Beta Subunit Transmembrane Domains Determine the Differential Ethanol Responses of β1- vs. β4-Containing BK Channels

Abstract

Ethanol (10-100 mM) usually decreases the steady-state activity (Po) of vascular smooth muscle BK channels while increasing Po of neuronal BK channels (Brodie et al., 2007). Native BK complexes consist of channel-forming α and tissue-specific, accessory β subunits. Four β subunit types were identified, with β1 and β4 prevailing in smooth muscle and neurons, respectively (Brenner et al., 2000). Remarkably, differential expression of β1- vs. β4-containing BK channels modifies Po adaptation to protracted ethanol exposure (Feinberg-Zadek et al., 2008; Martin et al., 2008). On the other hand, β1 is necessary for ethanol to inhibit vascular smooth muscle BK channels and the resulting vasoconstriction (Bukiya et al., 2009). To begin to address the mechanisms and protein regions contributing to differential ethanol actions on BK channels, we expressed channel-forming cbv1 subunits in absence and presence of β1 or β4. Then, we conducted G/Gmax-V plots from ionic currents in inside-out patches and evaluated their modification by acute ethanol exposure. As described for other BK channels (Orio et al., 2002), β1 but not β4 significantly reduced cbv1 V0.5 across 1-1,000 μM calcium. Ethanol (50 mM) decreased homomeric cbv1 V0.5 at low calcium while increasing V0.5 at high calcium, the “crossover” from current potentiation to inhibition occurring at 20 μM calcium. This crossover was left-shifted to 2 μM by β1 co-expression but unaltered by β4 co-expression. Therefore, β1 enables ethanol inhibition of current at calcium levels that are reached near the BK channel in myocytes. Evaluation of G/Gmax-V plots from currents mediated by cbv1±chimeric β1/β4 demonstrated that ethanol responses of complexes containing β chimeras that included β1 TM domains mimicked cbv1+wtβ1 responses. Conversely, ethanol responses of complexes containing β chimeras that included β4 TM domains mimicked cbv1+wtβ4 responses. Support:R37-AA011560 (AMD).