Mechanisms of permselectivity of connexin hemichannels to small molecules

Connexins can either that act as hemichannels, to facilitate ion and small molecule movement from the cytosol to the extracellular space or as gap junction channels to provide a pathway for solute exchange between cells. Connexins are ubiquitously expressed throughout the body and are implicated in a wide range of processes. The permselectivity of connexin hemichannels for small neurochemicals remains poorly understood. By coexpressing genetically encoded fluorescent sensors for ATP, glutamate and lactate with a range of connexins, we examined the ability of different hemichannels to permit release of these compounds under physiological conditions and in response to physiological stimuli (small changes in PCO ² and transmembrane depolarisation). We found that some connexin hemichannels were relatively non-selective (Cx26, Cx32, Cx43, Cx31.1) allowing passage of ATP, glutamate and lactate. By contrast other connexin hemichannels (Cx36, Cx46 and Cx50) were highly selective. Cx36 and Cx46 hemichannels allowed release of ATP, but not glutamate or lactate. The size of the permeating molecule cannot be the sole determinant of permselectivity. By contrast, Cx50 hemichannels permitted the release of lactate and glutamate but not ATP. We also found that the nature of the opening stimulus could alter the permselectivity of the hemichannel-for some of the relatively non-selective connexins, hemichannel opening via depolarisation was ineffective at allowing release of lactate. By performing a mutational analysis, informed by the differential selectivity of the closely related Cx46 and Cx50 hemichannels, we found that the charge on the N-terminus and N-terminus-TM2 interactions are key contributors to permselectivity for ATP.