Antioxidant defenses of flame scallop Ctenoides scaber (Born, 1778) exposed to the water-soluble fraction of used vehicle crankcase oils.

Used vehicle crankcase oils are a source of contamination in Caribbean marine environments and may alter the oxidative balance of organism that inhabiting coastal ecosystems. This paper aims to evaluate effects of a water-soluble fraction of used vehicle crankcase oils (WSF-UVCO) on the antioxidant responses of the flame scallop Ctenoides scaber. The organisms were exposed to ascending sublethal concentrations 0, 0.001, 0.01 and 0.1 % of WSF-UVCO in a static system of aquaria during one week. Subsequently activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S-transferase (GST) as well as concentrations of reduced glutathione (GSH) and thiobarbituric acid reactive substances (TBARS) were determined in the digestive gland, adductor muscle and gills. SOD, CAT, GST and TBARS increased in digestive gland of organisms exposed to WSF-UVCO at medium and highest concentrations, with a concomitant decrease in GPX and GR activities. In adductor muscle CAT decreased, but GR rose with exposure to 0.01 and 0.1 % WSF-UVCO; in gills, GST rose through all WSF-UVCO concentrations, and SOD, CAT and GR increased only at 0.1 %. The fluctuations in antioxidant enzymes and GST activities point out possible adjustments to control ROS production and detoxification of xenobiotics. These biochemical responses may guarantee the oxidative balance in flame scallop during short term exposure to low concentrations of WSF-UVCO. C. scaber appears suitable as an experimental organism for evaluating biological risks of sublethal exposure to hazardous xenobiotics in tropical marine environments.

Do differences in the activities of carbohydrate metabolism enzymes between Lake Whitefish ecotypes match predictions from transcriptomic studies?

Transcriptomic studies are facilitating the search for the molecular bases of adaptation in natural populations, but the impact of these differences in mRNA content on animal physiology are often unknown. One way to determine if molecular changes have the potential to influence animal physiology and performance is to test for correlated changes at higher levels of biological organization, including enzyme activity. Here, we measure the activities of carbohydrate metabolism enzymes to test if previously documented genetic and transcriptomic variation between 'dwarf' and 'normal' Lake Whitefish ecotypes are associated with corresponding changes in enzyme activity (measured as maximal rate, Vmax) in liver and skeletal muscle. We use laboratory-reared fish from the same populations as prior transcriptomic studies and find that white muscle mRNA content is a good predictor of glycolytic and glycogen metabolism enzyme activity, and dwarf whitefish have evolved higher activities than normal whitefish. However, the differences in hepatic mRNA content found between ecotypes in prior studies are not associated with comparable changes in enzyme activity. For example, dwarf whitefish have lower enzyme activities, but higher transcript abundances for two glycolytic enzymes compared to normal whitefish. Overall, we find that transcriptomic studies successfully highlight evolutionary variation in enzyme activities, but not always in the direction predicted, indicating that a variety of tissue-specific regulatory mechanisms contributed to the evolution of energy metabolism in Lake Whitefish.

Convergence in organ size but not energy metabolism enzyme activities among wild Lake Whitefish (Coregonus clupeaformis) species pairs.

The repeated evolution of similar phenotypes by similar mechanisms can be indicative of local adaptation, constraints or biases in the evolutionary process. Little is known about the incidence of physiological convergence in natural populations, so here we test whether energy metabolism in 'dwarf' and 'normal' Lake Whitefish evolves by similar mechanisms. Prior genomic and transcriptomic studies have found that divergence in energy metabolism is key to local adaptation in whitefish species pairs, but that distinct genetic and transcriptomic changes often underlie phenotypic evolution among lakes. Here, we predicted that traits at higher levels of biological organization, including the activities of energy metabolism enzymes (the product of enzyme concentration and turnover rate) and the relative proportions of metabolically active tissues (heart, liver, skeletal muscle), would show greater convergence than genetic and transcriptomic variation. We compared four whitefish species pairs and found convergence in organ size whereby all dwarf whitefish populations have a higher proportion of red skeletal muscle, three have relatively larger livers and two have relatively larger ventricles than normal fish. On the other hand, hepatic and muscle enzyme activities showed little convergence and were largely dependent on lake of origin. Only the most genetically divergent species pair (Cliff Lake) displayed white muscle enzyme activities matching results from laboratory-reared normal and dwarf whitefish. Overall, these data show convergence in the evolution of organ size, but not in the activities of candidate enzymes of energy metabolism, which may have evolved mainly as a consequence of demographic or ecological differences among lakes.

Adaptation and acclimation of aerobic exercise physiology in Lake Whitefish ecotypes (Coregonus clupeaformis).

The physiological mechanisms underlying local adaptation in natural populations of animals, and whether the same mechanisms contribute to adaptation and acclimation, are largely unknown. Therefore, we tested for evolutionary divergence in aerobic exercise physiology in laboratory bred, size-matched crosses of ancestral, benthic, normal Lake Whitefish (Coregonus clupeaformis) and derived, limnetic, more actively swimming "dwarf" ecotypes. We acclimated fish to constant swimming (emulating limnetic foraging) and control conditions (emulating normal activity levels) to simultaneously study phenotypic plasticity. We found extensive divergence between ecotypes: dwarf fish generally had constitutively higher values of traits related to oxygen transport (ventricle size) and use by skeletal muscle (percent oxidative muscle, mitochondrial content), and also evolved differential plasticity of mitochondrial function (Complex I activity and flux through Complexes I-IV and IV). The effects of swim training were less pronounced than differences among ecotypes and the traits which had a significant training effect (ventricle protein content, ventricle malate dehydrogenase activity, and muscle Complex V activity) did not differ among ecotypes. Only one trait, ventricle mass, varied in a similar manner with acclimation and adaptation and followed a pattern consistent with genetic accommodation. Overall, the physiological and biochemical mechanisms underlying acclimation and adaptation to swimming activity in Lake Whitefish differ.

Dietary fatty acid composition and the homeostatic regulation of mitochondrial phospholipid classes in red muscle of rainbow trout (Oncorhynchus mykiss).

Although dietary lipid quality markedly affects fatty acid (FA) composition of mitochondrial membranes from rainbow trout red muscle (Oncorhynchus mykiss), mitochondrial processes are relatively unchanged. As certain classes of phospholipids interact more intimately with membrane proteins than others, we examined whether specific phospholipid classes from these muscle mitochondria were more affected by dietary FA composition than others. To test this hypothesis, we fed trout with two diets differing only in their FA composition: Diet 1 had higher levels of 18:1n-9 and 18:2n-6 than Diet 2, while 22:6n-3 and 22:5n-6 were virtually absent from Diet 1 and high in Diet 2. After 5 months, trout fed Diet 2 had higher proportions of phosphatidylcholine (PC) and less phosphatidylethanolamine (PE) in mitochondrial membranes than those fed Diet 1. The FA composition of PC, PE and cardiolipin (CL) showed clear evidence of regulated incorporation of dietary FA. For trout fed Diet 2, 22:6n-3 was the most abundant FA in PC, PE and CL. The n-6 FA were consistently higher in all phospholipid classes of trout fed Diet 1, with shorter n-6 FA being favoured in CL than in PC and PE. Despite these marked changes in individual FA levels with diet, general characteristics such as total polyunsaturated FA, total monounsaturated FA and total saturated FA were conserved in PE and CL, confirming differential regulation of the FA composition of PC, PE and CL. The regulated changes of phospholipid classes presumably maintain critical membrane characteristics despite varying nutritional quality. We postulate that these changes aim to protect mitochondrial function.

Platelets release mitochondria serving as substrate for bactericidal group IIA-secreted phospholipase A2 to promote inflammation.

Mitochondrial DNA (mtDNA) is a highly potent inflammatory trigger and is reportedly found outside the cells in blood in various pathologies. Platelets are abundant in blood where they promote hemostasis. Although lacking a nucleus, platelets contain functional mitochondria. On activation, platelets produce extracellular vesicles known as microparticles. We hypothesized that activated platelets could also release their mitochondria. We show that activated platelets release respiratory-competent mitochondria, both within membrane-encapsulated microparticles and as free organelles. Extracellular mitochondria are found in platelet concentrates used for transfusion and are present at higher levels in those that induced acute reactions (febrile nonhemolytic reactions, skin manifestations, and cardiovascular events) in transfused patients. We establish that the mitochondrion is an endogenous substrate of secreted phospholipase A2 IIA (sPLA2-IIA), a phospholipase otherwise specific for bacteria, likely reflecting the ancestral proteobacteria origin of mitochondria. The hydrolysis of the mitochondrial membrane by sPLA2-IIA yields inflammatory mediators (ie, lysophospholipids, fatty acids, and mtDNA) that promote leukocyte activation. Two-photon microscopy in live transfused animals revealed that extracellular mitochondria interact with neutrophils in vivo, triggering neutrophil adhesion to the endothelial wall. Our findings identify extracellular mitochondria, produced by platelets, at the midpoint of a potent mechanism leading to inflammatory responses.

Scallops show that muscle metabolic capacities reflect locomotor style and morphology.

Although all scallops swim using their adductor muscle to close their valves, scallop species differ considerably in how they use their muscle during escape responses, in parallel with the striking interspecific differences in shell morphology. This provides an excellent opportunity to study links between muscle metabolic capacities and animal performance. We found that the capacity for anaerobic glycolysis and aerobic metabolism, as well as phosphoarginine levels in the phasic adductor muscle, differ with escape response strategy. Phosphoarginine contents were high in species that rely on phasic contractions (Amusium balloti, Placopecten magellanicus, and Pecten fumatus). Arginine kinase activities reflect reliance on rapid initial bursts of phasic contractions. Scallops that maintain their valves in a closed position for prolonged periods (P. fumatus, Mimachlamys asperrima, and Crassadoma gigantea) have high activities of enzymes of anaerobic glycolysis in their phasic adductor muscle. Myosin ATPase activity was lower in the nonswimming scallop, C. gigantea, than in swimming scallops. The different patterns and roles of swimming are reflected in interspecific differences in the biochemical attributes of the phasic adductor muscle. These patterns suggest coevolution of muscle metabolic capacities, patterns of adductor muscle use, and shell morphology in scallops.

Swimming away or clamming up: the use of phasic and tonic adductor muscles during escape responses varies with shell morphology in scallops.

The simple locomotor system of scallops facilitates the study of muscle use during locomotion. We compared five species of scallops with different shell morphologies to see whether shell morphology and muscle use change in parallel or whether muscle use can compensate for morphological constraints. Force recordings during escape responses revealed that the use of tonic and phasic contractions varied markedly among species. The active species, Amusium balloti, Placopecten magellanicus and Pecten fumatus, made more phasic contractions than the more sedentary species, Mimachlamys asperrima and Crassadoma gigantea. Tonic contractions varied considerably among these species, with the two more sedentary species often starting their response to the predator with a tonic contraction and the more active species using shorter tonic contractions between series of phasic contractions. Placopecten magellanicus made extensive use of short tonic contractions. Pecten fumatus mounted an intense series of phasic contractions at the start of its response, perhaps to overcome the constraints of its unfavourable shell morphology. Valve closure by the more sedentary species suggests that their shell morphology protects them against predation, whereas swimming by the more active species relies upon intense phasic contractions together with favourable shell characteristics.

Thermal acclimation, mitochondrial capacities and organ metabolic profiles in a reptile (Alligator mississippiensis).

Reptiles thermoregulate behaviourally, but change their preferred temperature and the optimal temperature for performance seasonally. We evaluated whether the digestive and locomotor systems of the alligator show parallel metabolic adjustments during thermal acclimation. To this end, we allowed juvenile alligators to grow under thermal conditions typical of winter and summer, providing them with seasonally appropriate basking opportunities. Although mean body temperatures of alligators in these groups differed by approximately 10°C, their growth and final anatomic status was equivalent. While hepatic mitochondria isolated from cold-acclimated alligators had higher oxidative capacities at 30°C than those from warm-acclimated alligators, the capacities did not differ at 20°C. Cold acclimation decreased maximal oxidative capacities of muscle mitochondria. For mitochondria from both organs and acclimation groups, palmitate increased oligomycin-inhibited respiration. GDP addition reduced palmitate-uncoupled rates more in liver mitochondria from warm- than cold-acclimated alligators. In muscle mitochondria, carboxyatractyloside significantly reduced palmitate-uncoupled rates. This effect was not changed by thermal acclimation. The aerobic capacity of liver, skeletal muscle and duodenum, as estimated by activities of cytochrome c oxidase (COX), increased with cold acclimation. At acclimation temperatures, the activities of COX and citrate synthase (CS) in these organs were equivalent. By measuring COX and CS in isolated mitochondria and tissue extracts, we estimated that cold acclimation did not change the mitochondrial content in liver, but increased that of muscle. The thermal compensation of growth rates and of the aerobic capacity of the locomotor and digestive systems suggests that alligators optimised metabolic processes for the seasonally altered, preferred body temperature. The precision of this compensatory response exceeds that typically shown by aquatic ectotherms whose body temperatures are at the mercy of their habitat.

Plasticity of oxidative metabolism in variable climates: molecular mechanisms.

Converting food to chemical energy (ATP) that is usable by cells is a principal requirement to sustain life. The rate of ATP production has to be sufficient for housekeeping functions, such as protein synthesis and maintaining membrane potentials, as well as for growth and locomotion. Energy metabolism is temperature sensitive, and animals respond to environmental variability at different temporal levels, from within-individual to evolutionary timescales. Here we review principal molecular mechanisms that underlie control of oxidative ATP production in response to climate variability. Nuclear transcription factors and coactivators control expression of mitochondrial proteins and abundance of mitochondria. Fatty acid and phospholipid concentrations of membranes influence the activity of membrane-bound proteins as well as the passive leak of protons across the mitochondrial membrane. Passive proton leak as well as protein-mediated proton leak across the inner mitochondrial membrane determine the efficacy of ATP production but are also instrumental in endothermic heat production and as a defense against reactive oxygen species. Both transcriptional mechanisms and membrane composition interact with environmental temperature and diet, and this interaction between diet and temperature in determining mitochondrial function links the two major environmental variables that are affected by changing climates. The limits to metabolic plasticity could be set by the production of reactive oxygen species leading to cellular damage, limits to substrate availability in mitochondria, and a disproportionally large increase in proton leak over ATP production.

Enzyme activity in the aestivating green-striped burrowing frog (Cyclorana alboguttata).

Green-striped burrowing frogs (Cyclorana alboguttata) can depress their resting metabolism by more than 80% during aestivation. Previous studies have shown that this species is able to withstand long periods of immobilisation during aestivation while apparently maintaining whole muscle mass and contractile performance. The aim of this study was to determine the effect of prolonged aestivation on the levels of metabolic enzymes (CCO, LDH and CS) in functionally distinct skeletal muscles (cruralis, gastrocnemius, sartorius, iliofibularis and rectus abdominus) and liver of C. alboguttata. CS activity was significantly reduced in all tissues except for the cruralis, gastrocnemius and the liver. LDH activity was significantly reduced in the sartorius and rectus abdominus, but remained at control (active) levels in the other tissues. CCO activity was significantly reduced in the gastrocnemius and rectus abdominus, and unchanged in the remaining tissues. Muscle protein was significantly reduced in the sartorius and iliofibularis during aestivation, and unchanged in the remaining muscles. The results suggest that the energy pathways involved in the production and consumption of ATP are remodelled during prolonged aestivation but selective. Remodelling and subsequent down-regulation of metabolic activity seem to target the smaller non-jumping muscles, while the jumping muscles retain enzyme activities at control levels during aestivation. These results suggest a mechanism by which aestivating C. alboguttata are able to maintain metabolic depression while ensuring that the functional capacity of critical muscles is not compromised upon emergence from aestivation.

Muscle metabolic capacities and plasma cortisol levels of the male three-spine stickleback Gasterosteus aculeatus: are there "femme fatale" or "macho male" effects?

To evaluate whether decreases in muscle metabolic capacities and increases in plasma cortisol explain the effects of neighboring conspecifics on male three-spine sticklebacks Gasterosteus aculeatus, we housed mature males alone, with a mature female, or with a rival mature male. The neighbors were separated from the focal male by a partition that allowed him to smell, see, and hear his neighbor. In the first experiment, focal males were allowed to reproduce, whereas in the second experiment, no reproduction occurred. Coloration and behaviors were monitored while the males tended their nests (or for the same period in the second experiment). The presence of a neighbor markedly affected the reproductive coloration of the focal male, with solitary males being less colorful than males housed with a rival male or a female. Solitary males showed greater aggression toward a model male stickleback than did males with neighbors. The presence of neighbors affected the anatomic and metabolic characteristics of the focal males primarily during nesting, when males housed with rival males had a lower hepatosomatic index and lower activities of mitochondrial and glycolytic enzymes in the axial muscle than did solitary males or males housed with females. Cortisol levels were highly variable in nesting males and did not differ with social condition but were higher in males that had been quick to construct their nests. On the other hand, when focal males were not provided with nesting material, solitary males tended to have lower cortisol levels than did males housed with rival males. While these results do not provide a mechanism for a "femme fatale" effect, they indicate that nesting males decrease metabolic status when housed with rival "macho males."

Effect of day length on oxidative capacities of mitochondria from red muscle of rainbow trout (Oncorhynchus mykiss).

In nature, seasons may be more reliably announced by changes in photoperiod than in temperature. To evaluate the role of day length in setting oxidative capacities of trout muscle mitochondria, we acclimated trout to summer (15 degrees C, 16L:8D), winter (5 degrees C, 8L:16D) and mixed conditions (15 degrees C, 8L:16D). Maximal oxidative capacities of isolated mitochondria at 5 and 15 degrees C were higher in mixed than summer conditions and higher again in winter conditions. At 5 degrees C, state 4 rates changed little with acclimation state whereas at 15 degrees C state 4 rates were lower in summer than in mixed or winter conditions. Using concentrations of the adenylate nucleotide translocase as the denominator for these rates gave much the same conclusions. By using inhibitors to block flux at specific points in the electron transport chain, we found that flux through Complexes II-IV was lowest in summer acclimated trout, increased upon acclimation to mixed and to winter conditions. Flux through complex IV was similar in trout acclimated to summer and mixed conditions, but increased significantly with acclimation to winter conditions. Flux through complex IV was 1.5 fold higher than state 3 rates for summer-acclimated trout but was similar to state 3 rates in trout acclimated to mixed or winter conditions. Our results indicate that a reduction in day length initiates increases in mitochondrial oxidative capacity typically associated with cold acclimation and that acclimation to both cold temperatures and short day lengths enhanced these changes. The overall similarity of the responses of state 3, of flux through complexes II-IV and of flux through complex IV suggests that a generalised mechanism such as changes in the phospholipid composition of the inner mitochondrial membrane may coordinate these changes.

Reduction of type IIb myosin and IIB fibers in tibialis anterior muscle of mini-muscle mice from high-activity lines.

Selective breeding of laboratory house mice (Mus domesticus) for high voluntary wheel running has generated four replicate lines that show an almost threefold increase in daily wheel-running distances as compared with four nonselected control lines. An unusual hindlimb "mini-muscle" phenotype (small muscles, increased mitochondrial enzyme levels, disorganized fiber distribution) has increased in frequency in two of the four replicate selected lines. The gene of major effect that accounts for this phenotype is an autosomal recessive that has been mapped to a 2.6335 Mb interval on MMU11, but not yet identified. This study examined the tibialis anterior muscle to determine whether changes in muscle fiber types could explain such modifications in muscle size and properties. Although selected and control lines did not exhibit systematic differences in the fiber types present in the tibialis anterior muscle, as assessed by electrophoresis of myosin heavy chains (MHC) and by histochemistry, mini-muscle mice lacked type IIB fibers and the corresponding MHCs. Mini-muscle tibialis show increased activities of hexokinase and citrate synthase compared with the normally sized muscles, likely the result of the modified fiber types in the muscle. The mini-muscle phenotype is the major means through which selective breeding for high wheel running has modified the functional capacities of the hindlimb muscles, as normally sized tibialis anterior muscles from control and selected lines did not show general differences in their enzymatic capacities, MHC profiles or fiber type composition, with the exception of an elevated hexokinase activity and a reduced GPa activity in the selected lines.

Paternal reproductive strategy influences metabolic capacities and muscle development of Atlantic salmon (Salmo salar L.) embryos.

Male Atlantic salmon follow a conditional strategy, becoming either "combatants" that undertake a seaward migration and spend at least a year at sea or "sneakers" that remain in freshwater and mature as parr. A variety of physiological indices showed significant but small differences between the offspring of males that use these two reproductive tactics. Offspring fathered by anadromous male Atlantic salmon (Salmo salar L.) showed greater muscular development and muscle metabolic capacities but lower spontaneous movements than those fathered by mature male parr. At hatch and at maximum attainable wet weight (MAWW), offspring fathered by anadromous males had higher activities of mitochondrial (cytochrome C oxidase and citrate synthase) and glycolytic (lactate dehydrogenase [LDH]) enzymes than progeny of mature male parr. Enzymatic profiles of progeny of anadromous fathers also suggested greater nitrogen excretion capacity (glutamate dehydrogenase) and increased muscular development (creatine kinase and LDH) than in the progeny of mature parr. At MAWW, juveniles fathered by mature parr made considerably more spontaneous movements, presumably increasing their energy expenditures. For juveniles fathered by anadromous males, total cross-sectional areas of white and red muscle at hatch were higher due to the greater number of large-diameter fibers. We suggest that the slightly lower metabolic capacities and muscular development of alevins fathered by mature parr could reflect differences in energy partitioning during their dependence on vitellus. Greater spontaneous movements of offspring of mature male parr could favor feeding and growth after the resorption of the vitellus.

Disturbance of social hierarchy by an invasive species: a gene transcription study.

BACKGROUND: Ecological and evolutionary changes in native populations facing invasion by exotic species are increasingly reported. Recently, it has been shown that competition with exotic rainbow trout (Oncorhynchus mykiss) disrupts dominance hierarchies within groups of native Atlantic salmon (Salmo salar). The genetic and molecular actors underlying phenotypic plasticity are poorly understood. METHODOLOGY: Here, we aimed at identifying the genetic and molecular actors contributing to this plastic loss of dominance hierarchies as well as at identifying genes implicated in behaviours related to social dominance. By using microarrays, we compared the genome-wide gene transcription profiles in brains of dominant versus subordinate juvenile Atlantic salmon in presence or absence of a competitive rainbow trout. PRINCIPAL FINDINGS: Adding the trout competitor resulted in dominant and subordinate salmon being more similar, both behaviourally and at the level of brain gene transcription patterns. Genes for which transcription levels differed between dominant and subordinate salmon in the absence of exotic trout were mainly over-expressed in dominant salmon and included genes implicated in protein turnover, neuronal structural change and oxygen transport. CONCLUSIONS/SIGNIFICANCE: Our study provides one of the few examples demonstrating a close interplay between behavioural plasticity and gene transcription, therefore contributing to the understanding of the molecular mechanisms underlying these processes in an ecologically relevant context.

Genetic consequences of interbreeding between farmed and wild Atlantic salmon: insights from the transcriptome.

Large annual escapees of farmed Atlantic salmon enhance the risk of extinction of wild populations through genetic and ecological interactions. Recently, we documented evolutionary change in gene transcription between farmed and wild Atlantic salmon after only five generations of artificial selection. While differences for most quantitative traits are expected to gradually dilute through repeated backcrossing to wild populations, the genetic basis of gene transcription has been shown to be largely nonadditive and hybrid crosses may display unexpected inheritance patterns. This makes it difficult to predict to what extent interbreeding between farmed and wild individuals will change the genetic makeup of wild salmon populations. Here, we compare the genome-wide gene transcription profiles of Norwegian wild salmon to that of a second generation hybrid cross [backcross: (Farmed X Wild) X Wild]. Over 6% (298, q-value < 0.01) of the detected genes exhibited highly significantly different transcription levels, and the range and average magnitude of those differences was strikingly higher than previously described between pure farmed and wild strains. Most differences appear to result from nonadditive gene interactions. These results suggest that interbreeding of fugitive farmed salmon and wild individuals could substantially modify the genetic control of gene transcription in natural populations exposed to high migration from fish farms, resulting in potentially detrimental effects on the survival of these populations. This further supports the idea that measures to considerably reduce the number of escaped farmed salmon and their reproduction in the wild are urgently needed.

Altered fibre types in gastrocnemius muscle of high wheel-running selected mice with mini-muscle phenotypes.

Selective breeding of mice for high voluntary wheel running has favoured characteristics that facilitate sustained, aerobically supported activity, including a "mini-muscle" phenotype with markedly reduced hind limb muscle mass, increased mass-specific activities of oxidative enzymes, decreased % myosin heavy chain IIb, and, in the medial gastrocnemius, reduced twitch speed, reduced mass-specific isotonic power, and increased fatigue resistance. To evaluate whether selection has altered fibre type expression in mice with either "mini" or normal muscle phenotypes, we examined fibre types of red and white gastrocnemius. In both the medial and lateral gastrocnemius, the mini-phenotype increased activities of oxidative enzymes and decreased activities of glycolytic enzymes. In red muscle samples, the mini-phenotype markedly changed fibre types, with the % type I and type IIA fibres and the surface area of type IIA fibres increasing; in addition, mice from selected lines in general had an increased % type IIA fibres and larger type I fibres as compared with mice from control lines. White muscle samples from mini-mice showed dramatic structural alterations, with an atypical distribution of extremely small, unidentifiable fibres surrounded by larger, more oxidative fibres than normally present in white muscle. The increased proportion of oxidative fibres and these atypical small fibres together may explain the reduced mass and increased mitochondrial enzyme activities in mini-muscles. These and previous results demonstrate that extension of selective breeding beyond the time when the response of the selected trait (i.e. distance run) has levelled off can still modify the mechanistic underpinnings of this behaviour.

Temperature adaptation in two bivalve species from different thermal habitats: energetics and remodelling of membrane lipids.

We compared lipid dynamics and the physiological responses of blue mussels Mytilus edulis, a cold-adapted species, and oysters Crassostrea virginica, a warmer-water species, during simulated overwintering and passage to spring conditions. To simulate overwintering, animals were held at 0 degrees C, 4 degrees C and 9 degrees C for 3 months and then gradually brought to and maintained at 20 degrees C for 5 weeks to simulate spring-summer conditions. Changes in lipid class and fatty acid composition were related to clearance rate and oxygen consumption. We found major differences between species in triglyceride (TAG) metabolism during overwintering. Mussels used digestive gland TAG stores for energy metabolism or reproductive processes during the winter, whereas oysters did not accumulate large TAG stores prior to overwintering. Mussel TAG contained high levels of 20:5n-3 compared to levels in oysters and in the diet. This may help to counteract the effect of low temperature by reducing the melting point of TAG and thus increasing the availability of storage fats at low temperature. Mussels seemed better able to mobilise 20:5n-3 and 18:4n-3 than other fatty acids. We also found that both bivalves underwent a major remodelling of membrane phospholipids. The unsaturation index decreased in the gills and digestive glands of both species during the early stages of warming, principally due to decreases in 22:6n-3 and 20:5n-3. In digestive glands, the unsaturation index did not increase with decreasing temperature beyond a threshold attained at 9 degrees C whereas a perfect negative relationship was observed in gills, as predicted by homeoviscous adaptation. The presence of digestive enzymes and acids in the digestive gland microenvironment may lead to specific requirements for membrane stability. That oysters had lower metabolic rates than mussels coincides with a lower unsaturation index of their lipids, as predicted by Hulbert's theory of membranes as metabolic pacemakers. Both species showed increased 20:4n-6 levels in their tissues as temperature rose, suggesting an increasing availability of this fatty acid for eicosanoid biosynthesis during stress responses. The contrast between the species in TAG dynamics and the similarity of their phospholipid remodelling emphasises the essential functional role of membrane phospholipid structure and the contrasting use of TAG by oysters and mussels during overwintering.

Thermal sensitivity of metabolic rates and swimming performance in two latitudinally separated populations of cod, Gadus morhua L.

Atlantic cod populations live in a wide thermal range and can differ genetically and physiologically. Thermal sensitivity of metabolic capacity and swimming performance may vary along a latitudinal gradient, to facilitate performance in distinct thermal environments. To evaluate this hypothesis, we compared the thermal sensitivity of performance in two cod stocks from the Northwest Atlantic that differ in their thermal experience: Gulf of St Lawrence (GSL) and Bay of Fundy (BF). We first compared the metabolic, physiological and swimming performance after short-term thermal change to that at the acclimation temperature (7 degrees C) for one stock (GSL), before comparing the performance of the two stocks after short-term thermal change. For cod from GSL, standard metabolism (SMR) increased with temperature, while active metabolism (AMR, measured in the critical swimming tests), EMR (metabolic rate after an exhaustive chase protocol), aerobic scope (AS) and critical swimming speeds (U (crit) and U (b-c)) were lower at 3 degrees C than 7 or 11 degrees C. In contrast, anaerobic swimming (sprint and burst-coasts in U (crit) test) was lower at 11 than 7 or 3 degrees C. Factorial AS (AMR SMR(-1)) decreased as temperature rose. Time to exhaustion (chase protocol) was not influenced by temperature. The two stocks differed little in the thermal sensitivities of metabolism or swimming. GSL cod had a higher SMR than BF cod despite similar AMR and AS. This led factorial AS to be significantly higher for the southern stock. Despite these metabolic differences, cod from the two stocks did not differ in their U (crit) speeds. BF cod were better sprinters at both temperatures. Cod from GSL had a lower aerobic cost of swimming at intermediate speeds than those from BF, particularly at low temperature. Only the activity of cytochrome C oxidase (CCO) in white muscle differed between stocks. No enzymatic correlates were found for swimming capacities, but oxygen consumption was best correlated with CCO activity in the ventricle for both stocks. Overall, the stocks differed in their cost of maintenance, cost of transport and sprint capacity, while maintaining comparable thermal sensitivities.