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Department of Physiology and Biophysics, Finch University of Health Sciences, North Chicago, Illinois 60064 USA
Correspondence: Address reprint requests to Lisa Ebihara MD/PhD, Dept. of Physiology and Biophysics, Finch University of Health Sciences/The Chicago Medical School, 3333 Green Bay Rd., North Chicago, IL 60064-3095. Tel.: 847-578-3424; Fax: 847-578-3265; E-mail: Lisa.Ebihara{at}finchcms.edu.
One of the most striking features of hemi-gap-junctional channels is that they are dramatically modulated by extracellular divalent cations. In this study, we characterized the effects of external divalent cations and voltage on macroscopic human connexin46 (hCx46) hemi-gap-junctional currents using the two-electrode voltage-clamp technique. Increasing extracellular magnesium resulted in a shift of the voltage dependence of activation to more positive potentials, a decrease in the maximum conductance, an acceleration of deactivation, and a slowing of activation. Hyperpolarizing the membrane potential could mimic the effect of raising external magnesium on the activation kinetics and maximum conductance. These results could be interpreted in terms of a sequential model of channel activation with two independent divalent cation binding sites. This model could also explain the effects of external calcium on hCx46 hemichannels. However, the apparent binding affinities for calcium were significantly higher than for magnesium. In addition, we identified a mutation in the first extracellular domain of hCx46 (hCx46*N63S) that resulted in hemichannels that showed increased sensitivity to magnesium blockade.
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