The Novel K_V7.2/K_V7.3 Channel Opener ICA-069673 Reveals Subtype-Specific Functional Roles in Guinea Pig Detrusor Smooth Muscle Excitability and Contractility

Abstract

The physiologic roles of voltage-gated K V 7 channel subtypes (K V 7.1-K V 7.5) in detrusor smooth muscle (DSM) are poorly understood. Here, we sought to elucidate the functional roles of K V 7.2/K V 7.3 channels in guinea pig DSM excitability and contractility using the novelWe employed a multilevel experimental approach using Western blot analysis, immunocytochemistry, isometric DSM tension recordings, fluorescence Ca 21 imaging, and perforated whole-cell patch-clamp electrophysiology. Western blot experiments revealed the protein expression of K V 7.2 and K V 7.3 channel subunits in DSM tissue. In isolated DSM cells, immunocytochemistry with confocal microscopy further confirmed protein expression for K V 7.2 and K V 7.3 channel subunits, where they localize within the vicinity of the cell membrane. ICA-069673 inhibited spontaneous phasic, pharmacologically induced, and nerve-evoked contractions in DSM isolated strips in a concentration-dependent manner. The inhibitory effects of ICA-069673 on DSM spontaneous phasic and tonic contractions were abolished in the presence of the K V 7 channel inhibitor XE991 [10,10-bis(4-pyridinylmethyl)-9 (10H)-anthracenone dihydrochloride]. Under conditions of elevated extracellular K 1 (60 mM), the effects of ICA-069673 on DSM tonic contractions were significantly attenuated. ICA-069673 decreased the global intracellular Ca 21 concentration in DSM cells, an effect blocked by the L-type Ca 21 channel inhibitor nifedipine. ICA-069673 hyperpolarized the membrane potential and inhibited spontaneous action potentials of isolated DSM cells, effects that were blocked in the presence of XE991. In conclusion, using the novel K V 7.2/ K V 7.3 channel activator ICA-069673, this study provides strong evidence for a critical role for the K V 7.2-and K V 7.3-containing channels in DSM function at both cellular and tissue levels.

The KV7 channel activator retigabine suppresses mouse urinary bladder afferent nerve activity without affecting detrusor smooth muscle K+ channel currents

“…; Provence et al . ), supporting the possibility of species‐dependent differences. However, no study has described Kv7 agonist‐ or antagonist‐sensitive currents in smooth muscle which have the properties (e.g.…”

“…pig and rat, whereas primarily K V 7.2 and K V 7.3 are reported in guinea pig UBSM (Svalø et al 2012;Anderson et al 2013;Provence et al 2015Provence et al , 2018. Expressed K V 7.2 and K V 7.3 channels exhibit a half-activation voltage of about −40 mV (Table 1), which is roughly the normal resting potential of UBSM as measured with microelectrodes in intact tissue (Heppner et al 1997;Takagi & Hashitani, 2016).…”

“…; Provence et al . ). However, to date, the site of action of these K V 7 modulators has been incompletely evaluated, leaving the door open to alternative interpretations of their effects on urinary bladder function.…”

“…XE-991 alone had no effect on TC amplitude, TC slope or afferent activity, but did cause a significant increase in TC frequency that was reversed by retigabine. as direct effects of K V 7 modulators on UBSM function (Rode et al 2010;Afeli et al 2013;Anderson et al 2013;Provence et al 2015). However, to date, the site of action of these K V 7 modulators has been incompletely evaluated, leaving the door open to alternative interpretations of their effects on urinary bladder function.…”

“…; Provence et al . , ). Expressed K V 7.2 and K V 7.3 channels exhibit a half‐activation voltage of about −40 mV (Table ), which is roughly the normal resting potential of UBSM as measured with microelectrodes in intact tissue (Heppner et al .…”