Salmon Liver Research Articles

Biliverdin reductase from the liver of Atlantic salmon (Salmo salar).

Biliverdin reductase was characterized and purified from the liver of Atlantic salmon (Salmo salar) using a novel enzymatic staining method. The properties of the enzyme are quite different from those of mammals. The purified enzyme is a monomeric protein with a molecular weight of approximately 68 kD and an isoelectric point of around 3.8. The enzyme can utilize both NADH and NADPH as coenzyme, but the kinetic properties of the NADH-dependent and the NADPH-dependent enzyme activities are different: K(m) value for biliverdin IXalpha is 0.6 microM in the NADPH system, while it is 6.8 microM in the NADH system. Both enzyme activities are inhibited by excess biliverdin IXalpha, but the NADPH-dependent enzyme activity is far more susceptible. The optimum pH for activity is 5.5 with NADPH and 6.0 with NADH. The optimum reaction temperature is 35 degrees C.

Characterization of two distinct aryl hydrocarbon receptor (AhR2) genes in Atlantic salmon ( Salmosalar) and evidence for multiple AhR2 gene lineages in salmonid fish

The aryl hydrocarbon receptor (AhR) mediates the toxicity of several environmental contaminants, e.g. 2,3,7,8-tetrachlorodibenzo- p-dioxin, and other halogenated hydrocarbons in vertebrates. This receptor initiates the transcription of several biotransformation enzymes, which in turn are responsible for causing severe harm to biological tissue. Here we describe the isolation and complete characterization of the first two AhR genes from the teleost fish Atlantic salmon ( Salmo salar). The predicted amino acid sequences contain regions characteristic of other vertebrate AhRs including basic helix-loop-helix (bHLH) and PER-ARNT-SIM (PAS) domains but show little similarity to other vertebrate AhRs across the C-terminal half. Furthermore, they do not contain distinct Q-rich domains as found in the mammalian AhR, which is in line with previously described fish AhR genes. The salmon cDNAs encode 1106 and 1107 putative residues, respectively, approximately 50 amino acids longer than previously characterized AhR genes. Phylogenetic analyses demonstrated that the two salmon AhR sequences cluster within the AhR subfamily of the bHLH-PAS family, in a clade containing fish AhR2 genes. Although the two AhR2 forms are 92% identical at the amino acid level, the distribution of sequence differences and the presence of both forms in 30 tested individuals suggest that they are not allelic but derived from separate loci. Interestingly, they are not orthologs of the rainbow trout ( Oncorhynchus mykiss) AhR2 α and β genes and the new salmon loci are therefore here designated AhR2 γ and AhR2 δ. In line with this, PCR with DNA from rainbow trout revealed a new trout AhR locus that was more similar to the two salmon genes than to the trout AhR2 α and β genes, suggesting that the rainbow trout possesses at least three distinct AhR2 genes. The presence of multiple AhR genes in these species is probably a consequence of the genome duplications that occurred in the early evolution of fish and later also specifically in the salmonid lineage. Reverse transcription-PCR analyses revealed that both AhR2 γ and AhR2 δ are transcribed in the liver, spleen and muscles of adult salmon.

Screening for cyanobacterial hepatotoxins in herring and salmon from the Baltic Sea

Nodularin, a cyclic pentapeptide hepatotoxin closely related to microcystins, is produced by the toxic brackish water cyanobacterium Nodularia spumigena which regularly forms blooms in the Baltic Sea. Baltic herring (Clupea harengus membras L.) and salmon (Salmo salar L.) were caught in the Baltic Sea during 1997 to assess levels of nodularin and microcystins resulting from the blooms. The total toxin content in herring muscle and salmon liver was measured with an enzyme-linked immunosorbent assay. Despite exceptionally heavy blooms of toxic Nodularia in the Baltic Sea during the summer of 1997, Baltic herring and salmon contained very low quantities of nodularin and microcystins (2.5–6.5 ng g–1 dry weight, not corrected for recovery, [30%]). These concentrations do not pose a health threat to humans, based on the World Health Organization value of tolerable daily intake. Moreover, the concentrations we found in salmon and herring were much lower than those reported for livers of European flounder (Platichthys flesus L.) or Atlantic cod (Gadus morhua L.) from the Baltic Sea during 1996–1999. We suspect that cyanobacterial hepatotoxins do not effectively accumulate in herring and salmon.

Quantitative PCR analysis of CYP1A induction in Atlantic salmon ( Salmo salar)

Environmental pollutants are hypothesized to be one of the causes of recent declines in wild populations of Atlantic salmon ( ̄ Salmo salar ) ̄ across Eastern Canada and the United States. Some of these pollutants, such as polychlorinated biphenyls and dioxins, are known to induce expression of the CYP1A subfamily of genes. We applied a highly sensitive technique, quantitative reverse transcription-polymerase chain reaction (RT-PCR), for measuring the levels of CYP1A induction in Atlantic salmon. This assay was used to detect patterns of CYP1A mRNA levels, a direct measure of CYP1A expression, in Atlantic salmon exposed to pollutants under both laboratory and field conditions. Two groups of salmon were acclimated to 11 and 17 °C, respectively. Each subject then received an intraperitoneal injection (50 mg kg −1) of either β-naphthoflavone (BNF) in corn oil (10 mg BNF ml −1 corn oil) or corn oil alone. After 48 h, salmon gill, kidney, liver, and brain were collected for RNA isolation and analysis. All tissues showed induction of CYP1A by BNF. The highest base level of CYP1A expression (2.56×10 10 molecules/μg RNA) was found in gill tissue. Kidney had the highest mean induction at five orders of magnitude while gill tissue showed the lowest mean induction at two orders of magnitude. The quantitative RT-PCR was also applied to salmon sampled from two streams in Massachusetts, USA. Salmon liver and gill tissue sampled from Millers River (South Royalston, Worcester County), known to contain polychlorinated biphenyls (PCBs), showed on average a two orders of magnitude induction over those collected from a stream with no known contamination (Fourmile Brook, Northfield, Franklin County). Overall, the data show CYP1A exists and is inducible in Atlantic salmon gill, brain, kidney, and liver tissue. In addition, the results obtained demonstrate that quantitative PCR analysis of CYP1A expression is useful in studying ecotoxicity in populations of Atlantic salmon in the wild.

Purification of serum precursor proteins to vitelline envelope (choriogenins) in masu salmon, Oncorhynchusmasou

Three vitelline envelope-related proteins (VERPs), very-high-molecular-weight VERP (vhVERP), high-molecular-weight VERP (hVERP) and low-molecular-weight VERP (lVERP) were purified from female masu salmon serum. The apparent molecular weights of vhVERP, hVERP and lVERP, in their native state, were 520, 88 and 54 kDa, respectively, by gel-filtration chromatography. Very-high-molecular-weight VERP comprises two subunits, corresponding to 175 and 126 kDa. On SDS-PAGE, hVERP and lVERP migrate at 53 and 47 kDa, respectively. Amino acid analysis of vhVERP and hVERP showed that they share a high content of glutamic acid and proline. By contrast, lVERP is rich in glutamic acid and asparatic acid. These features are in good agreement with the amino acid composition of the vitelline envelope. Immuno-biochemical analysis suggested that vhVERP is derived from hVERP by polymerization and/or aggregation. Antibodies against hVERP and lVERP specifically immunostained the vitelline envelope and liver of female masu salmon. In addition, both hVERP and lVERP were induced in the serum of estrogen-treated male fish. Taken together, it is suggested that hVERP and lVERP are homologous molecules with choriogenin H and choriogenin L in medaka, respectively. These results indicate that hVERP and lVERP are precursor proteins to the vitelline envelope (choriogenins) in masu salmon.

EST-based identification of genes expressed in the liver of adult Atlantic salmon ( Salmo salar)

A list of genes expressed in the liver of Atlantic salmon was compiled using the expressed sequence tag (EST) strategy. 733 ESTs, derived from 170 abundant and 563 rare mRNA encoding liver cDNA clones, were determined. Bioinformatic analysis revealed that 390 (53%) of the salmon liver ESTs could be ascribed to the transcriptional products of 93 identified genes including 7 previously described in the Atlantic salmon. The identified Atlantic salmon genes were classified with respect to cellular role which showed that 33 (36%) of the identified genes encoded proteins associated with primary liver functions such as transport, acute phase response, and blood clotting. Furthermore, comparative analysis revealed that 12 of the 16 salmon genes that were shown to encode abundant mRNA transcripts in liver had homologues that have also been shown to be highly expressed in mammalian liver systems. Finally, two cDNA variants corresponding to the two cDNA forms of the apolipoprotein A-I gene previously identified in rainbow trout were also found in Atlantic salmon.

Suppressive subtraction libraries to identify interferon-inducible genes in fish.

Previous attempts to identify genes in fish that respond to virus infection or interferon induction have not been particularly productive. Since these genes are very important in developing strategies to control disease outbreaks in aquaculture, we began a study of interferon-inducible genes in fish using suppressive subtraction hybridization to construct cDNA libraries enriched for interferon-inducible genes. Subtraction hybridization libraries were constructed with cDNA obtained from the kidney, spleen, and liver of Chinook salmon (Oncorhynchus tshawytscha) and staghorn sculpin (Hemilepidotus spinosus) before and after injection with poly IC, a potent interferon inducer. The "identified" genes in both cDNA libraries corresponded to previously identified genes of the fish complement system, the interferon-inducible proteins observed in mammalian cells, and the Vig-1 gene, identified in fish cells after infection with fish rhabdoviruses.

The effect of copper-treated net pens on farmed salmon ( Salmo salar) and other marine organisms and sediments

Samples of Atlantic salmon ( Salmo salar), saithe ( Pollacius virens), blue mussel ( Mytilus edulis), brown seaweed ( Ascophyllum nodosum) and sediment were collected from six different fish farms. Five of the farms used net pens treated with copper-containing coatings, whereas one farm did not use copper-containing coating (this was used as a reference location). Samples of muscle, liver and gills of Atlantic salmon and saithe, blue mussel and brown seaweed were freeze dried, homogenised, wet digested and analysed for copper by flame atomic absorption spectrometry. The results showed no significant differences in copper concentrations among the samples from the different locations. The copper contents of some of the samples appeared to be in the upper part of the normal concentration range. From a nutritional point of view, the use of copper-coatings on net pens did not affect the quality of the seafood products either within, or around the net pen.

Histone H1: An Antimicrobial Protein of Atlantic Salmon (Salmo salar)

Antimicrobial activity was detected in acid extracts of liver, intestine, and stomach of healthy Atlantic salmon (Salmo salar). An antimicrobial protein was isolated from salmon liver using acid extraction followed by ammonium sulfate precipitation, large-scale gel filtration chromatography, reverse-phase HPLC, and size exclusion HPLC. The salmon antimicrobial (SAM) protein was found to have a molecular mass of 20,734 Da by MALDI TOF mass spectrometry. Peptide mass fingerprinting and partial sequencing by tandem nanoelectrospray mass spectrometry identified the protein as histone H1. The protein had a minimal inhibitory concentration of 31 μg/mL against E. coli D31 in a plate clearing assay. The effect of the SAM protein on bacterial morphology was indistinguishable from that of (Ala-8,13,18)-magainin II, as shown by scanning electron microscopy, which suggests that the protein disrupts E. coli membranes in a manner similar to that of most antimicrobial peptides. This protein may act as an antimicrobial in vivo through active secretion or by release from cells during infection-related apoptosis.

The chum salmon insulin-like growth factor II promoter requires Sp1 for its activation by C/EBPβ

The chum salmon insulin-like growth factor II (IGF-II) gene is highly expressed in liver tissue. In this study we demonstrate that two transcription factors, Sp1 and C/EBPβ, are involved in the enhanced expression of the salmon IGF-II gene. The presence of the fish homolog for C/EBPβ in salmon liver RNA was confirmed by Northern blotting. The sIGF-II promoter was activated up to 20-fold by co-transfection with C/EBPβ. The functional importance of four out of the five putative C/EBPβ binding sites was demonstrated with mutational analysis in transient transfection assays. The transcription factor Sp1 binds to two sites within the salmon IGF-II promoter. Interestingly, mutation of the Sp1 binding sites decreases not only the basal IGF-II promoter activity but also the C/EBPβ-induced transactivation. These results demonstrate that liver-enriched C/EBPβ and ubiquitously expressed Sp1 each activate the sIGF-II promoter and that Sp1 is required for full transactivation of the sIGF-II gene by C/EBPβ. This suggests that C/EBPβ and Sp1 act in synergy.

Influence of dietary carbohydrate on antioxidant enzyme activities in liver of Atlantic salmon (Salmo salar L.)

Effects of dietary carbohydrates on hepatic antioxidant enzymeactivities were studied in Atlantic salmon (Salmo salarL.), fed diets containing no additional carbohydrate, or a low (15% addition)orhigh (30% addition) concentration of D-glucose or gelatinised potato starch.Addition of free glucose to the feeds resulted in glucose availability being 7and 18% when added at 15 and 30%, while the addition of gelatinised starch didnotseem to reduce starch availability. The dietary treatment groups (nocarbohydrate, 7 and 18% glucose or 15 and 30% gelatinised starch) were providedwith quantities of feed that supplied the same amounts of protein and lipid.There was a positive correlation between tiobarbituric acid-reactive substances(TBARS) in feeds and salmon liver. Liver glycogen concentrations increased withincreasing dietary glucose and starch, and the two higher levels of liverglycogen, found in groups G18 and S30, resulted in decreased activities ofcatalase, superoxide dismutase (t-SOD), and concentrations of glutathione. Thisindicates that there are links between carbohydrate metabolism and antioxidantsystems in salmon liver. There was no correlation between selenium dependentglutathione peroxidase (Se-GPX) activity and dietary carbohydrateconcentrations, but Se-GPX activity increased in response to increased TBARS.Catalase and t-SOD activities did not correlate with either TBARS values orSe-GPX activity.

A survey of genes in the Atlantic salmon (Salmo salar) as identified by expressed sequence tags

We describe the construction and quality analysis of six cDNA libraries from the liver, ovary, testis, brain, spleen and muscle tissues of adult Atlantic salmon. The cDNA libraries were then screened with total cDNA probes to catalogue clones representing the abundant and rare mRNA populations in each tissue. Subsequently, the 5′-terminal DNA sequences of 1152 cDNA clones, composed of 96 clones from each of the abundant and rare mRNA populations in the six tissues, were determined. Bioinformatic analysis revealed that 510 (50%) of the salmon expressed sequence tags (ESTs) of sufficient length showed significant homology to previously identified genes from salmonid and other species, while 517 (50%) of salmon ESTs were unidentified or novel. After accounting for multi-EST redundancy, the 510 identified ESTs provided DNA sequence markers for 178 salmon genes which are listed in terms of tissue of origin and mRNA abundance class.

Apparent digestibility coefficients and accumulation of astaxanthin E/ Z isomers in Atlantic salmon ( Salmo salar L.) and Atlantic halibut ( Hippoglossus hippoglossus L.)

Apparent astaxanthin (3,3′-dihydroxy-β,β-carotene-4,4′-dione) digestibility coefficients (ADC) and carotenoid compositions of the muscle, liver, whole kidney and plasma were compared in Atlantic salmon ( Salmo salar) and Atlantic halibut ( Hippoglossus hippoglossus) fed a diet supplemented with 66 mg astaxanthin kg −1 dry matter for 112 days. The astaxanthin source consisted of 75% all- E-, 3% 9 Z- and 22% 13 Z-astaxanthin, of (3 R,3′ R)-, (3 R,3′ S; meso)-, and (3 S,3′ S)-astaxanthin in a 1:2:1 ratio. The ADC of astaxanthin was significantly higher in Atlantic halibut than in Atlantic salmon after 56 and 112 days of feeding ( P<0.05). The ADC of all- E-astaxanthin was significantly higher than ADC of 9 Z-astaxanthin ( P<0.05). Considerably more carotenoids were present in all plasma and tissue samples of salmon than in halibut. Retention of astaxanthin in salmon muscle was 3.9% in salmon and 0 in halibut. All- E-astaxanthin accumulated selectively in the muscle of salmon, and in plasma of salmon and halibut compared with diet. 13 Z-astaxanthin accumulated selectively in liver and whole kidney of salmon and halibut, when compared with plasma. A reductive pathway for astaxanthin metabolism in halibut similar to that of salmon was shown by the presence of 3′,4′- cis and trans glycolic isomers of idoxanthin (3,3′,4′-trihydroxy-β,β-carotene-4′-one) in plasma, liver and whole kidney. In conclusion, the higher ADC of astaxanthin in halibut than Atlantic salmon may be explained by lower feed intake in halibut, and the lower retention of astaxanthin by a higher capacity to transform astaxanthin metabolically.

Multiple variants of the peroxisome proliferator-activated receptor ( PPAR) γ are expressed in the liver of Atlantic salmon ( Salmo salar)

A full-length cDNA encoding the peroxisome proliferator-activated receptor (PPAR) has for the first time been characterized from a fish species. The Atlantic salmon PPARγ cDNA of 2528 nucleotides (nt) was amplified from liver mRNA by reverse transcription (RT)-polymerase chain reaction (PCR). The deduced protein of 544 amino acids (aa) shares approximately 47% overall sequence identity with mammalian PPARγ. The N-terminal A/B region contains a repeated decapeptide motif and shows a low homology with other PPARs. In contrast, the central DNA-binding domain (DBD) and the C-terminal ligand-binding domain (LBD) show a high sequence identity to mammalian and Xenopus PPARγ. The salmon PPARγ LBD contains nine additional residues in a flexible loop that might affect ligand binding. Northern blot analysis of salmon liver RNA revealed a prominent transcript of about 1.7 kilo bases (kb), in addition to several mRNA species of about 2.4–2.6 kb, which is consistent with the presence of multiple putative polyadenylation sites in the 3′ untranslated region (UTR) of the 2528 nt long PPARγ cDNA. Two additional PPARγ cDNAs of 1719 and 2357 nt were then isolated. The 2357 nt long transcript encodes full-length PPARγ and seems to be ubiquitously expressed in salmon, whereas the liver-specific transcript of 1719 nt encodes a truncated variant of PPARγ. The truncated form lacks 39 C-terminal residues including the conserved activation function-2 (AF-2) motif, known to be associated with crucial cofactors. Three-dimensional modelling studies indicated that the C-terminal truncation would result in important alterations of the ligand-binding pocket. The presence of a truncated form with drastic changes in both ligand- and cofactor-binding sites is likely to modulate PPARγ activity in salmon liver.

Iron Ore Mines Leachate Potential for Oxyradical Production

The ecotoxicological effects of mining effluents is coming under much greater scrutiny. It appears necessary to explore possible health effects in association with iron ore mining effluents. The present results clearly demonstrate that iron-ore leachate is not an inert media but has the potential to induce lipid peroxidation. Peroxidation was assessed by measuring oxygen consumption in the presence of a reducing agent such as ascorbate or NADPH and a chelator such as EDTA. Labrador iron ore is an insoluble complex crystalline material containing a mixture of metals (Fe, Al, Ti, Mn, Mg,…, ) in contrast to the iron sources used for normal lipid peroxidation studies. The metal of highest percentage is iron (59.58%), a metal known to induce oxyradical production. Iron ore powder initiated ascorbic acid-dependent lipid peroxidation (nonenzymatic) in liposomes, lipids extracted from rat and salmon liver microsomes, and intact salmon liver microsomes. It also revealed an inhibitory effect of NADPH-dependent microsomes lipid peroxidation as well as on NADPH cytochrome c reductase activity. However, nonenzymatic peroxidation in rat liver microsomes was not significantly inhibited. Cytochrome P450 IA1- and IIB1-dependent enzymatic activities as well as P450 levels were not affected. The inhibition could be due to one of the other components of iron ore leachate (Mn, Al,…, ). These effects of iron-ore leachate indicate that a potential toxicity could be associated with its release into lakes. Further studies are necessary to explore in vivo effects on aquatic animals.

Hepatic glucose phosphorylating activities in perch (Perca fluviatilis) after different dietary treatments

Increased activity of hepatic glucose phosphorylation was observed in perch after feeding previously fasted fish. When a pellet diet containing 14% carbohydrate was given, most of the increased activity had a low affinity towards glucose ( S 0.5=19.5 mM) and resembled the mammalian glucokinase (Hexokinase IV or D) and the glucokinase-like activity previously observed in salmon liver. In addition, increased actitivy of a hexokinase with high affinity towards glucose ( K m=0.50 mM) was observed with the pellet diet. An increase in the activity of this hexokinase alone was observed when the fish were fed with filet of cod containing less than 0.2% carbohydrate. Perch with a very high hepatic glucokinase-like activity after eating the pellet diet had high activities of pyruvate kinase and glucose-6-phosphate dehydrogenase, indicating a high capacity of glycolysis and carbohydrate utilization. Simultaneously, the activity of glycogen phosphorylase was strongly reduced while the activity of fructose-1,6-bisphosphatase was not significantly changed. These observations were made with perch captured in the spawning season and brought to the laboratory. Assays of glucose phosphorylation in livers of perch eating the natural diet (insects) in the lake showed no glucokinase-like activity.

Metabolism of n-3 and n-6 fatty acids in Atlantic salmon liver: stimulation by essential fatty acid deficiency.

Oxidation, esterification, desaturation, and elongation of [1-14C]18:2n-6 and [1-14C]18:3n-3 were studied using hepatocytes from Atlantic salmon (Salmo salar L.) maintained on diets deficient in n-3 and n-6 polyunsaturated fatty acids (PUFA) or supplemented with n-3 PUFA. For both dietary groups, radioactivity from 18:3n-3 was incorporated into lipid fractions two to three times faster than from 18:2n-6, and essential fatty acids (EFA) deficiency doubled the incorporation. Oxidation to CO2 was very low and was independent of substrate or diet, whereas oxidation to acid-soluble products was stimulated by EFA deficiency. Products from 18:2n-6 were mainly 18:3n-6, 20:3n-6, and 20:4n-6, with minor amounts of 20:2n-6 and 22:5n-6. Products from 18:3n-3 were mainly 18:4n-3, 20:5n-3, and 22:6n-3, with small amounts of 20:3n-3. The percentage of 22:6n-3 in the polar lipid fraction of EFA-deficient hepatocytes was fourfold higher than in n-3 PUFA-supplemented cells. This correlated well with our other results obtained after abdominal injection of [1-14C]18:3n-3 and [1-14C]18:2n-6. In hepatocytes incubated with [4,5-3H]-22:6n-3, 20:5n-3 was the main product. This retroconversion was increased by EFA deficiency, as was peroxisomal betaoxidation activity. This study shows that 18:2n-6 and 18:3n-3 can be elongated and desaturated in Atlantic salmon liver, and that this conversion and the activity of retroconversion of very long chain PUFA is markedly enhanced by EFA deficiency.

Induction of hepatic estrogen receptor in juvenile Atlantic salmon in vivo by the environmental estrogen, 4-nonylphenol

Alkylphenol ethoxylate degradation products such as nonylphenol and octylphenol are shown to have estrogenic effects. Nonylphenol induces synthesis of vitellogenin (a precursor of egg yolk proteins) and zona radiata proteins (eggshell proteins) in juvenile and/or male fish. Little is known about the molecular mechanisms of estrogenicity of environmental chemicals such as nonylphenol. To study the mechanisms of estrogenic effects of 4-nonylphenol (NP), we examined its in vivo effects on the expression of the estrogen receptor (ER), vitellogenin (Vtg) and zona radiata protein (Zrp) genes in juvenile Atlantic salmon liver. We show that the ER mRNA synthesis is induced by NP in a dose-dependent manner in juvenile Atlantic salmon liver. The induction of the ER mRNA synthesis is followed by the induction of Zrp and Vtg mRNA synthesis. The ER transcripts reach peak levels earlier than the Zrp and Vtg mRNA and proteins, which is in agreement with the physiological effects of estradiol during zonagenesis and vitellogenesis. Various studies have also shown that NP competitively inhibits the binding of 17β-estradiol (E2) to ER. Our results further suggest that NP directly mimics E2 in inducing the ER, Zrp and Vtg genes in salmon liver.

The chum salmon IGF-II gene promoter is activated by hepatocyte nuclear factor 3β

IGF-II plays an important role in growth and development of vertebrates and is highly expressed in adult salmon liver. In the present study, we demonstrate that a liver-enriched transcription factor, hepatocyte nuclear factor 3β (HNF-3β), is an activator of the chum salmon IGF-II gene. Multiple binding sites for HNF-3β were identified within the 5′-UTR using electrophoretic mobility shift assays and mutation of these sites prevents binding of HNF-3β. In transient transfection assays it was shown that mutation of the HNF-3β binding sites results in a substantial decrease of HNF-3β-activated salmon IGF-II gene expression. This is the first identified transcription factor that is functionally involved in the regulation of fish IGF-II expression.

Immunohistochemical analysis of the vitellogenin response in the liver of Atlantic salmon exposed to environmental oestrogens

Induction of vitellogenin (Vtg) in oviparous vertebrates has been used as a biomarker of response for environmental oestrogens. This study reports the cellular localization of oestrogen- and xenoestrogen-induced Vtg synthesis in the liver of juvenile Atlantic salmon (Salmo salar). Paraffin-embedded liver sections were incubated with homologous monoclonal antibody against Atlantic salmon Vtg. Following intraperitoneal (ip) exposure of fish to estradiol-17beta E2; 5 mg kg-1 or 4-nonylphenol NP; 125 mg kg-1, Vtg induction was primarily demonstrated immunohistochemically in the cytoplasm of hepatocytes, endothelial cells and within hepatic sinusoids. Vtg staining of hepatocytes was not evenly distributed, as there was a high degree of polarization toward the sinusoid. The intensity of positive Vtg staining was stronger in the liver sections of E2-treated fish, compared with NP-treated fish. Hepatocytes of E2-, NP- and vehicle (control)-treated fish showed normal cellular structures, thus showing no evidence of histopathological changes. In parallel, indirect enzyme-linked immunosorbent assay (ELISA) and immunoblot analysis of plasma Vtg levels show significant induction of Vtg in E2- and NP-treated fish, as compared with untreated (control) fish. The present study demonstrates the applicability of immunohistochemistry in studies of cellular structures, processes and responses of fish exposure to oestrogen and oestrogen-mimicking compounds.