Sequence and structure comparisons between alpha and beta connexins, Cx43 and Cx26, revealed that Cx43 has a motif, the carbamylation motif, that confers CO 2 -sensitivity on a subset of beta connexins. By using a fluorescent dye loading assay, whole cell patch clamp recordings and real-time measurement of ATP release via GRAB ATP , we have demonstrated that Cx43 hemichannels open in a highly CO 2 -sensitive manner over the range 20–70 mmHg. Mutational analysis confirms that the equivalent residues to those in Cx26, known to be involved in mediating the effects of CO 2 on gating of hemichannels and gap junction channels, also mediate Cx43 hemichannel gating. These data predict that Cx43 will be partially open at resting physiological levels of PCO 2 . In acute mouse hippocampal slices, we have demonstrated a CO 2 -dependent enhancement of synaptic transmission that was blocked by the Cx43-selective mimetic peptide Gap26. Our data resolves an inconsistency in the literature between in v