Effects of Hg²⁺ and Cu²⁺ on the cytosolic Ca²⁺ level in molluscan blood cells evaluated by confocal microscopy and spectrofluorimetry

In the present work the effect of Hg²⁺ and Cu²⁺ on the level of cytosolic Ca²⁺ in mussel (Mytilus edulis L.) haemolymph cells were investigated by confocal microscopy and spectrofluorimetry utilizing the fluorescent dye Fluo3. In the blood cells of marine molluscs, exposure to Cu²⁺ and Hg²⁺ in the nanomolar and micromolar range causes a time-and concentration-dependent increase in the cytosolic Ca²⁺ level. Both the presence of a low-calcium containing medium and pretreatment of the cells with the channel blocker Verapamil greatly reduced the effects of higher (50 μM) Hg²⁺ concentrations, this indicating that Hg²⁺ enhances the influx of extracellular Ca²⁺ partly through activation of voltage-dependent Ca²⁺ channels. Low concentrations of Hg²⁺ (1 μM) and also of Cu²⁺ (0.5 μM), an "essential" element, were able to induce a sustained increase in cytosolic Ca²⁺, which was not affected either by Verapamil pretreatment or by lowering the extracellular calcium concentration. These data indicate that in mussel haemocytes heavy metal cations impair Ca²⁺ homeostasis not only by affecting Ca²⁺ channels, but also by interfering with other mechanisms of calcium transport across cellular membranes, such as the Ca²⁺-ATPases. The resulting increase in cytosolic Ca²⁺ could activate Ca-dependent processes which may be involved in many of the biochemical and physiological alterations observed in the cells of metal-exposed mussels. Specimens used in these experiments were collected from the river Linker near Plymouth, U.K. in June 1991.