thiourea and indo-1

Pulmonary veins (PVs) are the most important focus for generation of atrial fibrillation. The Na(+)/Ca(2+) exchange (NCX) current is important in PV electrical activity and cardiac glycosides-induced arrhythmias. The purpose of this study was to investigate whether KB-R7943, a NCX current blocker with preferential inhibition of the Ca(2+) influx, may alter PV electrophysiological characteristics and reduce glycoside-induced arrhythmogenicity.. Conventional microelectrodes were used to record the effects of KB-R7943 on action potentials and contractility in isolated rabbit PV tissue specimens with and without administration of ouabain. The ionic currents and intracellular calcium were studied in isolated single cardiomyocytes before and after KB-R7943 by the whole-cell patch clamp and indo-1 fluorimetric ratio techniques.. KB-R7943 (0, 3, 10, 30 microM) concentration-dependently prolonged APD(50) and APD(90) and decreased the PV firing rates (2.3 +/- 1.2 Hz, 2.1 +/- 1.2 Hz, 1.9 +/- 0.9 Hz, 1.7 +/- 1.1 Hz, n = 7, p < 0.05) and incidences of delayed afterdepolarizations (DADs). KB-R7943 (3, 30 microM) decreased transient inward currents, Ca(2+) transient and sarcoplasmic reticulum Ca(2+) content. Ouabain (0, 0.1, 1 microM) concentration-dependently increased the PV firing rates and DADs in PVs with spontaneous activity (n = 7) and induced nonsustained spontaneous activity (1 microM) in the PVs without spontaneous activity (n = 14). However, in the presence of KB-R7943 (30 microM), ouabain (1 microM) did not increase the PV firing rates or induce spontaneous activity in the PVs without spontaneous activity (n = 7).. KB-R7943 reduces the PV arrhythmogenic activity and prevents the ouabain-induced arrhythmogenicity. Our findings support the role of the NCX current in the PV electrical activity.

Topics: Action Potentials; Animals; Atrial Fibrillation; Calcium; Dose-Response Relationship, Drug; Fluorescent Dyes; Fluorometry; Indoles; Muscle, Smooth, Vascular; Myocytes, Cardiac; Ouabain; Patch-Clamp Techniques; Pulmonary Veins; Rabbits; Sarcoplasmic Reticulum; Sodium-Calcium Exchanger; Thiourea

Nitric oxide (NO) generated by skeletal muscle is believed to regulate force production but how this is achieved remains poorly understood. In the present work we tested the effects of NO synthase (NOs) inhibitors on membrane current and intracellular calcium in isolated skeletal muscle fibres from mouse, under voltage-clamp conditions. Resting [Ca(2+)] and [Ca(2+)] transients evoked by large depolarizations exhibited similar properties in control fibres and in fibres loaded with tenth millimolar levels of the NOs inhibitor N-nitro-L-arginine (L-NNA). Yet the voltage dependence of calcium release was found to be shifted by approximately 15 mV towards negative values in the presence of L-NNA. This effect could be reproduced by the other NOs inhibitor S-methyl-L-thiocitrulline (L-SMT). Separate experiments showed that the voltage dependence of charge movement and of the slow calcium current were unaffected by the presence of L-NNA, ruling out an effect on the voltage sensor. A negative shift in the voltage dependence of calcium release with no concurrent alteration in the properties of charge movement was also observed in fibres exposed to the oxidant H(2)O(2) (1 mM). Conversely the reducing agent dithiothreitol (10 mM) had no obvious effect on Ca(2+) release. Overall, the results indicate that physiological levels of NO exert a tonic inhibitory control on the activation of the calcium release channels. Changes in the voltage dependence of Ca(2+) release activation may be a ubiquitous physiological consequence of redox-related modifications of the ryanodine receptor.

Topics: Animals; Calcium; Citrulline; Enzyme Inhibitors; Hydrogen Peroxide; In Vitro Techniques; Indoles; Membrane Potentials; Mice; Mice, Inbred Strains; Muscle Fibers, Skeletal; Muscle, Skeletal; Nitric Oxide Synthase; Nitroarginine; Sarcoplasmic Reticulum; Thiourea

In most mammalian species, an increase in stimulation frequency (ISF) produces an increase in contractility (treppe phenomenon), which results from larger Ca2+ transients at higher frequencies, due to an increase in sarcoplasmic reticulum Ca2+ load and release. The present study attempts to elucidate the contribution of the Na(+)-Ca2+ exchanger (NCX) to this phenomenon. Isolated cat ventricular myocytes, loaded with [Ca2+]i- and [Na+]i-sensitive probes, were used to determine whether the contribution of the NCX to the positive inotropic effect of ISF is due to an increase in Ca2+ influx (reverse mode) and/or a decrease in Ca2+ efflux (forward mode) via the NCX, due to frequency-induced [Na+]i elevation, or whether it was due to the reduced time for the NCX to extrude Ca2+. The results showed that the positive intropic effect produced by ISF was temporally dissociated from the increase in [Na+]i and was not modified by KB-R7943 (1 or 5 microM), a specific blocker of the reverse mode of the NCX. Whereas the ISF from 10 to 30 beats min(-1) (bpm) did not affect the forward mode of the NCX (assessed by the time to half-relaxation of the caffeine-induced Ca2+ transient), the ISF to 50 bpm produced a significant reduction of the activity of the forward mode of the NCX, which occurred in association with an increase in [Na+]i (from 4.33+/-0.40 to 7.25+/-0.50 mM). However, both changes became significant well after the maximal positive inotropic effect had been reached. In contrast, the positive inotropic effect produced by ISF from 10 to 50 bpm was associated with an increase in diastolic [Ca2+]i, which occurred in spite of a significant increase in the relaxation rate and at a time at which no increases in [Na+]i were detected. The contribution of the NCX to stimulus frequency inotropy would therefore depend on a decrease in NCX-mediated Ca2+ efflux due to the reduced diastolic interval between beats and not on [Na+]i-dependent mechanisms.

Topics: Animals; Calcium; Calcium Signaling; Cardiotonic Agents; Cats; Cytosol; Electric Stimulation; Fluorescent Dyes; Heart Rate; In Vitro Techniques; Indoles; Myocardial Contraction; Ouabain; Papillary Muscles; Sodium; Sodium-Calcium Exchanger; Thiourea