Voltage-dependent facilitation of Cx46 hemichannels.

Gap junction channels are formed by two hemichannels in series (one from each neighboring cell), which are in turn connexin hexamers. Under normal conditions, hemichannels at the plasma membrane are mostly closed but can be opened by changes in membrane voltage, extracellular divalent ion concentration, phosphorylation, pH, and redox potential. Recently, interactions between channels have been found to modulate the activity of several ion channels, including gap junction channels. Here, we studied whether connexin46 (Cx46) hemichannels display such behavior. We studied the response of the Cx46 hemichannels expressed in Xenopus laevis oocytes to consecutive depolarization pulses. Hemichannels formed by wild-type Cx46 and a COOH-terminal domain truncation mutant (Cx46DeltaCT) were activated by voltage pulses. When the hemichannels were depolarized repeatedly from -60 mV to +80 mV, the amplitude of the outward and tail currents increased progressively with successive pulses. This phenomenon ("current facilitation") depended on the amplitude of the depolarization, reaching a maximum at approximately +60 mV in oocytes expressing Cx46, and on the interval between pulses, disappearing with intervals longer than about 20 s. The current facilitation was also present in oocytes expressing Cx46DeltaCT, ruling out a primary role of this domain in the facilitation. Nominal removal of divalent cations from the extracellular side caused maximal current activation of Cx46 and Cx46DeltaCT hemichannels and prevented facilitation. The results suggest that Cx46 hemichannels show a cooperative activation independent of their COOH-terminal domain.