Cardiac action potential (AP) is formed by many different ion currents. Abnormalities of these may result in cardiac arrhythmias leading to serious complications such as sudden cardiac death. Therefore, it has a significant importance to understand the role of each ion current. To study a given ion current, selective ion channel blockers are frequently used.
Our aim was to determine either 4,4-diisothiocyanostilbene-2,2-disulfonic acid (DIDS) or 9-anthracene carboxylic acid (9-AC) is more suitable to study calcium-activated chloride current (ICl(Ca)) contributing to early repolarization.
Whole-cell voltage-clamp was used to measure inward rectifier K+ current (IK1), rapid and slow components of delayed rectifier K+ currents (IKr and IKs) and L-type Ca2+ current (ICa,L). Sharp microelectrode technique was used to record APs. All experiments were done on enzymatically isolated canine left ventricular cells at 37 °C.
At fast stimulation rates, 0.5 mM 9-AC produced larger elevation in the plateau potential whereas the AP lengthened to the same extent by both drugs. While 0.2 mM DIDS resulted in a marked reduction of the maximal rate of depolarization, no such effect was seen with 9-AC. Early repolarization of the AP (phase-1) was reduced by both drugs, a greater reduction observed with DIDS during slow stimulation rates. All effects on AP of both drugs were reversible upon washout. Regardless of intracellular Ca2+ buffering, 9-AC- and DIDS-sensitive currents were identical in voltage-clamp condition. Increase in stimulation rate induced a correlating increase of calcium transients and DIDS-sensitive current. Increasing 9-AC concentrations from 0.1 to 0.5 mM in a cummulative manner a gradual reduction of phase-1 and lengthening of AP could be observed, while no further action could be detected at 1 mM. The half maximal effective concentration of 9-AC was 160 microM with a Hill coefficient of 2. At 0.5 mM, 9-AC did not affect ICa,L, IKs, IKr and IK1.
Based on these results, in voltage-clamp studies both DIDS and 9-AC proved equally useful but 9-AC proved superior than DIDS in AP studies to study the physiological role of ICl(Ca) due to the lack of Na+-channel inhibition.