Subcellular distribution of cadmium and its cellular ligands in mussel digestive gland cells as revealed by combined autometallography and X-ray microprobe analysis

Autometallography (AMG) and electron probe X-ray microanalysis (EPXMA) were applied in combination to determine the subcellular distribution of Cd and its subcellular ligands in the digestive gland cells of Cd-exposed mussels Mytilus galloprovincialis. Black silver deposits (BSD), which reveal the presence of metals when AMG is applied, were only localized in digestive cell lysosomes. Digestive cell cytoplasm and basophilic cells were devoid of BSD. EPXMA (static probe and X-ray mapping) indicated that Cd, S (possibly associated with metallothioneins or metallothionein-like proteins) and autometallographical Ag ions are co-localized within digestive cell lysosomes. In addition, Cd and S co-occur in the absence of Ag in the cytosol of digestive cells. AMG does not reveal the presence of the Cd 'pool' strongly bound to cytosolic Cd-metallothionein complexes; only 'free' Cd or Cd supposedly loosely bound to (semi) digested metallothionein within lysosomes was revealed. The levels of lysosomal Cd were indirectly quantified by stereology as the volume density of BSD (Vv(BSD)). Significantly higher values were recorded in Cd-exposed mussels compared with controls at all exposure times. However, VvBSD values were lower at days 7 and 21 than at day 1. This relative decrease in VvBSD reflected another (and confounding) response elicited by Cd-exposure in the digestive epithelium: the volume density of basophilic cells (Vv(BAS)) increased significantly as exposure progressed. Due to this cell-type replacement, the net accumulative capacity of the digestive epithelium decreases at long exposure times.