The present study reports the complete coding sequences for two paralogues for leptin (sLepA1 and sLepA2) and leptin receptor (sLepR) in Atlantic salmon. The deduced 171-amino acid (aa) sequence of sLepA1 and 175 aa sequence for sLepA2 shows 71.6% identity to each other and clusters phylogenetically with teleost Lep type A, with 22.4% and 24.1% identity to human Lep. Both sLep proteins are predicted to consist of four helixes showing strong conservation of tertiary structure with other vertebrates. The highest mRNA levels for sLepA1 in fed fish (satiation ration=100%) were observed in the brain, white muscle, liver, and ovaries. In most tissues sLepA2 generally had a lower expression than sLepA1 except for the gastrointestinal tract (stomach and mid-gut) and kidney. Only one leptin receptor ortholog was identified and it shares 24.2% aa sequence similarity with human LepR, with stretches of highest sequence similarity corresponding to domains considered important for LepR signaling. The sLepR was abundantly expressed in the ovary, and was also high in the brain, pituitary, eye, gill, skin, visceral adipose tissue, belly flap, red muscle, kidney, and testis. Fish reared on a rationed feeding regime (60% of satiation) for 10 months grew less than control (100%) and tended to have a lower sLepA1 mRNA expression in the fat-depositing tissues visceral adipose tissue (p<0.05) and white muscle (n.s.). sLepA2 mRNA levels was very low in these tissues and feeding regime tended to affect its expression in an opposite manner. Expression in liver differed from that of the other tissues with a higher sLepA2 mRNA in the feed-rationed group (p<0.01). Plasma levels of sLep did not differ between fish fed restricted and full feeding regimes. No difference in brain sLepR mRNA levels was observed between fish fed reduced and full feeding regimes. This study in part supports that sLepA1 is involved in signaling the energy status in fat-depositing tissues in line with the mammalian model, whereas sLepA2 may possibly play important roles in the digestive tract and liver. At present, data on Lep in teleosts are too scarce to allow generalization about how the Lep system is influenced by tissue-specific energy status and, in turn, may regulate functions related to feed intake, growth, and adiposity in fish. In tetraploid species like Atlantic salmon, different Lep paralogues seems to serve different physiological roles.

INTRODUCTIONAs a result of mining, forestry, waste disposal and fuel combustion, our environment is becoming increasingly contaminated with heavy metals. The aquatic environment receives waste products from such activities and may be the final depository for these anthropogenic ally remobilized heavy metals. In order to understand the impact of heavy metals on aquatic biota it is important to characterize the mechanisms available for aquatic life to transport, immobilize and excrete heavy metals.Relatively little is known about the specific mechanisms of uptake of metals in cells of non-mammalian vertebrates. This chapter attempts to assemble available information on fish. In general, metals absorbed across the gills or the intestinal wall are distributed via the circulation, bound to transport proteins, to different tissues of the body. Within the tissues metals can participate in the essential life functions, but can also exert toxic actions, or be detoxified by binding to the protein, metallothionein (MT). The intracellular levels of essential metal are regulated by transporters (which translocate metals across the plasma membrane) as well as by MT and other metal-binding proteins. Metals themselves cannot be metabolized (using the strict definition of the term) and can only be eliminated from the body by excretion.Metals that enter the body will react with different components of the cell (Chapter 1). The heavy metals are soft donors and will therefore readily bind to soft acceptors, such as sulfhydryl-groups. MT is a low molecular weight cytosolic, cysteine-rich protein that binds group lB and 2B heavy metals (Olsson and Raux, 1985). Within the cells of the body, MT is the major heavy metal-binding protein (Dunn et ai., 1987). Environmental exposure of fish to heavy metals has been shown to result in primarily renal and hepatic accu-

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The induction of metallothionein and vitellogenin synthesis in rainbow trout liver was studied after injection of oestradiol-17 beta alone or in combination with cadmium or zinc. Intraperitoneal injection of oestradiol-17 beta increased the liver somatic index, with subsequent induction of vitellogenin synthesis. Oestradiol-17 beta did not induce metallothionein synthesis. Injection of cadmium induced the synthesis of metallothionein mRNA and metallothionein. Injection of oestradiol-17 beta in combination with cadmium resulted in inhibition of transcription and translation of both vitellogenin and metallothionein. Chromatography of liver cytosols revealed that cadmium, when co-injected with oestradiol-17 beta, did not bind to metallothionein but would initially bind to high-molecular-mass (HMr) cytosolic proteins. In fish injected with cadmium in combination with oestradiol-17 beta, cadmium was gradually redistributed from HMr proteins to metallothionein. This resulted in induction of metallothionein synthesis and in binding of most of the cadmium to metallothionein. Induction of vitellogenin mRNA was observed 15 days after injection, as cadmium was being redistributed to newly synthesized metallothionein. These findings indicate that cadmium inhibits the transcription of vitellogenin. The binding of cadmium to these non-metallothionein proteins represses the induction of metallothionein and results in increased toxicity of the metal. Preinduction of metallothionein by zinc injections resulted in decreased cadmium sensitivity of the fish and a decrease in the repression of vitellogenin mRNA. Furthermore, a role for metallothionein in the detoxification of cadmium is indicated by the induction of vitellogenin synthesis that occurs once metallothionein has begun sequestering cadmium.

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