Temperature acclimation in respiratory and cytochrome c oxidase activity in common carp (Cyprinus carpio)

• 1.1. Common carp (Cyprinus carpio) exposed to experimental temperatures of 12, 18, 24, 30 or 36°C for a 4-week period were used to investigate the effect of temperature acclimation on the frequency of opercular movement (FOM), growth and cytochrome c oxidase (CCO) activity in heart, liver and muscle.
• 2.2. An exponential relationship between FOM and temperature after the first week (10₁₀ =1.76) disappeared after the second week.
• 3.3. The initially high FOM at temperatures of 30 or 36°C and the low FOM at 18 or 12°C changed over 4 weeks to approach the FOM of fish at 24°C.
• 4.4. This change in the relationship of FOM to temperature from highly dependent to independent appeared to be thermal compensation.
• 5.5. Heart and liver CCO activities were significantly affected by temperature, with the lowest activity at the approximate optimum temperature for growth, 24°C.
• 6.6. Highest CCO activities for heart and liver occurred at both the highest and lowest temperatures.
• 7.7. Among the three tissues, heart CCO activity was generally the highest and most affected by acclimation temperature.
• 8.8. Muscle tissue had the lowest CCO activity and was unaffected by temperature.
• 9.9. The high CCO activity at a cold acclimation of temperature 12°C was probably due to thermal compensation and the high activity at 36°C may have been a result of thermal stress.

     

Effect of methyl methacrylate on mitochondrial function and structure

• 1.1. Treatment of isolated rat liver mitochondria with methyl methacrylate (MM) produced membrane disruption as evidenced by the release of citrate synthase, and changes in the ultrastructure of mitochondria.
• 2.2. At concentration 0.1%, MM uncoupled oxidative phosphorylation as evidenced by stimulation of state 4 respiration supported either by pyruvate plus malate or succinate (+rotenone) and ATP-ase activity in intact mitochondria.
• 3.3. At concentration 1% MM stimulated ATP-ase activity in intact mitochondria and succinate (+rotenone) oxidation at state 4 and was without effect on this substrate oxidation at state 3.
• 4.4. MM inhibited pyruvate plus malate oxidation either at state 3 or in the presence of uncoupling agents.
• 5.5. MM inhibited the NADH oxidase of electron transport particles at a concentration which failed to inhibit either succinic oxidase or the NADH-ferricyanide reductase activity.
• 6.6. The data presented suggest that in the isolated mitochondria MM inhibits NADH oxidation in the vicinity of the rotenone sensitive site of complex I.
• 7.7. The general conclusion is that MM may block an electron transport and to uncouple oxidative phosphorylation in rat liver mitochondria. The overall in vitro effect would be to prevent ATP synthesis which could result in cell death under in vivo conditions.

     

Gender-related differences of mouse liver d-aspartate oxidase in the activity and response to administration of d-aspartate and peroxisome proliferators

• 1.1. The hepatic d-aspartate oxidase activity was found to be higher in female ddY and ICR mice than in their male counterparts. On the contrary, the free d-aspartate content in the liver was lower in female mice than in male mice, suggesting that d-aspartate is actually metabolized by d-aspartate oxidase in vivo.
• 2.2. Oral administration of d-aspartate to the animals increased the hepatic d-aspartate oxidase activity 2–3 fold in both genders without any significant difference in the rate of the increase between the genders.
• 3.3. Several peroxisomal enzyme activities other than d-aspartate oxidase examined were not affected by this treatment.
• 4.4. Experiments in vitro suggested that the increase in the d-aspartate activity might be explained in part by stabilization of the enzyme by d-aspartate.
• 5.5. The administration of clofibrate, a peroxisome proliferator, to male mice, increased the hepatic d-aspartate oxidase activity with a significant simultaneous decrease of d-aspartate content in the liver, in agreement with a possible role of the enzyme n vivo.
• 6.6. On the other hand, the administration of clofibrate or dehydroepiandrosterone to female mice decreased the d-aspartate oxidase activity.
• 7.7. The peroxisome proliferators were suggested to act to eliminate the gender difference of hepatic d-aspartate oxidase activity in mice.

     

Renal response to hypoxia in carp,Cyprinus carpio: Changes in glomerular filtration rate,urine and blood properties and plasma catecholamines of carp exposed to hypoxic conditions

• 1.1. Changes in glomerular nitration rate (GFR), urine and blood properties and plasma catecholamines of carp were investigated during and following hypoxia.
• 2.2. GFR and urine flow decreased with increased urinary concentrations of bio-components, except protein, in the course of hypoxia.
• 3.3. Decreases in blood pH, and increases in haematocrit value and plasma K⁺, Ca²⁺, Mg²⁺, inorganic phosphate (P_i), ammonia, lactic acid and catecholamines (CAs) were observed as hypoxia progressed.
• 4.4. Increased GFR and urine flow, and higher values for urinary components, except protein, compared with those of the control were found in the initial post-stress stage.
• 5.5. The possible significance of increased plasma CAs in relation to changes in renal function in hypoxic carp is discussed.

     

Purification and some properties of isocitrate dehydrogenase of a blowfly Aldrichina grahami

• 1.1. NADH-dependent isocitrate dehydrogenase has been purified 110-fold from the crude extract of the flight muscle mitochondria of Aldrichina grahami.
• 2.2. The purification procedure involved Triton X-100 treatment of isolated mitochondria, column chromatography on DEAE-cellulose, Affi-gel blue, and P-cellulose.
• 3.3. The purified enzyme was homogeneous by criteria of the polyacrylamide gel electrophoresis.
• 4.4. The enzyme of the blowfly contains more acidic amino acids and less hydrophobic amino acids than that of pig heart.
• 5.5. The molecular weight was determined to be 330,000 daltons. The subunit construction differs from ghat of mammalian isocitrate dehydrogenase.

     

Troponin C of the Antarctic icefish (Champsocephalus gunnari) white muscle

• 1.1. The troponin C (TN-C) from the Antarctic icefish (_{Champsocephalus gunnari}) has been isolated to homogeneity by a procedure involving extraction from acetone powder, DEAE-Sepharose 4B and AcA 54 column chromatography.
• 2.2. The calcium-induced conformational changes of apo TN-C have been studied by absorption difference spectroscopy, circular dichroism and intrinsic fluorescence.
• 3.3. The results indicate that the overall characteristics of icefish TN-C (such as amino acid composition, modifications of helix content and of the microenvironment of aromatic residues as a function of calcium binding) are quite similar to those of rabbit TN-C.
• 4.4. The intrinsic fluorescence properties are close to those reported for pike and carp TN-C.

     

Evidence for target site insensitivity to cyanide in Spodoptera eridania larvae

• 1.1. Isolated mitochondria from the whole southern armyworm larvae, Spodoptera eridania, show all of the characteristics of mammalian liver mitochondria, except for target site sensitivity to cyanide.
• 2.2. The armyworm larval mitochondria are 17 times less sensitive to cyanide when compared to rat liver mitochondria and cannot be completely inhibited with extremely large doses.
• 3.3. These data suggest the presence in the southern armyworm of either a cyanide-insensitive cytochrome oxidase, or the elaboration of cyanide-insensitive oxidative pathway reminiscent of an alternative oxidative pathway that is known to coexist in plants alongside the cyanide-sensitive pathway.

     

Further studies on the mechanism of action of gossypol on mitochondrial membrane

• 1.1. In order to explore the mechanism of inhibition of hydroxylases involved in steroidogenesis, by gossypol, we studied the effect of this drug on adrenal cortex mitochondria, and compared it with those on kidney and heart.
• 2.2. The uncoupler effect of gossypol (collapse of Δψ and Ca²⁺ efflux) was found to be lower in adrenal cortex mitochondria than in kidney and heart mitochondria.
• 3.3. Gossypol produced more extensive changes on the membrane lipidic matrix (increase in the order parameter for 5-doxylstearic acid) in adrenal cortex mitochondria than in the other mitochondria studied.
• 4.4. The results described above indicate that the mechanism of inhibition of gossypol of steroidogenic adrenal enzymes could be attributed to an alteration of the lipidic matrix which, in turn, modifies protein function.

     

A study of glycerolphosphate acyltransferase in rat liver mitochondria-submitochondrial localization and some properties of the solubilized enzyme

• 1.1. Glycerolphosphate acyltransferase (GPAT) was solubilized from the rat liver mitochondrial membranes using sodium cholate. Dithiothreitol was necessary to stabilize the solubilized enzyme on storage.
• 2.2. Unlike the enzyme in situ in mitochondrial membranes, the solubilized mitochondrial GPAT was susceptible to inhibition by N-ethylmaleimide; a property more characteristic of the distinct microsomal form of GPAT.
• 3.3. Solubilized mitochondrial GPAT retained its very high preference for saturated acyl-CoA substrate (palmitoyl-CoA) and had no activity whatever with any tested concentration of the unsaturated substrate oleoyl-CoA.
• 4.4. Solubilization increased the affinity of mitochondrial GPAT for palmitoyl-CoA whilst decreasing the K_m for glycerol phosphate.
• 5.5. After separation of liver mitochondrial outer and inner membranes and estimation of cross-contamination by appropriate markers it was concluded that the mitochondrial inner membrane contains significant GPAT activity. This was established with preparations from fed, 48 hr-starved and streptozotocin-diabetic rats.

     

Prostaglandin E1 and analogs of prostacyclin influencing superoxide production by the human neutrophil nadph oxidase system

• 1.1. The effects of prostaglandin (PG) E₁, and I₂ analogs (OP-41483 and OP-2507) on the Superoxide generation of human neutrophil NADPH oxidase (EC 1.6.99.6) in both whole-cell and cell-free systems were investigated.
• 2.2. In a whole-cell system, OP-2507 inhibited the Superoxide generation by neutrophils exposed to phorbol myristate acetate concentration-dependently through its superoxide-scavenging action.
• 3.3. The concentration of the drug required for 50% inhibition of the oxidase (IC₅₀) was 21 μM.
• 4.4. In a cell-free system, however, the drug in concentrations of < 100 μM did not inhibit the activation of NADPH oxidase by sodium dodecyl sulfate because of its inactivation by the detergent.
• 5.5. Although PGE₁ and OP-41483 did not inhibit the Superoxide production by stimulated neutrophils in a whole-cell system, they both inhibited the activation of NADPH oxidase in a cell-free system concentration-dependently, with IC₅₀ values of 44 and 170 μM, respectively.
• 6.6. In addition, in the cell-free system, the K_m value for NADPH of the oxidase was unchanged by PGE₁.
• 7.7. The results suggest that the PGI₂ analog, OP-2507, is a possible superoxide-scavenger and that PGE₁ inhibits the NADPH oxidase activation by sodium dodecyl sulfate in a cell-free system concentration-dependently.

     

Deleterious effects of disulfiram on the respiratory electron transport system of liver mitochondria

• 1.1. The mechanism of action of disulfiram on the respiratory electron transport system of the liver mitochondria was studied in vitro.
• 2.2. Disulfiram inhibited the respiration supported by malate-glutamate as well as succinate.
• 3.3. Mitochondrial respiration inhibition was dependent upon alteration of —SH groups.
• 4.4. The inhibitory action of disulfiram might be related to the crosslinking of several proteins of the inner mitochondrial membrane.
• 5.5. The effects described above could be attributed to disulfiram per se and not to the main metabolite diethyldithiocarbamate.

     

Effects of membrane-perturbing drugs and noradrenalin on cAMP accumulation and red cell water content in carp (Cyprinus carpio) red cells

• 1.1. Carp red cells were treated with drugs that affect the cell membranes. The water content of the cells and the accumulation of cAMP in the cells were measured in normoxia and in hypoxia using non-stimulated and adrenergically stimulated cells.
• 2.2. WGA, DIDS + CCCP and A23187 increased the water content of nonstimulated normoxic cells.
• 3.3. In hypoxia ouabain and DIDS + CCCP increased the water content but cytochalasin B, NPM, DIDS, CCCP and A23187 + CA²⁺ abolished the hypoxia-induced swelling.
• 4.4. Any membrane perturbation induced some cAMP formation, Sophora and Anquilla lectins being most potent.
• 5.5. Also in adrenergically stimulated cells, membrane perturbation generally increased cAMP formation.
• 6.6. However, cAMP accumulation diminished in cells treated with cytochalasin B, CCCP and DIDS + CCCP.
• 7.7. The adrenergic swelling of carp red cells was reduced in normoxia by DIDS. NPM and CCCP increased the adrenergic swelling in normoxia to hypoxic level.
• 8.8. In hypoxia WGA and Anquilla lectin decreased the swelling.

     

Rapid preparation of subsarcolemmal and interfibrillar mitochondrial subpopulations from cardiac muscle

• 1.1. Subsarcolemmal and interfibrillar mitochondria were prepared with complete recovery from rabbit and porcine heart muscle by upward-flotation during 60 sec of Percollö density gradient centrifugation.
• 2.2. Mitochondrial subpopulations were identified and characterized according to buoyant density, electron-microscopy, marker enzyme activities and respiratory performance.
• 3.3. ADP-induced state 3-respiration related to latent citrate synthase activity as a marker for structurally intact mitochondria was not significantly different in both mitochondrial subtypes.

     

Proteomics Analysis of Extracellular Matrix Remodeling During Zebrafish Heart Regeneration

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Highlights

• •We have developed a decellularization protocol for ECM protein enrichment.
• •We have characterized the proteome of adult zebrafish heart ECM.
• •We describe dynamic changes in heart ECM proteome during regeneration.
• •We describe changes in heart ECM stiffness during regeneration.

     

Effect of experimental ventilation of the skin on cutaneous oxygen uptake in the carp,Cyprinus carpio

• 1.1. Cutaneous O₂ uptake in the carp, Cyprinus carpio, was determined at various water flow rates across the skin (.V) ranging from 2.5 to 40 ml/min, using flow-through respirometers.
• 2.2. When thickness of water flow was 2mm, cutaneous O₂ uptake remained stable (about 3.8 nmol/cm²/min) at a .V of 20–40 ml/min and decreased with .V below 20 ml/min.
• 3.3. When thickness of water flow was 4 mm, cutaneous O₂ uptake decreased with .V below 40 ml/min.
• 4.4. Apparent water velocity (U') was calculated dividing .V by an area of a cross section of the water flow (0.5 and 1.0 cm² respectively). In both experiments, cutaneous O₂ uptake decreased with U' below 0.7 cm/sec.
• 5.5. This suggests that cutaneous O₂ uptake in the carp is limited at a low water velocity by a resistance of the hypoxic boundary layer.

     

Heart rate and its cholinergic control in the sole (Solea vulgaris), acclimatized to different temperatures

• 1.1. The effects of thermal acclimatization at 10 and 24°C on heart rate were investigated on unrestrained soles (Solea vulgaris).
• 2.2. The sensitivity of heart rate to temperature changes induced by temperature acclimatization was higher in cold-acclimatized than in warm-acclimatized soles.
• 3.3. Heart rate of cold-acclimatized fish to temperature changes was not affected by blocking the vagal tone with atropine.
• 4.4. After atropine treatment the ability of heart rate to show thermal compensation decreased in warm-acclimatized soles.
• 5.5. It is suggested that the vagus nerve can function differently at different temperatures.

     

Structural analogies between different redox enzymes inserted into biological membranes

• 1.1. We have compared the primary structure and the predicted secondary structure of subunit I (COI) of cytochrome oxidase with those of other integral redox enzymes which contain membrane-buried iron centres.
• 2.2. Some striking analogies have been found between the deduced transmembrane folding for COI and the known three-dimensional structure of the photosynthetic reaction centre of Rhodobacter sphaeroides.
• 3.3. These structural analogies are paralleled by a fundamental functional analogy between these two redox systems, since they both oxidize reduced cytochrome c at the positive side of the membrane, transferring electrons to membrane-buried metal centres.
• 4.4. A statistical evaluation has been performed on the amino acid composition of the peptides containing known histidine ligands of the membrane-buried iron in cytochrome b of the bc 1 complex and in the bacterial reaction centre.
• 5.5. This evaluation was then applied to the peptides which contain conserved histidines in subunit I of cytochrome oxidase, subunit that is known to bind both haem a and a3, indicating which of these histidines are the most likely ligands of the membrane-buried iron of the a-haems.
• 6.6. A sequence homology has been found between the known oxygen binding site and the haem binding peptide in cyt P450 and two peptides which are conserved in all the sequences of COI, thus indicatingt the possible oxygen catalytic site of cytochrome oxidase.

     

Characterization of two distinct latent proteinases associated with myofibrils of crucian carp (Carassius auratus cuvieri)

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• 1.1. Enzymatic properties of two distinct proteinases tightly associated with crucian carp myofibrils were characterized.
• 2.2. These proteinases were latent but activated at 50 and 60°C, respectively.
• 3.3. The optimum pH of 50°C-proteinase was neutral-alkaline, while that of 60°C-proteinase was weak acid-neutral pH.
• 4.4. Both proteinases required more than 1% NaCl for the activity, but 50°C-proteinase was partially inhibited at higher concentrations of NaCl.
• 5.5. Both proteinases were regarded as trypsin-like proteinases belonging to a serine proteinase family, but only 60°C-proteinase was sensitive to urea, n-butanol and iso-propanol.

     

Possible origin of extremely high contents of vitamin D3 in some kinds of fish liver

• 1.1. Comparative studies on the possible origin of extremely high contents of vitamin D₃ in some kinds of fish liver were performed.
• 2.2. Neither photochemical formation of vitamin D₃ in fish skin by solar radiation of 7-dehydrocholesterol (7-DHC) nor nonphotochemical enzymatic formation of vitamin D₃ from 7-DHC in fish liver was demonstrated as the origin of vitamin D₃.
• 3.3. On the other hand, when bastard halibuts and carps were farmed from fingerlings to adults with feedstuff's containing vitamin D₂ or D₃, significant amounts of the vitamins were accumulated in the fish liver.
• 4.4. The contents of vitamins D₂ and D₃ in bastard halibut liver increased according to the duration of farming and dose responses of the vitamins in carp liver were observed.
• 5.5. Significant amounts of vitamins D₂ and D₃ in phytoplankton and vitamin D₃ in Zooplankton and small fish were detected.
• 6.6. Therefore, we have concluded that the most probable origin of vitamin D₃ in fish liver is a result of food chains from plankton.

     

Lipid peroxidation affects catalytic properties of rat liver mitochondrial monoamine oxidases and their sensitivity to proteolysis

• 1.1. Lipid peroxidation (LPO) in rat liver mitochondria decreased the activity of monoamine oxidase (MAO) with physiological substrates serotonin and 2-phenylethylamine (by 15–30%) and induced deamination of glucosamine, which was highly sensitive to selective MAO A inhibitor pirlindole.
• 2.2. The LPO-induced changes in catalytic properties of MAOs are accompanied by their increased susceptibility to trypsinolysis, however sensitivity to inhibition by imipramine, chlorpromazine and spermine are insignificantly changed.
• 3.3. It is suggested that these results reflect LPO-induced conformational changes of enzyme molecules in membrane rather than their membrane topography.