title: Agonist-evoked calcium entry in vascular smooth muscle cells requires IP3 receptor-mediated activation of TRPC1.
abstract: Transient receptor potential canonical (TRPC) proteins have been proposed to function as plasma membrane Ca2+ channels activated by store depletion and/or by receptor stimulation. However, their role in the increase in cytosolic Ca2+ activated by contractile agonists in vascular smooth muscle is not yet elucidated. The present study was designed to investigate the functional and molecular properties of the Ca2+ entry pathway activated by endothelin-1 in primary cultured aortic smooth muscle cells. Measurement of the Ca2+ signal in fura-2-loaded cells allowed to characterize endothelin-1-evoked Ca2+ entry, which was resistant to dihydropyridine, and was blocked by 2-aminoethoxydiphenylborate (2-APB) and micromolar concentration of Gd3+. It was not activated by store depletion, but was inhibited by the endothelin ETA receptor antagonist BQ-123, and by heparin. On the opposite, thapsigargin-induced store depletion activated a Ca2+ entry pathway that was not affected by 2-APB, BQ-123 or heparin, and was less sensitive to Gd3+ than was endothelin-1-evoked Ca2+ entry. Investigation of the gene expression of TRPC isoforms by real-time RT-PCR revealed that TRPC1 was the most abundant. In cells transfected with TRPC1 small interfering RNA sequence, TRPC1 mRNA and protein expression were decreased by 72+/-3% and 86+/-2%, respectively, while TRPC6 expression was unaffected. In TRPC1 knockdown cells, both endothelin-1-evoked Ca2+ entry and store-operated Ca2+ entry evoked by thapsigargin were blunted. These results indicate that in aortic smooth muscle cells, TRPC1 is not only involved in Ca2+ entry activated by store depletion but also in receptor-operated Ca2+ entry, which requires inositol (1,4,5) triphosphate receptor activation.
authors.affiliation: Unit of Cellular Physiology, Université Catholique de Louvain, Bruxelles, Belgium.
authors.forename: Khalid
authors.initials: K
authors.lastname: Tai
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Hamaide
authors.forename: Huguette
authors.initials: H
authors.lastname: Debaix
authors.forename: Philippe
authors.initials: P
authors.lastname: Gailly
authors.forename: Maurice
authors.initials: M
authors.lastname: Wibo
authors.forename: Nicole
authors.initials: N
authors.lastname: Morel
mesh_terms: D000818:Animals; D000925:Anticoagulants / Q000008:administration & dosage / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D019332:Endothelin-1 / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation / Q000187:drug effects; D006493:Heparin / Q000008:administration & dosage / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000187:drug effects*; D008297:Male; D008845:Microinjections; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050052:TRPC Cation Channels / Q000819:agonists* / Q000235:genetics; D014162:Transfection; D014662:Vasoconstrictor Agents / Q000494:pharmacology
chemical_list: D000925:Anticoagulants; D002120:Calcium Channel Agonists; D019332:Endothelin-1; D053496:Inositol 1,4,5-Trisphosphate Receptors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D050052:TRPC Cation Channels; D014662:Vasoconstrictor Agents; C096351:transient receptor potential cation channel, subfamily C, member 1; D006493:Heparin; D002118:Calcium