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Calcium signals can be communicated between cells by the diffusion of a second messenger through gap junction channels or by the release of an extracellular purinergic messenger. We investigated the contribution of these two pathways in endothelial cell lines by photoliberating InsP₃ or calcium from intracellular caged precursors, and recording either the resulting intercellular calcium wave or else the released ATP with a luciferin/luciferase assay. Photoliberating InsP₃ in a single cell within a confluent culture triggered an intercellular calcium wave, which was inhibited by the gap junction blocker α-glycyrrhetinic acid (α-GA), the connexin mimetic peptide gap 26, the purinergic inhibitors suramin, PPADS and apyrase and by purinergic receptor desensitisation. InsP₃-triggered calcium waves were able to cross 20μm wide cell-free zones. Photoliberating InsP₃ triggered ATP release that was blocked by buffering …