CaMKII δ Met281/282 oxidation is not required for recovery of calcium transients during acidosis

• Published in: American journal of physiology. Heart and circulatory physiology Vol. 320; no. 3; pp. H1199 - H1212
• Main Authors: Kreitmeier, K G, Tarnowski, D, Nanadikar, M S, Baier, M J, Wagner, S, Katschinski, D M, Maier, L S, Sag, C M
• Format: Journal Article
• Language: English
• Published: United States 01.03.2021
• Subjects: Acidosis - complications; Acidosis - enzymology; Animals; Arrhythmias, Cardiac - enzymology; Arrhythmias, Cardiac - etiology; Arrhythmias, Cardiac - physiopathology; Biosensing Techniques; Calcium Signaling; Calcium-Binding Proteins - metabolism; Calcium-Calmodulin-Dependent Protein Kinase Type 2 - genetics; Calcium-Calmodulin-Dependent Protein Kinase Type 2 - metabolism; Female; Glutathione - metabolism; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Heart Rate; Hydrogen-Ion Concentration; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Myocardial Contraction; Myocytes, Cardiac - enzymology; Oxidation-Reduction; Phosphorylation; Reactive Oxygen Species - metabolism; redox signaling; CaMKII; excitation-contraction coupling; acidosis; calcium signaling
• ISSN: 0363-6135; 1522-1539
• DOI: 10.1152/ajpheart.00040.2020

CaMKII is needed for the recovery of Ca transients during acidosis but also mediates postacidic arrhythmias. CaMKIIδ can sustain its activity following Met281/282 oxidation. Increasing cytosolic Na during acidosis as well as postacidic pH normalization should result in prooxidant conditions within the cell favoring oxidative CaMKIIδ activation. We tested whether CaMKIIδ activation through Met281/282 oxidation is involved in recovery of Ca transients during acidosis and promotes cellular arrhythmias post-acidosis. Single cardiac myocytes were isolated from a well-established mouse model in which CaMKIIδ was made resistant to oxidative activation by knock-in replacement of two oxidant-sensitive methionines (Met281/282) with valines (MM-VV). MM-VV myocytes were exposed to extracellular acidosis (pH 6.5) and compared to wild type (WT) control cells. Full recovery of Ca transients was observed in both WT and MM-VV cardiac myocytes during late-phase acidosis. This was associated with comparably enhanced sarcoplasmic reticulum Ca load and preserved CaMKII specific phosphorylation of phospholamban at Thr17 in MM-VV myocytes. CaMKII was phosphorylated at Thr287, but not Met281/282 oxidized. In line with this, postacidic cellular arrhythmias occurred to a similar extent in WT and MM-VV cells, whereas inhibition of CaMKII using AIP completely prevented recovery of Ca transients during acidosis and attenuated postacidic arrhythmias in MM-VV cells. Using genetically altered cardiomyocytes with cytosolic expression of redox-sensitive green fluorescent protein-2 coupled to glutaredoxin 1, we found that acidosis has a reductive effect within the cytosol of cardiac myocytes despite a significant acidosis-related increase in cytosolic Na . Our study shows that activation of CaMKIIδ through Met281/282 oxidation is neither required for recovery of Ca transients during acidosis nor relevant for postacidic arrhythmogenesis in isolated cardiac myocytes. Acidosis reduces the cytosolic glutathione redox state of isolated cardiac myocytes despite a significant increase in cytosolic Na . Pharmacological inhibition of global CaMKII activity completely prevents recovery of Ca transients and protects from postacidic arrhythmias in MM-VV myocytes, which confirms the relevance of CaMKII in the context of acidosis. The current study shows that activation of CaMKIIδ through Met281/282 oxidation is neither required for CaMKII-dependent recovery of Ca transients during acidosis nor relevant for the occurrence of postacidic cellular arrhythmias. Despite a usually prooxidant increase in cytosolic Na , acidosis reduces the cytosolic glutathione redox state within cardiac myocytes. This novel finding suggests that oxidation of cytosolic proteins is less likely to occur during acidosis.