Increased sarcoplasmic reticulum calcium leak but unaltered contractility by acute CaMKII overexpression in isolated rabbit cardiac myocytes

The predominant cardiac Ca ²⁺/calmodulin-dependent protein kinase (CaMK) is CaMKIIδ. Here we acutely overexpress CaMKIIδ _C using adenovirus-mediated gene transfer in adult rabbit ventricular myocytes. This circumvents confounding adaptive effects in CaMKIIδ _C transgenic mice. CaMKIIδ _C protein expression and activation state (autophosphorylation) were increased 5- to 6-fold. Basal twitch contraction amplitude and kinetics (1 Hz) were not changed in CaMKIIδ _C versus LacZ expressing myocytes. However, the contraction-frequency relationship was more negative, frequency-dependent acceleration of relaxation was enhanced (τ _0.5Hz/τ _3Hz=2.14±0.10 versus 1.87±0.10), and peak Ca ²⁺ current (I _Ca) was increased by 31% (-7.1±0.5 versus -5.4±0.5 pA/pF, P<0.05). Ca ²⁺ transient amplitude was not significantly reduced (-27%, P=0.22), despite dramatically reduced sarcoplasmic reticulum (SR) Ca ²⁺ content (41%; P<0.05). Thus fractional SR Ca ²⁺ release was increased by 60% (P<0.05). Diastolic SR Ca ²⁺ leak assessed by Ca ²⁺ spark frequency (normalized to SR Ca ²⁺ load) was increased by 88% in CaMKIIδ _C 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 Ca ²⁺ leak may explain reduced SR Ca ²⁺ content, despite measured levels of SR Ca ²⁺-ATPaSe and Na ⁺/Ca ²⁺ exchange expression and function being unaltered. Ryanodine receptor (RyR) phosphorylation in CaMKIIδ _C myocytes was increased at both Ser2809 and Ser2815, but FKBP12.6 coimmunoprecipitation with RyR was unaltered. This shows for the first time that acute CaMKIIδ _C overexpression alters RyR function, leading to enhanced R Ca ²⁺ leak and reduced SR Ca ²⁺ content but without reducing twitch contraction and Ca ²⁺ transients. We conclude that this is attributable to concomitant enhancement of fractional SR Ca ²⁺ release in CaMKIIδ _C myocytes (ie, CaMKII-dependent enhancement of RyR Ca ²⁺ sensitivity during diastole and systole) and increased I _Ca.