Relatedness of hypoxia and hyperthermia tolerances in the Nile tilapia (Oreochromis niloticus) and their relationships with cardiac and gill traits.

In fish, thermal and hypoxia tolerances may be functionally related, as suggested by the oxygen- and capacity-limited thermal tolerance (OCLTT) concept, which explains performance failure at high temperatures due to limitations in oxygen delivery. In this study the interrelatedness of hyperthermia and hypoxia tolerances in the Nile tilapia (Oreochromis niloticus), and their links to cardiorespiratory traits were examined. Different groups of O. niloticus (n = 51) were subjected to hypoxia and hyperthermia challenges and the O2 tension for aquatic surface respiration (ASR pO2) and critical thermal maximum (CTmax) were assessed as measurement endpoints. Gill filament length, total filament number, ventricle mass, length and width were also measured. Tolerance to hypoxia, as evidenced by ASR pO2 thresholds of the individual fish, was highly variable and varied between 0.26 and 3.39 kPa. ASR events increased more profoundly as O2 tensions decreased below 2 kPa. The CTmax values recorded for the O. niloticus individuals ranged from 43.1 to 44.8 °C (Mean: 44.2 ± 0.4 °C). Remarkably, there was a highly significant correlation between ASR pO2 and CTmax in O. niloticus (r = -0.76, p < 0.0001) with ASR pO2 increasing linearly with decreasing CTmax. There were, however, no discernible relationships between the measured cardiorespiratory properties and hypoxia or hyperthermia tolerances. The strong relationship between hypoxia and hyperthermia tolerances in this study may be related to the ability of the cardiorespiratory system to provide oxygen to respiring tissues under thermal stress, and thus provides some support for the OCLTT concept in this species, at least at the level of the entire organism.

No additional stress of sublethal gas supersaturation in a landlocked population of Atlantic salmon (Salmo salar) exposed to environmental acidification.

The landlocked Atlantic salmon population "bleke" faces extinction due to environmental acidification (EA) and hydropower expansion in the Norwegian river Otra. Despite of restoration, unexpected mortality has been reported for this population, possibly due to gas bubble trauma (GBT) from gas supersaturation (GSS) downstream of hydroelectric plants, or EA induced aluminum toxicity. In this study, we applied the allostasis concept to investigate interactions between EA and GBT. This concept comprises additive effects of stressors, which can lead to allostatic overload. Stress coping mechanisms become maladaptive in such situations, which can be indicated by an inability to mount a proper cortisol response in fish. Fish were exposed to sublethal levels of simulated EA (SEA), GSS (a total gas pressure; TGP; of 110%) or a combination of these stressors for six days. Effects on allostatic load were subsequently investigated by assessing the cortisol response to an acute stress test. SEA increased cortisol responsiveness and GSS induced clinical signs of GBT, but no interacting effects between GSS and SEA were observed. This suggests that that 110% TGP did not have an additive effect on the allostatic load imposed by SEA.

Air-breathing behavior in Heterotis niloticus fingerlings: Response to changes in oxygen, temperature, and exercise regimes.

Air-breathing in fish is believed to have arisen as an adaptation to aquatic hypoxia. Although air-breathing has been widely studied in numerous fish species, little is known about the obligate air-breathing African bonytongue, Heterotis niloticus. We evaluated if abiotic factors and physical activity affect air-breathing patterns in fingerlings. The air-breathing frequency (fAB ) and behavioral responses of H. niloticus fingerlings were assessed in response to environmental oxygen, temperature, and exhaustion and activity in a series of experiments. The air-breathing behavior of H. niloticus fingerlings under optimum water conditions was characterized by swift excursions lasting less than 1 s to the air-water interface to gulp air. The intervals between air-breaths were highly variable, ranging from 3 to 259 s. Body size only slightly affected fAB , while hypoxia, hyperthermia, and exercise stress significantly increased fAB . Progressive hypoxia from 17.69 to 2.17 kPa caused a ~2.5-fold increase in fAB . Increasing temperatures to 27 and 32°C, from a baseline temperature of 22°C, significantly increased fAB from 0.4 ± 0.2 to 1.3 ± 0.5 and 1.6 ± 0.4 breaths min-1 , respectively. Lastly, following exhaustive exercise, fAB increased up to 3-fold. These observations suggest that H. niloticus fingerlings are very reliant on aerial oxygen, and their air-breathing behavior is sensitive to environmental changes and activity levels.

A novel method to estimate biofilm activity based on enzymatic oxygen release from hydrogen peroxide decomposition.

Biofilm is central for biological water treatment processes in recirculating aquaculture systems (RAS). A lack of suitable methods for quantifying biofilm activity, however, makes it difficult to assess and compare the microbial status of biofilm. This type of information of the biofilm will be useful to assess the colonization status of nitrifying biocarriers or to evaluate the effect of disinfectants on the biofilm activity. Here we introduce a novel assay for rapid assessment of microbial activities in the biofilm attached on bioelements from a RAS biofilter. The assay consisted of an intermittent respirometer platform where biofilter elements were exposed to 10 mg/L hydrogen peroxide (H2O2) for 1 h, following concurrent measurements of oxygen release from the decomposition of H2O2 caused by biofilm-associated enzymes. A different number of colonized, mature bioelements from a moving bed biofilter in a freshwater RAS were tested with repeated H2O2 exposure, and compared against their autoclaved forms. A substantial increase in dissolved oxygen (DO) concentration (0.92-2.31 mg O2/L) occurred with mature bioelements during 1 h of H2O2 exposure, compared to small amounts of DO release (≤0.27 mg O2/L) with autoclaved bioelements. This substantiates that H2O2 decomposition by biofilm is mainly governed by microbial enzymatic activities. A monomolecular model fitted well with the observed oxygen release profiles of tested mature bioelements after H2O2 exposure (R2 > 0.98). The kinetic rate constant of net oxygen release (k or , h-1) was proportional (R2 for linear fit = 0.99) to the number of mature bioelements tested. Repeated exposure of H2O2 to the same bioelements did not change k or , which indicates that 10 mg/L H2O2 with an exposure time of 1 h does not suppress enzymatic activity in biofilm. Our study provides a new rapid method that allows simple quantification of microbial activity in biofilm samples from aquaculture systems, which could potentially be also applied to study biofilm from wastewater treatment plants and other industries.

Combined antagonist treatment of glucocorticoid and mineralocorticoid receptor does not affect weight loss of fasting rainbow trout but inhibits a fasting-induced elevation of cortisol secretion.

The gastrointestinal system of fish reacts rapidly to food deprivation. The relative masses of digestive organs and activities of digestive enzymes decrease within days of fasting. This is believed to be an energy-conserving strategy as the metabolic cost of maintaining digestive capacity is high. Cortisol is known for its role in energy mobilization following stress exposure, and prolonged elevated cortisol levels have been shown to reduce growth rates in fish. Fish experiencing chronic cortisol elevations show structural changes to their digestive tissues and overall reductions in relative digestive tissue masses. In fish fasting for prolonged periods, circulating cortisol levels have been reported to be downregulated, upregulated, or unchanged compared to feeding fish. This study aimed to investigate if RU486 and spironolactone, antagonists of the glucocorticoid receptor (GR), and mineralocorticoid receptor (MR), respectively, alone or in combination affect circulating cortisol levels during prolonged starvation. In addition, we tested the effects of blocking GR and MR, on the down-regulation of relative digestive tissue mass during starvation, and its effects on weight loss. Three treatment groups of rainbow trout were intraperitoneally implanted with either GR, MR, or GR and MR blockers. A fourth group was implanted with cortisol, while a fifth group served as a control. All treatment groups were sampled over a course of four weeks of food deprivation and compared against each other and fed control fish at day 0 of the trial. Starvation for 2 weeks and longer significantly increased circulating cortisol levels in all groups except for the group implanted with GR and MR antagonists. Loss of body mass occurred most rapidly during the first week of starvation. Spironolactone treatment resulted in significantly reduced loss of mass during the first week, however, over the following weeks, no differences in mass loss were observed in the groups implanted with blockers, while cortisol-treated fish showed the highest decrease in body mass over time. Relative digestive tissue mass decreased in all groups but apparently, the fasting-induced elevation in plasma cortisol levels did not affect the relative weight loss of digestive tissues as no differences were observed between control fish and GR + MR antagonist treated fish. Very high cortisol levels caused by cortisol treatment however caused a faster decrease in the relative mass of some digestive organs, particularly the stomach.

Efficacy of H2O2 on the removal kinetics of H2S in saltwater aquaculture systems, and the role of O2 and NO3.

The occurrence of hydrogen sulfide (H2S) represents a challenge for recirculating aquaculture systems (RAS) under saline conditions. Even low concentrations of the toxic gas can result in sudden mass mortalities of fish, leading to large economic losses. There is an urgent need for efficient strategies to remove H2S, which can be applied effectively with a short response time, to prevent the risk of H2S-induced casualties. This study examines the kinetics of the two common oxidants applied to rearing water in a RAS facility; oxygen (O2) and hydrogen peroxide (H2O2) and evaluates their efficiency and applicability for the removal of H2S in an industrial RAS. Furthermore, we tested whether nitrate (NO3-) can be an oxygen donor in the chemical oxidation of H2S. The baseline oxidation rates of H2S by O2 were determined in air-equilibrated seawater (SW) and RAS water (RASW). The feasibility of using H2O2 as a practical treatment was evaluated by testing increasing H2O2 to H2S ratios in SW. In addition, RASW dilutions that yielded different concentrations of NO3- and total chemical oxygen demand (TCOD) were tested to identify their effects on H2S removal. The half-lives (t½) of H2S, derived from O2 oxidation rates, were considerably shorter in SW (118.5 ± 28.6 min) compared to RASW (168.0 ± 18.7 min). The addition of a 1:1 mole ratio of H2O2 to H2S, significantly increased the removal rate and decreased the half-life (t½) of H2S in SW to 29.5 ± 6.6 min. Further increasing H2O2:H2S ratios to 2:1 and 4:1, reduced t½ to 21.7 ± 5.2 and 17.4 ± 6.1 min, respectively. Similarly, a dosage of H2O2 at a ratio of 1:1 in RAS water resulted in a considerably shorter t½ of 86.1 ± 10.1 min. The influence of organic matter on the required H2O2 dose was demonstrated by the t½, which were reduced by 49% in RAS water and 75% in SW. NO3- was not found to be involved in the chemical removal of H2S. The results provide an improved understanding of the influence of RAS water chemistry and quality on H2S kinetics and the direct applicability of the kinetics for treating acute H2S levels in RAS to avoid mass mortalities. In conclusion, the addition of H2O2 is an efficient water treatment technology for H2S removal, and by adjusting H2O2 dosages accordingly to the concentrations of H2S and specific systems water parameters, a t½ <30 min can be achieved. Thus the technology is applicable in an industrial RAS, as a treatment process for acute levels of the hazardous gas H2S that is easily implemented, and safe for the fish.

Capacity for thermal adaptation in Nile tilapia (Oreochromis niloticus): Effects on oxygen uptake and ventilation.

Standard metabolic rate (SMR) and maximum metabolic rate (MMR) were determined for Nile tilapia acclimated to six different experimental temperatures from 18 °C to 38 °C. SMR increased exponentially with temperature, from 79.8 mg O2 kg-1 h-1 at 18 °C, to 255.1 mg O2 kg-1 h-1 at 38 °C (Q10 = 1.79). The main increase in Q10 occurred within the highest temperature range, whereas in the lower temperature from 18 °C to 22 °C, temperature did not significantly affect SMR. MMR showed a hyperbolic correlation with increasing temperature, rising from 240.5 mg O2 kg-1 h-1 at 18 °C to a peak of 373.8 mg O2 kg-1 h-1 at 30 °C, before decreasing again at higher temperatures. Absolute aerobic scope (AAS) peaked at 26.0 °C, which we conclude to be the optimal temperature for Nile tilapia. The optimal temperature range, defined as the thermal range where 80% or more of the metabolic scope (MS) can be maintained, occurred between 19.5 and 32.1 °C. The lower (TCMIN) and upper (TCMAX) critical temperatures occurred at 13.1 °C and 38.8 °C. Nile tilapia showed a 4-fold scope for increasing ventilation frequency from 24 opercular beats min-1 (OB min-1) during SMR at 18 °C, to a maximum of 100 OB min-1 which occurred during MMR at 34 °C. fV during MMR increased with temperature, but above 30 °C became uncoupled with MO2, as fish were unable to sustain their rates of oxygen consumption despite a high fV. There was a strong correlation between fV and SMR (r2 = 0.83) across all temperatures indicating that fV is a good predictor of SMR. However, the correlation between MMR and fV was weak (r2 = 0.06), due to a strong interacting effect of temperature. When selecting data from the thermal optimum range, a good correlation between fV and MO2 was obtained (r2 = 0.74).

Arapaima gigas maintains gas exchange separation in severe aquatic hypoxia but does not suffer branchial oxygen loss.

One of the most air-reliant obligate air-breathing fish is the South American Arapaima gigas, with substantially reduced gills impeding gas diffusion, thought to be a result of recurring aquatic hypoxia in its habitat. In normoxic water, A. gigas is reported to satisfy 70-80% of its O2 requirement from the air while excreting 60-90% of its CO2 to the water. If this pattern of gas exchange were to continue in severely hypoxic water, O2 loss at the gills would be expected. We hypothesized therefore that partitioning of CO2 would shift to the air phase in severe aquatic hypoxia, eliminating the risk of branchial O2 loss. By adapting a respirometer designed to measure aquatic MO2/MCO2, we were able to run intermittent closed respirometry on both water and air phase for both of these gasses as well as sample water for N-waste measurements (ammonia-N, urea-N) so as to calculate metabolic fuel utilization. In contrast to our prediction, we found that partitioning of CO2 excretion changed little between normoxia and severe hypoxia (83% versus 77% aquatic excretion, respectively) and at the same time there was no evidence of branchial O2 loss in hypoxia. This indicates that A. gigas can utilize distinct transfer pathways for O2 and CO2. Routine and standard MO2, N-waste excretion and metabolic fuel utilization did not change with water oxygenation. Metabolism was fuelled mostly by protein oxidation (53%), while carbohydrates and lipids accounted for 27% and 20%, respectively.

Prolonged cortisol elevation alters whole body and tissue metabolism in rainbow trout (Oncorhynchus mykiss).

Chronic elevation of circulating cortisol is known to have deleterious effects on fish, but information about the consequences of prolonged cortisol elevation on the metabolism of fish is scarce. To test the effects of chronic cortisol elevation on the aerobic performance of rainbow trout, we examined how two severities of chronically elevated plasma cortisol levels affected the oxygen uptake during rest and after exhaustive exercise using a high (HC) and a medium cortisol (MC) treatment. High cortisol doses significantly affected standard (SMR) and maximum metabolic rates (MMR) compared to control fish. In comparison, the medium cortisol treatment elevated maximum metabolic rates (MMR) but did not significantly influence SMR compared to a sham group (S) and control group (C). The medium cortisol treatment resulted in a significantly increased metabolic scope due to an elevation of MMR, an effect that was abolished in the HC group due to co-occuring elevations in SMR. The elevated SMR of the HC-treated fish could be explained by increased in vitro oxygen uptake rates (MO2) of specific tissues, indicating that the raised basal metabolism was caused, in part, by an increase in oxygen demand of specific tissues. Haematological results indicated an increased reliance on anaerobic metabolic pathways in cortisol-treated fish under resting conditions.

Hypoxia tolerance and metabolic coping strategies in Oreochromis niloticus.

The Nile tilapia (Oreochromis niloticus) is widely farmed in tropical and subtropical pond culture. O. niloticus is recognized as a species that is tolerant of hypoxic conditions, a trait that may largely be responsible for the success of this species in aquaculture. Until now, neither coping mechanisms nor a comparison of various indices of hypoxia tolerance to characterize the response to hypoxia, have been described. In the present study, Nile tilapia were subjected to hypoxia of increasing severity and duration to examine effects on metabolic rate (MO2) and post hypoxic oxygen debt. MO2 was measured during periods of severe hypoxia at 2.1 kPa O2 (10% oxygen saturation) lasting between 2 and 24 h at 27 °C. Hypoxia tolerance was assessed by determining the critical oxygen tension (Pcrit) and the pO2 at which loss of equilibrium (LOE) occurred. We show that the tolerance of Nile tilapia to severe hypoxia is largely achieved through a capacity for metabolic depression. Despite prolonged exposure to dissolved oxygen levels below Pcrit, the fish showed little excess post-hypoxic oxygen consumption (EPHOC) upon return to normoxic conditions. LOE did not occur until conditions became near-anoxic. Blood pH was not affected by severe hypoxia (2.1 kPa O2), but a significant acidosis occurred during LOE, accompanied by a significant elevation in lactate and glucose levels. The results from the present study indicate that Nile tilapia do not switch to anaerobic metabolism during hypoxia until pO2 falls below 2.1 kPa.

Growth, metabolism and respiration in Nile tilapia (Oreochromis niloticus) exposed to chronic or periodic hypoxia.

Tropical earthen ponds for extensive aquaculture are characterised by daily fluctuations in the availability of dissolved oxygen in the water. Primary production during the daytime ensures excess oxygen availability with oxygen partial pressures (pO2) exceeding 220 mmHg, while nocturnal respiration of fish, plankton and bacteria leads to nightly episodes of severe hypoxia (pO2 < 20 mmHg), often persisting for several hours. To investigate how oxygen availability affects feeding, growth, digestive performance, metabolism and behaviour in Nile tilapia (Oreochromis niloticus), a series of experiments were conducted under different oxygen regimes. To assess growth performance, triplicate groups of fish were held either under constant normoxia (pO2 17.4 ± 0.4 kPa), constant hypoxia (pO2 8.1 ± 0.6 kPa), or diel-cycling between normoxia (pO2 17.1 ± 0.6 kPa from 6 a.m. to 11 p.m.) and severe nocturnal hypoxia (0.4 ± 1.0 kPa from 11 p.m. to 6 a.m.). Chronic hypoxia led to significant affected feed intake and FCR, compared to the normoxic group, whereas nocturnal hypoxia was associated with a compensatory increase in appetite later in the day. Overall, this resulted in a significant increased feed intake compared to the normoxic group. Interestingly, exposure of fish to 6-h nocturnal hypoxia (diel-cycling hypoxia) for 9 weeks resulted in the best growth performance indicators among the treatment groups. Respirometry showed that tilapia respond to nocturnal hypoxia by metabolic depression, allowing them to return to normoxia with a modest oxygen debt. Behavioural observations revealed that aquatic surface respiration is employed when pO2 approaches 2.1 kPa.

Proactive coping style in early emerging rainbow trout carries a metabolic cost with no apparent return.

The timing with which salmonid larvae emerge from their gravel nests is thought to be correlated with a particular suite of behavioural and physiological traits that correspond to the stress coping style of the individual. Among these traits, aggressiveness, dominance and resilience to stress, are potentially interesting to exploit in aquaculture production. In the present study a series of experiments were performed, with the purpose of characterising behavioural, metabolic and production related traits in rainbow trout juveniles from different emergence fractions. Newly hatched rainbow trout were sorted according to their emergence time from an artificial redd. The early, middle, and late fractions were retained and assessed for their physiological response to stress, growth performance, metabolism, fasting tolerance, and potential for compensatory growth. The early emerging fraction showed proactive behavioural traits; they were faster to reappear following startling, showed a reduced cortisol response following stress, and a reduced metabolic cost of recovery. Emergence time was not correlated with any differences in standard or maximum metabolic rates, but was however, correlated with higher routine metabolic rates, as demonstrated by significantly bigger weight losses during fasting in the early emerging group. Growth rates and feed conversion efficiencies were not significantly different when fish were co-habitated under a restrictive feeding regime, suggesting that early emerging fish are not able to monopolise food resources. The intermediate emerging group, which makes up the bulk of a population and is often ignored, appears to possess the best growth performance traits, possibly because they do not expend excessive energy on dominance behaviour such as the early emerging group, while they are also not overly timid or stress prone such as the late emerging group.

Discontinuation of anti-VEGF cancer therapy promotes metastasis through a liver revascularization mechanism.

The impact of discontinuation of anti-VEGF cancer therapy in promoting cancer metastasis is unknown. Here we show discontinuation of anti-VEGF treatment creates a time-window of profound structural changes of liver sinusoidal vasculatures, exhibiting hyper-permeability and enlarged open-pore sizes of the fenestrated endothelium and loss of VE-cadherin. The drug cessation caused highly leaky hepatic vasculatures permit tumour cell intravasation and extravasation. Discontinuation of an anti-VEGF antibody-based drug and sunitinib markedly promotes liver metastasis. Mechanistically, host hepatocyte, but not tumour cell-derived vascular endothelial growth factor (VEGF), is responsible for cancer metastasis. Deletion of hepatocyte VEGF markedly ablates the 'off-drug'-induced metastasis. These findings provide mechanistic insights on anti-VEGF cessation-induced metastasis and raise a new challenge for uninterrupted and sustained antiangiogenic therapy for treatment of human cancers.

Innate immune defenses exhibit circadian rhythmicity and differential temporal sensitivity to a bacterial endotoxin in Nile tilapia (Oreochromis niloticus).

The present study investigated the daily dynamics of humoral immune defenses and the temporal influence in the sensitivity of these responses to a bacterial endotoxin in Nile tilapia (Oreochromis niloticus). The first experiment subjected the fish to two photoperiod conditions, 12L:12D (LD) and 0L:24D (DD), for 20 days to characterize the rhythms of humoral immunity. Serum alkaline phosphatase (ALP), lysozyme (LYZ), peroxidase (PER) and protease (PRO) exhibited significant rhythmicity under LD but not in DD. No significant rhythms were observed in esterase (ESA) and anti-protease (ANTI) in both photoperiod conditions. Fish reared under LD were subsequently subjected to DD while the group previously under DD was exposed to LD, and this carried on for 3 days before another set of samples was collected. Results revealed that the rhythms of LYZ, PER and PRO but not ALP persisted when photoperiod was changed from LD to DD. Nonetheless, immune parameters remained arrhythmic in the group subjected from DD to LD. Cluster analysis of the humoral immune responses under various light conditions revealed that each photic environment had distinct daily immunological profile. In the second experiment, fish were injected with bacterial endotoxin lipopolysaccharide (LPS) either at ZT3 (day) or at ZT15 (night) to evaluate the temporal sensitivity of humoral immunity to a pathogen-associated molecular pattern. The results demonstrated that responses to LPS were gated by the time of day. LPS significantly modulated serum ALP and ANTI activities but only when the endotoxin was administered at ZT3. Serum LYZ and PER were stimulated at both injection times but with differing response profiles. Modulated LYZ activity was persistent when injected at ZT3 but transient when LPS was applied at ZT15. The magnitude of LPS-induced PER activity was higher when the endotoxin was delivered at ZT3 versus ZT15. It was further shown that plasma cortisol was significantly elevated but only when LPS was administered at ZT3. On the other hand, plasma melatonin was significantly affected by LPS injection but only when exposed at ZT15. Taken together, this study shows that several key components of humoral immunity in tilapia exhibit circadian rhythms and adapt to photoperiodic changes. Further, results of the bacterial endotoxin challenge suggest that responsiveness of serum humoral factors to a biological insult is likely mediated by the time of day, highlighting the importance of circadian rhythm in the immunological functions of fish.

Fast-starting after a breath: air-breathing motions are kinematically similar to escape responses in the catfish Hoplosternum littorale.

Fast-starts are brief accelerations commonly observed in fish within the context of predator-prey interactions. In typical C-start escape responses, fish react to a threatening stimulus by bending their body into a C-shape during the first muscle contraction (i.e. stage 1) which provides a sudden acceleration away from the stimulus. Recently, similar C-starts have been recorded in fish aiming at a prey. Little is known about C-starts outside the context of predator-prey interactions, though recent work has shown that escape response can also be induced by high temperature. Here, we test the hypothesis that air-breathing fish may use C-starts in the context of gulping air at the surface. Hoplosternum littorale is an air-breathing freshwater catfish found in South America. Field video observations reveal that their air-breathing behaviour consists of air-gulping at the surface, followed by a fast turn which re-directs the fish towards the bottom. Using high-speed video in the laboratory, we compared the kinematics of the turn immediately following air-gulping performed by H. littorale in normoxia with those of mechanically-triggered C-start escape responses and with routine (i.e. spontaneous) turns. Our results show that air-breathing events overlap considerably with escape responses with a large stage 1 angle in terms of turning rates, distance covered and the relationship between these rates. Therefore, these two behaviours can be considered kinematically comparable, suggesting that air-breathing in this species is followed by escape-like C-start motions, presumably to minimise time at the surface and exposure to avian predators. These findings show that C-starts can occur in a variety of contexts in which fish may need to get away from areas of potential danger.

Dietary supplementation of essential fatty acids in larval pikeperch (Sander lucioperca); short and long term effects on stress tolerance and metabolic physiology.

The present study examined the effects of feeding pike perch larvae Artemia, enriched with either docosahexanoic acid (DHA), arachidonic acid (ARA), oleic acid (OA), olive oil (OO) or a commercial enrichment DHA Selco (DS) on tissue lipid deposition, stress tolerance, growth and development, and metabolic rate. There was higher tissue retention of ARA than DHA at comparable inclusion levels. No differences were observed between diets on the percentage contribution of ARA or DHA to the fatty acid profile of tissues (head and trunk). Total fatty acid content (mgg(-1)) was significantly higher in the head, reflecting its high content of neural tissue. Observations on larval erratic behaviour and mortality following exposure to salinity stress suggested that high inclusions levels of DHA had an alleviating effect, while ARA did not. Particularly larval groups reared for 16 days on diets enriched with OO and OA had mortality rates approaching 100% within two hours. Interestingly, this tendency, although not as pronounced, was also apparent in juvenile fish after 120 days of rearing on a common diet. Standard metabolic rate in larvae on an OO enriched diet was significantly elevated, but otherwise no groups had significant changes to their respiratory physiology. In addition to increased stress challenge sensitivity, early feeding with OA had long term impact on pike perch neural development indicated by a smaller brain size in juvenile fish. In conclusion, lack of DHA in the diet of pikeperch larvae suggests that this long chain polyunsaturated fatty acid is involved in processes that increase stress tolerance and that lack of dietary DHA in early larval stage caused increased stress sensitivity and long-term impaired neural development, while it does not appear to affect metabolic rate at rest.

Vascular arrangement and ultrastructure of the European eelpout Zoarces viviparus ovary: implications for maternal-embryonic exchange.

The structural basis for exchange between maternal serum and ovarian fluid in the viviparous teleost Zoarces viviparus was investigated. Casts of the ovarian vasculature showed that blood supply to the ovary is initially directed to the follicular appendages lining the ovarian wall through thick-walled muscular arteries running along the ovary wall and within the follicular appendages. The follicles had a rich capillary network with diffusion distances between maternal blood and ovarian fluid comparable to those found for gill epithelia, suggesting this is the primary site of gas exchange between maternal plasma and ovarian fluid. Follicular capillary beds were continuous with those in the ovary wall and were eventually drained by the ovarian and intestinal venous systems. The barrier between ovarian fluid and maternal blood consisted of the endothelial cells of the maternal blood vessels and a layer of epithelial cells lining the ovarian lumen, with an intermittent layer of loose connective fibers. Junctional complexes between cells were predominantly anchoring junctions with the occurrence of occasional occluding junctions, supporting the possibility of paracellular transport from maternal serum to ovarian fluid of small molecular weight compounds. Heavy investment in keratin filaments suggests that follicles are tissues of high structural integrity. Evidence for protein synthesis in the ovarian lining was found in the form of Golgi apparatus and rough endoplasmic reticulum. Although numerous cytoplasmic vacuoles and secretory granules were present in both epithelial and endothelial cells, the fate of synthesized protein remains to be determined.

Structural basis for control of secondary vessels in the long-finned eel Anguilla reinhardtii.

Histological sections of primary segmental arteries and associated interarterial anastomoses and secondary vessels from the long-finned eel Anguilla reinhardtii were examined by light and transmission electron microscopy. Interarterial anastomoses were found to originate from the primary vasculature as depressions through the tunica intima and media, from where they ran perpendicularly to the adventitial layer, before coiling extensively. From here the anastomoses travelled a relatively linear path in the outer margin of the adventitia to anastomose with a secondary vessel running in parallel with the primary counterpart. In contrast to findings from other species, secondary vessels had a structure quite similar to that of primary vessels; they were lined by endothelial cells on a continuous basement membrane, with a single layer of smooth muscle cells surrounding the vessel. Smooth muscle cells were also found in the vicinity of interarterial anastomoses in the adventitia, but these appeared more longitudinally orientated. The presence of smooth muscle cells on all aspects of the secondary circulation suggests that this vascular system is regulated in a similar manner as the primary vascular system. Because interarterial anastomoses are structurally integrated with the primary vessel from which they originate, it is anticipated that flow through secondary vessels to some extent is affected by the vascular tone of the primary vessel. Immunohistochemical studies showed that primary segmental arteries displayed moderate immunoreactivity to antibodies against 5-hydroxytryptamine and substance P, while interarterial anastomoses and secondary vessels showed dense immunoreactivity. No immunoreactivity was observed on primary or secondary arteries against neuropeptide Y or calcitonin gene-related peptide.