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The objective of this study was to investigate the role of palmitoylethanolamide (PEA) in the regulation of energy homeostasis in goldfish (Carassius auratus). We examined the effects of acute or chronic intraperitoneal treatment with PEA (20 μg·g -1 body weight) on parameters related to food intake and its regulatory mechanisms, locomotor activity, glucose and lipid metabolism, and the possible involvement of transcription factors and clock genes on metabolic changes in the liver. Acute PEA treatment induced a decrease in food intake at 6 and 8 h post-injection, comparable to that observed in mammals. This PEA anorectic effect in goldfish could be mediated through interactions with leptin and NPY, as PEA increased hepatic expression of leptin aI and reduced hypothalamic expression of npy. The PEA chronic treatment reduced weight gain, growth rate, and locomotor activity. The rise in glycolytic potential together with the increased potential of glucose to be transported into liver suggests an enhanced use of glucose in the liver after PEA treatment. In addition, part of glucose may be exported to be used in other tissues. The activity of fatty acid synthase (FAS) increased after chronic PEA treatment, suggesting an increase in the hepatic lipogenic capacity, in contrast with the mammalian model. Such lipogenic increment could be linked with the PEA-induction of REV-ERBα and BMAL1 found after the chronic treatment. As a whole, the present study shows the actions of PEA in several compartments related to energy homeostasis and feeding behavior, supporting a regulatory role for this N-acylethanolamine in fish., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Hypothalamic AMPK plays a major role in the regulation of whole body metabolism and energy balance. Present evidence has demonstrated that this canonical mechanism is evolutionarily conserved. Thus, recent data demonstrated that inhibition of AMPKα2 in fish hypothalamus led to decreased food intake and liver capacity to use and synthesize glucose, lipids, and amino acids. We hypothesize that a signal of abundance of nutrients from the hypothalamus controls hepatic metabolism. The vagus nerve is the most important link between the brain and the liver. We therefore examined in the present study whether surgical transection of the vagus nerve in rainbow trout is sufficient to alter the effect in liver of central inhibition of AMPKα2. Thus, we vagotomized (VGX) or not (Sham) rainbow trout and then intracerebroventricularly administered adenoviral vectors tagged with green fluorescent protein alone or linked to a dominant negative isoform of AMPKα2. The inhibition of AMPKα2 led to reduced food intake in parallel with changes in the mRNA abundance of hypothalamic neuropeptides [neuropeptide Y ( npy ), agouti-related protein 1 ( agrp1 ), and cocaine- and amphetamine-related transcript ( cartpt )] involved in food intake regulation. Central inhibition of AMPKα2 resulted in the liver having decreased capacity to use and synthesize glucose, lipids, and amino acids. Notably, these effects mostly disappeared in VGX fish. These results support the idea that autonomic nervous system actions mediate the actions of hypothalamic AMPKα2 on liver metabolism. Importantly, this evidence indicates that the well-established role of hypothalamic AMPK in energy balance is a canonical evolutionarily preserved mechanism that is also present in the fish lineage.

There is no evidence in fish brain demonstrating the existence of changes in lactate metabolism in response to alterations in glucose levels. We induced in rainbow trout through intraperitoneal (IP) treatments, hypoglycaemic or hyperglycaemic changes to assess the response of parameters involved in lactate metabolism in glucosensing areas like hypothalamus and hindbrain. To distinguish those effects from those induced by peripheral changes in the levels of metabolites or hormones, we also carried out intracerebroventricular (ICV) treatments with 2-deoxy-D-glucose (2-DG, a non-metabolizable glucose analogue thus inducing local glucopenia) or glucose. Finally, we also incubated hypothalamus and hindbrain in vitro in the presence of increased glucose concentrations. The changes in glucose availability were in general correlated to changes in the amount of lactate in both areas. However, when we assessed in these areas the response of parameters related to lactate metabolism, the results obtained were contradictory. The increase in glucose levels did not produce in general the expected changes in those pathways with only a minor increase in their capacity of lactate production. The decrease in glucose levels was, however, more clearly related to a decreased capacity of the pathways involved in the production and use of lactate, and this was especially evident after ICV treatment with 2-DG in both areas. In conclusion, the present results while addressing the existence of changes in lactate metabolism after inducing changes in glucose levels in brain glucosensing areas only partially support the possible existence of an astrocyte-neuron lactate shuttle in hypothalamus and hindbrain of rainbow trout relating glucose availability to lactate production and use.

In fish food intake is altered under stress conditions, and in a fish teleost model like rainbow trout food intake is associated with the activity of the glucosensor systems. Thus, we aimed to evaluate the possible interaction of stress with the response of glucosensor mechanisms in rainbow trout. Thus, we subjected rainbow trout (via intraperitoneal injections) to normoglycaemic (control), hypoglycaemic (4 mg.kg(-1) bovine insulin) or hyperglycaemic (500 mg.kg(-1) glucose body mass) conditions for 5 days under normal stocking density (NSD, 10 kg fish mass·m(-3)) or stress conditions induced by high stocking density (HSD, 70 kg fish mass·m(-3)). The experimental design was appropriate since hypoglycemia and hyperglycemia were observed in fish under NSD whereas in normoglycaemic fish HSD induced changes in stress-related parameters similar to those reported in fish literature, such as increased levels of cortisol and glucose in plasma and decreased levels of glycogen in liver. Food intake did not respond to changes in plasma glucose levels in fish under HSD conditions, in contrast with the decreased food intake observed when glucose levels increased in fish under NSD conditions. Moreover, the changes with the increase in plasma glucose levels in parameters involved in glucosensing in liver, Brockmann bodies (BB), hypothalamus, and hindbrain of fish in NSD either disappeared (DHAP and GAP levels, and GK, PK, and GPase activities in liver; glucose, DHAP and GAP levels in BB; glucose and DHAP levels, and GK and PK activities in hypothalamus; glycogen and DHAP levels, and GSase activity in hindbrain) or changed (cortisol levels in plasma; glycogen and GAP levels, and GSase and FBPase activities in liver; GK and PK activities in BB; GK and PK activities in hindbrain) in fish under HSD. Those changes suggest for the first time in fish the existence of an interaction between glucosensing capacity and stress. The readjustment in the activity of glucosensor systems is also associated with changes in food intake resulting in an inability of the fish to compensate with changes in food intake those of circulating glucose levels as observed in fish under non-stressed conditions., (Copyright © 2010 Elsevier Inc. All rights reserved.)

We hypothesize that variations in dietary carbohydrate levels produce changes in glucosensor parameters in previously characterized glucosensing areas (hypothalamus and hindbrain) related with the regulation of food intake of a carnivorous fish species like rainbow trout. Therefore, we fed trout with standard, carbohydrate-free (CF) or high-carbohydrate (HC) diets for 10 days to assess changes in glucosensing system and food intake. Fish fed CF diet displayed hypoglycemia and increased food intake. Fish fed a HC diet displayed hyperglycemia and decreased food intake. Changes in food intake due to dietary carbohydrates were accompanied in hypothalamus and hindbrain of fish fed with HC diet by changes in parameters involved in glucosensing, such as increased glucose, glucose 6-phosphate, and glycogen levels and increased glucokinase (GK), glycogen synthase, and pyruvate kinase activities as well as increased GK and GLUT2 expression. All those results address for the first time in fish, despite the relative intolerance to glucose of carnivorous species, that dietary carbohydrates are important regulators of the glucosensing system in carnivorous fish, suggesting that the information generated by this system can be associated with the changes observed in food intake.

We assessed the daily patterns of parameters involved in energy metabolism in liver, white muscle, and gills of rainbow trout. Where daily rhythms were found, we analyzed the potential influence of feeding. Immature rainbow trout were randomly distributed in 3 groups: fish fed for 7 days, fish fasted for 7 days, and fish fasted for 7 days and refed for 4 days. On sampling day, fish of fed and refed groups were fed at 11.00 h, and all fish were sampled from each treatment group using the following time schedule: 14.00, 18.00, 21.00, 00.00, 04.00, 07.00, 10.00 and 14.00 h. The results obtained from metabolic parameters can be grouped into four different categories, such as i) those displaying no daily changes in any group assessed in liver (acetoacetate and lactate levels), white muscle (protein levels, and low Km (glucose) hexokinase (HK) and HK-IV activities) and gills (protein levels), ii) those displaying no 24 h changes in fed fish but in refed or fasted fish in liver (glucose, glycogen, amino acid and protein levels, and HK-IV activity), white muscle (glycogen and amino acid levels) and gills (glucose levels), iii) those displaying 24 h changes that were apparently dependent on feeding since they disappear in fasted fish in liver (Low Km (glucose) HK, lactate dehydrogenase (LDH-O), glucose 6-phosphatase (G6Pase), fructose 1,6-bisphosphatase (FBPase) , alpha-glycerophosphate dehydrogenase (G3PDH), glutamate dehydrogenase (GDH) and aspartate aminotransferase (Asp-AT) activities), white muscle (glucose levels, and pyruvate kinase (PK), LDH-O, G3PDH and Asp-AT activities) and gills (glycogen and lactate levels, and Low Km (glucose) HK, HK-IV, LDH-O and Asp-AT activities), and iv) those parameters displaying 24 h changes apparently not dependent on feeding in liver (lactate levels and PK activity) and gills (amino acid levels, and PK and GDH activities). In general, most 24 h changes observed were dependent on feeding and can be also related to daily changes in activity.

We assessed the daily patterns of parameters involved in energy metabolism in plasma and brain of rainbow trout. Where daily rhythms were found, we analyzed the potential influence of feeding. Immature rainbow trout were randomly distributed in 3 groups: fish fed for 7 days, fish fasted for 7 days, and fish fasted for 7 days and refed for 4 days. On sampling day, fish of fed and refed groups were fed at 11.00 h, and all fish were sampled from each treatment group using the following time schedule: 14.00, 18.00, 21.00, 00.00, 04.00, 07.00, 10.00 and 14.00 h. The results obtained from metabolic parameters assessed in plasma and brain can be grouped into three different categories, such as (i) those displaying no 24 h changes in fed fish such as plasma lactate, protein or acetoacetate levels, as well as brain amino acid and protein levels, and lowKm(glucose) hexokinase, and aspartate aminotransferase activities, (ii) those displaying 24 h changes that were apparently dependent on feeding since they disappeared in fasted fish such as the case of plasma cortisol, glucose and triglyceride levels, as well as brain glycogen, glucose, and lactate levels, and pyruvate kinase and hexokinase IV activities, and (iii) those parameters displaying 24 h changes apparently not dependent on feeding such as plasma amino acids, brain acetoacetate levels as well as several enzyme activities measured in brain such as glucose 6-phosphate dehydrogenase, alpha-glycerophosphate dehydrogenase, glutamate dehydrogenase, and lactate dehydrogenase-oxidase. In general, 24 h changes dependent on feeding indicate an increased use of glucose in brain several hours post-feeding whereas those changes not dependent on feeding were characterized by reduced levels/activity at the night period suggesting a metabolic depression in brain during darkness.

The osmoregulatory and metabolic role of testosterone (T) in the euryhaline teleost Sparus auratus was examined. Fish were implanted with a slow-release coconut oil implant alone (control) or containing T (2 or 5microgg(-1) body weight) and sampled 1, 3, and 7 days after implantation. Gill Na(+),K(+)-ATPase activity increased in fish treated with the lower dose of T after 7 days of treatment. Kidney Na(+),K(+)-ATPase activity enhanced at first day post-implantation in the group treated with the higher dose of T but the values diminished by day 3. Plasma levels of metabolites (glucose, lactate, triglyceride, and protein) increased after T treatment. This higher availability of plasma metabolites was reflected in several metabolic changes within different tissues of T-treated fish such as (i) increased glycogen levels and capacity for gluconeogenesis, ketogenesis, glucose exporting, and amino acid catabolism in the liver, (ii) enhanced lipogenic capacity in the gills, (iii) increased glycogen levels and capacity for oxidizing amino acids in the kidney, and (iv) enhanced levels of glycogen, aceotacetate, glucose and triglycerides, and higher capacity of phosphorylating glucose in the brain. These results provide evidence regarding an osmoregulatory and metabolic role for T in S. auratus that could be related to changes in both processes during sexual maturation.

The gilthead sea bream (Sparus auratus) is an euryhaline fish where prolactin (PRL) and growth hormone (GH) play a role in the adaptation to different environmental salinities. To find out the role of these pituitary hormones in osmoregulation and energy metabolism, fish were implanted with slow release implants of ovine GH (oGH, 5 microg g(-1) body mass) or ovine prolactin (oPRL, 5 microg g(-1) body mass), and sampled 7 days after the start of the treatment. GH increased branchial Na(+),K(+)-ATPase activity and decreased sodium levels in line with its predicted hypoosmoregulatory action. GH had metabolic effects as indicated by lowered plasma protein and lactate levels, while glucose, triglycerides and plasma cortisol levels were not affected. Also, GH changed liver glucose and lipid metabolism, stimulated branchial and renal glucose metabolism and glycolytic activity, and enhanced glycogenolysis in brain. PRL induced hypernatremia. Furthermore, this hormone decreased liver lipid oxidation potential, and increased glucose availability in kidney and brain. Both hormones have opposite osmoregulatory effects and different metabolic effects. These metabolic changes may support a role for both hormones in the control of energy metabolism in fish that could be related to the metabolic changes occurring during osmotic acclimation.

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.

Changes in different osmoregulatory and metabolic parameters over time were assessed in gills, kidney, liver and brain of gilthead sea bream Sparus auratus transferred either from seawater (SW, 38 p.p.t.) to hypersaline water (HSW, 55 p.p.t.) or from SW to low salinity water (LSW, 6 p.p.t.) for 14 days. Changes displayed by osmoregulatory parameters revealed two stages during hyperosmotic and hypo-osmotic acclimation: (i) an adaptive period during the first days of acclimation (1-3 days), with important changes in these parameters, and (ii) a chronic regulatory period (after 3 days of transfer) where osmotic parameters reached homeostasis. From a metabolic point of view, two clear phases can also be distinguished during acclimation to hyperosmotic or hypo-osmotic conditions. The first one coincides with the adaptive period and is characterized by enhanced levels of plasma metabolites (glucose, lactate, triglycerides and protein), and use of these metabolites by different tissues in processes directly or indirectly involved in osmoregulatory work. The second stage coincides with the chronic regulatory period observed for the osmoregulatory parameters and is metabolically characterized in HSW-transferred fish by lower energy expenditure and a readjustment of metabolic parameters to levels returning to normality, indicative of reduced osmoregulatory work in this stage. In LSW-transferred fish, major changes in the second stage include: (i) decreased glycolytic potential, capacity for exporting glucose and potential for amino acid catabolism in liver; (ii) enhanced use of exogenous glucose through glycolysis, pentose phosphate and glycogenesis in gills; (iii) increased glycolytic potential in kidney; and (iv) increased glycogenolytic potential and capacity for use of exogenous glucose in brain.

The impact of different environmental salinities on the energy metabolism of gills, kidney, liver, and brain was assessed in gilthead sea bream (Sparus aurata) acclimated to brackish water [BW, 12 parts/thousand (ppt)], seawater (SW, 38 ppt) and hyper saline water (HSW, 55 ppt) for 14 days. Plasma osmolality and levels of sodium and chloride presented a clear direct relationship with environmental salinities. A general activation of energy metabolism was observed under different osmotic conditions. In liver, an enhancement of glycogenolytic and glycolytic potential was observed in fish acclimated to BW and HSW compared with those in SW. In plasma, an increased availability of glucose, lactate, and protein was observed in parallel with the increase in salinity. In gills, an increased Na+-K+-ATPase activity, a clear decrease in the capacity for use of exogenous glucose and the pentose phosphate pathway, as well as an increased glycolytic potential were observed in parallel with the increased salinity. In kidney, Na+-K+-ATPase activity and lactate levels increased in HSW, whereas the capacity for the use of exogenous glucose decreased in BW- and HSW- acclimated fish compared with SW-acclimated fish. In brain, fish acclimated to BW or HSW displayed an enhancement in their potential for glycogenolysis, use of exogenous glucose, and glycolysis compared with SW-acclimated fish. Also in brain, lactate and ATP levels decreased in parallel with the increase in salinity. The data are discussed in the context of energy expenditure associated with osmotic acclimation to different environmental salinities in fish euryhaline species.

To assess the role of noradrenaline (NA) as a possible regulator of brain energy metabolism in teleost fish, the impact of increased noradrenaline levels within the brain on several parameters of energy metabolism was assessed in rainbow trout brain. Accordingly, two different doses of noradrenaline, producing increases in brain NA levels comparable to those occurring in several physiological processes in nature, were selected. In a subsequent set of three different experiments, fish were intracerebroventricularly injected with 1 microL 100 g(-1) body weight of Cortland saline alone (control) or containing NA (5 nmol NA and 10 nmol NA); after 30 min, brain and plasma samples were taken to assess changes in parameters of energy metabolism due to NA treatment. The results obtained clearly show dose-dependent changes in NA-treated fish in several parameters, including decreased glycogen and ATP levels, increased lactate and pyruvate levels, decreased fructose 1,6-bisphosphatase activity, and increased pyruvate kinase and lactate dehydrogenase activities. Altogether, the present experiments show for the first time in a teleost fish evidence supporting that increased noradrenaline levels in the brain elicit metabolic changes in the brain (enhanced glycogenolysis and glycolysis), resulting in an increased energy demand. These metabolic changes may be related to those occurring under several physiological conditions in nature such as hypoxia, in which increased energy demand and increased noradrenaline levels occur in the brain simultaneously.

Gilthead seabream Sparus aurata were injected intraperitoneally with slow-release implants of coconut oil alone or containing cortisol (50 and 100 microg x g(-1) body weight), and sampled after two, five, and seven days to assess the simultaneous effects of cortisol on both osmoregulation and energy metabolism. Plasma cortisol levels increased in treated fish to 50-70 ng x ml(-1). An enhanced hypoosmoregulatory capacity of cortisol-implanted fish is suggested by the increase observed in gill Na+, K+-ATPase activity, and the decrease observed in plasma ion concentration (Na+ and Cl-) and osmolality. Cortisol also elicited metabolic changes in liver (increased gluconeogenic potential suggested by elevated FBPase activity, and decreased potential of glycolysis and pentose-phosphate shunt, suggested by the decreased activities of both PK and G6PDH) supporting changes in levels of plasma metabolites suitable for use in other tissues. Thus in this study, we demonstrate for the first time in fish that cortisol treatments elicit changes in the use of exogenous glucose in gills (decreased HK activity) and an increased glycolytic and glycogenic potential in brain (increased GPase, PK and PFK activities)., (Copyright 2003 Wiley-Liss, Inc.)

This review focuses on recent research on the metabolic function of fish brain. Fish brain is isolated from the systemic circulation by a blood-brain barrier that allows the transport of glucose, monocarboxylates and amino acids. The limited information available in fishes suggests that oxidation of exogenous glucose and oxidative phosphorylation provide most of the ATP required for brain function in teleosts, whereas oxidation of ketones and amino acids occurs preferentially in elasmobranchs. In several agnathans and benthic teleosts brain glycogen levels rather than exogenous glucose may be the proximate glucose source for oxidation. In situations when glucose is in limited supply, teleost brains utilize other fuels such as lactate or ketones. Information on use of lipids and amino acids as fuels in fish brain is scarce. The main pathways of brain energy metabolism are changed by several effectors. Thus, several parameters of brain energy metabolism have been demonstrated to change post-prandially in teleostean fishes. The absence of food in teleosts elicits profound changes in brain energy metabolism (increased glycogenolysis and use of ketones) in a way similar to that demonstrated in mammals though delayed in time. Environmental factors induce changes in brain energy parameters in teleosts such as the enhancement of glycogenolysis elicited by pollutants, increased capacity for anaerobic glycolysis under hypoxia/anoxia or changes in substrate utilization elicited by adaptation to cold. Furthermore, several studies demonstrate effects of melatonin, insulin, glucagon, GLP-1, cortisol or catecholamines on energy parameters of teleost brain, although in most cases the results are quite preliminary being difficult to relate the effects of those hormones to physiological situations. The few studies performed with the different cell types available in the nervous system of fish allow us to hypothesize few functional relationships among those cells. Future research perspectives are also outlined.

We hypothesize that a decrease in circulating levels of fatty acid (FA) in rainbow trout Oncorhynchus mykiss would result in the inhibition of putative hypothalamic FA sensing systems with concomitant changes in the expression of orexigenic and anorexigenic factors ultimately leading to a stimulation of food intake. To assess this hypothesis, we lowered circulating FA levels treating fish with SDZ WAG 994 (SDZ), a selective A1 adenosine receptor agonist that inhibits lipolysis. In additional groups, we also evaluated if the presence of intralipid was able to counteract changes induced by SDZ treatment, and the possible involvement of the hypothalamus-pituitary-interrenal (HPI) axis by treating fish with SDZ in the presence of metyrapone, which decreases cortisol synthesis in fish. The decrease in circulating levels of FA in rainbow trout induced a clear increase in food intake that was associated with the decrease of the anorexigenic potential in hypothalamus (decreased POMC-A1 and CART mRNA abundance), and with changes in several parameters related to putative FA-sensing mechanisms in hypothalamus. Intralipid treatment counteracted these changes. SDZ treatment also induced increased cortisol levels and the activation of different components of the HPI axis whereas these changes disappeared in the presence of intralipid or metyrapone. These results suggest that the HPI axis is involved in a counter-regulatory response in rainbow trout to restore FA levels in plasma. [ABSTRACT FROM AUTHOR]

In a previous study, we provided evidence for the presence in hypothalamus and Brockmann bodies (BB) of rainbow trout Oncorhynchus mykiss of sensing systems responding to changes in levels of oleic acid (long-chain fatty acid, LCFA) or octanoic acid (medium-chain fatty acid, MCFA). Since those effects could be attributed to an indirect effect, in the present study, we evaluated in vitro if hypothalamus and BB respond to changes in FA in a way similar to that observed in vivo. In a first set of experiments, we evaluated in hypothalamus and BB exposed to increased oleic acic or octanoic acid concentrations changes in parameters related to FA metabolism, FA transport, nuclear receptors and transcription factors, reactive oxygen species (ROS) effectors, components of the KATP channel, and (in hypothalamus) neuropeptides related to food intake. In a second set of experiments, we evaluated in hypothalamus the response of those parameters to oleic acid or octanoic acid in the presence of inhibitors of fatty acid sensing components. The responses observed in vitro in hypothalamus are comparable to those previously observed in vivo and specific inhibitors counteracted in many cases the effects of FA. These results support the capacity of rainbow trout hypothalamus to directly sense changes in MCFA or LCFA levels. In BB increased concentrations of oleic acid or octanoic acid induced changes that in general were comparable to those observed in hypothalamus supporting direct FA sensing in this tissue. However, those changes were not coincident with those observed in vivo allowing us to suggest that the FA sensing capacity of BB previously characterized in vivo is influenced by other neuroendocrine systems. [ABSTRACT FROM AUTHOR]

Abstract: To assess the effects of the polycyclic aromatic hydrocarbons (PAHs) β-naphthoflavone (β-NF) and benzo(a)pyrene (BaP) on liver intermediary metabolism and plasma steroid hormones, immature female rainbow trout (Oncorhynchus mykiss) were intraperitoneally injected (2μlg−1) with vegetable oil alone (control) or containing β-NF or BaP (10mgkg−1) and returned to their tanks; 3, 24, and 72h after injection, 11 fish were sampled from each group. On each sampling time, plasma hormone levels (cortisol and 17β-estradiol) and metabolic parameters in plasma (glucose, lactate, and α-amino acid levels) and liver (glycogen, glucose, lactate, and α-amino acid levels, and HK, GK, PK, LDH, G6Pase, G6PDH, FBPase, GDH, Asp-AT, and HOAD activities) were assessed. Changes described for hormonal systems resulted in an increase in plasma levels of cortisol after 24 and 72h of treatment with both PAHs whereas no changes were noticed for 17β-estradiol levels. Changes in intermediary metabolism described effects in several pathways due to treatment with both PAHs. These changes can be summarized as increased glucose and lactate levels in plasma, and increased glycogenolysis and gluconeogenesis in liver after 24 and 72h of treatment with both PAHs. Furthermore, β-NF treatment stimulated amino acid catabolism in liver. These metabolic changes can be associated with increased levels of plasma cortisol, and suggest a different metabolic behavior depending on PAHs. [Copyright &y& Elsevier]

Abstract: To assess the effects of naphthalene on liver intermediary metabolism and plasma steroid hormones, immature female rainbow trout (Oncorhynchus mykiss), in a first experiment, were intraperitoneally injected (2μLg−1) with vegetable oil alone (control) or containing naphthalene (10 and 50mgkg−1) and returned to their tanks. At 1, 3, and 6h after injection, eight fish were sampled from each group. A second experiment was similarly designed but used fish intraperitoneally implanted (10μLg−1) with slow-release coconut oil implants alone (control) or containing naphthalene at doses of 10 and 50mgkg−1 body weight that were sampled 1, 3, and 5 days after injection. At each sampling time, plasma hormone levels (cortisol and 17β-estradiol) and metabolic parameters in plasma (glucose and lactate) and liver (glucose, lactate, and glycogen levels and HK, GK, GPase, GDH, FBPase, and PK activities) were assessed. Changes described for both hormonal systems resulted in a decrease in plasma levels of cortisol and 17β-estradiol. Changes observed in intermediary metabolism described effects in several pathways of liver energy metabolism due to naphthalene. These changes can be summarized as increased glycogenolysis, use of exogenous glucose, and glycolysis and decreased gluconeogenesis. The increased energy production in liver suggested by these changes can be related to the increased detoxification activity known to occcur in liver after PAH exposure, and can be also related directly or indirectly to the changes observed in the levels of plasma steroids. [Copyright &y& Elsevier]

Animals can habituate to certain repeated stressors and reduce the physiological response that such stressor evoked initially. Studies related to stress habituation in fish are scarce and the available data differ depending on the species and on the type, duration and severity of the stressor. The main objective of this study was to investigate the stress response of juvenile Senegalese sole ( Solea senegalensis ) submitted to repeated chasing stress for 3 days previous to the experiment in order to evaluate the occurrence of habituation to those stress conditions in this fish species. Thus, five different experimental groups were evaluated: not stressed fish (control, C), fish stressed only on the experimental day (ST/naïve), and fish stressed on the experimental day and on the 3 previous days: during the day (ST/Dt), at night (ST/Nt) or both (ST/Dt + Nt). Parameters related to primary and secondary responses to chasing were evaluated in plasma, liver and brain. Chasing in ST/naïve group induced incremented values of plasma cortisol, glucose and lactate but no changes in catecholamine levels compared to controls. In trained fish, higher cortisol but decreased glucose, lactate and catecholamine levels were observed after stress compared to controls and to ST/naïve groups. In the liver, stress did not induce any changes with respect to controls whereas ST/Dt and ST/Dt + Nt showed lower values of glucose and glycogen than stressed naïve fish. In the brain, ST/naïve group presented no significant changes in serotonergic activity. However, incremented serotonergic activity was detected in fish previously trained. Furthermore, dopaminergic activity decreased in diurnal trained and nocturnal trained groups with respect to ST/naïve fish. Crh expression in hypothalamus was higher in ST/naïve fish but not in fish submitted to repeated stress compared to controls. In summary, it seems that there was no habituation to the repeated acute stress protocol in Solea senegalensis in terms of serotonergic activity and cortisol release during the physiological stress response. However, the decreased levels of plasma catecholamines and energy metabolites, and of the hypothalamic crh mRNA abundance and dopaminergic activity, indicate a modulation of the stress response in trained fish. Altogether, the results suggest that either the chasing stressor was too strong or the training period too short for the animals to habituate, indicating that repeated chasing within short periods should be avoided when manipulating fish in order to keep proper welfare conditions in this species. [ABSTRACT FROM AUTHOR]

Abstract: To elucidate the short-term time-course of liver metabolic response in rainbow trout to acute handling stress we subjected rainbow trout to 5min chasing and obtained samples 0 to 480min post-stress. Levels of cortisol, glucose and lactate were measured in plasma, whereas metabolite levels, enzyme activities, mRNA abundance of parameters related to energy metabolism, and glucocorticoid receptors were assessed in liver. Acute stress affected many parameters related to energy metabolism, with most of them turning back to normal levels after 480min. In general, the present results support the existence of two stages in the short-term time-course of metabolic response to handling stress. A first stage occurring few minutes post-stress (15–45min), was characterized by increased mobilization of liver glycogen resulting in increased production of endogenous glucose, reduced use of exogenous glucose and reduced lipogenic potential. A second stage, occurring 60–120min post-stress onwards was characterized by the recovery of liver glycogen levels, the increased capacity of liver for releasing glucose, and the recovery of lipogenic capacity whereas no changes were noted in gluconeogenic potential, which probably needs longer time periods to become enhanced. [Copyright &y& Elsevier]

Abstract: The aim of the present study was to obtain evidence for the possible modulatory effect of leptin on glucosensing capacity in hypothalamus and hindbrain of rainbow trout. In a first experiment, trout were injected ICV with saline alone or containing increased doses of leptin (0.3–30μgμl−1). Leptin induced in general in both hypothalamus and hindbrain dose-dependent changes in parameters related to glucosensing (increased glycogenic and glycolytic potentials together with increased GK activity, and increased mRNA levels of genes involved in glucosensing response) compatible with those occurring under hyperglycemic conditions, a situation that is known to produce anorexia. The anorectic action of leptin in our experimental conditions was observed in a second experiment. The specificity of leptin action was tested in a third experiment in which trout were injected ICV with saline, or leptin alone, or leptin plus agents known to inhibit leptin signaling pathways in mammals. The results obtained suggest that the central action of leptin on glucosensing system can be related to the JAK/STAT and IRS-PI(3)K pathways. Finally, we also provide evidence for a peripheral effect of central leptin treatment (increased liver glycogenolytic potential), which could be associated with increased sympathetic activity. [Copyright &y& Elsevier]

Abstract: The gilthead sea bream Sparus aurata is a euryhaline and euritherm species with the capacity of living under different environmental conditions of salinity and temperature. The influence of acclimation to different environmental salinities (5, 38 and 55‰) and temperatures (12°, 19° and 26°C) for seven weeks was analyzed in plasma and tissues (liver, gills and kidney) of gilthead sea bream assessing levels of metabolites and enzyme activities related to energy metabolism. Changes observed in specimens acclimated to different environmental salinities agree with previous results reported for this species. The temperature alone did also affect metabolic parameters in a way similar to that previously described. A significant interaction of salinity with temperature was found in most parameters assessed in tissues suggesting that the metabolic effects of salinity are different depending on the temperature of acclimation. The interactions were different among tissues and parameters displaying different patterns of changes. In general, the acclimation to extreme temperatures (especially low) alters the metabolic responses to different salinities thus suggesting that the energy demand of increased osmoregulatory work is not so important under temperature conditions different from those commonly found in nature and in those used in culture. [Copyright &y& Elsevier]

Abstract: The purpose of the study was to assess in female fish the possible interaction between treatment with a polycyclic aromatic hydrocarbon (PAH) like naphthalene and the onset of vitellogenesis. In a first experiment, female rainbow trout (Oncorhynchus mykiss) at stages 2–3 (previtellogenesis) or 4 (early vitellogenesis) were intraperitoneally injected (2 μl g−1) with vegetable oil alone (control) or containing naphthalene (50 mg kg−1) to be sampled 3 h later. A second experiment was similarly designed but using fish intraperitoneally implanted (10 μl g−1) with slow-release coconut oil implants alone (control) or containing 50 mg naphthalene kg−1 body mass that were sampled 3 days after injection. On each sampling time, plasma levels of cortisol and 17β-estradiol, and several metabolic parameters in plasma, liver and gonad were assessed. In controls, early vitellogenic fish compared with previtellogenic fish displayed changes that in some cases are confirmatory of previous studies whereas in other cases provide new information in plasma (increased amino acid levels), liver (decreased capacity for exporting glucose and reduced amino acid levels) and gonad (decreased amino acid levels). Naphthalene treatment produced in previtellogenic fish decreased 17β-estradiol levels in plasma, increased plasma glucose or decreased liver gluconeogenic capacity whereas no major effects were noticed on parameters involved in lipid, amino acid and lactate metabolism. Differential effects of naphthalene treatment were noticed in early vitellogenic fish such as decreased 17β-estradiol and glucose levels in plasma, increased hexokinase and glucokinase and lack of changes in fructose 1,6-bisphosphatase activities in liver, and a lower decrease of amino acid levels in gonad. Those alterations produced by naphthalene treatment resulted in a decreased capacity for covering the energy demand of vitellogenesis in liver and gonad that could contribute to a delay and/or impairment of the onset of maturation. [Copyright &y& Elsevier]

Abstract: The influence of arginine vasotocin (AVT) on osmoregulation and metabolism in gilthead sea bream Sparus auratus was evaluated by two experimental approaches. In the first, seawater (SW, 36ppt)-acclimatized fish were injected intraperitoneally with vehicle (vegetable oil) or two doses of AVT (0.5 and 1μg/g body weight). Twenty-four hours later, eight fish from each group were sampled; the remaining fish were transferred to low saline water (LSW, 6ppt, hypoosmotic test), SW (transfer control), and hypersaline water (HSW, 55ppt, hyperosmotic test). After another 24h (48-h post-injection), fish were sampled. The only significant effect observed was the increase of sodium levels in AVT-treated fish transferred to HSW. In the second experiment, fish were injected intraperitoneally with slow-release vegetable oil implants (mixture 1:1 of coconut oil and seeds oil) alone or containing AVT (1μg/g body weight). After 3 days, eight fish from each group were sampled; the remaining fish were transferred to LSW, SW, and HSW as above, and sampled 3 days later (i.e. 6 days post-injection). In the AVT-treated group transferred from SW to SW, a significant increase vs. control was observed in gill Na+,K+-ATPase activity. Kidney Na+,K+-ATPase activity decreased in the AVT-treated group transferred to LSW and no changes were observed in the other groups. These osmoregulatory changes suggest a role for AVT during hyperosmotic acclimation based on changes displayed by gill Na+,K+-ATPase activity. AVT treatment increased plasma cortisol levels in fish transferred to LSW and HSW. In addition, AVT treatment affected parameters of carbohydrate, lipid, amino acid, and lactate metabolism in plasma and tissues (gills, kidney, liver, and brain). The most relevant effects were the increased potential of liver for glycogen mobilization and glucose release resulting in increased plasma levels of glucose in AVT-treated fish transferred to LSW and HSW. These changes may be related to the energy repartitioning process occurring during osmotic adaptation of S. auratus to extreme environmental salinities and could be mediated by increased levels of cortisol in plasma. [Copyright &y& Elsevier]

Abstract: The influence of three different environmental salinities (seawater, SW:38 ppt salinity; brackish water, BW:12 ppt; and low salinity water, LSW:6 ppt salinity) on the growth, osmoregulation and metabolism of young gilthead sea bream (Sparus aurata L.) was studied over a period of 100 days. 480 inmature fish (20 g mean body weight) were randomly divided into six tanks of 2500 l (80 fish per tank) and maintained under three different salinities (38 ppt, 12 ppt and 6 ppt) in an open system. Every three weeks, 10 fish from each tank were anesthetized, weighed and lenghed. At the end of experiment, 10 fish from each tank were anesthetized, weighed and sampled for plasma, brain, gill and liver. Gill Na+, K+-ATPase activity, plasma osmolality, ions (sodium and chloride), glucose, lactate, protein and triglyceride, and hepatosomatic index were examined. In addition, levels of glycogen, lactate, ATP and activities of potential regulatory enzymes (hexokinase, pyruvate kinase, glycogen phosphorylase, and glucose 6-phosphate dehydrogenase) were assesed in liver, brain, and gill. BW-acclimated fish showed a better growth with respect to SW- or LSW-acclimated fish (12>38>6 ppt). The same relationship was observed for weight gain and specific growth rate. Osmoregulatory parameters in plasma (osmolality, Na+ and Cl− levels) were similar in SW- and BW-acclimated fish but significantly higher than those of LSW-acclimated fish. Gill Na+, K+-ATPase activity showed lower values in intermediate salinity (6>38>12 ppt). No changes were observed in metabolic parameters analyzed in plasma, whereas only minor changes were observed in metabolic parameters of liver, gills and brain that could be correlated with the higher growth rates observed in fish acclimated to BW, which do not allow us to attribute the best growth rate observed at 12 ppt to lower metabolic rates in that salinity. [Copyright &y& Elsevier]