenon would allow the CA birds to reduce the energetic cost of body temperature maintenance under fasting. By analysing ultradian regulation of body temperature, we describe a means by which an endotherm appears to lower thermoregulatory costs in response to the combined stressors of cold and fasting.

Proline as a fuel for insect flight: enhancing carbohydrate oxidation in hymenopterans.

Bees are thought to be strict users of carbohydrates as metabolic fuel for flight. Many insects, however, have the ability to oxidize the amino acid proline at a high rate, which is a unique feature of this group of animals. The presence of proline in the haemolymph of bees and in the nectar of plants led to the hypothesis that plants may produce proline as a metabolic reward for pollinators. We investigated flight muscle metabolism of hymenopteran species using high-resolution respirometry performed on permeabilized muscle fibres. The muscle fibres of the honeybee, Apis mellifera, do not have a detectable capacity to oxidize proline, as those from the migratory locust, Locusta migratoria, used here as an outgroup representative. The closely related bumblebee, Bombus impatiens, can oxidize proline alone and more than doubles its respiratory capacity when proline is combined with carbohydrate-derived substrates. A distant wasp species, Vespula vulgaris, exhibits the same metabolic phenotype as the bumblebee, suggesting that proline oxidation is common in hymenopterans. Using a combination of mitochondrial substrates and inhibitors, we further show that in B. impatiens, proline oxidation provides reducing equivalents and electrons directly to the electron transport system. Together, these findings demonstrate that some bee and wasp species can greatly enhance the oxidation of carbohydrates using proline as fuel for flight.

Lipid-induced thermogenesis is up-regulated by the first cold-water immersions in juvenile penguins.

The passage from shore to marine life is a critical step in the development of juvenile penguins and is characterized by a fuel selection towards lipid oxidation concomitant to an enhancement of lipid-induced thermogenesis. However, mechanisms of such thermogenic improvement at fledging remain undefined. We used two different groups of pre-fledging king penguins (Aptenodytes patagonicus) to investigate the specific contribution of cold exposure during water immersion to lipid metabolism. Terrestrial penguins that had never been immersed in cold water were compared with experimentally cold-water immersed juveniles. Experimentally immersed penguins underwent ten successive immersions at approximately 9-10 °C for 5 h over 3 weeks. We evaluated adaptive thermogenesis by measuring body temperature, metabolic rate and shivering activity in fully immersed penguins exposed to water temperatures ranging from 12 to 29 °C. Both never-immersed and experimentally immersed penguins were able to maintain their homeothermy in cold water, exhibiting similar thermogenic activity. In vivo, perfusion of lipid emulsion at thermoneutrality induced a twofold larger calorigenic response in experimentally immersed than in never-immersed birds. In vitro, the respiratory rates and the oxidative phosphorylation efficiency of isolated muscle mitochondria were not improved with cold-water immersions. The present study shows that acclimation to cold water only partially reproduced the fuel selection towards lipid oxidation that characterizes penguin acclimatization to marine life.

Ontogeny of non-shivering thermogenesis in Muscovy ducklings (Cairina moschata).

In precocial birds, developing the capacity for early regulatory thermogenesis appears as a fundamental prerequisite for survival and growth in cold environments. However, the exact nature of these processes has not been thoroughly investigated. Several bird species, such as Muscovy ducks (Cairina moschata), develop muscular non-shivering thermogenesis when chronically exposed to cold. The aim of this study was to investigate the age-dependent development of non-shivering thermogenesis in ducklings reared either at thermoneutrality (25°C) or in the cold (4°C). Non-shivering thermogenesis was assessed weekly by simultaneously measuring whole body metabolic heat production and electromyographic activity during shivering at different temperatures ranging from 29°C to 0°C. We found that ducklings reared at thermoneutrality displayed a capacity for non-shivering thermogenesis during the first month of post-hatching life. This thermogenic mechanism increased further in ducklings chronically exposed to a cold environment, but it decreased over time when birds were kept in a thermoneutral environment.

Early septic shock induces loss of oxidative phosphorylation yield plasticity in liver mitochondria.

We aimed to study the change in mitochondrial oxidative phosphorylation efficiency occurring at the early stage of septic shock in an experimental model. Thirty-six male Wistar rats were divided into two groups. In the first group, a cecal ligation and puncture (CLP) was carried out to induce septic shock for 5 h. The second group includes sham-operated rats and constitutes the control group. Blood gas analysis, alanine amino transferase, and lactic acid dosages were assayed 5 h after surgery. Liver mitochondria were isolated for in vitro functional characterization, including mitochondrial respiratory parameters, oxidative phosphorylation efficiency, oxi-radical production, membrane potential, and cytochrome c oxidase activity and content. Liver interleukin 1ß (IL-1ß) and tumor necrosis α mRNA levels were determined. Septic shock induced a severe hypotension occurring 180 min after CLP in association with a metabolic acidosis, an increase in plasma alanine amino transferase, liver IL-1ß gene expression, and mitochondrial reactive oxygen species production. The rates of mitochondrial oxygen consumption and the activity and content of cytochrome c oxidase were significantly decreased while no alterations in the oxidative phosphorylation efficiency and inner membrane integrity were found. These results show that contrary to what was expected, liver mitochondria felt to adjust their oxidative phosphorylation efficiency in response to the decrease in the mitochondrial oxidative activity induced by CLP. This loss of mitochondrial bioenergetics plasticity might be related to mitochondrial oxidative stress and liver cytokines production.

Lactate kinetics of rainbow trout during graded exercise: do catheters affect the cost of transport?

Changes in lactate kinetics as a function of exercise intensity have never been measured in an ectotherm. Continuous infusion of a tracer is necessary to quantify rates of lactate appearance (Ra) and disposal (Rd), but it requires double catheterization, which could interfere with swimming. Using rainbow trout, our goals were to: (1) determine the potential effects of catheters and blood sampling on metabolic rate (O2), total cost of transport (TCOT), net cost of transport (NCOT) and critical swimming speed (Ucrit), and (2) monitor changes in lactate fluxes during prolonged, steady-state swimming or graded swimming from rest to Ucrit. This athletic species maintains high baseline lactate fluxes of 24 µmol kg(-1) min(-1) that are only increased at intensities >2.4 body lengths (BL) s(-1) or 85% Ucrit. As the fish reaches Ucrit, Ra is more strongly stimulated (+67% to 40.4 µmol kg(-1) min(-1)) than Rd (+41% to 34.7 µmol kg(-1) min(-1)), causing a fourfold increase in blood lactate concentration. Without this stimulation of Rd during intense swimming, lactate accumulation would double. By contrast, steady-state exercise at 1.7 BL s(-1) increases lactate fluxes to ~30 µmol kg(-1) min(-1), with a trivial mismatch between Ra and Rd that only affects blood concentration minimally. Results also show that the catheterizations and blood sampling needed to measure metabolite kinetics in exercising fish have no significant impact on O2 or TCOT. However, these experimental procedures affect locomotion energetics by increasing NCOT at high speeds and by decreasing Ucrit.

Metabolic response to lipid infusion in fasting winter-acclimatized king penguin chicks (Aptenodytes patagonicus).

During the cold austral winter, king penguin chicks are infrequently fed by their parents and thus experience severe nutritional deprivation under harsh environmental conditions. These energetic constraints lead to a range of energy sparing mechanisms balanced by the maintenance of efficient thermogenic processes. The present work investigated whether the high thermogenic capacities exhibited by winter-acclimatized king penguin chicks could be related to an increase in lipid substrate supply and oxidation in skeletal muscle, the main site of thermogenesis in birds. To test this hypothesis, we examined i) the effect of an experimental rise in plasma triglyceride on the whole metabolic rate in winter-acclimatized (WA) and de-acclimatized king penguin chicks kept at thermoneutrality (TN), and ii) investigated the fuel preference of muscle mitochondria. In vivo, a perfusion of a lipid emulsion induced a small 10% increase of metabolic rate in WA chicks but not in TN group. In vitro, the oxidation rate of muscle mitochondria respiring on lipid-derived substrate was +40% higher in WA chicks than in TN, while no differences were found between groups when mitochondria oxidized carbohydrate-derived substrate or succinate. Despite an enhanced fuel selection towards lipid oxidation in skeletal muscle, a rise of circulating lipids per se was not sufficient to fully unravel the thermogenic capacity of winter-acclimatized king penguin chicks.

Selective upregulation of lipid metabolism in skeletal muscle of foraging juvenile king penguins: an integrative study.

The passage from shore to marine life of juvenile penguins represents a major energetic challenge to fuel intense and prolonged demands for thermoregulation and locomotion. Some functional changes developed at this crucial step were investigated by comparing pre-fledging king penguins with sea-acclimatized (SA) juveniles (Aptenodytes patagonicus). Transcriptomic analysis of pectoralis muscle biopsies revealed that most genes encoding proteins involved in lipid transport or catabolism were upregulated, while genes involved in carbohydrate metabolism were mostly downregulated in SA birds. Determination of muscle enzymatic activities showed no changes in enzymes involved in the glycolytic pathway, but increased 3-hydroxyacyl-CoA dehydrogenase, an enzyme of the ß-oxidation pathway. The respiratory rates of isolated muscle mitochondria were much higher with a substrate arising from lipid metabolism (palmitoyl-L-carnitine) in SA juveniles than in terrestrial controls, while no difference emerged with a substrate arising from carbohydrate metabolism (pyruvate). In vivo, perfusion of a lipid emulsion induced a fourfold larger thermogenic effect in SA than in control juveniles. The present integrative study shows that fuel selection towards lipid oxidation characterizes penguin acclimatization to marine life. Such acclimatization may involve thyroid hormones through their nuclear beta receptor and nuclear coactivators.