Oxidative stress and antioxidant defense responses by goldfish tissues to acute change of temperature from 3 to 23 °C

The effects of a rapid transfer from a low (3 °C) to a warm (23 °C) temperature on oxidative stress markers and antioxidant defenses were studied in the brain, liver and kidney of the goldfish, Carassius auratus. Cold-acclimated fish were acutely moved to 23 °C and sampled after 1, 6, 12, 24, 48 or 120 h of warm temperature exposure. Lipid peroxide levels increased quickly during the first few hours at 23 °C, but thiobarbituric acid-reactive substances changed little. Protein carbonyl content was reduced by 20-40% in the liver over the entire experimental course, but increased transiently in the kidney. The content of high-molecular mass thiols decreased by two-thirds in the brain and was affected slightly i

Effect of hypoxia on the activity and binding of glycolytic and associated enzymes in sea scorpion tissues

The effect of hypoxia on the levels of glycogen, glucose and lactate as well as the activities and binding of glycolytic and associated enzymes to subcellular structures was studied in brain, liver and white muscle of the teleost fish, Scorpaena porcus. Hypoxia exposure decreased glucose levels in liver from 2.53 to 1.70 µmol/g wet weight and in muscle led to its increase from 3.64 to 25.1 µmol/g wet weight. Maximal activities of several enzymes in brain were increased by hypoxia: hexokinase by 23%, phosphoglucoisomerase by 47% and phosphofructokinase (PFK) by 56%. However, activities of other enzymes in brain as well as enzymes in liver and white muscle were largely unchanged or decreased during experimental hypoxia. Glycolytic enzymes in all three tissues were partitioned between soluble and particulate-bound forms. In several cases, the percentage of bound enzymes was reduced during hypoxia; bound aldolase in brain was reduced from 36.4 to 30.3% whereas glucose-6-phosphate dehydrogenase fell from 55.7 to 28.7% bound. In muscle PFK was reduced from 57.4 to 41.7% bound. Oppositely, the proportion of bound aldolase and triosephosphate isomerase increased in hypoxic muscle. Phosphoglucomutase did not appear to occur in a bound form in liver and bound phosphoglucomutase disappeared in muscle during hypoxia exposure. Anoxia exposure also led to the disappearance of bound fructose-1,6-bisphosphatase in liver, whereas a bound fraction of this enzyme appeared in white muscle of anoxic animals. The possible function of reversible binding of glycolytic enzymes to subcellular structures as a regulatory mechanism of carbohydrate metabolism is discussed

Influence of exercise on the distribution of enzymes in trout white muscle and kinetic properties of AMP-deaminase from free and bound fractions

Effects of exercise on the distribution of phosphofructokinase (PFK), fructose-1,6-biphosphatase (FBPase), and AMP-deaminase between free and particulate-bound fractions was analyzed in white skeletal muscle of rainbow trout Oncorhynchus mykiss. With a widely used technique for the separation of free and bound enzyme fractions (homogenization in low ionic strength, high sucrose buffer), the data showed that the amount of bound PFK increased from 64 to 95% during burst swimming whereas other enzymes were unaffected. Since this data for AMP-deaminase contrasted with earlier reports, different methods of separating free and bound enzyme were evaluated. A clear effect of exercise on AMP-deaminase binding occurred when high ionic strength media (either KCl or KF) were used; in extraction media containing 150 mM KCl, the percent bound rose from 30% in controls to 97% after 1 min burst swimming. Exercise also produced stable changes to AMP-deaminase kinetic properties, including for the bound enzyme (compared with the free) a 2-fold higher Km AMP, a 3-fold higher Ki for inorganic phosphate, and a 60% increase in Ka ADP after 1 min burst exercise. The data suggest that AMP-deaminase in working skeletal muscle is subject to combined controls by allosteric effectors, post-translational modification, and distribution between free and bound states

Effect of exercise on the properties of AMP-deaminase from trout white muscle

AMP-deaminase was purified to homogeneity from white skeletal muscle of control (resting) and exercised (1 min burst swimming) rainbow trout, Oncorhynchus mykiss. The enzyme showed a subunit molecular weight of 71,600 ± 550 kD, a Km AMP of 1.6-1.8 mM at pH 7, and was affected by allosteric inhibitors (GTP, IMP) amd activators (ADP, ATP). AMP-deaminase was inhibited by MgSO4 but activated by low concentrations of NaCl and KCl (100-150 mM); higher KCl was inhibitory. Exercise resulted in a stable modification of some properties (possibly via reversible phosphorylation); I50 values for IMP decreased by 65% and activation energies (from Arrhenius plots) changed significantly. Other properties were affected by assay pH: Km AMP decreased by 50% and Ka, ADP decreased by 70% when pH was lowered from pH 7.3 (typical of resting muscle) to pH 6.6 (muscle pH after exhaustive exercise). The data suggest that a stable modification of AMP-deaminase during exercise, coupled with effects of reduced cytosolic pH, could enhance enzyme function in the rapid conversion of AMP to IMP in working fish muscle

Influence of exercise on the activity and the distribution between free and bound forms of glycolytic and associated enzymes in tissues of horse mackerel

The effects of short-term burst (5 min at 1.8 m/s) swimming and long-term cruiser (60 min at 1.2 m/s) swimming on maximal enzyme activities and enzyme distribution between free and bound states were assessed for nine glycolytic and associated enzymes in tissues of horse mackerel, Trachurus mediterraneus ponticus. The effects of exercise were greatest in white muscle. The activities of phosphofructokinase (PFK), pyruvate kinase (PK), fructose-1,6-bisphosphatase (FBPase), and phosphoglucomutase (PGM) all decreased to 47, 37, 37 and 67%, respectively, during 60-min exercise and all enzymes except phosphoglucoisomerase (PGI) and PGM showed a change in the extent of binding to subcellular particulate fractions during exercise. In red muscle, exercise affected the activities of PGI, FBPase, PFK, and lactate dehydrogenase (LDH) and altered percent binding of only PK and LDH. In liver, exercise increased the PK activity 2.3-fold and reduced PGI 1.7-fold only after 5 min of exercise but altered the percent binding of seven enzymes. Fewer effects were seen in brain, with changes in the activities of aldolase and PGM and in percent binding of hexokinase, PFK and PK. Changes in enzyme activities and in binding interactions with subcellular particulate matter appear to support the altered demands of tissue energy metabolism during exercise

Activity and properties of phosphofructokinase in white muscle of horse mackerel Trachurus mediterraneus under swimming stimulation

The activity and properties of phosphofructokinase (PFK) in tissues of horse mackerel which was swimming at burst regimen for 5 min and at cruiser one for 60 min have been investigated. In white muscle the PFK activity increased 1,6-fold after burst swimming and Hill's coefficient rose as well and decreased 3-fold after cruiser one. Swimming did not change the half-maximal saturation constant for both substrates and inhibition constants for ATP and citrate. In the preparations from white muscle of fish which were stimulated by burst swimming the PFK activity at physiological pH values (6,0-7,0) was higher comparing with one from the control group and after cruiser swimming. Incubation of preparations at 45 °C decreased the activity of PFK in control and cruiser swimming groups (to 61-67% of initial level) and increased it after burst swimming (1,3-fold). The mechanisms involving in stable modification of PFK under different swimming regimens are discussed

Induction of oxidative stress in Rana ridibunda during recovery from winter hibernation

The effect of transfer from winter hibernation at 5°C to a warm temperature (20°C for 1 or 24 h) on oxidative stress and antioxidant defenses was assessed in the frog, Rana ridibunda. The temperature increase had little effect on the levels of thiobarbituric acid reactive substances but carbonylprotein levels (581, 740, 1270 and 614 pmol/mg protein in brain, liver, kidney and muscle, respectively, at 5°C) rose 3.2-fold in brain and 2.6-fold in liver after transfer to 20°C. The switch to 20°C also affected the activities of antioxidant enzymes: catalase in liver, kidney and muscle rose 33%, 55% and 126%, respectively, whereas superoxide dismutase increased 2.4-fold in liver and 2.5-fold in muscle. Glutathione reductase and glucose-6-phosphate dehydrogenase activities were also affected in some organs. The data show that recovery from winter hibernation in frogs causes increased oxidative stress and stimulates an elevation of organ antioxidant defenses

The effect of potassium dichromate on free radical processes in goldfish: Possible protective role of glutathione

The effects of 96 h exposure to Cr6+ (added as potassium dichromate) on the status of antioxidant defenses and markers of oxidative damage were evaluated in three tissues of goldfish, Carassius auratus. Fish exposure to high dichromate concentrations, 10 and 50 mg/l, increased protein carbonyl levels in brain and liver, but not in kidney. Chromium exposure also increased concentrations of lipid peroxides in brain (at 5 mg/l) and liver (10 mg/l), but not in kidney. The concentrations of reduced glutathione (GSH) were higher in the liver of goldfish treated with 5-50 mg/l Cr6+ than in controls, but in kidney only the 5 mg/l-treated group showed increased GSH levels. Dichromate at 1 mg/l increased the concentration of oxidized glutathione (GSSG) in liver and kidney by 80% and 60%, respectively, whereas at 10 and 50 mg/l the levels of GSSG decreased by 50% in kidney. These results indicate that the dichromate concentrations used induced oxidation of lipids and proteins in goldfish tissues in a concentration- and tissue-specific manner. Also, the redox status of fish tissues was affected in a concentration- and tissue-specific manner. The activities of glutathione reductase increased in all three tissues in response to dichromate treatment, increasing by ∼2-fold in brain and liver in goldfish treated with 50 mg/l Cr6+. Dichromate treatment did not change the activities of SOD, catalase or GST in brain, but reduced the activities of SOD in liver and kidney, and catalase in liver. The results suggest that the glutathione system may be responsible for protecting against the deleterious effects of dichromate in fish and indicate the possible development of an adaptive response during the 96 h treatment with the toxicant