The mechanism of lysosome activation by 17-estradiol has been studied in mussel blood cells. Cell treatment with estradiol induced a sustained increase of cytosolic free Ca²⁺ that was completely prevented by preincubating the cells with the Ca²⁺ chelator BAPTA-AM. Estradiol treatment was also followed by destabilization of the lysosomal membranes, as detected in terms of the lysosomes' increased permeability to neutral red. The effect of estradiol on lysosomes was almost completely prevented by preincubation with the inhibitor of cytosolic Ca²⁺-dependent PLA₂ (cPLA₂), arachidonyl trifluoromethyl ketone (AACOCF3), and was significantly reduced by preincubation with BAPTA-AM. In contrast, it was virtually unaffected by preincubation with the inhibitor of Ca²⁺-independent PLA₂, (E)-6-(bromomethylene)tetrahydro-3-(1-naphtalenyl)-2H-pyran-2-one (BEL). The Ca²⁺ ionophore A-23187 yielded similar effects on [Ca²⁺]_i and lysosomes. Exposure to estradiol also resulted in cPLA₂ translocation from cytosol to membranes, lysosome enlargement, and increased protein degradation. These results suggest that the destabilization of lysosomal membranes following cell exposure to estradiol occurs mainly through a Ca²⁺-dependent mechanism involving activation of Ca²⁺-dependent PLA₂. This mechanism promotes lysosome fusion and catabolic activities and may mediate short-term estradiol effects.