scholarly journals Selective inhibition of phosphodiesterase 1 relaxes urinary bladder smooth muscle: role for ryanodine receptor-mediated BK channel activation

2012 ◽  
Vol 303 (10) ◽  
pp. C1079-C1089 ◽  
Author(s):  
Wenkuan Xin ◽  
Rupal P. Soder ◽  
Qiuping Cheng ◽  
Eric S. Rovner ◽  
Georgi V. Petkov
Keyword(s):  
Membrane Potential ◽  
Guinea Pig ◽  
Muscle Force ◽  
Selective Inhibition ◽  
Bk Channels ◽  
Bk Channel ◽  
Channel Activation ◽  
Contraction Amplitude ◽  
Intracellular Ca ◽  
Camp Levels

The large conductance voltage- and Ca2+-activated K+ (BK) channel is a major regulator of detrusor smooth muscle (DSM) excitability and contractility. Recently, we showed that nonselective phosphodiesterase (PDE) inhibition reduces guinea pig DSM excitability and contractility by increasing BK channel activity. Here, we investigated how DSM excitability and contractility changes upon selective inhibition of PDE type 1 (PDE1) and the underlying cellular mechanism involving ryanodine receptors (RyRs) and BK channels. PDE1 inhibition with 8-methoxymethyl-3-isobutyl-1-methylxanthine (8MM-IBMX; 10 μM) increased the cAMP levels in guinea pig DSM cells. Patch-clamp experiments on freshly isolated DSM cells showed that 8MM-IBMX increased transient BK currents and the spontaneous transient hyperpolarization (STH) frequency by ∼2.5- and ∼1.8-fold, respectively. 8MM-IBMX hyperpolarized guinea pig and human DSM cell membrane potential and significantly decreased the intracellular Ca2+ levels in guinea pig DSM cells. Blocking BK channels with 1 μM paxilline or inhibiting RyRs with 30 μM ryanodine abolished the STHs and the 8MM-IBMX inhibitory effects on the DSM cell membrane potential. Isometric DSM tension recordings showed that 8MM-IBMX significantly reduced the spontaneous phasic contraction amplitude, muscle force integral, duration, frequency, and tone of DSM isolated strips. The electrical field stimulation-induced DSM contraction amplitude, muscle force integral, and duration were also attenuated by 10 μM 8MM-IBMX. Blocking BK channels with paxilline abolished the 8MM-IBMX effects on DSM contractions. Our data provide evidence that PDE1 inhibition relaxes DSM by raising cellular cAMP levels and subsequently stimulates RyRs, which leads to BK channel activation, membrane potential hyperpolarization, and decrease in intracellular Ca2+ levels.

scholarly journals Large-conductance voltage- and Ca2+-activated K+ channel regulation by protein kinase C in guinea pig urinary bladder smooth muscle

2014 ◽  
Vol 306 (5) ◽  
pp. C460-C470 ◽  
Author(s):  
Kiril L. Hristov ◽  
Amy C. Smith ◽  
Shankar P. Parajuli ◽  
John Malysz ◽  
Georgi V. Petkov
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Guinea Pig ◽  
Bk Channels ◽  
Bk Channel ◽  
K Channel ◽  
Channel Activity ◽  
Open Probability ◽  
Channel Regulation ◽  
Pkc Activation

Large-conductance voltage- and Ca2+-activated K+ (BK) channels are critical regulators of detrusor smooth muscle (DSM) excitability and contractility. PKC modulates the contraction of DSM and BK channel activity in non-DSM cells; however, the cellular mechanism regulating the PKC-BK channel interaction in DSM remains unknown. We provide a novel mechanistic insight into BK channel regulation by PKC in DSM. We used patch-clamp electrophysiology, live-cell Ca2+ imaging, and functional studies of DSM contractility to elucidate BK channel regulation by PKC at cellular and tissue levels. Voltage-clamp experiments showed that pharmacological activation of PKC with PMA inhibited the spontaneous transient BK currents in native freshly isolated guinea pig DSM cells. Current-clamp recordings revealed that PMA significantly depolarized DSM membrane potential and inhibited the spontaneous transient hyperpolarizations in DSM cells. The PMA inhibitory effects on DSM membrane potential were completely abolished by the selective BK channel inhibitor paxilline. Activation of PKC with PMA did not affect the amplitude of the voltage-step-induced whole cell steady-state BK current or the single BK channel open probability (recorded in cell-attached mode) upon inhibition of all major Ca2+ sources for BK channel activation with thapsigargin, ryanodine, and nifedipine. PKC activation with PMA elevated intracellular Ca2+ levels in DSM cells and increased spontaneous phasic and nerve-evoked contractions of DSM isolated strips. Our results support the concept that PKC activation leads to a reduction of BK channel activity in DSM via a Ca2+-dependent mechanism, thus increasing DSM contractility.

scholarly journals BK Channel Gating Mechanisms: Progresses Toward a Better Understanding of Variants Linked Neurological Diseases

2021 ◽  
Vol 12 ◽  
Author(s):  
Jianmin Cui
Keyword(s):  
Neurological Disorders ◽  
Recent Progress ◽  
Molecular Mechanisms ◽  
Neurological Diseases ◽  
Bk Channels ◽  
Bk Channel ◽  
Channel Activation ◽  
Gating Mechanisms ◽  
Intracellular Ca ◽  
Atomistic Structure

The large conductance Ca2+-activated potassium (BK) channel is activated by both membrane potential depolarization and intracellular Ca2+ with distinct mechanisms. Neural physiology is sensitive to the function of BK channels, which is shown by the discoveries of neurological disorders that are associated with BK channel mutations. This article reviews the molecular mechanisms of BK channel activation in response to voltage and Ca2+ binding, including the recent progress since the publication of the atomistic structure of the whole BK channel protein, and the neurological disorders associated with BK channel mutations. These results demonstrate the unique mechanisms of BK channel activation and that these mechanisms are important factors in linking BK channel mutations to neurological disorders.

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

2014 ◽  
Vol 306 (1) ◽  
pp. C45-C58 ◽  
Author(s):  
John Malysz ◽  
Serge A. Y. Afeli ◽  
Aaron Provence ◽  
Georgi V. Petkov
Keyword(s):  
Smooth Muscle ◽  
Urinary Bladder ◽  
Guinea Pig ◽  
Open Channel ◽  
Bk Channels ◽  
Bk Channel ◽  
Muscle Involvement ◽  
Whole Cell ◽  
Intracellular Ca ◽  
Probability Number

Mechanisms underlying ethanol (EtOH)-induced detrusor smooth muscle (DSM) relaxation and increased urinary bladder capacity remain unknown. We investigated whether the large conductance Ca2+-activated K+ (BK) channels or L-type voltage-dependent Ca2+ channels (VDCCs), major regulators of DSM excitability and contractility, are targets for EtOH by patch-clamp electrophysiology (conventional and perforated whole cell and excised patch single channel) and isometric tension recordings using guinea pig DSM cells and isolated tissue strips, respectively. EtOH at 0.3% vol/vol (∼50 mM) enhanced whole cell BK currents at +30 mV and above, determined by the selective BK channel blocker paxilline. In excised patches recorded at +40 mV and ∼300 nM intracellular Ca2+ concentration ([Ca2+]), EtOH (0.1–0.3%) affected single BK channels (mean conductance ∼210 pS and blocked by paxilline) by increasing the open channel probability, number of open channel events, and open dwell-time constants. The amplitude of single BK channel currents and unitary conductance were not altered by EtOH. Conversely, at ∼10 μM but not ∼2 μM intracellular [Ca2+], EtOH (0.3%) decreased the single BK channel activity. EtOH (0.3%) affected transient BK currents (TBKCs) by either increasing frequency or decreasing amplitude, depending on the basal level of TBKC frequency. In isolated DSM strips, EtOH (0.1–1%) reduced the amplitude and muscle force of spontaneous phasic contractions. The EtOH-induced DSM relaxation, except at 1%, was attenuated by paxilline. EtOH (1%) inhibited L-type VDCC currents in DSM cells. In summary, we reveal the involvement of BK channels and L-type VDCCs in mediating EtOH-induced urinary bladder relaxation accommodating alcohol-induced diuresis.

BK channel activation by L-type Ca2+ channels CaV1.2 and CaV1.3 during the subthreshold phase of an action potential

2021 ◽  
Author(s):  
Amber E Plante ◽  
Joshua P Whitt ◽  
Andrea L. Meredith
Keyword(s):  
Membrane Potential ◽  
Action Potential ◽  
Firing Rate ◽  
Low Voltage ◽  
Bk Channels ◽  
Bk Channel ◽  
Action Potential Firing ◽  
Channel Activation ◽  
Potential Firing ◽  
Current Activation

Mammalian circadian (24-hour) rhythms are timed by the pattern of spontaneous action potential firing in the suprachiasmatic nucleus (SCN). This oscillation in firing is produced through circadian regulation of several membrane currents, including large-conductance Ca2+- and voltage-activated K+ (BK) and L-type Ca2+ channel (LTCC) currents. During the day, steady-state BK currents depend mostly on LTCCs for activation, while at night, they depend predominantly on RyRs. However, the contribution of these Ca2+ channels to BK channel activation during action potential firing has not been thoroughly investigated. In this study, we used a pharmacological approach to determine that both LTCCs and RyRs contribute to the baseline membrane potential of SCN action potential waveforms, as well as action potential-evoked BK current, during the day and night, respectively. Since the baseline membrane potential is a major determinant of circadian firing rate, we focused on the LTCCs contributing to low voltage activation of BK channels during the subthreshold phase. For these experiments, two LTCC subtypes found in SCN (CaV1.2 and CaV1.3) were co-expressed with BK channels in heterologous cells, where their differential contributions could be separately measured. CaV1.3 channels produced currents that were shifted to more hyperpolarized potentials compared to CaV1.2, resulting in increased subthreshold Ca2+ and BK currents during an action potential command. These results show that while multiple Ca2+ sources in SCN can contribute to the activation of BK current during an action potential, specific BK-CaV1.3 partnerships may optimize the subthreshold BK current activation that is critical for firing rate regulation.

Functional coupling between the Na+/Ca2+ exchanger and nonselective cation channels during histamine stimulation in guinea pig tracheal smooth muscle

2007 ◽  
Vol 293 (1) ◽  
pp. L191-L198 ◽  
Author(s):  
Paola Algara-Suárez ◽  
Catalina Romero-Méndez ◽  
Tom Chrones ◽  
Sergio Sánchez-Armass ◽  
Ulises Meza ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Force Measurement ◽  
Isometric Force ◽  
Sustained Response ◽  
Functional Coupling ◽  
Channel Activation ◽  
Histamine Stimulation ◽  
Nonselective Cation Channels ◽  
Intracellular Ca

Airway smooth muscle (ASM) contracts partly due to an increase in cytosolic Ca2+. In this work, we found that the contraction caused by histamine depends on external Na+, possibly involving nonselective cationic channels (NSCC) and the Na+/Ca2+ exchanger (NCX). We performed various protocols using isometric force measurement of guinea pig tracheal rings stimulated by histamine. We observed that force reached 53 ± 1% of control during external Na+ substitution by N-methyl-d-glucamine+, whereas substitution by Li+ led to no significant change (91 ± 1%). Preincubation with KB-R7943 decreased the maximal force developed (52.3 ± 5.6%), whereas preincubation with nifedipine did not (89.7 ± 1.8%). Also, application of the nonspecific NCX blocker KB-R7943 and nifedipine on histamine-precontracted tracheal rings reduced force to 1 ± 3%, significantly different from nifedipine alone (49 ± 6%). Moreover, nonspecific NSCC inhibitors SKF-96365 and 2-aminoethyldiphenyl borate reduced force to 1 ± 1% and 19 ± 7%, respectively. Intracellular Ca2+ measurements in isolated ASM cells showed that KB-R7943 and SKF-96365 reduced the peak and sustained response to histamine (0.20 ± 0.1 and 0.19 ± 0.09 for KB-R, 0.43 ± 0.16 and 0.47 ± 0.18 for SKF, expressed as mean of differences). Moreover, Na+-free solution only inhibited the sustained response (0.54 ± 0.25). These data support an important role for NSCC and NCX during histamine stimulation. We speculate that histamine induces Na+ influx through NSCC that promotes the Ca2+ entry mode of NCX and CaV1.2 channel activation, thereby causing contraction.

Na+/Ca2+ exchange and its role in intracellular Ca2+ regulation in guinea pig detrusor smooth muscle

2001 ◽  
Vol 280 (5) ◽  
pp. C1090-C1096 ◽  
Author(s):  
C. Wu ◽  
C. H. Fry
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Guinea Pig ◽  
Outward Current ◽  
Detrusor Smooth Muscle ◽  
Small Rise ◽  
Intracellular Ca ◽  
Net Flux ◽  
Isolated Smooth Muscle Cells

The role of Na+/Ca2+ exchange in regulating intracellular Ca2+ concentration ([Ca2+]i) in isolated smooth muscle cells from the guinea pig urinary bladder was investigated. Incremental reduction of extracellular Na+ concentration resulted in a graded rise of [Ca2+]i; 50–100 μM strophanthidin also increased [Ca2+]i. A small outward current accompanied the rise of [Ca2+]i in low-Na+ solutions (17.1 ± 1.8 pA in 29.4 mM Na+). The quantity of Ca2+ influx through the exchanger was estimated from the charge carried by the outward current and was ∼30 times that which is necessary to account for the rise of [Ca2+]i, after correction was made for intracellular Ca2+ buffering. Ca2+ influx through the exchanger was able to load intracellular Ca2+ stores. It is concluded that the level of resting [Ca2+]i is not determined by the exchanger, and under resting conditions (membrane potential −50 to −60 mV), there is little net flux through the exchanger. However, a small rise of intracellular Na+ concentration would be sufficient to generate significant net Ca2+ influx.

scholarly journals BK Channels in Rat and Human Pulmonary Smooth Muscle Cells are BKα-β1 Functional Complexes Lacking the Oxygen-Sensitive Stress Axis Regulated Exon Insert

2016 ◽  
Vol 6 (4) ◽  
pp. 563-575 ◽  
Author(s):  
Neil D. Detweiler ◽  
Li Song ◽  
Samantha J. McClenahan ◽  
Rachel J. Versluis ◽  
Sujay V. Kharade ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Splice Variants ◽  
Muscle Cells ◽  
Bk Channels ◽  
Bk Channel ◽  
Stress Axis ◽  
Human Donor ◽  
Intracellular Ca ◽  
Oxygen Sensitive

A loss of K+ efflux in pulmonary arterial smooth muscle cells (PASMCs) contributes to abnormal vasoconstriction and PASMC proliferation during pulmonary hypertension (PH). Activation of high-conductance Ca2+-activated (BK) channels represents a therapeutic strategy to restore K+ efflux to the affected PASMCs. However, the properties of BK channels in PASMCs—including sensitivity to BK channel openers (BKCOs)—are poorly defined. The goal of this study was to compare the properties of BK channels between PASMCs of normoxic (N) and chronic hypoxic (CH) rats and then explore key findings in human PASMCs. Polymerase chain reaction results revealed that 94.3% of transcripts encoding BKα pore proteins in PASMCs from N rats represent splice variants lacking the stress axis regulated exon insert, which confers oxygen sensitivity. Subsequent patch-clamp recordings from inside-out (I-O) patches confirmed a dense population of BK channels insensitive to hypoxia. The BK channels were highly activated by intracellular Ca2+ and the BKCO lithocholate; these responses require BK α-β1 subunit coupling. PASMCs of CH rats with established PH exhibited a profound overabundance of the dominant oxygen-insensitive BKα variant. Importantly, human BK (hBK) channels in PASMCs from human donor lungs also represented the oxygen-insensitive BKα variant activated by BKCOs. The hBK channels showed significantly enhanced Ca2+ sensitivity compared with rat BK channels. We conclude that rat BK and hBK channels in PASMCs are oxygen-insensitive BK α-β1 complexes highly sensitive to Ca2+ and the BKCO lithocholate. BK channels are overexpressed in PASMCs of a rat model of PH and may provide an abundant target for BKCOs designed to restore K+ efflux.

scholarly journals Transduction of Voltage and Ca2+ Signals by Slo1 BK Channels

Physiology ◽  
2013 ◽  
Vol 28 (3) ◽  
pp. 172-189 ◽  
Author(s):  
T. Hoshi ◽  
A. Pantazis ◽  
R. Olcese
Keyword(s):  
Gate Voltage ◽  
Bk Channels ◽  
Activation Mechanism ◽  
Channel Activation ◽  
Regulatory Factors ◽  
Voltage Sensors ◽  
Voltage Gated ◽  
Intracellular Ca ◽  
Biophysical Mechanism ◽  
Allosteric Model

Large-conductance Ca2+- and voltage-gated K+ channels are activated by an increase in intracellular Ca2+ concentration and/or depolarization. The channel activation mechanism is well described by an allosteric model encompassing the gate, voltage sensors, and Ca2+ sensors, and the model is an excellent framework to understand the influences of auxiliary β and γ subunits and regulatory factors such as Mg2+. Recent advances permit elucidation of structural correlates of the biophysical mechanism.

scholarly journals Molecular mechanism of BK channel activation by the smooth muscle relaxant NS11021

2020 ◽  
Vol 152 (6) ◽  
Author(s):  
Michael E. Rockman ◽  
Alexandre G. Vouga ◽  
Brad S. Rothberg
Keyword(s):  
Smooth Muscle ◽  
Muscle Relaxant ◽  
Time Course ◽  
Bk Channels ◽  
Bk Channel ◽  
Neurological Deficits ◽  
Open Probability ◽  
Channel Activation ◽  
Voltage Sensors ◽  
Smooth Muscle Relaxant

Large-conductance Ca2+-activated K+ channels (BK channels) are activated by cytosolic calcium and depolarized membrane potential under physiological conditions. Thus, these channels control electrical excitability in neurons and smooth muscle by gating K+ efflux and hyperpolarizing the membrane in response to Ca2+ signaling. Altered BK channel function has been linked to epilepsy, dyskinesia, and other neurological deficits in humans, making these channels a key target for drug therapies. To gain insight into mechanisms underlying pharmacological modulation of BK channel gating, here we studied mechanisms underlying activation of BK channels by the biarylthiourea derivative, NS11021, which acts as a smooth muscle relaxant. We observe that increasing NS11021 shifts the half-maximal activation voltage for BK channels toward more hyperpolarized voltages, in both the presence and nominal absence of Ca2+, suggesting that NS11021 facilitates BK channel activation primarily by a mechanism that is distinct from Ca2+ activation. 30 µM NS11021 slows the time course of BK channel deactivation at −200 mV by ∼10-fold compared with 0 µM NS11021, while having little effect on the time course of activation. This action is most pronounced at negative voltages, at which the BK channel voltage sensors are at rest. Single-channel kinetic analysis further shows that 30 µM NS11021 increases open probability by 62-fold and increases mean open time from 0.15 to 0.52 ms in the nominal absence of Ca2+ at voltages less than −60 mV, conditions in which BK voltage sensors are largely in the resting state. We could therefore account for the major activating effects of NS11021 by a scheme in which the drug primarily shifts the pore-gate equilibrium toward the open state.

scholarly journals Spontaneous Transient Outward Currents Arise from Microdomains Where BK Channels Are Exposed to a Mean Ca2+ Concentration on the Order of 10 μM during a Ca2+ Spark

2002 ◽  
Vol 120 (1) ◽  
pp. 15-27 ◽  
Author(s):  
Ronghua ZhuGe ◽  
Kevin E. Fogarty ◽  
Richard A. Tuft ◽  
John V. Walsh
Keyword(s):  
Voltage Dependence ◽  
Ryanodine Receptors ◽  
Bk Channels ◽  
Bk Channel ◽  
Channel Activation ◽  
Membrane Area ◽  
Outward Currents ◽  
Cell Recording ◽  
Excised Patches ◽  
Spontaneous Transient Outward Currents

Ca2+ sparks are small, localized cytosolic Ca2+ transients due to Ca2+ release from sarcoplasmic reticulum through ryanodine receptors. In smooth muscle, Ca2+ sparks activate large conductance Ca2+-activated K+ channels (BK channels) in the spark microdomain, thus generating spontaneous transient outward currents (STOCs). The purpose of the present study is to determine experimentally the level of Ca2+ to which the BK channels are exposed during a spark. Using tight seal, whole-cell recording, we have analyzed the voltage-dependence of the STOC conductance (g(STOC)), and compared it to the voltage-dependence of BK channel activation in excised patches in the presence of different [Ca2+]s. The Ca2+ sparks did not change in amplitude over the range of potentials of interest. In contrast, the magnitude of g(STOC) remained roughly constant from 20 to −40 mV and then declined steeply at more negative potentials. From this and the voltage dependence of BK channel activation, we conclude that the BK channels underlying STOCs are exposed to a mean [Ca2+] on the order of 10 μM during a Ca2+ spark. The membrane area over which a concentration ≥10 μM is reached has an estimated radius of 150–300 nm, corresponding to an area which is a fraction of one square micron. Moreover, given the constraints imposed by the estimated channel density and the Ca2+ current during a spark, the BK channels do not appear to be uniformly distributed over the membrane but instead are found at higher density at the spark site.

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