QUANTIFICATION OF METALLOTHIONEINS IN THE COMMON ASTEROID ASTERIAS-RUBENS (ECHINODERMATA) EXPOSED EXPERIMENTALLY OR NATURALLY TO CADMIUM

Uptake and intracellular fate of Cd was assessed in the pyloric caeca of the common asteroid Asterias rubens exposed experimentally or natur ally to Cd. Cd partitioning among intracellular metal-binding pools wa s studied by graphite furnace atomic absorption spectrometry of heat-t reated cytosolic fractions which had been separated by gel filtration chromatography. Low molecular mass proteins bound > 80% of cytosolic C d and appeared to become saturated with the metal after experimental e xposure to 20 mu g Cd l(-1) for 10-20 days. These proteins presented s everal features of metallothioneins (MTs), namely molecular mass (13-2 2 kDa), heat-stability (90 degrees C, 10 min), high Cd content, and hi gh thiolic content as determined by differential pulse polarography. A nalysis of chromatographic fractions indicated that the majority of th iolic groups (66-73%) present in heat-treated cytosol was associated w ith the MT pool. Subsequent determinations of MT concentrations were m ade using whole cytosol extracts after correcting for interference fro m thiolic groups present in residual, heat-stable, high molecular mass proteins. Asteroids collected from unpolluted sites in SW England, SW Netherlands and SW Norway contained basal levels of 2.5-4.5 mg MT g(- 1) dw. Asteroids experimentally exposed to Cd (20 mu g Cd l(-1)) respo nded by a 2-fold increase in the production of MT after 30 days. Turno ver of MTs was rapid in A. rubens and probably accounted for the low n et increase in protein and characteristic rapid loss of Cd from the py loric caeca. Alkaline phosphatase (AP) activity was measured as a mark er of the functional state of the pyloric caeca. It was not significan tly affected during Cd exposure in the laboratory. Asteroids collected in the heavy metal polluted Sorfjord (SW Norway) contained significan tly higher MT concentrations (5-5.6 mg MT g(-1) dw) and significantly lower AP activity in the most heavily contaminated region, though thes e two parameters were not correlated.