Abstract

Blebbistatin (BS) is a recently discovered inhibitor of the myosin II isoform and has been adopted as the mechanical uncoupler of choice for optical mapping, because previous studies suggest that BS has no significant cardiac electrophysiological effects in a number of species. The aim of this study was to determine whether BS affects cardiac electrophysiology in isolated New Zealand White rabbit hearts. Langendorff-perfused hearts (n= 39) in constant-flow mode had left ventricular monophasic action potential duration (MAPD) measured at apical and basal regions during constant pacing (300 ms cycle length). Standard action potential duration restitution was obtained using the single extrastimulus method with measurement of the maximal restitution slope. Ventricular fibrillation threshold was measured as the minimal current inducing sustained ventricular fibrillation with burst pacing (30 stimuli, at 30 ms intervals). Optical action potentials were recorded using the voltage-sensitive dye di-4-ANEPPS. Measurements were taken at baseline and after 60 min perfusion with BS (5 μm). Blebbistatin significantly prolonged left ventricular apical (mean ± SEM; from 129.9 ± 2.9 to 170.7 ± 4.1 ms, P < 0.001, n= 8) and basal MAPD (from 135.0 ± 2.3 to 163.3 ± 5.6 ms, P < 0.001) and effective refractory period (from 141.3 ± 4.8 to 175.6 ± 3.7 ms, P < 0.001) whilst increasing the maximal slope of restitution (apex, from 0.79 ± 0.09 to 1.57 ± 0.16, P < 0.001; and base, from 0.71 ± 0.06 to 1.44 ± 0.24, P < 0.001) and ventricular fibrillation threshold (from 5.3 ± 1.1 to 17.0 ± 2.9 mA, P < 0.001). In other hearts, blebbistatin significantly prolonged optically recorded action potentials (from 136.5 ± 6.3 to 173.0 ± 7.9 ms, P < 0.05, n= 4). In control experiments, the increase of MAPD with blebbistatin was present whether the hearts were perfused in constant-pressure mode (n= 5) or in unloaded conditions (n= 5). These data show that blebbistatin significantly affects cardiac electrophysiology. Its use in optical mapping studies should be treated with caution.

Highlights

  • Left ventricular pressure slowly decreased over a 5 h period by ∼15%, whilst Perfusion pressure (PP) and heart rate (HR; from 152.9 ± 9.5 to 137.7 ± 7.3 beats min−1, P > 0.05) did not change significantly

  • Blebbistatin led to a gradual decrease in Left ventricular pressure (LVP), and left ventricular developed pressure was abolished within 20 min

  • After 60 min perfusion with blebbistatin, systolic LVP decreased significantly from a baseline of 88.0 ± 6.3 to 18.8 ± 2.9 mmHg (P < 0.001, n = 8), whilst end-diastolic pressure increased from 4.8 ± 0.6 to 18.4 ± 2.9 mmHg (P < 0.001, n = 8)

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Summary

Objectives

The aim of this study was to determine whether BS affects cardiac electrophysiology in isolated New Zealand White rabbit hearts. The aim of the present study was to use non-optical mapping techniques to measure the effects of blebbistatin on ventricular electrophysiological properties in isolated, Langendorff-perfused rabbit hearts

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