Apoptosis is a process characterized by complex intracellular signaling pathways. However, intercellular communication is also proven to be an important factor in determining the degree of cell death. The latter can be mediated by the diffusion of molecules through connexin (Cx) channels, i.e. gap junctions channels (GJs), that connect the cytoplasm of adjacent cells, and hemichannels, half of a GJ, that form a paracrine conduit between the cytoplasm and the extracellular environment. The purpose of this study was to characterize the role of both Cx channels in the spread of apoptosis and to identify possible signaling molecules.

We used an in vitro experimental setup which consisted of loading a fine localized area of an adherent C6 glioma cell culture, stably transfected with Cx43 (C6Cx43), with the apoptotic agent Cytochrome C (CytC) using an in situ electroporation technique. This resulted in spread of apoptosis to cells, which did not initially contain the exogenous CytC. We distinguished two different zones: a band like area right next to the electroporated area, and clusters and individual apoptotic cells further away. The role of the Cx channels was further studied by comparing with the wild type culture and by applying several GJ and/or hemichannel blockers. The results lead to the conclusion that GJs mediate spread of apoptosis to neighbouring cells that border the electroporation area while hemichannels are suggested to play a role in apoptosis in more distant cells. The cell permeable calcium chelator BAPTA-AM reduced the cell death mediated by GJs as well as hemichannels, suggesting that calcium is involved in both processes.