Bs-452682-2 the impaired intracellular calcium buffering in atrial myocytes of patients with long-standing persistent atrial fibrillation is mediated by reduced cardiac troponin c levels

Decreased levels of myofilament proteins and impaired atrial Ca2+ homeostasis contribute to atrial hypocontractility and thromboembolic events in long-standing persistent atrial fibrillation (AF). The myofilament protein cardiac troponin C (cTnC) is known to be the major cytosolic Ca2+ buffer, but its role in AF-associated Ca2+ handling abnormalities remains poorly investigated. This study aimed to clarify the influence of cTnC on impaired intracellular Ca2+ signalling in AF. Right atrial myocytes from sinus rhythm (Ctrl) and AF patients, as well as atrial induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) with knocked down cTnC levels (si-cTnC) were investigated by simultaneously measuring membrane currents (patch-clamp technique) and cytosolic Ca2+ (Fluo-3 AM and Fluo-4 AM). Myofilament protein levels were determined using immunoblotting. Cardiac troponin C protein levels were reduced in atrial tissues of AF patients. By integrating Na+-Ca2+-exchange current (INCX) during caffeine-induced Ca2+ release, SR Ca2+ content was determined and similar in both groups. Buffering parameters (maximum buffer capacity, Bmax, and dissociation constant of buffers, Kd) were obtained by fitting a hyperbolic function on buffering curves generated from INCX and the caffeine-induced Ca2+ transient. However, intracellular calcium buffering was reduced in AF, presumably due to the reduced cardiac troponin C. (Figures A and B). Interestingly, the reduction of cTnC levels using small interfering RNA (siRNA) caused the same alterations in intracellular Ca2+ buffering in atrial iPSC-CM. (Figure C). In these modelled cells, a higher Ca2+ spark frequency was observed as opposed to the control group (Figure D). Diminished cTnC levels underlie the reduced intracellular Ca2+ buffering observed in AF. By promoting spontaneous SR Ca2+ release events that could trigger delayed afterdepolarizations, reduced cTnC levels could contribute to the arrhythmogenic substrate promoting AF.