The effects of arachidonic acid on the endocrine and osmoregulatory response of tilapia (Oreochromis mossambicus) acclimated to seawater and subjected to confinement stress.

In previous studies in freshwater tilapia (Oreochromis mossambicus), dietary supplementation with arachidonic acid (ArA; 20:4n - 6) had considerable, opposing effects on the main ion-transporting enzyme Na(+)/K(+)-ATPase in gills and kidneys and changed the release of osmoregulatory hormones, such as cortisol. The present study was performed to assess the influence of dietary ArA on (1) the osmoregulatory capacity of tilapia acclimated to seawater (SW) (34‰) and (2) the osmoregulatory imbalance associated with acute stress. The increased ambient salinity was associated with significant alterations in the tissue fatty acid composition, particularly the n - 6 polyunsaturated fatty acids (PUFAs). Tissue levels of ArA were further increased as a result of dietary supplementation, whereas docosahexaenoic acid (DHA, 22:6n - 3) and eicosapentaenoic acid (EPA, 20:5n - 3) decreased in gills and kidneys. Basal plasma cortisol as well as lactate levels were elevated in the ArA-supplemented SW-acclimated tilapia compared with the control group. The 5 min of confinement (transient stress) increased plasma cortisol, glucose, and lactate levels with significantly higher levels in ArA-supplemented tilapia. Confinement was also associated with significantly elevated plasma osmolality, sodium, chloride, and potassium levels. ArA-supplemented tilapia showed markedly lower ionic disturbances after confinement, suggesting that dietary ArA can attenuate the hydromineral imbalance associated with acute stress. These results emphasize the involvement of ArA and/or its metabolites in the endocrine and osmoregulatory processes and the response to confinement stress.

Dietary supplementation with arachidonic acid in tilapia (Oreochromis mossambicus) reveals physiological effects not mediated by prostaglandins.

This study aims to clarify the role of the polyunsaturated fatty acid arachidonic acid (ArA, 20:4n-6) in the stress response of Mozambique tilapia (Oreochromis mossambicus). ArA is converted into eicosanoids, including prostaglandins, which can influence the response to stressors. Tilapia, a species able to form ArA from its precursor, was supplemented with ArA for 18 days, after which they were confined for 5 min. Acetylsalicylic acid (ASA, COX-inhibitor) was subsequently administered to distinguish ArA-mediated effects from enhanced prostaglandin E(2) (PGE(2)) synthesis. ArA supplemented fish had higher ArA levels in gills and kidneys, and these levels were further enhanced after ASA treatment. Levels of total monounsaturated and polyunsaturated fatty acids as well as docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and ArA, were altered 24h after confinement, particularly in the kidneys. ArA supplementation had no effect on basal cortisol levels, while ArA + ASA reduced basal cortisol levels. ArA + ASA augmented the cortisol response to confinement. The combination of ArA + ASA also elevated plasma basal prolactin (tPRL)(177) and 3,5,3'-triiodothyronine (T(3)) levels. Neither ArA nor ASA affected the stress-associated increases in plasma glucose and lactate. Na(+), K(+)-ATPase activity in the gills was reduced after ArA supplementation and was even further suppressed by subsequent ASA treatment. In an additional feeding trial, ArA supplementation enhanced the renal Na(+), K(+)-ATPase activity. In vitro, ArA was a potent inhibitor of the Na(+), K(+)-ATPase activity of gill and kidney homogenates. In contrast, PGE(2) had no effect on branchial ATPase, whereas the effect on renal ATPase activity was concentration dependent. Modifying the dietary intake of ArA alters the response of tilapia to an acute stressor and influences osmoregulatory processes and it is unlikely that these effects are due to an enhanced production of prostaglandins.

Arachidonic acid reduces the stress response of gilthead seabream Sparus aurata L.

In this study the influence of the dietary level of the fatty acid arachidonic acid (ArA, 20:4n-6) was determined on the acute stress response and osmoregulation of adult gilthead seabream Sparus aurata L. Seabream were fed a diet containing either 0.9% or 2.4% of total fatty acids as ArA for 18 days before being subjected to a 5 min period of net confinement. Prior to this stressor, a subgroup of fish from both dietary treatment groups was treated with acetylsalicylic acid (ASA), an irreversible blocker of cyclooxygenase (COX). This would indicate whether any effects were caused by an enhanced synthesis of prostaglandins derived from ArA. The highest ArA levels were found in the kidneys, and these were further enhanced by dietary ArA-supplementation. In gill tissues, there were significant changes in all selected fatty acid classes 24 h after confinement, except for the docosahexaenoic acid (DHA, 22:6n-3): eicosapentaenoic acid (EPA, 20:5n-3) ratio. ArA feeding strongly reduced the cortisol response to confinement, which was partially counteracted by ASA treatment. ArA also attenuated the stress-associated increase in plasma osmolality and, in combination with ASA, enhanced the osmolality and plasma chloride levels, but reduced plasma sodium levels after confinement. Furthermore, ArA enhanced the branchial Na(+), K(+)-ATPase activity both before and after confinement, whereas feeding ASA diminished this effect. It appeared that the effects of ArA-supplementation could not always be ascribed to an increase in prostaglandin synthesis. It is advisable to determine the long-term effects of replacing fish oils in commercial diets with vegetable oils that contain no long-chain fatty acids, particularly in carnivorous/marine species with low fatty acid elongation and desaturation activities. The effects of a low dietary intake of ArA (and other polyunsaturated fatty acids) should be studied over a longer term, taking into account any consequences for the health of the fish.

Lipid digestion in turbot (Scopthalmus maximus) 11: Lipolysis in vitro of (14)C-labelled triacylglycerol, cholesterol ester and phosphatidylcholine by digesta from different segments of the digestive tract.

Preparations of digesta from the stomach, foregut, hindgut and rectum of turbot (Scophthalmus maximus) were assayed for the ability to hydrolyse glycerol tri[1-(14)C]oleate ((14)C-TAG), 1,2-di[1-(14)C]palmitoyl L-3-phosphatidylcholine ((14)C-PC) and cholesterol [1-(14)]oleate ((14)C-CE) over 1,2,3 and 17h. In the assay of foregut digesta with (14)C-TAG substrate, 37.8% of the total radioactivity was found in the FFA class after the first hour of incubation. This value increased to a maximum of 68.5% of the available label after 17h incubation. Over the same time the proportion of radioactivity in diacylglycerols (DAG, 31.6%-7.4%) decreased while that in the monoacylglycerols increased (MAG, 14.0%-22.3%). In assays of digesta from the hindgut and rectum, after 1 h of incubation, the proportion of radioactivity recovered in FFA represented 64.9% and 74.8%, respectively, whereas the proportions in both DAG and MAG decreased with incubation time. Similarly to (14)C-TAG, the highest rate of lipolytic hydrolysis of (14)C-CE occurred in digesta from the posterior digestive tract where the proportions of radioactivity recovered in FFA of the hindgut (50.0%) and rectum (81.9%) preparations were substantially higher than those of the stomach (3.5%) and the foregut (14.4%) after 1h. With (14)C-PC as substrate the levels of radiolabelled FFA in both the foregut and the hindgut (2.4% and 7.6%, respectively) were markedly lower than the 37.5% in the rectum. The results suggest that the posterior digestive tract is very active in non-specific and phospholipid lipolysis and a region where the major part of lipid digestion takes place.

Lipid digestion in turbot (Scophthalmus maximus). I: Lipid class and fatty acid composition of digesta from different segments of the digestive tract.

The lipid content and lipid composition of digesta from the stomach, foregut, hindgut and rectum of juvenile turbot fed a commercial diet were determined in order to examine the process of lipid digestion in this species. The moisture content of the digesta increased along the digestive tract from 71.5% in the stomach to 89.6% in the rectum. The lipid content of the digesta increased initially from 15.7% of the dry weight in the stomach to 36.1% in the foregut but thereafter decreased through 23.2% in the hindgut to 9.1% in the rectum. The proportion of triacylglycerols (TAG) in the total lipid of the digesta decreased from 63% in the stomach to 17.4% in the rectum whereas that of free fatty acids (FFA) increased from 10016 to 48.9%. The highest proportions of monoacylglycerols (MAG), diacylglycerols (DAG) and most phospholipids were observed in the lipid of the hindgut digesta. In addition, a fall in levels of neutral and phospholipid classes as digesta moved from hindgut to rectum signified absorption.Analysis of the fatty acid composition of the lipid classes TAG, DAG and MAG suggest a polyunsaturated fatty acid specificity for hydrolysis may exist. Saturated, monounsaturated and polyunsaturated fatty acids (PUFA) accounted for 17.9%, 45.4% and 37.0%, respectively of the FFA present in the foregut whereas the corresponding values for the rectum were 32.6%, 51.9% and 16.3%. Overall, the results suggest a PUFA specifity for hydrolysis may exist alongside the positional non-specific lipolytic activity associated with the hindgut regions of the digestive tract of turbot and that PUFA, released by lipolysis are more effectively absorbed from the digesta than monounsaturated and saturated fatty acids.

The effect of dietary lecithin and lipase, as a function of age, on n-9 fatty acid incorporation in the tissue lipids of Sparus aurata larvae.

The present study tested the effect of dietary lecithin and exogenous lipase on the incorporation of oleic acid in the tissue lipids of gilthead seabream larvae (Sparus aurata). Two of four microdiets were prepared by the addition of [(14)C]oleic acid as free fatty acid (FFA) to diets containing either 5% cuttlefish liver oil (CLO) or 5% soybean lecithin. Glycerol tri[1-(14)C]oleate was similarly incorporated in two other diets identical in lipid (4% cuttlefish liver oil, 1% soybean lecithin) and non-lipid composition but differed in that one contained a supplement of 0.05% porcine lipase. The effect of these diets was tested by following the incorporation of the label (dpm/mg larvae DBW) in the neutral and phospholipid fractions of seabream larvae at four different ages (21, 27, 32 and 45 days after hatching).A significant (p<0.05) effect of dietary lecithin on the incorporation of labelled FFA in both larval neutral and phospholipid fractions was demonstrated at all ages. This was particularly pronounced during early development (day 21) where fish fed the lecithin supplement incorporated 6.75 times more label than the diet containing [(14)C]oleic acid in CLO. The dietary lecithin enhancing effect diminished with age but was still significant at day 45 (2.17 times more label). In addition, the label was considerably higher in the phospholipid fraction compared to the neutral lipid, reflecting the high demand for membrane synthesis during rapid growth. Lecithin fed larvae demonstrated a higher consumption rate and efficiency of incorporation than fish consuming the cuttlefish liver oil diet, suggesting an emulsifying function for dietary phospholipid.In contrast, the supplementation with lipase showed a clear effect only in older fish where 45 day old larvae fed the lipase diet demonstrated a 3.42 times increase in radioactivity in their tissue lipids. This late lipase response may be the result of an insufficient level of dietary lecithin (M) and a short intestinal length being ineffective, in the early larval stages, in incorporating labelled free fatty acid from dietary glycerol tri[1-(14)C]oleate breakdown.