title: TRPC3 regulates the automaticity of embryonic stem cell-derived cardiomyocytes.
abstract: BACKGROUND
  Cardiac pacemaking is a complex phenomenon that is not completely understood. Canonical transient receptor potential isoform 3 (TRPC3) channel is a cation channel that permeates both Ca(2+) and Na(+). TRPC3 was previously found to express in adult cardiomyocytes. However, its role in cardiac pacemaking is unexplored. Here we used mouse embryonic stem cell-derived cardiomyocytes (mESC-CMs) to investigate whether TRPC3 regulates the spontaneous automaticity and the underlying mechanism involved.
  
  
  RESULTS
  Immunocytochemistry results showed that TRPC3 is expressed at the T-tubules of mESC-CMs. Whole-cell patch clamping showed that single mESC-CMs contain TRPC3 current. Confocal Ca(2+) imaging showed that the TRPC3-specific blocker Pyr3 decreased Ca(2+) transients and local Ca(2+) release (LCR) of mESC-CMs. Combined current and voltage clamp recordings from the same cell showed that reducing the TRPC3 current, either by Pyr3 or a dominant negative (loss-of-function) construct of TRPC3, decreased the pacemaker activity of mESC-CMs as reflected by a decrease in action potential rate, a depolarized maximum diastolic potential and a decrease in slope of phase 4 diastolic depolarization. Furthermore, decreasing the TRPC3 current diminished, while increasing the TRPC3 current augmented the sodium-calcium exchanger (NCX) current in mESC-CMs. Lastly, decrease in TRPC3 current decreased the phosphorylation of ryanodine receptor isoform 2 at Ser2809 and phospholamban at Thr17.
  
  
  CONCLUSIONS
  TRPC3 positively regulates diastolic depolarization of spontaneous action potential by increasing LCR and NCX current and therefore is an important determinant in pacemaking of mESC-CMs.
authors.affiliation: School of Life Sciences, The Chinese University of Hong Kong, Hong Kong.
authors.forename: Zenghua
authors.initials: Z
authors.lastname: Qi
authors.affiliation: School of Life Sciences, The Chinese University of Hong Kong, Hong Kong.
authors.forename: Chun Kit
authors.initials: CK
authors.lastname: Wong
authors.affiliation: School of Life Sciences, The Chinese University of Hong Kong, Hong Kong.
authors.forename: Chi Ho
authors.initials: CH
authors.lastname: Suen
authors.affiliation: School of Life Sciences, South China Normal University, Guangzhou, PR China.
authors.forename: Jinzhao
authors.initials: J
authors.lastname: Wang
authors.affiliation: School of Life Sciences, South China Normal University, Guangzhou, PR China.
authors.forename: Cheng
authors.initials: C
authors.lastname: Long
authors.affiliation: Department of Physiology, Justus Liebig University Giessen, Germany.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Sauer
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong.
authors.forename: Xiaoqiang
authors.initials: X
authors.lastname: Yao
authors.affiliation: School of Life Sciences, The Chinese University of Hong Kong, Hong Kong; Key Laboratory for Regenerative Medicine, Ministry of Education, The Chinese University of Hong Kong, Hong Kong; State Key Laboratory of Agrobiotechnology, The Chinese University of Hong Kong, Hong Kong; Centre for Novel Biomaterials, The Chinese University of Hong Kong, Hong Kong. Electronic address: fayetsang@cuhk.edu.hk.
authors.forename: Suk Ying
authors.initials: SY
authors.lastname: Tsang
mesh_terms: D000200:Action Potentials; D000818:Animals; D053595:Embryonic Stem Cells; D051379:Mice; D032383:Myocytes, Cardiac / Q000502:physiology*; D050052:TRPC Cation Channels / Q000502:physiology*
chemical_list: D050052:TRPC Cation Channels; C104603:TRPC3 cation channel
keywords: Calcium; Canonical transient receptor potential isoform 3; Embryonic stem cell-derived cardiomyocytes; Phase 4 diastolic depolarization; Spontaneous action potential