diacetylmonoxime and Arrhythmias--Cardiac

Unlike other excitation-contraction uncouplers, blebbistatin has few electrophysiological side effects and has gained increasing acceptance as an excitation-contraction uncoupler in optical mapping experiments. However, the possible role of blebbistatin in ventricular arrhythmia has hitherto been unknown. Furthermore, experiments with blebbistatin and 2,3-butanedione monoxime (BDM) offer an opportunity to assess the contribution of dynamic instability and wavelength of impulse propagation to the induction and maintenance of ventricular arrhythmia. Recordings of monophasic action potentials were used to assess effects of blebbistatin in Langendorff-perfused rabbit hearts (n = 5). Additionally, panoramic optical mapping experiments were conducted in rabbit hearts (n = 7) that were sequentially perfused with BDM, then washed out, and subsequently perfused with blebbistatin. The susceptibility to arrhythmia was investigated using a shock-on-T protocol. We found that 1) application of blebbistatin did not change action potential duration (APD) restitution; 2) in contrast to blebbistatin, BDM flattened APD restitution curve and reduced the wavelength; and 3) incidence of sustained arrhythmia was much lower under blebbistatin than under BDM (2/123 vs. 23/99). While arrhythmias under BDM were able to stabilize, the arrhythmias under blebbistatin were unstable and terminated spontaneously. In conclusion, the lower susceptibility to arrhythmia under blebbistatin than under BDM indicates that blebbistatin has less effects on arrhythmia dynamics. A steep restitution slope under blebbistatin is associated with higher dynamic instability, manifested by the higher incidence of not only wave breaks but also wave extinctions. This relatively high dynamic instability leads to the self-termination of arrhythmia because of the sufficiently long wavelength under blebbistatin.

Topics: Action Potentials; Animals; Anti-Arrhythmia Agents; Arrhythmias, Cardiac; Cardiac Pacing, Artificial; Diacetyl; Disease Models, Animal; Excitation Contraction Coupling; Heart Conduction System; Heterocyclic Compounds, 4 or More Rings; Perfusion; Rabbits; Time Factors; Voltage-Sensitive Dye Imaging

Arrhythmias are benign or lethal, depending on their sustainability and frequency. To determine why lethal arrhythmias are prone to occur in diseased hearts, usually characterized by nonuniform muscle contraction, we investigated the effect of nonuniformity on sustainability and frequency of triggered arrhythmias.. Force, membrane potential, and intracellular Ca(2+) concentration ([Ca(2+)](i)) were measured in 51 rat ventricular trabeculae. Nonuniform contraction was produced by exposing a restricted region of muscle to a jet of 20 mmol/L 2,3-butanedione monoxime (BDM) or 20 mumol/L blebbistatin. Sustained arrhythmias (>10 seconds) could be induced by stimulus trains for 7.5 seconds only with the BDM or blebbistatin jet (100 nmol/L isoproterenol, 1.0 mmol/L [Ca(2+)](o), 24 degrees C). During sustained arrhythmias, Ca(2+) surges preceded synchronous increases in [Ca(2+)](i), whereas the stoppage of the BDM jet made the Ca(2+) surges unclear and arrested sustained arrhythmias (n=6). With 200 nmol/L isoproterenol, 2.5 mmol/L [Ca(2+)](o), and the BDM jet, lengthening or shortening of the muscle during sustained arrhythmias accelerated or decelerated their cycle in both the absence (n=10) and presence (n=10) of 100 mumol/L streptomycin, a stretch-activated channel blocker, respectively. The maximum rate of force relaxation correlated inversely with the change in cycle lengths (n=14; P<0.01). Sustained arrhythmias with the BDM jet were significantly accelerated by 30 mumol/L SCH00013, a Ca(2+) sensitizer of myofilaments (n=10).. These results suggest that nonuniformity of muscle contraction is an important determinant of the sustainability and frequency of triggered arrhythmias caused by the surge of Ca(2+) dissociated from myofilaments in cardiac muscle.

Topics: Actin Cytoskeleton; Animals; Arrhythmias, Cardiac; Calcium Signaling; Diacetyl; Heart; Heart Ventricles; Heterocyclic Compounds, 4 or More Rings; Membrane Potentials; Myocardial Contraction; Rats

We examined whether non-uniform muscle contraction affects delayed afterdepolarizations (DADs) by dissociating Ca(2+) from myofilaments within the border zone (BZ) between contracting and stretched regions.. Force, sarcomere length (SL), membrane potential, and [Ca(2+)](i) dynamics were measured in 31 ventricular trabeculae from rat hearts. Non-uniform muscle contraction was produced by exposing a restricted region of muscle to a jet of solution containing 20 mmol/L 2,3-butanedione monoxime (BDM). DADs were induced by 7.5 s-2 Hz stimulus trains at an SL of 2.0 microm (24 degrees C, [Ca(2+)](o) 2.0 mmol/L). The BDM jet enhanced DADs (n = 6, P < 0.05) and aftercontractions (n = 6, P < 0.05) with or without 100 micromol/L streptomycin and occasionally elicited an action potential. A stretch pulse from an SL of 2.0 microm to 2.1 or 2.2 microm during the last stimulated twitch of the trains accelerated Ca(2+) waves in proportion to the increment of force by the stretch (P < 0.01) with or without streptomycin. In the presence of 1 mmol/L caffeine, rapid shortening of the muscle after the stretch pulse increased [Ca(2+)](i) within the BZ, whose amplitude correlated with the increment of force by the stretch (n = 15, P < 0.01).. These results suggest that non-uniform muscle contraction can enhance DADs by dissociating Ca(2+) from myofilaments within the BZ and thereby cause triggered arrhythmias.

Topics: Animals; Arrhythmias, Cardiac; Calcium; Calcium Channel Blockers; Diacetyl; In Vitro Techniques; Membrane Potentials; Myocardial Contraction; Myocardium; Rats

Ca2+ waves underlying triggered propagated contractions (TPCs) are initiated in damaged regions in cardiac muscle and cause arrhythmias. We studied Ca2+ waves underlying TPCs in rat cardiac trabeculae under experimental conditions that simulate the functional nonuniformity caused by local mechanical or ischemic local damage of myocardium. A mechanical discontinuity along the trabeculae was created by exposing the preparation to a small jet of solution with a composition that reduces excitation-contraction coupling (ECC) in myocytes within that segment. The jet solution contained either caffeine (5 mmol/L), 2,3-butanedione monoxime (BDM; 20 mmol/L), or low Ca2+ concentration ([Ca2+]; 0.2 mmol/L). Force was measured with a silicon strain gauge and sarcomere length with laser diffraction techniques in 15 trabeculae. Simultaneously, [Ca2+]i was measured locally using epifluorescence of Fura-2. The jet of solution was applied perpendicularly to a small muscle region (200 to 300 microm) at constant flow. When the jet contained caffeine, BDM, or low [Ca2+], during the stimulated twitch, muscle-twitch force decreased and the sarcomeres in the exposed segment were stretched by shortening normal regions outside the jet. Typical protocols for TPC induction (7.5 s-2.5 Hz stimulus trains at 23 degrees C; [Ca2+]o=2.0 mmol/L) reproducibly generated Ca2+ waves that arose from the border between shortening and stretched regions. Such Ca2+ waves started during force-relaxation of the last stimulated twitch of the train and propagated (0.2 to 2.8 mm/sec) into segments both inside and outside of the jet. Arrhythmias, in the form of nondriven rhythmic activity, were induced when the amplitude of the Ca2+-wave was increased by raising [Ca2+]o. Arrhythmias disappeared rapidly when uniformity of ECC throughout the muscle was restored by turning the jet off. These results show, for the first time, that nonuniform ECC can cause Ca2+ waves underlying TPCs and suggest that Ca2+ dissociated from myofilaments plays an important role in the initiation of Ca2+ waves.

Topics: Animals; Arrhythmias, Cardiac; Caffeine; Calcium; Diacetyl; Heart; Myocardial Contraction; Myocytes, Cardiac; Rats; Rats, Inbred BN; Sarcomeres; Sarcoplasmic Reticulum

Chemical uncouplers diacetyl monoxime (DAM) and cytochalasin D (cyto-D) are used to abolish cardiac contractions in optical studies, yet alter intracellular Ca(2+) concentration ([Ca(2+)](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-[beta-[2-(di-n-butylamino)-6-naphthyl]vinyl]pyridinium (di-4-ANEPPS) to map [Ca(2+)](i) transients (emission wavelength = 585 +/- 20 nm) and action potentials (APs) (emission wavelength > 610 nm; excitation wavelength = 530 +/- 20 nm). Confocal images showed that rhod-2 is primarily in the cytosol. DAM (15 mM) and cyto-D (5 microM) increased AP durations (APD(75) = 20.0 +/- 3 to 46.6 +/- 5 ms and 39.9 +/- 8 ms, respectively, n = 4) and refractory periods (45.14 +/- 12.1 to 82.5 +/- 3.5 ms and 78 +/- 4.24 ms, respectively). Cyto-D reduced conduction velocity by 20% within 5 min and DAM by 10% gradually in 1 h (n = 5 each). Uncouplers did not alter the direction and gradient of repolarization, which progressed from apex to base in 15 +/- 3 ms. Peak systolic [Ca(2+)](i) increased with cyto-D from 743 +/- 47 (n = 8) to 944 +/- 17 nM (n = 3, P = 0.01) but decreased with DAM to 398 +/- 44 nM (n = 3, P < 0.01). Diastolic [Ca(2+)](i) was higher with cyto-D (544 +/- 80 nM, n = 3) and lower with DAM (224 +/- 31, n = 3) compared with controls (257 +/- 30 nM, n = 3). DAM prolonged [Ca(2+)](i) transients at 75% recovery (54.3 +/- 5 to 83.6 +/- 1.9 ms), whereas cyto-D had no effect (58.6 +/- 1.2 ms; n = 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.

Topics: Action Potentials; Animals; Arrhythmias, Cardiac; Calcium; Cytochalasin D; Diacetyl; Electrophysiology; Fluorescent Dyes; Heart; Heterocyclic Compounds, 3-Ring; In Vitro Techniques; Kinetics; Mice; Mice, Inbred Strains; Myocardial Contraction; Nucleic Acid Synthesis Inhibitors; Organ Preservation Solutions; Perfusion

Whether 2,3-butanedione monoxime (BDM, < or = 5mmol/l) suppresses primarily crossbridge cycling or total Ca(2+) handling in the blood-perfused whole heart remains controversial. Although BDM seems to suppress primarily total Ca(2+) handling in canine hearts, more evidence is lacking. We therefore analyzed the cardiac mechanoenergetics, namely, E(max) (contractility), PVA (total mechanical energy), and O(2) consumption of canine BDM-treated hearts by our recently developed integrative method to assess myocardial total Ca(2+) handling. This method additionally required the internal Ca(2+) recirculation fraction. We obtained this from the beat constant of the exponential decay component of the postextrasystolic potentiation. Our analysis indicated significant decreases in both internal Ca(2+) recirculation fraction and total Ca(2+) handling in the BDM-treated heart, but virtually no change in the reactivity of E(max) to total Ca(2+) handling. This result corroborates the view that BDM suppresses primarily total Ca(2+) handling rather than crossbridge cycling in the canine blood-perfused heart.

Topics: Animals; Arrhythmias, Cardiac; Calcium; Diacetyl; Dogs; Enzyme Inhibitors; In Vitro Techniques; Myocardial Contraction; Myocardium; Oxygen Consumption; Sarcoplasmic Reticulum; Ventricular Function, Left