Stable expression of the human large‐conductance Ca2+‐activated K+ channel α‐ and β‐subunits in HEK293 cells

摘要

>We have generated HEK293 cell lines stably expressing high levels of either the human BK channel α-subunit alone or the BK channel α-subunit and β-subunit together. For co-expression a plasmid with three expression cassettes was constructed. Patch-clamp recordings on inside-out patches from the transfected cells resulted in macroscopic currents reflecting the expression of 200–800 BK channels per patch. No decrease in channel expression could be detected in cells grown for more than 50 passages. The α-subunit when expressed alone conducted currents which were sensitive to intracellular Ca2+ in the physiological range. In the presence of the β-subunit the steady-state activation curves were shifted by −20 to −30 mV and channel deactivation kinetics were slowed. The BK channel opener NS1608 (10 μM) shifted the steady-state activation curves for the α-subunit as well as for the αβ-subunits by −40 to −50 mV.