Measurement of Ca2+ Transients in NRCs

The measurement of the fluorometric Ca²⁺ transients in NRCs was performed as described by Kirschmer et al. (16 (link)). Briefly, cells were cultured on cover slides coated with laminin (Roche) and loaded for 20 min at room temperature with Fura-2 (2 mM) in a Ca²⁺-free normal Tyrode solution (140 mM NaCl, 4 mM KCl, 1 mM MgCl₂, 5 mM Hepes, 10 mM glucose, pH 7.4). Ca²⁺ transients were measured in normal Tyrode solution supplemented with 1 mM CaCl₂ at a pacing frequency of 1 Hz using the “Myocyte and Contractility System” from Ionoptix. Data were corrected for background fluorescence 340/380 and analyzed using the IonWizard 6.3 software (Ionoptix).