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The normal variations in hepatic levels of metallothionein, zinc and copper were studied during an annual reproductive cycle in rainbow trout of both sexes. In female fish, the total hepatic zinc levels closely followed the estradiol-17β and the LSI levels. Hence, the zinc levels rose in September, peaked in December and dropped in January. No distinct peaks were, however, observed in the whole-liver copper content. The hepatic metallothionein levels in female fish began to increase at the onset of exogenous vitellogenesis. Maximum levels were reached after estradiol-17β and LSI levels had dropped in January. In male fish no distinct peaks in either zinc or copper levels were observed. The metallothionein levels increased somewhat during the time of spermatogenesis. It is suggested that metallothionein may regulate the hepatic zinc distribution during the annual reproductive cycle in female rainbow trout, thereby ensuring the organism of a control mechanism to keep the pool of available zinc at an appropriate level.

An homologous radioimmunoassay for brown trout vitellogenin (VTG) was developed. Intact VTG, isolated from juvenile brown trout by selective precipitation and anion exchange chromatography was labelled with Na(125)I, with 1,3,4,6-tetrachloro-3α,6α-diphenylglycoluril (Iodogen) as the oxidizing agent. Incorporation of Na(125)I into VTG was higher than 75% and there was little degradation of the labelled protein. Labelled VTG eluted at the same position as unlabelled, purified brown trout VTG when analyzed by gel filtration on Sepharose 6B. Antisera with high titers, i.e. 1∶250 000, against brown trout VTG were raised in rabbits. The sensitivity of the assay was 5 ng VTG/ml and the standard curve was linear between 10 and 100 ng VTG/ml. Plasma from maturing female brown trout, as well as estradiol-treated and untreated juvenile brown trout diluted parallel to the standard curve, while plasma from maturing female rainbow trout and estradiol-treated arctic charr diluted non-parallel to the standard curve for brown trout VTG. Purified rainbow trout VTG and plasma from maturing female rainbow trout diluted parallel to each other, but with lower sensitivity than for brown trout VTG. Determinations of protein-bound phosphorus in the plasma of estradiol-treated juvenile brown trout correlated well with the RIA determinations of VTG. Repeated freezing and thawing of plasma samples yielded up to a hundred-fold increase in the apparent VTG level, while storage of a plasma sample for one year at -20°C did not affect the VTG level as measured by RIA.

Rainbow trout were exposed to 200 μg cadmium/l in the water during four months at 6-10°C. The liver, kidney and gills were analyzed for cadmium, copper, zinc, metallothionein and metallothionein mRNA. Cadmium accumulated in all three organs and reached the highest concentration in the kidney. The tissue zine and copper concentrations showed no major alterations during the experiment. The cytosolic distribution of cadmium, copper and zinc was followed during four months of exposure by Sephadex G-75 chromatography. It was found that cadmium was predominantly associated with proteins of an apparent molecular vieght of 10,000 daltons. These proteins were further identified as metallothioneins after fast protein liquid chromatography on a Mono-Q column. The metallothionein concentration was significantly higher in liver of exposed fish than in control fish after only one month. The kidneys reached significantly elevated levels of metallothionein in the exposed group after three months. In the gills, elevated metallothionein concentrations were observed after four months of exposure. After four months of exposure, the metallothionein mRNA content of liver and kidney was analyzed using a rainbow trout anti-sense RNA probe. Elevated MT mRNA levels were observed in both kidney and liver. These results demonstrate thatde novo synthesis of metallothionein is induced by cadmium in rainbow trout after exposure to the metalvia water.