Alteration of mitochondrial membrane potential (DELTA_PSI_m) and phosphatidylserine translocation as early indicators of heavy metal-induced apoptosis in the earthworm Eisenia hortensis

The effects of the heavy metals cadmium and copper (50-500 ìM) on the apoptotic events involving changes in mitochondrial membrane potential ( m) and phosphatidylserine (PS) translocation were investigated in the immune cells (celomocytes) of the earthworm Eisenia hortensis. Using the fluorescent probe JC-1, loss of membrane potential due to depolarization was detected in a greater proportion of cases when induced by cadmium compared to copper (58.7 % vs. 37 %) and at a lower concentration (50 ìM vs. 125ìM). With the use of the general caspase inhibitor Z-VAD-fmk, PS translocation detected by annexin V-FITC was found to be caspase-dependent when induced by cadmium at 125-250 ìM but not at 50 ìM or 500 ìM; a high proportion of earthworms (60 %) exhibited inhibitory effects. Additionally, the collapse in membrane potential and PS translocation were found to strongly correlate (r > 0.5) in 89 % of cases when induced by cadmium and copper. Thus, heavy metals appear to induce death in celomocytes of E. hortensis through apoptosis by means of caspase dependent pathways