title: Steady-state modulation of voltage-gated K+ channels in rat arterial smooth muscle by cyclic AMP-dependent protein kinase and protein phosphatase 2B.
abstract: Voltage-gated potassium channels (Kv) are important regulators of membrane potential in vascular smooth muscle cells, which is integral to controlling intracellular Ca2+ concentration and regulating vascular tone. Previous work indicates that Kv channels can be modulated by receptor-driven alterations of cyclic AMP-dependent protein kinase (PKA) activity. Here, we demonstrate that Kv channel activity is maintained by tonic activity of PKA. Whole-cell recording was used to assess the effect of manipulating PKA signalling on Kv and ATP-dependent K+ channels of rat mesenteric artery smooth muscle cells. Application of PKA inhibitors, KT5720 or H89, caused a significant inhibition of Kv currents. Tonic PKA-mediated activation of Kv appears maximal as application of isoprenaline (a β-adrenoceptor agonist) or dibutyryl-cAMP failed to enhance Kv currents. We also show that this modulation of Kv by PKA can be reversed by protein phosphatase 2B/calcineurin (PP2B). PKA-dependent inhibition of Kv by KT5720 can be abrogated by pre-treatment with the PP2B inhibitor cyclosporin A, or inclusion of a PP2B auto-inhibitory peptide in the pipette solution. Finally, we demonstrate that tonic PKA-mediated modulation of Kv requires intact caveolae. Pre-treatment of the cells with methyl-β-cyclodextrin to deplete cellular cholesterol, or adding caveolin-scaffolding domain peptide to the pipette solution to disrupt caveolae-dependent signalling each attenuated PKA-mediated modulation of the Kv current. These findings highlight a novel, caveolae-dependent, tonic modulatory role of PKA on Kv channels providing new insight into mechanisms and the potential for pharmacological manipulation of vascular tone.
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom.
authors: Jennifer L
authors: JL
authors: Brignell
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom.
authors: Matthew D
authors: MD
authors: Perry
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom.
authors: Carl P
authors: CP
authors: Nelson
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom; Department of Cancer Studies and Molecular Medicine, Leicester Royal Infirmary, Leicester, United Kingdom.
authors: Jonathon M
authors: JM
authors: Willets
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom.
authors: R A John
authors: RA
authors: Challiss
authors: Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, United Kingdom.
authors: Noel W
authors: NW
authors: Davies
mesh_terms: D054758:A Kinase Anchor Proteins / Q000378:metabolism; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D019703:Calcineurin / Q000378:metabolism*; D002227:Carbazoles / Q000494:pharmacology; D021941:Caveolae / Q000187:drug effects / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D016048:Dideoxyadenosine / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects; D007545:Isoproterenol / Q000494:pharmacology; D008297:Male; D008638:Mesenteric Arteries / Q000166:cytology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism*; D024642:Potassium Channels, Voltage-Gated / Q000378:metabolism*; D011758:Pyrroles / Q000494:pharmacology; D017208:Rats, Wistar
chemical_list: D054758:A Kinase Anchor Proteins; D002227:Carbazoles; D024642:Potassium Channels, Voltage-Gated; D011758:Pyrroles; D016048:Dideoxyadenosine; C057416:KT 5720; D017868:Cyclic AMP-Dependent Protein Kinases; D019703:Calcineurin; D000262:Adenylyl Cyclases; D007545:Isoproterenol