SkM1 and Cx32 improve conduction in canine myocardial infarcts yet only SkM1 is antiarrhythmic

Abstract

Aims: Reentry accounts for most life-threatening arrhythmias, complicating myocardial infarction, and therapies that consistently prevent reentry from occurring are lacking. In this study, we compare antiarrhythmic effects of gene transfer of green fluorescent protein (GFP; sham), the skeletal muscle sodium channel (SkM1), the liver-specific connexin (Cx32), and SkM1/Cx32 in the subacute canine infarct.

Methods and results: Immediately after ligation of the left anterior descending artery, viral constructs were implanted in the epicardial border zone (EBZ). Five to 7 days later, efficient restoration of impulse propagation (narrow QRS and local electrogram duration) occurred in SkM1, Cx32, and SkM1/Cx32 groups (P< 0.05 vs. GFP). Programmed electrical stimulation from the EBZ induced sustained ventricular tachycardia (VT)/ventricular fibrillation (VF) in 15/22 GFP dogs vs. 2/12 SkM1, 6/14 Cx32, and 8/10 SkM1/Cx32 (P< 0.05 SkM1 vs. GFP). GFP, SkM1, and SkM1/Cx32 had predominantly polymorphic VT/VF, whereas in Cx32 dogs, monomorphic VT predominated (P< 0.05 for Cx32 vs. GFP). Tetrazolium red staining showed significantly larger infarcts in Cx32- vs. GFP-treated animals (P< 0.05).

Conclusion: Whereas SkM1 gene transfer reduces the incidence of inducible VT/VF, Cx32 therapy to improve gap junctional conductance results in larger infarct size, a different VT morphology, and no antiarrhythmic efficacy.

Figure 1

Immunohistochemistry of the EBZ. Positive SkM1 (green) and Cx32 (red) staining was detected in animals that received the corresponding adenovirus. Nuclei were stained blue using DAPI. Bars represent 50 µm.

Figure 2

Typical recordings of lead II ECG (upper tracings) and local EGs (lower tracings)—in the healthy myocardium (left) and in EBZs (right).

Figure 3

QRS duration during normal and premature stimulation. (A) Measurements of QRS duration during stimulation from the paraseptal region (PS; left) and from the EBZ/injection region (right). Electrical stimulation was applied as described in Section 2. Some animals could not be included in this analysis because they fibrillated before completion of the protocol (PS: Ad-GFP n= 18, Ad-SkM1 n= 10, Ad-Cx32 n= 11, and Ad-SkM1/Cx32 n= 8; EBZ: Ad-GFP n= 9, Ad-SkM1 n= 10, Ad-Cx32 n= 8, and Ad-SkM1/Cx32 n= 5). *P< 0.05.

Figure 4

Incidence and phenotype of induced VT. (A) Typical examples of the phenotypic response to PES. (B) Percentage of inducible VT/VF (left panel) and percentage of polymorphic VT/VF vs. monomorphic VT (right panel). Absolute numbers of animals with VT/VF and total number of animals studied; and the number of animals with monomorphic VT and the total number of animals with VT/VF are presented within the associated bars. *P< 0.05 vs. Ad-GFP; ⁺P< 0.05 vs. Ad-SkM1.

Figure 5

Response to capture and slowing of pacing rate vs. infarct size. (A) Induction of a monomorphic VT in a Cx32-treated animal (left), initiation of electrical pacing (middle), and reemergence of sinus rhythm (right) which occurred after slowing the rate of electronic pacing. (B) Summary data on infarct sizes. (C) Subanalysis of infarct sizes in animals with and without inducible VT/VF and polymorphic VT/VF vs. monomorphic VT. Shown are the analyses of all animals (left) or the group of GFP-treated animals (right). The number of animals tested within the other groups was too low to allow for a meaningful breakdown and/or statistical analysis (data not shown). *P< 0.05 vs. Ad-GFP; ⁺P< 0.05 vs. no VT/VF; ^†P< 0.05 vs. monomorphic VT.

Figure 6

(Upper) Confirmation of functional presence of Cx32. (A) Representative isochronal maps of EBZ obtained from animals injected with Ad-GFP or Ad-Cx32. As per Section 2, normal pH is 7.4 and low pH is 6.0. (B) Summary data on conduction velocity (CV) indicating faster impulse propagation in the Ad-Cx32 group (n= 14) vs. the Ad-GFP group (n= 10)—both in normal and low pH; *P< 0.05. (C) Summary data on AP parameters. RP, resting potential; APD, AP duration. (Lower) Comparison of SkM1 and SkM1/Cx32 effects to speed conduction and increase V_max. (D) Typical isochronal maps of EBZ obtained from animals injected with Ad-GFP, Ad-SkM1, or Ad-SkM1/Cx32. In this protocol, extracellular K⁺ was increased from 4 to 7 mmol/L (see Section 2). (E) Summary data on CV. (F) Summary data on AP parameters; note that all groups significantly depolarized upon application of high K⁺. (G) Higher membrane responsiveness (V_max vs. MP) curves in Ad-SkM1 and Ad-SkM1/Cx32-injected preparations vs. Ad-GFP. Ad-GFP, n= 12; Ad-SkM1, n= 8; Ad-SkM1/Cx32, n= 10. ^†P< 0.05 vs. K⁺ = 4 mM/L; *P< 0.05 vs. Ad-GFP.