The impact of social status on the erythrocyte beta-adrenergic response in rainbow trout, Oncorhynchus mykiss

The objective of the present investigation was to determine whether chronic increases in circulating cortisol concentrations, resulting from the occupation of subordinate status in rainbow trout social hierarchies, resulted in an enhancement of the erythrocyte adrenergic response. Rainbow trout (Oncorhynchus mykiss) were confined in fork length matched pairs for 6 h, 18 h, 48 h or 5-7 days, and social status was assigned through observations of behaviour. Erythrocyte adrenergic responsiveness, determined in vitro as changes in water content following incubation with the beta-adrenoreceptor agonist isoproterenol, was significantly greater in subordinate than dominant fish at 48 h of social interactions but not after 5-7 days, nor when assessed as changes in extracellular pH (pHe). However, the activity of the Na+/H+ exchanger (beta-NHE), assessed in vitro as the pHe change following incubation with the permeable cyclic AMP analogue 8-bromo-cyclic AMP, was significantly lower in subordinate fish. The number of erythrocyte membrane-bound adrenergic receptors (Bmax) was significantly higher in subordinate than dominant fish at 48 h, but had decreased by 5-7 days to a value that was not significantly different from that for dominant fish. The apparent dissociation constant (KD) of these receptors was not significantly impacted by either social status or interaction time. Finally, the relative expressions of beta-3b adrenergic receptor (AR) and beta-NHE mRNA were determined using real-time PCR and were found to be minimally affected by social rank. Relative to a control group, beta-3b AR mRNA was significantly up-regulated in both dominant and subordinate trout at all time periods, whereas the expression of beta-NHE was in general significantly down-regulated. Unlike the situation in rainbow trout treated with exogenous cortisol, elevations in circulating cortisol resulting from low social status did not "pre-adapt" the erythrocyte adrenergic response, but rather may have served to offset the potentially adverse effects elicited by plasma catecholamines, which were elevated during social hierarchy formation.     

Effects of autonomic blockade on acute thermal tolerance and cardioventilatory performance in rainbow trout,Oncorhynchus mykiss

Predicted future increases in global temperature may impose challenges for ectothermic animals like fish, but the physiological mechanisms determining the critical thermal maximum (CT_max) are not well understood. One hypothesis suggests that impaired cardiac performance, limited by oxygen supply, is an important underlying mechanism. Since vagal bradycardia is suggested to improve cardiac oxygenation and adrenergic stimulation may improve cardiac contractility and protect cardiac function at high temperatures, we predicted that pharmacological blockade of cardiac autonomic control would lower CT_max. Rainbow trout was instrumented with a flow probe and a ventilation catheter for cardioventilatory recordings and exposed to an acute thermal challenge until CT_max following selective pharmacological blockade of muscarinic or β-adrenergic receptors.Contrary to our prediction, CT_max (~26 °C) was unchanged between treatments. While β-adrenergic blockade reduced heart rate it did not impair cardiac stroke volume across temperatures suggesting that compensatory increases in cardiac filling pressure may serve to maintain cardiac output. While warming resulted in significant tachycardia and increased cardiac output, a high cholinergic tone on the heart was observed at temperatures approaching CT_max. This may represent a mechanism to maintain scope for heart rate and possibly to improve myocardial contractility and oxygen supply at high temperatures. This is the first study evaluating the importance of autonomic cardiac control on thermal tolerance in fish. While no effects on CT_max were observed, this study raises important questions about the underlying mechanisms determining thermal tolerance limits in ectothermic animals.     

The recovery of locomotory activity following exhaustive exercise in juvenile rainbow trout (Oncorhynchus mykiss)

This study investigated the recovery of locomotory activity in exhausted juvenile rainbow trout (Oncorhynchus mykiss, approximately 6-10-cm fork length) in response to two conditions: (1) direct transfer to a range of velocities (0-15 cm s(-1)) in a swim flume (forced swimming) and (2) direct transfer to a pool downstream of a swim channel where a choice of velocities was presented: 2-3 cm s(-1) in the lower half of the pool, a range of velocities from 7 to 40 cm s(-1) in the upper half the pool near the channel entrance, and a velocity of 57 cm s(-1) in a swim channel emptying into the pool (volitional swimming). Exhausted trout showed a pronounced delay in the recovery of normal locomotory activity. With forced swimming, the time required to resume swimming was inversely proportional to water velocity. At 15 cm s(-1), almost all exhausted fish recovered immediately, whereas it took about 1 h for recovery at a current of 5 cm s(-1). In contrast, nonexhausted fish responded to imposed velocity with immediate rheotactic responses (orientation and station holding) at all test velocities. In voluntary swim trials, exhausted trout showed a marked preference for holding station in current in the downstream pool (approximately 11 cm s(-1)) but took, on average, 2 h longer than nonexhausted fish to make transits in the swim channel. Moreover, their ground speed in the swim channel was significantly slower. We conclude that swimming performance is impaired for at least 6 h by exhaustive exercise. Maladaptive behaviors during this time include a preference for current near the surface over cover and a reduced capacity for burst activity, both of which would translate into greater predation risk and reduced ability to forage.     

Exhaustive exercise and the cellular stress response in rainbow trout, Oncorhynchus mykiss

The goal of this study was to examine the cellular response to exhaustive exercise in male and female rainbow trout to determine if HSPs are involved in the early stages of the recovery process. Levels of HSPs and key metabolic parameters were measured in white muscle, heart plasma, and blood plasma throughout 6 h of recovery from exhaustive burst exercise. Plasma creatine kinase (CK) was also quantified as an indicator of exercise-induced tissue damage. The observed trends in ATP and lactate were consistent with established patterns of exhaustion and the beginnings of metabolic recovery. However, no upregulation of hsp70, hsp30, or hsp90 was evident in heart or muscle tissue of males or females, and plasma CK measurements suggest that tissue damage was minimal. Our results indicate that hsp70, hsp30, and hsp90 are not part of the early recovery process from burst exercise in fish, perhaps due to the maintenance of core temperatures as well as a lack of exercise-induced tissue damage.     

Effect of induced triploidy on fin regeneration of juvenile rainbow trout, Oncorhynchus mykiss

Triploidy was induced in the rainbow trout in order to evaluate whether the altered numbers and sizes of triploid cells could modify fin regeneration. Amputation of one lobe of the tail fin of diploid and triploid juveniles resulted in regeneration for experimentals and controls. Nevertheless, both rate and frequency of regeneration in triploids were significantly increased as compared with diploids, a fact that can be attributed to the increased nuclear and cellular volume in a wide range of tissues, whereas the cell numbers were reduced. These data suggest that a great deal of interesting and important research could be done using triploid animals as experimental models for studying the regeneration of appendages.     

Antibacterial proteins in rainbow trout, Oncorhynchus mykiss

Antibacterial proteins are an important part of the innate immune system for all animals. They have been extensively studied in mammals, amphibians and invertebrates, but have received only scant attention in fish. Their expression and processing, however, provide a way of monitoring defence vigour during development or with seasonal changes in physiology. The aim of the present work was to identify and characterise antibacterial proteins in rainbow trout. In vitro analyses of extracts of the peripheral blood leucocytes, head kidney leucocytes and mucus from adult unstimulated (non-immune) fish showed marked antibacterial activity against Gram positive bacteria. Fractionation by ion exchange chromatography and RP-HPLC of head kidney extracts showed the presence of two forms of lysozyme but no constitutively expressed antimicrobial proteins of < 10 kDa. By contrast, chromatographic analyses of mucus revealed at least four antibacterial proteins. Two are conventional lysozymes, a third is an unusual lysozyme-like protein with a low isoelectric point, and the fourth is a highly hydrophobic, cationic peptide of c. 3 kDa.     

Exhaustive exercise does not affect the preferred temperature for recovery in juvenile rainbow trout (Oncorhynchus mykiss)

We tested the hypothesis that juvenile rainbow trout (Oncorhynchus mykiss) would select a temperature colder than their acclimation temperature (16 deg +/-1 deg C) to minimize postexhaustive exercise metabolic demands and enhance oxygen availability. After an initial 3-h exploratory period in a thermal gradient (6 degrees -25 degrees C), fish selected a temperature of approximately 14 degrees C and had a baseline exploratory swimming activity of approximately 60 cm min(-1). Subsequently, experimental (chased) fish were individually removed, exhaustively exercised for 1.5 min, and replaced. Both control (unchased) and experimental fish were allowed to explore the thermal gradient for another 2 h. Immediately after being chased, trout had a metabolic profile that was consistent with being exhausted; levels of plasma and muscle lactate were 4.38+/-0.25 mmol L(-1) and 28.0+/-2.0 mmol kg(-1), respectively, and levels of muscle glycogen, adenosine triphosphate, and phosphocreatine were 3.89+/-0.95, 4.23+/-0.62, and 3.07+/-0.73 mmol kg(-1), respectively. Although exploratory swimming activity of the chased fish was significantly lower (by 81%) as compared with control fish during the first 5 min postchase, differences in the mean, median, and mode values for selected temperatures during the next 2 h were neither large (<1 degrees C) nor significant (P>0.05). Contrary to our initial hypothesis, these findings suggest that juvenile rainbow trout do not select a colder temperature to decrease metabolic rate following exhaustive exercise. Instead, rainbow trout selected a temperature marginally cooler than their acclimation temperature (16 degrees C) regardless of whether they had been previously exhausted.     

虹鳟对时间和地点的学习记忆(英文)

为确定虹鳟(Oncorhynchus mykiss)是否能将日投放两次食物的不同时间和地点联系起来,在30天内,于09:00-10:00将食饵投放在水族箱一侧,于15:00-16:00将食饵投放在水族箱另一侧.在第21天和28天,不放食饵以确定虹鳟对时间-地点的学习记忆.结果表明,虹鳟将不同时间和地点联系起来以获取食物,并且在水族箱两个投放食饵之处均表现出预取食活动,提示该物种具有控制时间-地点学习任务的内在机制.     

The effect of β-adrenergic blockade on the recovery process after strenuous exercise in the rainbow trout, Salmo gairdneri Richardson

Trout fitted with arterial catheters were subjected to 6 min of strenuous exercise, injected with either saline (controls) or the β-adrenergic antagonist propranolol, and monitored over the following 8-h recovery period. Control responses were very similar to those previously reported, except for much higher resting and post-exercise plasma catecholamine levels, and less marked RBC pHi regulation, perhaps due to season (February–May). Trout subjected to prior β-blockade would not exercise. Trout β-blocked immediately after exercise showed a much higher incidence of mortality during the recovery period, but accompanying symptoms were similar to those previously documented in control trout dying after exercise. Specific effects of post-exercise β-blockade seen in both survivors and mortalities were a sustained elevation of arterial Pco₂ and an inhibition of blood glucose elevation. There were negligible effects on RBC pHi and volume regulation, blood metabolic acid and lactate dynamics, or plasma ion changes. The results provide little support for the hypothesis that β-adrenergic actions of plasma catecholamines are intimately involved in post-exercise recovery, but must be considered in the context of the 'winter' trout, where β-responses may be diminished.     

Culturing and characterization of astrocytes isolated from juvenile rainbow trout (Oncorhynchus mykiss)

An access to brain cell cultures from fish would enable screening of possible neurotoxic chemicals contaminating the aquatic environment. In the present study, a protocol for a successful routine isolation and culturing of brain cells from juvenile rainbow trout was worked out. The coating material was shown to be of importance for cell proliferation. Cells grow better on a surface coated with laminin than on those coated with poly-L-lysine (PLL), poly-D-lysin (PDL) or poly-L-ornithine (PLO). The best cell growth was obtained on double-coated surfaces (PLL, PDL or PLO plus laminin). On such a culture substrate and with a seeding density of 1 x 10(7) cells/cm(2) confluence was obtained within 3-4 weeks at an incubation temperature of 18 degrees C. Approximately 95% of the cells were identified as astrocytes on the basis of a positive staining with antibodies against the astrocyte specific glial protein (GFAP). No oligodendrocytes or fibroblasts were identified in the cultures, and despite several efforts, neurons did not grow under the culture conditions used. When challenged with ligands known to awake a calcium transient in mammalian astrocytes, 44% of the cells responded to ATP with an increase in [Ca 2+](i), 38% to norepinephrine, 27% to 5-hydroxytryptamine, 7% to histamine and 6% to glutamate. Kainate, quisqualate and gamma-aminobutyric acid did not awake a calcium transient in the cells. Using a proper protocol, it is thus quite easy to get an almost pure culture of astrocyte, whereas neurones proved to very difficult to culture.     

The effect of Cu on gill and esophagus lipid metabolism in the rainbow trout Oncorhynchus mykiss

Rainbow trout Oncorhynchus mykiss were exposed to 0, 100, 300 and 800 μgl⁻¹ ambient Cu in brackish water (BW) for 4 days at 13 °C and subsequently transferred to either clean BW, clean fresh water (FW) or clean seawater (SW) at 16 °C. After incubation with ³²P-phosphate and ¹⁴C-acetate added as precursors to the water the fish showed a degradation, depending on previous [CU], of ³²P- and ¹⁴C-labelled gill membrane phospholipids if they had been transferred to SW or remained in BW. Corresponding experiments where the fish were exposed to Cu in BW for 12 days showed a similar subsequent degradation in SW and BW of both gill and esophagus membrane phospholipids, however to a much lesser degree in gill tissue than after 4 days. Plasma Na⁺ was similarly reduced by up to 8%, depending on previous ambient Cu, but in this case only after transfer to FW and only after 4 days of exposure. Both the effect on membrane lipid metabolism and plasma Na⁺ thus showed acclimation to ambient Cu but there was apparently no direct correlation between the two different types of observed changes in membrane function.     

Effects of some pesticides on the vital organs of juvenile rainbow trout (Oncorhynchus mykiss)

Gill, trunk kidney, spleen, and liver of rainbow trout (Oncorhynchus mykiss) were examined after exposure to different sublethal concentrations of carbosulfan (25, 50 and 200 μg L⁻¹), propineb (3, 6 and 24 mg L⁻¹), and benomyl (2, 5 and 20 mg L⁻¹) for 14 days. Lesions were observed in gill, trunk kidney, spleen, and liver of rainbow trout exposed to either concentration of pesticides. The most important lesions were determined in the highest concentrations of pesticides. Lamellar fusion, lamellar hyperplasia, epithelial lifting, vacuolization of epithelial tissue, epithelial necrosis, hypertrophy and sloughing of epithelium were observed on fish exposed to carbosulfan, propineb and benomyl. Fish had cell necrosis, degeneration and oedemas in liver, trunk kidney and spleen. None of these lesions were seen in control fish.     

Early biochemical effects of Microcystis aeruginosa extracts on juvenile rainbow trout (Oncorhynchus mykiss)

Microcystins (MC) are usually the predominant cyanotoxins associated with cyanobacterial blooms in natural surface waters. These toxins are well-known hepatotoxic agents that proceed by inhibiting protein phosphatase in aquatic biota; recent studies have also reported oxidative stress and disruption of ion regulation in aquatic organisms. In the present study, young trout (Oncorhynchus mykiss) were exposed to crude extracts of Microsystis aeruginosa for four days at 15 °C. The level of microcystins was calculated to confirm the presence of toxins in these crude extracts: 0, 0.75, 1.8 and 5 μg/L. Protein phosphatase measured in the liver increased by at least 3-fold and is significantly as a result of exposure to these sublethal concentrations of crude extract, his indicates an early defense response against protein phosphatase inhibition from cyanotoxins. This was corroborated by the decreased phosphate content in proteins found in the liver and brain. No increase in glutathione-S transferase (GST) activity was observed and lipid peroxidation was unaffected in both liver and brain tissue exposed to the cyanobacterial extracts. The data revealed that the proportion of the reduced (metal-binding) form of metallothionein (MT) decreased by two-fold relative to the control group (with a concomitant increase in the proportion of the oxidized form). The level of phosphate associated with MT increased by 1.5-fold at the highest concentration of crude extract. Acetylcholinesterase (AChE) activity in brain tissue was decreased after exposure to the highest concentration of crude extract, suggesting a slowdown in neural activity. However, no biotransformation processes or detoxification of GST was triggered. Our findings show early sign of biochemical effects of MC-LR in young trout.     

A blue variant in the rainbow trout, Oncorhynchus mykiss Walbaum

A blue variant of the rainbow trout, which appeared in a French fish farm, displayed an iridescent body color that was cobalt blue on the back, lighter on the undersides, and silvery on the belly and which held up to adult stage. This color was supposed to result from a Tyndall effect involving a structural arrangement of melanin pigments because it disappeared when it was associated with a depigmenting gene. This blue variant appeared to be governed by an autosomal recessive gene. Blue fry survival and body weight were about 25% less than those of wild-type sibs, but no major problem was observed in further breeding performances, including reproduction. These features do not correspond with those of the blue variants previously described in the rainbow trout.     

Effects of dietary thymol‐carvacrol on growth performance,hematological parameters and tissue composition of juvenile rainbow trout,Oncorhynchus mykiss

Growth performance, carcass quality, survival and hematological responses of Oncorhynchus mykiss juveniles (initial weight 8.4 ± 0.1 g) fed diets containing thymol‐carvacrol powder at the levels of 0, 1.0, 2.0, 3.0 g kg^?1 were tested. Thymol‐carvacrol powder originated from Origanum vulgare, a Mediterranean plant, added to diets. Each diet was fed to triplicate groups of fish for 45 days. Fish fed diets containing thymol‐carvacrol had significantly higher final weight and growth than the control group. Food conversion ratio in fish fed diets containing 2.0 and 3.0 g kg^?1 thymol‐carvacrol was statistically better than in other treatments. Survival was not different among all treatments. The number of lymphocytes increased when thymol‐carvacrol was used at higher levels. Furthermore, whole body lipid content was higher in fish fed 1.0 and 2.0 g kg^?1 thymol‐carvacrol than the other groups, but body protein in the group fed 3.0 g kg^?1 was higher than in other groups. Also, body ash in control and 1.0 g kg^?1 was higher than in other groups. Whole body dry matter was not affected by dietary treatments. These results indicated that dietary administration of thymol‐carvacrol can influence some growth, hematological parameters and tissue composition in rainbow trout juveniles.     

Social context, competitive interactions and the dynamic nature of antipredator responses of juvenile rainbow trout Oncorhynchus mykiss

The effects of food ration and social context, as well as possible interactions, on the antipredator behaviour of juvenile rainbow trout Onchorhynchus mykiss were investigated in a pair of laboratory studies. In experiment 1, pairs of dominant and subordinate O. mykiss were exposed to conspecific alarm cues when maintained under high or low food rations. Under high food rations, dominant individuals responded to predation risk, whereas subordinates used the opportunity to feed. Under low food ration, however, the opposite pattern was observed, where subordinates responded to predation cues and dominants did not. Experiment 2 consisted of performing the same experiment, however separating the dominant and subordinate O. mykiss 3 h before testing. When tested separately, dominant and subordinate individuals did not differ in their responses to alarm cues, regardless of food ration. These results demonstrate that there is a complex interaction between current energy status and social context on decision making by prey animals.     

Exposure to a contextually neutral stressor potentiates fear conditioning in juvenile rainbow trout,Oncorhynchus mykiss

Organisms faced with stressors deploy a suite of adaptive responses in the form of behavioral, physiological and cognitive modifications to overcome the challenge. Interactive effects of these responses are known to influence learning and memory processes and facilitation is thought to be dependent, in part, upon contextual relevance of the stressor to the learning task. Predation is one such stressor for prey animals, and their ability to manage reliable information about predators is essential for adaptive antipredator strategies. Here, we investigated (i) the influence cortisol has on the ability of juvenile rainbow trout to learn and retain conditioned antipredator responses to predatory cues, and (ii) whether conditioned behavioral and physiological responses to predator cues are fixed or deployed in a threat-sensitive manner. Trout were fed cortisol-coated pellets minutes prior to a conditioning event where they were exposed to novel predator odor paired with chemical alarm cues (unconditioned stimulus). We tested for conditioned responses by exposing trout to predator cues after 2, 4 or 10 days and subsequently documented physiological and behavioral responses. Both control and cortisol-fed trout learned the predator odor and responded 2 and 4 days post conditioning. However, at 10 days only cortisol-fed trout maintained strong behavioral responses to predator cues. Interestingly, we failed to find conditioned physiological responses to predator odor despite the presence of threat-sensitive cortisol responses to the unconditioned stimulus. Our findings suggest cortisol exposure prior to predator-learning may enhance retention of conditioned responses, even without a contextual link between stressor source and learning task.     

Interaction between dietary calcium supplementation and chronic waterborne zinc exposure in juvenile rainbow trout, Oncorhynchus mykiss

This study investigated the effects of dietary Ca2+ on branchial Ca2+ and Zn2+ uptake, new and total zinc accumulation in target tissues (gill, liver and kidney), calcium and zinc homeostasis, and acute tolerance to waterborne zinc in fish chronically exposed to waterborne zinc. Juvenile rainbow trout (Oncorhynchus mykiss) were maintained on a calcium-enriched diet [41.2 mg vs. 21.2 mg (control) calcium/g dry wt. of food] and chronic waterborne zinc exposure (2.3 micromol/L), both separately and in combination, for 28 days. Calcium-supplemented diet in the absence of waterborne zinc significantly reduced branchial Ca2+ and Zn2+ influx rates, and new and total zinc accumulations in target tissues relative to control. However it did not protect against the acute zinc challenge. In contrast, waterborne zinc exposure significantly increased branchial Ca2+ and Zn2+ influx rates, new and total zinc concentrations in target tissues, and acute zinc tolerance relative to control. Interestingly, no such changes in any of these parameters were recorded in fish treated simultaneously with elevated dietary Ca2+ and waterborne zinc, except acute zinc tolerance which was highest among all the treatments. Thus, we conclude that the interactions between elevated dietary Ca2+ and waterborne zinc can protect freshwater fish against waterborne zinc toxicity.