AB207. Constitutive PKA activity is essential for maintaining the excitability and contractility in guinea pig urinary bladder smooth muscle: central role played by the BK channel

Abstract: The large conductance voltage- and Ca-activated K(BK) channel is a critical regulator of urinary bladder smooth muscle (UBSM) excitability and contractility in many species. BK channel activation can hyperpolarize the cell resting membrane potential (RMP), decrease the intracellular Ca level, and attenuate UBSM cell excitability, therefore cause UBSM relaxation. Elevation of cellular cAMP levels by β-adrenergic receptor activation or phosphodiesterase (PDE) inhibition suppresses guinea pig UBSM excitability and contractility via BK channel activation. The current study aimed at determining the mechanism by which basal PKA, without elevation of cellular cAMP levels, controls UBSM excitability, and contractility via BK channel activation. UBSM strips (—2-4 mm wide and —5-8 mm long) were harvested from guinea pig bladder following removing urothelium. We used perforated patch-clamp and line-scanning confocal techniques on freshly-isolated guinea pig UBSM cells, and isometric tension recordings of UBSM isolated strips. Our data show that PKA inhibitors, H-89, PKI 14-22, or KT-5720: (I) reduced Ca spark frequency, eliminated spontaneous transient BK channel currents, abolished the transient hyperpolarizations, and depolarized the cell RMP; (II) increased the spontaneous phasic contraction amplitude, force, duration, and decreased the phasic contraction frequency; (III) in the presence of PKA inhibitor, H-89, PDE1 inhibition with 8MM-IBMX did not affect BK channel activity or DSM contractility. Using a selective PKG inhibitor, DT-2, we provided direct evidence at cellular level that basal PKG does not play a role in the regulation of BK channel activity in UBSM cells. In conclusion, basal PKA plays an essential role in maintaining BK channel activity, and thereby controlling UBSM excitability and contractility.

Keywords: Detrusor; PKA; H-89; KT-5720; PKI 14-22