title: Molecular and pharmacological characteristics of transient voltage-dependent K+ currents in cultured human pulmonary arterial smooth muscle cells.
abstract: The A-type voltage-dependent K(+) current (I(A)) has been identified in several types of smooth muscle cells including the pulmonary artery (PA), but little is known about the pharmacological and molecular characteristics of I(A) in human pulmonary arterial smooth muscle cells (hPASMCs). We investigated I(A) expressed in cultured PASMCs isolated from the human main pulmonary artery, using patch-clamp techniques, reverse transcriptase-polymerase chain reaction (RT-PCR), quantitative real-time RT-PCR and immunocytochemical studies. With high EGTA and ATP in the pipette, the outward currents were dominated by a transient K(+) current (I(A)), followed by a relatively small sustained outward current (I(K)). I(A) was inhibited by 4-aminopyridine (4-AP) concentration-dependently, and could be separated pharmacologically into two components by tetraethylammonium (TEA) sensitivity. A component was sensitive to TEA, and the second component was insensitive to TEA. I(A) was inhibited by blood depressing substrate (BDS)-II, a specific blocker of K(V)3.4 subunit, and phrixotoxin-II, a specific blocker of K(V)4.2 and 4.3. Flecainide inhibited I(A) concentration-dependently, but it inhibited it preferentially in the presence of TEA (TEA-insensitive I(A)). Systematic screening of expression of K(V) genes using RT-PCR showed the definite presence of transcripts of the I(A)-encoding genes for K(V)3.4, K(V)4.1, K(V)4.2 and K(V)4.3 as well as the I(K)-encoding genes for K(V)1.1, K(V)1.5 and K(V)2.1. The real-time RT-PCR analysis showed that the relative abundance of the encoding genes of I(A) alpha-subunit and K(V) channel-interacting proteins (KChIPs) was K(V)4.2 > K(V)3.4 > K(V)4.3 (long) > K(V)4.1, and KChIP3 >> KChIP2, respectively. The presence of K(V)3.4, K(V)4.2 and K(V)4.3 proteins was also demonstrated by immunocytochemical studies, and confirmed by immunohistochemical staining using intact human PA sections. These results suggest that I(A) in cultured hPASMCs consists of two kinetically and pharmacologically distinct components, probably K(V)3.4 and K(V)4 channels.
authors.affiliation: Department of Cardiovascular & Respiratory Medicine, University of Tokyo, Bunkyo-ku, Japan.
authors.forename: Haruko
authors.initials: H
authors.lastname: Iida
authors.forename: Taisuke
authors.initials: T
authors.lastname: Jo
authors.forename: Kuniaki
authors.initials: K
authors.lastname: Iwasawa
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Morita
authors.forename: Hisako
authors.initials: H
authors.lastname: Hikiji
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Takato
authors.forename: Teruhiko
authors.initials: T
authors.lastname: Toyo-Oka
authors.forename: Ryozo
authors.initials: R
authors.lastname: Nagai
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Nakajima
mesh_terms: D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D007150:Immunohistochemistry; D008564:Membrane Potentials / Q000187:drug effects*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology; D024642:Potassium Channels, Voltage-Gated / Q000187:drug effects* / Q000235:genetics* / Q000502:physiology; D011651:Pulmonary Artery / Q000187:drug effects / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D002135:Calcium-Binding Proteins; D026902:Potassium Channel Blockers; D024642:Potassium Channels, Voltage-Gated; D012333:RNA, Messenger