title: Membrane coordination of receptors and channels mediating the inhibition of neuronal ion currents by ADP.
abstract: ADP and other nucleotides control ion currents in the nervous system via various P2Y receptors. In this respect, Cav2 and Kv7 channels have been investigated most frequently. The fine tuning of neuronal ion channel gating via G protein coupled receptors frequently relies on the formation of higher order protein complexes that are organized by scaffolding proteins and harbor receptors and channels together with interposed signaling components. However, ion channel complexes containing P2Y receptors have not been described. Therefore, the regulation of Cav2.2 and Kv7.2/7.3 channels via P2Y1 and P2Y12 receptors and the coordination of these ion channels and receptors in the plasma membranes of tsA 201 cells have been investigated here. ADP inhibited currents through Cav2.2 channels via both P2Y1 and P2Y12 receptors with phospholipase C and pertussis toxin-sensitive G proteins being involved, respectively. The nucleotide controlled the gating of Kv7 channels only via P2Y1 and phospholipase C. In fluorescence energy transfer assays using conventional as well as total internal reflection (TIRF) microscopy, both P2Y1 and P2Y12 receptors were found juxtaposed to Cav2.2 channels, but only P2Y1, and not P2Y12, was in close proximity to Kv7 channels. Using fluorescence recovery after photobleaching in TIRF microscopy, evidence for a physical interaction was obtained for the pair P2Y12/Cav2.2, but not for any other receptor/channel combination. These results reveal a membrane juxtaposition of P2Y receptors and ion channels in parallel with the control of neuronal ion currents by ADP. This juxtaposition may even result in apparent physical interactions between receptors and channels.
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria.
authors.forename: Hend
authors.initials: H
authors.lastname: Gafar
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria.
authors.forename: Manuel
authors.initials: M
authors.lastname: Dominguez Rodriguez
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria.
authors.forename: Giri K
authors.initials: GK
authors.lastname: Chandaka
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria.
authors.forename: Isabella
authors.initials: I
authors.lastname: Salzer
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria. stefan.boehm@meduniwien.ac.at.
authors.forename: Stefan
authors.initials: S
authors.lastname: Boehm
authors.affiliation: Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, Schwarzspanierstrasse 17/I, A-1090, Vienna, Austria.
authors.forename: Klaus
authors.initials: K
authors.lastname: Schicker
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism*; D020864:Calcium Channels, N-Type / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D007473:Ion Channels / Q000378:metabolism*; D051658:KCNQ2 Potassium Channel / Q000378:metabolism; D051659:KCNQ3 Potassium Channel / Q000378:metabolism; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009474:Neurons / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D058472:Receptors, Purinergic P2Y1 / Q000378:metabolism*; D058925:Receptors, Purinergic P2Y12 / Q000378:metabolism*
chemical_list: C500545:CACNA1B protein, human; D020864:Calcium Channels, N-Type; D007473:Ion Channels; D051658:KCNQ2 Potassium Channel; D051659:KCNQ3 Potassium Channel; C548576:P2RY12 protein, human; D058472:Receptors, Purinergic P2Y1; D058925:Receptors, Purinergic P2Y12; D000244:Adenosine Diphosphate
keywords: Calcium channel; FRET; Kv7 channel; P2Y receptors; TIRF