The patch clamp technique underwent continual advancement and developed numerous variants in cardiac electrophysiology since its introduction in the late 1970s. In the beginning, the capability of the technique was limited to recording one single current from one cell stimulated with a rectangular command pulse. Since that time, the technique has been extended to record multiple currents under various command pulses including action potential. The current review summarizes the development of the patch clamp technique in cardiac electrophysiology with special focus on the potential applications in integrative physiology.

The objective of this work is to examine the contribution of late Na(+) current (INa,L) to the cardiac action potential (AP) and arrhythmogenic activities. In spite of the rapidly growing interest toward this current, there is no publication available on experimental recording of the dynamic INa,L current as it flows during AP with Ca(2+) cycling. Also unknown is how the current profile changes when the Ca(2+)-calmodulin dependent protein kinase II (CaMKII) signaling is altered, and how the current contributes to the development of arrhythmias. In this study we use an innovative AP-clamp Sequential Dissection technique to directly record the INa,L current during the AP with Ca(2+) cycling in the guinea pig ventricular myocytes. First, we found that the magnitude of INa,L measured under AP-clamp is substantially larger than earlier studies indicated. CaMKII inhibition using KN-93 significantly reduced the current. Second, we recorded INa,L together with IKs, IKr, and IK1 in the same cell to understand how these currents counterbalance to shape the AP morphology. We found that the amplitude and the total charge carried by INa,L exceed that of IKs. Third, facilitation of INa,L by Anemone toxin II prolonged APD and induced Ca(2+) oscillations that led to early and delayed afterdepolarizations and triggered APs; these arrhythmogenic activities were eliminated by buffering Ca(2+) with BAPTA. In conclusion, INa,L contributes a significantly large inward current that prolongs APD and unbalances the Ca(2+) homeostasis to cause arrhythmogenic APs.

The role of Ca²⁺-activated Cl^- current (I_Cl(Ca)) in cardiac arrhythmias is still controversial. It can generate delayed afterdepolarizations in Ca²⁺-overloaded cells while in other studies incidence of early afterdepolarization (EAD) was reduced by I_Cl(Ca). Therefore our goal was to examine the role of I_Cl(Ca) in spatial and temporal heterogeneity of cardiac repolarization and EAD formation. Experiments were performed on isolated canine cardiomyocytes originating from various regions of the left ventricle; subepicardial, midmyocardial and subendocardial cells, as well as apical and basal cells of the midmyocardium. I_Cl(Ca) was blocked by 0.5mmol/L 9-anthracene carboxylic acid (9-AC). Action potential (AP) changes were tested with sharp microelectrode recording. Whole-cell 9-AC-sensitive current was measured with either square pulse voltage-clamp or AP voltage-clamp (APVC). Protein expression of TMEM16A and Bestrophin-3, ion channel proteins mediating I_Cl(Ca), was detected by Western blot. 9-AC reduced phase-1 repolarization in every tested cell. 9-AC also increased AP duration in a reverse rate-dependent manner in all cell types except for subepicardial cells. Neither I_Cl(Ca) density recorded with square pulses nor the normalized expressions of TMEM16A and Bestrophin-3 proteins differed significantly among the examined groups of cells. The early outward component of I_Cl(Ca) was significantly larger in subepicardial than in subendocardial cells in APVC setting. Applying a typical subepicardial AP as a command pulse resulted in a significantly larger early outward component in both subepicardial and subendocardial cells, compared to experiments when a typical subendocardial AP was applied. Inhibiting I_Cl(Ca) by 9-AC generated EADs at low stimulation rates and their incidence increased upon beta-adrenergic stimulation. 9-AC increased the short-term variability of repolarization also. We suggest a protective role for I_Cl(Ca) against risk of arrhythmias by reducing spatial and temporal heterogeneity of cardiac repolarization and EAD formation.