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Plasma membrane calcium ATPase 1 (PMCA1, Atp2b1) is emerging as a key contributor to cardiac physiology, involved in calcium handling and myocardial signalling. In addition, genome wide association studies have associated PMCA1 in several areas of cardiovascular disease including hypertension and myocardial infarction. Here, we investigated the role of PMCA1 in basal cardiac function and heart rhythm stability. Cardiac structure, heart rhythm and arrhythmia susceptibility were assessed in a cardiomyocyte-specific PMCA1 deletion (PMCA1 CKO ) mouse model. PMCA1 CKO mice developed abnormal heart rhythms related to ventricular repolarisation dysfunction and displayed an increased susceptibility to ventricular arrhythmias. We further assessed the levels of cardiac ion channels using qPCR and found a downregulation of the voltage-dependent potassium channels, K v 4.2, with a corresponding reduction in the transient outward potassium current which underlies ventricular repolarisation in the murine heart. The changes in heart rhythm were found to occur in the absence of any structural cardiomyopathy. To further assess the molecular changes occurring in PMCA1 CKO hearts, we performed proteomic analysis. Functional characterisation of differentially expressed proteins suggested changes in pathways related to metabolism, protein-binding, and pathways associated cardiac function including β-adrenergic signalling. Together, these data suggest an important role for PMCA1 in basal cardiac function in relation to heart rhythm control, with reduced cardiac PMCA1 expression resulting in an increased risk of arrhythmia development., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Isoform 4 of the plasma membrane calcium/calmodulin dependent ATPase (PMCA4) has recently emerged as an important regulator of several key pathophysiological processes in the heart, such as contractility and hypertrophy. However, direct monitoring of PMCA4 activity and assessment of calcium dynamics in its vicinity in cardiomyocytes are difficult due to the lack of molecular tools. In this study, we developed novel calcium fluorescent indicators by fusing the GCaMP2 calcium sensor to the N-terminus of PMCA4 to generate the PMCA4-GCaMP2 fusion molecule. We also identified a novel specific inhibitor of PMCA4, which might be useful for studying the role of this molecule in cardiomyocytes and other cell types. Using an adenoviral system we successfully expressed PMCA4-GCaMP2 in both neonatal and adult rat cardiomyocytes. This fusion molecule was correctly targeted to the plasma membrane and co-localised with caveolin-3. It could monitor signal oscillations in electrically stimulated cardiomyocytes. The PMCA4-GCaMP2 generated a higher signal amplitude and faster signal decay rate compared to a mutant inactive PMCA4(mut)GCaMP2 fusion protein, in electrically stimulated neonatal and adult rat cardiomyocytes. A small molecule library screen enabled us to identify a novel selective inhibitor for PMCA4, which we found to reduce signal amplitude of PMCA4-GCaMP2 and prolong the time of signal decay (Tau) to a level comparable with the signal generated by PMCA4(mut)GCaMP2. In addition, PMCA4-GCaMP2 but not the mutant form produced an enhanced signal in response to β-adrenergic stimulation. Together, the PMCA4-GCaMP2 and PMCA4(mut)GCaMP2 demonstrate calcium dynamics in the vicinity of the pump under active or inactive conditions, respectively. In summary, the PMCA4-GCaMP2 together with the novel specific inhibitor provides new means with which to monitor calcium dynamics in the vicinity of a calcium transporter in cardiomyocytes and may become a useful tool to further study the biological functions of PMCA4. In addition, similar approaches could be useful for studying the activity of other calcium transporters during excitation-contraction coupling in the heart., (© 2013.)