Coupling asymmetry of heterotypic connexin 45/connexin 43-EGFP gap junctions: Properties of fast and slow gating mechanisms

Although fast and slow gating mechanisms have been described in gap junctions (GJs), their relative contributions to dependence on transjunctional voltage, V_j, is still unclear. We used cell lines expressing wild-type connexin 45 (Cx45) and connexin 43 fused with enhanced green fluorescent protein (Cx43-EGFP) to examine mechanisms of gating in homo- and heterotypic GJs formed of these connexins. Macroscopically Cx45/Cx45 channels show high sensitivity to V_i. Cx45 channels demonstrate two types of gating: fast transitions between open and residual states and slow transitions between open and completely closed states. Single-channel conductance of the Cx45 channel is ≈32 pS for the open state and ≈4 pS for the residual state. Cx45/Cx43-EGFP heterotypic junctions exhibit very asymmetrical Vj gating with the maximum junctional conductance shifted to Vj positive on the Cx45 side. Conductance of single Cx45/Cx43-EGFP channels is ≈55 pS for the open state and ≈4 pS for the residual state, values consistent with the simple-series connection of Cx45 and Cx43-EGFP hemichannels. At Vi = 0, the slow gate of many Cx45 hemichannels is closed in both homotypic Cx45/Cx45 and heterotypic Cx45/Cx43-EGFP junctions. Fast and slow Vj gates of both Cx45 and Cx43 hemichannels close for relative negativity at their cytoplasmic end. Coupling mediated by Cx45/Cx43-EGFP junctions can exhibit asymmetry that can be strongly modulated by small changes in difference of holding potentials. Cx45/Cx43 junctions are likely to be found in brain and heart and may mediate rectifying electrical transmission or modulatable chemical communication.