Effect of total body lipid on the toxicity and toxicokinetics of pentachlorophenol in rainbow trout (Oncorhynchus mykiss)

Juvenile rainbow trout were fed high fat (21%) or low fat (13%) practical diets for 11 wk in order to establish a significant difference in total body lipid; a difference of 3% was obtained. An acute lethality test (144 h) was performed on the two diet groups using pentachlorophenol (PCP). Elimination rate constants (k₂) and acute lethality thresholds were estimated by fitting a one-compartment, first-order bioconcentration model to both the LC50/time data, and the time-to-death/water-concentration data. Estimates using these two methods were, respectively, 2.32 and 2.29 d⁻¹ for the high fat treatment, and 2.84 and 2.66 d⁻¹ for the low fat treatment. There was no significant difference in the resistance of fish to PCP due to lipid or weight as determined by a regression of time-to-death on these variables. Whole-body PCP levels, were measured in individual fish that died during the bioassay. Mean lethal body residues were found to range from 0.08 to 0.15 mmol PCP kg⁻¹ and significant differences were found between both diet treatments and exposure concentrations. Analysis of covariance on the lethal residue data showed that lipid levels interacted significantly with exposure concentration, suggesting that the fish do not act as an ideal single compartment. Correction of PCP residues for fish lipid content showed that some of the variation in lethal body burdens could be accounted for by lipid content. In fish which survived the lethality bioassay at 91 μg 1⁻¹, lipid levels were significantly decreased compared to initial levels. At the 91 1⁻¹ μg exposure concentration PCP residues were not correlated with the body lipid content.     

Xanthate effects on cadmium intracellular distribution in rainbow trout (Oncorhynchus mykiss) gills

The uptake of ¹⁰⁹cadmium through perfused rainbow trout gills in the presence of xanthates was studied, and the subcellular distribution of cadmium in perfused gill tissue was determined. Pnenol absorption was also studied because xanthates form hydrophobic Cd complexes with a log P_{octanol/water} similar to that of phenol.

1. Xanthate concentrations higher than 10⁻⁵ M increased the rate of cadmium transfer through the gills and cadmium retention in gill tissue. Cadmium was present as a hydrophobic complex at this and higher xanthate concentrations.

2. A redistribution of cadmium from metallothionein to high molecular weight cadmium binding fractions occurred in the presence of 10⁻⁴ M xanthate.

3. The rate of phenol transfer across the gill epithelium was much higher than the rate of cadmium transfer regardless of whether xanthate was present. The rate of phenol transfer stabilized much faster than the rate of cadmium transfer irrespective of whether xanthate was present, indicating that different uptake mechanisms were involved.

We conclude that in the presence of xanthate concentrations higher than 10⁻⁵ M cadmium is taken up as a hydrophobic Cd(xanthate)₂ complex by the epithelial cells. Within the cell the complex dissociates, and the metal ion is bound to intracellular cadmium-binding ligands. The metal is probably translocated through the basolateral membrane as a free ion.     

Renal responses to acute lead waterborne exposure in the freshwater rainbow trout (Oncorhynchus mykiss)

The possible nephrotoxic effects of waterborne lead exposure (as Pb(NO₃)₂) were investigated in the freshwater rainbow trout (Oncorhynchus mykiss). Kidney lead accumulation was time-dependent, increasing upon exposure to 0.57 ± 0.01 mg dissolved Pb L⁻¹ for up to 96 h with a significantly higher burden occurring in the posterior kidney compared to the anterior segment. Urine analyses in trout exposed to 1.20 ± 0.09 mg dissolved Pb L⁻¹ revealed a significant increase in urinary lead excretion rate throughout 96 h of exposure. Urine flow rate and glomerular filtration rate (GFR) were not impacted with the exception of a significant decrease in GFR from 84 to 96 h in lead-exposed trout. Urine pH decreased significantly over time in lead-exposed fish. Correspondingly, urine ammonia excretion rate showed a marked increase from 48 h onwards. In experimental fish, urine glucose excretion was significantly greater by 96 h while urine lactate, urea and protein excretion were not significantly altered by lead exposure. The urine excretion rate of Ca²⁺ increased significantly by approximately 43% after only 24 h of lead exposure, and was maintained at a higher rate than controls for up to 96 h. Magnesium excretion increased in a time-dependent fashion, reaching a two- to three-fold rise by 96 h. In contrast, rates of Na⁺ and Cl⁻ excretion were decreased in experimental fish by approximately 30% by 48 h, this trend continuing for the duration of lead-exposure. There were no changes in any of these parameters in similarly treated control fish. Clearance ratio analyses indicated progressive decreases in the net reabsorption efficiencies of the renal system for Ca²⁺, Mg²⁺, Pb, and glucose, suggesting that the active tubular transport mechanisms for these substances were inhibited by lead exposure, while Na⁺, K⁺, Cl⁻, lactate, and protein reabsorptions were unaffected. Net ammonia secretion increased. We conclude that changes in renal function both reflect and help to minimize some of the associated disturbances in systemic physiology. Lead-induced ionoregulatory toxicity in rainbow trout, particularly the disturbance of Ca²⁺ homeostasis, is not exclusively a branchial phenomenon, but is in part a result of disruption of ionoregulatory mechanisms at the kidney. This action of lead outside the gills is critical to consider when developing guidelines for water quality.     

Experimentally determined blood and water flow limitations for uptake of hydrophobic compounds using perfused gills of rainbow trout (Oncorhynchus mykiss): Allometric applications

The influence of water and blood flows on the uptake of a number of hydrophobic compounds was investigated using perfused gills of rainbow trout. For all compounds studied, the uptake rate constants increased with water flow between 0.045 and 0.52 L min⁻¹ kg⁻¹, and remained constant at higher flow. The uptake rate constants did not change when blood flow decreased from 10 to 4.4 mL min⁻¹ kg⁻¹, while they doubled when blood flow increased from 10 to 20 mL min⁻¹ kg⁻¹. Both water and blood flows thus influence the uptake of hydrophobic chemicals. Using allometric relations, it can be established that the water flow will limit the uptake of hydrophobic chemicals for fish weighing more than 5 g. The flow of water will not limit uptake in fish < 5 g, irrespective of physiological conditions and oxygen concentration. At low oxygen concentration, which will increase the water flow in large fish, the uptake of hydrophobic chemicals may increase with a factor of 5 or more. Increasing the blood flow may maximally increase the uptake of hydrophobic chemicals two-fold, in small as well as in large fish.     

Histological alterations in gills and liver of rainbow trout (Oncorhynchus mykiss) after exposure to the antibiotic oxytetracycline

The aim of this study was to investigate the histopathological effects of oxytetracycline (OTC) on the gill and liver tissues of rainbow trouts (Oncorhynchus mykiss) following acute (96 h: 0.005–50 mg/L) and chronic (28 days: 0.3125–5 μg/L) exposures. Results suggest the existence of a cause-and-effect relationship between the exposure to OTC and tissue damage. Most predominant disorders observed in gills were progressive (e.g. hypertrophy of mucous cells and hyperplasia of epithelial cells) in acute exposure and regressive (e.g. lamellar fusion, epithelial lifting of lamellae and some changes in tissue architecture) in chronic exposure. However, only the acute exposure was responsible for a significant increase of the total gill pathological index. PAGE index, reflecting the extent of gill tissue available for gas exchanges in fish, remained unchanged for both exposures. In liver, circulatory (e.g. hemorrhage and increase of sinusoidal space), regressive (e.g. pyknotic nucleus, vacuolization and hepatocellular degenerations) and progressive (e.g. hypertrophy of hepatocytes) changes were observed, but just after acute exposure. After chronic exposure, only inflammatory changes (e.g. leucocytes infiltration) were observed. Following both exposures, a significant increase of the total liver pathological index was recorded. Despite the increase of the histological damage in individuals exposed to OTC, lesions observed were of minimal or moderate pathological importance, non-specific and reversible. The data gathered following acute and chronic exposures also suggest the onset of adaptive mechanisms of fish, namely for longer exposure periods. Furthermore the observed histological alterations appear to be result of several physio-metabolic disorders consequence of the biochemical and molecular modes of action of OTC.     

Inhibitory effects of ammonia and urea on gill carbonic anhydrase enzyme activity of rainbow trout (Oncorhynchus mykiss)

The effects of ammonia and urea on branchial carbonic anhydrase (CA) enzyme which plays a key role in ionoregulation, osmoregulation and acid-base balance of rainbow trout (Oncorhynchus mykiss) were investigated. CA activity of the control group for ammonia and urea was determined as 1285.7 ± 67.9 and 1261.7 ± 60.8 EU/mg protein, respectively. The CA enzyme activities of the other groups were measured at 1, 2 and 3 h after ammonia and urea applications. The corresponding activities of ammonia were 774.9 ± 68.8, 732.1 ± 48.6 and 768.1 ± 59.5 EU/mg protein, respectively and that of urea were 769.3 ± 58.9, 638.2 ± 47.7 and 1108.1 ± 61.1 EU/mg protein, respectively. The differences between the initial CA activities for the controls was not significantly (P > 0.01). The CA activities were significantly (P < 0.01) inhibited both in ammonia and urea group. However, the ammonia inhibited more than urea since there was significant differences between final values of gill CA activities.     

Effects of atrazine on hepatic metabolism and endocrine homeostasis in rainbow trout (Oncorhynchus mykiss)

The herbicide atrazine (ATZ) is one of the most widely used pesticides in the world and is now under scrutiny for its alleged capacity to disrupt the endocrine system. Exhibiting negligible interaction with the estrogen receptor (ER), ATZ's mode of action remains to be elucidated. ATZ may act as an inducer of the enzyme aromatase, which converts androgens to estrogens, although other mechanisms should also be taken into consideration such as impairment of hepatic metabolism. Therefore we administered juvenile rainbow trout (Oncorhynchus mykiss) a dose of either 2 or 200 μg ATZ/kg, or of carrier control phosphate buffered saline (PBS) and we measured plasma concentrations of testosterone (T), 17beta-estradiol (E2) and vitellogenin (Vtg) 6 days after exposure. Simultaneously we analyzed hepatic gene expression of cytochrome P450 (CYP) 1A and pi-class glutathione S-transferase (GST-P), and catalase (CAT) activity. Although sex steroid levels showed no significant alterations, we found a dose-dependent increase in Vtg and a concomitant decrease in CYP1A. There was no effect of ATZ on GST-P mRNA levels but GST-P was positively correlated with CYP1A. Also, CYP1A was negatively correlated with liver CAT and E2, and varied with T concentrations in a hormetic manner. The results showed that ATZ can alter hepatic metabolism, induce estrogenic effects and oxidative stress in vivo, and that these effects are linked.     

Effects of long-term nonylphenol exposure on gonadal development and biomarkers of estrogenicity in juvenile rainbow trout Oncorhynchus mykiss

Environmental pollutants with estrogenic activity including nonylphenol (NP) have the potential to alter gonadal development and reproduction of wild fish. To investigate the estrogenic action of environmentally relevant concentrations of NP, rainbow trout (Oncorhynchus mykiss) were continuously exposed during the embryonic, larval and juvenile life stage to 1.05 and 10.17 μg/l NP for 1 year, and sexual differentiation, vitellogenin (VG), VG mRNA, and zona radiata protein (ZRP) expression were examined after that period. The applied NP concentrations did not affect mortality and hatching rates, and did not have an influence on the body weight of 1-year-old fish. No occurrence of testis–ova was observed and sex-ratios of NP exposed groups of fish were unchanged when compared with control groups. The induction of VG and ZRP expression was a more sensitive reaction to the presence of NP than the formation of testis–ova and the reversal of sex. Increased VG expression in trout liver occurred already at 1.05 μg/l NP, whereas VG mRNA levels, quantified by competitive RT-PCR, were not significantly elevated in NP exposed fish. ZRP contents were significantly higher at 10.17 μg/l NP. Since induction of VG did not occur in all fish exposed to 1 or 10 μg/l NP and ZRP induction did not occur in all fish exposed to 10 μg/l, some individuals may be more affected by exposure to NP than others. This study demonstrates that NP concentrations typically found in sewage treatment effluents and some rivers do not affect sexual differentiation in rainbow trout, but induce VG and ZRP expression in the liver of exposed fish.     

Bioaccumulation and biotransformation of chiral triazole fungicides in rainbow trout (Oncorhynchus mykiss)

There are very little data on the bioaccumulation and biotransformation of current-use pesticides (CUPs) despite the fact that such data are critical in assessing their fate and potential toxic effects in aquatic organisms. To help address this issue, juvenile rainbow trout (Oncorhynchus mykiss) were exposed to dietary concentrations of a mixture of chiral triazole fungicides (bromuconazole, cyproconazole, metconazole, myclobutanil, penconazole, propiconazole, tebuconazole, tetraconazole, and triadimefon) and a chiral legacy pesticide [-hexachlorocyclohexane (-HCH)] to study the bioaccumulation and biotransformation of these CUPs. Fish accumulated all triazoles rapidly during the 8 day uptake phase, and was followed by rapid elimination, which was estimated by taking accelerated sampling times during the 16 day depuration period. Half-lives (t_1/2s) and times to 95% elimination (t₉₅s) ranged from 1.0 to 2.5 and 4.5 to 11.0 days, respectively. Chiral analysis suggested no significant selectivity in biotransformation for most of the compounds based on statistically unaltered enantiomer fractions (EFs) in the fish compared to food values; exceptions were a change in EF of myclobutanil and changes in diastereomer fractions (DFs) of propiconazole and cyproconazole. No biotransformation was observed for -HCH based on consistent EFs in the fish throughout the experiment and a t_1/2 (15.8 days) that fell within the 95% confidence interval of a log K_ow–log t_1/2 relationship developed for assessing biotransformation of organic contaminants. This relationship did show that biotransformation accounted for the majority (ranging from 59.9 to 90.4%) of the elimination for all triazoles, and that triazole compounds with oxygen and hydroxyl functional groups were more easily biotransformed. This research indicated that chiral analysis may potentially miss biotransformation of CUPs and other potential non-persistent organic contaminants and shows the utility of the log K_ow–log t_1/2 relationship as a mechanistic tool for quantifying biotransformation. Based on the rapid biotransformation of the triazoles, future research should focus on formation of metabolites and their fate and possible effects in the environment.     

An in vitro biotic ligand model (BLM) for silver binding to cultured gill epithelia of freshwater rainbow trout (Oncorhynchus mykiss)

"Reconstructed" gill epithelia on filter supports were grown in primary culture from dispersed gill cells of freshwater rainbow trout (Oncorhynchus mykiss). This preparation contains both pavement cells and chloride cells, and after 7-9 days in culture, permits exposure of the apical surface to true freshwater while maintaining blood-like culture media on the basolateral surface, and exhibits a stable transepithelial resistance (TER) and transepithelial potential (TEP) under these conditions. These epithelia were used to develop a possible in vitro version of the biotic ligand model (BLM) for silver; the in vivo BLM uses short-term gill binding of the metal to predict acute silver toxicity as a function of freshwater chemistry. Radio-labeled silver ((110m)Ag as AgNO(3)) was placed on the apical side (freshwater), and the appearance of (110m)Ag in the epithelia (binding) and in the basolateral media (flux) over 3 h were monitored. Silver binding (greater than the approximate range 0-100 mug l(-1)) and silver flux were concentration-dependent with a 50% saturation point (apparent K(d)) value of about 10 mug l(-1) or 10(-7) M, very close to the 96-h LC50 in vivo in the same water chemistry. There were no adverse effects of silver on TER, TEP, or Na(+), K(+)-ATPase activity, though the latter declined over longer exposures, as in vivo. Silver flux over 3 h was small (<20%) relative to binding, and was insensitive to water chemistry. However, silver binding was decreased by elevations in freshwater Na(+) and dissolved organic carbon (humic acid) concentrations, increased by elevations in freshwater Cl(-) and reductions in pH, and insensitive to elevations in Ca(2+). With the exception of the pH response, these effects were qualitatively and quantitatively similar to in vivo BLM responses. The results suggest that an in vitro BLM approach may provide a simple and cost-effective way for evaluating the protective effects of site-specific waters.     

The organochlorine o,p'-DDD disrupts the adrenal steroidogenic signaling pathway in rainbow trout (Oncorhynchus mykiss)

The mechanisms of action of o,p'-DDD on adrenal steroidogenesis were investigated in vitro in rainbow trout (Oncorhynchus mykiss). Acute exposures to o,p'-DDD inhibited ACTH-stimulated cortisol secretion while cell viability decreased significantly only at the highest concentration tested (200 microM o,p'-DDD). Stimulation of cortisol secretion with a cAMP analogue (dibutyryl-cAMP) was inhibited at a higher concentration than that needed to inhibit ACTH-stimulated cortisol synthesis in cells exposed to o,p'-DDD. Forskolin-stimulated cortisol secretion and cAMP production, and NaF-stimulated cAMP production were inhibited in a concentration-dependent manner by o,p'-DDD. In contrast, basal cortisol secretion was stimulated while basal cAMP production was unaffected by o,p'-DDD. Pregnenolone-stimulated cortisol secretion was enhanced by o,p'-DDD at a physiologically relevant pregnenolone concentration, while o,p'-DDD inhibited cortisol secretion when a pharmacological concentration of pregnenolone was used. Our results suggest that the cAMP generation step is a target in o,p'-DDD-mediated disruption of ACTH-stimulated adrenal steroidogenesis in rainbow trout but that other downstream targets such as steroidogenic enzymes responsible for cortisol synthesis might also be affected.     

Molecular dosimetry of DNA adducts in rainbow trout (Oncorhynchus mykiss) exposed to benzo(a)pyrene by different routes

 Farm raised rainbow trout (Oncorhynchus mykiss) were exposed by various routes to benzo(a)pyrene (BP) as a representative carcinogenic polycyclic aromatic hydrocarbon (PAH). Following exposure of fish to the chemical by intraperitoneal (i.p.) injection, ³²P-postlabelling studies indicated that non-feral trout were relatively resistant to the formation of BP-DNA adducts in liver. No adducts were detected in fish exposed to single doses (20 mg/kg) of BP. Multiple exposures (e.g. 2×25 mg/kg) were necessary in order for adducts to be detected, indicating that induction of the metabolising enzymes required for the bioactivation of BP is necessary. These studies provided reference information on DNA adducts for comparison with data from subsequent experiments at environmentally realistic low level exposures. Two types of low level aquatic exposure were carried out. The first procedure exposed fish for 30 days to a nominally constant low level (1.2 and 0.4 μg/l) of a homogeneous dispersion of BP in water, to simulate low level aquatic environmental exposures. Following ³²P-postlabelling analysis of the liver DNA of exposed fish, BP-DNA adducts were not detected. In the second procedure, fish were exposed to a constant low level of BP (ca. 0.5 μg/l) for 15 days then to a pulse (60 μg/l) which was allowed to naturally decline (to ca. 2 μg/l) during a further 15 days. Following this exposure, significant levels of BP-DNA adducts were detected in livers of trout. The effect of dietary exposures was investigated by feeding trout a diet containing either 58 μg or 288 μg BP per day for 6 days, equivalent to total doses of 43 mg/kg and 216 mg/kg. In both cases BP-DNA adducts were detected in livers of exposed fish. The results provide useful information on the types of exposures to PAHs which may pose a genotoxic risk to fish in the environment. Received: 5 April 1994 / Accepted: 31 May 1994     

Effects of salinity on aldicarb toxicity in juvenile rainbow trout (Oncorhynchus mykiss) and striped bass (Morone saxatilis x chrysops).

Fluctuations in several environmental variables, such as salinity, can influence the interactions between organisms and pollutants in aquatic organisms, and, therefore, affect the toxicity of xenobiotics. In this study, after 2 species of fish, rainbow trout (Oncorhynchus mykiss) and hybrid striped bass (Morone saxatilis x chrysops) were acclimated to 4 salinity regimens of 1.5, 7, 14, and 21 ppt for 1 week and then exposed to 0.5 mg/l aldicarb. Mortality, brain, and muscle cholinesterase levels were measured after 96 h. Rates of (14)C-aldicarb sulfoxide formation were determined in kidney (trout only), liver, and gill microsomes from each species acclimated to the 4 salinity regimens. Salinity significantly enhanced aldicarb toxicity, cholinesterase inhibition, and (14)C-aldicarb sulfoxide formation in rainbow trout but not in striped bass. In vitro incubations with (14)C-aldicarb and the cytochrome P450 (CYP) inhibitor, N-benzylimidazole, did not significantly alter aldicarb sulfoxide formation in tissue microsomes from either species of fish, indicating CYP did not contribute to aldicarb sulfoxidation. Salinity increased flavin-containing monooxygenase (FMO) mRNA expression and catalytic activities in microsomes of liver, gill, and kidney of rainbow trout, which was consistent with the salinity-induced enhancement of aldicarb toxicity. Salinity did not alter FMO mRNA expression and catalytic activities in striped bass, which was also consistent with the lack of an effect of salinity on aldicarb toxicity in this species. These results suggest that salinity-mediated enhancement of aldicarb toxicity is species-dependent, and at least partially due to the salinity-related upregulation of FMOs, which, in turn, increases the bioactivation of aldicarb to aldicarb sulfoxide, which is a more potent inhibitor of cholinesterase than aldicarb.     

Role of glutathione redox cycle and catalase in defense against oxidative stress induced by endosulfan in adrenocortical cells of rainbow trout (Oncorhynchus mykiss)

The role of antioxidants in maintaining the functional integrity of adrenocortical cells during in vitro exposure to endosulfan, an organochlorine pesticide, was investigated in rainbow trout (Oncorhynchus mykiss). Aminotriazole (ATA), an inhibitor of catalase (CAT), L-buthionine sulfoximine (L-BSO), an inhibitor of glutathione (GSH) synthesis, and N-acetyl cysteine (NAC), a glutathione precursor, were used to investigate the role of CAT and GSH redox cycle in protection against the adrenal toxicity of endosulfan, a pesticide that impairs cell viability (LC50 366 microM) and cortisol secretion (EC50 19 microM) in a concentration-related manner. Pretreatment with ATA and L-BSO enhanced the toxicity of endosulfan (LC50 and EC50, respectively, 302 and 2.6 microM with ATA, 346 and 3.1 microM with L-BSO), while pretreatment with NAC had no significant effect on cell viability and increased the EC50 of endosulfan to 51 microM. CAT activity was significantly reduced following exposure to endosulfan when cells were pretreated with ATA. Pretreatment with L-BSO significantly decreased glutathione peroxidase (GPx) activity and reduced glutathione (GSH) levels in a concentration-related manner following exposure to endosulfan, while GSH levels were significantly higher in NAC pretreated cells compared to untreated cells. Finally, pretreatment with ATA and L-BSO increased, while pretreatment with NAC decreased, lipid hydroperoxides (LOOH) levels. CAT, GPx, and GSH were identified as important antioxidants in maintaining the function and integrity of rainbow trout adrenocortical cells and ATA, L-BSO, and NAC were identified as effective modulators of CAT and GSH redox cycle. Moreover, this study suggests that the glutathione redox cycle may be more efficient than catalase in protecting adrenocortical cells against endosulfan-induced oxidative stress.     

Effects of anesthesia (tricaine methanesulfonate,MS222) on liver biotransformation in rainbow trout (Oncorhynchus mykiss)

The effect of tricaine methanesulfonate (MS222) on rainbow trout liver biotransformation rates was investigated with a microsomal model; an in vitro preparation that can be employed with or without the use of an anaesthetic. Two experimental sets of rainbow trout microsomes were tested; one representing in vivo or surgical tricaine exposures and the other representing in vitro tissue/organ collection tricaine exposures. Microsomal incubations were performed on these two experimental groups with phenol as substrate to assess the effects of tricaine on Phase I (ring-hydroxylation) and II (glucuronidation) liver biotransformation by monitoring production of hydroquinone (HQ), catechol (CAT), and phenylglucuronide (PG). The use of a 2-h 100 mg/l exposure of tricaine for surgical anesthesia with or without 24-h recovery did not significantly (P< or =0.05) affect rates of phenol (Phase I and II) biotransformation rates; nor, did the 5-min 300 mg/l tricaine exposure for isolated organ/tissue collection significantly (P< or =0.05) affect phenol (Phase I and II) biotransformation rates. There were also no significant statistical differences (P< or =0.05) in P450 protein levels, or 7-ethoxyresorufin-O-deethylase (EROD) activity in these microsomal assays between any of the tricaine treated rainbow trout and controls.     

Features of cadmium and calcium uptake and toxicity in rainbow trout (Oncorhynchus mykiss) mitochondria

Molecular features of cadmium (Cd) and calcium (Ca) uptake and toxicity in rainbow trout liver mitochondria were studied using modulators of mitochondrial permeability transition pore (MPTP), mitochondrial calcium uniporter (MCU) and rapid uptake mode (RaM). Malate-glutamate energized mitochondria were exposed to 20μM Cd and 50μM Ca, singly and in combination, with and without addition of ruthenium red (RR), cyclosporin A (CsA), bongkrekic acid (BKA) or dithiothreitol (DTT). State 3 mitochondrial respiration was inhibited by 50% by either Cd or Ca, and by 70% when the two cations were added simultaneously. All the modulators tested reduced the inhibition of state 3 respiration with DTT completely reversing the Cd effect. While state 4 respiration was unaffected by Ca and/or Cd, 1.5-3 fold stimulation was observed on addition of the modulators. Uncoupler-stimulated respiration was inhibited by Cd, Ca and Cd+Ca with complete (DTT) and partial (RR, CsA, BKA) protection of the Cd and Cd+Ca effects. All the modulators completely reversed the Ca-induced inhibition. Swelling, the hallmark of MPTP, measured following incubation of mitochondria with 0-100μM of the two cations, singly and in combination, was abolished by all the modulators. Overall these data show the existence of membrane channels in rainbow trout liver mitochondria with some characteristics similar to mammalian MPTP, MCU and RaM. Moreover, entry of Ca and Cd into mitochondria is important in the toxicity of these cations.     

Mechanisms of fenthion activation in rainbow trout (Oncorhynchus mykiss) acclimated to hypersaline environments

Previous studies in rainbow trout have shown that acclimation to hypersaline environments enhances the toxicity to thioether organophosphate and carbamate pesticides. In order to determine the role of biotransformation in this process, the metabolism of the thioether organophosphate biocide, fenthion was evaluated in microsomes from gills, liver and olfactory tissues in rainbow trout (Oncorhynchus mykiss) acclimated to freshwater and 17‰ salinity. Hypersalinity acclimation increased the formation of fenoxon and fenoxon sulfoxide from fenthion in liver microsomes from rainbow trout, but not in gills or in olfactory tissues. NADPH-dependent and independent hydrolysis was observed in all tissues, but only NADPH-dependent fenthion cleavage was differentially modulated by hypersalinity in liver (inhibited) and gills (induced). Enantiomers of fenthion sulfoxide (65% and 35% R- and S-fenthion sulfoxide, respectively) were formed in liver and gills. The predominant pathway of fenthion activation in freshwater appears to be initiated through initial formation of fenoxon which may be subsequently converted to the most toxic metabolite fenoxon R-sulfoxide. However, in hypersaline conditions both fenoxon and fenthion sulfoxide formation may precede fenoxon sulfoxide formation. Stereochemical evaluation of sulfoxide formation, cytochrome P450 inhibition studies with ketoconazole and immunoblots indicated that CYP3A27 was primarily involved in the enhancement of fenthion activation in hypersaline-acclimated fish with limited contribution of FMO to initial sulfoxidation.     

Dietary exposure of rainbow trout (Salmo gairdneri) to Great Lakes coho salmon (Oncorhynchus kisutch Walbaum). Bioaccumulation of halogenated aromatic hydrocarbons and host resistance studies

Great Lakes coho salmon (Oncorhynchus kisutch walbaum) contain a gradient of halogenated aromatic hydrocarbons (HAHs) naturally bioaccumulated from their environment which have immunomodulatory potential. Juvenile rainbow trout (Salmo gairdneri) were fed diets of coho salmon from Lakes Michigan and Ontario, the Pacific Ocean and control trout chow, for a 20 weak period. The body burdens of organochlorines bioaccumulated in rainbow trout were similar to levels in the dietary coho salmon. The effect on natural resistance was assessed by challenge with a titrated dose of Vibrio anguillarum (VA-58). The ability to mount a protective immune response was determined by immunization with a VA-58 bacterin followed by challenge with virulent VA-58. These parameters of host resistance of rainbow trout were not compromised following dietary exposure to Great Lakes coho salmon.     

Dietary toxicity of single-walled carbon nanotubes and fullerenes (C60) in rainbow trout (Oncorhynchus mykiss)

The objective of this investigation was to compare the toxicity of two manufactured carbon nanomaterials (CNs) to determine if shape influenced toxicity. Juvenile rainbow trout Oncorhynchus mykiss were fed a control diet (no CN addition), or a diet supplemented with 500 mg single-walled carbon nanotubes (SWCNT) kg(-1) or 500 mg C(60) kg(-1) for six weeks. Fish growth, haematology, tissue ion concentrations, histopathology, osmoregulation, and biochemistry were evaluated. At week 4, but not on weeks 2 and 6, significant elevation in brain TBARS (an indication of lipid peroxidation) was observed in fish exposed to SWCNTs (16.2 ± 1.38 nmol mg(-1) protein) compared to the control (9.11 ± 0.81 nmol mg(-1) protein) and fish exposed to C(60) (8.28 ± 0.56 nmol mg(-1) protein). No other significant treatment-related differences were observed. Results indicate that dietary exposure to SWCNTs and C(60) in rainbow trout did not result in overt toxicity.