Gastro-intestinal transport of calcium and cadmium in fresh water and seawater acclimated trout (Oncorhynchus mykiss)

Transport of calcium (Ca) and cadmium (Cd) was examined along the gastro-intestinal tract (GIT) of freshwater and seawater Oncorhynchus mykiss irideus (FWT and SWTies respectively) using in vitro and in vivo experiments. Based on known physiological differences between FWT and SWT which aid in regulating ion levels and osmolarity, we hypothesized that SWT would have lower rates of Ca uptake. Also, we predicted that Cd rates would also be lower because Cd is known to share a common transport mechanism with Ca. Kinetics of Ca and Cd transport were determined using mucosal salines of varying concentrations [1, 10, 30, 60, and 100 (mmol L^− 1 for Ca, μmol L^− 1 for Cd)]. Linear and saturating relationships were found for Ca for FWT and SWT, but overall SWT had lower rates. Linear and/or saturating relationships were also found for Cd uptake, but rates varied little between fish types. Elevated Ca had no inhibitory effect on Cd transport, and Ca channel blockers nifedipine and verapamil had little effect on Ca or Cd uptake. However, lanthanum reduced Ca transport into some compartments. A 21 day in vivo feeding experiment was also performed where FWT and SWT were exposed to control diets or Cd-spiked diets (552 μg Cd g^− 1 food). Whole body Cd uptake between fish types was similar, but the majority of Cd in SWT remained in the posterior intestine tissue, while FWT transported more Cd through their gut wall. Overall it appears that large differences in Ca and Cd uptake between FWT and SWT exist, with SWT generally having lower rates.     

Induction of cytochrome P450 1A and DNA damage in isolated rainbow trout (Oncorhynchus mykiss) hepatocytes by 2,3,7,8 tetrachlorodibenzo-p-dioxin

Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on hepatocytes isolated from immature rainbow trout (Oncorhynchus mykiss) by collagenase perfusion were investigated with respect to induction of cytochrome P450 1A (CYP1A) enzyme activities and protein contents as well as DNA damage. Exposure of primary rainbow trout hepatocytes to TCDD resulted in increased CYP1A contents, as determined by immunoblotting, enhanced activities of 7-ethoxyresorufin-O-deethylase (EROD) and increased DNA damage as determined by the comet assay. By means of electron microscopy, no symptoms of cytotoxicity could be observed except for slight increases of lysosomal components and the smooth endoplasmic reticulum. Whereas CYP1A contents constantly increased over the duration of the entire experiment, EROD activities remained constant from day 3 of exposure to 1 nM TCDD; maximum induction of CYP1A activities was reached with 0.1 nM TCDD after 5 days. DNA damage increased in a time- and dose-dependent fashion until day 3. After 5 days, DNA damage was less pronounced, and the number of damaged nuclei declined in all TCDD concentrations. Since TCDD has been shown to not directly react with DNA, metabolism of TCDD or TCDD-induced changes in other metabolic pathways are suspected to result in the production of DNA-reactive (endogenous) substances.     

Tissue damage in organic rainbow trout muscle investigated by proteomics and bioinformatics

The response to tissue damage is a complex process, which involves the coordinated regulation of multiple proteins to ensure tissue repair. In order to investigate the effect of tissue damage in a lower vertebrate, samples were taken from rainbow trout (Oncorhynchus mykiss) at day 7 after damage and proteins were separated using 2DE. The experimental design included two groups of rainbow trout, which were fed organic feed either with or without astaxanthin. In total, 96 proteins were found to be affected by tissue damage, clearly demonstrating in this lower vertebrate the complexity and magnitude of the cellular response, in the context of a regenerative process. Using a bioinformatics approach, the main biological function of these proteins were assigned, showing the regulation of proteins involved in processes such as apoptosis, iron homeostasis, and regulation of muscular structure. Interestingly, it was established that exclusively within the astaxanthin feed group, three members of the annexin protein family (annexin IV, V, and VI) were regulated in response to tissue damage.     

Strain variability in fatty acid composition of Chattonella marina (Raphidophyceae) and its relation to differing ichthyotoxicity toward rainbow trout gill cells

Lipid profiles of three strains (Mexico, Australia, Japan) of Chattonella marina (Subrahmanyan) Hara et Chihara were studied under defined growth (phosphate, light, and growth phase) and harvest (intact and ruptured cells) conditions. Triacylglycerol levels were always <2%, sterols <7%, free fatty acids varied between 2 and 33%, and polar lipids were the most abundant lipid class (>51% of total lipids). The major fatty acids in C. marina were palmitic (16:0), eicosapentaenoic (EPA, 20:5ω3), octadecatetraenoic (18:4ω3), myristic (14:0), and palmitoleic (16:1ω7c) acids. Higher levels of EPA were found in ruptured cells (21.4–29.4%) compared to intact cells (8.5–25.3%). In general, Japanese N‐118 C. marina was the highest producer of EPA (14.3–29.4%), and Mexican CMCV‐1 the lowest producer (7.9–27.1%). Algal cultures, free fatty acids from C. marina, and the two aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal (suspected fatty acid‐derived products) were tested against the rainbow trout fish gill cell line RTgill‐W1. The configuration of fatty acids plays an important role in ichthyotoxicity. Free fatty acid fractions, obtained by base saponification of total lipids from C. marina showed a potent toxicity toward gill cells (median lethal concentration, LC₅₀ (at 1 h) of 0.44 μg · mL^?1 in light conditions, with a complete loss of viability at >3.2 μg · mL^?1). Live cultures of Mexican C. marina were less toxic than Japanese and Australian strains. This difference could be related to differing EPA content, superoxide anion production, and cell fragility. The aldehydes 2E,4E‐decadienal and 2E,4E‐heptadienal also showed high impact on gill cell viability, with LC₅₀ (at 1 h) of 0.34 and 0.36 μg · mL^?1, respectively. Superoxide anion production was highest in Australian strain CMPL01, followed by Japanese N‐118 and Mexican CMCV‐1 strains. Ruptured cells showed higher production of superoxide anion compared to intact cells (e.g., 19 vs. 9.5 pmol · cell^?1 · hr^?1 for CMPL01, respectively). Our results indicate that C. marina is more ichthyotoxic after cell disruption and when switching from dark to light conditions, possibly associated with a higher production of superoxide anion and EPA, which may be quickly oxidized to produce more toxic derivates, such as aldehydes.     

Trophic interference by Salmo trutta on Aplochiton zebra and Aplochiton taeniatus in southern Patagonian lakes

The length and mass ratio, diet and isotopic composition of Aplochiton zebra and Aplochiton taeniatus inhabiting a Salmo trutta‐invaded and a S. trutta‐free lake in southern Patagonia were compared. Results indicate that S. trutta exercises important trophic interference over A. zebra and A. taeniatus, causing changes in their dietary composition by reducing the consumption of winged Diptera through changes in feeding behaviours that involve jumping out of the water. This effect is significantly higher in A. zebra than in A. taeniatus a species that has a highly specialized diet. The dietary changes of A. zebra and A. taeniatus in sympatry with S. trutta lead to an impoverishment of their isotopic nitrogen signals (δ¹⁵N), suggesting a reduction of their trophic position. In the case of A. zebra, this translates into a significant decrease in its body condition factor. Such interference could lead to a population decline of this species and would explain the current distribution range decline and allopatry with S. trutta in fluvial systems.     

Dietary Modifications of Phospholipid Composition and Biophysical Properties of the Brush Border Membrane Along the Trout Intestine

Brush border membranes (BBM) are isolated from middle and posterior intestine of trout fed either an essential fatty acid-rich diet or a saturated one. The different phospholipid classes are separated, and their fatty acid composition is determined. Fluorescence anisotropy studies are performed using two lipid fluorophores, namely diphenylhexatriene (DPH) and trimethyl-aminodiphenylhexatriene (TMA-DPH). The results indicate that the usual parameters affecting the lipid fluidity such as the phospholipid:protein (PL:PROT), cholesterol:phospholipid (CHOL:PL), and sphingomyelin:phosphatidylcholine (SP:PC) ratios and the unsaturation of the acyl chains are sufficient to explain the fluidity values determined using DPH, but not those obtained with TMA-DPH as a probe. This fluorophore is assessed to be localized only in the external leaflet of the membrane. Hence, it will be affected by the composition of the major phospholipids of this leaflet, sphingomyelin and phosphatidylcholine.     

Copper exposure and the degeneration of olfactory receptors in rainbow trout (Oncorhynchus mykiss)

Abstract

Sublethal concentrations of copper in water cause the degeneration of olfactory receptors in rainbow trout (Oncorhynchus mykiss). Receptor cell loss has been correlated to the loss of olfaction in fish and may cause difficulties in olfactory mediated behaviors such as migration. This study investigated the effects of three levels of copper (100, 75 and 50 mg L^?1) on the olfactory epithelium of rainbow trout. Twenty fish randomly allocated between three exposure groups and one control were exposed for 24 hours under static renewal conditions. Light and scanning electron microscopic observations of olfactory tissue were taken to determine the extent of degeneration of receptors. In addition, levels of copper and zinc in the brain tissues were analyzed to determine if the olfactory route was a significant route of copper exposure and transfer to fish brain tissue. Results indicate that degeneration of receptors is related to the concentration of copper. Levels of copper in brain were found to be below detection of the instrument. Levels of zinc were extremely variable ranging from 52 to 132 ng zinc g^?1 brain tissue.     

Purification and Characterization of Glycoprotein from the Skin Mucus of the Rainbow Trout,Salmo gaivdneri

Mucus glycoprotein (RGP) was purified and characterized from the skin mucus of rainbow trout, Salmo gairdneri. RGP was found to contain 30.1% NeuAc, 26.0% GalNAc, 5.0% Gal, and 26.0% amino acids. The protein moiety of RGP is very rich in Thr (32.4 mol%). Neither NeuGc nor KDN (2-keto-3-deoxy-d-glycero-d-galacto-nononic acid) was found in RGP. Alkaline borohydride treatment of RGP yielded a major disaccharide alditol, NeuAcα2→6GalNAc-ol and more than 4 minor oligosaccharide alditols including NeuAc→(GalNAcα1→)GalNAc-ol. It was evident that an average RGP molecule has approximately 500 NeuAc-containing oligosaccharide chains, which are attached to the Thr and Ser residues of the protein moiety and spaced at an average of 3 amino acids apart.     

Oscillations in Superoxide Anion Release by Polymorphonuclear Leukocytes and its Inhibition by Human Saliva

The release of superoxide anion (O 2 -) by inflammatory polymorphonuclear leukocytes (PMN), obtained from the intraperitoneal cavity of mice, was investigated during two different times of the day. NaF-stimulated PMN at nigth-phase showed greater release of O 2 - than at day. Addition of human saliva in the culture medium dramatically inhibits O 2 - generation by both day- and night-phase cells. Also, the inhibitory effect of diurnal saliva on the cell activation is higher than the nocturnal saliva.     

Diurnal rhythms of plasma growth hormone,somatostatin, thyroid hormones,cortisol and glucose concentrations in rainbow trout,Oncorhynchus mykiss,during progressive food deprivation

Abstract

The diurnal patterns of changes in plasma cortisol, growth hormone (GH), somatostatin‐14 (SRIF), thyroid hormones (L‐thyroxine, T₄ and triiodo‐L‐thyronine, T₃) and glucose were investigated in rainbow trout, Oncorhynchus mykiss, both during single meal‐feeding and during a progressive fast of 13 weeks. All measured variables exhibited a diurnal pattern in fed rainbow trout, most of which appeared to be correlated with the time of feeding (30–60 min after the onset of light), while additional changes, associated with the scotophase were also found for cortisol. Although fasting had no affect on mean daily plasma cortisol or SRIF concentrations, there was a progressive increase in mean daily plasma GH concentrations and a progressive decrease in mean daily plasma thyroid hormone and glucose concentrations associated with fasting. However, for GH, significant changes were not evident until week 10 of the fast. In addition, fasting appeared to phase‐shift the diurnal patterns of plasma GH, cortisol and glucose concentrations; the consequence of such a shift is that conclusions as to the effects of fasting, if based on a single time of sampling, may be flawed.