MicroRNA miR-133a-3p Facilitates Adrenergic Proarrhythmic Ectopy in Rat Pulmonary Vein Myocardium by Increasing cAMP Content.

Cardiac-specific microRNA miR-133a-3p modulates adrenergic signaling. Adrenergic receptors and their intracellular pathways are the key players in proarrhythmic ectopy derived from the myocardial sleeves of the pulmonary veins. We studied the effect of miR-133a-3p on ectopy induced by norepinephrine in myocardial tissue of rat pulmonary veins. Using microelectrode technique, we revealed facilitation of proarrhythmic pattern of spontaneous bursts of action potentials induced by norepinephrine in tissue preparations of the pulmonary veins isolated from rats in 24 h after injection of a transfection mixture containing miR-133a-3p (1 mg/kg) in vivo. According to ELISA data, the cAMP level in the pulmonary vein myocardium of rats receiving miR-133a-3p was 2-fold higher than in control animals. Bioinformatic analysis showed that mRNA of protein phosphatases and some phosphodiesterases are most probable targets of miR-133a-3p. The proarrhythmic effect of miR-133a-3p can be related to inhibition of the expression of phosphodiesterases accompanied by cAMP accumulation and increased intracellular ß-adrenergic signaling.

Role of Muscarinic M1, M2, and M3 Receptors in the Regulation of Electrical Activity of Myocardial Tissue of Caval Veins during the Early Postnatal Ontogeny.

We studied the influence of blockers of muscarinic M1, M2, and M3 receptors on the effect of acetylcholine in the myocardial tissue of caval veins in rats at the early stage of ontogeny. The experiments were performed on isolated preparations of the right superior vena cava working under their own rhythm. Action potentials were recorded using the standard microelectrode technique. Acetylcholine (1 µM) suppressed automatic activity in the superior vena cava myocardium. Preliminary perfusion of the preparation with non-selective blocker atropine (1 µM) completely abolished the effect of acetylcholine, treatment with M2 receptor blocker AQ-RA 741 (1 µM) led to partial suppression of the effect of acetylcholine. Blockers of M1 and M3 receptors pirenzepine (1 µM) and 4DAMP (0.1 µM) did not suppress the effect of acetylcholine. Thus, the effect of acetylcholine is predominantly realized via M2 receptors, but M3 receptors can also partially mediate its effect in the superior vena cava myocardium in rats at the early stages of ontogeny.

Extracellular NAD+ Suppresses Adrenergic Effects in the Atrial Myocardium of Rats during the Early Postnatal Ontogeny.

The effects of sympathetic cotransmitter NAD+ (10 µM) on bioelectric activity of the heart under conditions of adrenergic stimulation were studied on isolated spontaneously contracting preparations (without stimulation) of the right atrium from 2-7-day-old rats. Action potentials were recorded in the working myocardium using standard microelectrode technique. Perfusion of the right atrium with norepinephrine solution (1 µM) altered the configuration and significantly lengthened the action potentials. NAD + against the background of norepinephrine stimulation significantly decreased the duration of action potentials, in particular, at 25% repolarization. The effect of purine compounds NAD + , ATP, and adenosine on bioelectrical activity of the heart of newborn rats was studied under basal conditions (without norepinephrine stimulation). The effect of NAD + against the background of adrenergic stimulation was more pronounced than under basal conditions and was probably determined by suppression of ICaL, which can be the main mechanism of NAD + action on rat heart.

Diadenosine Polyphosphates Suppress the Effects of Sympathetic Nerve Stimulation in Rabbit Heart Pacemaker.

The modulatory influence of diadenosine tetraphosphate (Ap4A) and diadenosine pentaphosphate (Ap5A) on the effect of intramural autonomic nerve stimulation in isolated rabbit sinoatrial node were examined. Electrical activity of the sinoatrial node was recorded intracellularly. Against the background of blockade of adrenergic effects with propranolol (3×10-6 M) or in preparations isolated 2 h after injection of reserpine (2 mg/kg), nerve stimulation induced short-term membrane hyperpolarization and diminished the sinus node firing rate. These phenomena were not affected by Ap4A or Ap5A (10-5 M). Under the action of atropine (3×10-6 M) that completely eliminated the cholinergic influences, nerve stimulation enhanced the sinus node firing rate by 17.30±3.45% from the initial rate. Both Ap4A and Ap5A moderated the stimulation-induced elevation of firing rate to 9.9±2.8 and 10.5±2.9%, respectively. The data suggest that diadenosine polyphosphates significantly modulate the sympathetic influences on the heart rhythm, but have no effect on the parasympathetic control over activity of sinoatrial node.

Effect of Purine Co-Transmitters on Automatic Activity Caused by Norepinephrine in Myocardial Sleeves of Pulmonary Veins.

We studied the effect of extracellular purine nucleotides (NAD+ and ATP) on spontaneous arrhythmogenic activity caused by norepinephrine in myocardial sleeves of pulmonary veins. In pulmonary veins, NAD+ and ATP reduced the frequency of action potentials and their duration at regular type of spontaneous activity caused by norepinephrine. NAD+ and ATP lengthened the intervals between spike bursts at periodic (burst) type of spontaneous activity. In addition, ATP shortened the duration of spike bursts and the number of action potentials in the "bursts" caused by norepinephrine in the pulmonary veins. It was hypothesized that NAD+ and ATP attenuate the effects of sympathetic stimulation and when released together with norepinephrine from sympathetic endings in vivo, probably, reduce arrhythmogenic activity in myocardial sleeves of pulmonary veins.

Effect of Exogenous Extracellular Nicotinamide Adenine Dinucleotide (NAD⁺) on Bioelectric Activity of the Pacemaker and Conduction System of the Heart.

In rat sinoatrial node, NAD(+) (10 µM) reduced the rate of spontaneous action potentials, duration of action potentials, and the velocity of slow diastolic depolarization, but the rate of action potential front propagation increases. In passed rabbit Purkinje fibers, NAD(+) (10 µM) reduced the duration of action potentials. Under conditions of spontaneous activity of Purkinje fibers, NAD(+) reduced the fi ring rate and the rate of slow diastolic depolarization. The effects of extracellular NAD(+) on bioelectric activity of the pacemaker (sinoatrial node) and conduction system of the heart (Purkinje fibers) are probably related to activation of P1 and P2 purinoceptors.

[Influence exogenous nicotinamide adenine dinucleotide (NAD+) on contractile and bioelectric activity of the rat heart].

This study is aimed to the investigation of the nicotinamide adenine dinucleotide (NAD+) effects and mechanisms of action in a heart. NAD+ (mcM) induces multiphase alternation of contractile activity of isolated rat heart: short positive inotropic action is followed by a negative inotropic phase. NAD+ (1-100 mcM) induces decreasing of action potential duration (APD) in rat atrial myocardium (from 45 +/- 0.82 ms in control experiments to 39 +/- 1.05 (n = 8) and 32 +/- 2 (n = 8) during application of 10 and 100 mcM of NAD+, respectively). Significant APD increase (from 45 +/- 0.82 ms to 74 +/- 1.89 (n = 8) ms) was observed during washing out of NAD+ (100 mcM). ATP or adenosine was unable to increase APD both during application or washing out. NAD+ induced APD decrease was not suppressed by P1-antagonist theophylline. P1-purinoreceptor and metabolite independent direct action of NAD+ in rat heart is suggested. Activation of P2X or P2Y receptors, cyclic ADP-ribose accumulation in cardiomyocytes is proposed as a main mechanism of NAD(+)-induced effects in the heart.

[Hypothermia induced alteration of refractoriness in the ventricular myocardium of ground souirrel Citellus undulatus].

Bioelectrical activity and refractoriness in ventricular myocardium of the hibernator--ground squirrel Citellus undulatus were investigated during hypothermia. Experiments were performed with use of isolated, perfused preparations of papillary muscle from right ventricular. Preparations were obtained from hibernating (HS), summer active (SAS) squirrels and from rats. Bioelectrical activity was registered using the standard microelectrode technique at 37-17 degrees C. Action potentials duration (APD), refractoriness duration (RD) and the velocity of the action potential wave front (dV/dt) were estimated. Hypothermia induced APD and RD prolongation were demonstrated in all groups of experimental animals. However, normalized RD was significantly longer in the HS group during hypothermia than in SAS and rats. Ratio of RD to APD in HS group exceeds unity at 17 degrees C, which allows to suggest so called "postrepolarization refractoriness" during hypothermia. Also, HS reveal more prominent preservation of dV/dt during hypothermia than SAS and rat. Significant prolongation of RD and maintenance of normal excitation conduction during hypothermia probably plays essential role in hibernators resistivity to cold induced arrhythmias.

[Influence of new III class antiarrhythmic drug Niferidil (RG-2) on bioelectrical activity of rat pulmonary veins myocardium].

Myocardial cells in pulmonary veins are thought to play a major role in the initiation and maintenance of atrial arrhythmias, including atrial fibrillation. In experiments on rat pulmonary veins, antiarrhythmic drug niferidil (RG-2) produced increase of APD90% and functional refractory period and decrease of action potential amplitude and slope of APD restitution curve. Thus niferidil (RG-2) exerts stronger action on pulmonary veins than left atrium. This can take important part in Niferidil (RG-2) antiarrhythmic activity.

[Excitation conduction in the rat pulmonary veins myocardium assessed by the optical mapping].

In this study, conduction of excitation in the rat left atrial and pulmonary veins myocardium was assessed by the optical mapping technique. Rat atrial myocardium and pulmonary vein myocardium demonstrated weak differences in the activation time and conduction velocity. Maximal conduction velocity in atrial myocardium estimated by the optical mapping was 84 +/- 14 cm/s, the same parameter in pulmonary vein myocardium was 71 +/- 11 cm/s. Period of refractoriness in the atria was significantly shorter (44 +/- 3 ms) than in pulmonary vein myocardium (60 +/- 3 ms). Despite the differences in the period of refractoriness excitation wavelength in the pulmonary veins and atrial myocardium was very similar (43 +/- 7 and 37 +/- 5 mm, respectively). Conduction of excitation in rat atria was characterized by high level of "linearity". Regions with essential decrease of conduction velocity and complete abruption of conduction (conduction blocks) were observed in pulmonary vein myocardium. It was suggested that high histological heterogeneity and intercellular electrical resistance played an important role in conduction abnormalities in rat pulmonary veins.

[Ionic mechanisms of action of class III antiarrhythmic drugs].

In this review we discuss mechanisms of antiarrhythmic and adverse proarrhythmic action of class III drugs. Special attention is given to ionic currents and channels which determine specific features of their effects (IKr, IKa, IKur). We consider general patterns of changes of bioelectrical activity in atria and ventricles leading to development of arrhythmias or stabilization of rhythm. We also discuss value of QT interval as predictor of torsade de pointes. Perspectives and limitations of development of novel class III antiarrhythmic drugs are discussed as well. We present consideration of efficacy and mechanisms of action of such compounds as dronedarone and vernacalant suggested for termination of atrial fibrillation and maintenance of sinus rhythm. Special attention is given to RG 2 - a novel compound with class III activity.

[Investigation of pacemaker shift in the rabbit sinoatrial node using the optical mapping technique].

Changes of the activation sequence in the rabbit sinoatrial node under the influence of low temperature and I(f) selective blocker ivabradine have been studied using the optical mapping technique. Both factors caused a shift of the pacemaker within the sinoatrial node region. These results are compared with the data obtained recently in the investigation of pacemaker shift under the influence of cholinergic and adrenergic factors. Possible mechanisms of the pacemaker shift are discussed. The suppression of electric activity in the central part of the sinoatrial node during the action of acetylcholine, which is called cholinergic inexcitability, may be considered as one of the mechanisms of the pacemaker shift. It is shown that the main cause of cholinergic inexcitability is the activation of potassium acetylcholine-dependent current I(KACh).

[Modern conceptions of the atrial fibrillation development. The role of the myocardial sleeves in the pulmonary veins].

Atrial fibrillation (AF) is the most common supraventricular cardiac arrhythmia. In this review several conceptions focused on the mechanisms of the AF initiation are discussed. At present time viewpoint that the ectopical activity in the pulmonary vien myocardial sleeves (PVs) account for AF in prevailing. PVs myocardium has been the subject of many anatomical and physiological investigations. PVs myocardium differs from left atria tissue and has many moprhological properties that make in convenient substrate for AF initiation and maintenance. PVs cardiomyocytes were shown to have great variability of electrophysiological properties (action potential duration, resting potential, upstroke velocity, etc.). Attempt to discuss afterdepolarization, triggered activity and abnormal automaticity as initiators of AF in PVs was made. It was shown that as in experimental condition, as in vivo in PVs can exist er-entry. Possibly, re-entry from PVs could be the one mechanism by which AF is initiated. In review big attention to the innervations of PVs and role of the sympathetic and the parasympathetic nerves in PVs ectopical activity is paid. Combined influence of autonomic nerves may be critical to initiating AF in PVs. Pharmacological intervention as a possible way to suppress or prevent the activity in the PVs that leads to AF is discussed.

[Changes in the activation sequence in the rabbit sinoatrial node induced by adrenergic stimulation].

We used high resolution optical mapping for the study of changes of activation sequence of a rabbit sinoatrial node induced by adrenergic stimulation during natural and paced rhythm. Activation of adrenoreceptors with isoproterenol (10, 100 nanoM, 1 microM) as well as stimulation of intramural postganglionic sympathetic nerves caused pacemaker migration within sinoatrial node and increase of the rate of generation of excitation. Pacemaker migration in cases of pronounced acceleration of rhythm could proceed in two stages. Termination of adrenergic influences has been followed by restoration of initial chronotopography of excitation of sinoatrial node.

[The cholinergic non-excitability phenomenon in the atrial myocardium of lower vertebrates].

Changes of electric activity induced by acetylcholine were studied in atrial myocardium of fishes (cod and carp) and reptilians (lizard and grass-snake). Standart microelectrode technique and novel method of optical mapping were used in the study. Acetylcholine (1-50 microM) provoked decrease of the action potential amplitude down to full inhibition of electrical activity in wide regions of atrium of cod and carp. We define this phenomenon as cholinergic inexcitability. In other regions excitation persisted even during action of 500 microM acetylcholine. In atria of lizard and grass-snake acetylcholine caused shortening of action potential without changes in it's amplitude. Local cholinergic inexcitability, shown in the atrial myocardium of fishes, is quite similar to the phenomenon, that was described earlier in the atria of frogs. It presents the heart of fish as an interesting model for study of mechanisms of cholinergic atrial arrhythmias initiation.

[Changes of activation sequence in a rabbit sinoatrial node induced by cholinergic influences].

We used high resolution optical mapping for the study of activation sequence in a rabbit sinoatrial node in normal state and during artificial stimulation as well as changes of excitation chronotopography under cholinergic influences. In the norm excitation originates in central region of sinoatrial node. During acetylcholine action (10, 50, 100 M) and activation of intramural parasympathetic nerves suppression of electrical activity in the zone of initial pacemaker localization occurs. Suppression of activation is associated with pacemaker migration within sinoatrial node and lowering of the rate of generation of excitation. Termination of cholinergic influences has been followed by restoration of initial chronotopography of excitation. During artificial stimulation of sinoatrial node nonexitable (refractory) area has been formed in the region of initial localization of the natural pacemaker.

[Influence of ADP-ribose, AMP and adenosine on bioelectric activity of hibernating ground squirrel atrium and papillary muscle].

The aim of work was to investigate effects of adenosine, AMP and ADP-ribose (1x10(-5)) on bioelectric activity of atrium and papillary muscle of nonhibernating (rat) and hibernating (Yakutian ground squirrel) animals. Action potential (AP) was registered with use of standard microelectrode technique. AP duration (APD) at level of 90% repolarisation in rat atrium in control experiments was 30+/-5 ms, APD at level of 50% repolarisation was 12+/-2 ms. APD at level of 90% repolarisation in rat papillary muscle was 56+/-7 ms, at level of 50% repolarisation was 18+/-2 ms. APD at level of 90% repolarisation in ground squirrel atrium was 77+/-6, APD at level of 50% repolarisation was 38+/-6 ms. APD at level of 90% repolarisation in ground squirrel papillary muscle was 105+/-9 ms, APD at level of 50% repolarisation was 42+/-8 ms. Purine nucleotides and nucleoside, that were tested in work, except ADP-ribose, act as inhibitory factors and decrease APD both in rat and hibernating ground squirrel heart. ADP-ribose decreases APD in papillary muscle of hibernator but did not in its atrium. In ground squirrel atrium AMP and adenosine decrease APD at level of 50% repolarisation by 10+/-3% and 18+/-3% respectively. AMP and adenosine decrease APD at level of 90% repolarisation by 9+/-2% and 11+/-2% respectively. In ground squirrel papillary muscle ADP-ribose, AMP and adenosine decrease APD at level of 50% repolarisation by 26+/-8%, 23+/-8% and 26+/-7%. ADP-ribose, AMP and adenosine decrease APD at level of 90% repolarisation by 12+/-3%, 10+/-3%, 13+/-3%. Thus, decrease of APD in ground squirrel papillary muscle at level of 90% repolarisation during nucleotides and adenosine action was 2-2.5 fold less, than the rat.

[The influence of purine nucleotides and adenosine on bioelectrical activity of bat (Pipistrellus nathusii) heart].

The aim of work was to investigate effects of adenosine, AMP, GMP and ADP-ribose on bioelectric activity of bat heart. Purine nucleotides decreased action potential duration at level of 90% (APD90) repolarization in bat ventricular myocardium. When preparation of right ventricle was paced with frequency of 6 Hz, APD50 and APD90 were 7 +/- 2 ms and 60 +/- 5 ms, respectively. Adenosine decreased APD90 by 50 +/- 10% (n=6), AMP - by 30 +/- 10% (n=6), GMP - by 38 +/- 5% (n=6), ADP-ribose - by 24 +/- 6% (n=6) (concentration of all compounds - 10 ). Effects of purine nucleotides and nucleoside in bat ventricular myocardium exceed effects of same compounds in rat and ground squirrel ventricular myocardium. Inhibitory effects of purine nucleotides and adenosine in bat heart could be mediated by A1 adenosine receptors.