Intracellular Ca 2+ overload, prolongation of the action potential duration (APD), and downregulation of inward rectifier potassium (I K1 ) channel are hallmarks of electrical remodeling in cardiac hypertrophy and heart failure (HF). We hypothesized that enhancement of I K1 currents is a compensation for I K1 deficit and a novel modulation for cardiac Ca 2+ homeostasis and pathological remodeling. In adult Sprague-Dawley (SD) rats in vivo , cardiac hypertrophy was induced by isoproterenol (Iso) injection (i.p., 3 mg/kg/d) for 3, 10, and 30 days. Neonatal rat ventricular myocytes (NRVMs) were isolated from 1 to 3 days SD rat pups and treated with 1 μmol/L Iso for 24 h in vitro . The effects of zacopride, a selective I K1 /Kir2.1 channel agonist, on cardiac remodeling/hypertrophy were observed in the settings of 15 μg/kg in vivo and 1 μmol/L in vitro . After exposing to Iso for 3 days and 10 days, rat hearts showed distinct concentric hypertrophy and fibrosis and enhanced pumping function ( P < 0.01 or P < 0.05), then progressed to dilatation and dysfunction post 30 days. Compared with the age-matched control, cardiomyocytes exhibited higher cytosolic Ca 2+ ( P < 0.01 or P < 0.05) and lower SR Ca 2+ content ( P < 0.01 or P < 0.05) all through 3, 10, and 30 days of Iso infusion. The expressions of Kir2.1 and SERCA2 were downregulated, while p -CaMKII, p -RyR2, and cleaved caspase-3 were upregulated. Iso-induced electrophysiological abnormalities were also manifested with resting potential (RP) depolarization ( P < 0.01), APD prolongation ( P < 0.01) in adult cardiomyocytes, and calcium overload in cultured NRVMs ( P < 0.01). Zacopride treatment effectively retarded myocardial hypertrophy and fibrosis, preserved the expression of Kir2.1 and some key players in Ca 2+ homeostasis, normalized the RP ( P < 0.05), and abbreviated APD ( P < 0.01), thus lowered cytosolic [Ca 2 + ] i ( P < 0.01 or P < 0.05). I K1 channel blocker BaCl 2 or chloroquine largely reversed the cardioprotection of zacopride. We conclude that cardiac electrical remodeling is concurrent with structural remodeling. By enhancing cardiac I K1 , zacopride prevents Iso-induced electrical remodeling around intracellular Ca 2+ overload, thereby attenuates cardiac structural disorder a...
Arrhythmogenesis in acute myocardial infarction (MI) is associated with depolarization of resting membraine potential (RMP) and decrease of inward rectifier potassium current (IK1) in cardiomyocytes. However, clinical anti-arrhythmic agents that primarily act on RMP by enhancing the IK1 channel are not currently available. We hypothesized that zacopride, a selective and moderate agonist of the IK1/Kir2.1 channels, prevents and cures acute ischemic arrhythmias. To test this viewpoint, adult Sprague-Dawley (SD) rats were subjected to MI by ligating the left main coronary artery. The antiarrhythmic effects of zacopride (i.v. infusion) were observed in the settings of pre-treatment (zacopride given 3 min prior to coronary occlusion), post-treatment (zacopride given 3 min after coronary occlusion) and therapeutic treatment (zacopride given 30 s after the onset of the first sustained ventricular tachycardia (VT)/ventricular fibrillation (VF) post MI). In all the three treatment modes, zacopride (15 μg/kg) inhibited MI-induced ventricular tachyarrhythmias, as shown by significant decreases in the premature ventricular contraction (PVC) and the duration and incidence of VT or VF. In Langendorff perfused rat hearts, the antiarrhythmic effect of 1 μmol/L zacopride were reversed by 1 μmol/L BaCl2, a blocker of IK1 channel. Patch clamp results in freshly isolated rat ventricular myocytes indicated that zacopride activated the IK1 channel and thereby reversed hypoxia-induced RMP depolarization and action potential duration (APD) prolongation. In addition, zacopride (1 μmol/L) suppressed hypoxia- or isoproterenol- induced delayed afterdepolarizations (DADs). In Kir2.x transfected Chinese hamster ovary (CHO) cells, zacopride activated the Kir2.1 homomeric channel but not the Kir2.2 or Kir2.3 channels. These results support our hypothesis that moderately enhancing IK1/Kir2.1 currents as by zacopride rescues ischemia- and hypoxia- induced RMP depolarization, and thereby prevents and cures acute ischemic arrhythmias. This study brings a new viewpoint to antiarrhythmic theories and provides a promising target for the treatment of acute ischemic arrhythmias.
We recently reported that zacopride is a selective inward rectifier potassium current (I K1) channel agonist, suppressing ventricular arrhythmias without affecting atrial arrhythmias. The present study aimed to investigate the unique pharmacological properties of zacopride. The whole-cell patch-clamp technique was used to study I K1 currents in rat atrial myocytes and Kir2.x currents in human embryonic kidney (HEK)-293 cells transfected with inward rectifier potassium channel (Kir)2.1, Kir2.2, Kir2.3, or mutated Kir2.1 (at phosphorylation site S425L). Western immunoblots were performed to estimate the relative protein expression levels of Kir2.x in rat atria and ventricles. Results showed that zacopride did not affect the IK1 and transmembrane potential of atrial myocytes. In HEK293 cells, zacopride increased Kir2.1 homomeric channels by 40.7%±9.7% at -50 mV, but did not affect Kir2.2 and Kir2.3 homomeric channels, and Kir2.1-Kir2.2, Kir2.1-Kir2.3 and Kir2.2-Kir2.3 heteromeric channels. Western immunoblots showed that similar levels of Kir2.3 protein were expressed in rat atria and ventricles, but atrial Kir2.1 protein level was only 25% of that measured in the ventricle. In addition, 5-hydroxytryptamine (5-HT)3 receptor was undetectable, whereas 5-HT4 receptor was weakly expressed in HEK293 cells. The Kir2.1-activating effect of zacopride in these cells was abolished by inhibition of protein kinase A (PKA), but not PKC or PKG. Furthermore, zacopride did not activate the mutant Kir2.1 channel in HEK293 cells but selectively activated the Kir2.1 homomeric channel via a PKA-dependent pathway, independent to that of the 5-HT receptor.
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