Environmental variation associated with overwintering elicits marked metabolic plasticity in a temperate salmonid, Salvelinus fontinalis.

Poleward winters commonly expose animals, including fish, to frigid temperatures and low food availability. Fishes that remain active over winter must therefore balance trade-offs between conserving energy and maintaining physiological performance in the cold, yet the extent and underlying mechanisms of these trade-offs are not well understood. We investigated the metabolic plasticity of brook char (Salvelinus fontinalis), a temperate salmonid, from the biochemical to whole-animal level in response to cold and food deprivation. Acute cooling (1°C day-1) from 14°C to 2°C had no effect on food consumption but reduced activity by 77%. We then assessed metabolic performance and demand over 90 days with exposure to warm (8°C) or cold winter (2°C) temperatures while fish were fed or starved. Resting metabolic rate (RMR) decreased substantially during initial cooling from 8°C to 2°C (Q10=4.2-4.5) but brook char exhibited remarkable thermal compensation during acclimation (Q10=1.4-1.6). Conversely, RMR was substantially lower (40-48%) in starved fish, conserving energy. Thus, the absolute magnitude of thermal plasticity may be masked or modified under food restriction. This reduction in RMR was associated with atrophy and decreases in in vivo protein synthesis rates, primarily in non-essential tissues. Remarkably, food deprivation had no effect on maximum oxygen uptake rates and thus aerobic capacity, supporting the notion that metabolic capacity can be decoupled from RMR in certain contexts. Overall, our study highlights the multi-faceted energetic flexibility of Salvelinus spp. that likely contributes to their success in harsh and variable environments and may be emblematic of winter-active fishes more broadly.

Resilience and phenotypic plasticity of Arctic char (Salvelinus alpinus) facing cyclic hypoxia: insights into growth, energy stores and hepatic metabolism.

Arctic char (Salvelinus alpinus) is facing the decline of its southernmost populations due to several factors including rising temperatures and eutrophication. These conditions are also conducive to episodes of cyclic hypoxia, another possible threat to this species. In fact, lack of oxygen and reoxygenation can both have serious consequences on fish as a result of altered ATP balance and an elevated risk of oxidative burst. Thus, fish must adjust their phenotype to survive and equilibrate their energetic budget. However, their energy allocation strategy could imply a reduction in growth which could be deleterious for their fitness. Although the impact of cyclic hypoxia is a major issue for ecosystems and fisheries worldwide, our knowledge on how salmonid deal with high oxygen fluctuations remains limited. Our objective was to characterize the effects of cyclic hypoxia on growth and metabolism in Arctic char. We monitored growth parameters (specific growth rate, condition factor), hepatosomatic and visceral indexes, relative heart mass and hematocrit of Arctic char exposed to 30 days of cyclic hypoxia. We also measured the hepatic protein synthesis rate, hepatic triglycerides as well as muscle glucose, glycogen and lactate and quantified hepatic metabolites during this treatment. The first days of cyclic hypoxia slightly reduce growth performance with a downward trend in specific growth rate in mass and condition factor variation compared to the control group. This acute exposure also induced a profound metabolome reorganization in the liver with an alteration of amino acid, carbohydrate and lipid metabolisms. However, fish rebalanced their metabolic activities and successfully maintained their growth and energetic reserves after 1 month of cyclic hypoxia. These results demonstrate the impressive ability of Arctic char to cope with its changing environment but also highlight a certain vulnerability of this species during the first days of a cyclic hypoxia event.

Age-related flexibility of energetic metabolism in the honey bee Apis mellifera.

The mechanisms that underpin aging are still elusive. In this study, we suggest that the ability of mitochondria to oxidize different substrates, which is known as metabolic flexibility, is involved in this process. To verify our hypothesis, we used honey bees (Apis mellifera carnica) at different ages, to assess mitochondrial oxygen consumption and enzymatic activities of key enzymes of the energetic metabolism as well as ATP5A1 content (subunit of ATP synthase) and adenylic energy charge (AEC). We also measured mRNA abundance of genes involved in mitochondrial functions and the antioxidant system. Our results demonstrated that mitochondrial respiration increased with age and favored respiration through complexes I and II of the electron transport system (ETS) while glycerol-3-phosphate (G3P) oxidation was relatively decreased. In addition, glycolytic, tricarboxylic acid cycle and ETS enzymatic activities increased, which was associated with higher ATP5A1 content and AEC. Furthermore, we detected an early decrease in the mRNA abundance of subunits of NADH ubiquinone oxidoreductase subunit B2 (NDUFB2, complex I), mitochondrial cytochrome b (CYTB, complex III) of the ETS as well as superoxide dismutase 1 and a later decrease for vitellogenin, catalase and mitochondrial cytochrome c oxidase subunit 1 (COX1, complex IV). Thus, our study suggests that the energetic metabolism is optimized with aging in honey bees, mainly through quantitative and qualitative mitochondrial changes, rather than showing signs of senescence. Moreover, aging modulated metabolic flexibility, which might reflect an underpinning mechanism that explains lifespan disparities between the different castes of worker bees.

Social experience influences thermal sensitivity: lessons from an amphibious mangrove fish.

Understanding the factors affecting the capacity of ectothermic fishes to cope with warming temperature is critical given predicted climate change scenarios. We know that a fish's social environment introduces plasticity in how it responds to high temperature. However, the magnitude of this plasticity and the mechanisms underlying socially modulated thermal responses are unknown. Using the amphibious hermaphroditic mangrove rivulus fish Kryptolebias marmoratus as a model, we tested three hypotheses: (1) social stimulation affects physiological and behavioural thermal responses of isogenic lineages of fish; (2) social experience and acute social stimulation result in distinct physiological and behavioural responses; and (3) a desensitization of thermal receptors is responsible for socially modulated thermal responses. To test the first two hypotheses, we measured the temperature at which fish emerged from the water (i.e. pejus temperature) upon acute warming with socially naive isolated fish and with fish that were raised alone and then given a short social experience prior to exposure to increasing temperature (i.e. socially experienced fish). Our results did not support our first hypothesis as fish socially stimulated by mirrors during warming (i.e. acute social stimulation) emerged at similar temperatures to isolated fish. However, in support of our second hypothesis, a short period of prior social experience resulted in fish emerging at a higher temperature than socially naive fish suggesting an increase in pejus temperature with social experience. To test our third hypothesis, we exposed fish that had been allowed a brief social interaction and naive fish to capsaicin, an agonist of TRPV1 thermal receptors. Socially experienced fish emerged at significantly higher capsaicin concentrations than socially naive fish suggesting a desensitization of their TRPV1 thermal receptors. Collectively, our data indicate that past and present social experiences impact the behavioural response of fish to high temperature. We also provide novel data suggesting that brief periods of social experience affect the capacity of fish to perceive warm temperature.

Transcriptional and metabolomic investigation of the stress response in snow crab during simulated transport condition (Chionoecetes opilio).

The molecular mechanisms underlying the stress response are poorly described in crustaceans. This includes the snow crab (Chionoecetes opilio), a commercially important stenotherm species distributed throughout the northern hemisphere. A better understanding of the stress response in C. opilio is desperately needed for commercial and conservation purposes. The purpose of this study was to investigate the transcriptional and metabolomic response of C. opilio exposed to stressors. Crabs were randomly assigned to 24 or 72 h treatment groups where they were exposed to conditions simulating live transport (handling and air exposure). A control group was kept in cold (2 °C) and well­oxygenated saltwater. The hepatopancreas of the crabs was sampled to perform RNA-sequencing and high-performance chemical isotope labeling metabolomics. Differential gene expression analyses showed that classic crustaceans' stress markers, such as crustacean hyperglycemic hormones and heat shock proteins, were overexpressed in response to stressors. Tyrosine decarboxylase was also up-regulated in stressed crabs, suggesting an implication of the catecholamines tyramine and octopamine in the stress response. Deregulated metabolites revealed that low oxygen was an important trigger in the stress response as intermediate metabolites of the tricarboxylic acid cycle (TCA) accumulated. Lactate, which accumulated unevenly between crabs could potentially be used to predict mortality. This study provides new information on how stressors affect crustaceans and provides a basis for the development of stress markers in C. opilio.

Taurine depletion impairs cardiac function and affects tolerance to hypoxia and high temperatures in brook char (Salvelinus fontinalis).

Physiological and environmental stressors can cause osmotic stress in fish hearts, leading to a reduction in intracellular taurine concentration. Taurine is a ß-amino acid known to regulate cardiac function in other animal models but its role in fish has not been well characterized. We generated a model of cardiac taurine deficiency (TD) by feeding brook char (Salvelinus fontinalis) a diet enriched in ß-alanine, which inhibits cardiomyocyte taurine uptake. Cardiac taurine levels were reduced by 21% and stress-induced changes in normal taurine handling were observed in TD brook char. Responses to exhaustive exercise and acute thermal and hypoxia tolerance were then assessed using a combination of in vivo, in vitro and biochemical approaches. Critical thermal maximum was higher in TD brook char despite significant reductions in maximum heart rate. In vivo, TD brook char exhibited a lower resting heart rate, blunted hypoxic bradycardia and a severe reduction in time to loss of equilibrium under hypoxia. In vitro function was similar between control and TD hearts under oxygenated conditions, but stroke volume and cardiac output were severely compromised in TD hearts under severe hypoxia. Aspects of mitochondrial structure and function were also impacted in TD permeabilized cardiomyocytes, but overall effects were modest. High levels of intracellular taurine are required to achieve maximum cardiac function in brook char and cardiac taurine efflux may be necessary to support heart function under stress. Taurine appears to play a vital, previously unrecognized role in supporting cardiovascular function and stress tolerance in fish.

The ovine conceptus utilizes extracellular serine, glucose, and fructose to generate formate via the one carbon metabolism pathway.

Highly proliferative cells rely on one carbon (1C) metabolism for production of formate required for synthesis of purines and thymidine for nucleic acid synthesis. This study was to determine if extracellular serine and/or glucose and fructose contribute the production of formate in ovine conceptuses. Suffolk ewes (n = 8) were synchronized to estrus, bred to fertile rams, and conceptuses were collected on Day 17 of gestation. Conceptuses were either snap frozen in liquid nitrogen (n = 3) or placed in culture in medium (n = 5) containing either: 1) 4 mM D-glucose + 2 mM [U-13C]serine; 2) 6 mM glycine + 4 mM D-glucose + 2 mM [U-13C]serine; 3) 4 mM D-fructose + 2 mM [U-13C]serine; 4) 6 mM glycine + 4 mM D-fructose + 2 mM [U-13C]serine; 5) 4 mM D-glucose + 4 mM D-fructose + 2 mM [U-13C]serine; or 6) 6 mM glycine + 4 mM D-glucose + 4 mM D-fructose + 2 mM [U-13C]serine. After 2 h incubation, conceptuses in their respective culture medium were homogenized and the supernatant analyzed for 12C- and 13C-formate by gas chromatography and amino acids by high performance liquid chromatography. Ovine conceptuses produced both 13C- and 12C-formate, indicating that the [U-13C]serine, glucose, and fructose were utilized to generate formate, respectively. Greater amounts of 12C-formate than 13C-formate were produced, indicating that the ovine conceptus utilized more glucose and fructose than serine to produce formate. This study is the first to demonstrate that both 1C metabolism and serinogenesis are active metabolic pathways in ovine conceptuses during the peri-implantation period of pregnancy, and that hexose sugars are the preferred substrate for generating formate required for nucleotide synthesis for proliferating trophectoderm cells.

Pyoluteorin and 2,4-diacetylphloroglucinol are major contributors to Pseudomonas protegens Pf-5 biocontrol against Botrytis cinerea in cannabis.

Pseudomonas protegens Pf-5 is an effective biocontrol agent that protects many crops against pathogens, including the fungal pathogen Botrytis cinerea causing gray mold disease in Cannabis sativa crops. Previous studies have demonstrated the important role of antibiotics pyoluteorin (PLT) and 2,4-diacetylphloroglucinol (DAPG) in Pf-5-mediated biocontrol. To assess the potential involvement of PLT and DAPG in the biocontrol exerted by Pf-5 against B. cinerea in the phyllosphere of C. sativa, two knockout Pf-5 mutants were generated by in-frame deletion of genes pltD or phlA, required for the synthesis of PLT or DAPG respectively, using a two-step allelic exchange method. Additionally, two complemented mutants were constructed by introducing a multicopy plasmid carrying the deleted gene into each deletion mutant. In vitro confrontation assays revealed that deletion mutant ∆pltD inhibited B. cinerea growth significantly less than wild-type Pf-5, supporting antifungal activity of PLT. However, deletion mutant ∆phlA inhibited mycelial growth significantly more than the wild-type, hypothetically due to a co-regulation of PLT and DAPG biosynthesis pathways. Both complemented mutants recovered in vitro inhibition levels similar to that of the wild-type. In subsequent growth chamber inoculation trials, characterization of gray mold disease symptoms on infected cannabis plants revealed that both ∆pltD and ∆phlA significantly lost a part of their biocontrol capabilities, achieving only 10 and 19% disease reduction respectively, compared to 40% achieved by inoculation with the wild-type. Finally, both complemented mutants recovered biocontrol capabilities in planta similar to that of the wild-type. These results indicate that intact biosynthesis pathways for production of PLT and DAPG are required for the optimal antagonistic activity of P. protegens Pf-5 against B. cinerea in the cannabis phyllosphere.

In Tuber Biocontrol of Potato Late Blight by a Collection of Phenazine-1-Carboxylic Acid-Producing Pseudomonas spp.

Phenazine-1-carboxylic acid (PCA) produced by plant-beneficial Pseudomonas spp. is an antibiotic with antagonistic activities against Phytophthora infestans, the causal agent of potato late blight. In this study, a collection of 23 different PCA-producing Pseudomonas spp. was confronted with P. infestans in potato tuber bioassays to further understand the interaction existing between biocontrol activity and PCA production. Overall, the 23 strains exhibited different levels of biocontrol activity. In general, P. orientalis and P. yamanorum strains showed strong disease reduction, while P. synxantha strains could not effectively inhibit the pathogen's growth. No correlation was found between the quantities of PCA produced and biocontrol activity, suggesting that PCA cannot alone explain P. infestans' growth inhibition by phenazine-producing pseudomonads. Other genetic determinants potentially involved in the biocontrol of P. infestans were identified through genome mining in strains displaying strong biocontrol activity, including siderophores, cyclic lipopeptides and non-ribosomal peptide synthase and polyketide synthase hybrid clusters. This study represents a step forward towards better understanding the biocontrol mechanisms of phenazine-producing Pseudomonas spp. against potato late blight.

Comparative Genomics of Potato Common Scab-Causing Streptomyces spp. Displaying Varying Virulence.

Common scab of potato causes important economic losses worldwide following the development of necrotic lesions on tubers. In this study, the genomes of 14 prevalent scab-causing Streptomyces spp. isolated from Prince Edward Island, one of the most important Canadian potato production areas, were sequenced and annotated. Their phylogenomic affiliation was determined, their pan-genome was characterized, and pathogenic determinants involved in their virulence, ranging from weak to aggressive, were compared. 13 out of 14 strains clustered with Streptomyces scabiei, while the last strain clustered with Streptomyces acidiscabies. The toxicogenic and colonization genomic regions were compared, and while some atypical gene organizations were observed, no clear correlation with virulence was observed. The production of the phytotoxin thaxtomin A was also quantified and again, contrary to previous reports in the literature, no clear correlation was found between the amount of thaxtomin A secreted, and the virulence observed. Although no significant differences were observed when comparing the presence/absence of the main virulence factors among the strains of S. scabiei, a distinct profile was observed for S. acidiscabies. Several mutations predicted to affect the functionality of some virulence factors were identified, including one in the bldA gene that correlates with the absence of thaxtomin A production despite the presence of the corresponding biosynthetic gene cluster in S. scabiei LBUM 1485. These novel findings obtained using a large number of scab-causing Streptomyces strains are challenging some assumptions made so far on Streptomyces' virulence and suggest that other factors, yet to be characterized, are also key contributors.

Reduced Shmt2 Expression Impairs Mitochondrial Folate Accumulation and Respiration, and Leads to Uracil Accumulation in Mouse Mitochondrial DNA.

BACKGROUND: Adequate cellular thymidylate (dTMP) pools are essential for preservation of nuclear and mitochondrial genome stability. Previous studies have indicated that disruption in nuclear dTMP synthesis leads to increased uracil misincorporation into DNA, affecting genome stability. To date, the effects of impaired mitochondrial dTMP synthesis in nontransformed tissues have been understudied. OBJECTIVES: This study aimed to determine the effects of decreased serine hydroxymethyltransferase 2 (Shmt2) expression and dietary folate deficiency on mitochondrial DNA (mtDNA) integrity and mitochondrial function in mouse tissues. METHODS: Liver mtDNA content, and uracil content in liver mtDNA, were measured in Shmt2+/- and Shmt2+/+ mice weaned onto either a folate-sufficient control diet (2 mg/kg folic acid; C) or a modified diet lacking folic acid (0 mg/kg folic acid) for 7 wk. Shmt2+/- and Shmt2+/+ mouse embryonic fibroblast (MEF) cells were cultured in defined culture medium containing either 0 or 25 nM folate (6S-5-formyl-tetrahydrofolate, folinate) to assess proliferative capacity and mitochondrial function. Chi-square tests, linear mixed models, and 2-factor ANOVA with Tukey post hoc analyses were used to analyze data. RESULTS: Shmt2 +/- mice exhibited a 48%-67% reduction in SHMT2 protein concentrations in tissues. Interestingly, Shmt2+/- mice consuming the folate-sufficient C diet exhibited a 25% reduction in total folate in liver mitochondria. There was also a >20-fold increase in uracil in liver mtDNA in Shmt2+/- mice consuming the C diet, and dietary folate deficiency also increased uracil content in mouse liver mtDNA from both Shmt2+/+ and Shmt2+/- mice. Furthermore, decreased Shmt2 expression in MEF cells reduced cell proliferation, mitochondrial membrane potential, and oxygen consumption rate. CONCLUSIONS: This study demonstrates that Shmt2 heterozygosity and dietary folate deficiency impair mitochondrial dTMP synthesis in mice, as evidenced by the increased uracil in mtDNA. In addition, Shmt2 heterozygosity impairs mitochondrial function in MEF cells. These findings suggest that elevated uracil in mtDNA may impair mitochondrial function.

Contrasting strategies of hypoxic cardiac performance and metabolism in cichlids and armoured catfish.

The heart of tropical fishes is a particularly useful model system in which to investigate mechanisms of hypoxic tolerance. Here we focus on insights gained from two groups of fishes, cichlids and armoured catfishes. Cichlids respond to hypoxia by entering a sustained hypometabolism with decreased heart performance to match whole animal circulatory needs. Heart rate is decreased along with protein turnover to reduce adenosine triphosphate demand. This occurs despite the inherent capacity for high levels of cardiac power development. Although highly hypoxic tolerant at the whole animal level, the heart of cichlids does not have high constitutive activities of glycolytic enzymes compared to other species. Information is conflicting with respect to changes in glycolytic gene expression and enzyme activity following hypoxic exposure with some studies showing increases and others decreases. In contrast to cichlids, species of armoured catfish, that are routinely exposed to water of low oxygen content, do not display hypoxic bradycardia. Under hypoxia there are early changes in glucose trafficking suggestive of activation of glycolysis before lactate accumulation. Thereafter, heart glycogen is mobilized and lactate accumulates in both heart and blood, in some species to very high levels. Heart performance under hypoxia is enhanced by defense of intracellular pH. A functional sarcoplasmic reticulum and binding of hexokinase to the outer mitochondrial membrane may also play a role in cardioprotection. Maintenance of heart performance under hypoxia may relate to a tradeoff between air breathing via a modified stomach and circulatory demands for digestion.

Chronic social stress alters protein metabolism in juvenile rainbow trout, Oncorhynchus mykiss.

When confined in pairs, juvenile rainbow trout (Oncorhynchus mykiss) form dominance hierarchies in which subordinate fish exhibit characteristic physiological changes including reduced growth rates and chronically elevated plasma cortisol concentrations. We hypothesized that alterations in protein metabolism contribute to the reduced growth rate of socially stressed trout, and predicted that subordinate trout would exhibit reduced rates of protein synthesis coupled with increases in protein degradation. Protein metabolism was assessed in dominant and subordinate fish after 4 days of social interaction, and in fish that were separated after 4 days of interaction for a 4 days recovery period, to determine whether effects on protein metabolism recovered when social stress was alleviated. Protein metabolism was assessed in liver and white muscle by measuring the fractional rate of protein synthesis and markers of protein degradation. In the white muscle of subordinate fish, protein synthesis was inhibited and activities of the ubiquitin-proteasome pathway (UPP) and the autophagy lysosomal system (ALS) were elevated. By contrast, the liver of subordinate fish exhibited increased rates of protein synthesis and activation of the ALS. When allowed to recover from chronic social stress for 4 days, differences in protein metabolism observed in white muscle of subordinate fish during the interaction period disappeared. In liver, protein synthesis returned to baseline levels during recovery from social stress, but markers of protein degradation did not. Collectively, these data support the hypothesis that inhibition of muscle protein synthesis coupled with increases in muscle protein breakdown contribute to the reduced growth rates of subordinate rainbow trout.

Thermal tolerance and fish heart integrity: fatty acids profiles as predictors of species resilience.

The cardiovascular system is a major limiting system in thermal adaptation, but the exact physiological mechanisms underlying responses to thermal stress are still not completely understood. Recent studies have uncovered the possible role of reactive oxygen species production rates of heart mitochondria in determining species' upper thermal limits. The present study examines the relationship between individual response to a thermal challenge test (CTmax), susceptibility to peroxidation of membrane lipids, heart fatty acid profiles and cardiac antioxidant enzyme activities in two salmonid species from different thermal habitats (Salvelinus alpinus, Salvelinus fontinalis) and their hybrids. The susceptibility to peroxidation of membranes in the heart was negatively correlated with individual thermal tolerance. The same relationship was found for arachidonic and eicosapentaenoic acid. Total H2O2 buffering activity of the heart muscle was higher for the group with high thermal resistance. These findings underline a potential general causative relationship between sensitivity to oxidative stress, specific fatty acids, antioxidant activity in the cardiac muscle and thermal tolerance in fish and likely other ectotherms. Heart fatty acid profile could be indicative of species resilience to global change, and more importantly the plasticity of this trait could predict the adaptability of fish species or populations to changes in environmental temperature.

Activation of oxygen-responsive pathways is associated with altered protein metabolism in Arctic char exposed to hypoxia.

Fish exposed to fluctuating oxygen concentrations often alter their metabolism and/or behaviour to survive. Hypoxia tolerance is typically associated with the ability to reduce energy demand by supressing metabolic processes such as protein synthesis. Arctic char is amongst the most sensitive salmonid to hypoxia, and typically engage in avoidance behaviour when faced with lack of oxygen. We hypothesized that a sensitive species will still have the ability (albeit reduced) to regulate molecular mechanisms during hypoxia. We investigated the tissue-specific response of protein metabolism during hypoxia. Little is known about protein degradation pathways during hypoxia in fish and we predict that protein degradation pathways are differentially regulated and play a role in the hypoxia response. We also studied the regulation of oxygen-responsive cellular signalling pathways [hypoxia inducible factor (HIF), unfolded protein response (UPR) and mTOR pathways] since most of what we know comes from studies on cancerous mammalian cell lines. Arctic char were exposed to cumulative graded hypoxia trials for 3 h at four air saturation levels (100%, 50%, 30% and 15%). The rate of protein synthesis was measured using a flooding dose technique, whereas protein degradation and signalling pathways were assessed by measuring transcripts and phosphorylation of target proteins. Protein synthesis decreased in all tissues measured (liver, muscle, gill, digestive system) except for the heart. Salmonid hearts have preferential access to oxygen through a well-developed coronary artery, therefore the heart is likely to be the last tissue to become hypoxic. Autophagy markers were upregulated in the liver, whereas protein degradation markers were downregulated in the heart during hypoxia. Further work is needed to determine the effects of a decrease in protein degradation on a hypoxic salmonid heart. Our study showed that protein metabolism in Arctic char is altered in a tissue-specific fashion during graded hypoxia, which is in accordance with the responses of the three major hypoxia-sensitive pathways (HIF, UPR and mTOR). The activation pattern of these pathways and the cellular processes that are under their control varies greatly among tissues, sometimes even going in the opposite direction. This study provides new insights on the effects of hypoxia on protein metabolism. Adjustment of these cellular processes is likely to contribute to shifting the fish phenotype into a more hypoxia-tolerant one, if more than one hypoxia event were to occur. Our results warrant studying these adjustments in fish exposed to long-term and diel cycling hypoxia.

Interrelationship Between Contractility, Protein Synthesis and Metabolism in Mantle of Juvenile Cuttlefish (Sepia officinalis).

Young juvenile cuttlefish (Sepia officinalis) can grow at rates as high as 12% body weight per day. How the metabolic demands of such a massive growth rate impacts muscle performance that competes for ATP is unknown. Here, we integrate aspects of contractility, protein synthesis, and energy metabolism in mantle of specimens weighing 1.1 g to lend insight into the processes. Isolated mantle muscle preparations were electrically stimulated and isometric force development monitored. Preparations were forced to contract at 3 Hz for 30 s to simulate a jetting event. We then measured oxygen consumption, glucose uptake and protein synthesis in the hour following the stimulation. Protein synthesis was inhibited with cycloheximide and glycolysis was inhibited with iodoacetic acid in a subset of samples. Inhibition of protein synthesis impaired contractility and decreased oxygen consumption. An intact protein synthesis is required to maintain contractility possibly due to rapidly turning over proteins. At least, 41% of whole animal MO2 is used to support protein synthesis in mantle, while the cost of protein synthesis (50 µmol O2 mg protein-1) in mantle was in the range reported for other aquatic ectotherms. A single jetting challenge stimulated protein synthesis by approximately 25% (2.51-3.12% day-1) over a 1 h post contractile period, a similar response to that which occurs in mammalian skeletal muscle. Aerobic metabolism was not supported by extracellular glucose leading to the contention that at this life stage either glycogen or amino acids are catabolized. Regardless, an intact glycolysis is required to support contractile performance and protein synthesis in resting muscle. It is proposed that glycolysis is needed to maintain intracellular ionic gradients. Intracellular glucose at approximately 3 mmol L-1 was higher than the 1 mmol L-1 glucose in the bathing medium suggesting an active glucose transport mechanism. Octopine did not accumulate during a single physiologically relevant jetting challenge; however, octopine accumulation increased following a stress that is sufficient to lower Arg-P and increase free arginine.

Differences in mitochondrial efficiency explain individual variation in growth performance.

The physiological causes of intraspecific differences in fitness components such as growth rate are currently a source of debate. It has been suggested that differences in energy metabolism may drive variation in growth, but it remains unclear whether covariation between growth rates and energy metabolism is: (i) a result of certain individuals acquiring and consequently allocating more resources to growth, and/or is (ii) determined by variation in the efficiency with which those resources are transformed into growth. Studies of individually housed animals under standardized nutritional conditions can help shed light on this debate. Here we quantify individual variation in metabolic efficiency in terms of the amount of adenosine triphosphate (ATP) generated per molecule of oxygen consumed by liver and muscle mitochondria and examine its effects, both on the rate of protein synthesis within these tissues and on the rate of whole-body growth of individually fed juvenile brown trout (Salmo trutta) receiving either a high or low food ration. As expected, fish on the high ration on average gained more in body mass and protein content than those maintained on the low ration. Yet, growth performance varied more than 10-fold among individuals on the same ration, resulting in some fish on low rations growing faster than others on the high ration. This variation in growth for a given ration was related to individual differences in mitochondrial properties: a high whole-body growth performance was associated with high mitochondrial efficiency of ATP production in the liver. Our results show for the first time, to our knowledge, that among-individual variation in the efficiency with which substrates are converted into ATP can help explain marked variation in growth performance, independent of food intake. This study highlights the existence of inter-individual differences in mitochondrial efficiency and its potential importance in explaining intraspecific variation in whole-animal performance.

Diel cycling hypoxia enhances hypoxia tolerance in rainbow trout (Oncorhynchus mykiss): evidence of physiological and metabolic plasticity.

Many fish naturally encounter a daily cycle of hypoxia, but it is unclear whether this exposure hardens hypoxia-intolerant fish to future hypoxia or leads to accumulated stress and death. The rainbow trout (Oncorhynchus mykiss) is a putatively hypoxia-sensitive species found in rivers and estuaries that may routinely experience hypoxic events. Trout were exposed to one of four 135 h treatments in a swim-tunnel respirometer: (1) air-saturated control (20.7 kPa PO2 ); (2) diel cycling O2 (20.7-4.2 kPa PO2  over 24 h); (3) acute hypoxia (130 h at 20.7 kPa PO2  followed by 5 h at 4.2 kPa PO2 ); and (4) the mean oxygen tension (12.4 kPa PO2 ) experienced by the diel cycled fish. Some responses were similar in diel O2 cycled and mean PO2 -treated fish, but overall, exposure to ecologically representative diel hypoxia cycles improved hypoxia tolerance. Diel hypoxia-induced protective responses included increased inducible HSP70 concentration and mean corpuscular hemoglobin concentration, as well as reduced plasma cortisol. Acclimation to diel hypoxia allowed metabolic rates to decline during hypoxia, reduced oxygen debt following subsequent exposures, and allowed fish to return to an anabolic phenotype. The data demonstrate that acute diel cycling hypoxia improves hypoxia tolerance in previously intolerant fish through the activation of cellular protective mechanisms and a reduction in metabolic O2 requirements.

Transcriptome and proteome analyses to investigate the molecular underpinnings of cold response in the Colorado potato beetle, Leptinotarsa decemlineata.

The Colorado potato beetle (Leptinotarsa decemlineata (Say)) is an insect that can cope with prolonged periods of low temperatures exposure. The molecular changes required to adapt to such conditions have not been thoroughly investigated in this insect. The current work aims at characterizing deregulated transcripts and proteins in adult L. decemlineata exposed to 15 °C and -5 °C using RNA-sequencing-based transcriptomics and liquid chromatography tandem mass spectrometry (LC-MS/MS)-based proteomics approaches, respectively. RNA-sequencing highlighted the differential expression of several transcripts, including ubiquilin-1 and ubiquitin carboxyl-terminal hydrolase 5, in insects submitted to low temperatures when compared with control insects. In addition, proteomics approach detected 2840 proteins in cold-exposed beetles including elevated levels for 409 proteins and reduced levels for 200 proteins. Cuticular proteins CP1, CP4, CP5 and CP7 as well as eukaryotic translation initiation factor 4B were notable proteins with elevated levels in cold insects. Functional analysis of targets modulated at low temperatures using DAVID indicated processes likely affected under cold conditions including select metabolic cascades and RNA-associated processes. Overall, this work presents molecular candidates impacted by low temperatures exposure in L. decemlineata and builds on the current knowledge associated with response to these conditions in this insect.

Reversion to developmental pathways underlies rapid arm regeneration in juvenile European cuttlefish, Sepia officinalis (Linnaeus 1758).

Coleoid cephalopods, including the European cuttlefish (Sepia officinalis), possess the remarkable ability to fully regenerate an amputated arm with no apparent fibrosis or loss of function. In model organisms, regeneration usually occurs as the induction of proliferation in differentiated cells. In rare circumstances, regeneration can be the product of naïve progenitor cells proliferating and differentiating de novo . In any instance, the immune system is an important factor in the induction of the regenerative response. Although the wound response is well-characterized, little is known about the physiological pathways utilized by cuttlefish to reconstruct a lost arm. In this study, the regenerating arms of juvenile cuttlefish, with or without exposure at the time of injury to sterile bacterial lipopolysaccharide extract to provoke an antipathogenic immune response, were assessed for the transcription of early tissue lineage developmental genes, as well as histological and protein turnover analyses of the resulting regenerative process. The transient upregulation of tissue-specific developmental genes and histological characterization indicated that coleoid arm regeneration is a stepwise process with staged specification of tissues formed de novo, with immune activation potentially affecting the timing but not the result of this process. Together, the data suggest that rather than inducing proliferation of mature cells, developmental pathways are reinstated, and that a pool of naïve progenitors at the blastema site forms the basis for this regeneration.