Chronic Cadmium Exposure can Alter Energy Allocation to Physiological Functions in the Shrimp Penaeus vannamei.

Environmental stressors in aquatic organisms can be assessed using a bioenergetic approach based on the evaluation of changes in their physiological parameters. We evaluated the chronic effects of cadmium (Cd2+) on the energy balance as well as the survival, growth, metabolism, nitrogen excretion, hepatosomatic index, oxidized energy substrate, and osmoregulation of the shrimp Penaeus vannamei with the hypothesis that the high energy demand related to the homeostatic regulation of Cd2+could disrupt the energy balance and as a consequence, their physiological functions. The shrimp exposed to Cd2+ had higher mortality (30%), directed more energy into growth (33% of energy intake), ingested 10% more energy, and defecated less than control animals. Cd2+ exposure caused a tendency to decrease metabolism and ammonia excretion but did not alter the hepatosomatic index, type of energy substrate oxidized, and the hyperosmorregulatory pattern of the species. The Cd+2 exposure may have induced a trade-off response because there was a growth rate increase accompanied by increased mortality.

Tissue Accumulation and the Effects of Long-Term Dietary Copper Contamination on Osmoregulation in the Mudflat Fiddler Crab Minuca rapax (Crustacea, Ocypodidae).

We examined copper accumulation in the hemolymph, gills and hepatopancreas, and hemolymph osmolality, Na+ and Cl- concentrations, together with gill Na+/K+-ATPase and carbonic anhydrase activities, after dietary copper delivery (0, 100 or 500 Cu µg g-1) for 12 days in a fiddler crab, Minuca rapax. In contaminated crabs, copper concentration decreased in the hemolymph and hepatopancreas, but increased in the gills. Hemolymph osmolality and gill Na+/K+-ATPase activity increased while hemolymph [Na+] and [Cl-] and gill carbonic anhydrase activity decreased. Excretion likely accounts for the decreased hemolymph and hepatopancreas copper titers. Dietary copper clearly affected osmoregulatory ability and hemolymph Na+ and Cl- regulation in M. rapax. Gill copper accumulation decreased carbonic anhydrase activity, suggesting that dietary copper affects acid-base balance. Elevated gill Na+/K+-ATPase activity appears to compensate for the ion-regulatory disturbance. These effects of dietary copper illustrate likely impacts on semi-terrestrial species that feed on metal-contaminated sediments.

Osmoregulatory disturbance in Neotropical fish exposed to the crude extracts of the cyanobacterium, Radiocystis fernandoi.

Blooms of cyanobacteria, a common event in eutrophic environments, result in the release of potentially toxic substances into the water. The cyanobacterium Radiocystis fernandoi produces microcystin (MC) and other peptides that may disturb homeostasis. This study evaluated the effect of intraperitoneal injections containing the crude extract (CE) of R. fernandoi strain R28 on the gills and kidneys of neotropical fish, Piaractus mesopotamicus, 3, 6 and 24 h post-injection. CE contained MC-RR, MC-YR and minor other oligopeptides. Plasma ions and the activities of the enzymes PP1 and PP2A, Na+/K+-ATPase (NKA), H+-ATPase (HA) and carbonic anhydrase (CA) were determined and morphological changes in both the gills and kidneys were characterized. Compared to controls, the concentration of Na+ within the plasma of P. mesopotamicus decreased after treatment with CE 3 h post treatment and increased after 24 h; the concentration of K+ decreased after 6 h. The activity of the endogenous PP1 and PP2A was unchanged in the gills and was inhibited in the kidneys 6 h after i.p. injection. In the gills, NKA activity increased after 3 h and decreased 6 h post i.p. exposure. Further, NKA activity did not differ from the controls 24-h post injection. In the kidneys, NKA, HA and CA activities were unaffected by treatment. The mitochondria-rich cell (MRC) density in the gills decreased after 3 h in the filament and 3 and 6 h in the lamellae and was restored to the control levels 24 h post-exposure. Filament epithelial hyperplasia and hypertrophy, lamellar atrophy and rupture of the lamellar epithelium were the most common effects of treatment in the gills. No histopathological changes occurred in the kidneys. This study demonstrates that a single dose of toxic CE from R. fernandoi can cause a transitory ion imbalance in P. mesopotamicus which is related to the changes in MRC levels and NKA activity. Ionic balance was recovered 24 h post i.p. injection, however, morphological changes that occurred in the gills took a longer amount of time to return to normal. To conclude, the effects of components contained within the CE of R. fernandoi may be harmful to P. mesopotamicus. In particular, the recovery of ionic regulation depends on MRC responses and histopathological changes produced by CE may affect gas exchange and other gill functions.

Low salinity negatively affects early larval development of Nile tilapia, Oreochromis niloticus: insights from skeletal muscle and molecular biomarkers.

The present study evaluated the effects of low salinity on the early larval development of Oreochromis niloticus, specifically histological damage to white muscle, morphology of the yolk-sac surface and trunk area, and molecular expression of apoptosis and cell proliferation biomarkers. Newly hatched larvae were submitted to four salinity treatments for a period of 48 or 72 h, in duplicate: (S0) freshwater, (S2) 2 g l-1, (S4) 4 g l-1, and (S6) 6 g l-1NaCl. Larval development was examined using histology, electron microscopy, enzyme-linked immunosorbent assay (ELISA), and morphometry. At the yolk-sac surface, larvae of S4 and S6 displayed alterations to the apical opening of chloride cells that may be related to osmotic expenditure caused by the increased salinity. Caspase-3 expression did not differ significantly among treatments, however significantly lower proliferating cell nuclear antigen (PCNA) expression (P < 0.05) suggested minor cell proliferation in larvae of S4 and S6 compared with S0 and S2. Furthermore, there was a significant reduction in both trunk area and percentage of normal white muscle fibres (WF) in larvae of S4 and S6. Vacuolated areas and myofibrils concentrated at the cell periphery and found in the white muscle from larvae exposed to saline environments suggested disturbance to muscle development. Oedema and mononuclear infiltrate were also observed in the white muscle of S4 and S6 larvae. Together these results indicated that treatments with 4 and 6 g l-1 NaCl may cause osmoregulation expenditure, morphological alterations to the yolk-sac surface and histological damage to skeletal muscle that negatively affected the early larval development of O. niloticus.

Combined effects of waterborne copper exposure and salinity on enzymes related to osmoregulation and ammonia excretion by blue crab Callinectes sapidus.

Copper is essential, but can be toxic to aquatic organisms when present in high concentrations. In freshwater crustaceans, copper inhibits enzymes related to ionic and osmoregulation and to the ammonia efflux, that leads to Na+ imbalance and inhibition of ammonia excretion. In the animals inhabiting estuarine or seawater, mechanisms of copper toxicity is not clear, but had been described as disruption of ionregulation and metabolism. To clarify the mechanism of copper toxicity in crustaceans inhabiting variable salinity, this work investigated whether copper affects ammonia excretion and enzymes used for ammonia balance and osmoregulation in the blue crab Callintectes sapidus acclimated to salinity 2 and 30 ppt. To achieve this, juveniles of the blue crab were exposed to 63.5 µg/L of copper at both salinities for 96 h. This is an environmentally realistic copper concentration. Results of ammonia efflux, free amino acids and Na+ concentrations in hemolymph, Na+/K+-ATPase, H+-ATPase and, carbonic anhydrase (CA) activities in gills were consistent with the osmoregulatory pattern adopted by the blue crab, which hyperosmoregulates at salinity 2 ppt and osmoconforms at 30 ppt. At 30 ppt copper reduced free amino acid in hemolymph of crabs, suggesting an effect of the metal on osmotic performance. At 2 ppt, copper significantly increased the H+-ATPase activity involved in ammonia excretion. This may be a compensatory response of crabs to maintain low levels of ammonia in their hemolymph; which can be increased by copper exposure. Results presented here are useful for the improvement of the Biotic Ligand Model (BLM) to predict copper toxicity for saltwater environments.

Release of taurine and glutamate contributes to cell volume regulation in human retinal Müller cells: differences in modulation by calcium.

Neuronal activity in the retina generates osmotic gradients that lead to Müller cell swelling, followed by a regulatory volume decrease (RVD) response, partially due to the isoosmotic efflux of KCl and water. However, our previous studies in a human Müller cell line (MIO-M1) demonstrated that an important fraction of RVD may also involve the efflux of organic solutes. We also showed that RVD depends on the swelling-induced Ca2+ release from intracellular stores. Here we investigate the contribution of taurine (Tau) and glutamate (Glu), the most relevant amino acids in Müller cells, to RVD through the volume-regulated anion channel (VRAC), as well as their Ca2+ dependency in MIO-M1 cells. Swelling-induced [3H]Tau/[3H]Glu release was assessed by radiotracer assays and cell volume by fluorescence videomicroscopy. Results showed that cells exhibited an osmosensitive efflux of [3H]Tau and [3H]Glu (Tau > Glu) blunted by VRAC inhibitors 4-(2-butyl-6,7-dichloro-2-cyclopentylindan-1-on-5-yl)-oxybutyric acid and carbenoxolone reducing RVD. Only [3H]Tau efflux was mainly dependent on Ca2+ release from intracellular stores. RVD was unaffected in a Ca2+-free medium, probably due to Ca2+-independent Tau and Glu release, but was reduced by chelating intracellular Ca2+. The inhibition of phosphatidylinositol-3-kinase reduced [3H]Glu efflux but also the Ca2+-insensitive [3H]Tau fraction and decreased RVD, providing evidence of the relevance of this Ca2+-independent pathway. We propose that VRAC-mediated Tau and Glu release has a relevant role in RVD in Müller cells. The observed disparities in Ca2+ influence on amino acid release suggest the presence of VRAC isoforms that may differ in substrate selectivity and regulatory mechanisms, with important implications for retinal physiology. NEW & NOTEWORTHY The mechanisms for cell volume regulation in retinal Müller cells are still unknown. We show that swelling-induced taurine and glutamate release mediated by the volume-regulated anion channel (VRAC) largely contributes the to the regulatory volume decrease response in a human Müller cell line. Interestingly, the hypotonic-induced efflux of these amino acids exhibits disparities in Ca2+-dependent and -independent regulatory mechanisms, which strongly suggests that Müller cells may express different VRAC heteromers formed by the recently discovered leucine-rich repeat containing 8 (LRRC8) proteins.

Effects of the recombinant crustacean hyperglycemic hormones rCHH-B1 and rCHH-B2 on the osmo-ionic regulation of the shrimp Litopenaeus vannamei exposed to acute salinity stress.

The Pacific white shrimp Litopenaeus vannamei is a euryhaline organism that copes with salinity fluctuations in the environment; therefore, its osmotic and ionic regulation abilities are vital. Osmoregulation may be controlled by the crustacean hyperglycemic hormone (CHH), a neuropeptide mainly expressed in the eyestalks. In L. vannamei, CHH-B1 and CHH-B2 are CHH isoforms isolated from the eyestalks whose expression is influenced by environmental salinity. It has been suggested that they are involved in the response to salinity stress. To clarify this, we investigated the effect of the recombinant peptides, rCHH-B1 and rCHH-B2, on the osmo-ionic regulation of shrimp acutely exposed to different salinity conditions (8, 26 and 45‰). Both rCHHs promoted differential effects on the osmoregulatory capacity (OC) and the ionoregulatory capacity (IC) for hemolymph Na+ and Cl- during iso-osmotic (26‰) and hyper-osmotic (45‰) transfers. These changes were linked to the changes observed in Na+/K+ ATPase and carbonic anhydrase gene expression in gills, especially under high salinity conditions, suggesting that the hormones may regulate the expression of these genes. Glucose and protein levels measured during acute salinity transfer suggest their roles as sources of metabolic energy for osmotic regulation or as organic osmolytes. These results taken together suggest that both the CHH-B1 and CHH-B2 peptides are important regulators of the physiological response of L. vannamei to acute salinity fluctuations.

Histological alterations in gills of Macrobrachium amazonicum juveniles exposed to ammonia and nitrite.

Aquaculture has shown great growth in the last decades. Due to the restrictions on water use, production systems are becoming increasingly more intensive, raising concerns about the production water quality. Macrobrachium amazonicum is among the freshwater prawn species with favorable characteristics for production and possibility of intensification. Nitrogen compounds such as ammonia and nitrite affect the health of aquatic organisms since they quickly reach toxic concentrations. These compounds can also cause damage to the gill structure, leading to hypoxia in tissues, affecting acid-base balance, osmoregulation (salt absorption) and ammonia excretion, decreasing the immune capacity of the animal and, in extreme cases, cause death. The aim of this study was to assess histological changes in the gills of Macrobrachium amazonicum juveniles subjected to different concentrations of total ammonia and nitrite. The prawns were subjected to different concentrations of those compounds and their gills were removed and preserved for histological analysis. The gills were assessed for changes according to the Organ Index (Iorg) and, for each change, an importance factor (w) was attributed according to the degree of reversibility and applied according to the degree of extension or frequency of the damage. The damage to the gills in the treatments with 100% mortality, both for ammonia and nitrite, corresponded to the high occurrence of progressive, regressive, circulatory, and inflammation damages. The other treatments (which caused less mortality) had mainly inflammation and regressive damages, whose occurrence increased according to the increase in ammonia and nitrite concentration. The histological analysis confirmed that the higher the total ammonia and nitrite concentrations, the larger the damages caused to the gill structure and that lower nitrite concentrations caused similar damages to those caused by higher total ammonia concentrations, which reflects the lower capacity M. amazonicum has to tolerate nitrite.

Effect of polychlorinated biphenyls on osmoregulatory response and apoptosis in GIFT tilapia, Oreochromis niloticus.

In the present study, GIFT tilapia Oreochromis niloticus were exposed to polychlorinated biphenyls (PCBs) for 7, 14, and 21 days. Over the duration of the exposure, genotoxicity and the activity of Na+/K+-ATPase (NKA) and Ca+/Mg+-ATPase (CMA) were measured in the gill, kidney, and intestine, to evaluate changes in osmoregulatory response in O. niloticus. Our results showed significant decreases in organic NKA (except in gill tissues after 0.5 mg/L PCB-exposure) and CMA activity. The results of the genotoxicity assay showed significant increases in atp1a1a, nkcc2 (only in gill tissue), and fxyd7 (except after 21 days of 5 mg/L PCB exposure). We found significant increases in caspase proteins in the liver in the 5-mg/L PCB exposure group, and the transcripts showed dose-dependent increases between treatment groups over the exposure duration. This study presents evidence that chronic exposure to PCB could result in organic osmoregulatory response and hepatic apoptosis in GIFT tilapia.

Acute effects of cadmium on osmoregulation of the freshwater teleost Prochilodus lineatus: enzymes activity and plasma ions.

Cadmium (Cd) is a trace element that is very toxic to fish. It is commonly found in surface waters contaminated with industrial effluents. When dissolved in water, Cd can rapidly cause physiological changes in the gills and kidneys of freshwater fish. The objective of this study was to evaluate the acute effects of Cd on the osmoregulation of the Neotropical fish Prochilodus lineatus. Juvenile fish were exposed to Cd at two concentrations [1 (Cd1) and 10 (Cd10) µgL(-1)] for 24 and 96h. The effects of Cd were evaluated through the analysis of ions (Na(+), K(+), Ca(2+), and Cl(-)) and plasma osmolality, and by measuring the activities of enzymes involved in osmoregulation obtained from the gills and kidney. Fish exposed to Cd for 24 and 96h showed a decrease in Na(+)/K(+)-ATPase activity in the gills and kidney. The activity of carbonic anhydrase decreased in the gills after 24h and in both tissues after 96h of Cd exposure. The gill Ca(2+)-ATPase activity also decreased with Cd exposure, with a concomitant drop in the plasma concentration of Ca(2+). Despite the hypocalcemia, there were no changes in the concentration of the ions Na(+), K(+), and Cl(-) or in plasma osmolality. Among the enzymes involved in ion transport, H(+)-ATPase was the only enzyme that showed increased activity in gills, whereas its activity in kidney remained unchanged. The results of this study demonstrate that waterborne Cd at the maximum concentrations set by Brazilian guidelines for freshwater affects the gills and kidney functions of P. lineatus. Acute exposure to Cd resulted in the decrease of the activity of enzymes, which culminated with the loss of the fish's ability to regulate the levels of calcium in the blood, leading to hypocalcemia.

Nickel exposure promotes osmoregulatory disturbances in Oreochromis niloticus gills: histopathological and energy dispersive spectrometry analysis.

Water is an essential factor for maintaining the vital functions of living beings. Nickel is the 24th most abundant element on Earth; it is a heavy metal that is genotoxic and mutagenic in its chloride form. Due to industrial use, its concentration in surface sediments increased considerably. Fish develop characteristics that make them excellent experimental models for studying aquatic toxicology. They are particularly useful because they can alert of the potential danger of chemical substances or environmental pollution. Due to water quality impairment and because there are few published studies that relate nickel to tissue alteration, this study aimed to examine the consequences of nickel in an aquatic environment. For this analysis, individuals of Oreochromis niloticus were exposed for 96 h to three different concentrations of nickel dissolved in water according to the standard established by Brazilian law and compared them to a control group. After exposure, the gills were analyzed using X-ray microanalysis, ultramorphology, and histological and histochemical analysis. The results demonstrated that all the concentrations used in the experiment altered the histophysiology of the individuals exposed. In conclusion, the nickel presents a toxic potential to fish, even at the lowest concentration tested, which is equivalent to half of the concentration allowed by law. The CONAMA resolution should be revised for this parameter because of the interference of this metal in the histophysiology of the tested organism.

The effect of salinity on osmoregulation and development of the juvenile fat snook, Centropomus parallelus (POEY).

Eurihaline fish support waters with different salt concentration. However, numerous studies have shown that salinity can affect fish development. Thus, the effect of salinity change from 20 to 5 and 35 on survival, weight, length, gill chloride cell ultrastructure and gill Na+, K+ ATPase activity was evaluated in Centropomus parallelus following short-term (6, 24 and 96 hours) and long-term exposure (30 and 60 days). Salinity did not affect C. parallelus survival, final weight and length. The quantity of chloride cells increased visibly at salinities of 5 and 35, with the cells exhibiting the typical features of uptake and secretory cells, respectively. Na+, K+ ATPase activity in the gill of the C. parallelus was significantly greater at a salinity of 5 than at a salinity of 20 or 35 after 96 hours, but not after 30 or 60 days. These results indicate that salinity change from high to low salt water induces gill chloride cell and Na+, K+ ATPase activity adaptations after short-term exposure. However, after long-term exposure at salinity 5, gill Na+, K+ ATPase activity is no more necessary at high levels. The increase in salinity to 35 does not induce significant change in gills. Juveniles of C. parallelus may thus be capable of acclimating to salinities of 5 to 35 for 60 days without significant effects on development.

The effect of salinity on osmoregulation and development of the juvenile fat snook, Centropomus parallelus (POEY) / O efeito da salinidade sobre a osmoregulacao e o desenvolvimento do juvenil de robalo-peva, Centropomus parallelus (POEY)

Eurihaline fish support waters with different salt concentration. However, numerous studies have shown that salinity can affect fish development. Thus, the effect of salinity change from 20 to 5 and 35 on survival, weight, length, gill chloride cell ultrastructure and gill Na+, K+ ATPase activity was evaluated in Centropomus parallelus following short-term (6, 24 and 96 hours) and long-term exposure (30 and 60 days). Salinity did not affect C. parallelus survival, final weight and length. The quantity of chloride cells increased visibly at salinities of 5 and 35, with the cells exhibiting the typical features of uptake and secretory cells, respectively. Na+, K+ ATPase activity in the gill of the C. parallelus was significantly greater at a salinity of 5 than at a salinity of 20 or 35 after 96 hours, but not after 30 or 60 days. These results indicate that salinity change from high to low salt water induces gill chloride cell and Na+, K+ ATPase activity adaptations after short-term exposure. However, after long-term exposure at salinity 5, gill Na+, K+ ATPase activity is no more necessary at high levels. The increase in salinity to 35 does not induce significant change in gills. Juveniles of C. parallelus may thus be capable of acclimating to salinities of 5 to 35 for 60 days without significant effects on development.

Peixes eurihalinos suportam águas com diferentes concentrações de sal. Contudo, muitos estudos têm mostrado que a salinidade pode afetar o desenvolvimento do peixe. Portanto, o efeito da mudança de salinidade de 20 para 5 e 35 na taxa de sobrevivência, peso, comprimento, morfologia das células de cloreto branquiais e atividade da Na+, K+ ATPase foram avaliadas no Centropomus parallelus após curto (6, 24 e 96 horas) e longo tempo de exposição (30 e 60 dias). A salinidade não afetou a sobrevivência, o peso e comprimento final do robalo-peva. A quantidade de células de cloreto aumentou visivelmente nas salinidades 5 e 35, exibindo morfologias típicas de células que absorvem e secretam sal, respectivamente. A atividade da Na+, K+ ATPase nas brânquias do C. parallelus foi significativamente maior na salinidade 5 do que nas salinidades 20 ou 35 após 96 horas, mas não após 30 e 60 dias. Esses resultados indicam que a mudança de alta para baixa salinidade provoca adaptações nas células de cloreto e na atividade da Na+, K+ ATPase branquial em curto prazo. Contudo, após longa exposição na salinidade 5, a alta atividade da Na+, K+ ATPase branquial não é mais necessária. O aumento de salinidade para 35 não induz mudanças significativas nas brânquias. Portanto, juvenis de C. parallelus possuem a capacidade de aclimatação nas salinidades de 5 a 35 semefeitos significativos no desenvolvimento após 60 dias.