title: Orai1 and TRPC1 Proteins Co-localize with CaV1.2 Channels to Form a Signal Complex in Vascular Smooth Muscle Cells.
abstract: Voltage-dependent CaV1.2 L-type Ca2+channels (LTCC) are the main route for calcium entry in vascular smooth muscle cells (VSMC). Several studies have also determined the relevant role of store-operated Ca2+channels (SOCC) in vascular tone regulation. Nevertheless, the role of Orai1- and TRPC1-dependent SOCC in vascular tone regulation and their possible interaction with CaV1.2 are still unknown. The current study sought to characterize the co-activation of SOCC and LTCC upon stimulation by agonists, and to determine the possible crosstalk between Orai1, TRPC1, and CaV1.2. Aorta rings and isolated VSMC obtained from wild type or smooth muscle-selective conditional CaV1.2 knock-out (CaV1.2KO) mice were used to study vascular contractility, intracellular Ca2+mobilization, and distribution of ion channels. We found that serotonin (5-HT) or store depletion with thapsigargin (TG) enhanced intracellular free Ca2+concentration ([Ca2+]i) and stimulated aorta contraction. These responses were sensitive to LTCC and SOCC inhibitors. Also, 5-HT- and TG-induced responses were significantly attenuated in CaV1.2KOmice. Furthermore, hyperpolarization induced with cromakalim or valinomycin significantly reduced both 5-HT and TG responses, whereas these responses were enhanced with LTCC agonist Bay-K-8644. Interestingly, in situ proximity ligation assay revealed that CaV1.2 interacts with Orai1 and TRPC1 in untreated VSMC. These interactions enhanced significantly after stimulation of cells with 5-HT and TG. Therefore, these data indicate for the first time a functional interaction between Orai1, TRPC1, and CaV1.2 channels in VSMC, confirming that upon agonist stimulation, vessel contraction involves Ca2+entry due to co-activation of Orai1- and TRPC1-dependent SOCC and LTCC.
authors.affiliation: From the Departamento de Fisiología Médica y Biofísica and Groupo de Fisiopatología Cardiovascular, Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41013 Sevilla, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Ávila-Medina
authors.affiliation: Groupo de Fisiopatología Cardiovascular, Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41013 Sevilla, Spain.
authors.forename: Eva
authors.initials: E
authors.lastname: Calderón-Sánchez
authors.affiliation: From the Departamento de Fisiología Médica y Biofísica and.
authors.forename: Patricia
authors.initials: P
authors.lastname: González-Rodríguez
authors.affiliation: the Department of Neurophysiology and Neuropharmacology, Center for Physiology and Pharmacology, Medical University of Vienna, 1090 Wien, Austria, and.
authors.forename: Francisco
authors.initials: F
authors.lastname: Monje-Quiroga
authors.affiliation: the Departamento de Fisiología, Universidad de Extremadura, 10071 Cáceres, Spain.
authors.forename: Juan Antonio
authors.initials: JA
authors.lastname: Rosado
authors.affiliation: From the Departamento de Fisiología Médica y Biofísica and.
authors.forename: Antonio
authors.initials: A
authors.lastname: Castellano
authors.affiliation: Groupo de Fisiopatología Cardiovascular, Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41013 Sevilla, Spain.
authors.forename: Antonio
authors.initials: A
authors.lastname: Ordóñez
authors.affiliation: From the Departamento de Fisiología Médica y Biofísica and Groupo de Fisiopatología Cardiovascular, Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, 41013 Sevilla, Spain, tasmani@us.es.
authors.forename: Tarik
authors.initials: T
authors.lastname: Smani
mesh_terms: D000818:Animals; D001011:Aorta / Q000166:cytology / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D000071740:ORAI1 Protein / Q000235:genetics / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*; D014661:Vasoconstriction / Q000502:physiology
chemical_list: C514350:CACNA1C protein, mouse; D020746:Calcium Channels, L-Type; D046912:Multiprotein Complexes; D000071740:ORAI1 Protein; C520430:Orai1 protein, mouse; D050052:TRPC Cation Channels; C096351:transient receptor potential cation channel, subfamily C, member 1; D012701:Serotonin; D002118:Calcium
keywords: CaV1.2; CaV1.2 channel; Orai1; TRPC1; Vascular tone regulation; calcium; calcium release-activated calcium channel protein 1 (ORAI1); excitation-contraction coupling (E-C coupling); ion channel; vascular smooth muscle cells