Der einfluß der temperatur auf thermostabilität,isoenzym-muster und reaktionskinetik der laktat-dehydrogenase aus fischen

Methodological problems complicate investigations on thermostability of lactate dehydrogenase (LDH). It is difficult to demonstrate a correlation between adaptation-temperature (AT) and LDH thermostability. Heat-inactivation characteristics change completely if diluted or undiluted tissue extracts are heated. In purified LDH (purchased from Boehringer, Mannheim, FRG), additions such as casein, bovine-serum albumin, NADH and pyruvate — even in small concentrations — can alter considerably the degree of heat resistance. It LDH activity is measured as a function of experimental temperature (ET) according to the composition of the actual test mixture (e.g. altered pyruvate concentration), a different temperature optimum is found. If tissue extracts containing unpurified enzymes are used, the accompanying substances act on the enzyme and modify its properties. Thus, possible influence of AT on enzyme properties can be concealed (suppressed, over-emphasized). In Idus idus acclimated to 10° or 20°C, brain, gill, gut and white dorsal muscle reveal identical LDH-isoenzyme patterns. However, liver-LDH shows a pattern dependent on the AT. A total of 11 bands with LDH activity were found. In 10°C fishes, the Isoenzymes 1, 3, 6 and 7 are especially active. However, 20°C fishes show marked activity of Isoenzymes 5 and 8, and a reduced activity of Isoenzyme 7. According to their electrophoretic mobility, the particular isoenzymes of LDH of white dorsal muscle of I. idus or Rhodeus amarus can be clearly distinguished. The ATs 10° or 20°C do not influence the dependence of reaction order on ET: this is not true for the velocity constant.     

Lactate and succinate oxidoreductases in marine invertebrates

Nineteen species of littoral marine invertebrates, representing Porifera, Cnidaria, Ctenophora, Brachiopoda, Mollusca, and Arthropoda, were studied with respect to the ability of tissue extracts to catalyze the lactate and succinate dehydrogenase reactions in both directions. Pyruvate reductase (PR) activity varied tremendously with species, from 0.014 mole/min/g of tissue in the etenophore Mnemiopsis leidyi to 145 -moles/min in leg muscle of the horseshoe crab Limulus polyphemus, a 10,000-fold range. Lactate dehydrogenase (LD) activity varied from 0.010 in the ctenophore to 2.91 in the squid Loligo pealei, a 300-fold range. The ratio PR/LD, indicating the probability of lactic acid production, was 1,968 in muscle of the flounder, the only vertebrate studied. It was very much lower in all invertebrates; in a brachiopod, Terebratulina septentrionalis, the ratio was only 0.68. Fumarate reductase (FR) and succinate dehydrogenase (SD) activities varied less widely. The extremes of the ratio FR/SD, indicating the probability of succinic acid production, were 0.24 in the clam Mercenaria mercenaria, and 7.6 in the oyster Crassostrea virginica. PR/LD appears related to the capacity for vigorous muscular activity, sustaining rapid movement of larger animals, and FR/SD appears related to tolerance of temporary anaerobiosis, such as found in sessile animals that close their valves tightly on exposure to air during low tide.     

Lactate formation in Callianassa californiensis and Upogebia pugettensis (Crustacea: Thalassinidea)

The ability to accumulate lactate as a result of laboratory exposure to anoxia was examined in 2 species of mud-dwelling shrimp, the ghost shrimp Callianassa californiensis and the mud shrimp Upogebia pugettensis. Hemolymph lactate accumulated to a much greater degree in the mud shrimp, even though the overall levels of lactate dehydrogenase activity were similar in tissues of the two species. Carbohydrate reserves, estimated as glycogen, were not significantly depleted in muscle or midgut gland of either species as a result of anoxic stress. The results are discussed in relation to possible metabolic strategies employed to cope with the hypoxic environment of these crustaceans.     

Sodium cyanide-induced modulations in the activities of some oxidative enzymes and metabolites in the fingerlings of Cyprinus carpio (Linnaeus)

Carp fingerlings exposed to a sublethal concentration (0.5?mg?L^?1) of sodium cyanide showed a steady decrement over a 7-day period in respiratory rate, rise in lactate dehydrogenase (LDH), and fall in succinate dehydrogenase (SDH) activities followed by variations in lactic and pyruvate levels. Changes in these enzyme activities might be due to impaired oxidative metabolism and severe cellular damage leading to the release of these enzymes. Decline in the activities of SDH and LDH clearly represents a shift from aerobic to anaerobic metabolism as evidenced by elevated lactate and decline in pyruvate levels. The shift to anaerobic metabolism is also reflected by severe drop in the respiratory rate of the fish. This may be a consequence of the blockage of electron transfer from cytochrome c oxidase to molecular oxygen, thus ceasing cellular respiration and it can lead to cellular hypoxia even in the presence of normal hemoglobin oxygenation. Hence, we indirectly reconfirm the inhibition of oxidative metabolism by sodium cyanide. Alterations in behavioral pattern induced by sublethal sodium cyanide exposure may be due to the combination of cytotoxic hypoxia with lactate acidosis, which depresses the central nervous system (CNS); as the brain is the most sensitive site to anoxia, it results in impaired CNS function.     

Enzyme activities of fish skeletal muscle and brain as influenced by depth of occurrence and habits of feeding and locomotion

Activities of lactate dehydrogenase (LDH), pyruvate kinase (PK), malate dehydrogenase (MDH) and citrate synthase (CS) were measured in the white skeletal muscle of marine fishes having different depths of occurrence and different feeding and locomotory strategies. There were significant depth-related differences in the two glycolytic enzymes, LDH and PK. LDH activity was most variable, and differed by 3 orders of magnitude between the most active shallow-living species and certain deep-sea fishes likely to have only minimal capacities for active locomotion. Superimposed on the depth-related patterns was a high degree of interspecific variation (up to 20-fold) in enzymic activity among species from any given range of depth of occurrence. Variation of both LDH and PK activities, noted for shallow- and deep-living fishes, seems to be largely accounted for by differences in feeding habits and locomotory performance. Active pelagic swimmers have much higher activities of LDH and PK than, for example, deep-living sit-and-wait predators. Benthopelagic fishes like rattails and the sablefish have the highest activities found among deep-living fishes, suggesting that these species engage in relatively active food-searching behavior compared to most other deep-sea fishes. The activity of CS, an enzyme of the citric acid cycle and an indicator of aerobic metabolism, varied little among species. Thus, the large interspecific variation in glycolytic potential (LDH and PK) among species is not associated with a similar variation in aerobic metabolism of white muscle. The much higher and more variable activity of MDH relative to CS suggests that, in addition to its function in the citric acid cycle, MDH may play an important role in redox balance in fish white muscle. In a comparison of white muscle composition between the shallow- and deep-living species, water content did not differ significantly, but protein content was significantly higher in shallow- than in deep-living fishes (211 and 130 mg g^-1 wet wt of muscle, respectively). The differences in muscle protein content are small relative to the differences between shallow- and deep-living species in LDH, PK and MDH activities. Thus, depthrelated differences in muscle enzymic activity are caused by factors other than enzyme dilution. Enzyme activities (LDH, PK and CS) in brain tissue were relatively constant among species regardless of depth of occurrence or feeding and locomotory habits. Habitat and lifestyle do not seem to influence the demands for overall metabolic function in brain. The utility of muscle enzymic activity data for making predictions about the ecological characteristics of difficult-to-observe, deep-living, fishes is discussed.     

Molecular mechanisms of adaptation to low temperature in marine poikilotherms. Some regulatory properties of dehydrogenases from two arctic species

The properties of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase from gill tissue of the tanner crab Chionocetes bairdi, and lactate dehydrogenase (LDH) and glyceraldehyde dehydrogenase from skeletal muscle of C. bairdi and the yellowfin sole Limanda aspera were examined over the physiological temperature range of the animals. Both animals were obtained in the Bering Sea in winter, and their enzymes appear to be remarkably cold-adapted. Affinity of sole LDH for substrate appears to increase with decreasing temperature, thus keeping reaction rate essentially independent of temperature at physiological concentrations of the substrate. Calculated values of activation energy are low, in keeping with the argument that organisms from cold environments have enzymes with a reduced energy of activation. In addition, Hill plots of the substrate saturation curves for lactate dehydrogenase from muscle of sole indicate that there is a facilitation of allosteric behaviour at low temperatures. Maximum affinity of sole LDH for substrate in the absence of univalent cations occurs at 3°C, while in the presence of 150 mN K⁺, it occurs between 0° to-2°C. The effects of Mg²⁺ on enzyme activity appear to be determined by concentration of substrate and temperature. Thus, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase are stimulated more effectively by Mg²⁺ at low temperature and at low substrate levels whereas, at high concentrations of substrate, they are relatively independent of the bivalent cation. All four dehydrogenases are affected by the univalent cations Na⁺, K⁺ and NH₄ ⁺ in a manner which appears to be determined, in part at least, by concentration of substrate and by temperature. These findings suggest mechanisms for the maintenance and regulation of enzyme activity in poikilothermic tissues at low and changing temperatures.     

Depth zonation and metabolic adaptation in Dover sole,Microstomus pacificus,and other deep-living flatfishes: factors that affect the sole

Flatfishes of Monterey Bay, central California, undergo species replacements with increasing depth along a transect from 100 m on the continental shelf down to a depth of 1400 m on the continental slope. The Dover sole, Microstomus pacificus, differs from the other local flatfish species by undergoing an extensive ontogenetic vertical migration, occupying all depth zones at different life stages, and having its maximum spawning biomass in the oxygen minimum zone between 600 and 1000 m. Size-activity relationships and depth-activity relationships for the glycolytic enzyme lactate dehydrogenase (LDH) and for two enzymes associated with aerobic metabolism, malate dehydrogenase and citrate synthase (CS), were examined in white-muscle tissue of shallow-living, deep-living and ontogenetically-migrating species. Scaling coefficients (b) for weight-specific enzyme activity (log activity)=a+b (log wet weight), varied in sign as well as magnitude for fishes living at different depths. In the shallow-living California halibut Paralichthys californicus, LDH scaled positively (0.39) and CS scaled negatively (-0.15) with size, a pattern observed previously for most shallow-water fish species. The permanently deep-living species, the deepsea sole Embassichthys bathybius, differed in that both LDH and CS scaled strongly negative (-2.0 and-1.5, respectively). For the ontogenetically migrating Dover sole Microstomus pacificus, there was a shelf-slope transition. For the shelf specimens (200 m), LDH scaled positive (0.11) and CS negative (-0.29) and for the slope specimens (400 m), LDH scaled negative (-0.65) and CS strongly negative (-0.63). Rex sole, Glyptocephalus zachirus, showed a similar shelf-slope transition. Intraspecific depth-enzyme activity differences were not incremental, but changed abruptly between the continental shelf stations (100 to 200 m) and the continental slope (400 to 1400 m). Based on comparisons with laboratory-maintained individuals, the decline in the metabolic capacity of the white muscle of Dover sole is a phenotypic response to the low food and oxygen conditions of the continental slope. Contrary to expectation, anaerobic capacity (LDH activity) decreased in response to low oxygen conditions, suggesting that in a permanently hypoxic environment such as the oxygen minimum zone the metabolic strategy may be to not incur an oxygen debt that would be difficult to pay back.     

Activities of metabolic enzymes in the deep-water crabsChaceon fenneri andC. quinquedens and the shallow-water crabCallinectes sapidus

The activities of 18 enzymes were measured in gill, hepatopancreas and muscle tissue of the deep-water crabsChaceon fenneri andC. quinquedens and the shallow-water crabCallinectes sapidus collected from the Gulf of Mexico in January 1989. The activities of catabolic enzymes were correlated in general with the known metabolic rates of the three species. Activities were much higher inC. sapidus than inChaceon fenneri andC. quinquedens. In some cases,C. quinquedens had higher activities thanC. fenneri. The activities of enzymes of amino acid metabolism (glutamate dehydrogenase and alanine aminotransferase) were higher inC. quinquedens, which had high hemolymph [ammonia] and ammonia excretion rates. The activity of lactate dehydrogenase (LDH) ofC. fenneri andC. quinquedens were correlated with the two species' abilities to withstand hypoxia. The more hypoxiatolerant species,C. quinquedens, had higher activity of LDH in its muscles than didC. fenneri.     

Metabolic and molecular stress responses of the gilthead seabream Sparus aurata during long-term exposure to increasing temperatures

Tolerance to a changing climate regime and persistence in the natural environment depends on the limited capacity to acclimate to changing temperatures. The present study aimed to identify and characterize thermal limits of the Mediterranean fish Sparus aurata as well as the processes providing heat protection during exposure to high temperatures. Processes studied included heat shock protein expression, protein kinase activity and metabolic adjustments. Molecular responses were addressed through the expression of Hsp70 and Hsp90 and the phosphorylation of stress-activated protein kinases, p38 mitogen-activated protein kinase (p38 MAPK) and cJun-N-terminal kinases (JNKs). Thermal impacts on metabolic capacities were assessed by studying the maximum activities of citrate synthase (CS), malate dehydrogenase (MDH) and 3-hydroxyacyl CoA dehydrogenase (HOAD) as well as pyruvate kinase (PK) and lactate dehydrogenase (L-LDH). The expression of Hsp70 and hsp90 was activated when the fish were exposed to temperatures beyond 20°C. Increased phosphorylation of p38 MAPK and JNKs indicated the parallel activation of MAPK signaling cascades and the potential involvement of MAPKs in the induction of Hsp genes. Exposure to extreme temperatures beyond 24°C caused an increase in the enzymatic activity of PK and LDH indicating an enhanced glycolytic potential.     

Depth-related enzymic activities in muscle,brain and heart of deep-living pelagic marine teleosts

The activities [units^-1 wet weight tissue] of lactate dehydrogenase (LDH), pyruvate kinase (PK), malate dehydrogenase (MDH) and isocitrate dehydrogenase (IDH) in white skeletal muscle, brain and heart of 24 pelagic teleost fishes were determined. In addition, for several of the foregoing species, citrate synthase (CS) activities were examined in white skeletal muscle. In muscle, the activities of all these enzymes decrease exponentially with increasing minimum depth of occurrence of the species; this decrease closely parallels the decrease in respiratory rate found previously for these same species. The decline in enzymic activity with increasing minimum depth of occurrence is much greater than the decline in body protein content of the whole fishes, suggesting a disproportionately rapid fall in enzyme concentration in comparison to contractile and structural protein concentrations. The similar reductions in activities of both glycolytic (LDH and PK) and citric acid cycle (CS, MDH and IDH) enzymes with depth indicate that both standard and active metabolisms of deeper-dwelling species are reduced relative to shallower-dwelling forms. There is no suggestion of increased anaerobic capacity with depth or in relation to species, occurrence in the oxygen minimum layer. In brain and heart, there is no significant decrease in enzymic activity with increasing minimum depth of occurrence. These two tissues have similar capacities for metabolism in most fishes, when comparisons are based on enzymic activity per gram wet weight of tissue.     

Effects of nutrition and temperature on metabolic enzyme activities in larval and juvenile red drum,Sciaenops ocellatus,and lane snapper,Lutjanus synagris

The metabolic enzyme activities were determined in larvae of red drum, Sciaenops ocellatus, and lane snapper, Lutjanus synagris, to determine the effect of temperature and nutrition on metabolic enzyme activities and to evaluate if metabolic enzyme activities are useful in assessing the feeding condition of larval fish. During experiments conducted during the spring of 1990, lactate dehydrogenase (LDH) activities in both red drum and lane snapper were approximately an order of magnitude lower than values typical for adult fish; LDH and citrate synthase (CS) activities increased during early developmental stages, but nutritional effects were apparent. Clear differences (up to 4-fold) between well-fed and starving fish were evident in both LDH and CS activity in red drum. Differences between well-fed and poorly fed larvae were evident until 9 d after hatching. Lane snapper larvae reared at a 25°C had significantly lower LDH activities than larvae reared at 28°C.     

Metabolic responses of the squat lobster (Pleuroncodes monodon) larvae to low oxygen concentration

Squat lobster populations found in the Humboldt Current System over the continental shelf from ~28 to 37°S release pelagic larvae in sub-surface cold (~11 °C) hypoxic waters. Larvae subsequently spread throughout the water column encountering both normoxic and hypoxic conditions. We analyzed some short- and long-term responses of Pleuroncodes monodon larval metabolism to hypoxia at 11 °C. Routine and postprandial aerobic respiration rates were lower in hypoxia than in normoxia for all zoeal stages. Zoea V oxyconformed, while megalopae oxyregulated down to very low oxygen concentrations. Throughout zoea I development, the rate of nitrogen (protein) accumulation in zoea I was lower, and C:N ratios were higher under hypoxic conditions than in normoxia. Citrate synthase (CS) and malate dehydrogenase (MDH) apparent specific activities (as indicators of aerobic and metabolic potentials, respectively) decreased and remained at the same level, respectively, throughout zoea I reared under hypoxic conditions. Anaerobic to aerobic potential (lactate dehydrogenase (LDH)/CS) was higher in organisms reared under hypoxia, and MDH/LDH potential ratios were characteristic of organisms tolerant to hypoxia. In spite of P. monodon zoea endurance and metabolic adaptations to decreasing oxygen tensions, intense hypoxia as such of their release site would affect their overall condition especially toward the end of the molt cycle. Our results indicate the importance of considering the interaction between environmental oxygen variability and recruitment success.     

Biochemical indices of aerobic and anaerobic capacity in muscle tissues of California elasmobranch fishes differing in typical activity level

Biochemical indices of aerobic and anaerobic metabolic capacity were measured in white myotomal muscle of eight marine elasmobranch fish species representing a broad range of locomotor performance, and in red myotomal muscle and heart of three of those species. The objectives were to determine if metabolic capacities vary with typical fish activity level, to compare the endothermic mako shark with ectothermic pelagic sharks, and to compare elasmobranchs with teleosts in order to test the hypothesis that elasmobranchs have lower aerobic capacities, metabolic rates, and swimming speeds. In white myotomal muscle, activities of the enzymes citrate synthase (an index of aerobic capacity), pyruvate kinase, and lactate dehydrogenase (LDH, an index of anaerobic capacity) covaried with typical activity level, and the ability to tolerate intracellular acidification (nonbicarbonate buffering capacity) corresponded with LDH activity. Enzyme activities in red muscle and heart did not show a consistent pattern with respect to fish activity. In comparison with ectothermic sharks, the mako shark had greater aerobic and anaerobic capacities in white muscle, but no significant differences were found in red muscle or heart. This pattern has also been found in teleosts. Thus, endothermic fishes elevate the temperature of red muscle, a tissue specialized for high aerobic performance, whereas white muscle biochemical characteristics are adjusted to support high rates of contraction both aerobically and anaerobically. Muscle enzymic activities of elasmobranchs and teleosts with comparable locomotor habits are similar, thus refuting the hypothesis that elasmobranchs are sluggish, with lower metabolic capacities than teleosts.     

Effect of dissolved oxygen level on respiratory metabolism, nutritional physiology, and immune condition of southern king crab Lithodes santolla (Molina, 1782) (Decapoda, Lithodidae)

Episodes of hypoxia are common in the marine environment, and their ecological effects depend, in part, on their severity and duration. Many species of decapod crustaceans reside in areas with fluctuating oxygen regimens. Physiological mechanisms enhance the ability of these crustaceans to cope with acute episodes of hypoxia. Southern king crab, Lithodes santolla, fishery is important in the south of South America, and some data describe fishing zones with low dissolved oxygen (DO) levels (3.5 mgO₂ l⁻¹, i.e., 8.3 kPa). Our main objective was to evaluate the effect of dissolved oxygen level on respiratory metabolism, nutritional physiology, and immunological condition of L. santolla juveniles. Individual animals were exposed for 10 days to different oxygen tensions (2.1, 4.2, 8.5, 12.7, and 21.1 kPa) to quantify the oxygen consumption rate; thereafter, blood oxyhemocyanin (Hc), protein concentration, as well as hemocytes, were sampled. Freeze-dried animals were dissected, and digestive gland metabolites (glycogen, protein, glucose, cholesterol, acylglycerol, and lactate) and digestive enzyme activity (general protease, trypsin, and chymotrypsin), as well as gill lactate dehydrogenase (LDH) activity, were quantified. In the present study, Lithodes santolla showed a critical oxygen tension between 4 and 9 kPa, indicating that this crab species is more sensitive to DO than other crustacean species. Protein and Hc concentrations followed a similar pattern to that of oxygen consumption. Digestive gland glycogen and protein concentration did not change after 10 days at different oxygen exposures, but glucose, cholesterol, and acylglycerol concentrations decreased linearly and proportionally to the available oxygen in the water. As in other decapods, chymotrypsin showed over 90% of the total quantified proteases activity. Chymotrypsin activity together with total proteases and trypsin was not affected by the environmental oxygen tension. Gill LDH and digestive gland lactate followed a similar increase at lower environmental oxygen tension but dropped sharply at the lowest tension (2.1 kPa). Dissolved oxygen affected also the immune system through reduction of hemocytes. This could provide a critical window for opportunistic pathogens to become established when crabs are exposed to hypoxic conditions. L. santolla juveniles show a moderate tolerance to low oxygen availability by modifying the concentration of hemolymph proteins, mainly OxyHc, some digestive gland metabolites, and by activating the anaerobic metabolism. This allows L. santolla juveniles to inhabit temporarily low oxygen zones in the deep ocean and suggests an advantage for culture conditions.     

Effects of long-term acclimation to environmental hypercapnia on extracellular acid–base status and metabolic capacity in Mediterranean fish <Emphasis Type="Italic">Sparus aurata</Emphasis>

In the context of future scenarios of anthropogenic CO₂ accumulation in marine surface waters, the present study addresses the effects of long-term hypercapnia on a Mediterranean fish, Sparus aurata. By equilibration with elevated CO₂ levels seawater pH was lowered to a value of 7.3, close to the maximum pH drop expected in marine surface waters from atmospheric CO₂ accumulation. Intra- and extracellular acid–base parameters as well as changes in enzyme profiles were studied in red and white muscles and the heart under both normocapnia and hypercapnia. The activities of pyruvate kinase (PK), lactate dehydrogenase (L-LDH), citrate synthase (CS), malate dehydrogenase and and 3-hydroxyacyl CoA dehydrogenase (HOAD) reflect the pathways and capacity of oxidative processes in metabolism. Long-term hypercapnia caused a transient reduction in blood plasma pH (pH_e) as well as in intracellular pH (pH_i). Compensation of the acidosis occurred through increased plasma and cellular bicarbonate levels. Changes in enzymatic activities, especially the increase in the activity of L-LDH, paralleled by a drop in CS activity in white and red muscles reflect a shift from aerobic to anaerobic pathways of substrate oxidation during long-term acclimation under hypercapnia. The present results suggest that moderate environmental hypercapnia changes the metabolic profile in tissues of S. aurata. Consequences for slow processes like growth and reproduction potential as well as potential harm at population, species and ecosystem levels require further investigation.     

Morpho-histological and enzymatic alterations in earthworms Drawida willsi and Lampito mauritii exposed to urea,phosphogypsum and paper mill sludge

The effects of varying concentrations of urea, phosphogypsum and paper mill sludge (PMS) on the morphology, histology, tissue protein content, lipid peroxidation (LPX), activities of lactate dehydrogenase (LDH), acetylcholinesterase (AChE) and catalase in earthworms Drawida willsi and Lampito mauritii have been studied over an exposure period of 24?hr. Integumentary lesions, clitellar swelling and loss of pigmentations were found to be major morpho-pathological changes in the worms. Histology indicated cuticular damage, ruptured epithelium and muscle fibres with accumulation of cellular debris. Lowest tissue protein content (57.02?±?4.02?mg/g tissue) and highest LPX (0.113?±?0.04, 0.137?±?0.08?nmol/mg protein) were noticed in D. willsi at a high concentration of PMS, whereas highest tissue protein content was observed in L. mauritii (115.32?±?7.18?mg/g tissue) with the same treatment. In both the species, LDH activity was minimum at a high concentration of urea (0.172?±?0.02; 0.247?±?0.08?U/mg protein). AChE activity was highest (0.099?±?0.002?U/mg protein) at a high concentration of PMS in D. willsi, whereas catalase activity was the maximum (0.338?±?0.02?U/mg protein) at high concentrations of PMS in L. mauritii. The study indicated that morpho-histological and enzymatic alterations in these earthworms exposed to agrochemicals could be useful biomarkers to evaluate soil toxicity.     

Biochemical adaptations of the mudskipper Boleophthalmus boddaerti to a lack of oxygen

Specimens of the mudskipper Boleophthalmus boddaerti, collected along the estuarine canal at Pasir Ris, Singapore between April 1987 and December 1989, were examined in the laboratory. After being exposed to environmental hypoxia for 6 h, no accumulation of lactate, alanine or succinate was observed in the muscle tissue of B. boddaerti. In addition, the blood lactate content and muscle creatine phosphate (CrP), adenosine triphosphate (ATP) and glycogen contents were not significantly different from those of the control. During normoxic recovery, oxygen debt repayment was only a small fraction of the oxygen deficit incurred during the 6 h of hypoxic exposure. Therefore, it would appear that B. boddaerti coped with environmental hypoxia by undergoing metabolic rate reduction. After anoxic exposure, the CrP, ATP and adenosine diphosphate (ADP) contents in the muscle tissue of B. boddaerti were significantly lower than the corresponding control values. Lactate accumulated in the blood and muscle tissue of the anoxic fish though the muscle energy charge and glycogen content remained constant. Hence, B. boddaerti was able to cope with such a period of anoxia without increasing its glycolytic rate. Succinate, which was not detected in the muscle tissue of the control and hypoxia-exposed fish, accumulated in significant quantities in the muscle tissue of B. boddaerti exposed to environmental anoxia and functional hypoxia. In the case of functional hypoxia, there was a significant decrease in the muscle glycogen content, and the muscle lactate content increased 17.3-fold, indicating that glycolysis was activated to provide the source of energy during muscular exercise.     

Superoxide dismutase in the marine sponge <Emphasis Type="Italic">Cliona celata</Emphasis>

The aim of this work is to investigate the activity of the antioxidant enzyme superoxide dismutase in the cosmopolitan sponge Cliona celata (Grant, 1826), since this enzyme has been described as a useful biomarker for marine pollution in other marine invertebrates. The quantification of the catalytic activity for superoxide dismutase is quite complex because its substrate is an unstable free radical. Several methods have been developed for this enzymatic activity determination; most of them are based on inhibition of a redox reaction involving the superoxide radical. In this work, two methods are compared, for crude sponge extracts, as far as repeatability, reproducibility and sensibility are concerned. The adrenaline oxidation method seems to be the most adequate for these determinations. Statistical treatment of the data indicates that the reference value for the specific superoxide dismutase activity in C. celata should be in the interval [0–535.5] U/mg of total protein in wet tissues, for normal populations.     

Comparative effects of five chelating agents on testicular toxicity in mice induced by acute exposure to cadmium

The aim of this study was to develop new antidotes for cadmium (Cd) since this metal is known to produce mammalian toxicity. N-p-hydroxymethylbenzyl-D-glucamine dithiocarbamate (HBGD), N-benzyl-D-glucamine dithiocarbamate (BGD), diethyldithiocarbamate (DDTC), 2,3-dimercaptopropanol (BAL), and ethylenediamine-tetraacetic acid (EDTA) were studied for their ability to inhibit the adverse effects induced by Cd on mouse testes. The parameters examined included concentrations of Cd, calcium (Ca), iron (Fe), and zinc (Zn) in testes, lipid peroxidation (LPO) levels in testes, lactate dehydrogenase (LDH) activity in serum and reproductive ability of male mice. Mice injected intraperitoneally (ip) with CdCl₂ (2.5?mgCd?kg^?1) after 30?min or 24?h, were then injected ip with chelating agents (400?µmol?kg^?1). Cd increased the concentrations of testicular Ca, Cd, Fe, Zn, and LPO levels as well as the activity of LDH in serum. HBGD and BGD effectively prevented the increase in above indices, and improved the reproductive ability weakened by exposure to Cd. The results suggested that HBGD and BGD are more effective detoxificants in the case of testicular toxicity in mice induced by acute exposure to Cd.     

Esterase heterogeneity and an investigation into racial differences in the cirripede Balanus balanoides using acrylamide gel electrophoresis

Four groups of bands showing esterase activity were found after electrophoretic separation of extracts of Balanus balanoides (L.) prosoma on polyacrylamide gels. Each group was distinguishable from other groups by mobility, reaction with specific substrates, and susceptibility to various inhibitors. Groups designated BbEII and BbEIII showed cholinesterase activity, BbEIV showed carboxylesterase activity, and BbEI possible arylesterase activity. Polymorphisms were found within Groups BbEIII and BbEI. The frequency distribution of BbEIV polymorphs in a sample from the Menai Straits, UK, fitted the Hardy-Weinberg predicted values for 4 co-dominant alleles. A sample from St. Andrews, New Brunswick, Canada, however, did not fit the Hardy-Weinberg prediction. No variation between individuals was found when malate dehydrogenase (MDH) and lactate dehydrogenase (LDH) isozymes were examined. Hence, these systems are unsuitable for testing racial differences. The results for the cholinesterase and arylesterase isozymes substantiate the view, originally based on differences in reproductive phenology and egg size, that genetically separate races exist on either side of the Atlantic Ocean.