title: Increased sarcoplasmic reticulum calcium leak but unaltered contractility by acute CaMKII overexpression in isolated rabbit cardiac myocytes.
abstract: The predominant cardiac Ca2+/calmodulin-dependent protein kinase (CaMK) is CaMKIIdelta. Here we acutely overexpress CaMKIIdeltaC using adenovirus-mediated gene transfer in adult rabbit ventricular myocytes. This circumvents confounding adaptive effects in CaMKIIdeltaC transgenic mice. CaMKIIdeltaC protein expression and activation state (autophosphorylation) were increased 5- to 6-fold. Basal twitch contraction amplitude and kinetics (1 Hz) were not changed in CaMKIIdeltaC versus LacZ expressing myocytes. However, the contraction-frequency relationship was more negative, frequency-dependent acceleration of relaxation was enhanced (tau(0.5Hz)/tau(3Hz)=2.14+/-0.10 versus 1.87+/-0.10), and peak Ca2+ current (ICa) was increased by 31% (-7.1+/-0.5 versus -5.4+/-0.5 pA/pF, P<0.05). Ca2+ transient amplitude was not significantly reduced (-27%, P=0.22), despite dramatically reduced sarcoplasmic reticulum (SR) Ca2+ content (41%; P<0.05). Thus fractional SR Ca2+ release was increased by 60% (P<0.05). Diastolic SR Ca2+ leak assessed by Ca2+ spark frequency (normalized to SR Ca2+ load) was increased by 88% in CaMKIIdeltaC versus LacZ myocytes (P<0.05; in an multiplicity-of-infection-dependent manner), an effect blocked by CaMKII inhibitors KN-93 and autocamtide-2-related inhibitory peptide. This enhanced SR Ca2+ leak may explain reduced SR Ca2+ content, despite measured levels of SR Ca2+-ATPase and Na+/Ca2+ exchange expression and function being unaltered. Ryanodine receptor (RyR) phosphorylation in CaMKIIdeltaC myocytes was increased at both Ser2809 and Ser2815, but FKBP12.6 coimmunoprecipitation with RyR was unaltered. This shows for the first time that acute CaMKIIdeltaC overexpression alters RyR function, leading to enhanced SR Ca2+ leak and reduced SR Ca2+ content but without reducing twitch contraction and Ca2+ transients. We conclude that this is attributable to concomitant enhancement of fractional SR Ca2+ release in CaMKIIdeltaC myocytes (ie, CaMKII-dependent enhancement of RyR Ca2+ sensitivity during diastole and systole) and increased ICa.
authors.affiliation: Abteilung Kardiologie & Pneumologie/Herzzentrum, Georg-August-Universität Göttingen, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Kohlhaas
authors.forename: Tong
authors.initials: T
authors.lastname: Zhang
authors.forename: Tim
authors.initials: T
authors.lastname: Seidler
authors.forename: Darya
authors.initials: D
authors.lastname: Zibrova
authors.forename: Nataliya
authors.initials: N
authors.lastname: Dybkova
authors.forename: Astrid
authors.initials: A
authors.lastname: Steen
authors.forename: Stefan
authors.initials: S
authors.lastname: Wagner
authors.forename: Lu
authors.initials: L
authors.lastname: Chen
authors.forename: Joan Heller
authors.initials: JH
authors.lastname: Brown
authors.forename: Donald M
authors.initials: DM
authors.lastname: Bers
authors.forename: Lars S
authors.initials: LS
authors.lastname: Maier
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000502:physiology; D002135:Calcium-Binding Proteins / Q000378:metabolism; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D005260:Female; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000201:enzymology / Q000502:physiology*; D010766:Phosphorylation; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000502:physiology; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000502:physiology; D000099211:Phospholamban
chemical_list: D002118:Calcium; D020746:Calcium Channels, L-Type; D002135:Calcium-Binding Proteins; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D019837:Ryanodine Receptor Calcium Release Channel; D019831:Sodium-Calcium Exchanger; D000099211:Phospholamban