The gap junction modifier ZP1609 decreases cardiomyocyte hypercontracture following ischaemia/reperfusion independent from mitochondrial connexin 43.

Abstract

Dysregulation of gap junction-mediated cell coupling contributes to development of arrhythmias and myocardial damage after ischaemia/reperfusion (I/R). Connexin 43 (Cx43) is present at ventricular gap junctions and also in the mitochondria of cardiomyocytes. The dipeptide (2S, 4R)-1-(2-aminoacetyl)-4-benzamidopyrrolidine-2-carboxylic acid (ZP1609) has antiarrhythmic properties and reduces infarct size when given at reperfusion. However, it is unclear, whether ZP1609 targets Cx43-containing mitochondria and affects cardiomyocyte hypercontracture following I/R. We studied the effects of ZP1609 on the function of murine sub-sarcolemmal mitochondria (SSM, containing Cx43) and interfibrillar mitochondria (IFM, lacking Cx43). Murine isolated cardiomyocytes were subjected to simulated I/R without and with ZP1609 (applied during I/R or at the onset of reperfusion only), and the number of cardiomyocytes undergoing hypercontracture was quantified. Biochemical pathways targeted by ZP1609 in cardiomyocytes were analysed. ZP1609 inhibited ADP-stimulated respiration and ATP production in SSM and IFM. ROS formation and calcium retention capacities in SSM and IFM were not affected by ZP1609, whereas potassium uptake was enhanced in IFM. The number of rod-shaped cardiomyocytes was increased by ZP1609 (10μM) when administered either during I/R or reperfusion. ZP1609 altered the phosphorylation of proteins contributing to the protection against I/R injury. ZP1609 reduced mitochondrial respiration and ATP production, but enhanced potassium uptake of IFM. Additionally, ZP1609 reduced the extent of cardiomyocytes undergoing hypercontracture following I/R. The protective effect was independent of mitochondrial Cx43, as ZP1609 exerts its effects in Cx43-containing SSM and Cx43-lacking IFM.