Prolonged β-adrenergic stimulation disperses ryanodine receptor clusters in cardiomyocytes and has implications for...

Ryanodine receptors (RyRs) exhibit dynamic arrangements in cardiomyocytes, and we previously showed that 'dispersion' of RyR clusters disrupts Ca²⁺ homeostasis during heart failure (HF) (Kolstad et al., eLife, 2018). Here, we investigated whether prolonged β-adrenergic stimulation, a hallmark of HF, promotes RyR cluster dispersion and examined the underlying mechanisms. We observed that treatment of healthy rat cardiomyocytes with isoproterenol for 1 hr triggered progressive fragmentation of RyR clusters. Pharmacological inhibition of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) reversed these effects, while cluster dispersion was reproduced by specific activation of CaMKII, and in mice with constitutively active Ser2814-RyR. A similar role of protein kinase A (PKA) in promoting RyR cluster fragmentation was established by employing PKA activation or inhibition. Progressive cluster dispersion was linked to declining Ca²⁺ spark fidelity and magnitude, and slowed release kinetics from Ca²⁺ propagation between more numerous RyR clusters. In healthy cells, this served to dampen the stimulatory actions of β-adrenergic stimulation over the longer term and protect against pro-arrhythmic Ca²⁺ waves. However, during HF, RyR dispersion was linked to impaired Ca²⁺ release. Thus, RyR localization and function are intimately linked via channel phosphorylation by both CaMKII and PKA, which, while finely tuned in healthy cardiomyocytes, underlies impaired cardiac function during pathology.