Effects of mechanical uncouplers, diacetyl monoxime, and cytochalasin-D on the electrophysiology of perfused mouse hearts

Abstract

concentration ([Ca 2ϩ ]i) handling and vulnerability to arrhythmias in a species-dependent manner. The effects of uncouplers were investigated in perfused mouse hearts labeled with rhod-2/AM or 4-[␤-[2-(di-n-butylamino)-6-naphthyl]vinyl]pyridinium (di-4-ANEPPS) to map [Ca 2ϩ ]i transients (emission wavelength ϭ 585 Ϯ 20 nm) and action potentials (APs) (emission wavelength Ͼ 610 nm; excitation wavelength ϭ 530 Ϯ 20 nm). Confocal images showed that rhod-2 is primarily in the cytosol. DAM (15 mM) and cyto-D (5 M) increased AP durations (APD 75 ϭ 20.0 Ϯ 3 to 46.6 Ϯ 5 ms and 39.9 Ϯ 8 ms, respectively, n ϭ 4) and refractory periods (45.14 Ϯ 12.1 to 82.5 Ϯ 3.5 ms and 78 Ϯ 4.24 ms, respectively). Cyto-D reduced conduction velocity by 20% within 5 min and DAM by 10% gradually in 1 h (n ϭ 5 each). Uncouplers did not alter the direction and gradient of repolarization, which progressed from apex to base in 15 Ϯ 3 ms. Peak systolic [Ca 2ϩ ]i increased with cyto-D from 743 Ϯ 47 (n ϭ 8) to 944 Ϯ 17 nM (n ϭ 3, P ϭ 0.01) but decreased with DAM to 398 Ϯ 44 nM (n ϭ 3, P Ͻ 0.01). Diastolic [Ca 2ϩ ]i was higher with cyto-D (544 Ϯ 80 nM, n ϭ 3) and lower with DAM (224 Ϯ 31, n ϭ 3) compared with controls (257 Ϯ 30 nM, n ϭ 3). DAM prolonged [Ca 2ϩ ]i transients at 75% recovery (54.3 Ϯ 5 to 83.6 Ϯ 1.9 ms), whereas cyto-D had no effect (58.6 Ϯ 1.2 ms; n ϭ 3). Burst pacing routinely elicited long-lasting ventricular tachycardia but not fibrillation. Uncouplers flattened the slope of AP restitution kinetic curves and blocked ventricular tachycardia induced by burst pacing. optical action potentials; action potentials; intracellular calcium; restitution kinetics MOLECULARLY ENGINEERED MICE have been extensively used to genetically alter a specific component of a complex signaling process and to develop models of human diseases. Transgenic mice are used as models for various cardiac diseases and offer an effective strategy to elucidate the mechanisms underlying long QT-related arrhythmias, metabolic diseases, and the pathology of heart failure (29). A limitation of mouse models is the small size of the heart, making it difficult to study changes in contractility, electrophysiology, and vulnerability to arrhythmias in intact hearts. The challenge of measuring changes in cardiac phenotype has been partly overcome by applying optical technique to map electrical activity, but a major technical difficulty in the application of optical techniques to measure action potentials (APs) and intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) transients has been the distortion of the signals by movement due to muscle contractions.Several approaches have been used to reduce movement artifacts: 1) perfusion in Ca 2ϩ -free Tyrode solution to abolish contractions, an approach applicable to amphibian hearts (38); 2) design perfusion chambers to mechanically stabilize the heart (18, 37); 3) perfusion with an inhibitor of L-type voltage- Chemical uncouplers can potentially provide a practical approach to block movement artifacts and have been used to...

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Imaging cellular signals in the heart in vivo : Cardiac expression of the high-signal Ca 2+ indicator GCaMP2

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