title: Constitutive PKA activity is essential for maintaining the excitability and contractility in guinea pig urinary bladder smooth muscle: role of the BK channel.
abstract: The elevation of protein kinase A (PKA) activity activates the large-conductance voltage- and Ca(2+)-activated K(+) (BK) channels in urinary bladder smooth muscle (UBSM) cells and consequently attenuates spontaneous phasic contractions of UBSM. However, the role of constitutive PKA activity in UBSM function has not been studied. Here, we tested the hypothesis that constitutive PKA activity is essential for controlling the excitability and contractility of UBSM. We used patch clamp electrophysiology, line-scanning confocal and ratiometric fluorescence microscopy on freshly isolated guinea pig UBSM cells, and isometric tension recordings on freshly isolated UBSM strips. Pharmacological inhibition of the constitutive PKA activity with H-89 or PKI 14-22 significantly reduced the frequency and amplitude of spontaneous transient BK channel currents (TBKCs) in UBSM cells. Confocal and ratiometric fluorescence microscopy studies revealed that inhibition of constitutive PKA activity with H-89 reduced the frequency and amplitude of the localized Ca(2+) sparks but increased global Ca(2+) levels and the magnitude of Ca(2+) oscillations in UBSM cells. H-89 abolished the spontaneous transient membrane hyperpolarizations and depolarized the membrane potential in UBSM cells. Inhibition of PKA with H-89 or KT-5720 also increased the amplitude and muscle force of UBSM spontaneous phasic contractions. This study reveals the novel concept that constitutive PKA activity is essential for controlling localized Ca(2+) signals generated by intracellular Ca(2+) stores and cytosolic Ca(2+) levels. Furthermore, constitutive PKA activity is critical for mediating the spontaneous TBKCs in UBSM cells, where it plays a key role in regulating spontaneous phasic contractions in UBSM.
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina; and.
authors.forename: Wenkuan
authors.initials: W
authors.lastname: Xin
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina; and Department of Urology, Fourth Hospital of China Medical University, Shenyang, China.
authors.forename: Ning
authors.initials: N
authors.lastname: Li
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina; and.
authors.forename: Qiuping
authors.initials: Q
authors.lastname: Cheng
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina; and.
authors.forename: Vitor S
authors.initials: VS
authors.lastname: Fernandes
authors.affiliation: Department of Drug Discovery and Biomedical Sciences, South Carolina College of Pharmacy, University of South Carolina, Columbia, South Carolina; and petkov@cop.sc.edu.
authors.forename: Georgi V
authors.initials: GV
authors.lastname: Petkov
mesh_terms: D000818:Animals; D020013:Calcium Signaling; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006168:Guinea Pigs; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009119:Muscle Contraction / Q000187:drug effects; D053580:Muscle Strength; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D013997:Time Factors; D001743:Urinary Bladder / Q000187:drug effects / Q000201:enzymology*
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D047428:Protein Kinase Inhibitors; D017868:Cyclic AMP-Dependent Protein Kinases
keywords: Ca2+ sparks; H-89; PKI 14–22; large-conductance voltage- and Ca2+-activated K+ channels; protein kinase A