Effects of copper on the acute cortisol response and associated physiology in rainbow trout

The aim of this study was to determine the effects of chronic waterborne copper (Cu) exposure on the acute stress-induced cortisol response and associated physiological consequences in rainbow trout (Oncorhynchus mykiss). Trout were exposed to 30 μg Cu/L in moderately hard water (120 mg/L as CaCO(3)) for 40 days, following which time the acute cortisol response was examined with a series of stressors. At 40 days, a 65% increase in Cu was observed in the gill, but no accumulation was observed in the liver, brain or head kidney. Stressors such as air exposure or confinement did not elicit an increase in circulating cortisol levels for Cu-exposed fish, in contrast to controls. However, this inhibitory effect on the acute cortisol response appeared to have few implications on the ability of Cu-exposed fish to maintain ion and carbohydrate homeostasis. For example, plasma Na(+), Ca(2+) and glucose levels as well as hepatic glycogen levels were the same post-stress in control and Cu-exposed fish. Trout were also challenged with exposure to 50% seawater for 48 h, where Cu-exposed trout maintained plasma Na(+), glucose and hepatic glycogen levels. However, Cu-exposed fish experienced decreased plasma K(+) levels throughout the Cu exposure and stress tests. In conclusion, chronic Cu exposure resulted in the abolition of an acute cortisol response post-stress. There was no Cu accumulation in the hypothalamus-pituitary-interrenal axis (HPI axis) suggesting this was not a direct toxic effect of Cu on the cortisol regulatory pathway. However, the lack of an acute cortisol response in Cu-exposed fish did not impair the ability of the fish to maintain ion and carbohydrate homeostasis. This effect on cortisol may be a strategy to reduce costs during the chronic stress of Cu exposure, and not endocrine disruption as a result of toxic injury.     

A note on copper bioaccumulation in Mozambique tilapia,Oreochromis mossambicus (Osteichthyes: Cichlidae)

Copper (Cu) is one of the most commonly reported metal pollutants in African water bodies, but there are few studies on African freshwater fish species of copper accumulation and copper toxicity. Adult O. mossambicus were exposed to 0mg l^?1 (control) and 0.75mg l^?1 Cu for 96h and 0 (control), 0.11, 0.29 and 0.47mg l^?1 copper for 64 days. Samples of liver and gills were collected after 96h, and after 1, 32 and 64 days, respectively. There were significant differences in the mean Cu accumulation values in the liver and gills between the control and the Cu-exposed fish after the 96-h exposure. In fish exposed to 0.11 and 0.29mg l^?1 Cu for 64 days there was an increase in copper level in the tissues. In fish exposed to 0.47mg l^?1 Cu the concentration in the gill and liver tissue did not increase between Day 1 and Day 32. At this time, Cu accumulation in the liver was higher than for fish exposed to 0.11 and 0.29mg l^?1 Cu for 64 days. Exposure to approximately 0.47mg l^?1 Cu for more than 32 days induced mortality.     

Changes in tissue adenylates and water content of bluegill, Lepomis macrochirus, exposed to copper

Bluegill were exposed to copper (2–0 mg/1⁻¹ unfiltered) for 24, 48, 72 or 96 h at 24° C. This concentration approximated the 96 h LC20. Water content of liver increased 4–8% after 24 h exposure and muscle 4–6% by 48 h. These changes persisted throughout the 96 h experiments. Copper exposed fish exhibited decreases in muscle ATP, ADP, total adenylates, and energy charge by 48 h. There was a trend toward recovery of ATP with further exposure. Liver ATP of exposed fish was lower than controls at all intervals with the greatest difference evident at 48 h. No significant changes in brain adenylates were observed. Muscle and liver lactic acid was unchanged at 48 h exposure, therefore tissue hypoxia was not the cause of the adenylate changes. It was concluded that copper causes decreases in muscle and liver ATP several days before probable death of copper exposed fish and the changes seen are the result of dilution by increased tissue water, and the copper acting on certain cellular enzymes involved in detoxification and energy metabolism.     

Shifts in a fish's resource holding power during a contact paired interaction: the influence of a copper-contaminated diet in rainbow trout

The influence of sublethal chronic dietary copper (Cu) exposure on the dominant-subordinate relationship between pairs of juvenile rainbow trout (Oncorhynchus mykiss) was examined. Fish were fed either a normal (11 mg Cu kg(-1) food) or Cu-contaminated (721 mg Cu kg(-1) food) diet for 8 wk. Paired interactions were observed--control versus control, Cu-exposed versus Cu-exposed, and control versus Cu-exposed fish--using a computer-aided video tracking system to measure duration of interactions, total distance moved, and the number of encounters during each contest. In concurrence with game theory, each interaction became escalated with a lesser size disparity between contestants. However, in Cu-exposed versus Cu-exposed fish interactions, the dominant-subordinate relationship was decided sooner and with less aggression than a control versus control fish interaction with fish of a similar relative body mass disparity. During control versus Cu-exposed fish interactions, control fish would normally dominate interactions (12 out of 16 bouts) unless the Cu-exposed fish had a 15% body mass advantage. Muscle glycogen and lactate levels after each contest reflected the duration of bouts and winners of the contests, irrespective of Cu exposure. We conclude that Cu-contaminated fish are less able to compete and have lower resource holding power than controls and will withdraw from a contest at a lower level of interaction, unless a size advantage in the Cu-exposed fish increases the probability of winning.     

The effect of sub-lethal exposure to copper and the time course of recovery in clean water on biochemical changes in juvenile fish (Acipenser persicus)

This study reports the 96-h LC₅₀ value and tissue copper (Cu) levels and biochemical changes in juvenile fish (Acipenser persicus) exposed to 0.026?mg/l ambient Cu for 1, 7 and 14?days. It then examined the recovery of the same parameters after placing the juvenile fish in clean water for a further period of 28?days. The intestine, kidney and gill Cu levels, plasma glucose, total protein, triglyceride, cortisol, triiodothyronine and thyroxine concentrations, liver protein contents, liver catalase, superoxide dismutase (SOD) and glutathione S-transferase activities were studied. The 96-h LC₅₀ value of Cu was 0.502?mg/l for juvenile A. persicus. The results indicate that Cu exposure produced significant accumulations of Cu in gills and kidney over the treatment time. Sublethal dose of Cu resulted in a short-term increase in plasma glucose, total protein and cortisol levels that decreased with time. After the 28-day recovery phase, there were significant differences in kidney Cu levels and triglyceride concentrations as well as SOD activities between recovery fish treatments and their control groups on day 42. The 28-day recovery phase caused significant decreases in total protein levels and SOD activities of Cu-exposed fish on day 42 compared to day 14. The results suggest that 28?days are insufficient for complete recovery to Cu exposure by juveniles and a longer period would be required for full recovery. Moreover, the study showed that the recovery phase following Cu exposure could change biochemical parameters to levels that are not close to those seen during exposure or control levels.     

Effect of Hypoxia on Physiological-Biochemical Blood Parameters in Some Marine Fish

Changes of hematocrit, hemoglobin, and blood plasma glucose and Na⁺ and K⁺ content in response to hypoxia were studied in three Black Sea fish species. It was shown that in response to hypoxia in low-mobile rock perch Scorpaena porcus L., hematocrit and the blood plasma glucose level increased, while the content of K⁺ in erythrocytes decreased and the content of Na⁺ increased. In moderately mobile sea carp Diplodus annularis L. autogenic hypoxia caused a rise of hematocrit and blood plasma glucose. In actively swimming jack mackerel Trachurus mediterraneus ponticus Aleev, only considerable increase of Na⁺ content was revealed in hypoxia. The obtained results indicate that fish with different mobility under hypoxic conditions use different adaptation mechanisms. The value and direction of changes of the chosen parameters can be used to determine resistance of fish to oxygen deficit.     

Influence of euthanasia method on blood and gill variables in normoxic and hypoxic Gulf killifish Fundulus grandis

In many experiments, euthanasia, or humane killing, of animals is necessary. Some methods of euthanasia cause death through cessation of respiratory or cardiovascular systems, causing oxygen levels of blood and tissues to drop. For experiments where the goal is to measure the effects of environmental low oxygen (hypoxia), the choice of euthanasia technique, therefore, may confound the results. This study examined the effects of four euthanasia methods commonly used in fish biology (overdose of MS‐222, overdose of clove oil, rapid cooling and blunt trauma to the head) on variables known to be altered during hypoxia (haematocrit, plasma cortisol, blood lactate and blood glucose) or reflecting gill damage (trypan blue exclusion) and energetic status (ATP, ADP and ATP:ADP) in Gulf killifish Fundulus grandis after 24 h exposure to well‐aerated conditions (normoxia, 7·93 mg O₂ l^?1, c. 150 mm Hg or c. 20 kPa) or reduced oxygen levels (0·86 mg O₂ l^?1, c. 17 mm Hg or c. 2·2 kPa). Regardless of oxygen treatment, fish euthanized by an overdose of MS‐222 had higher haematocrit and lower gill ATP:ADP than fish euthanized by other methods. The effects of 24 h hypoxic exposure on these and other variables, however, were equivalent among methods of euthanasia (i.e. there were no significant interactions between euthanasia method and oxygen treatment). The choice of an appropriate euthanasia method, therefore, will depend upon the magnitude of the treatment effects (e.g. hypoxia) relative to potential artefacts caused by euthanasia on the variables of interest.     

The influence of environmental temperature and oxygen concentration on the recovery of largemouth bass from exercise: implications for live–release angling tournaments

The impact of variation in water temperature and dissolved oxygen on recovery of largemouth bass Micropterus salmoides from exercise was examined. For this, largemouth bass were first exercised and recovered for either 1, 2 or 4 h at ambient water temperatures (25° C) in fully oxygenated water. Results showed that exercise forced fish to utilize anaerobic metabolism to meet energy demands, and resulted in reductions in anaerobic energy stores adenosine triphosphate (ATP), Phosphocreatine (PCr) and glycogen. Exercise also resulted in a seven‐fold increase in lactate within white muscle. After 2 h of recovery in oxygenated water at acclimation temperature, physiological recovery from exercise was under way, and by 4 h most variables examined had returned to control levels. Next, largemouth bass were exercised at ambient temperatures and recovered for 2 h in environments with either elevated temperature (32° C), reduced temperature (14 and 20° C), hypoxia or hyperoxia. Both elevated and reduced temperature impaired recovery of tissue lactate and tissue ATP relative to fish recovered in water at acclimation temperature, while hyperoxic water impaired recovery of tissue ATP. Moderately hypoxic waters impaired the recovery of plasma glucose, plasma lactate and tissue PCr relative to fish recovered in fully oxygenated water. Results from this study are discussed in the context of critical oxygen and temperature guidelines for largemouth bass. In addition, several recommendations are made concerning remedial treatments used in livewells (tanks) during angling tournaments when fish are recovering from exercise associated with angling.     

1H-NMR study of the metabolome of a moderately hypoxia-tolerant fish, the common carp (Cyprinus carpio)

All 20.000 different fish species vary greatly in their ability to tolerate and survive fluctuating oxygen concentrations in the water. Especially fish of the genus Carassius, e.g. the crucian carp and the goldfish, exhibit a remarkable tolerance to limited/absent oxygen concentrations. The metabolic changes of anoxia-tolerant crucian carp were recently studied and published. Contrary to crucian carp, the hypoxia-tolerant common carp cannot survive a complete lack of oxygen (anoxia). Therefore, we studied the ¹H-NMR-based metabolomics of brain, heart, liver and white muscle extracts of common carp, subjected to anoxia (0 mg O₂ l^?1) and hypoxia (0.9 mg O₂ l^?1) at 5 °C. Specifically, fish were exposed to normoxia (i.e. 9 mg O₂ l^?1; controls 24 h, 1 week and 2 weeks), acute hypoxia (24 h), chronic hypoxia (1 week) and chronic hypoxia (1 week) with normoxic reoxygenation (1 week). Additionally, we also investigated the metabolic responses of fish to anoxia for 2 h. Both anoxia and hypoxia significantly changed the tissue levels of standard energy metabolites as lactate, glycogen, ATP/ADP and phosphocreatine. Remarkably, anoxia induced increased lactate levels in all tissues except for the heart whereas hypoxia resulted in decreased lactate concentrations in all tissues except for brains. Furthermore, hypoxia and anoxia influenced amino acids (alanine, valine/(iso)leucine) and neurotransmitters levels (GABA, glutamate). Lastly, we also detected ‘other’ i.e. previously not reported compounds to play a role in the present context. Scyllo-inositol levels changed significantly in heart, liver and muscle, providing novel insights into the anoxia/hypoxic responses of the common carp.     

An in situ study of rock bass (Ambloplites rupestris) physiology: effect of season and mercury contamination

Selected physiological and biochemical variables were examined in rock bass, Ambloplites rupestris, which were collected on five different sampling dates from an area of chronic mercury contamination and a reference site on the South River, Virginia.The onset of spawning represented the most significant seasonal influence in the physiological profile of the fish, with elevations in hematocrit, hemoglobin, plasma protein, and plasma glucose. Sex-related differences in plasma calcium, liver glycogen and liver ascorbic acid were also unique to the period. Female rock bass had significantly higher levels of liver glutathione than did males on all but one of the sampling dates, although the cause of this difference is not clear.Rock bass from the mercury contaminated site had an average muscle mercury concentration of 1.37 mg Hg g^–1, and an average liver mercury concentration of 2.86 mg Hg g^–1. These levels were approximately an order of magnitude greater than those found in the tissues of the reference fish which averaged 0.165 and 0.101 mg Hg g^–1 in muscle and liver respectively. In July 1987, mercury concentrations in the liver of both reference and contaminated fish increased significantly, possibly the result of greater uptake of the metal through increased feeding or changes in the mercury level of selected prey items. Rock bass collected from the two sites in July also had significantly different levels of liver glutathione: reference fish exhibited an elevation and contaminated fish a depression. When fish from the two sampling stations received a 96-hr exposure to 150 µg HgCl₂ in the laboratory, both groups exhibited elevated liver mercury and decreased liver glutathione. Mercury levels in the gall bladders of the exposed fish were also elevated, suggesting that glutathione may have been lost through excretion with the metal in the bile.On the whole, physiological differences between the two groups of rock bass were limited, indicating that exposure to the mercury is not having a significant impact on the rock bass from the contaminated area. This is further supported by field examination of the fish and comparison of condition indices from rock bass previously taken from the same two stations.Those factors which significantly altered the physiology of the rock bass were unique to certain times of the year, indicating that the most appropriate sampling approach in future studies is one which examines a number of variables over a range of environmental conditions.     

Altered burst swimming in rainbow trout Oncorhynchus mykiss exposed to natural and synthetic oestrogens

Juvenile rainbow trout Oncorhynchus mykiss were exposed to two concentrations each of 17β‐oestradiol (E2; natural oestrogen hormone) or 17α‐ethinyl oestradiol (EE2; a potent synthetic oestrogen hormone) to evaluate their potential effects on burst‐swimming performance. In each of six successive burst‐swimming assays, burst‐swimming speed (U_burst) was lower in fish exposed to 0·5 and 1 µg l^?1 E2 and EE2 for four days compared with control fish. A practice swim (2 days prior to exposure initiation) in control fish elevated initial U_burst values, but this training effect was not evident in the 1 µg l^?1 EE2‐exposed fish. Several potential oestrogen‐mediated mechanisms for U_burst reductions were investigated, including effects on metabolic products, osmoregulation and blood oxygen‐carrying capacity. Prior to burst‐swimming trials, fish exposed to E2 and EE2 for 4 days had significantly reduced erythrocyte numbers and lower plasma glucose concentrations. After six repeated burst‐swimming trials, plasma glucose, lactate and creatinine concentrations were not significantly different among treatment groups; however, plasma Cl^? concentrations were significantly reduced in E2‐ and EE2‐treated fish. In summary, E2 and EE2 exposure altered oxygen‐carrying capacity ([erythrocytes]) and an osmoregulatory‐related variable ([Cl^?]), effects that may underlie reductions in burst‐swimming speed, which will have implications for fish performance in the wild.     

Bioenergetic targeting during organ preservation: (31)P magnetic resonance spectroscopy investigations into the use of fructose to sustain hepatic ATP turnover during cold hypoxia in porcine livers

During liver preservation, ATP supplies become depleted, leading to loss of cellular homeostatic controls and a cascade of ensuing harmful changes. Anaerobic glycolysis is unable to prolong ATP production for a significant period because of metabolic blockade. Our aim was to promote glycolysis during liver cold hypoxia by supplying fructose as an additional substrate, compared to supplementation with an equivalent concentration of glucose. Porcine livers (two groups; n = 5 in each) were retrieved by clinical harvesting techniques and subjected to two cycles of cold hypoxia and oxygenated hypothermic reperfusion. In the second cycle of reperfusion, the perfusate was supplemented with either 10 mmol/L glucose (Group 1) or 10 mmol/L fructose (Group 2). During reperfusion in both groups, similar levels of ATP were detected by phosphorus magnetic resonance spectroscopy ((31)P MRS). However, during subsequent hypoxia, ATP was detected for much longer periods in the fructose-perfused group. The rate of ATP loss was sevenfold slower during hypoxia in the presence of fructose than in the presence of glucose (ATP consumption of -7.2 x 10(-3)% total (31)P for Group 1 versus -1.0 x 10(-3)% total (31)P for Group 2; P < 0. 001). The changes in ATP were mirrored by differences in other MRS-detectable intermediates; e.g., inorganic phosphate was significantly higher during subsequent hypoxia in Group 1 (45.7 +/- 2.7% total (31)P) than in Group 2 (33.7 +/- 1.1% total (31)P; P < 0. 01). High-resolution MRS of liver tissue extracts demonstrated that fructose was metabolized mainly via fructose 1-phosphate. We conclude that fructose supplied by brief hypothermic perfusion may improve the bioenergetic status of cold hypoxic livers by sustaining anaerobic glycolysis via a point of entry into the pathway that is different from that for glucose.     

Influence of available dietary carbohydrate content on tolerance of waterborne copper by rainbow trout,Salmo gairdneri Richardson*

The effect of increased available dietary carbohydrate on the tolerance of waterborne copper by rainbow trout was investigated. Trout were reared for six to eight weeks on one of four isocaloric and isonitrogenous diets which varied in available carbohydrate content due to either processing (extrusion v. steam-pelleting) or supplementation with cerelose (0 or 21%). The incipient lethal level (ILL) of copper for trout reared on the low-available-carbohydrate steam-pelleted diet was 350 μg I^?1, significantly higher than the 276 μg I^?1 shown for trout reared on the high-available-carbohydrate extruded diet. The ILL of copper for the low-carbohydrate group (0% cerelose) was 408 μg I^?1, significantly higher than the ILL of 246 μg I^?1 of trout reared on the high-carbohydrate diet (21% cerelose). Increased available dietary carbohydrate resulted in increased liver glycogen content and liver-body weight ratios and decreased liver protein content. Significant correlations were evident between waterborne copper tolerance and liver glycogen, liver-body weight ratios and liver protein. Relative to those on high carbohydrate diets, livers of fish on low-carbohydrate diets contained higher concentrations of copper after seven days of exposure to 0 or 196 μg I^?1 of copper. The elevated liver glycogen levels, resulting from increased dietary carbohydrate, are thought to have impaired liver function, thus reducing copper tolerance.     

Welfare status of cultured seabass (Dicentrarchus labrax L.) and seabream (Sparus aurata L.) assessed by blood parameters and tissue characteristics

Seabass (SBS) and seabream (SBM) juveniles were reared with the goal of obtaining three different final densities (SINT = 40 kg m^?3; INT = 20 kg m^?3; SEM = 0.2 kg m^?3) to ascertain the effects thereof on the welfare of the fish. Significant blood metabolites and hepatic glycogen were determined every 3 months and at harvest. Trials lasted 18 months in seabass and 17 months in seabream. At the end of experiment the main biometric productive parameters and quality of body composition were also recorded. Regarding intensively reared seabass (SINT‐SBS, INT‐SBS), the plasma triglycerids, total cholesterol and transaminases (AST, ALT) were always significantly higher than in semi‐intensively maintained fish (SEM‐SBS). At the final sampling in the SINT‐SBS batch, the total protein and glucose were also markedly increased. Conversely, at harvest the liver glycogen content decreased in SINT‐SBS (34 ± 8 mg g^?1 liver) with respect to INT‐SBS (57 ± 12 mg g^?1 liver) and SEM‐SBS (63 ± 11 mg g^?1 liver). No differences among groups were observed for creatine kinase (CK) and lactate dehydrogenase (LDH). With regard to seabream, SINT‐SBM and INT‐SBM constantly showed plasma triglycerids and total cholesterol as being significantly higher when compared to SEM‐SBM. In the final two blood samplings, SINT‐SBM exhibited the most elevated values for LDH. At harvest, AST, ALT, total protein and glucose markedly increased in SINT‐SBM, whereas liver glycogen content was reduced (22.5 ± 9 mg g^?1 liver), more than in INT‐SBM (70 ± 16 mg g^?1 liver) and SEM‐SBM (75 ± 20 mg g^?1 liver). In both seabass and seabream, body composition was very similar in the different stocking densities, except for total cholesterol. Total n‐3 polyunsaturated fatty acid (PUFA) in seabream was significantly different from fish of the semi‐intensive groups; however, nutritional values and fatty acid profiles were equally good. The intermediate final density of seabass and seabream at 20 kg m^?3 seemed to give the best results in terms of their well being when compared to fish reared at 40 kg m^?3. The absence of differences in blood metabolites and hepatic glycogen levels between the intensive batch and the semi‐intensive groups until harvest was a reference to the positive status of the fish. A density of 20 kg m^?3 can be considered acceptable for farm strategy planning for raising healthy on‐growing seabass and seabream juveniles.     

Hepatic copper-transporting ATPase ATP7B: function and inactivation at the molecular and cellular level

Copper-transporting ATPase ATP7B (Wilson disease protein) is a member of the P-type ATPase family with characteristic domain structure and distinct ATP-binding site. ATP7B plays a central role in the regulation of copper homeostasis in the liver by delivering copper to the secretory pathway and mediating export of excess copper into the bile. The dual function of ATP7B in hepatocytes is coupled with copper-dependent intracellular relocalization of the transporter. The final destination of ATP7B in hepatocytes during the copper-induced trafficking process is still under debate. We show the results of immunocytochemistry experiments in polarized HepG2 cells that support the model in which elevated copper induces trafficking of ATP7B to sub-apical vesicles, and transiently to the canalicular membrane. In Atp7b ^-/- mice, an animal model of Wilson disease, both copper delivery to the trans-Golgi network and copper export into the bile are disrupted despite large accumulation of copper in the cytosol. We review the biochemical and physiological changes associated with Atp7b inactivation in mouse liver and discuss the pleiotropic consequences of the common Wilson disease mutation, His1069Gln.     

Dietary fructose vs glucose lowers copper solubility in the digesta in the small intestine of rats

The hypothesis was tested that dietary fructose vs glucose lowers copper solubility in the digesta in the small intestine of rats, which in turn causes a decreased copper absorption. Male rats were fed adequate-copper (5 mg Cu/kg) diets containing either fructose or glucose (709.4 g monosaccharide/kg) for a period of 5 wk. Fructose vs glucose significantly lowered copper concentrations in plasma and the liver, but did not alter hepatic copper mass. Fructose feeding resulted in a significantly lesser intestinal solubility of copper as based on either a smaller soluble fraction of copper in the liquid phase of small intestinal contents or a lower copper concentration in the liquid phase. The latter fructose effect can be explained by the observed fructose-induced increase in volume of liquid phase of intestinal digesta. After administration of a restricted amount of diet extrinsically labeled with⁶⁴Cu, rats fed fructose also had significantly lower soluble⁶⁴Cu fraction in the digesta of the small intestine. Although this study shows that fructose lowered intestinal copper solubility, only a slight reduction of apparent copper absorption was observed. It is suggested that the fructose-induced lowering of copper status in part counteracted the fructose effect on copper absorption at the level of the intestinal lumen.     

Food deprivation alters osmoregulatory and metabolic responses to salinity acclimation in gilthead sea bream Sparus auratus

The influence of acclimation to different environmental salinities (low salinity water, LSW; seawater, SW; and hyper saline water, HSW) and feeding conditions (fed and food deprived) for 14 days was assessed on osmoregulation and energy metabolism of several tissues of gilthead sea bream Sparus auratus. Fish were randomly assigned to one of six treatments: fed fish in LSW, SW, and HSW, and food-deprived fish in LSW, SW, and HSW. After 14 days, plasma, liver, gills, kidney and brain were taken for the assessment of plasma osmolality, plasma cortisol, metabolites and the activity of several enzymes involved in energy metabolism. Food deprivation abolished or attenuated the increase in gill Na⁺,K⁺-ATPase activity observed in LSW- and HSW-acclimated fish, respectively. In addition, a linear relationship between renal Na⁺,K⁺-ATPase activity and environmental salinity was observed after food deprivation, but values decreased with respect to fed fish. Food-deprived fish acclimated to extreme salinities increased production of glucose through hepatic gluconeogenesis, and the glucose produced was apparently exported to other tissues and served to sustain plasma glucose levels. Salinity acclimation to extreme salinities enhanced activity of osmoregulatory organs, which is probably sustained by higher glucose use in fed fish but by increased use of other fuels, such as lactate and amino acids in food-deprived fish.     

Effects of a Single Therapeutic Dose of Glycerol on Cerebral Metabolism in the Brains of Young Mice: Possible Increase in Brain Glucose Transport and Glucose Utilization

Abstract: This is a study of the effects of a single “therapeutic” dose of glycerol [2 g(22 mmol)/kg i.p.] on brain carbohydrate and energy metabolism in normal nursing weanling mice. Findings were correlated with brain water and electrolyte content and with metabolite changes in plasma, red blood cells, and liver. Plasma glycerol levels peaked at 21 mM 7.5 min after injection and returned to the control value, 0.16 mM, by 2 h. Plasma Na⁺ concentration decreased and plasma protein increased for as long as 2 h after injection. Although red blood cells were freely permeable to glycerol, there was no evidence for glycerol metabolism in these cells. Glycerol levels in liver paralleled those in plasma. Glycerol injection increased liver glucose concentration 23% and doubled hepatic glycerol-1-phosphate levels. Liver ATP levels were reduced 24% after glycerol injection. Brain water concentration was significantly reduced from 7.5 min to 30 min after glycerol injection; brain Na⁺ and K⁺ levels were unchanged. There was no evidence for glycerol entry into brain (the amount detected in brain tissue could be explained by the glycerol content in the blood of the brain). While plasma glucose increased 33%, brain glucose increased 87%. Concomitantly there were statistically significant increases in fructose-1,6-diphosphate, lactate, α-ketoglutarate, and malate levels. The disproportionately high brain glucose value suggests increased transport of glucose from the blood to the brain. Increases in fructose-1,6-diphosphate, lactate, α-ketoglutarate, and malate are compatible with an increased metabolic flux in the glycolytic pathway and Krebs citric acid cycle. As has been previously shown for urea and/or mannitol, these changes may result from the effects of the hyperosmolar glycerol solution on the blood-brain barrier and on cerebral glucose utilization. The sustained lowering of plasma Na⁺ concentration after a single “therapeutic” glycerol injection suggests a need for monitoring plasma Na⁺ levels in the clinical situation. Possible lowering of hepatic ATP levels by the use of glycerol in humans is another concern.     

Exposure to waterborne copper and high temperature induces the formation of reactive oxygen species and causes mortality in the Amazonian fish <Emphasis Type="Italic">Hoplosternum littorale</Emphasis>

Hoplosternum littorale is an Amazon fish that lives in urban areas surrounded by polluted igarapés, where elevated copper concentrations eventually occur. The central goal of this study was to evaluate the associated effects of high temperature and copper contamination on survival time and biochemical responses of the Amazonian fish species H. littorale. We exposed fish to two nominal dissolved copper concentrations (50 and 500 µg l^?1) and combined temperatures of 28 and 34°C. Our findings showed that the combination of these variables affects the survival time of this species. The activity of the biotransformation enzymes ethoxyresorufin-O-deethylase and glutathione-S-transferase showed no alterations in fish within all treatments. The increase of reactive oxygen species and the decrease in potential total antioxidant capacity promoted the imbalance in the antioxidant system. An induction in superoxide dismutase activity occurred in fish exposed to copper concentrations of 50 and 500 µg l^?1 at both temperatures, suggesting liver impairments. Thus, we suggest that H. littorale is sensitive to copper, and this sensitivity is increased further with exposure to high temperatures, particularly in the survival time and reactive oxygen species formation of this fish species.