Disruption of metallothionein expression with antisense oligonucleotides abolishes protection against cadmium cytotoxicity in molluscan hemocytes

The relationship between metallothionein (MT) induction and cytotoxicity wa s examined in isolated oyster hemocytes exposed in vitro to cadmium, copper , and zinc. In all cases MT induction increased to peak levels with increas ed metal dose, then declined with continued increases in dose. The effectiv eness of these metals as inducers of MT was cadmium > copper > zinc, with c admium the most effective inducer in magnitude of induction and sensitivity to dose. Cytotoxicity of the metals, based on decreases in lysosomal neutr al red retention times, was copper > cadmium > zinc. Zinc was relatively no ntoxic and a poor inducer of MT. Cadmium, which was intermediate in toxicit y, required concentrations in excess of that causing peak induction of MT t o have substantial effects on lysosomal membrane integrity. In contrast, co pper was highly cytotoxic at concentrations below that which resulted in pe ak MT induction. This reversal in the rank order of cadmium and copper sugg ested that the toxicity of cadmium was tempered by the protective effects o f MT. This was verified by disrupting MT expression with antisense phosphor othioate-substituted oligodeoxynucleotides. Administration of 2.5 muM CdCl2 , which induced MT but had no effect on lysosomal membrane stability when a dministered alone, reduced neutral red retention time to 41% of control lev els when administered in the presence of antisense. The resulting toxicity was greater than that caused by a comparable concentration of copper. The f indings represented evidence that expression of MT protects against cadmium toxicity in an estuarine mollusc.