CaMKII inhibition targeted to the sarcoplasmic reticulum inhibits frequency-dependent acceleration of relaxation and Ca2+ current facilitation

Abstract

Cardiac Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) in heart has been implicated in Ca 2+ current (I Ca ) facilitation, enhanced sarcoplasmic reticulum (SR) Ca2+ release and frequency dependent acceleration of relaxation (FDAR) via enhanced SR Ca 2+ uptake. However, questions remain about how CaMKII may work in these three processes. Here we tested the role of CaM-KII in these processes using transgenic mice (SR-AIP) that express four concatenated repeats of the CaMKII inhibitory peptide AIP selectively in the SR membrane. Wild type mice (WT) and mice expressing AIP exclusively in the nucleus (NLS-AIP) served as controls. Increasing stimulation frequency produced typical FDAR in WT and NLS-AIP, but FDAR was markedly inhibited in SR-AIP. Quantitative analysis of cytosolic Ca 2+ removal during [Ca 2+ ] i decline revealed that FDAR is due to an increased apparent V max of SERCA. CaMKII dependent RyR phosphorylation at Ser2815 and SR Ca 2+ leak were both decreased in SR-AIP vs. WT. This decrease in SR Ca 2+ leak may partly balance the reduced SERCA activity leading to relatively unaltered SR-Ca 2+ load in SR-AIP vs. WT myocytes. Surprisingly, CaMKII regulation of the L-type Ca 2+ channel (I Ca facilitation and recovery from inactivation) was abolished by the SR-targeted CaM-KII inhibition in SR-AIP mice. Inhibition of CaMKII effects on I Ca and RyR function by the SR-localized AIP places physical constraints on the localization of these proteins at the junctional microdomain. Thus SR-targeted CaMKII inhibition can directly inhibit the activation of SR Ca 2+ uptake, SR Ca 2+ release and I Ca by CaMKII, effects which have all been implicated in triggered arrhythmias.

Aerobic interval training enhances cardiomyocyte contractility and Ca2+ cycling by phosphorylation of CaMKII and Thr-17 of phospholamban

“…This hypothesis was supported by the effect of the specific CaMK-inhibitor AIP, which was able to substantially blunt the exercise training-induced increase in fractional shortening and Ca 2+ transient amplitude. Previous work has shown that CaMKII has a series of targets in the cell, including the L-type Ca 2+ channel, the ryanodine receptor 2 (RyR2), and SERCA-2a [8,9]. Hence, inhibiting CaMK with AIP may have limited the inotropic response of exercise by acting on a series of sites in the excitation-contraction coupling process that collectively abolished the training-induced effects.…”

“…Mainly, PLB is phosphorylated by cAMP-dependent protein kinase A (PKA) at serine (Ser)-16 and by Ca 2+ /calmodulin dependent kinase II (CaMKII) at threonine (Thr)-17 [8]. There are also indications of a PLB-independent action of CaMKII upon SERCA-2a that appears to increase diastolic SR Ca 2+ re-uptake [9].…”

“…Furthermore, CaMKII has been functionally implicated in the mediation of frequency-dependent acceleration of relaxation (FDAR) that contributes to optimal diastolic ventricular filling at increased heart rates [9], even in PLB-knockout mice [11].…”

Determinants of frequency-dependent contraction and relaxation of mammalian myocardium

“…These studies, using PLB KO animals, indicated that FDAR was not dependent on PLB but rather depended on CaMKII activity. In a recent study [40] the role of CaMKII was tested using TG mice that express four concatenated repeats of the CaMKII inhibitory peptide AIP (Auto Inhibitor Peptide). These mice show a moderate reduction in cardiac function and a decreased response to β-adrenergic stimulation.…”

“…As a result of these factors, the calcium decline may critically and predominantly modulate relaxation in unloaded myocytes, and thus manipulation of calcium decline will directly affect relaxation kinetics when load is absent. Indeed, in the unloaded isolated myocyte at sub-physiological pacing frequencies, inhibition of CAMKII targeted to the SR blunts FDAR [17,40]. However, relaxation of the myocardium under loaded conditions as it occurs in vivo is likely governed in a significantly different manner.…”