Intracellular calcium dynamics, shortened action potential duration, and late-phase 3 early afterdepolarization in Langendorff-perfused rabbit ventricles

Abstract

Introduction: To elucidate the mechanism of late-phase 3 early after depolarization (EAD) in ventricular arrhythmogenesis, we hypothesized that intracellular calcium (Ca(i) ) overloading and action potential duration (APD) shortening may promote late-phase 3 EAD and triggered activity, leading to development of ventricular fibrillation (VF).

Methods and results: In isolated rabbit hearts, we performed microelectrode recording and simultaneous dual optical mapping of transmembrane potential (V(m) ) and Ca(i) transient on left ventricular endocardium. An I(KATP) channel opener, pinacidil, was used to abbreviate APD. Rapid pacing was then performed. Upon abrupt cessation of rapid pacing with cycle lengths of 60-200 milliseconds, there were APD(90) prolongation and the corresponding Ca(i) overloading in the first postpacing beats. The duration of Ca(i) transient recovered to 50% (DCaT(50) ) and 90% (DCaT(90) ) in the first postpacing beats was significantly longer than baseline. Abnormal Ca(i) elevation coupled with shortened APD produced late-phase 3 EAD induced triggered activity and VF. In additional 6 preparations, the heart tissues were treated with BAPTA-AM, a calcium chelator. BAPTA-AM significantly reduced the maximal Ca(i) amplitude (26.4 ± 3.5% of the control; P < 0.001) and the duration of Ca(i) transients in the mapped region, preventing the development of EAD and triggered activity that initiated VF.

Conclusions: I (KATP) channel activation along with Ca(i) overloading are associated with the development of late-phase 3 EAD and VF. Because acute myocardial ischemia activates the I(KATP) channel, late-phase 3 EADs may be a mechanism for VF initiation during acute myocardial ischemia.

Figure 1

A: Cut-open LV preparation in Langendorff-perfused isolated rabbit hearts. LA: left atrium; RCA: right coronary artery; LV endo: left ventricle endocardium; RV: right ventricle; a: anterior papillary muscle; b: posterior papillary muscle. B: Optical recording traces of V_m and Ca_i of LV endocardium at pacing cycle length (PCL) of 250 ms. APD₅₀: action potential duration measured to 50% repolarization; APD₉₀: action potential duration measured to 90% repolarization; DCaT₅₀: duration of Ca_i transient measured to 50% repolarization; DCaT₉₀: duration of Ca_i transient measured to 90% repolarization. C: Effects of pinacidil on APD₅₀, APD₉₀, DCaT₅₀, and DCaT₉₀ as a function of pacing cycle length between 100 and 350 ms. D: Comparison of the APD and DCaT in control and pinacidil at PCL of 180 ms. ^*p<0.0001; ^{^}p<0.001

Figure 2

A: Optical signals of V_m and Ca_i of beats following pause after pacing with pinacidil. Rapid pacing with pinacidil produced an APD prolongation of first post-pacing beat. Blue traces indicate the morphology of action potentials of the last beat in the pacing train. S, pacing stimulus. B: Pinacidil induced a longer pause following pacing train. ^*p<0.001. C: Effects of pinacidil on APD₅₀, APD₉₀, DCaT₅₀, and DCaT₉₀ of first post-pacing beat when PCL decreased from 350 to 100 ms. Rapid pacing under pinacidil produced a prolongation of APD₉₀ and increase of DCaT₅₀ and DCaT₉₀. ^*p<0.001; ^{^}p<0.01 as compared to pacing cycle length greater than 250 ms.

Figure 3

A: Typical V_m and Ca_i signals of late-phase 3 EAD induced triggered activity and ventricular fibrillation. The late phase 3 EAD is the delayed repolarization that occurred before the dashed line on the V_m tracing. PCL: 90 ms. S: pacing stimulus. B: V_m and Ca_i recordings for VF onset induced by EADs after post-pacing pause. PCL at 70 ms. Crosse indicates the origin of the EAD in the mapped region.

Figure 4

Glass microelectrode recordings of transmembrane potential (TMP) of single myocyte in LV endocardium of isolated rabbit hearts. Blue traces indicate the TMP recording of the last paced beat in the train. A: control study. B and C: pinacidil infusion to produce APD₉₀ prolongation in first post-pacing beat (B) and late-phase 3 EAD induced triggered activities and VF (C). PCL at 120 ms. D: comparison of the post-pacing pause duration in control, EAD, and VF induction. ^*p<0.0001

Figure 5

A: BAPTA-AM significantly reduced the maximal Ca_i amplitudes normalized to baseline. B: V_m and Ca_i optical signal recordings after BAPTA-AM perfusion showed no APD prolongation in first post-pacing beats. Blue traces indicate the action potential recording of the last paced beats in the train. C: Glass microelectrode TMP recording in BAPTA-AM study. D: Effect of BAPTA-AM on post-rapid pacing APD₉₀. Optical imaging data (n=6). E: BAPTA-AM significantly reduced the corresponding Ca_i increase in first post-pacing beat. The Ca_i increase magnitude was normalized to the Ca_i level in last pacing beat. ^* p<0.001.