Effects of stress on growth, cortisol and glucose levels in non-domesticated Eurasian perch (Perca fluviatilis) and domesticated rainbow trout (Oncorhynchus mykiss)

Eurasian perch (Perca fluviatilis) is a promising aquaculture candidate, but the growth performance of this non-domesticated species may be negatively affected by its stress responsiveness to intensive culture conditions. To evaluate this potential problem, juvenile Eurasian perch were exposed to a standardized handling stressor twice a week for an 8-week period. A similar study was conducted on domesticated rainbow trout (Oncorhynchus mykiss) for comparison of intra- and inter-specific differences. The stressed fish of both species showed lower body growth than the non-stressed control fish, however, the final mean body mass was 35.4% lower in the stressed Eurasian perch than in the non-stressed controls, compared to 22.8% difference between the two groups in rainbow trout. The stress responsiveness was examined by comparing the post-stress cortisol and glucose levels in repeatedly stressed fish and fish exposed to the stressor only once. The cortisol stress response in both species strongly indicated a habituation to the repeated stressor. Thus, repeatedly stressed Eurasian perch reached maximum cortisol levels of 130 ng/mL after 0.5 h compared to 200 ng/mL in the fish stressed once, while considerably smaller differences in cortisol levels were shown between the repeatedly and single stressed rainbow trout. Rainbow trout also showed lower post-stress glucose levels in the repeatedly stressed fish compared to the single stressed fish. In contrast, the glucose levels in both groups of Eurasian perch increased abruptly after stress treatment and remained elevated at approximately 19 mM for 6 h; levels were three times as high as the peak levels 3 h post-stress in rainbow trout. Together, the habituation of the stress response shown in both species did not eliminate the growth difference found in the repeatedly stressed fish versus the control fish. Further, the lower growth performance of Eurasian perch compared to rainbow trout could partly be due to the increased energy consumption in the more stress responsive Eurasian perch.     

Modifying the physiological responses to handling stress in beluga sturgeon Huso huso (Linnaeus, 1758); Interactive effects of feeding time and dietary fat

The present study investigated the interaction of feeding times with two dietary fat levels on physiological responses to handling stress in juvenile Beluga sturgeon. Fish were fed with two different diets (high energy; HE: 24% fat and low energy; LE: 12% fat) for 8 weeks at two feeding times; 09:00 and 16:00 (during the day) or 21:00 and 04:00 (during the night). At the end of the trial, blood samples were taken and the resting fish were held in a net out of water for 90 s as the handling stressor. Three hours after application of stress, post‐stress blood was taken. Cortisol, glucose, and lactate concentrations were considered as stress indicators. The mean values of cortisol and lactate levels did not reveal a significant difference between pre‐ and 3‐hr post‐stress samples, but the average concentration of glucose showed a significant difference. Cortisol, lactate and glucose concentrations were not influenced by the difference in the diets. Moreover, the cortisol and lactate concentrations were not affected by the different feeding times, while glucose levels were significantly affected by the feeding times with the lowest level in fish fed during the day. With respect to stress indicators, the results revealed that feeding times affected pre‐ and post‐stress secondary response to handling stress, but the rate and magnitude of metabolites (cortisol, glucose and lactate) were not affected by dietary fat levels. Therefore, it is necessary to examine the best feeding times with the interaction of feed ingredients in sturgeon farming.     

Effect of fasting on body composition and responses to stress in sunshine bass

The integrated responses of the hormonal regulation of growth and stress in sunshine bass (Morone chrysops X Morone saxatilis) as regulated by feed deprivation were investigated. Groups of fish were fed 1.5% of the body weight per day or offered no feed for 4 weeks. Another group of fish was not fed for 3 weeks and feed was offered during the fourth week. Fish in each group were sampled immediately before or after a 15-min low water confinement stressor after each week of the experiment. Liver mass and liver glycogen content were decreased after one week of fasting and remained low until the end of the study. However, both recovered after a week of refeeding. Intraperitoneal fat was significantly lower after two weeks of fasting and did not recover after a week of refeeding. None of these components were affected by confinement stress. Plasma glucose in unstressed fish was generally unaffected by fasting or refeeding; however, plasma glucose increased after confinement stress in fed but not in fasted fish. The cortisol stress response was unaltered by fasting and remained robust. Plasma IGF-I generally decreased in fasted fish but was not significantly lower than fed fish until the fourth week. A week of refeeding did not restore plasma IGF-I concentrations. Plasma IGF-I concentrations were higher in confinement stressed fed fish after two and four weeks but were unchanged in the fourth week. There was no change in the plasma IGF-I concentrations in fasted or refed fish due to the stress. Liver weight and liver glycogen were essentially depleted after 2 weeks of fasting. The reduction of liver glycogen greatly reduced the glucose response to stress; however, the cortisol stress response was maintained for at least four weeks of fasting. Intraperitoneal fat was decreased very little after 4 weeks of fasting. Plasma IGF-I concentrations were reduced only after 3 weeks of fasting.     

Responses to handling and confinement stressors in juvenile great sturgeon Huso huso

The effects of acute stressors on physiological responses of juvenile great sturgeon or beluga Huso huso L. were investigated in two experiments. In the first experiment, fish were handled by placing them in containers at either low density (LD, one fish l^?1) or high density (HD, four fish l^?1) for 60 s. Concentrations of plasma cortisol, glucose and lactate were determined from blood collected at 0, 1, 3, 6 and 12 h after application of the stressor. Plasma cortisol concentrations increased after the disturbance in H. huso from both handling treatments, but changes were not significant. Plasma glucose rose significantly by 22·9 and 31·6% in LD and HD handling treatments, respectively, after 3 h. Significant increases in plasma lactate occurred within 1 h in both treatment groups, but that of the HD group was much higher. In the second experiment, fish were held at two different densities, LD (2 kg m^?2 tank bottom surface area) and HD (7 kg m^?2), for 8 weeks and then subjected to an aerial emersion handling stressor in a net for 60 s; blood samples were taken before handling (resting, 0 h) and at 1, 3, 6 and 9 h after handling. Plasma cortisol increased significantly in fish from the HD treatment from 8·8 ± 0·3 to 19·2 ± 2·4 ng ml^?1 (mean ±s.e. ) by 1 h after stress, but post‐handling changes in the LD group were not significant. Significant increases in both plasma glucose and lactate were observed by 1 h in both treatment groups, with peak levels of plasma glucose evident at 3 h [69·4 ± 2·9 and 60·9 ± 1·7 mg dl^?1 (mean ±s.e .) in LD and HD groups, respectively]. Plasma glucose levels were significantly higher in the LD group than in the HD group at 3 and 6 h. Post‐handling haemoglobin content increased by 1 h and white blood cell numbers were reduced by 3 and 6 h in the HD treatment group compared with resting values, but changes in these blood features in the LD group were not significant. Acute handling did not affect haematocrit in either treatment. The results suggest that H. huso is relatively resistant to handling and confinement, and could tolerate normal hatchery practices associated with aquaculture. Because changes in cortisol concentrations were relatively low compared with those in most teleosts, glucose and lactate concentrations may be more useful as stress indicators in juvenile H. huso. This study also demonstrated that prior exposure to a chronic stressor, specifically high stocking density, could alter the physiological response to subsequent acute handling in H. huso.     

Changes in free and total plasma cortisol levels in juvenile haddock (Melanogrammus aeglefinus) exposed to long-term handling stress

We measured changes in free and total plasma cortisol levels, plasma glucose, gill hsp70 levels, and growth in haddock (Melanogrammus aeglefinus) subjected to a long-term handling stress (15 s out of water, each day, for 4 weeks), and the effect of this long-term stress on the ability of haddock to respond to an acute stressor. The acute stressor was a single handling stress, and fish were sampled at 1, 6, and 12 h post-stress. During the long-term stress study, free and total plasma cortisol levels increased significantly (10-fold) in the stressed group after the second week. However, the percentage of free cortisol was already significantly elevated by the first week (control 17%, stressed 55%), and remained high during the second week (control 35% and stressed 65%). After 3 and 4 weeks of handling, both free and total cortisol declined in stressed fish to levels that were not significantly different from pre-stress values. Control fish grew significantly more than stressed fish (by 32% and 18%, respectively) over the 4 week study, and condition factor only increased in control fish. Although fish from the control group showed elevated total plasma cortisol levels (to 47 ng mL(-1)) 1 h after the acute stress, and the levels in stressed fish were comparable to those for the control fish, no significant increase in plasma cortisol was measured in the group subjected to the long-term stress. Free plasma cortisol levels did not increase significantly in either group following the acute stress. However, free plasma cortisol levels were significantly higher in long-term stress group, as compared with the control group, at 6 h post-stress. Plasma glucose and gill hsp70 levels were not altered by either the long-term stress or acute stressor. Our data indicate that cortisol (free and total), but not glucose or hsp70, appears to be adequate to assess short- and long-term stress in haddock.     

Dietary oils mediate cortisol kinetics and the hepatic mRNA expression profile of stress-responsive genes in gilthead sea bream (Sparus aurata) exposed to crowding stress. Implications on energy homeostasis and stress susceptibility

Juveniles of gilthead sea bream were fed with plant protein-based diets with fish oil (FO diet) or vegetable oils (66VO diet) as dietary lipid sources. No differences in growth performance were found between both groups, and fish with an average body mass of 65–70 g were crowded (90–100 kg/m³) to assess the stress response within the 72 h after the onset of stressor. The rise in plasma cortisol and glucose levels was higher in stressed fish of group 66VO (66VO-S) than in FO group (FO-S), but the former stressed group regained more quickly the cortisol resting values of the corresponding non-stressed diet group. The cell–tissue repair response represented by derlin-1, 75 kDa glucose-regulated protein and 170 kDa glucose-regulated protein was triggered at a lower level in 66VO-S than in FO-S fish. This occurred in concert with a long-lasting up-regulation of glucocorticoid receptors, antioxidant enzymes, enzyme subunits of the mitochondrial respiratory chain, and enzymes involved in tissue fatty acid uptake and β-oxidation. This gene expression pattern allows a metabolic phenotype that is prone to “high power” mitochondria, which would support the replacement of fish oil with vegetable oils when theoretical requirements in essential fatty acids for normal growth are met by diet.     

Recovery from acute, chronic and transport stress in the pot-bellied seahorse Hippocampus abdominalis

Recovery from acute and chronic stress in the pot-bellied seahorse Hippocampus abdominalis under aquaculture conditions was investigated in this study to evaluate (a) whether these relatively slow and sedentary fish develop the 'fight or flight' response and (b) the impact of transportation stress on pot-bellied seahorse physiology. Fish were exposed to either a control treatment, an acute stressor (air exposure for 60 s) or a chronic stressor (confinement or transportation). Plasma concentrations of cortisol, glucose and lactate did not increase in response to an acute stressor, suggesting that adrenergic activation was absent. In contrast, chronic stress initiated increases in cortisol (77 ng ml⁻¹) and glucose (7 mM), which both returned to pre-stress concentrations (4 ng ml⁻¹, 4 mM respectively) within the subsequent 6 h. Recovery from chronic stress thus fitted the teleost paradigm, i.e. plasma levels of cortisol and glucose returned to pre-stress values in c . 6 h during recovery from a chronic stressor. The seemingly rapid return to homeostasis suggests that special precautions, over and above normal procedures, may not be required for long-distance transportation of pot-bellied seahorses.     

Variations of Some Physiological and Immunological Parameters in Siberian Sturgeon (Acipenser baerii,Brandt, 1869) Subjected to an Acute Stressor

This study was carried out to investigate the effect of an acute stressor on the variation of some physiological and immunological parameters of Siberian sturgeon (Acipenser baerii) juveniles. Fish, reared in 3 tanks for 10 weeks, were used for this study. The acute stress of fish consisted of 2 min of air exposure stress. Plasma levels of cortisol, glucose, and lactate as well as lysozyme activity in plasma were measured before stress and 1 hr, 3 hr, 6 hr, 9 hr, 12 hr, and 24 hr after stress. The plasma cortisol significantly increased in the highest level 1 hr after stress, yet it gradually declined after 3 hr. The glucose significantly increased only 1 hr after stress. There was no significant difference between plasma lactate prestress and poststress. Moreover, lysozyme activity was enhanced by stress, thus reaching the highest level 9 hr after stress. The results of this study indicate that Siberian sturgeon not only have a rapid response to acute stress, but also a great capacity for recovery from stress, thus returning physiological parameters to prestress levels after 6 hr.     

Hematology and recovery response in jacundá, Crenicichla saxatilis (Linnaeus, 1758) after short‐term handling stress

The objective of this study was to evaluate the hematological response of ringtail pike cichlid ornamental fish (Crenicichla saxatilis) during the recovery period after short‐term stress. The fish were previously submitted to the stress of chasing, capture and air exposure. Assayed were 24 C. saxatilis (85.2 ± 61.6 g) in three groups of eight fish; after 0.5, 6 and 24 h recovery, blood samples were collected. The total erythrocyte, relative thrombocyte and differential leukocyte counts as well as total hemoglobin, hematocrit, glucose, total plasma protein and the red blood cells (RBC) indices of mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and MCH concentration (MCHC) were determined. Stress responses were observed after 0.5 h, although hyperglycemia remained constant during the experiment. Total lymphocyte and hemoglobin values decreased after 0.5 h in the recovery period. An increase of neutrophils and monocytosis was observed after 0.5 and 6 h, respectively. The MCHC remained stable until after 0.5 h, then varied from this time forward. MCV, MCH and erythrocyte numbers oscillated throughout the experiment. Intense stress was observed in the studied C. saxatilis, with most hematological variables not returning to their initial levels after 24 h.     

Cold shocks: a stressor for common carp

The stress response of common carp Cyprinus carpio was studied by evaluating plasma cortisol, glucose and lactate after single or multiple rapid temperature drops (ΔT: 7, 9 or 11° C). All three amplitudes used induced a significant rise in plasma cortisol levels. Peaks occurred within 20 min after onset of the cold shock. No stress-related secondary metabolic changes were observed in any of the experiments described: plasma glucose levels remained unaffected and plasma lactate levels dropped. Carp of 60 days old showed a significant stress response, although plasma cortisol levels were lower than those observed in carp of 120 days. Furthermore, fish that had experienced multiple cold shocks showed an overall lower cortisol response than fish experiencing a single cold shock, indicating that habituation to this stressor occurred.     

Impairment of the stress response in matrinxã juveniles (Brycon amazonicus) exposed to low concentrations of phenol

In this study we measured plasma cortisol, plasma glucose, plasma sodium and potassium, and liver and gill hsp70 levels in juvenile matrinxã (Brycon amazonicus) subjected to a 96 h exposure to phenol (0, 0.2, and 2.0 ppm), and the effect of this exposure on their ability to respond to a subsequent handling stress. Fish were sampled prior to initiation of exposure and 96 h, and at 1, 6, 12, and 24 h post-handling stress. During the 96 h exposure, plasma cortisol and glucose levels remained unchanged in all treatments. While plasma sodium levels were significantly reduced in all groups, plasma potassium levels only decreased in fish exposed to 0 and 0.2 ppm of phenol. Liver hsp70 levels decreased significantly at 96 h in fish exposed to 2.0 ppm of phenol. All groups, except fish exposed to 0.2 ppm of phenol, were able to increase plasma cortisol and glucose levels after handling stress. Fish exposed to 2.0 ppm of phenol showed decreased gill and liver hsp70 levels after the handling stress. Our data suggest that exposure to phenol may compromise the ability of matrinxã to elicit physiological responses to a subsequent stressor.     

Temperature affects physiological stress responses to acute confinement in sunshine bass (Morone chrysops x Morone saxatilis)

Sunshine bass (Morone chrysopsxMorone saxatilis) were subjected to a 15-min low-water confinement stressor at temperatures ranging from 5 to 30 degrees C. Physiological responses were evaluated by measuring hematocrit, and plasma chloride, glucose and cortisol. Fish acclimated to 30 degrees C had initial glucose concentrations of 3.13 mM (564 mg/L) which were significantly lower than in fish acclimated to 5 and 10 degrees C (4.32 and 4.82 mM or 779 and 868 mg/l, respectively). Fish survived the conditions imposed at every temperature except 30 degrees C, where 15 out of 42 fish died during the stress and recovery protocol. The general pattern was an initial increase in hematocrit, followed by a delayed decrease in hematocrit and chloride, and an increase in plasma glucose and cortisol. In general, fish stressed at temperatures below 20 degrees C had lower and more delayed changes in plasma glucose and cortisol than fish tested at 20, 25 and 30 degrees C. Initial cortisol concentrations were 65 ng/ml and increased to above 200 ng/ml in fish held at 20 degrees C and above. At the higher temperatures, glucose concentrations were twice the initial concentration after stress and cortisol changes were four to five times the initial concentration after the stress. Quantitative responses for glucose and cortisol were moderate and recovery rapid in fish stressed at 10 and 15 degrees C; therefore, this range of water temperature is recommended when handling sunshine bass.     

Daily Rhythms in the Hypothalamus-Pituitary-Interrenal Axis and Acute Stress Responses in a Teleost Flatfish,Solea senegalensis

The endocrine axis controlling the stress response displays daily rhythms in many factors such as adrenal sensitivity and cortisol secretion. These rhythms have mostly been described in mammals, whereas they are poorly understood in teleost fish, so that their impact on fish welfare in aquaculture remains unexplored. In the present research, the authors investigated the daily rhythms in the hypothalamus-pituitary-interrenal (HPI) axis in the flatfish Solea senegalensis, which has both scientific and commercial interest. In a first experiment, hypothalamic expression of corticotropin-releasing hormone (crh) and its binding protein (crhbp), both pituitary proopiomelanocortin A and B (pomca and pomcb) expression, as well as plasma cortisol, glucose, and lactate levels were analyzed throughout a 24-h cycle. All variables displayed daily rhythms (cosinor, p?<?.05), with acrophases varying depending on the factor analyzed: crh and cortisol peaked at the beginning of the dark phase (zeitgeber time [ZT]?=?14.5 and 14.4?h, respectively), pomca and pomcb as well as glucose at the beginning of the light phase (ZT?=?1.2, 2.4, and 3.4?h, respectively), and crhbp and lactate at the end of the dark phase (ZT?=?22.3 and 23.0?h, respectively). In a second experiment, the influence of an acute stressor (30 s of air exposure), applied at two different time points (ZT 1 and ZT 13), was tested. The stress response differed depending on the time of day, showing higher cortisol values (96.2?±?10.7?ng/mL) when the stressor was applied at ZT 1 than at ZT 13 (52.6?±?11.1?ng/mL). This research describes for the first time the daily rhythms in endocrine factors of the HPI axis of the flatfish S. senegalensis, and the influence of daytime on the stress responses. A better knowledge of the chronobiology of fish provides a helpful tool for understanding the circadian physiology of the stress response, and for designing timely sound protocols to improve fish welfare in aquaculture. (Author correspondence: jflopez@um.es)     

Cohesive social behaviour shortens the stress response: the effects of conspecifics on the stress response in lake sturgeon Acipenser fulvescens

An examination was made of whether social interactions can have a beneficial effect through the attenuation of the stress response in a social species. In the first experiment, one larger (mean ± s . e . 194·0 ± 12·5 g) and seven smaller (32·0 ± 2·6 g) juvenile lake sturgeon Acipenser fulvescens were placed in tanks to determine whether a classic dominance effect would be established based on body size ( n = 6). Large fish did not establish a territory or aggressively interact with smaller fish, as there were no significant differences in nearest-neighbour distances and an absence of aggressive behaviour (biting, chasing and pushing). In the second experiment, it was hypothesized that the presence of conspecifics would have a beneficial effect through an attenuation of the stress response. Fish in groups or isolation were stressed by a brief aerial exposure (30 s), and blood plasma was measured at regular time intervals (0, 20, 40, 60, 120 and 240 min) following the stressor via an implanted cannula ( n = 9–11). The presence of conspecifics did not affect the peak cortisol response, however, the overall cortisol response was shorter in duration compared to fish in isolation. Furthermore, secondary stress variables (plasma ions and glucose) showed differences between fish in groups and isolation. The results of these experiments suggest that social interaction plays an important and beneficial role in regulating the stress response in cohesive social species such as A. fulvescens .     

Rates of cortisol increase and decrease in channel catfish and sunshine bass exposed to an acute confinement stressor

Channel catfish and sunshine bass were exposed to a low-water stress event and allowed to recover in fresh water or a solution of metomidate (dl-1-(1-phenylethyl)-5-(metoxycarbonyl) imidazole hydrochloride), which inhibits the synthesis of cortisol. Change in time of plasma cortisol was used as an index of cortisol secretion and clearance. Plasma cortisol and glucose increased during the exposure to low-water stress in both fish, but the changes of both plasma components were more dramatic in sunshine bass. Exposure to metomidate during recovery resulted in a short-term increase in plasma glucose but differences between controls and metomidate-exposed fish were relatively minor thereafter. Cortisol began to decrease in catfish immediately after the removal of the stress but continued to increase for 15 min in sunshine bass recovering in fresh water and for 5 min in bass recovering in metomidate. Catfish recovering in fresh water had a cortisol elimination rate of -1.28 ng/mL/min compared with -2.45 ng/mL/min for fish recovering in metomidate (P>0.05) while sunshine bass recovering in fresh water had an elimination rate of -6.96 ng/mL/min compared with -4.50 ng/mL/min for fish recovering in metomidate (P>0.05). These data indicate that the rapid decrease of plasma cortisol after removal of the stressor is due to an almost immediate decrease of secretion, tissue uptake and a rapid renal loss due to the absence of a plasma binding protein.     

Chronic stress in pregnant guinea pigs (<Emphasis Type="Italic">Cavia aperea f. porcellus</Emphasis>) attenuates long-term stress hormone levels and body weight gain,but not reproductive output

Stress, when extreme or chronic, can have a negative impact on health and survival of mammals. This is especially true for females during reproduction when self-maintenance and investment in offspring simultaneously challenge energy turnover. Therefore, we investigated the effects of repeated stress during early- and mid-gestation on the maternal stress axis, body weight gain and reproductive output. Female guinea pigs (Cavia aperea f. porcellus, n = 14) were either stressed (treatment: exposure to strobe light in an unfamiliar environment on gestational day -7, 0, 7, 14, 21, 28, 35, 42) or left completely undisturbed (control) throughout pregnancy. Females of both groups received the same respective diets, and reproductive parameters were evaluated upon parturition. Additionally, hormonal data were obtained from blood and feces. The stress exposure induced a significant increase in plasma cortisol concentrations during the afternoon. In contrast to this short-term response in plasma cortisol concentrations, we found no significant differences in the levels of cortisol metabolites in feces collected after stress exposure between groups and even significantly decreased levels of fecal cortisol metabolites on non-stress days over time in treatment females. Among treatment females, gain in body weight was attenuated over gestation and body weight was lower compared to control females during lactation, especially in cases of large litter sizes. No differences could be seen in the reproductive parameters. We conclude that repeated stress exposure with strobe light during early- and mid-gestation results in a down-regulation of the hypothalamic–pituitary–adrenal axis and lower weight gain in treatment females, but has no effect on reproductive output.     

Endocrine and metabolic responses to stress in a laboratory population of the tropical damselfish Acanthochromis polyacanthus

The effects of confinement and exercise on the stress response of the spiny damselfish Acanthochromis polyacanthus were investigated in a laboratory stock of fish. Cultured spiny damselfish had basal plasma cortisol values (<16 ng ml⁻¹) similar to those found in wild fish, and basal plasma glucose and lactate levels that were similar to those found in other teleosts. Plasma cortisol concentrations increased in response to stress with a latency period of 5–10 min. Removal of the stressor resulted in partial recovery of cortisol levels by 24 h. Plasma glucose levels increased in response to stress in all experiments with significant increases occurring within 15 min of the imposition of stress. Elevations in plasma glucose concentrations were not initially reflected in changes in liver or muscle glycogen content, with significant reductions in liver glycogen concentrations only occurring in response to extended periods of stress. In contrast to many temperate species, plasma lactate concentrations did not consistently increase in response to stress, suggesting that the stress response in spiny damselfish is not strongly characterized by anaerobiosis.