Alcohol modulation of BK channel gating depends on β subunit composition

Guruprasad Kuntamallappanavar, Alejandro Dopico

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

In most mammalian tissues, Ca2+ i/voltage-gated, large conductance K+ (BK) channels consist of channel-forming slo1 and auxiliary (β1-β4) subunits. When Ca2+ i (3-20 μM) reaches the vicinity of BK channels and increases their activity at physiological voltages, β1- and β4-containing BK channels are, respectively, inhibited and potentiated by intoxicating levels of ethanol (50 mM). Previous studies using different slo1s, lipid environments, and Ca2+ i concentrations-all determinants of the BK response to ethanol-made it impossible to determine the specific contribution of β subunits to ethanol action on BK activity. Furthermore, these studies measured ethanol action on ionic current under a limited range of stimuli, rendering no information on the gating processes targeted by alcohol and their regulation by βs. Here, we used identical experimental conditions to obtain single-channel and macroscopic currents of the same slo1 channel ("cbv1" from rat cerebral artery myocytes) in the presence and absence of 50 mM ethanol. First, we assessed the role five different β subunits (1,2,2-IR, 3-variant d, and 4) in ethanol action on channel function. Thus, two phenotypes were identified: (1) ethanol potentiated cbv1-,cbv1+β3-, and cbv1+β4-mediated currents at low Ca2+ i while inhibiting current at high Ca2+ i, the potentiation-inhibition crossover occurring at 20 μM Ca2+ i; (2) for cbv1+β1, cbv1+wt β2, and cbv1+β2-IR, this crossover was shifted to ~3 μM Ca2+ i. Second, applying Horrigan-Aldrich gating analysis on both phenotypes, we show that ethanol fails to modify intrinsic gating and the voltage-dependent parameters under examination. For cbv1, however, ethanol (a) drastically increases the channel's apparent Ca2+ affinity (nine-times decrease in Kd) and (b) very mildly decreases allosteric coupling between Ca2+ binding and channel opening (C). The decreased Kd leads to increased channel activity. For cbv1+β1, ethanol (a) also decreases Kd, yet this decrease (two times) is much smaller than that of cbv1; (b) reduces C; and (c) decreases coupling between Ca2+ binding and voltage sensing (parameter E). Decreased allosteric coupling leads to diminished BK activity. Thus, we have identified critical gating modifications that lead to the differential actions of ethanol on slo1 with and without different β subunits.

Original languageEnglish (US)
Pages (from-to)419-440
Number of pages22
JournalJournal of General Physiology
Volume148
Issue number5
DOIs
StatePublished - Jan 1 2016

Fingerprint

Large-Conductance Calcium-Activated Potassium Channels
Ethanol
Alcohols
Phenotype
Cerebral Arteries
Muscle Cells

All Science Journal Classification (ASJC) codes

  • Physiology

Cite this

Alcohol modulation of BK channel gating depends on β subunit composition. / Kuntamallappanavar, Guruprasad; Dopico, Alejandro.

In: Journal of General Physiology, Vol. 148, No. 5, 01.01.2016, p. 419-440.

Research output: Contribution to journalArticle

@article{d49e20cd01b24b99bc041ca5268d675b,
title = "Alcohol modulation of BK channel gating depends on β subunit composition",
abstract = "In most mammalian tissues, Ca2+ i/voltage-gated, large conductance K+ (BK) channels consist of channel-forming slo1 and auxiliary (β1-β4) subunits. When Ca2+ i (3-20 μM) reaches the vicinity of BK channels and increases their activity at physiological voltages, β1- and β4-containing BK channels are, respectively, inhibited and potentiated by intoxicating levels of ethanol (50 mM). Previous studies using different slo1s, lipid environments, and Ca2+ i concentrations-all determinants of the BK response to ethanol-made it impossible to determine the specific contribution of β subunits to ethanol action on BK activity. Furthermore, these studies measured ethanol action on ionic current under a limited range of stimuli, rendering no information on the gating processes targeted by alcohol and their regulation by βs. Here, we used identical experimental conditions to obtain single-channel and macroscopic currents of the same slo1 channel ({"}cbv1{"} from rat cerebral artery myocytes) in the presence and absence of 50 mM ethanol. First, we assessed the role five different β subunits (1,2,2-IR, 3-variant d, and 4) in ethanol action on channel function. Thus, two phenotypes were identified: (1) ethanol potentiated cbv1-,cbv1+β3-, and cbv1+β4-mediated currents at low Ca2+ i while inhibiting current at high Ca2+ i, the potentiation-inhibition crossover occurring at 20 μM Ca2+ i; (2) for cbv1+β1, cbv1+wt β2, and cbv1+β2-IR, this crossover was shifted to ~3 μM Ca2+ i. Second, applying Horrigan-Aldrich gating analysis on both phenotypes, we show that ethanol fails to modify intrinsic gating and the voltage-dependent parameters under examination. For cbv1, however, ethanol (a) drastically increases the channel's apparent Ca2+ affinity (nine-times decrease in Kd) and (b) very mildly decreases allosteric coupling between Ca2+ binding and channel opening (C). The decreased Kd leads to increased channel activity. For cbv1+β1, ethanol (a) also decreases Kd, yet this decrease (two times) is much smaller than that of cbv1; (b) reduces C; and (c) decreases coupling between Ca2+ binding and voltage sensing (parameter E). Decreased allosteric coupling leads to diminished BK activity. Thus, we have identified critical gating modifications that lead to the differential actions of ethanol on slo1 with and without different β subunits.",
author = "Guruprasad Kuntamallappanavar and Alejandro Dopico",
year = "2016",
month = "1",
day = "1",
doi = "10.1085/jgp.201611594",
language = "English (US)",
volume = "148",
pages = "419--440",
journal = "Journal of General Physiology",
issn = "0022-1295",
publisher = "Rockefeller University Press",
number = "5",

}

TY - JOUR

T1 - Alcohol modulation of BK channel gating depends on β subunit composition

AU - Kuntamallappanavar, Guruprasad

AU - Dopico, Alejandro

PY - 2016/1/1

Y1 - 2016/1/1

N2 - In most mammalian tissues, Ca2+ i/voltage-gated, large conductance K+ (BK) channels consist of channel-forming slo1 and auxiliary (β1-β4) subunits. When Ca2+ i (3-20 μM) reaches the vicinity of BK channels and increases their activity at physiological voltages, β1- and β4-containing BK channels are, respectively, inhibited and potentiated by intoxicating levels of ethanol (50 mM). Previous studies using different slo1s, lipid environments, and Ca2+ i concentrations-all determinants of the BK response to ethanol-made it impossible to determine the specific contribution of β subunits to ethanol action on BK activity. Furthermore, these studies measured ethanol action on ionic current under a limited range of stimuli, rendering no information on the gating processes targeted by alcohol and their regulation by βs. Here, we used identical experimental conditions to obtain single-channel and macroscopic currents of the same slo1 channel ("cbv1" from rat cerebral artery myocytes) in the presence and absence of 50 mM ethanol. First, we assessed the role five different β subunits (1,2,2-IR, 3-variant d, and 4) in ethanol action on channel function. Thus, two phenotypes were identified: (1) ethanol potentiated cbv1-,cbv1+β3-, and cbv1+β4-mediated currents at low Ca2+ i while inhibiting current at high Ca2+ i, the potentiation-inhibition crossover occurring at 20 μM Ca2+ i; (2) for cbv1+β1, cbv1+wt β2, and cbv1+β2-IR, this crossover was shifted to ~3 μM Ca2+ i. Second, applying Horrigan-Aldrich gating analysis on both phenotypes, we show that ethanol fails to modify intrinsic gating and the voltage-dependent parameters under examination. For cbv1, however, ethanol (a) drastically increases the channel's apparent Ca2+ affinity (nine-times decrease in Kd) and (b) very mildly decreases allosteric coupling between Ca2+ binding and channel opening (C). The decreased Kd leads to increased channel activity. For cbv1+β1, ethanol (a) also decreases Kd, yet this decrease (two times) is much smaller than that of cbv1; (b) reduces C; and (c) decreases coupling between Ca2+ binding and voltage sensing (parameter E). Decreased allosteric coupling leads to diminished BK activity. Thus, we have identified critical gating modifications that lead to the differential actions of ethanol on slo1 with and without different β subunits.

AB - In most mammalian tissues, Ca2+ i/voltage-gated, large conductance K+ (BK) channels consist of channel-forming slo1 and auxiliary (β1-β4) subunits. When Ca2+ i (3-20 μM) reaches the vicinity of BK channels and increases their activity at physiological voltages, β1- and β4-containing BK channels are, respectively, inhibited and potentiated by intoxicating levels of ethanol (50 mM). Previous studies using different slo1s, lipid environments, and Ca2+ i concentrations-all determinants of the BK response to ethanol-made it impossible to determine the specific contribution of β subunits to ethanol action on BK activity. Furthermore, these studies measured ethanol action on ionic current under a limited range of stimuli, rendering no information on the gating processes targeted by alcohol and their regulation by βs. Here, we used identical experimental conditions to obtain single-channel and macroscopic currents of the same slo1 channel ("cbv1" from rat cerebral artery myocytes) in the presence and absence of 50 mM ethanol. First, we assessed the role five different β subunits (1,2,2-IR, 3-variant d, and 4) in ethanol action on channel function. Thus, two phenotypes were identified: (1) ethanol potentiated cbv1-,cbv1+β3-, and cbv1+β4-mediated currents at low Ca2+ i while inhibiting current at high Ca2+ i, the potentiation-inhibition crossover occurring at 20 μM Ca2+ i; (2) for cbv1+β1, cbv1+wt β2, and cbv1+β2-IR, this crossover was shifted to ~3 μM Ca2+ i. Second, applying Horrigan-Aldrich gating analysis on both phenotypes, we show that ethanol fails to modify intrinsic gating and the voltage-dependent parameters under examination. For cbv1, however, ethanol (a) drastically increases the channel's apparent Ca2+ affinity (nine-times decrease in Kd) and (b) very mildly decreases allosteric coupling between Ca2+ binding and channel opening (C). The decreased Kd leads to increased channel activity. For cbv1+β1, ethanol (a) also decreases Kd, yet this decrease (two times) is much smaller than that of cbv1; (b) reduces C; and (c) decreases coupling between Ca2+ binding and voltage sensing (parameter E). Decreased allosteric coupling leads to diminished BK activity. Thus, we have identified critical gating modifications that lead to the differential actions of ethanol on slo1 with and without different β subunits.

UR - http://www.scopus.com/inward/record.url?scp=84995877262&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84995877262&partnerID=8YFLogxK

U2 - 10.1085/jgp.201611594

DO - 10.1085/jgp.201611594

M3 - Article

C2 - 27799321

AN - SCOPUS:84995877262

VL - 148

SP - 419

EP - 440

JO - Journal of General Physiology

JF - Journal of General Physiology

SN - 0022-1295

IS - 5

ER -