Remodeling of sarcoplasmic reticulum Ca ²⁺ uptake in cardiac Purkinje cells after ischemic myocardial infarction in various large mammalian species and humans

After myocardial infarction (MI), increased spontaneous sarcoplasmic reticulum (SR)-Ca²⁺ releases depolarize the membrane and trigger action potentials (APs) in cardiac Purkinje cells (Pcells). This abnormal Ca²⁺-activity is involved in ventricular fibrillation. Spontaneous Ca²⁺-transients analysis suggested that intensification of SR-Ca²⁺ uptake accounts for the abnormal SR-Ca²⁺ release in post-MI Pcells. Increased SR-Ca²⁺-pump (SERCA) density, phospholamban (PLB)-dependent Ca²⁺-pump activation, and modification of the pump Ca²⁺-transport properties can mediate an increase in SR-Ca²⁺ uptake. We examined whether Pcells of ischemic hearts show signs of these alterations, hence supporting the hypothesis of post-MI increase in SR-Ca²⁺-uptake. Pcells were prepared from hearts with and without MI in dogs, sheep, pigs, and humans. The distribution of SR-Ca²⁺ pumps and phosphorylated forms of PLB, pPLBSer16 and pPLBThr17, was captured by specific immunofluorescence and confocal microscopy. Protein and transcript levels of sarco/endoplasmic reticulum calcium ATPase isoform 2 (SERCA2) subisoforms were measured in Purkinje fibers and myocardium by Western blot (WB) and reverse transcription quantitative polymerase chain reaction (RT-qPCR), respectively. In normal hearts, Ca²⁺ pumps and PLB antibodies colocalized throughout Pcells. After MI, Ca²⁺ pump staining exhibited larger intensity in peripheral compared with central regions of Pcells. Phosphorylated PLB staining was unchanged, indicating no alteration of the pump-β-adrenergic regulation after MI. Expression of the regular cardiac pump, SERCA2a, was preserved. However, the emergence of another pump, SERCA2b, was found after MI. The addition of SERCA2b to the existing SERCA2a expression increased the total pump density, which was consistent with an augmentation of SR-Ca²⁺-uptake in Pcells after MI. After MI, the peripheral region of Pcells seems to express the SERCA2b pump subisoform, which is consistent with larger pump density and intensification of SR-Ca²⁺ uptake.<b>NEW & NOTEWORTHY</b> A gradient in the density of SR-Ca²⁺ pumps appears from the center to the periphery of Purkinje cells (Pcells) after MI. We found that this post-MI rearrangement could result from the peripheral expression of SERCA2b pump, which is absent in healthy hearts. The additional expression of SERCA2b to the existing cardiac pump SERCA2a, and possibly more efficient Ca²⁺-transport properties of SERCA2b, are consistent with the proarrhythmic elevation of SR-Ca²⁺ uptake previously proposed in Pcells after MI.