Abstract
SummaryAcute RyR2 activation by exchange protein directly activated by cAMP (Epac) reversibly perturbs myocyte Ca2+ homeostasis, slows myocardial action potential conduction, and exerts pro‐arrhythmic effects. Loose patch‐clamp studies, preserving in vivo extracellular and intracellular conditions, investigated Na+ current in intact cardiomyocytes in murine atrial and ventricular preparations following Epac activation. Depolarising steps to varying test voltages activated typical voltage‐dependent Na+ currents. Plots of peak current against depolarisation from resting potential gave pretreatment maximum atrial and ventricular currents of −20.23 ± 1.48 (17) and −29.8 ± 2.4 (10) pA/μm2 (mean ± SEM [n]). Challenge by 8‐CPT (1 μmol/L) reduced these currents to −11.21 ± 0.91 (12) (P < .004) and −19.3 ± 1.6 (11) pA/μm2 (P < .04) respectively. Currents following further addition of the RyR2 inhibitor dantrolene (10 μmol/L) (−19.91 ± 2.84 (13) and −26.6 ± 1.7 (17)), and dantrolene whether alone (−19.53 ± 1.97 (8) and −27.6 ± 1.9 (14)) or combined with 8‐CPT (−19.93 ± 2.59 (12) and −29.9 ± 2.5(11)), were indistinguishable from pretreatment values (all P >> .05). Assessment of the inactivation that followed by applying subsequent steps to a fixed voltage 100 mV positive to resting potential gave concordant results. Half‐maximal inactivation voltages and steepness factors, and time constants for Na+ current recovery from inactivation in double‐pulse experiments, were similar through all the pharmacological conditions. Intracellular sharp microelectrode membrane potential recordings in intact Langendorff‐perfused preparations demonstrated concordant variations in maximum rates of atrial and ventricular action potential upstroke, (dV/dt)max. We thus demonstrate an acute, reversible, Na+ channel inhibition offering a possible mechanism for previously reported pro‐arrhythmic slowing of AP propagation following modifications of Ca2+ homeostasis, complementing earlier findings from chronic alterations in Ca2+ homeostasis in genetically‐modified RyR2‐P2328S hearts.
Highlights
Cardiac arrhythmias result from disruptions in the normal excitable activity propagating through successive structures in the heart
Reductions of Na+ current of the kind described above have been associated with corresponding reductions in maximum action potential upstroke rate,max in turn resulting in the pro-arrhythmic slowing of action potentials (AP) conduction[32] reported in previous explorations of effects of exchange protein directly activated by cAMP (Epac) modulation.[19]
In comparison with (i) untreated atrial (Figure 7) and ventricular cardiomyocytes (Figure 8), (ii) 8-CPT challenge reducedmax. This was reversed by further addition of (iii) dantrolene, which when applied by itself (iv) gavemax resembling those of untreated hearts
Summary
Cardiac arrhythmias result from disruptions in the normal excitable activity propagating through successive structures in the heart. Reductions of Na+ current of the kind described above have been associated with corresponding reductions in maximum action potential upstroke rate, (dV/dt)max in turn resulting in the pro-arrhythmic slowing of AP conduction[32] reported in previous explorations of effects of Epac modulation.[19] Figures 7 and 8 illustrate typical traces of atrial and ventricular action potential waveforms in response to regular a Control Significant differences in both atrial and ventricular (dV/dt)max values were detected amongst findings obtained in the absence (atria: 219.56 ± 9.64 V/s [n = 23]; ventricles: 174.11 ± 6.21 V/s [n = 17]), and following challenge with 8-CPT before (atria: 170.97 ± 9.78 V/s [n = 20]; ventricles: 128.64 ± 5.26 V/s [n = 14]), and following further addition of dantrolene (atria: 206.74 ± 10.47 V/s [n = 22]; ventricles: 170.90 ± 4.16 V/s [n = 20]) and of dantrolene applied alone (atria: 213.48 ± 6.39 V/s [n = 29]; ventricles: 189.70 ± 9.28 V/s [n = 20]) (atria: F = 5.41; P = .0018; ventricles: F = 12.98; P = 8.8 × 10−7).
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