Effects of Heavy Metal on Rat Liver Microsomal Ca”-ATPase and Ca2+ Sequestering

In isolated hepatic microsomal vesicles the heavy metals Cd*+, Cu’+, and Zn2+ inhibit Ca2+ uptake and evoke a prompt efflux of Ca2+ from preloaded vesicles in a dose-dependent manner. N-Ethylmaleimide also inhibits Ca2+ uptake and causes Ca2+ release, but it is less effective in these respects than the heavy metals. Measurement of mannose-6-phosphatase activity indicate that the heavy metal-induced Ca2+ efflux is not caused by a general increase in membrane permeability. Heavy metals also inhibit the Ca2+-ATPase activity and the formation of the phosphorylated intermediate of the enzyme. In contrast, the sulfhydryl modifying reagent, N-ethylmaleimide inhibits the Ca2+-ATPase activity while it has a relatively small effect on Ca2+ release. Thus, the effects of these agents on Ca2+ sequestering and Ca2+-ATPase activity are not strictly proportional. The sulfhydryl group reducing agent dithiothreitol protects the microsomes from the effects of heavy metals, while glutathione is less protective. Addition of vanadate to vesicles, at a concentration which completely blocked the activity of the Ca2+ATPase, resulted in a small and slow release of the accumulated Ca2+. Subsequent additions of heavy metals evoked a massive Ca2+ release. Thus, the effects of heavy metals on Ca2+ efflux cannot be due entirely to their inhibition of the Ca” pump. The heavy metal-induced Ca2+ efflux is not inhibited either by ruthenium red or tetracaine.