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Requirement for functional BK channels in maintaining oscillation in venomotor tone revealed by species differences in expression of the ?1 accessory subunits.

ABSTRACT: We determined the possible role of large-conductance Ca2+-activated K (BK) channels in regulation of venous tone in small capacitance veins and blood pressure. In rat mesenteric venous smooth muscle cells (MV SMC), BK channel ?- and ?1-subunits were coexpressed, unitary BK currents were detected, and single-channel currents were sensitive to voltage and [Ca2+]i. Rat MV SMCs displayed Ca sparks and iberiotoxin-sensitive spontaneous transient outward currents. Under resting conditions in vitro, rat MV exhibited nifedipine-sensitive spontaneous oscillatory constrictions. Blockade of BK channels by paxilline and Ca2+ sparks by ryanodine constricted rat MV. Nifedipine caused venodilation and blocked paxilline-induced, KCl-induced (20 mM), and BayK8644-induced contraction. Acute inhibition of BK channels with iberiotoxin in vivo increased blood pressure and reduced venous capacitance, measured as an increase in mean circulatory filling pressure in conscious rats. BK channel ?-subunits and L-type Ca2+ channel ?1-C subunits are expressed in murine MV. However, these channels are not functional because murine MV lack nifedipine-sensitive basal tone and rhythmic constrictions. Murine MV were also insensitive to paxilline, ryanodine, KCl, and BayK8644, consistent with our previous studies showing that murine MV do not have BK ?1-subunits. These data show that not only there are species-dependent properties in ion channel control of venomotor tone but also BK channels are required for rhythmic oscillations in venous tone.

SUBMITTER: Xu H

PROVIDER: S-EPMC3248636 | biostudies-literature | 2012 Jan

REPOSITORIES: biostudies-literature

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Requirement for functional BK channels in maintaining oscillation in venomotor tone revealed by species differences in expression of the β1 accessory subunits.

Xu Hui H Kandlikar Sachin S SS Westcott Erika B EB Fink Gregory D GD Galligan James J JJ

Journal of cardiovascular pharmacology 20120101 1

We determined the possible role of large-conductance Ca2+-activated K (BK) channels in regulation of venous tone in small capacitance veins and blood pressure. In rat mesenteric venous smooth muscle cells (MV SMC), BK channel α- and β1-subunits were coexpressed, unitary BK currents were detected, and single-channel currents were sensitive to voltage and [Ca2+]i. Rat MV SMCs displayed Ca sparks and iberiotoxin-sensitive spontaneous transient outward currents. Under resting conditions in vitro, ra ...[more]

PMID: 21885988

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Project description:Voltage/Ca²?(i)-gated, large conductance K+ (BK) channels result from tetrameric association of ? (slo1) subunits. In most tissues, BK protein complexes include regulatory ? subunits that contain two transmembrane domains (TM1, TM2), an extracellular loop, and two short intracellular termini. Four BK ? types have been identified, each presenting a rather selective tissue-specific expression profile. Thus, BK ? modifies current phenotype to suit physiology in a tissue-specific manner. The smooth muscle-abundant BK ?1 drastically increases the channel's apparent Ca²?(i) sensitivity. The resulting phenotype is critical for BK channel activity to increase in response to Ca2+ levels reached near the channel during depolarization-induced Ca2+ influx and myocyte contraction. The eventual BK channel activation generates outward K+ currents that drive the membrane potential in the negative direction and eventually counteract depolarization-induced Ca2+ influx. The BK ?1 regions responsible for the characteristic phenotype of ?1-containing BK channels remain to be identified. We used patch-clamp electrophysiology on channels resulting from the combination of smooth muscle slo1 (cbv1) subunits with smooth muscle-abundant ?1, neuron-abundant ?4, or chimeras constructed by swapping ?1 and ?4 regions, and determined the contribution of specific ?1 regions to the BK phenotype. At Ca2+ levels found near the channel during myocyte contraction (10 µM), channel complexes that included chimeras having both TMs from ?1 and the remaining regions ("background") from ?4 showed a phenotype (V(half), ?(act), ?(deact)) identical to that of complexes containing wt ?1. This phenotype could not be evoked by complexes that included chimeras combining either ?1 TM1 or ?1 TM2 with a ?4 background. Likewise, ? "halves" (each including ?1 TM1 or ?1 TM2) resulting from interrupting the continuity of the EC loop failed to render the normal phenotype, indicating that physical connection between ?1 TMs via the EC loop is also necessary for proper channel function.

Project description:Arachidonic acid (AA) inhibits the activity of several different voltage-gated Ca(2+) channels by an unknown mechanism at an unknown site. The Ca(2+) channel pore-forming subunit (Ca(V)alpha(1)) is a candidate for the site of AA inhibition because T-type Ca(2+) channels, which do not require accessory subunits for expression, are inhibited by AA. Here, we report the unanticipated role of accessory Ca(V)beta subunits on the inhibition of Ca(V)1.3b L-type (L-) current by AA. Whole cell Ba(2+) currents were measured from recombinant channels expressed in human embryonic kidney 293 cells at a test potential of -10 mV from a holding potential of -90 mV. A one-minute exposure to 10 microM AA inhibited currents with beta(1b), beta(3), or beta(4) 58, 51, or 44%, respectively, but with beta(2a) only 31%. At a more depolarized holding potential of -60 mV, currents were inhibited to a lesser degree. These data are best explained by a simple model where AA stabilizes Ca(V)1.3b in a deep closed-channel conformation, resulting in current inhibition. Consistent with this hypothesis, inhibition by AA occurred in the absence of test pulses, indicating that channels do not need to open to become inhibited. AA had no effect on the voltage dependence of holding potential-dependent inactivation or on recovery from inactivation regardless of Ca(V)beta subunit. Unexpectedly, kinetic analysis revealed evidence for two populations of L-channels that exhibit willing and reluctant gating previously described for Ca(V)2 channels. AA preferentially inhibited reluctant gating channels, revealing the accelerated kinetics of willing channels. Additionally, we discovered that the palmitoyl groups of beta(2a) interfere with inhibition by AA. Our novel findings that the Ca(V)beta subunit alters kinetic changes and magnitude of inhibition by AA suggest that Ca(V)beta expression may regulate how AA modulates Ca(2+)-dependent processes that rely on L-channels, such as gene expression, enzyme activation, secretion, and membrane excitability.

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Requirement for functional BK channels in maintaining oscillation in venomotor tone revealed by species differences in expression of the ?1 accessory subunits.

Publications

Requirement for functional BK channels in maintaining oscillation in venomotor tone revealed by species differences in expression of the β1 accessory subunits.

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