BK channel regulation by phosphodiesterase type 1: a novel signaling pathway controlling human detrusor smooth muscle function

Large-conductance Ca2+-activated K+ (BK) channels are critical regulators of detrusor smooth muscle (DSM) function. We aimed to investigate phosphodiesterase type 1 (PDE1) interactions with BK channels in human DSM to determine the mechanism by which PDE1 regulates human urinary bladder physiology. A combined electrophysiological, functional, and pharmacological approach was applied using human DSM specimens obtained from open bladder surgeries. The perforated whole cell patch-clamp technique was used to record transient BK currents (TBKCs) and the cell membrane potential in freshly isolated human DSM cells in combination with the selective PDE1 inhibitor, 8-methoxymethyl-3-isobutyl-1-methylxanthine (8MM-IBMX). Isometric DSM tension recordings were used to measure spontaneous phasic and electrical field stimulation-induced contractions in human DSM isolated strips. Selective pharmacological inhibition of PDE1 with 8MM-IBMX (10 μM) increased TBKC activity in human DSM cells, which was abolished by subsequent inhibition of protein kinase A (PKA) with H-89 (10 μM). The stimulatory effect of 8MM-IBMX on TBKCs was reversed upon activation of muscarinic acetylcholine receptors with carbachol (1 μM). 8MM-IBMX (10 μM) hyperpolarized the DSM cell membrane potential, an effect blocked by PKA inhibition. 8MM-IBMX significantly decreased spontaneous phasic and nerve-evoked contractions of human DSM isolated strips. The results reveal a novel mechanism that pharmacological inhibition of PDE1 attenuates human DSM excitability and contractility by activating BK channels via a PKA-dependent mechanism. The data also suggest interactions between PDE1 and muscarinic signaling pathways in human DSM. Inhibition of PDE1 can be a novel therapeutic approach for the treatment of overactive bladder associated with detrusor overactivity.

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Large-conductance voltage- and Ca2+-activated K+ channels regulate human detrusor smooth muscle function

AJP Cell Physiology ◽

10.1152/ajpcell.00495.2010 ◽

2011 ◽

Vol 301 (4) ◽

pp. C903-C912 ◽

Cited By ~ 72

Author(s):

Kiril L. Hristov ◽

Muyan Chen ◽

Whitney F. Kellett ◽

Eric S. Rovner ◽

Georgi V. Petkov

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Specific Inhibitor ◽

Bk Channels ◽

Bk Channel ◽

Bladder Function ◽

Detrusor Smooth Muscle ◽

Patch Clamp Technique ◽

Whole Cell ◽

Human Detrusor

The large-conductance voltage- and Ca2+-activated K+ (BK) channel is expressed in many smooth muscle types, but its role in human detrusor smooth muscle (DSM) is unclear. With a multidisciplinary approach spanning channel molecules, single-channel activity, freshly isolated human DSM cells, intact DSM preparations, and the BK channel specific inhibitor iberiotoxin, we elucidated human DSM BK channel function and regulation. Native human DSM tissues were obtained during open surgeries from patients with no preoperative history of overactive bladder. RT-PCR experiments on single human DSM cells showed mRNA expression of BK channel α-, β1-, and β4-subunits. Western blot and immunocytochemistry confirmed BK channel α, β1, and β4 protein expression. Native human BK channel properties were described using the perforated whole cell configuration of the patch-clamp technique. In freshly isolated human DSM cells, BK channel blockade with iberiotoxin inhibited a significant portion of the total voltage step-induced whole cell K+ current. From single BK channel recordings, human BK channel conductance was calculated to be 136 pS. Voltage-dependent iberiotoxin- and ryanodine-sensitive transient BK currents were identified in human DSM cells. In current-clamp mode, iberiotoxin inhibited the hyperpolarizing membrane potential transients and depolarized the cell resting membrane potential. Isometric DSM tension recordings revealed that BK channels principally control the contractions of isolated human DSM strips. Collectively, our results indicate that BK channels are fundamental regulators of DSM excitability and contractility and may represent new targets for pharmacological or genetic control of urinary bladder function in humans.

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Modulation of aortic smooth muscle cell membrane potential by extracellular calcium.

Hypertension ◽

10.1161/01.hyp.14.5.549 ◽

1989 ◽

Vol 14 (5) ◽

pp. 549-555 ◽

Cited By ~ 7

Author(s):

C E Palant ◽

N Stern ◽

A Meyer ◽

M L Tuck ◽

D B Lee ◽

...

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Cell Membrane ◽

Smooth Muscle Cell ◽

Muscle Cell ◽

Extracellular Calcium ◽

Cell Membrane Potential ◽

Aortic Smooth Muscle Cell ◽

Aortic Smooth Muscle ◽

Muscle Cell Membrane

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Pressure-induced smooth muscle cell depolarization in pulmonary arteries from control and chronically hypoxic rats does not cause myogenic vasoconstriction

Journal of Applied Physiology ◽

10.1152/japplphysiol.00819.2004 ◽

2005 ◽

Vol 98 (3) ◽

pp. 1119-1124 ◽

Cited By ~ 23

Author(s):

Jay S. Naik ◽

Scott Earley ◽

Thomas C. Resta ◽

Benjimen R. Walker

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Cell Membrane ◽

Pulmonary Vascular Resistance ◽

Vascular Resistance ◽

Pulmonary Arteries ◽

Barometric Pressure ◽

Cell Membrane Potential ◽

Intraluminal Pressure ◽

Dose Dependent

Chronic obstructive pulmonary diseases, as well as prolonged residence at high altitude, can result in generalized airway hypoxia, eliciting an increase in pulmonary vascular resistance. We hypothesized that a portion of the elevated pulmonary vascular resistance following chronic hypoxia (CH) is due to the development of myogenic tone. Isolated, pressurized small pulmonary arteries from control (barometric pressure ≅ 630 Torr) and CH (4 wk, barometric pressure = 380 Torr) rats were loaded with fura 2-AM and perfused with warm (37°C), aerated (21% O2-6% CO2-balance N2) physiological saline solution. Vascular smooth muscle (VSM) intracellular Ca2+ concentration ([Ca2+]i) and diameter responses to increasing intraluminal pressure were determined. Diameter and VSM cell [Ca2+]i responses to KCl were also determined. In a separate set of experiments, VSM cell membrane potential responses to increasing luminal pressure were determined in arteries from control and CH rats. VSM cell membrane potential in arteries from CH animals was depolarized relative to control at each pressure step. VSM cells from both groups exhibited a further depolarization in response to step increases in intraluminal pressure. However, arteries from both control and CH rats distended passively to increasing intraluminal pressure, and VSM cell [Ca2+]i was not affected. KCl elicited a dose-dependent vasoconstriction that was nearly identical between control and CH groups. Whereas KCl administration resulted in a dose-dependent increase in VSM cell [Ca2+]i in arteries taken from control animals, this stimulus elicited only a slight increase in VSM cell [Ca2+]i in arteries from CH animals. We conclude that the pulmonary circulation of the rat does not demonstrate pressure-induced vasoconstriction.

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Cytosolic Calcium Oscillations in Smooth Muscle Cells

Physiology ◽

10.1152/physiologyonline.2000.15.1.50 ◽

2000 ◽

Vol 15 (1) ◽

pp. 50-55 ◽

Cited By ~ 4

Author(s):

Jean-Pierre Savineau ◽

Roger Marthan

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Cell Membrane ◽

Smooth Muscle Cells ◽

Inositol Trisphosphate ◽

Muscle Cells ◽

Cytosolic Calcium ◽

Calcium Oscillations ◽

Membrane Receptors ◽

Cell Membrane Potential

In a variety of smooth muscle cells, agonists activating membrane receptors induce oscillations in the cytoplasmic Ca2+ concentration via an inositol trisphosphate-activated mechanism. Ca2+ oscillations participate in the control of cell membrane potential and the tone of smooth muscle. There is evidence that alterations in Ca2+ oscillations modulate smooth muscle responsiveness.

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A synthetic transmembrane segment derived from TRPV4 channel self-assembles into potassium-like channels to regulate vascular smooth muscle cell membrane potential

Journal of Materials Chemistry B ◽

10.1039/c3tb21645d ◽

2014 ◽

Vol 2 (24) ◽

pp. 3809-3818 ◽

Cited By ~ 14

Author(s):

Zhiqiang Yu ◽

Jie Li ◽

Jinhang Zhu ◽

Min Zhu ◽

Feifei Jiang ◽

...

Keyword(s):

Smooth Muscle ◽

Membrane Potential ◽

Cell Membrane ◽

Vascular Smooth Muscle ◽

Smooth Muscle Cell ◽

Muscle Cell ◽

Vascular Smooth Muscle Cell ◽

Cell Membrane Potential ◽

Trpv4 Channel ◽

Muscle Cell Membrane

A synthetic K+-like channel mediates K+outward flow to regulate vascular smooth muscle cell membrane potential, blood vessel tone and blood pressure.

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Oscillatory Membrane Potential Response to Glucose in Islet β-Cells: A Comparison of Islet-Cell Electrical Activity in Mouse and Rat

Endocrinology ◽

10.1210/en.2006-0424 ◽

2006 ◽

Vol 147 (10) ◽

pp. 4655-4663 ◽

Cited By ~ 48

Author(s):

Jocelyn E. Manning Fox ◽

Armen V. Gyulkhandanyan ◽

Leslie S. Satin ◽

Michael B. Wheeler

Keyword(s):

Insulin Secretion ◽

Membrane Potential ◽

Cell Membrane ◽

Electrical Activity ◽

Cell Membrane Potential ◽

Dependent Manner ◽

Β Cells ◽

Action Potential Firing ◽

Patch Clamp Technique ◽

Elevated Glucose

In contrast to mouse, rat islet β-cell membrane potential is reported not to oscillate in response to elevated glucose despite demonstrated oscillations in calcium and insulin secretion. We aim to clarify the electrical activity of rat islet β-cells and characterize and compare the electrical activity of both α- and β-cells in rat and mouse islets. We recorded electrical activity from α- and β-cells within intact islets from both mouse and rat using the perforated whole-cell patch clamp technique. Fifty-six percent of both mouse and rat β-cells exhibited an oscillatory response to 11.1 mm glucose. Responses to both 11.1 mm and 2.8 mm glucose were identical in the two species. Rat β-cells exhibited incremental depolarization in a glucose concentration-dependent manner. We also demonstrated electrical activity in human islets recorded under the same conditions. In both mouse and rat α-cells 11 mm glucose caused hyperpolarization of the membrane potential, whereas 2.8 mm glucose produced action potential firing. No species differences were observed in the response of α-cells to glucose. This paper is the first to demonstrate and characterize oscillatory membrane potential fluctuations in the presence of elevated glucose in rat islet β-cells in comparison with mouse. The findings promote the use of rat islets in future electrophysiological studies, enabling consistency between electrophysiological and insulin secretion studies. An inverse response of α-cell membrane potential to glucose furthers our understanding of the mechanisms underlying glucose sensitive glucagon secretion.

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Suppression of human detrusor smooth muscle excitability and contractility via pharmacological activation of large conductance Ca2+-activated K+channels

AJP Cell Physiology ◽

10.1152/ajpcell.00417.2011 ◽

2012 ◽

Vol 302 (11) ◽

pp. C1632-C1641 ◽

Cited By ~ 41

Author(s):

Kiril L. Hristov ◽

Shankar P. Parajuli ◽

Rupal P. Soder ◽

Qiuping Cheng ◽

Eric S. Rovner ◽

...

Keyword(s):

Smooth Muscle ◽

Electrical Field Stimulation ◽

Bk Channels ◽

Bk Channel ◽

Resting Membrane Potential ◽

Force Frequency ◽

Detrusor Smooth Muscle ◽

Patch Clamp Technique ◽

Whole Cell ◽

Current Clamp Mode

Overactive bladder syndrome is frequently associated with increased detrusor smooth muscle (DSM) contractility. We tested the hypothesis that pharmacological activation of the large-conductance voltage- and Ca2+-activated K+(BK) channel with NS-1619, a selective BK channel opener, reduces the excitability and contractility of human DSM. We used the amphotericin-perforated whole cell patch-clamp technique on freshly isolated human DSM cells, live-cell Ca2+imaging, and isometric DSM tension recordings of human DSM strips obtained from open bladder surgeries. NS-1619 (30 μM) significantly increased the amplitude of the voltage step-induced whole cell BK currents, and this effect was abolished by pretreatment with 200 nM iberiotoxin (IBTX), a selective BK channel inhibitor. In current-clamp mode, NS-1619 (30 μM) significantly hyperpolarized the resting membrane potential, and the hyperpolarization was reversed by IBTX (200 nM). NS-1619 (30 μM) significantly decreased the intracellular Ca2+level in isolated human DSM cells. BK channel activation with NS-1619 (30 μM) significantly inhibited the amplitude, muscle force, frequency, duration, and tone of the spontaneous phasic and pharmacologically induced DSM contractions from human DSM isolated strips. IBTX (200 nM) suppressed the inhibitory effects of NS-1619 on spontaneous contractions. The amplitude of electrical field stimulation (0.5–50 Hz)-induced contractions was significantly reduced by NS-1619 (30 μM). Our data suggest that pharmacological activation of BK channels could represent a novel treatment option to control bladder dysfunction in humans.

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A novel approach allows identification of K channels in the lateral membrane of rat CCD

AJP Renal Physiology ◽

10.1152/ajprenal.1994.266.5.f813 ◽

1994 ◽

Vol 266 (5) ◽

pp. F813-F822 ◽

Cited By ~ 16

Author(s):

W. H. Wang ◽

C. M. McNicholas ◽

A. S. Segal ◽

G. Giebisch

Keyword(s):

Membrane Potential ◽

Cell Membrane ◽

Cell Membrane Potential ◽

K Channel ◽

Dependent Manner ◽

Patch Clamp Technique ◽

K Channels ◽

Lateral Membrane ◽

Novel Approach ◽

Voltage Dependent

We have developed a novel approach to study K channels in the lateral membrane of principal cells (PC) in rat cortical collecting ducts (CCD). The technique consists of 1) exposing the CCD apical membrane, 2) removing the intercalated cells adjoining a PC by gentle suction through a pipette, and 3) applying patch-clamp technique to the lateral membrane of PC. Functional viability of the PC was confirmed by three indexes: 1) maintenance of physiological cell membrane potentials (-85 +/- 3 mV); 2) depolarization of the cell membrane potential with 1 mM Ba2+; and 3) hyperpolarization of the cell potential with 0.1 mM amiloride. Two types of K channels were identified: a low-conductance K channel and an intermediate-conductance K channel. In cell-attached patches the slope conductance of the low-conductance K channel was 27 pS and that of the intermediate-conductance K channel was 45 pS. The open probability (Po) of the 27-pS K channel was 0.81 +/- 0.02 and was not voltage dependent. In contrast, the Po of the 45-pS K channel was 0.23 +/- 0.01 at the spontaneous cell membrane potential and was increased by hyperpolarization. In addition, decrease of the bath pH from 7.4 to 6.7 reduced the 27-pS K channel current amplitude in a voltage-dependent manner, but the Po was not affected. Finally, two time constants were required to fit open- and closed-time histograms of both populations of K channels. Application of 1 mM Ba2+ completely blocked these K channels. We conclude that two types of K channel are present in the basolateral membrane of PC.

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Inhibition of phosphodiesterases relaxes detrusor smooth muscle via activation of the large-conductance voltage- and Ca2+-activated K+ channel

AJP Cell Physiology ◽

10.1152/ajpcell.00432.2011 ◽

2012 ◽

Vol 302 (9) ◽

pp. C1361-C1370 ◽

Cited By ~ 20

Author(s):

Wenkuan Xin ◽

Qiuping Cheng ◽

Rupal P. Soder ◽

Georgi V. Petkov

Keyword(s):

Smooth Muscle ◽

Guinea Pig ◽

Bk Channels ◽

Bk Channel ◽

Dependent Manner ◽

Detrusor Smooth Muscle ◽

Channel Activity ◽

Patch Clamp Technique ◽

Membrane Hyperpolarization ◽

Pde Inhibition

Detrusor smooth muscle (DSM) exhibits increased spontaneous phasic contractions under pathophysiological conditions such as detrusor overactivity (DO). Our previous studies showed that activation of cAMP signaling pathways reduces DSM contractility by increasing the large-conductance voltage- and Ca2+-activated K+ (BK) channel activity. Here, we tested the hypothesis whether inhibition of phosphodiesterases (PDEs) can reduce guinea pig DSM excitability and contractility by increasing BK channel activity. Utilizing isometric tension recordings of DSM isolated strips and the perforated patch-clamp technique on freshly isolated DSM cells, we examined the mechanism of DSM relaxation induced by PDE inhibition. Inhibition of PDEs by 3-isobutyl-1-methylxanthine (IBMX), a nonselective PDE inhibitor, significantly reduced DSM spontaneous and carbachol-induced contraction amplitude, frequency, duration, muscle force integral, and tone in a concentration-dependent manner. IBMX significantly reduced electrical field stimulation-induced contractions of DSM strips. Blocking BK channels with paxilline diminished the inhibitory effects of IBMX on DSM contractility, indicating a role for BK channels in DSM relaxation mediated by PDE inhibition. IBMX increased the transient BK currents (TBKCs) frequency by ∼3-fold without affecting the TBKCs amplitude. IBMX increased the frequency of the spontaneous transient hyperpolarizations by ∼2-fold and hyperpolarized the DSM cell resting membrane potential by ∼6 mV. Blocking the BK channels with paxilline abolished the IBMX hyperpolarizing effects. Under conditions of blocked Ca2+ sources for BK channel activation, IBMX did not affect the depolarization-induced steady-state whole cell BK currents. Our data reveal that PDE inhibition with IBMX relaxes guinea pig DSM via TBKCs activation and subsequent DSM cell membrane hyperpolarization.

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