beta-adrenergic regulation of late Na+ current during cardiac action potential is mediated by both PKA and CaMKII

Late Na+ current (I-NaL) significantly contributes to shaping cardiac action potentials (APs) and increased I-NaL is associated with cardiac arrhythmias. beta-adrenergic receptor (beta AR) stimulation and its downstream signaling via protein kinase A (PKA) and Ca2+/calmodulin-dependent protein kinase II (CaMKII) pathways are known to regulate INaL. However, it remains unclear how each of these pathways regulates I-NaL during the AP under physiological conditions. Here we performed AP-clamp experiments in rabbit ventricular myocytes to delineate the impact of each signaling pathway on INaL at different AP phases to understand the arrhythmogenic potential. During the physiological AP (2 Hz, 37 degrees C) we found that I-NaL had a basal level current independent of PKA, but partially dependent on CaMKII. beta AR activation (10 nM isoproterenol, ISO) further enhanced I-NaL via both PKA and CaMKII pathways. However, PKA predominantly increased I-NaL early during the AP plateau, whereas CaMKII mainly increased I-NaL later in the plateau and during rapid repolarization. We also tested the role of key signaling pathways through exchange protein activated by cAMP (Epac), nitric oxide synthase (NOS) and reactive oxygen species (ROS). Direct Epac stimulation enhanced INaL similar to the beta AR-induced CaMKII effect, while NOS inhibition prevented the beta AR-induced CaMKII-dependent INaL enhancement. ROS generated by NADPH oxidase 2 (NOX2) also contributed to the ISO-induced I-NaL activation early in the AP. Taken together, our data reveal differential modulations of I-NaL by PKA and CaMKII signaling pathways at different AP phases. This nuanced and comprehensive view on the changes in INaL during AP deepens our understanding of the important role of I-NaL in reshaping the cardiac AP and arrhythmogenic potential under elevated sympathetic stimulation, which is relevant for designing therapeutic treatment of arrhythmias under pathological conditions.