Altered stress responses in rainbow trout following a dietary administration of cortisol and β-napthoflavone

Previous studies have demonstrated that fish inhabiting polluted waterways often have an impaired stress response at the organismal level. Given the possible link between the organismal (i.e. cortisol) and cellular (i.e. heat shock proteins; hsp) stress responses, we conducted this study to examine the ability of rainbow trout to respond to a 2 h, +14 °C heat stress (HS) challenge following a 28 d, sub-chronic exposure to increased concentrations of cortisol (5 mg kg⁻¹ b.w.), β-napthoflavone (bnf; 50 mg kg⁻¹ b.w.), and a combination of both (mixture), through the diet (1.5% b.w. every 48 h). While control fish responded to the HS by significantly increasing components of their organismal (cortisol, glucose, and lactate) and cellular (hepatic hsp70 protein) stress responses 6 and 24 h post HS, cortisol-, bnf-, and mixture-fed fish had impaired stress responses at both levels of organization. Additionally, hepatic hsp70 levels were significantly reduced 6 h post HS in cortisol-fed fish. While bnf-fed fish had significantly higher EROD activity, cortisol potentiated EROD activity in the mixture-fed fish. Similarly, plasma cortisol concentrations in the mixture-fed fish were significantly lower relative to cortisol-fed fish. These data are the first to indicate that sub-chronically stressed fish can have impaired stress responses at both the organismal and cellular levels. These findings raise questions regarding: (a) the universal and simple applicability of biological indicators of stress in fish; (b) the possible functional relationship between these two levels of stress responses; and (c) the importance of hsps in the generalized stress response of the whole organism. This revised version was published online in August 2006 with corrections to the Cover Date.     

Stress hormones and the cellular stress response in salmonids

The relationship between stress protein (SP) levels and the hormonal stress response in salmonids was examined through the measurement of gill SP70 and SP30 levels together with plasma cortisol, glucose and ion concentrations, in response to handling stress (45 s holding in a net), intraperitoneal cortisol implants, and heat shock (+10 °C). Handling and cortisol implants resulted in increased plasma cortisol and glucose levels. Heat shock following handling reduced plasma [Na⁺] below that observed in response to the handling stress alone, and heat shock following cortisol implant significantly lowered both plasma [Cl^–] and [Na⁺] below that of fish receiving the cortisol implant alone. Increased SP70 levels occurred 1 h following the 2 h +10 °C heat shock. Handling the fish prior to the application of heat shock suppressed the increase of SP70 levels in the gills. However, increased plasma cortisol concentrations alone did not attenuate gill tissue SP70 increase caused by heat shock. Physiological (10^–7 M) and pharmacological (10^–5 M) concentrations of adrenaline caused increased levels of SP70 in hepatocytes. Addition of the -blocker propanolol blocked this response to adrenaline. The results indicate that handling procedures do not result in an increase of hsp30 or hsp70 and may suppress hsp synthesis under certain circumstances.     

Effects of Pro-Tex on zebrafish (Danio rerio) larvae,adult common carp (Cyprinus carpio) and adult yellowtail kingfish (Seriola lalandi)

Aquaculture practices bring several stressful events to fish. Stressors not only activate the hypothalamus–pituitary–interrenal-axis, but also evoke cellular stress responses. Up-regulation of heat shock proteins (HSPs) is among the best studied mechanisms of the cellular stress response. An extract of the prickly pear cactus (Opuntia ficus indica), Pro-Tex, a soluble variant of TEX-OE^®, may induce expression of HSPs and reduce negative effects of cellular stress. Pro-Tex therefore is used to ameliorate conditions during stressful aquaculture-related practices. We tested Pro-Tex in zebrafish (Danio rerio), common carp (Cyprinus carpio L.) and yellowtail kingfish (Seriola lalandi) exposed to aquaculture-relevant stressors (thermal stress, net confinement, transport) and assessed its effects on stress physiology. Heat shock produced a mild increase in hsp70 mRNA expression in 5-day-old zebrafish larvae. Pro-Tex increased basal hsp70 mRNA expression, but decreased heat-shock-induced expression of hsp70 mRNA. In carp, Pro-Tex increased plasma cortisol and glucose levels, while it did not affect the mild stress response (increased plasma cortisol and glucose) to net confinement. In gills, and proximal and distal intestine, stress increased hsp70 mRNA expression; in the distal intestine, an additive enhancement of hsp70 mRNA expression by Pro-Tex was seen under stress. In yellowtail kingfish, Pro-Tex reduced the negative physiological effects of transport more efficiently than when fish were sedated with AQUI-S^®. Overall, our data indicate that Pro-Tex has protective effects under high levels of stress only. As Pro-Tex has potential for use in aquaculture, its functioning and impact on health and welfare of fish should be further studied.     

Stress response of juvenile rainbow trout (<Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>)to chemical cues released from stressed conspecifics

The objective of this study was to determine whether exposure of rainbow trout (Oncorhynchus mykiss) to water containing a stressed trout or skin extract from stressed and non-stressed trout would elicit a stress response in conspecifics. Juvenile rainbow trout were exposed for 1 hour to water containing a stressed fish, homogenized skin extracts from a non-stressed fish, skin extract from a stressed fish and water with none of these factors. The stress response was measured over a 24-h period (1, 6, 12, 24 h after exposure). Plasma cortisol levels increased at 12 h in fish exposed to water from a stressed fish and skin extract from a stressed fish. Plasma glucose and hepatic hsp70 levels were not affected by treatments. The results suggest that rainbow trout elicit a stress response when exposed to stress-related alarm cues released from conspecifics.     

Indicators of reproductive performance in rainbow trout Oncorhynchus mykiss (Walbaum) selected for high and low responsiveness to stress

A number of indicators of reproductive performance were assessed in rainbow trout divergently selected on the basis of their plasma cortisol response to a standardized stressor. For both male and female rainbow trout Oncorhynchus mykiss (Walbaum), body weight was significantly related to stress responsiveness. High‐responding (HR) females were significantly heavier than low‐responding (LR) females overall, and HR males were significantly heavier than LR males overall. There was no significant difference in the specific growth rate (SGR) of HR and LR female fish during the experimental period, whereas in contrast the mean SGR of HR males was significantly greater than that of LR males. The divergent confinement‐induced levels of blood cortisol in HR and LR groups were stable for more than 1 year after selection. There was no significant difference in the extent to which confinement stress reduced blood oestradiol‐17β levels in HR and LR females, despite a large difference in relative blood cortisol levels. The onset and rate of ovulation was similar within the HR and LR groups. Differences in fecundity, relative fecundity and egg volume and weight were wholly attributable to the different mean body weights of the HR and LR fish. There was no difference between the sperm counts of HR and LR males. There was significantly higher mortality among eggs derived from HR parents overall between fertilization and hatch, but maximum mean mortality did not exceed 12%. Mean time to eyeing in the HR groups was 224 degree days, and in the LR groups was 244 degree days. Hatching commenced at 344 and 347 degree days in the HR and LR groups, respectively, and was complete within 412 and 416 degree days respectively. Overall, selection for stress responsiveness was associated with no significant benefits or costs in reproductive performance.     

Effect of some repetitive factors on turbot stress response

The stress response of the turbot, Scopthalmus maximus (L.), to repetitive factors including netting, air exposure, blood sampling and hand-stripping, were tested using two different tank sizes as well as two blood sampling techniques. Exposure of juvenile fish to air for 1-4 min had no immediate effect on plasma cortisol concentrations or haematocrit values. Similarly, the serial netting of immature fish from tanks did not significantly modify plasma cortisol concentrations, haematocrit or osmolarity values. Hand-stripping of mature males was more disturbing than air exposure. The cumulative effect on plasma cortisol levels and osmolarity of stress factors such as netting, air exposure, blood sampling and stripping applied simultaneously to mature males in a 16 m3 tank and repeated twice daily for several days was recorded. The level of cortisol increased from 5 ng ml-1 to 300 ng ml-1 after 10 days of treatment, while an osmoregulatory imbalance and fish death were observed. Moreover, adaptation of fish to smaller tanks seemed to improve the increased plasma cortisol levels and death rate. Reduction in the number of stress factors applied greatly decreased both the range of physiological responses and the death rate. In order to avoid a cumulative stress response, handling of fish should therefore be reduced to a minimum.     

Change in blood stress and antioxidant markers and hydromineral balance of common carp (Cyprinus carpio) anaesthetized with citronellal and linalool: Comparison with eugenol

The present study aimed to investigate the effects of anaesthesia with citronellal and linalool on stress, antioxidant and hydromineral responses of common carp. Eugenol was used as control anaesthetic, as it is a common anaesthetic in aquaculture. The fish (110 ± 5.65 g) were exposed to 406 mg/L citronellal, 982 mg/L linalool or 43 mg/L eugenol within 5 min (short‐term anaesthesia) or 10 and 20 mg/L of each anaesthetic for 3 hr (long‐term exposure), before blood sampling. The results showed that the short‐term anaesthesia had no significant effects on plasma lactate, sodium, chloride and phosphorus levels and blood catalase (CAT) and superoxide dismutase activities. The short‐term anaesthesia with linalool led to higher stress responses (increased cortisol and glucose levels) and oxidative stress (increased malondialdehyde [MDA] level and decreased total antioxidant capacity [TAC] and glutathione peroxidase [GPx] activity) compared to the eugenol and citronellal. Citronellal group had significantly higher cortisol and lower TAC level and GPx activity compared to eugenol group. Under the long‐term exposure, increase in anaesthetic concentration led to significant increase in plasma cortisol, glucose and phosphorus levels. The linalool‐treated fish had significantly higher cortisol and MDA levels compared to the other groups. The eugenol group had significantly higher lactate and phosphorus and lower TAC levels compared to the other groups. Although citronellal causes slight increase in plasma cortisol level, it induces lower tissue damages compared to eugenol. Thus, citronellal might be an alternative anaesthetic for carp anaesthesia based on the present tested parameters.     

The ability of clove oil and MS-222 to minimize handling stress in rainbow trout (Oncorhynchus mykiss Walbaum)

The effects of 60‐mg L^?1 clove oil and 60‐mg L^?1 tricaine methanesulphonate (MS‐222) on the blood chemistry of rainbow trout were compared after exposure to handling stress via caudal puncture blood sampling. Fish sampled by caudal puncture and subsequently exposed to anaesthetics showed a typical handling stress response over a 48‐h period. There were no significant differences between the responses of fish exposed to equal concentrations of clove oil and MS‐222, with the following exceptions: the blood glucose at full anaesthesia, and lactate at full recovery increased significantly in the clove oil‐exposed fish. In a subsequent experiment, the stress response observed in fish sampled by caudal puncture and exposed to clove oil and MS‐222 was compared with a non‐anaesthetized control group. The increases in plasma cortisol levels were significantly lower at recovery in fish treated with either anaesthetic compared with the control fish. Fish exposed to MS‐222 had significantly higher cortisol levels at 1 h. These findings show that few differences exist between the anaesthetic effects of clove oil and MS‐222 on the physiological response of fish to stress. However, clove oil is more effective at reducing the short‐term stress response induced by handling and blood sampling, and is recommended as an alternative fish anaesthetic.     

黄条鰤hsp70基因克隆及其在早期生长发育过程中的表达调控特性

热休克蛋白(heat shock proteins, HSPs)在鱼类的应激与免疫反应中发挥重要的生理调控作用,HSP70是该家族的重要成员。为探讨热休克蛋白在大洋性经济鱼类黄条鰤 (Seriola aureovittata)生长发育中的生理作用,本研究克隆获得了黄条鰤hsp70基因的全长cDNA序列,并采用定量PCR技术测定了其组织分布及在早期生长发育过程中的表达特征。结果显示,黄条鰤 hsp70基因的cDNA序列全长为2 332 bp,其中,5′-UTR长度为187 bp,ORF长度为1 920 bp,3′-UTR长度为225 bp,编码639个氨基酸,蛋白质分子量为70.1 kDa,等电点为5.16。黄条鰤 hsp70 mRNA的组织表达具有性别二态性差异,其中,在雌性鳃、心、脾脏和卵巢组织中显著高表达(P<0.05),且以卵巢中表达量最高;雄性垂体、鳃、头肾和精巢组织显著高表达(P<0.05),且以鳃中表达量最高。胚胎发育过程的表达检测显示,在卵裂前的受精卵中可检测到hsp70的表达,表明其具有亲本遗传的特性。同时,在胚胎发育过程的各个时期都可检测到hsp70 mRNA的表达,且在低囊胚期之前的各发育阶段一直保持较低表达水平,在原肠前期开始显著上调表达(P<0.05),其后保持相对较高表达水平,至胚胎孵化出膜期达峰值。在仔稚幼鱼中,hsp70 mRNA在初孵仔鱼和1 d仔鱼中高表达,其后在4 d仔鱼中显著降低(P<0.05),其后显著上调表达,至15 d仔鱼达峰值,其后在20 d仔鱼显著下降,并在25 d后稚鱼和幼鱼中保持相对较低表达水平。研究结果可为深入认识黄条鰤hsp70基因的结构特征、发生发育及其早期生长发育阶段的表达调控功能提供依据。     

Effects of dietary inclusions of red beet and betaine on the acute stress response and muscle lipid peroxidation in rainbow trout

This study evaluates the effects of red beet (RB) and betaine on rainbow trout submitted to an acute stress challenge. A control diet was compared with four experimental diets in which red beet (14 and 28%) and betaine (0.9 and 1.63%) were incorporated in different concentrations according to a factorial design. Cortisol in plasma and fin, glucose and lactate plasma levels, and malondialdehide (MDA) in muscle were all measured before the stress challenge and 30 min and 6 and 12 h after the stress challenge as parameters to determine the diet effects. RB and betaine had no effect on cortisol, glucose, and MDA basal levels. However, lactate basal levels were significantly lower on fish fed with RB and betaine. Thirty minutes after the stress challenge, there was a significant increase in plasma and fin cortisol, glucose and lactate concentrations, although fish fed with diets containing RB and betaine showed significantly higher plasma cortisol values. MDA values of fish fed with 14% RB and 0.9% betaine were significantly higher than MDA values from fish fed with 28% RB and 1.63% betaine. After 6 and 12 h, plasma and fin cortisol and lactate levels recovered in a similar trend. Glucose plasma levels recovered in almost all groups 12 h after the stress. Also, MDA values recovered basal levels after 6 and 12 h. RB and betaine did not enhance the tolerance to the stress challenge compared to the control group, although the presence of these ingredients had no negative effect on any of the stress indicators.     

Physiological stress responses in spotted wolffish (Anarhichas minor) subjected to acute disturbance and progressive hypoxia

The spotted wolffish (Anarhichas minor) is considered a promising species for coldwater aquaculture. It is a sedentary, bottom-dwelling fish which exhibits a calm and “non-stressed” behaviour in captivity. There are, however, no reports on the physiological responses to stressors in this species. In the present study we investigated primary (cortisol secretion) and secondary (glucose mobilization) responses to common aquaculture stressors like disturbance and hypoxia. Pre-stress plasma cortisol levels were within those generally considered representative for unstressed fish (~ 10 ng ml^− 1), whereas basal glucose levels were unusually low (0.3–0.4 mM). After exposure to a short, but severe disturbance challenge (emptying the tank of water for 10 min), the increase in plasma cortisol level was slow and relatively weak, reaching a peak level of 25 ng ml^− 1 4 to 8 h after disturbance. When the fish were exposed to a gradually decreasing oxygen level in the tank, a significantly elevated plasma cortisol level (35 ng ml^− 1) was seen in the fish that remained in the tank until oxygen saturation had decreased to 20% oxygen saturation after 2.5 h. However, a two-fold higher plasma cortisol level (~ 70 ng ml^− 1) was seen in all fish exposed to reduced oxygen levels (60, 40 and 20% oxygen saturation) after 3.5 h recovery in normoxic water. Plasma glucose levels showed only moderate increases (~ 70%) following disturbance and hypoxia challenges. An in vivo injection of ACTH caused a strong elevation of plasma cortisol (peak level ~ 170 ng ml^− 1), demonstrating a high capacity for interrenal steroidogenesis in the spotted wolffish. The slow and relatively weak cortisol response to stressors, and low plasma glucose levels, may relate to the sedentary lifestyle of the spotted wolffish. The stress-response is characterised by a passive (reactive) coping style, which is considered adaptive for farming of this species.     

Effects of hormonal manipulation on stress responses in male and female broodstocks of pikeperch Sander lucioperca

This study was designed to determine the effects of hormonal manipulation on stress responses in female and male pikeperch. Two-year-old cultured female and male broodstocks with an average weight of 337.4 ± 20.1 (mean ± SE; n = 16) and 318.7 ± 15.1 g (n = 16), respectively, were randomly allocated into four hormonal treatments each containing 4 fish. Two sexual groups of 16 fish for each gender were considered. Sexually mature male and female pikeperch were injected with either physiological saline solution (as control group), common carp pituitary extract (CPE), human chorionic gonadotropin (hCG) and or luteinizing hormone-releasing hormone analog (LHRHa₂). The blood samples were taken before hormonal injection and after ovulation and spermiation. Then the plasma levels of stress indices (cortisol, glucose, and lactate) were determined. The results showed that all CPE-, HCG-, and LHRHa_2- injected males produced sperm. In females treated with CPE and hCG, three of four ovulated, but none of LHRHa_2- and saline-injected fish spawned. Significant changes in cortisol, glucose, and lactate levels were observed among the females injected with different hormones. Plasma cortisol and glucose levels increased significantly in males injected with CPE and females injected with hCG, but no significant change was observed in lactate levels before and after hormonal induction. Comparison of two sexes revealed significant differences in glucose levels for females in some groups before injection, while CPE-injected sexes showed significant changes in cortisol and lactate concentrations. The results indicated that the induction of ovulation or spermiation stimulated stress responses especially in female pikeperch, and therefore, all the procedures should be made to minimize the disturbance during the artificial spawning.     

Cellular responses to temperature stress in steelhead trout (Onchorynchus mykiss) parr with different rearing histories

This study compared the heat-shock response and metabolic energy status in hatchery-raised and two groups of wild-caught steelhead trout parr collected from the Navarro River watershed, California. Wild parr were from coastal and inland sites with different thermal regimes. Fish were exposed in the laboratory to 25 ± 0.2°C for 2-h and the heat-shock response was assessed in muscle tissue via induction of heat-shock proteins (hsps) 63, 72, 78, and 89. Metabolic measurements included muscle phosphocreatine (PCr), ATP, ADP, and AMP, and hepatic glycogen. Inland and coastal fish overlapped considerably with regard to their hsp responses and energetic endpoints, but hatchery fish were distinct in the biochemical patterns they exhibited. Hsp expression levels after temperature shock were significantly lower in hatchery than in wild fish. Hatchery fish also had significantly lower hepatic glycogen and higher muscle ADP, ATP, and PCr concentrations than wild fish. Coastal and inland steelhead did not differ significantly with regard to peak hsp72 and hsp89 levels or to concentrations of energy metabolites. However, fish from the warm-water, inland site expressed significantly less hsp63, maintained higher basal levels of hsp72, and induced hsp89 more slowly than fish from the cold-water, coastal site. Discriminant function analysis revealed that hatchery fish can be distinguished from wild Navarro River fish with 84.9% certainty using the following function: f(x) = − 43.6 + 0.14(Gly) + 4.1(PCr) + 186.4(AMP) + 80.8(ADP) − 0.14(hsp63) + 0.005(hsp72). This study demonstrates that within a single species, rearing conditions or genetic variation can influence an organism’s disposition and cellular response to thermal stress. Extrapolation of results from laboratory studies on hatchery fish to wild fish may therefore not be possible, and caution must be used when interpreting hsp data obtained for wild fish with different thermal histories.     

Physiological Stress Responses of Cutthroat Trout to Loading by Fish Pump,Conveyor, or Dip Net

The physiological stress response of cutthroat trout, Oncorhynchus clarki, to three methods of loading into truck tanks for transport was evaluated. All loading methods caused a significant decrease in chloride and increase in plasma cortisol levels, relative to the baseline sample of unstressed fish. Based on plasma chloride changes, the conveyor method appeared the least stressful to the fish, while fish pump and dip netting were not significantly different from each other. Differences in plasma cortisol responses suggested that the conveyor and dip net method were less stressful than the fish pump. Plasma glucose concentrations in the fish were not elevated after loading and did not differ among loading methods. Overall, the conveyor method was the least stressful method of loading, but the differences were not sufficient magnitude to preferentially use one method over another.     

Size‐dependent stress response in juvenile Arctic charr (Salvelinus alpinus) under prolonged predator conditioning

Predator conditioning can be used to improve post‐release antipredator recognition of hatchery‐reared salmonids. However, possible negative stress‐related effects of prolonged predator conditioning on juvenile fish physiology are poorly understood. We studied the effects of prolonged (91 days) predator odour exposure on whole‐body cortisol level and spleen size in six full‐sib families of juvenile hatchery‐bred Arctic charr (Salvelinus alpinus). Chemical cues from water containing charr‐fed pikeperch (Sander lucioperca) were used as the predator exposure stimuli and lakewater was used as a chemical control. Our study revealed that juvenile body cortisol levels post‐predator conditioning were affected by treatment, fish size and their interaction. Importantly, among the smaller (i.e. slowest growing) charr, the predator‐exposed fish had higher cortisol levels than control fish, while the opposite pattern was true for the larger fish. These results suggest that chemical cues from charr‐fed predators induce a prolonged stress response in juvenile charr. As prolonged predation exposure seems to elevate stress levels in a size‐dependent manner, the larger, faster growing fish could possibly have intrinsically lower stress responses to predation threats than smaller, slower growing fish. Possible coupling between stress sensitivity and growth requires further attention due to the likely implications for the management of unintended domestication among captive‐reared salmonids.     

Whole-body corticosteroid and plasma cortisol concentrations in larval and juvenile Atlantic cod Gadus morhua L. following acute stress

Methods were developed to assess whole‐body immunoreactive corticosteroid concentrations (IRC) and plasma levels of cortisol in Atlantic cod subjected to several common, acute stressors. A measurable corticosteroid stress response was observed at the first sampling in whole bodies of 8‐day post‐hatch (dph) larvae. Two groups of juveniles (～5 and 30 g) were subjected to a 30 s net stressor and whole‐body IRC and plasma cortisol levels were determined. Post‐stressor IRC in smaller fish rose approximately 14‐fold, peaked at 1 h, were sustained for 3–6 h and returned to pre‐stressor levels within 24 h. Post‐stressor plasma cortisol levels in larger fish rose approximately 18‐fold, peaked at 0.5–1 h, were sustained for 1–3 h and then returned to near pre‐stressor levels after 24 h. Immunoreactive corticosteroid concentrations appeared to remain elevated longer than plasma cortisol levels suggesting that steroids other than cortisol were contributing to total immunoreactivity in homogenates. Juveniles exposed to either a grading procedure or high density transport had maximal IRC and plasma cortisol levels within 90 min which returned to pre‐stressor levels within 24–72 h. A reduction in water salinity (20 g L^?1) did not moderate the corticosteroid response in juveniles. The results show that Atlantic cod respond to common, acute stressors in a manner similar to other teleosts. Whole‐body homogenates can be used to identify changes in IRC in response to acute stress in cod with the caveat that recovery IRC may differ from plasma cortisol concentrations.     

Effects of iodized feed on stress modulation in steelhead trout, Oncorhynchus mykiss (Walbaum)

In the aquaculture industry, the physiological systems of fish can be chronically stressed by various biological, chemical and physical factors. Chronic stress leads to a decrease in the overall health and growth of the fish, making them more prone to diseases. Dietary iodine has been shown to reduce this stress response in chickens and increase disease resistance in dairy cattle, but the mechanisms by which iodine affects stress resistance and immunocompetency is not completely understood and has not been extensively studied in fish. This study investigated the effects of iodized feed as a nutritional supplement in relation to stress modulation in steelhead trout (Oncorhynchus mykiss). We investigated effects on primary, secondary and tertiary stress responses by measuring plasma levels of cortisol, glucose and thyroid hormones as well as haematocrit percentage and average growth rate in steelhead trout. Iodine‐supplemented fish, on average, had lower levels of plasma cortisol and glucose and lower packed cell volumes than fish fed with regular commercial feed (P<0.05). Iodine‐supplemented fish also showed higher levels of thyroid hormones and exhibited better growth over the period of the experiment (P<0.05). It was concluded that, used with other husbandry practices, use of iodine‐supplemented feed could lead to better host defence, growth and survival in fish raised in the aquaculture industry.     

Effects of fasting on metabolic and immunological responses of common carp (Cyprinus carpio) to a further acute stress

Common carp were fasted for 0, 3 and 14 days prior to exposure to a 2‐hr crowding stress. Serum cortisol, glucose, lactate, _{T4, T3}, serum alternative complement (ACH50), lysozyme, total immunoglobulin (Ig) and blood WBC were determined before and 0, 6 and 24 hr after the crowding stress. Three days of fasting led to a significant decrease in serum levels of lactate and thyroid hormones. Fourteen days of fasting led to initiation of physiological process to maintain serum glucose by increase in serum cortisol level and lactate utilization. These fish had lower levels of thyroid hormones suggesting suppression of basal metabolism. The fish also had lower WBC and serum lysozyme and total Ig levels indicating immunosuppression. After the crowding stress, the fish fasted for 3 days had significantly lower lactate, thyroid hormones and total Ig levels. The fish fasted for 14 days had higher cortisol but lower glucose and thyroid hormones levels suggesting impairment in metabolic responses to the crowding stress. Immunological and health indicators including WBC, and ACH50, lysozyme and total Ig of these fish were significantly lower than the other groups after the crowding stress. Overall, the results show that a 14‐day fasting leads to stress responses, reduction in metabolism and immunosuppression. Moreover, 14 days of fasting lead to greater cortisol response to a further crowding stress, which induces marked immunosuppression and fallen welfare.     

Salt as a stress response mitigator of matrinxã, Brycon cephalus (Günther), during transport

The present study evaluated the physiological responses of matrinxã, Brycon cephalus (Günther) submitted to transport stress under the influence of sodium chloride. Different salt concentrations (0.0%, 0.1%, 0.3% and 0.6%) were added to four 200-L plastic tanks. Each tank was stocked with 30 fish (mean weight 1.0 ± 0.2 kg) and transported for 4 h. Blood was sampled prior to transport and immediately after and 24 and 96 h after transport. Plasma cortisol and glucose and serum sodium and potassium, plasma chloride and ammonia were analysed. Changes in plasma cortisol were observed immediately after transportation, except in fish transported in 0.3% and 0.6% salt. Twenty-four hours later, this hormone had returned to its initial level in all fish. Blood glucose was not changed in fish treated with 0.6% salt immediately after transport, and returned to the initial level within 96 h after the other treatments. All treatments resulted in lower levels of plasma chloride after transport, except for fish treated with 0.6% salt, with fish treated with 0.0% and 0.3% salt recovering 24 h later. Serum sodium decreased immediately after transport only in the control fish, returning to the initial level 24 h later. The results indicate that treatment with 0.6% NaCl reduces most of the physiological responses of matrinxã to the stress of transport.     

Cortisol response of golden shiners (Notemigonus crysoleucas) fed diets differing in lipid content

The purpose of this study was to characterize the plasma cortisol response in golden shiners (Notemigonus crysoleucas) under crowding stress, and to determine whether dietary lipid affects the cortisol response. The main differences between the diets were the relative amounts and types of fatty acids of the n-3 and n-6 families. In experiment 1, triplicate groups of golden shiners were fed diets with 4 or 13% menhaden fish oil, or 4 or 13% poultry fat at 3% body weight for 4 weeks before the crowding stress. In experiment 2, fish were fed diets with 8% lipid from soybean oil (SBO), cod liver oil (CLO), (SBO+CLO), or cottonseed oil (CSO) for 6 weeks before the stress test. Plasma cortisol was measured initially, two hours after the stressor was applied, and two hours after the stressor was removed. The overall pattern of the cortisol response was similar to that seen in other warmwater fishes, but the rate and magnitude of the response were not affected by diet in either experiment. Golden shiners have an intermediate cortisol response compared to other fish that have been exposed to similarly stressful conditions.