A Positive Modulator of K<sub>Ca</sub>2 and K<sub>Ca</sub>3 Channels, 4,5-Dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), Inhibits Bladder Afferent Firing in Vitro and Bladder Overactivity in Vivo

Hougaard, Charlotte; Fraser, M. O.; Chien, Chin-Cheng; Bookout, Angie L.; Katofiasc, Mary; Jensen, Bo; Rode, Frederik; Bitsch-Nørhave, J.; Teuber, Lene; Thor, Karl B.; Strøbæk, Dorte; Burgard, Edward C.; Rønn, Lars Christian B.

doi:10.1124/jpet.108.143123

Cited by 34 publications

(52 citation statements)

References 27 publications

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“…It has previously been reported that Tl 1 influx can be monitored in FLIPR assays to detect activation of K 1 channels such as K v 7.2 and K Ca 2.3 (Weaver et al, 2004). In addition, our earlier studies demonstrated the capacity to detect activators of another Ca 21 -activated K 1 channel (hK Ca 3.1) in a similar assay (Hougaard et al, 2009;Jørgensen et al, 2013). The BK Tl 1 influx assay described in the current study appears to be highly suitable for detecting new smallmolecule BK activators.…”

Section: Discussionmentioning

confidence: 89%

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

Nausch

Rode

Jørgensen

et al. 2014

J Pharmacol Exp Ther

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Large-conductance Ca 21 -activated K 1 channels (BK, K Ca 1.1, MaxiK) are important regulators of urinary bladder function and may be an attractive therapeutic target in bladder disorders. In this study, we established a high-throughput fluorometric imaging plate reader-based screening assay for BK channel activators and identified a small-molecule positive modulator, NS19504 (5-[(4-bromophenyl)methyl]-1,3-thiazol-2-amine), which activated the BK channel with an EC 50 value of 11.0 6 1.4 mM. Hit validation was performed using high-throughput electrophysiology (QPatch), and further characterization was achieved in manual whole-cell and inside-out patch-clamp studies in human embryonic kidney 293 cells expressing hBK channels: NS19504 caused distinct activation from a concentration of 0.3 and 10 mM NS19504 left-shifted the voltage activation curve by 60 mV. Furthermore, whole-cell recording showed that NS19504 activated BK channels in native smooth muscle cells from guinea pig urinary bladder. In guinea pig urinary bladder strips, NS19504 (1 mM) reduced spontaneous phasic contractions, an effect that was significantly inhibited by the specific BK channel blocker iberiotoxin. In contrast, NS19504 (1 mM) only modestly inhibited nerve-evoked contractions and had no effect on contractions induced by a high K 1 concentration consistent with a K 1 channel-mediated action. Collectively, these results show that NS19504 is a positive modulator of BK channels and provide support for the role of BK channels in urinary bladder function. The pharmacologic profile of NS19504 indicates that this compound may have the potential to reduce nonvoiding contractions associated with spontaneous bladder overactivity while having a minimal effect on normal voiding.

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Section: Discussionmentioning

confidence: 89%

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

Nausch

Rode

Jørgensen

et al. 2014

J Pharmacol Exp Ther

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“…6,7-Dichloro-1H-indole-2,3-dione-3-oxime (NS309), a potent SK/IK channel activator (Strøbaek et al, 2004), increases bladder capacity and decreases voiding frequency in rats (Pandita et al, 2006), but its extremely short half-life precludes in vivo application. 4,5-Dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), another SK/IK channel activator, inhibits DSM carbachol-induced contractions in vitro and bladder overactivity in vivo (Hougaard et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Pharmacological Activation of Small Conductance Calcium-Activated Potassium Channels with Naphtho[1,2-d]thiazol-2-ylamine Decreases Guinea Pig Detrusor Smooth Muscle Excitability and Contractility

Parajuli

Soder

Hristov

et al. 2011

J Pharmacol Exp Ther

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Small conductance Ca 2ϩ -activated K ϩ (SK) and intermediate conductance Ca 2ϩ -activated K ϩ (IK) channels are thought to be involved in detrusor smooth muscle (DSM) excitability and contractility. Using naphtho[1,2-d]thiazol-2-ylamine (SKA-31), a novel and highly specific SK/IK channel activator, we investigated whether pharmacological activation of SK/IK channels reduced guinea pig DSM excitability and contractility. We detected the expression of all known isoforms of SK (SK1-SK3) and IK channels at mRNA and protein levels in DSM by singlecell reverse transcription-polymerase chain reaction and Western blot. Using the perforated patch-clamp technique on freshly isolated DSM cells, we observed that SKA-31 (10 M) increased SK currents, which were blocked by apamin (1 M), a selective SK channel inhibitor. In current-clamp mode, SKA-31 (10 M) hyperpolarized the cell resting membrane potential, which was blocked by apamin (1 M) but not by 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34) (1 M), a selective IK channel inhibitor. SKA-31 (10 nM-10 M) significantly inhibited the spontaneous phasic contraction amplitude, frequency, duration, and muscle force in DSM isolated strips. The SKA-31 inhibitory effects on DSM contractility were blocked by apamin (1 M) but not by TRAM-34 (1 M), which did not per se significantly affect DSM spontaneous contractility. SK channel activation with SKA-31 reduced contractions evoked by electrical field stimulation. SKA-31 effects were reversible upon washout. In conclusion, SK channels, but not IK channels, mediate SKA-31 effects in guinea pig DSM. Pharmacological activation of SK channels reduces DSM excitability and contractility and therefore may provide a novel therapeutic approach for controlling bladder dysfunction.

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“…Bladder ring preparations were obtained from six female Sprague-Dawley rats ( ∼ 200 g) [7]. Bladder samples were obtained from nine female pigs (6 months, 75-95 kg).…”

Section: Methodsmentioning

confidence: 99%

“…4,5-dichloro-1,3-diethyl-1,3-dihydrobenzoimidazol-2-one (NS4591) is a new positive modulator of SK/IK channels that has been shown to inhibit the number of action potentials in bladder primary afferent neurons generated by suprathreshold depolarizing pulses. NS4591 also reduced carbachol-induced twitches in rat bladder rings in an apamin (a specific inhibitor of SK channels) -sensitive manner and, in vivo , NS4591 inhibited detrusor overactivity induced by capsaicin and acetic acid in rats and cats, respectively [7].…”

Section: Introductionmentioning

confidence: 92%

“…Therefore, K + channels are potential targets for pharmacological treatment of detrusor overactivity, even though experiences with openers of K ATP and large-conductance Ca -activated K + channels have attracted interest as putative targets for treatment of detrusor overactivity. Activation of SK/IK channels has been shown to reduce detrusor contractility and alleviate detrusor overactivity in animal models [6,7]. 4,5-dichloro-1,3-diethyl-1,3-dihydrobenzoimidazol-2-one (NS4591) is a new positive modulator of SK/IK channels that has been shown to inhibit the number of action potentials in bladder primary afferent neurons generated by suprathreshold depolarizing pulses.…”

Section: Introductionmentioning

confidence: 99%

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Effect of the SK/IK channel modulator 4,5‐dichloro‐1,3‐diethyl‐1,3‐dihydro‐benzoimidazol‐2‐one (NS4591) on contractile force in rat, pig and human detrusor smooth muscle

Nielsen¹,

Rode²,

Rahbek³

et al. 2011

BJU International

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minimum force between oscillations by 30 ± 5% ( n = 9) in pig detrusor strips ( P < 0.005). In the presence of 10 μ M NS4591, carbachol (1 μ M) induced rhythmic contractions with an amplitude and normalized mean power frequency (nmeanPF) of 8.4 ± 5.1% and 0.11 ± 0.06 mN root mean square (rms) Hz ( n = 12), respectively, vs. 21 ± 3.4% and 0.17 ± 0.04 mN rms Hz in control strips ( n = 13). Apamin induced 6-and 11-fold increases in amplitude and nmeanPF vs. 1.3-and 2-fold increases in control strips.• In human detrusor strips ( n = 15), the cumulative addition of NS4591 (1-30 μ M) significantly reduced the amplitude by 69 ± 11%, the nmeanPF by 78 ± 6% and the minimum force between carbachol-induced oscillations by 59 ± 5% ( P < 0.008). The addition of apamin (0.3 μ M) before application of 1 μ M carbachol abolished the effects of NS4591 on amplitude and partially abolished its effect on nmeanPF by 41 ± 7%, vs. a 78 ± 6% reduction in the absence of apamin ( n = 8).• In spontaneously active detrusor preparations, NS4591 reduced or abolished contractions.• Furthermore, NS4591 (10 μ M) decreased the carbachol-induced increase in the fura-2 ratio by 43 ± 3% compared with control ( n = 12) ( P < 0.03). CONCLUSIONS• The SK/IK channel modulator NS4591 inhibits EFS-and carbachol-induced contractions in rat, pig and human detrusor muscle.• NS4591 may have therapeutic potential for treatment of detrusor overactivity. KEYWORDS detrusor, SK channels, NS4591, OABWhat's known on the subject? and What does the study add? Increased urinary bladder detrusor smooth muscle phasic contractility has been suggested to be associated with idiopathic bladder overactivity. Small conductance Ca2 + -activated K + (SK 1-3) channels have attracted considerable interest as putative target for new therapeutic strategy for treating overactive bladder. These channels play an important role in regulating the function and activity of urinary bladder smooth muscle (UBSM), and the loss of SK channel function has been shown to increase UBSM excitability and contractility. However, it is not known whether activation of SK channels has the converse effect of reducing UBSM excitability and contractility. In this paper, we investigated this possibility in the rat, pig and human UBSM by using the novel SK channel opener NS4591. These studies demonstrate that the SK channel modulator NS4591 has a potential role as a pharmacological tool to study the involvement of SK (and IK) channels in mammals and rodents urinary bladder. NS4591 may have therapeutic potential for treatment of detrusor overactivity. OBJECTIVE • To investigate the importance of small (SK)-and intermediate (IK)-conductance Ca2+ -activated K + channels on bladder function, by studying the effects of 4,5-dichloro-1,3-diethyl-1,3-dihydrobenzoimidazol-2-one (NS4591), a new modulator of SK/IK channels, on contractions induced by electrical field stimulation (EFS) and carbachol in rat, pig and human detrusor. PATIENTS AND METHODS• Detrusor biopsies were obtained from rats, pigs and male patients unde...

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A Positive Modulator of K_Ca2 and K_Ca3 Channels, 4,5-Dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), Inhibits Bladder Afferent Firing in Vitro and Bladder Overactivity in Vivo

Cited by 34 publications

References 27 publications

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

Pharmacological Activation of Small Conductance Calcium-Activated Potassium Channels with Naphtho[1,2-d]thiazol-2-ylamine Decreases Guinea Pig Detrusor Smooth Muscle Excitability and Contractility

Effect of the SK/IK channel modulator 4,5‐dichloro‐1,3‐diethyl‐1,3‐dihydro‐benzoimidazol‐2‐one (NS4591) on contractile force in rat, pig and human detrusor smooth muscle

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A Positive Modulator of KCa2 and KCa3 Channels, 4,5-Dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), Inhibits Bladder Afferent Firing in Vitro and Bladder Overactivity in Vivo

Cited by 34 publications

References 27 publications

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

NS19504: A Novel BK Channel Activator with Relaxing Effect on Bladder Smooth Muscle Spontaneous Phasic Contractions

Pharmacological Activation of Small Conductance Calcium-Activated Potassium Channels with Naphtho[1,2-d]thiazol-2-ylamine Decreases Guinea Pig Detrusor Smooth Muscle Excitability and Contractility

Effect of the SK/IK channel modulator 4,5‐dichloro‐1,3‐diethyl‐1,3‐dihydro‐benzoimidazol‐2‐one (NS4591) on contractile force in rat, pig and human detrusor smooth muscle

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A Positive Modulator of K_Ca2 and K_Ca3 Channels, 4,5-Dichloro-1,3-diethyl-1,3-dihydro-benzoimidazol-2-one (NS4591), Inhibits Bladder Afferent Firing in Vitro and Bladder Overactivity in Vivo