Sedentary ageing remodels the structure–function relationships that underlie calcium signalling at super-resolution scales in the heart

Intracellular calcium handling underpins excitation–contraction coupling in the heart, and its dysfunction has been linked to both mechanical and electrical impairments. Age-related remodelling of the structure–function relationships that underpin calcium spark morphology and dynamics may therefore be causatively linked to associated cardiac dysfunction. We aim to uncover age-dependent features of calcium spark morphology and the distribution of type-2 ryanodine receptor (RyR) clusters, responsible for intracellular calcium release. Isolated ventricular cardiomyocytes were obtained from male Wistar rats at 6, 18 and 24 months of age. Subcellular structure was assessed using expansion microscopy to localize RyR, and calcium sparks were recorded in the absence and presence of 1 µM isoprenaline using spinning disk confocal microscopy. Super-resolution imaging revealed that both aged points were associated with a greater degree of cluster fragmentation, sparseness and structural disorder than the 6-month group, leading to an increased prevalence of remodelled phenotype cells in the aged groups. The 18-month group exhibited the most substantial remodelling of many properties, including the prevalence of remodelled cells, indicating either a genuine biphasic progression of remodelling with age or the divergence of structurally healthy and structurally remodelled groups with age. Moreover, ageing enhanced the impact of β-adrenergic stimulation on calcium spark morphology, increasing spark amplitude, full-width half maximum and spark mass. Our study reveals that RyR cluster properties are non-linearly remodelled with age, with RyR phosphorylation leading to remodelling of calcium spark morphology with age, which may contribute to reduced contractile performance and increased arrhythmia incidence.