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Funding Acknowledgements Supported by the ÚNKP-19-3 New National Excellence program of the Ministry for Innovation and Technology Introduction and aims Adaptation of the human heart to physical activity is a complex mechanism that includes the change of heart rate, morphology of the action potential (AP) among others. Stimulation of β-adrenergic receptors (β-AR) causes the shortening of the AP duration of ventricular cardiomyocytes. This is caused by the regulation of the potassium currents by the β-adrenergic signaling pathway. Our aim was to investigate the role of protein kinase A (PKA) and calcium/calmodulin-dependent protein kinase II (CaMKII) in the regulation of the slow component (IKs) of the delayed rectifier potassium current under β-AR activation. Methods Our experiments were performed on isolated canine cardiomyocytes from the left ventricle. The IKs current profile was determined under a ventricular AP. We used "AP voltage clamp" conditions in six experimental groups: Control (CTRL), β-AR stimulation with isoproterenol (ISO), CaMKII inhibition with KN-93 (KN-93), PKA inhibition with H-89 (H-89) β-AR stimulation with inhibited CaMKII (KN-93 + ISO), β-AR stimulation with inhibited PKA (H-89 + ISO). β-AR stimulation with inhibited CaMKII and PKA (KN-93 + H-89 + ISO) Results The highest current density of IKs was approximately 6 times higher and the charge delivered by IKs was about 8 times larger in the ISO group than in CTRL or KN-93 conditions. In the KN-93 + ISO group, IKs amplitude was about 60% smaller and delivered about half the total charge compared to the ISO group. In the H‑89 + ISO group, IKs was about 30% smaller and delivered 40% less total charge than in the ISO group. In the KN-93 + H-89 + ISO group the IKs did not changed sicnificantly. Conclusion Based on our results, CaMKII plays an important role in regulating IKs by β-AR stimulation.

The role of transient receptor potential melastatin 4 (TRPM4) channels has been frequently tested using their inhibitor 9-phenanthrol in various cardiac preparations; however, the selectivity of the compound is uncertain. Therefore, in the present study, the concentration-dependent effects of 9-phenanthrol on major ionic currents were studied in canine isolated ventricular cells using whole-cell configuration of the patch-clamp technique and 10 mM BAPTA-containing pipette solution to prevent the Ca2+-dependent activation of TRPM4 channels. Transient outward (Ito1), rapid delayed rectifier (IKr), and inward rectifier (IK1) K+ currents were suppressed by 10 and 30 μM 9-phenanthrol with the blocking potency for IK1 < IKr < Ito1 and partial reversibility. L-type Ca2+ current was not affected up to the concentration of 30 μM. In addition, a steady outward current was detected at voltages positive to –40 mV in 9-phenanthrol, which was larger at more positive voltages and larger 9-phenanthrol concentrations. Action potentials were recorded using microelectrodes. Maximal rate of depolarization, phase-1 repolarization, and terminal repolarization were decreased and the plateau potential was depressed by 9-phenanthrol (3–30 μM), congruently with the observed alterations of ionic currents. Significant action potential prolongation was observed by 9-phenanthrol in the majority of the studied cells, but only at 30 μM concentration. In conclusion, 9-phenanthrol is not selective to TRPM4 channels in canine ventricular myocardium; therefore, its application as a TRPM4 blocker can be appropriate only in expression systems but not in native cardiac cells.

Although late sodium current (INa-late) has long been known to contribute to plateau formation of mammalian cardiac action potentials, lately it was considered as possible target for antiarrhythmic drugs. However, many aspects of this current are still poorly understood. The present work was designed to study the true profile of INa-late in canine and guinea pig ventricular cells and compare them to INa-late recorded in undiseased human hearts. INa-late was defined as a tetrodotoxin-sensitive current, recorded under action potential voltage clamp conditions using either canonic- or self-action potentials as command signals. Under action potential voltage clamp conditions the amplitude of canine and human INa-late monotonically decreased during the plateau (decrescendo-profile), in contrast to guinea pig, where its amplitude increased during the plateau (crescendo profile). The decrescendo-profile of canine INa-late could not be converted to a crescendo-morphology by application of ramp-like command voltages or command action potentials recorded from guinea pig cells. Conventional voltage clamp experiments revealed that the crescendo INa-late profile in guinea pig was due to the slower decay of INa-late in this species. When action potentials were recorded from multicellular ventricular preparations with sharp microelectrode, action potentials were shortened by tetrodotoxin, which effect was the largest in human, while smaller in canine, and the smallest in guinea pig preparations. It is concluded that important interspecies differences exist in the behavior of INa-late. At present canine myocytes seem to represent the best model of human ventricular cells regarding the properties of INa-late. These results should be taken into account when pharmacological studies with INa-late are interpreted and extrapolated to human. Accordingly, canine ventricular tissues or myocytes are suggested for pharmacological studies with INa-late inhibitors or modifiers. Incorporation of present data to human action potential models may yield a better understanding of the role of INa-late in action potential morphology, arrhythmogenesis, and intracellular calcium dynamics.

Intracellular calcium concentration ([Ca