Forced expression of gap junction proteins, connexins, enables gap junction-deficient cell lines to propagate intercellular calcium waves. Here, we show that ATP secretion from the poorly coupled cell lines, C6 glioma, HeLa, and U373 glioblastoma, is potentiated 5-to 15-fold by connexin expression. ATP release required purinergic receptoractivated intracellular Ca 2؉ mobilization and was inhibited by Cl ؊ channel blockers. Calcium wave propagation also was reduced by purinergic receptor antagonists and by Cl ؊ channel blockers but insensitive to gap junction inhibitors. These observations suggest that cell-to-cell signaling associated with connexin expression results from enhanced ATP release and not, as previously believed, from an increase in intercellular coupling.