Ethanol Modulates BK_CaChannels by Acting as an Adjuvant of Calcium

Abstract

Ethanol modulation of calcium-and voltage-gated potassium (slo1) channels alters neuronal excitability, cerebrovascular tone, brain function, and behavior, yet the mechanism of this modulation remains unknown. Using patch-clamp electrophysiology on recombinant BK Ca channels cloned from mouse brain and expressed in Xenopus laevis oocytes, we demonstrate that ethanol, even at concentrations maximally effective to modulate BK Ca channel function (100 mM), fails to gate the channel in absence of activating calcium. Moreover, ethanol does not modify intrinsic, voltage-or physiological magnesium-driven gating. The alcohol works as an adjuvant of calcium by selectively facilitating calcium-driven gating. This facilitation, however, renders differential ethanol effects on channel activity: potentiation at low (Ͻ10 M) and inhibition at high (Ͼ10 M) calcium, this dual pattern remaining largely unmodified by coexpression of brain slo1 channels with the neuronally abundant BK Ca channel ␤ 4 subunit. Calcium recognition by either of the slo1 high-affinity sensors (calcium bowl and RCK1 Asp362/ Asp367) is required for ethanol to amplify channel activation by calcium. The Asp362/Asp367 site, however, is necessary and sufficient to sustain ethanol inhibition. This inhibition also results from ethanol facilitation of calcium action; in this case, ethanol favors channel dwelling in a calcium-driven, lowactivity mode. The agonist-adjuvant mechanism that we advance from the calcium-ethanol interaction on slo1 might be applicable to data of ethanol action on a wide variety of ligandgated channels.

Ethanol-mediated relaxation of guinea pig urinary bladder smooth muscle: involvement of BK and L-type Ca2+ channels

“…To our knowledge, no previous study investigated the effects of EtOH on DSM electrical properties with any configuration of the patch-clamp technique. In non-DSM cells, similar to our findings, acute EtOH exposure increased the whole cell BK currents in a subset of dorsal root ganglion neurons (15) and recombinant BK channels (mSlo) expressed in Xenopus laevis oocytes (30). The latter study also demonstrated the dependency of EtOH-mediated responses on intracellular Ca 2ϩ levels.…”

“…The effects of EtOH on the open dwell time in DSM patches were different from those described previously. Unlike in the recombinant cells expressing mSlo in oocytes, where either decreases in mean open dwell time constant or increases in the longest time constant were seen at ϳ0.3% (ϳ50 mM) (8,30), in DSM patches at ϳ300 nM free [Ca 2ϩ ] we report increases for both shorter and longer time constants (Table 2). We also noted an increase in the proportion of dwell time events favoring the shorter time constant, in contrast to recombinant mSlo channels (8,30), as well as an overall increase in the incidence of BK channel openings.…”

“…The evidence for their involvement has been obtained in studies investigating neuronal or cardiovascular cells (4,9). Additional insights have been provided in recombinant expression systems (10,30). In contrast, relatively little work has been carried out in the urinary bladder.…”

“…In the presence of ϳ2 or 10 M intracellular free [Ca 2ϩ ], it was not possible to reliably calculate open dwell time constants due to frequent simultaneous BK channel openings and high P o . Other investigators using excised patches derived from neuronal, recombinant, and GH3 cells with the [Ca 2ϩ ] ranging from 100 nM to 10 M reported EtOH-induced enhancements in the P o , increases in the open dwell time constant, and decreases in the closed dwell time constant without changing the singlechannel conductance (8,15,26,30). In our study, in excised patches derived from DSM cells at ϳ300 nM intracellular free [Ca 2ϩ ] where the P o was low, we also identified an enhancement of the P o , an increase in the open dwell time constants and no change in single BK channel conductance.…”

Low-Dose Ethanol Preconditioning Protects Against Oxygen-Glucose Deprivation/Reoxygenation-Induced Neuronal Injury By Activating Large Conductance, Ca2+-Activated K+ Channels In Vitro

“…Liu et al found that ethanol fails to gate the channel in the absence of activating Ca 2? , indicating that ethanol modulates the BK Ca channel by acting as an adjuvant of Ca 2? [46]. In Xenopus oocytes, gintonin-mediated BK Ca channel activation is blocked by a PKC inhibitor [47].…”