Enzymes In White Muscle Research Articles

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.

Air breathing and aquatic gas exchange during hypoxia in armoured catfish.

Air breathing in fish is commonly believed to have arisen as an adaptation to aquatic hypoxia. The effectiveness of air breathing for tissue O2 supply depends on the ability to avoid O2 loss as oxygenated blood from the air-breathing organ passes through the gills. Here, we evaluated whether the armoured catfish (Hypostomus aff. pyreneusi)-a facultative air breather-can avoid branchial O2 loss while air breathing in aquatic hypoxia, and we measured various other respiratory and metabolic traits important for O2 supply and utilization. Fish were instrumented with opercular catheters to measure the O2 tension (PO2) of expired water, and air breathing and aquatic respiration were measured during progressive stepwise hypoxia in the water. Armoured catfish exhibited relatively low rates of O2 consumption and gill ventilation, and gill ventilation increased in hypoxia due primarily to increases in ventilatory stroke volume. Armoured catfish began air breathing at a water PO2 of 2.5kPa, and both air-breathing frequency and hypoxia tolerance (as reflected by PO2 at loss of equilibrium, LOE) was greater in individuals with a larger body mass. Branchial O2 loss, as reflected by higher PO2 in expired than in inspired water, was observed in a minority (4/11) of individuals as water PO2 approached that at LOE. Armoured catfish also exhibited a gill morphology characterized by short filaments bearing short fused lamellae, large interlamellar cell masses, low surface area, and a thick epithelium that increased water-to-blood diffusion distance. Armoured catfish had a relatively low blood-O2 binding affinity when sampled in normoxia (P50 of 3.1kPa at pH 7.4), but were able to rapidly increase binding affinity during progressive hypoxia exposure (to a P50 of 1.8kPa). Armoured catfish also had low activities of several metabolic enzymes in white muscle, liver, and brain. Therefore, low rates of metabolism and gill ventilation, and a reduction in branchial gas-exchange capacity, may help minimize branchial O2 loss in armoured catfish while air breathing in aquatic hypoxia.

The effect of prolonged exercise training on swimming performance and the underlying biochemical mechanisms in juvenile common carp (Cyprinus carpio)

To investigate the effect of prolonged exercise training on swimming performance and the underlying biochemical mechanisms in juvenile common carp (Cyprinus carpio), we measured the critical swimming speed (Ucrit), the excess post-exercise oxygen consumption (EPOC), the activity of red and white muscle enzymes [pyruvate kinase (PK), lactate dehydrogenase (LDH) and citrate synthase (CS)], the tissue substrates (glycogen and glucose content of muscle and liver) and metabolite (the lactate content of plasma and muscle) content of exercise-trained (60% Ucrit for 4weeks) and non-trained fish. We also measured the biochemical indices of both trained and non-trained fish immediately after Ucrit, after exhaustive exercise and 1h after exhaustive exercise. The aerobic swimming performance, as indicated by Ucrit, increased significantly after exercise training, most likely because of the higher tissue metabolic capacity, as suggested by the higher CS activity in the red muscle tissue, and the higher energy store and more efficient substrate utilization, as suggested by higher liver and muscle glycogen contents at rest but lower tissue glycogen contents after Ucrit. The lower lactate content after Ucrit is most likely because of higher aerobic metabolic capacity, and (or) the clearance rate of lactate in trained fish may also contribute to improved aerobic swimming performance. Compared to Ucrit, exhaustive exercise elicited higher plasma and muscle lactate contents. The anaerobic metabolic performance is not affected by the exercise training, as suggested by the EPOC. However, trained fish did show higher lactate clearance rates, as suggested by lower muscle lactate content after a 1h recovery period following exhaustive exercise compared to non-trained fish. Furthermore, trained fish decreased their liver and muscle glycogen contents more profoundly after exhaustive exercise, suggesting that training can improve the substrate utilization during anaerobic exercise.

The correlation between fish growth and several biochemical characteristics with reference to the steelhead Parasalmo mykiss Walb.

Correlation analysis demonstrated a statistically significant correlation of linear-weight characteristics of the steelhead rainbow trout (cultivated steelhead form) with an RNA/DNA ratio and the expression level of the gene encoding cytochrome c oxidase (CCO) in two-year-old individuals (1+) as well as the expression level of the gene encoding the myosin heavy chain (MyHC) and activities of the enzymes CCO and lactate dehydrogenase (LDH) in muscles and 1-glycerophosphate dehydrogenase (1-GPDH) and glucose-6-phosphate dehydrogenase (G6PDH) in the liver of two- and three-year-old individuals (1+ and 2+). With age, the correlation of 1-GPDH and G6PDH activities in the rainbow trout liver with the fish body weight increased, whereas their correlations with the body length reduced. The age- and sex-related distinctions in the MyHC gene expression and activities of the white muscle enzyme LDH and the liver enzymes 2-GPDH and G6PDH were detected in rainbow trout of both age cohorts.

Muscle enzymes reveal walleye (Sander vitreus) are less active when larger prey (cisco,Coregonus artedi) are present

Optimal foraging behaviour in walleye (Sander vitreus) was tested in eight lakes: four containing large prey (cisco, Coregonus artedi) and four without cisco. All eight lakes contained small prey (yellow perch, Perca flavescens). Activity costs and growth potential of walleye were estimated using white muscle enzymes. Anaerobic capacity, measured by lactate dehydrogenase (LDH), increased with walleye size and was lower in lakes with cisco. Aerobic capacity, measured by citrate synthase (CS), decreased with walleye size only in lakes without cisco. Growth potential of walleye, estimated by nucleoside diphosphokinase (NDPK), increased with size only in lakes with cisco. Overall, when cisco were available walleye were less active, able to maintain aerobic capacities, and increased their growth potential as they grew larger. Yellow perch LDH, CS, and NDPK were lower in lakes with cisco. This suggests that yellow perch were less active in lakes where walleye had an alternative large prey species, but yellow perch had higher growth potential when they were the only prey, reflecting the advantage of growth beyond edible sizes. This study reveals that there are physiological benefits for both predators and prey in communities with a wider range of prey sizes.

High Swimming and Metabolic Activity in the Deep‐Sea EelSynaphobranchus kaupiiRevealed by Integrated In Situ and In Vitro Measurements

Several complementary studies were undertaken on a single species of deep-sea fish (the eel Synaphobranchus kaupii) within a small temporal and spatial range. In situ experiments on swimming and foraging behaviour, muscle performance, and metabolic rate were performed in the Porcupine Seabight, northeast Atlantic, alongside measurements of temperature and current regime. Deep-water trawling was used to collect eels for studies of animal distribution and for anatomical and biochemical analyses, including white muscle citrate synthase (CS), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), and pyruvate kinase (PK) activities. Synaphobranchus kaupii demonstrated whole-animal swimming speeds similar to those of other active deep-sea fish such as Antimora rostrata. Metabolic rates were an order of magnitude higher (31.6 mL kg(-1) h(-1)) than those recorded in other deep-sea scavenging fish. Activities of CS, LDH, MDH, and PK were higher than expected, and all scaled negatively with body mass, indicating a general decrease in muscle energy supply with fish growth. Despite this apparent constraint, observed in situ burst or routine swimming performances scaled in a similar fashion to other studied species. The higher-than-expected metabolic rates and activity levels, and the unusual scaling relationships of both aerobic and anaerobic metabolism enzymes in white muscle, probably reflect the changes in habitat and feeding ecology experienced during ontogeny in this bathyal species.

Changes in lactate dehydrogenase and malate dehydrogenase activities during hypoxia and after temperature acclimation in the armored fish, Rhinelepis strigosa (Siluriformes, Loricariidae).

Lactate (LDH) and malate dehydrogenase (MDH) of white skeletal muscle of fishes acclimated to 20, 25 and 30 degrees C and thereafter submitted to hypoxia were studied in different substrate concentrations. Significant differences for LDH and MDH of white muscle enzyme activities are described here for the first time in Rhinelepis strigosa of fishes acclimated to 20 degrees C and submitted to hypoxia for six hours. LDH presented a significant decrease in enzyme affinity for pyruvate in acute hypoxia, for fishes acclimated to 20 degrees C and an increase for fishes acclimated to 30 degrees C.

Bathymetric demography, enzyme activity patterns, and bioenergetics of deep-living scorpaenid fishes (genera Sebastes and Sebastolobus):paradigms revisited

Previous metabolic studies of scorpaenid fishes have measured interand intra-specific diffcrt,nccs in physiological and biochemical capacities in fishes llving at different depths. Small sample sizes and undocumented provenance of the samplesstudied have not allowed a full examination of the environmental b a s t for metabolic adaptation. The objective of this study was to determine the bathymetric demography of 22 scorpaenid fishes of the genera Sebastes and Sebastolobus, and to examine intera l n t r a s p e c i f ~ c hanges in metabolic capacity in 4 of them (Sebastes goodei, Sebastes diploproa, Sebastolobrrs alascanus and Sel~astolobus altivelis) representative of different environmental conditions. Weattempted to collect samples that spanned the tull depth and size ranges of each species. We examined changes in mefabolic capacity as it was affected by size, depth ot occurrence, and the existence of an oxygen minimum zone (OMZ) centered at 800 m. Lactate dehydrogenasc (LDH) was measured as an indicator of glycolytic capacity. Citrate synthase (CS), a Krebs cycle enzyme, was measured as an indicator 01 aerobic capacity, Midate dehydrogenase (MDH), an enzyme which helps maintain redox balance between the mltochondrion and cytosol, was also used as an indicator of aerobic capacity. When normal17ed to a common size. S. goodei, S. diploproa, S. alascanus and S. altivelis displayed an inter-specific pattern of declining metabolic activity with depth of occurrence. Intra-specific size scaling of glycolytic capacity (LDH) was positive in the shallow-livlng S. goodei. However, no scaling occurred in S diploproa and S. alascanus, and negative scaling of glycolytic activity occurred in the deep-living S. altivehs. MDH did not scale in S. goodei and S. diploproa but was negative in the 2 deeper-living Sebastolobus species. CS activity scaled negatively in S. goodei, S. alascanus, and S. altivelis, but the deep-watc!r specialist, S. altivelis, had the steepest slope. S. altivehs was the only species where similar-sized individuals could be compared over a wide depth range. There was no elvvdtion of LDH, or increase in anaerobic poise, in response to the hypoxia of the OMZ. Rather, all 3 enzyme activities were highest at 600 m and generally declined with ~ncreaslng depth. Contrary to the existing paradigm, S. alascanus and S. altivelis were not found to be shallowand deep-living congeners. Rather. S. altivelis is a permanent resident of deep water, whlle S. alascanus settles in shallow water then migrates to deep water with the onset of sexual maturity. When living in deep water, the supposed shallow water species. S. alascanus, maintains hlgher activities of all enzymes and grows an order of magnitude larger (>5000 vs <500 g) . In deep-water slope habitats expected patterns of allometric scaling of white muscle enzyme activities appear to be altered by the limitations of food and oxygen availability, a pattern previously observed in flatfishes.

Effects of Feeding and Food Deprivation on Oxygen Consumption, Muscle Protein Concentration and Activities of Energy Metabolism Enzymes in Muscle and Brain of Shallow-Living (Scorpaena Guttata) and Deep-Living (Sebastolobus Alascanus) Scorpaenid Fishes

ABSTRACT The effects of feeding and fasting were examined on the deep-living short-spine thornyhead (Sebastolobus alascanus) and the confamilial shallow-living spotted scorpionfish (Scorpaena guttata) to determine whether the low metabolic rate of the deeper-living species was in part a consequence of food deprivation in its habitat. Laboratory acclimation for periods of 90–115 days under either ad libitum feeding or complete fasting did not lead to similar rates of respiration in individuals of the two species held under identical conditions. Respiration of fish fed ad libitum was 52 % (S. guttata) or 68% (S. alascanus) higher than for fasted fish of the same species. Furthermore, the metabolic rates of freshly collected specimens of S. alascanus resembled those of laboratory-fasted fish. In white skeletal muscle, both total protein concentration and the activities of four enzymes of ATP metabolism, lactate dehydrogenase (LDH) and pyruvate kinase (PK) of glycolysis, malate dehydrogenase (MDH) and citrate synthase (CS, a citric acid cycle indicator), were lower in S. alascanus than in S. guttata. Within a species, protein concentration and activities of the four enzymes in white muscle, but not in brain, were higher in fed than in starved fish, although these differences were greater in S. alascanus than in S. guttata. During fasting, LDH and PK activity in white muscle of S. alascanus decreased much more than MDH and CS activity; decreases in enzyme activities in red muscle were smaller than those in white muscle. Activities of enzymes in white skeletal muscle of field-collected S. alascanus generally resembled those of the fasted specimens. In contrast, red muscle of field-collected S. alascanus, compared with that of either fed or starved laboratory-held specimens, had a highly glycolytic poise (high LDH and PK activities relative to MDH and CS activities), which may suggest that muscle enzyme activities in the field-collected fish reflect adaptation to the low oxygen level in its adult habitat, the oxygen minimum layer. The strong correlations found between tissue biochemical properties and respiration rate allow us to develop a predictive index for metabolic rate from simple biochemical analyses, e.g. white muscle protein content or CS activity. We conclude that the low metabolic rate of S. alascanus is due to at least four depth-related factors: reduced abundance of food, low temperature, low ambient oxygen concentration and darkness, which may select for reduced locomotory activity.

Does the aerobic capacity of fish muscle change with growth rates?

To ascertain whether growth rate modifies the oxidative capacity of fish white muscle, we examined the effects of individual growth rate on the activities of four mitochondrial enzymes in white muscle of the fast growing Atlantic cod,Gadus morhua. Growth rates were individually monitored in cod held at three acclimation temperatures during experiments repeated in four seasons. The size dependence of citrate synthase (CS), cytochrome C oxidase (CCO) and β-hydroxyacyl CoA dehydrogenase (HOAD) activities was established using wild cod ranging from 115 to 17,350 g. Given their negative allometry, CS and CCO activities in the experimental cod were corrected to those expected for a 1.2 kg animal. HOAD activities did not change with size. The specific activities of CCO and CS were positively correlated with growth rate. However, for both enzymes, season explained more of the variability than growth rate or temperature. Season was the only factor to significantly affect the activity of HOAD, while temperature and season interacted to determine glutamate dehydrogenase activity. CS activity was positively correlated with the initial condition of the cod, which differed among the seasons. The other enzymes did not show this relationship. The independent changes of these enzymes suggest that mitochondria undergo qualitative modifications with changes in growth rate, season and size. Although growth rate and the activities of CCO and CS are positively correlated, the activity of the mitochondrial enzymes is more affected by size, physical condition and season.

Gluconeogenesis in trout (Oncorhynchus mykiss) white muscle: purification and characterization of fructose-1,6-bisphosphatase activity in vitro

Studies of the enzyme fructose-1,6-bisphosphatase (FBPase) of rainbow trout (Oncorhynchus mykiss) have been undertaken in order to illuminate aspects of skeletal muscle gluconeogenesis in these animals. Maximal activities in crude homogenates of several organs suggest that the liver possesses the greatest FBPase activity on a unit g(-1) tissue basis but that the white muscle, owing to its bulk, contributes substantially to whole body FBPase activity. Studies of fructose-6-phosphate-1-kinase (PFK) and FBPase in crude homogenates of several organs suggests an important role for intracellular pH in regulating the relative carbon flux through the FBPase/PFK locus in vivo. Furthermore, a three-step purification scheme is described for trout white muscle FBPase by which a stable and homogeneous (by SDS PAGE) enzyme preparation (isoelectric point = 7.2; molecular weight = 37.6 kd) was obtained. Kinetic studies of the purified enzyme were undertaken at 20°C under conditions reflective of "rest" and "exercise/recovery" intramuscular pH in vivo. Affinity for substrate (F-1,6-P2) was increased (Km = 6.88 versus 2.44 μmol 1-(-1) as was enzyme activity when pH was lowered from 7.0 to 6.5. Various inhibitor metabolites are identified including F-2,6-P2 (mixed-type inhibitor, Ki = 0.201 μmol 1(-1), pH 7.0) and AMP (non-competitive inhibitor, Ki = 0.438 μmol 1(-1), pH 7.0). Inhibition by F-2,6-P2 was strongly alleviated by a reduction in pH from 7.0 to 6.5 (I50 increased from 0.14 to 0.32 μmol 1(-1)). AMP on the other hand was a more potent inhibitor at pH 6.5 but this inhibition was totally reversed under conditions of citrate, NH4 (+) and AMP typical of muscle during recovery from exercise in vivo. In purified white muscle enzyme preparations, FBPase demonstrated maximal activity at pH 6.5 whereas the optimal pH of PFK was 7.0 or greater. Indeed, it appears from these in vitro data that regulation by metabolite levels as well as pH are required for net FBPase flux in vivo. It is concluded, therefore that trout white muscle FBPase demonstrates the potential to play an important enzymatic role in the control of intramuscular gluconeogenesis in these animals. The results are discussed in relation to present knowledge regarding the metabolic responses of trout white muscle to, and its subsequent recovery from, exhaustive exercise.

Scaling of atp-supplying enzymes, myofibrillar proteins and buffering capacity in fish muscle: Relationship to locomotory habit

ABSTRACT Pelagic fishes with an ability to swim in strong bursts have previously been shown to have large size-dependent increases (positive allometric scaling exponents) in the activities of glycolytic enzymes in white skeletal muscle. This scaling of glycolytic activity has been hypothesized to provide the anaerobic power supporting the size-independence of relative burst swimming speeds (body lengths s−1) in these fishes. This paper presents tests of several predictions of this hypothesis, using different-sized individuals of two pelagic teleosts, the kelp bass (Paralabrax clathratus) and the freshwater rainbow trout (Salmo gairdneri), and a flatfish, the Dover sole (Microstomus pacificus). In the two pelagic species, an increase in body size was accompanied by an increase in activities in white muscle (i.u. g wet mass of muscle−1) of lactate dehydrogenase (LDH), an indicator of potential for anaerobic glycolysis, and creatine phosphokinase (CPK), an enzyme that helps maintain stable ATP concentration during muscular activity. Activities of citrate synthase (CS), an indicator of the potential for aerobic metabolism, decreased with size. In the flatfish, activities of all enzymes in white muscle decreased with body size, a trend proposed to reflect lack of adaptive value of strong burst swimming ability in this benthic fish. Activities of LDH and CS were size-independent in brain of flatfish, indicating that the scaling patterns observed in the muscle of this species were related to muscle function, not to common, organism-wide changes with size. In white muscle of P. clathratus, total protein and soluble protein concentrations and buffering capacity increased with body size in parallel, but myofibrillar protein was size-independent. These results suggest that the capacity for anaerobically powered work and the maximal potential to generate force scale only modestly in relation to total body mass and therefore do not appear to be functionally related to the pattern of glycolytic scaling. Thus, these data support the hypothesis that the functional role of the strongly positive scaling of glycolytic enzymes in the white muscle of pelagic fish is to provide increased power during burst swimming in larger-sized fishes.

Lactate dehydrogenase of goldfish red and white muscle in relation to thermal accumulation

1. 1. Activities of the red and white muscle LDH from 8°C-acclimated goldfish were about three times higher than those acclimated to 28°C. 2. 2. Isozyme composition and some kinetic properties of the red muscle LDH differed from those of the white muscle enzyme. 3. 3. The amount of red muscle as well as LDH activity tended to increase during cold acclimation.

Starvation and the activities of glycolytic and gluconeogenic enzymes in skeletal muscles and liver of the plaice,Pleuronectes platessa

Changes in body index parameters and liver, red muscle and white muscle enzyme profiles have been determined in fed and four month starved plaice,Pleuronectes platessa. The results are compared to other vertebrates to estimate specific tissue metabolic patterns and changes in these patterns with starvation. 1. Liver demonstrates the lowest glycolytic but highest gluconeogenic capacity of the three tissues. Red muscle has little, if any, gluconeogenic potential, based upon low activities of phosphoenol pyruvate carboxykinase and glucose-6-phosphatase and no detectable activities of pyruvate carboxylase. Plaice white skeletal muscle has the highest glycolytic potential of the tissues studied. 2. Plaice starved for four months demonstrate significant reductions in liver-somatic index and red muscle-somatic index, and increases in tissue water contents (Table 1). Enzyme activities generally decline in both muscle types, but are maintained in the liver (Table 2). Activities of liver soluble phosphoenol pyruvate carboxykinase increase by approximately 8-fold, suggesting that the enzymic response to starvation in plaice is similar to that of mammals. 3. These results suggest that starvation in plaice is associated with both a decrease in spontaneous activity and metabolic capacity of skeletal muscles, and an enhanced potential for liver gluconeogenesis. Also, it is possible that the precursors for liver gluconeogenesis do not form pyruvate as an intermediate step.

In vitro studies of skeletal muscle membranes. Adenylate cyclase of fast and slow twitch muscle and the effects of denervation.

Sarcolemmal membranes were prepared from slow-twitch (red) and fast-twitch (white) skeletal muscle of the rat. A sensitive adenylate cyclase assay was used and basal, fluoride- and catecholamine-stimulated activities measured. The greater in vivo sensitivity of red muscle to the effects of catecholamines correlates, in the present study, with approximately a twofold stimulation of its sarcolemmal adenylate cyclase with isoproterenol (10 micronm). The white muscle enzyme, on the other hand, is only minimally stimulated (20%) at the same concentration of beta-adrenergic agonist. Fast-twitch muscle is known to be physiologically insensitive to catecholamine in vivo. A course of sciatic nerve denervation was followed to further distinguish these two metabolic types of skeletal muscle and their respective adenylate cyclases. The slow-twitch muscle enzyme activities were completely and permanently lost on denervation. The white muscle enzyme, however, recovered almost completely after an initial reduction in specific activity the first week. Interestingly, the NaF-stimulated activity lagged behind both the basal and hormone-stimulated activities of the white muscle enzyme, in returning to control levels. The activities of cyclic nucleotide phosphodiesterase were evaluated in homogenates of the two muscle types in innervated rats and following denervation, in order to further define the neural influence on skeletal muscle cyclic nucleotide metabolism. The results suggest that the motor nerve may regulate some of the metabolic properties of slow-twitch muscle (which may involve cyclic AMP) by controlling the responsiveness of its sarcolemmal-bound adenylate cyclase system.

Serum and tissue enzymes and adrenocortical responses in rats exercised at intervals after cold acclimation.

Tolerance of rats to exercise at room temperature was studied at various intervals after cold acclimation. Survival was reduced when rats were exercised immediately after removal from the cold, but improved as time out of the cold was extended, attaining 100% survival after 24 h. The levels of serum enzymes, white muscle enzymes, and plasma corticosterone were greater in cold-acclimated rats exercised at 0 h or 6 h after removal from the cold than in unacclimated exercised rats. By 12–24 h after removal from the cold differences between the two groups of rats were less marked or absent. It is apparent that the greater response of cold-acclimated rats to exercise is lessened within 12 h and that a normal response is present within 24 h.

Tolerance of unacclimated and cold-acclimated rats to exercise in th cold: serum, red and white muscle enzymes, and histological changes.

Unacclimated and cold-acclimated rats were exercised for 3 h, 5 h, or 9 h in a cold room maintained at 1.7 °C. The cold-acclimated rats tolerated these exercise periods, but two-thirds of the unacclimated rats died during 9 h exercise. In red and white muscle the intermediate exercise interval (5 h) induced significantly greater increases in the activities in muscle of creatinephosphokinase and glycolytic enzymes of unacclimated rats, while during 9 h exercise enzyme activity declined in muscles of unacclimated rats and increased in cold-acclimated ones. The rise in serum enzyme activity during exercise was consistently greater in unacclimated than in cold-acclimated rats. Apparently the reduction in exercise tolerance was associated with and may have been in part due to loss of enzyme content and activity in muscles. Collectively, these and other biochemical responses suggested that homeostatic mechanisms had been exhausted in the rats dually stressed by cold exposure and exercise. Except for the activities of aldolase and the ratio of lactic dehydrogenase to alpha-glycerophosphate dehydrogenase, those enzymes associated with "aerobic" function (transaminases) showed the predominant changes in red muscle, and those associated with "anaerobic" function (glycolytic enzymes) the predominant changes in white muscle. The greater responses of the glycolytic enzymes in a predominantly "aerobic" tissue suggest that the biochemical adaptability of red muscle is greater than that of white muscle.