α1-adrenoceptor stimulation potentiates L-type Ca 2+ current through Ca2+/calmodulin-dependent PK II (CaMKII) activation in rat ventricular myocytes

Jin O-Uchi, Kimiaki Komukai, Yoichiro Kusakari, Toru Obata, Kenichi Hongo, Hiroyuki Sasaki, Satoshi Kurihara

Research output: Contribution to journalArticlepeer-review

48 Scopus citations

Abstract

α1-Adrenoceptor stimulation (α1ARS) modulates cardiac muscle contraction under physiological conditions by means of changes in Ca2+ current through L-type channels (ICa,L) and Ca2+ sensitivity of the myofilaments. However, the cellular mechanisms of α1ARS are not fully clarified. In this study, we investigated the role of Ca2+/calmodulin-dependent PK II (CaMKII) in the regulation of ICa,L during α1ARS in isolated adult rat ventricular myocytes by using the perforated patch-clamp technique. CaMKII inhibition with 0.5 μM KN-93 abolished the potentiation in I Ca,L observed during α1ARS by 10 μM phenylephrine. In the presence of PKC inhibitor (10 μM chelerythrine), the potentiation of ICa,L by phenylephrine also disappeared. In Western immunoblotting analysis, phenylephrine (≥ μM) increased the amount of autophosphorylated CaMKII (active CaMKII) significantly, and this increase was abolished by CaMKII inhibition or PKC inhibition. Also, we investigated changes in the subcellular localization of active CaMKII by using immunofluorescence microscopy and immunoelectron microscopy. Before α1ARS, active CaMKII was exclusively located just beneath the plasmalemma. However, after α1ARS, active CaMKII was localized close to transverse tubules, where most of L-type Ca2+ channels are located. From these results, we propose that CaMKII, which exists near transverse tubules, is activated and phosphorylated by α1ARS and that CaMKII activation directly potentiates ICa,L in rat ventricular myocytes.

Original languageEnglish (US)
Pages (from-to)9400-9405
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number26
DOIs
StatePublished - Jun 28 2005
Externally publishedYes

Keywords

  • Cardiac muscle
  • PKC
  • Perforated patch-clamp
  • Phenylephrine

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