Copper induced upregulation of apoptosis related genes in zebrafish (Danio rerio) gill

Copper (Cu) is an essential micronutrient that, when present in high concentrations, becomes toxic to aquatic organisms. It is known that Cu toxicity may induce apoptotic cell death. However, the precise mechanism and the pathways that are activated, in fish, are still unclear. Thus, this study aimed to assess which apoptotic pathways are triggered by Cu, in zebrafish (Danio rerio) gill, the main target of waterborne pollutants. Fish where exposed to 12.5 and 100 μg/L of Cu during 6, 12, 24 and 48 h. Fish gills were collected to TUNEL assay and mRNA expression analysis of selected genes by real time PCR. An approach to different apoptosis pathways was done selecting p53, caspase-8, caspase-9 and apoptosis inducing factor (AIF) genes. The higher incidence of TUNEL-positive cells, in gill epithelia of the exposed fish, proved that Cu induced apoptosis. The results suggest that different apoptosis pathways are triggered by Cu at different time points of the exposure period, as the increase in transcripts was sequential, instead of simultaneous. Apoptosis seems to be initiated via intrinsic pathway (caspase-9), through p53 activation; then followed by the extrinsic pathway (caspase-8) and finally by the caspase-independent pathway (AIF). A possible model for Cu-induce apoptosis pathways is proposed.