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

Abstract

<jats:p>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 Ca<jats:sup>2+</jats:sup>-activated K<jats:sup>+</jats:sup>(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 Ca<jats:sup>2+</jats:sup>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 Ca<jats:sup>2+</jats:sup>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.</jats:p>

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

“…For example, the BK channel blockade with either iberiotoxin or charybdotoxin prolonged the action potentials correlating with enhanced contractility (16,18), whereas the BK channel opener NS11021 reduced action potential generation and DSM contractility in guinea pigs (29). In human DSM, the selective BK channel inhibitor iberiotoxin and activator NS1619 increased and decreased, respectively, the contractility of tissue strips and excitability of freshly isolated DSM cells confirming the important regulatory role of this K ϩ channel subtype (22,25). Key characteristics of the BK channels are 1) their large conductance whereby the opening or closure of few channels has pronounced effects on cellular excitability; 2) a dual sensitivity to metabolic factors (e.g., Ca 2ϩ and cAMP-pathways) and membrane voltage; and 3) a close functional interrelationship with VDCCs, which are the primary regulators of contractility mediating the influx of Ca 2ϩ to initiate DSM contractions (3,24,44,46,54).…”

“…Among them, the large-conductance voltage-and Ca 2ϩ -activated K ϩ channels (also known as BK, Slo, or K Ca 1.1 channels) and L-type voltage-dependent Ca 2ϩ channels (VDCCs) are recognized as key regulators of excitability and contractility of DSM (44). Supporting data, provided for DSM studies using rat, guinea pig, mouse, and importantly human tissues and cells (5,17,18,22,24,25,46), demonstrate the role of BK channels in the regulation of the resting membrane potential, modulation of the repolarization phase of DSM action potentials, and generation of spontaneous transient BK currents (TBKCs), also known as spontaneous transient outward currents. For example, the BK channel blockade with either iberiotoxin or charybdotoxin prolonged the action potentials correlating with enhanced contractility (16, 18), whereas the BK channel opener NS11021 reduced action potential generation and DSM contractility in guinea pigs (29).…”

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

“…1). The activation of BK channels hyperpolarizes the human DSM cell membrane potential (13,28). The pharmacological inhibition of constitutive PKA activity abolishes TBKCs and depolarizes DSM cell membrane potential (27) (Figs.…”

“…The amphotericin-B perforated whole cell patch-clamp technique was used for electrophysiological recordings in freshly isolated DSM single cells as previously described (11)(12)(13)17). Patch-clamp recordings were conducted using an Axopatch 200B amplifier system and Digidata 1440A controlled with pCLAMP 10.2 software (Molecular Devices, Union City, CA).…”

“…Isometric contractions of human DSM isolated strips were measured using a Myomed myograph system (MED Associates, St. Albans, VT) as previously described (11)(12)(13)17). Briefly, DSM strips were secured to isometric force displacement transducers and were suspended in temperature-controlled (37°C) water-jacketed tissue baths containing 10 ml physiological saline solution, which was prepared daily and contained (in mM) 119 NaCl, 4.7 KCl, 24 NaHCO 3, 1.2 KH2PO4, 2.5 CaCl2, 1.2 MgSO4, and 11 D-glucose, and was aerated with 95% O2-5% CO2 to obtain pH 7.4.…”