title: Local coupling of TRPC6 to ANO1/TMEM16A channels in smooth muscle cells amplifies vasoconstriction in cerebral arteries.
abstract: Anoctamin-1 [ANO1, also known as transmembrane protein 16A (TMEM16A)] is a Ca(2+)-activated Cl(-) channel expressed in arterial myocytes that regulates membrane potential and contractility. Signaling mechanisms that control ANO1 activity in arterial myocytes are poorly understood. In cerebral artery myocytes, ANO1 channels are activated by local Ca(2+) signals generated by plasma membrane nonselective cation channels, but the molecular identity of these proteins is unclear. Arterial myocytes express several different nonselective cation channels, including multiple members of the transient receptor potential receptor (TRP) family. The goal of this study was to identify localized ion channels that control ANO1 currents in cerebral artery myocytes. Coimmunoprecipitation and immunofluorescence resonance energy transfer microscopy experiments indicate that ANO1 and canonical TRP 6 (TRPC6) channels are present in the same macromolecular complex and localize in close spatial proximity in the myocyte plasma membrane. In contrast, ANO1 is not near TRPC3, TRP melastatin 4, or inositol trisphosphate receptor 1 channels. Hyp9, a selective TRPC6 channel activator, stimulated Cl(-) currents in myocytes that were blocked by T16Ainh-A01, an ANO1 inhibitor, ANO1 knockdown using siRNA, and equimolar replacement of intracellular EGTA with BAPTA, a fast Ca(2+) chelator that abolishes local Ca(2+) signaling. Hyp9 constricted pressurized cerebral arteries, and this response was attenuated by T16Ainh-A01. In contrast, T16Ainh-A01 did not alter depolarization-induced (60 mM K(+)) vasoconstriction. These data indicate that TRPC6 channels generate a local intracellular Ca(2+) signal that activates nearby ANO1 channels in myocytes to stimulate vasoconstriction.
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Qian
authors.initials: Q
authors.lastname: Wang
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: M Dennis
authors.initials: MD
authors.lastname: Leo
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Damodaran
authors.initials: D
authors.lastname: Narayanan
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee.
authors.forename: Korah P
authors.initials: KP
authors.lastname: Kuruvilla
authors.affiliation: Department of Physiology, University of Tennessee Health Science Center, Memphis, Tennessee jjaggar@uthsc.edu.
authors.forename: Jonathan H
authors.initials: JH
authors.lastname: Jaggar
mesh_terms: D000818:Animals; D000075369:Anoctamin-1; D065096:Calcium Chelating Agents / Q000494:pharmacology; D020013:Calcium Signaling; D002536:Cerebral Arteries / Q000378:metabolism; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism*; D034622:RNA Interference; D017207:Rats, Sprague-Dawley; D050052:TRPC Cation Channels / Q000819:agonists / Q000378:metabolism*; D046509:Tissue Culture Techniques; D014162:Transfection; D014661:Vasoconstriction / Q000187:drug effects; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: C552942:ANO1 protein, rat; D000075369:Anoctamin-1; D065096:Calcium Chelating Agents; D018118:Chloride Channels; D050052:TRPC Cation Channels; C492145:Trpc6 protein, rat; D014662:Vasoconstrictor Agents
keywords: anoctamin-1 channel; arterial smooth muscle cell; transient receptor potential channel; vasoconstriction