Poster Presentation 2014 International Biophysics Congress

Effects of calmodulin on calcium activation of cardiac ryanodine receptor  (#385)

Ye Win Oo 1 , D.F. vanHelden 1 , M.S. Imtiaz 1 , D.R. Laver 1
  1. The University of Newcastle, Callaghan, NSW, Australia

During excitation-contraction coupling in cardiac muscle, depolarization of the sarcolemma causes the opening of voltage-gated (L-type) Ca2+ channels and the influx of Ca2+ into the cell1.  The subsequent rise in cytoplasmic [Ca2+] activates Ca2+ release channels (ryanodine receptors, RyR2) in the sarcoplasmic reticulum (SR, the Ca2+ store)1. Ca2+ release via RyR2 is regulated by Ca2+ and Mg2+ in the cytoplasm and SR lumen. Calmodulin (CaM, MW 16.7 kDa) is a cytosolic protein that can regulate cellular events by directly binding to proteins, including RyR22. CaM regulation of RyR2 was recently shown to be important normal cell function because mutations in CaM led to sudden death in humans3. CaM is normally bound to RyR2 in the cell and its dissociation causes increased SR Ca2+ release4.  However, when RyR2 are isolated from cells and examined by single channel recording in artificial lipid bilayers, CaM is not usually present because it dissociates from the RyR2 within minutes. Therefore, this study was designed to examine the effects of CaM by adding it back to RyR2 in lipid bilayers and measuring its effect on RyR2 regulation by Ca2+ and Mg2+. The open probability of RyR2 was 0.10 ± 0.02 (n=8) at 100 nM [Ca2+]c and 0.77 ± 0.1 (n=7) at 100 mM [Ca2+]c.  CaM (0.5 mM) reduced open probability of RyR2 by 63% ± 9.01 (n=8) at 100 nM [Ca2+]c and by 24% ± 6.46 (n=7) at 100 mM. The reductions in activity were mediated by reduced mean open time and increased mean closed time. 

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