Impaired Ca2+ cycling of nonischemic myocytes contributes to sarcomere dysfunction early after myocardial infarction

Abstract

Changes in the nonischemic remote myocardium of the heart contribute to left ventricular dysfunction after ischemia and reperfusion (I/R). Understanding the underlying mechanisms early after I/R is crucial to improve the adaptation of the viable myocardium to increased mechanical demands. Here, we investigated the role of myocyte Ca²⁺ handling in the remote myocardium 24 h after 60 min LAD occlusion. Cardiomyocytes isolated from the basal noninfarct-related parts of wild type mouse hearts demonstrated depressed beat-to-beat Ca²⁺ handling. The amplitude of the Ca²⁺ transients as well as the kinetics of Ca²⁺ transport were reduced by up to 25%. These changes were associated with impaired sarcomere contraction. While expression levels of Ca²⁺ regulatory proteins were unchanged in remote myocardium compared to the corresponding regions of sham-operated hearts, mobility shift analyses of phosphorylated protein showed 2.9 ± 0.4-fold more unphosphorylated phospholamban (PLN) monomers, the PLN species that inhibits the Ca²⁺ ATPase SERCA2a (P ≤ 0.001). Phospho-specific antibodies revealed normal phosphorylation of PLN at T17 in remote myocardium, but markedly reduced phosphorylation at its PKA-dependent phosphorylation site, S16 (P ≤ 0.01). The underlying cause involved enhanced activity of protein phosphatases, particularly PP2A (P ≤ 0.01). In contrast, overall PKA activity was normal. The PLN interactome, as determined by co-immunoprecipitation and mass spectrometry, and the phosphorylation state of PKA targets other than PLN were also unchanged. Isoproterenol enhanced cellular Ca²⁺ cycling much stronger in remote myocytes than in healthy controls and improved sarcomere function. We conclude that the reduced phosphorylation state of PLN at S16 impairs myocyte Ca²⁺ cycling in the remote myocardium 24 h after I/R and contributes to contractile dysfunction.

Keywords: Myocardial infarction; Myocyte calcium cycling; Phospholamban; Remote myocardium; SERCA2a; Sarcomere function.