Effect of diethylamine analog of ethmozine on parameters of sodium current in isolated rat cardiomyocytes

Voltage clamp experiments were made on ezymically isolated and internally perfused rat cardiac cells. The effect of a diethylamine analog of ethmozine (DAAE) on sodium current (INa) was tested when the drug was applied inside or outside the cell. It was found that the effect of DAAE (8 X 10(-6) g/ml) on INa was asymmetrical: after DAAE addition outside the cell, the amplitude of INa was effectively suppressed. Thus, 5 minutes after DAAE action the maximal value of INa in a voltage-current relationship was 20% of the control value without significant changes in the kinetics of INa. When the DAAE was added inside the cell preferentially, the inactivation time constant was increased without significant changes in the amplitude of the maximal INa. The same results were obtained with pronase (1 mg/ml) added inside the cell. It was supposed that as compared to ethmozine, the DAAE possesses a supplementary binding site on the cardiac cell membrane possibly linked to the structures responsible for inactivation processes.     

Effect of ethmozine on action potentials and myocardial contraction in guinea pigs

Ethmozine decreased the maximum rate of action potential rise (Vmax) in a dose-dependent manner. Using the Scatchard plot the apparent dissociation constant was calculated to be 1.52 X 10(-5) g/ml. Ethmozine also decreased the force of contraction in the concentration range between 1 X 10(-6) and 1 X 10(-4) g/ml with the apparent dissociation constant obtained from the Scatchard plot being equal to 1.48 X 10(-5) g/ml. The linear correlation coefficient between the decrease in Vmax and the decrease in the force of contraction was found to be equal to 0.998. Negative inotropic action of ethmozine was less pronounced when the stimulation frequency had been switched from 0.8 to 0.1 Hz. The decrease in Vmax under the action of ethmozine (3 X 10(-5) g/ml) was diminished from 56 +/- 7% (0.8 Hz) to only 3 +/- 8% (0.1 Hz). This was accompanied by the decrease in the negative inotropic effect: from 58 +/- 9% (0.8 Hz) to 16 +/- 15% (0.1 Hz). It was assumed that the negative inotropic action of ethmozine was mediated by the Na--Ca exchange, which was inhibited by the decrease of the intracellular Na+ concentration due to the blockade of sodium channels by ethmozine.     

Effect of tetrodotoxin and ethmozine on arrhythmias of a dog heart isolated in the late stage of experimental myocardial infarct

Dog hearts with ventricular extrasystole that developed 24 hours after coronary artery occlusion were isolated and perfused with blood from support dogs. After heart isolation the rhythm disturbances persisted regardless the decreased frequency of the ventricular beats. Administration of tetrodotoxin (4 X 10-8--10-7 g/ml) and ethmozine (3--5X X10-5 g/ml) abolished ventricular arrhythmias and restored the sinus rhythm. Potential mechanisms of the increased susceptibility of ischemic myocardial fibers to tetrodotoxin and antiarrhythmic drugs are discussed.     

Frequency-dependent blockade of sodium channels in isolated rat myocardial cells by the anti-arrhythmia agent ethmozine

A patch-clamp method was used to study the effects of the phenotiazine antiarrhythmic drug ethmozine (E) on the fast sodium inward current (INa) in freshly isolated heart muscle cells of adult rats. At a concentration of 10(-5) M E caused INa inhibition that could be enhanced by increasing the frequency of depolarization. This inhibition was reversible. After the termination of repetitive depolarization the amplitude of INa recovered with a time constant of about 10 sec. These findings may help to explain the therapeutic efficiency of E in high frequency cardiac rhythm disturbances.     

Influence of ethacizin (the diethyl analog of ethmozine) on phase-dependent parasympathetic effects on the heart

The influence of ethacizin (a diethylamine analog of ethmozine) (1.10(-7)-1.10(-6) g/ml) upon the phase-dependent chronotropic parasympathetic effects was studied on the perfused frog heart. The vagolytic influence of ethacizin (5.10(-7) and 1.10(-6) g/ml) was detected; the concentration of 1.10(-7) g/ml was found ineffective. The vagolytic effect consisted of a decreased maximum of phase-dependent effect, reduced latency and time required for the manifestation of the maximum increase. The period of inhibitory vagal stimulus effectiveness did not change significantly.     

Stimulus-dependent blockade of sodium channels of the Ranvier node membrane by the phenothiazine derivative ethmozine

The action of the antiarrhythmic drug ethmozine on sodium channels of the membrane was studied in experiments on single from Ranvier nodes by the voltage clamp method. Application of ethmozine to both the outer and the inner side of the membrane reduced the amplitude of the sodium current INa; the kinetics of this current and steady-state inactivation of the sodium channels were unchanged. Tonic and phasic (transient, stimulus-dependent) components can be distinguished in the ethmozine block of the sodium current. Tonic blockage of the sodium current develops slowly and can be potentiated by high-frequency stimulation of the membrane. The possible nature of the tonic block is discussed. The stimulus-dependent blockade of the sodium current deepens with an increase in the frequency and amplitude of depolarizing stimuli. Prolonged membrane depolarization does not evoke any additional blocking of the sodium current. It is concluded that the stimulus-dependent blockade is due to interaction between ethomizine and open sodium channels. Modification of the channels by batrachotoxin (preventing inactivation of the sodium channels) makes them insensitive to ethmozine. Increasing the potassium ion concentration on the outer side of the membrane was found to reduce the tonic effect of ethmozine and to potentiate the stimulus-dependent blockade. The action of ethmozine was compared with the effects of tertiary and quaternary local anesthetics.A. V. Vishnevskii Institute of Surgery, Academy of Medical Sciences of the USSR, Moscow. Translated from Neirofiziologiya, Vol. 13, No. 4, pp. 380–389, July–August, 1981.     

Comparison of the electrophysiological effect of amiodarone, lidocaine and quinidine on rat ventricular cells.

The effects of 20 microM each of amiodarone, lidocaine and quinidine on action potential and membrane currents were studied in rat ventricular cells. At a stimulation frequency of 0.1 Hz, quinidine prolonged the action potential duration (APD50) from 120 +/- 26 to 660 +/- 8 msec and increased the time to peak (Tp) amplitude from 7 +/- 1 msec to 32 +/- 6 msec. Lidocaine shortened APD50 from 123 +/- 15 to 83 +/- 6 msec without altering Tp. Amiodarone changed neither APD50 nor Tp. Voltage clamp study revealed that quinidine inhibited sodium inward current (INa) even when this current was elicited by depolarizing pulses at 0.1 Hz from a holding potential of -90 mV. For amiodarone and lidocaine, the inhibition was observed when INa was elicited from a holding potential of -70 mV. A frequency-dependent inhibition of INa by amiodarone and lidocaine was observed at frequencies higher than 1 Hz. Quinidine showed this inhibition even at 1 Hz. In correlation with the stronger frequency dependent inhibition of INa, a greater delay of the recovery and increase of the non-recovery fraction of INa was induced by quinidine. For lidocaine and amiodarone, only the recovery time constant was delayed. In cells treated with sea anemone toxin (ATX, 0.2 microM), APD50 was prolonged to 4-5 sec in 5 min. Quinidine, but not amiodarone, completely reversed the effect of ATX. Quinidine showed use-dependent inhibition of INa in these ATX-treated cells. Amiodarone, however, did not show this inhibition. It is likely that amiodarone suppresses INa by delaying the recovery of INa instead of blocking the open-state Na(+)-channels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Antagonists of calmodulin delay injury development in the severely ischemic perfused working rat heart

The effects of calmodulin antagonists trifluoperazine (TFP) and calmidazolium (CMZ) and of ethmozine (a phenothiazine without anticalmodulin activity) on the postischemic recovery in the perfused working rat hearts were studied. In the hearts subjected to 25 min zero-flow ischemia coronary flow, cardiac output, MVO2 and external work recovered to about 50% of the preischemic values during 40 min of reperfusion. TFP (5 x 10(-7) M and 10(-6) M) or CMZ (10(-7) M) improved the functional recovery to 75-94% whereas 5 x 10(-7) M ethmozine was not effective. In all experimental groups a prolongation of the ischemic period caused a progressive deterioration of the functional recovery while the total postischemic LDH release showed an initial gradual rise followed by a later decay. TFP and CMZ prolonged the time-to-half decay of the hemodynamic functions (tHF50) by 4-7 min and the time-to-peak of total LDH release (tLDHmax) by 5-10 min. In the hearts subjected to 0.2 ml/min low-flow ischemia tHF50 and tLDHmax were increased to 40 min, CMZ prolonged these times by further 5-10 min. Thus, TFP and CMZ delayed the development of the myocardial ischemic injury. Although other interpretations are possible, our data are consistent with the hypothesis that calmodulin-sensitive process is involved in the ischemic damage of the myocardium.     

The vagus-inhibiting action of alpha-endorphin]

An intravenous bolus injection of 0.1 ml alpha-endorphin (1 x 10(-8)-1 x 10(-4) g/ml) didn't change the heart rate in frogs. The parasympathetic bradycardia induced by the peripheral vagus stimulation was decreased by alpha-endorphin. This vago-inhibitory action was dose-dependent (1 x 10(-5)-1 x 10(-4) g/ml). The maximal inhibitory action was watched in 4-8 and 9-15 minutes after bolus injection of alpha-endorphin in concentration of 1 x 10(-5) and 1 x 10(-4) g/ml accordingly.     

Influence of light on membrane potential of Neurospora crassa cells

In experiments with left atrial and right ventricular tissues fast sodium current (INa) was decreased by substituting sucrose for sodium in Tyrode's solution and with antiarrhythmic drugs--INa blockers (lidocaine, diphenylhydantoine and ethmozine). It was shown that INa decrease results in the growth of refractoriness (R) of the cardiac tissues. The hypothesis is advanced that the R growth caused by INa decrease is one of the mechanisms of selective sensitiveness of ischemic heart tissues to antiarrhythmic drugs--INa blockers.     

Epithelium modulates the potency of vasoactive intestinal peptide in the guinea pig.

The purpose of the study was to evaluate the importance of the epithelium in determining the potency of exogenous vasoactive intestinal peptide (VIP) in inhibiting responses of isolated guinea pig trachea to vagal stimulation. Isolated innervated tracheal preparations (n = 56) were mounted in glass organ baths in Krebs-Henseleit (K-H) solution at 37 degrees C and gassed with 95% O2-5% CO2. The inside of the trachea was separately perfused with K-H solution at 1 ml/min. The vagal nerve trunks were stimulated (20 V, 1-ms pulses, 10-s trains) at low (0.5 Hz) and high frequency (15 Hz) alternately, and the contractile responses were measured as increases in intratracheal pressures. VIP (10(-8)-10(-7) M) inhibited responses to both high- and low-frequency stimulation. VIP was more potent in inhibiting contractions when administered to the outside than the inside surface of the trachea, and disruptionon of the epithelium abolished this difference. The endopeptidase inhibitors phosphoramidon and thiorphan (5 x 10(-6) M) potentiated the action of VIP. These data indicate that the epithelium reduces the efficacy of VIP. We suggest that the epithelium is a site of degradation of VIP by endopeptidase and may also be a diffusion barrier.     

Inhibiting action of anti-arrhythmia agents of the phenothiazine series--etmozin and its diethylamino analog--on platelet aggregation and metabolism of endogenous arachidonic acid

Effect of the cardiotropic drugs of the phenothiazine series ethmozine, and its diethylamine analogue (DAAE), on platelet aggregation and formation of arachidonic acid metabolites has been studied. Both drugs inhibit the ADP-induced aggregation in the platelet-rich plasma. Ethmozine inhibits only the second (irreversible) wave of aggregation, while DAAE inhibits both the first (reversible) and the second one. 50% inhibition (ID50) of the second wave of aggregation is observed at the following concentrations of the two agents: 300-500 micrograms/ml (ethmozine) and 20 micrograms/ml (DAAE). DAAE completely inhibits the irreversible aggregation of platelets washed off plasma, induced by arachidonic acid (ID50 approximately 30 micrograms/ml) and Ca2+-ionophore A23187 (ID approximately 55 micrograms/ml); the aggregation, induced by thrombin is inhibited by 80-90% (ID approximately 130 micrograms/ml). Formation of arachidonic acid metabolites in platelets effected by these inducers was measured by the accumulation of malondialdehyde (MDA). DAAE fails to inhibit MDA formation induced by exogenous arachidonic acid, but completely prevents the synthesis of MDA induced by A23187 and thrombin. These data suggest that DAAE inhibits the release of endogenous arachidonic acid from membrane phospholipids catalysed by phospholipase A2, but does not affect its subsequent metabolic transformations. In all probability, ethmozine and DAAE, just as other phenothiazines, affect platelets via the inhibition of Ca2+-calmodulin-dependent reactions and processes.     

Ionic mechanism of the Woodwors staircase

Analysis of single-chamber model of electromechanical coupling in the myocardial cell has shown that Woodwors staircase can be imitated in two cases: 1) stationary input current Ca2+ strongly exceeds the potential-dependent uptake of Ca2+ into the cell through the sarcolemma; 2) the action potential (AP) is shortened abruptly with an increase of the myocardium stimulation frequency. The experiments performed on a fragment of the frog heart ventricle supported the conclusions of the model. Blocking of Ca-channels with nifedipine (10(-6) g/mol) at the background of isotonic substitution of 70% of NaCl resulted in the development of "negative staircase" with an increase of stimulation rhythm. An abrupt shortening of AP after rest at joint action of adrenaline (10(-6) g/ml) and blocker of Ca-channels D-600 (10(-6) g/ml) was accompanied by Woodwors staircase.     

Effect of ozone exposure on contractile activity and chemoreactivity of uterus horns longitudinal muscles of nonpregnant rats

With the purpose of studying the mechanism of ozone action on uterus smooth muscles it was investigated the influence of ozone-content (approximately 0.50 mkg/ml) Krebs' solution or its 10- and 100-fold dissolution on contractile activity and beta-adrenoreactivity of 56 longitudinal strips of uterus horns of 17 nonpregnant rats. Ozone at concentration approximately 0.50 mkg/ml (but not in concentration of approximately 0.05 and approximately 0.005 mkg/ml) reversibly raised frequency, amplitude and total contractile activity of intact myometrium strips, and also fast and reversibly reducel its beta-adrenoreactivity, i.e. decreased of inhibitory action of adrenaline (10(-8), 10(-7), 10(-6) g/ml), but did not change uterostimulatory effect of acetylcholine (10(-6) g/ml) and oxiyocin (5 x 10(-4) ME/ml), what evident about specificity of ozone beta-adrenoblokate effect. Ozone (approximately 0.50 and 0.05 mkg/ml) did not change ov value of potassium contracture of myometrium strips which was depolarized by hyperpotassium (60 mM KCL) Krebs' solution, but reduced inhibitory action of adrenaline (10(-8) g/ml). The question is being discussed about mechanisms of ozone beta-adrenoblocade actions, about clinical role of this phenomenon, and the possibility of using beta-adrenoreceptor sensibilizators direct action (histidine, tryptophan, tyrosine, trimetazidin and mildronat) at ozonotherapy with the purpose reduction of its negative effects.     

Effects of dried bonito (katsuobushi) and captopril, an angiotensin I-converting enzyme inhibitor, on rat isolated aorta: a possible mechanism of antihypertensive action

In order to elucidate the mechanism of the antihypertensive action of dried bonito (katsuobushi), we compared the effects of dried bonito extracts with those of captopril, an angiotensin I-converting enzyme (ACE) inhibitor, on aorta preparations isolated from rats. Dried bonito extracts (3 x 10(-4) to 3 x 10(-3) g/ml) more potently relaxed contractions induced by norepinephrine (10(-7) M) than contractions induced by KCl (55.9 mM). Dried bonito extracts (3 x 10(-3) g/ml) slightly inhibited 10(-7) M angiotensin I-induced contractions. In contrast, captopril (10(-8) to 10(-7) M) did not affect 10(-7) M norepinephrine- or 55.9 mM KCl-induced contractions, but a higher concentration of captopril (10(-6) M) very slightly relaxed it. Captopril (10(-8) to 10(-6) M) markedly inhibited 10(-7) M angiotensin I-induced contractions in a concentration-dependent manner. These results suggest that antihypertensive mechanism of action induced by dried bonito involves direct action on vascular smooth muscle in addition to ACE-inhibitory activity.     

Effect of monensin on the specific activity of ammonia production by ruminal bacteria and disappearance of amino nitrogen from the rumen.

When unadapted mixed ruminal bacteria (312 mg of protein per liter) were treated with monensin (5 mM) in vitro, the rates of ammonia production from enzymatic digests of casein, gelatin, and soy protein (0.5 g of N per liter) were decreased from 46 +/- 2 to 24 +/- 1, 20 +/- 1 to 7 +/- 1, and 40 +/- 2 to 18 +/- 2 nmol/mg of protein per min, respectively. Monensin also caused a decrease in ammonia production in vivo. Nonlactating dairy cows which were fed 0.56 kg of timothy hay 12 times per day had a steady-state ruminal ammonia concentration of 2.7 +/- 0.1 mM, and the ammonia concentration decreased to 1.2 +/- 0.2 mM when monensin (350 mg/day) was added to the diet. The decrease in ammonia production was associated with a 10-fold reduction (4.1 x 10(6) versus 4.2 x 10(5)/ml) in the most probable number of ammonia-producing ruminal bacteria that could use protein hydrolysate as an energy source. Monensin had little effect on the most probable number of carbohydrate-utilizing ruminal bacteria (6.5 versus 7.0 x 10(8)/ml). The addition of protein hydrolysates (560 g) to the rumen caused a rapid increase in the ammonia concentration, but this increase was at least 30% lower when the animals were fed monensin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of endogenous opioids in ischemic preconditioning but not in short-term hibernation in pigs

Endogenous opioids are involved in ischemic preconditioning (IP) in several species. Whether or not opioids are important for IP and short-term myocardial hibernation (STMH) in pigs is currently unknown. In 34 enflurane-anesthetized pigs, the left anterior descending coronary artery was flow constantly perfused. Subendocardial blood flow (Endo), infarct size (IS; percent area at risk), and the free energy change of ATP hydrolysis (DeltaG) were determined. After 90-min severe ischemia and 120-min reperfusion, IS averaged 28.3 +/- 5.4% (means +/- SE) (n = 8; Endo: 0.047 +/- 0.009 ml. min(-1) x g(-1)). IP by 10-min ischemia and 15-min reperfusion reduced IS to 9.9 +/- 3.8% (P < 0.05, n = 8; Endo: 0.044 +/- 0.009 ml. min(-1) x g(-1)). After naloxone (1 mg/kg iv followed by 2 microg x kg(-1) x min(-1)), IS averaged 25.8 +/- 7.0% (n = 6; Endo: 0.039 +/- 0.008 ml x min(-1) x g(-1)) without and 24.7 +/- 4.7% (n = 6; Endo: 0.044 +/- 0.006 ml x min(-1) x g(-1)) with IP. At 5-min moderate ischemia in the presence of naloxone, Endo decreased from 0.90 +/- 0.07 to 0.28 +/- 0.03 ml x min(-1) x g(-1)and DeltaG decreased from -58.6 +/- 1.0 to -52.6 +/- 0.4 kJ/mol. Prolongation of ischemia to 90 min did not alter Endo, but DeltaG recovered toward control values (57.7 +/- 1.1 kJ/mol), and the myocardium remained viable. These responses are identical to those of nonnaloxone-treated pigs. Endogenous opioids are involved in IP but not in STMH in pigs.     

Co-culture of human monocytes and thyrocytes in bicameral chamber: monocyte-derived IL-1alpha impairs the thyroid epithelial barrier

Pro-inflammatory cytokines are important mediators in tissue responses to a wide range of endogenous (e.g. autoantigens) and exogenous (e.g. infections, wounds, biomaterials) stimuli. The complex interactions taking place between different cell types in such processes are difficult to examine in vivo. Here we studied the effect of human monocytes on thyroid epithelial cells co-cultured in bicameral chambers. Freshly isolated monocytes (1x10(6)/ml) added to the basal compartment reduced the transepithelial resistance (from 300-600 to <100 Omega.cm(2)) and caused a disruption of the tight junctions in apically grown thyrocyte monolayers after co-culture for 24 h. The barrier function was further attenuated by monocytes exposed to lipopolysaccharide (10 microg/ml) or polystyrene microspheres (size: 3 microm; 1x10(7)/ml). Loss of transepithelial resistance was accompanied by release of interleukin 1alpha (maximally 550 pg/ml) from the monocytes. Conversely, the resistance remained high when co-cultures were simultaneously incubated with neutralizing anti-human interleukin 1alpha antibodies. The results show that the integrity of cultured thyroid epithelium is impaired by monocytes without requirement of direct cell-to-cell contact. This action, mediated by interleukin-1alpha, suggests a mechanism by which hidden (lumenal) autoantigens might be exposed to interstitial antigen-presenting cells in autoimmune thyroid disease. In perspective, the model provides a tool in which humoral and cell-cell dependent processes generated by bioactive agents and particulate materials, for instance, during the healing and repair of tissue around biomaterials and hybrid implants, can be selectively examined.     

Effect of diethylamine analog of ethmozine on parameters of fast sodium current in normal and depolarized myocardial fibers

The effect of a diethylamine analog of ethmozine (DAAE) on fast sodium current of normal and depolarized frog atrial trabeculae was studied by means of the double sucrose gap technique. The depolarization of the fibers was produced both by increasing extracellular potassium concentration up to 8 +/- 9 mM and by current passing. The resting potential of normal fibers was within the range of 75-80 mV, and the depolarized one was within 65 +/- 60 mV. DAAE (8 X 10(-7) g/ml) reduced sodium conductivity, slowed inactivation and reactivation of fast sodium current and shifted the steady-state activation curve (h infinity) to a more negative potential, but the steady-state activation curve (m infinity) to a more positive potential. All these effects were expressed more considerably in the depolarized fibers. The depressing ability of DAAE is assumed to be due to high concentration of the drug in the membrane owing to its high solubility in lipids. The slowing of sodium reactivation and inactivation shows the existence of a receptor for DAAE linked with a h-gate. Prolonged antiarrhythmic action is accounted for by a very slow recovery of sodium current after the drug superfusion. A stronger effect of DAAE on sodium current in the depolarized fibers is likely to point to its selective action on ischemic tissue.     

Effect of chromium ions on the function of neuromuscular junctions

It has been shown on neuro-muscular preparations of frog sartorius muscle that chromium ions in the concentrations 1-4 x 10(-6) g/ml strengthen spontaneous and evoked transmitter release. Cr3+ ions in the concentrations above 4 x 10(-6) g/ml decrease the membrane potential of muscle fibres, decrease the quantum content of the end plate potentials. Experiments on a single Ranvier node have shown that Cr3+ ions decrease the amplitude, increase the rate and duration of the action potential of a nerve fibre. It is concluded that chromium ions produce a pronounced effect on synaptic transmission, which differs significantly from the action of manganese, cobalt and nickel ions.