Cortisol treatment improves the development of hypoosmoregulatory mechanisms in the euryhaline rainbow trout,Salmo gairdneri

The effect of cortisol on osmoregulatory parameters was studied in rainbow trout, (Salmo gairdneri), kept in freshwater (FW) and/or transferred to seawater (SW). Repeated injections of 20 μg cortisol/g fish stimulated gill and gut Na⁺/K⁺-ATPase activity and reduced plasma Na⁺ and Cl⁻ levels after 2 weeks of treatment in FW-adapted fish. Cortisol doses of 0.05 and 1.0 μg/g were without effect. Repeated injections of 10 μg cortisol/g stimulated gill Na⁺/K⁺-ATPase activity and reduced plasma Na+ and Cl⁻ levels in fish in FW, and significantly improved ion regulation after their transfer to 28SW. Higher doses of cortisol (10 and 20 μg/g) induced hyperglycemia, whereas low doses (0.05 and 1.0 μg/g were without effect or induced hypoglycemia. Plasma glucose levels decreased in cortisol-treated fish transferred to SW, whereas transient hyperglycemia was seen in the control fish.     

The effect of dietary sodium chloride on some osmoregulatory parameters of the teleost, Oreochromis niloticus, after transfer from freshwater to seawater

The aim of this work was to determine the effects of supplemental dietary sodium chloride on salt water acclimation of tilapia Oreochromis niloticus. Fish were fed a basal diet supplemented with NaCl (8%) during three weeks in fresh water (FW) and then transferred to salt water (SW) at 15 and 20. Changes in plasma osmolality, chloride ion concentration (Cl^–), plasma level of cortisol and gill Na⁺, K⁺-ATPase activity were measured at 6, 12, 24, 48, 72 and 168 h after transfer to 15SW, while the higher strength SW group (20) was only monitored up to 24 h. Morphological changes in the gill mitochondria-rich (MR) cells were examined in relation to environmental salinity. The changes associated with dietary NaCl were sporadic and of small magnitude. The plasma osmolality and Cl^– increased immediately after transfer up to 12–24 h, but fish fed dietary salt (S) showed lower values than the control group (C). The S group showed higher plasma levels of cortisol than the control, which maintained its initial levels during the experiment. Gill Na⁺, K⁺-ATPase activity of the S group began to increase in the first hours after transfer, reaching maximum at 12 h and returned to basal level at 24 h, while the control group maintained basal levels. The differences between gill Na⁺, K⁺-ATPase activity of S and C fish were significant (p < 0.05) at 12 h. Transmission electron microscopy (TEM) revealed that MR cells in SW show more mitochondria and a more developed tubular system arising from the basolateral membrane. The MR cells of both groups frequently formed a multicellular complex in SW, consisting of a main MR and one or more accessory cells. Such complexes are rarely observed in FW. Some MR cells of fish fed supplemented dietary salt displayed convex apical membrane in FW.     

Enhanced hypoosmoregulatory response to growth hormone after cortisol treatment in immature rainbow trout,Salmo gairdneri

The growth-independent effect of ovine growth hormone (oGH) and oGH + cortisol treatment on seawater (SW) adaptation in immature rainbow trout, Salmo gairdneri was investigated. Fish were injected every second day with saline, 2.0 μg oGH/g or 2.0 μg oGH + 8.0 μg cortisol/g for a maximum of 8 injections in freshwater (FW). Subgroups were transferred to 28‰ SW after 4 or 8 injections, and changes in plasma Na⁺ and Cl⁻, muscle water content and gill Na⁺/K⁺-ATPase activity were measured. In both of the hormone-treated groups retained in FW, gill Na⁺/K⁺-ATPase activity and interlamellar chloride cell density increased. The effects were most pronounced in the oGH + cortisol group after 2 weeks of treatment. After transfer to SW most of the control fish died due to the osmotic stress, whereas in the hormone-treated groups, mortality was low and there was a positive correlation between pretransfer gill Na⁺/K⁺-ATPase and the ability to maintain ionic-osmotic homeostasis after SW transfer. After two weeks of oGH + cortisol treatment, gill Na⁺/K⁺-ATPase activity was maximal. In contrast, after SW transfer, Na⁺/K⁺-ATPase activity increased further in the oGH-treated group. This group regulated ionic-osmotic parameters less effectively than the oGH + cortisol-treated group. The data indicate that GH and cortisol are important hormones in the regulation of hypoosmoregulatory mechanisms in S. gairdneri.     

Physiology of seawater acclimation in the striped bass,Morone saxatilis (Walbaum)

Several experiments were performed to investigate the physiology of seawater acclimation in the striped bass, Morone saxatilis. Transfer of fish from fresh water (FW) to seawater (SW; 31–32 ppt) induced only a minimal disturbance of osmotic homeostasis. Ambient salinity did not affect plasma thyroxine, but plasma cortisol remained elevated for 24h after SW transfer. Gill and opercular membrane chloride cell density and Na⁺,K⁺-ATPase activity were relatively high and unaffected by salinity. Average chloride cell size, however, was slightly increased (16%) in SW-acclimated fish. Gill succinate dehydrogenase activity was higher in SW-acclimated fish than in FW fish. Kidney Na⁺, K⁺-ATPase activity was slightly lower (16%) in SW fish than in FW fish. Posterior intestinal Na⁺,K⁺-ATPase activity and water transport capacity (J_v) did not change upon SW transfer, whereas middle intestinal Na⁺,K⁺-ATPase activity increased 35% after transfer and was correlated with an increase in J_v (110%). As salinity induced only minor changes in the osmoregulatory organs examined, it is proposed that the intrinsic euryhalinity of the striped bass may be related to a high degree of “preparedness” for hypoosmoregulation that is uncommon among teleosts studied to data.     

Response of rainbow trout gill (Na++K+)-ATPase and chloride cells to T3 and NaCl administration

With the aim of comparing the effects of oral T₃ and NaCl administration on trout hypoosmoregulatory mechanisms, three groups of rainbow trout (Oncorhynchus mykiss Walbaum) held in freshwater (FW) were fed a basal diet (C), the same diet containing 8.83 ppm of 3,5,3-triiodo-L-thyronine (T₃) (T) or 10% (w/w) NaCl (N) respectively for 30 d. They were then transferred to brackish water (BW) for 22 d and fed on diet C. Gill (Na⁺+K⁺)-ATPase activity and its dependence on ATP, Na⁺ and pH, number of gill chloride cells (CC), serum T₃ level as well as fish growth, condition factor (K) and mortality were evaluated. During the FW phase, as compared to C trout, T trout showed a two fold higher serum T₃ level, had unchanged gill (Na⁺+K⁺)-ATPase activity and increased CC number, whereas N trout showed higher gill (Na⁺+K⁺)-ATPase activity and CC number. At the end of the experiment the enzyme activity was in the order T>N>C groups and all groups showed similar CC number. Both treatments changed the enzyme activation kinetics by ATP and Na⁺. A transient increase in K value occurred in N group during the period of salt administration. In BW, T and N groups had higher and lower survival than C group respectively. Other parameters were unaffected by the treatments. This trial suggests that T₃ administration promotes the development of hypoosmoregulatory mechanisms of trout but it leaves the (Na⁺+K⁺)-ATPase activity unaltered till the transfer to a hyperosmotic environment.     

Influence of cortisol, growth hormone, insulin-like growth factor I and 3,3′,5-triiodo-l-thyronine on hypoosmoregulatory ability in the euryhaline teleost Fundulus heteroclitus

The capacity of cortisol, ovine growth hormone (oGH), recombinant bovine insulin-like growth factor I (rbIGF-I) and 3,3,5-triiodo-l-thyronine (T₃) to increase hypoosmoregulatory capacity in the euryhaline teleost Fundulus heteroclitus was examined. Fish acclimated to brackish water (BW, 10 ppt salinity) were injected with a single dose of hormone suspended in oil and transferred to seawater (SW, 35 ppt salinity) 10 days post-injection. Fish were sampled 24 h after transfer and plasma osmolality and gill Na⁺, K⁺-ATPase activity were examined. Transfer from BW to SW induced significantly increased plasma osmolality but not gill Na⁺, K⁺-ATPase activity. Cortisol (50 g g^–1 body weight) improved the ability to maintain plasma osmolality and to increase gill Na⁺, K⁺-ATPase activity. oGH (5 g g^–1 body weight) also increased hypoosmoregulatory ability and gill Na⁺, K⁺-ATPase activity. A cooperation between oGH and cortisol was observed in increasing hypoosmoregulatory ability but not in increasing gill Na⁺, K⁺-ATPase activity. rbIGF-I (0.5 g g^–1 body weight) alone was without effect in increasing salinity tolerance or gill Na⁺, K⁺-ATPase activity. rbIGF-I and oGH showed a positive interaction in increasing salinity tolerance, but not gill Na⁺, K⁺-ATPase activity. Treatment with T₃ (5 g g^–1 body weight) alone did not increase salinity tolerance or gill Na⁺, K⁺-ATPase activity, and there was no consistent significant interaction between cortisol and T₃ or between GH and T₃. The results confirm the classical role of cortisol as a seawater-adapting hormone and indicate an interaction between cortisol and the GH/IGF-I axis during seawater acclimation of Fundulus heteroclitus.     

Effects of 17β-estradiol and 4-nonylphenol on smoltification and vitellogenesis in Atlantic salmon (Salmo salar)

The impact of 17-estradiol (E₂) and the putative estrogenic compound, 4-nonylphenol (4-NP), on smoltification and vitellogenesis in Atlantic salmon (Salmo salar) was investigated during a 30 day period starting late April. Three groups of fresh water (FW) fish (1 year old, mixed sexes, average weight 23 g) were injected once a week with 50 µg (0.18 µmol) 17-estradiol, 3 mg (13.6 µmol) 4-nonylphenol dissolved in peanut oil, or peanut oil alone as control. Every ten days, subgroups were challenged with 28 ppt seawater (SW) for 24h, and sampled together with subgroups of FW fish. Treatment effects were examined on vitellogenic and osmoregulatory parameters. E₂ and 4-NP treatment increased the total calcium and protein level in plasma and the hepatosomatic index of FW fish, both indicating an activated vitellogenesis in the liver. The presence of vitellogenin in the plasma of 4-NP- and E₂-treated groups was further indicated by the appearance of a high molecular weight vitellogenin band (550 kDa) in electropherograms produced by native gel electrophoresis. This band appeared in exactly the same position in both the E₂- and the 4-NP-treated groups but could not be detected in controls. During the 30 day treatment period, control fish approached the peak of smoltification, as indicated by a distinct silvery appearance, decreasing condition factor, increasing levels of gill Na⁺,K⁺-ATPase and improved hypoosmoregulatory performance in the SW-challenge test. Both E₂ and 4-NP treatments significantly inhibited the progress of smoltification, as judged by a significant reduction of gill Na⁺,K⁺-ATPase activity, relative -subunit Na⁺,K⁺-ATPase mRNA expression, gill chloride cell density and a poorer hypoosmoregulatory performance of treated fish. The impaired SW-tolerance of E₂- and 4-NP-treated fish was strongly correlated with a decreased gill Na⁺,K⁺-ATPase activity. Despite a difference in relative potency, the present study shows that 4-nonylphenol and 17-estradiol may have qualitatively similar inhibitory effects on smoltification and hypoosmoregulatory physiology of Atlantic salmon. Both 4-NP and E₂ activated the vitellogenic system, and the study supports the hypothesis that sexual maturation and smoltification are antagonistic, developmental phenomena in salmon. It is suggested that the presence of estrogenic compounds in the environment may negatively influence smoltification and migration in wild stocks of salmon.     

Cortisol stimulates intestinal fluid uptake in Atlantic salmon (Salmo salar) in the post-smolt stage

The fluid uptake rate of the posterior intestine of salmonids increases during the parr-smolt transformation. Intestinal fluid uptake in post-smolt Atlantic salmon was investigated after treatment with cortisol and growth hormone (GH), alone or together. Two replicate experiments were conducted in August 1991 and August 1992. Cortisol was emulsified in vegetable shortening and vegetable oil (1:1) and implanted into the peritoneal cavity. GH was administered as intraperitoneal injections in a saline vehicle on days 0 and 2. On days 5 and 6, plasma cortisol levels, gill Na⁺,K⁺-ATPase activity, andin vitro measurements of fluid transport rate (J_v) across the posterior intestine were measured. Implants of cortisol elevated the plasma cortisol levels within a physiological range, and resulted in elevated gill Na⁺,K⁺-ATPase activity, as expected. The fluid uptake rate across the posterior intestine was roughly doubled by cortisol treatment. GH treatment did not affect intestinal fluid transport, gill Na⁺,K⁺-ATPase activity, or plasma cortisol concentrations. The seawater-adapting increase in the rate of fluid uptake by the posterior intestine of smolting salmon is probably stimulated by elevated plasma cortisol concentrations.     

Phenotypic plasticity in gene expression and physiological response in red drum <Emphasis Type="Italic">Sciaenops ocellatus</Emphasis> exposed to a long-term freshwater environment

Red drum (Sciaenops ocellatus) is a euryhaline fish commonly found in the Gulf of Mexico and along the Atlantic coast of North America. Because of high commercial demand and its euryhaline characteristics, aquaculture of this species has diversified from marine to low-salinity aquaculture systems. In recent years, interest in the feasibility of producing red drum in inland freshwater systems has grown and this prompted us to investigate its osmoregulatory capacity after rearing for 8 months in a freshwater aquaculture system. We compared the activities of several genes and enzymes involved in the osmoregulatory process in freshwater-acclimatized (FW) and seawater (SW) red drum. The gene expression profiles were variable: the expression of genes encoding Na⁺/K⁺-ATPase (NKA) and the cystic fibrosis transmembrane regulator (CFTR) was slightly higher in SW than FW fish, while phosphoenolpyruvate carboxykinase (PEPCK) and the glucocorticoid receptor messenger RNA (mRNA) levels were higher in FW red drum. The total plasma K concentration was 60.3% lower, and gill NKA activity was 63.5% lower in FW than in SW fish. PEPCK activity was twofold higher in FW than in SW red drum. Similarly, liver glycogen was 60% higher in FW fish. In summary, both gene expression and the enzyme activity data support the phenotypic plasticity of red drum and suggest that the limited capacity for ion homeostasis observed, in particular the low plasma K concentration, was due to the composition of freshwater and does not necessarily reflect a physiological inability to osmoregulate.     

Effects of sulfide on K+ flux pathways in red blood cells of crucian carp and rainbow trout

The effect of sulfide on K⁺ influx pathways was measured in red blood cells (RBCs) of sulfide-sensitive rainbow trout (Oncorhynchus mykiss) and sulfide-tolerant crucian carp (Carassius carassius). In trout RBCs, maximal inhibition of Na⁺, K⁺-ATPase was attained at 10 mol l^–1 sulfide and amounted to 32% without being influenced by pH between 6.7 and 8.3. Ouabain-resistant K⁺ influx in the absence and presence of sulfide was insignificant at pH values between 6.7 and 7.7. At higher pH values ouabain-resistant K⁺ influx increased, but was inhibited to about 15% by 30 mol l^–1 sulfide. In RBCs of crucian carp neither Na⁺, K⁺-ATPase nor ouabain-resistant K⁺ influx were affected by sulfide concentrations up to 850 mol l^–1. Differences in sulfide-sensitivity of K⁺ influx between both species can be based upon different properties of the membrane transporter themselves. The reduced Na⁺, K⁺-ATPase activity in trout RBCs may also result from a slightly reduced (by 9%) ATP level after sulfide exposure. In addition, intracellular sulfide concentrations were higher in trout RBCs as compared to crucian carp. In trout, intracellular sulfide concentrations reached extracellular levels within 5 min of incubation whereas sulfide concentrations in crucian carp RBCs remained about 2-fold lower than extracellular concentrations. Although the physiological basis of sulfide-insensitive K⁺ influx in crucian carp RBCs is currently unknown it may contribute to the extremely high sulfide-tolerance of this species.