[Influence of mitochondrial oxidative reactions on myocardial contractility. Effects of Ni2+].

Inotropic effects of Ni2+ and mitochondrial oxidative readions were accordingly tested by the use of frog myocardial preparations, excised from left frog ventricle, and rat heart mitochondria (RHM). Amplitude of the cardiac contraction was increased in the presence of 10-200 microM Ni2+ in dose-dependent manner. It was found that Ni2+ is not toxic for RHM. State 4 in a KCI medium was stimulated by 100 microM Ni2+. At the same time, Ni2+ did not affected state 3 and 2,4-dinitrophenol-stimulated respiration of RHM. Our findings allow to be supposed that Ni2+-induced increase in the amplitude of the cardiac contraction can be affected by energetic state of RHM.

[T-channels and Na+,Ca2+-exchangers as components of the Ca2+-system of the myocardial activity regulation of the frog Rana temporaria].

Earlier we have shown that regulation of rhythm and strength of the frog heart contractions, mediated by transmitters of the autonomic nervous system, is of the Ca2+-character. In the present work, we studied chrono- and inotropic effect of verapamil--an inhibitor of Ca2+-channels of the L-type, of nickel chloride--an inhibitor of Ca2+-channels of the T-type, and of Na+,Ca2+-exchangers as well as of adrenaline and acetylcholine (ACh) after nickel chloride. It has been found that the intracardially administered NiCl2 at a dose of 0.01 microg/kg produced a sharp fall of amplitude of action potential (AP) and an almost twofold deceleration of heart rate (HR). The intracardiac administration of NiCl2 (0.01 microg/kg) on the background of action of verapamil (6 mg/kg, i/m) led as soon as after 3 min to even more prominent HR deceleration and to further fall of the AP amplitude by more than 50% as compared with norm. The intracardiac administration of adrenaline (0.5 mg/kg) partly restored the cardiac activity. However, preservation of the myocardium electrical activity in such animals was brief and its duration did not exceed several minutes. Administration of Ni2+ on the background of acetylcholine (3.6 mg/kg) led to almost complete cessation of cardiac activity. As soon as after 3 min after injection of this agent the HR decreased to 2 contractions/min. On EG, the 10-fold fall of the AP amplitude was recorded. The elucidate role of extra- and intracellular Ca2+ in regulation of heart contractions, isometric contraction of myocardium preparations was studied in response to action of NiCl2 (10-200 microM), verapamil (70 microM), adrenaline (5 microM), and acetylcholine (0.2 microM) after NiCl2. It is found that Ni2+ caused a dose-dependent increase of the muscle contraction amplitude. Minimal change of the contraction amplitude (on average, by 14.9% as compared with control) was recorded at a Ni2+ concentration of 100 microM. An increase of Ni2+ in the sample to 200 microM increased the cardiac contraction strength, on average, by 41%. The negative inotropic action of verapamil was essentially reduced by 100 microM Ni2+. Adrenaline added to the sample after Ni2+ produced stimulating effect on the cardiac muscle, with and almost twofold rise of the contraction amplitude. ACh (0.2 microM) decreased the cardiac contraction amplitude, on average, by 56.3%, whereas Ni2+ (200 microM) administered after ACh not only restored, but also stimulated partly the myocardial work. Within several parts of percent there was an increase of such isometric contraction parameters as amplitude of the muscle-developed effort, maximal rate, maximal acceleration, time of semirise and semifall. The obtained experimental results indicate that the functional activity of the frog pacemaker and contractile cardiomyocytes is regulated by the Ca2+-dependent mechanisms. Structure of these mechanisms includes the potential-controlled L- and T-channels of the plasma membrane as well as Na+,Ca2+-exchangers characteristic exclusively of contractile cardiomyocytes. The existence of these differences seems to be due to the cardiomyocyte morphological peculiarities that appeared in evolution at the stage of the functional cell specialization.

[Peculiarities of Ca2+ regulation of functional activity of myocardium of frog Rana temporaria].

To elucidate role of intra- and extracellular Ca2+ in regulation of rhythm and strength of frog heart contractions, there were studied ECC and isometric contraction of myocardium preparations in response to verapamil, adrenaline, and blockers of alpha- and beta-adrenoreceptors. It has been shown that after an intramuscular injection of verapamil (6 mg/kg), bradycardia develops, the heart rate (HR) decreasing by 50-70 %. Further, the cardiac arrest occurred; however, administration to the animals of adrenaline (100 mg/kg) restored the cardiac rhythm for a short while. After an intramuscular injection of adrenaline at doses of 0.1-10 mg/kg, no essential changes were observed in the potential action amplitude and HR; an increase of the administered adrenalin concentration to 100 mg/kg was not accompanied by the cardiac rhythm stimulation, as this takes place in homoiothermal animals and human; on the contrary, an essential HR deceleration was revealed. Phentolamine (5 mg/kg) gradually decelerated HR rhythm by 32-45 %. The potential amplitude changed insignificantly. A subsequent intracardiac injection of adrenaline (100 mg/kg) on the background of block of alpha-adrenoreceptors produced acceleration of the rhythm (by 13-21%) and fall of the electrogram amplitude. These results can indicate that in the frog heart, phentolamine interacts predominanty with alpha-adrenoreceptors. An intracardiac administration of propranolol (1 mg/kg) to frogs promoted inhibition of beta-adrenergic receptors and produced a gradual cardiac rhythm deceleration. In experiments on assessment of verapamil effect on the character of contractions this preparation at a concentration of 150 microM was established to produce a significant dosedependent decrease of the contraction strength. A rise of verapamil concentration in the sample to 200 microM led to a decrease of the amplitude, on average, by 68-70 % and in individual preparations -- by 80-85 %; however, administration into the sample of adrenaline (10 microM) restored the cardiac contraction strength. Adrenaline (1 nM--100 microM) increased markedly the contraction amplitude. Phentolamine (10 microM) did not inhibit transmission of contractile signal to cardiomyocytes; this was manifested in that the contraction amplitude after addition of adrenaline (10 microM) into the sample was approximately the same as in the sample containing no phentolamine. Propranolol (10 microM) eliminated the stimulatory action of adrenaline (10 microM). The results of these experiments indicate that in the frog ventricular cardiomyocytes the main adrenaline acceptors are beta-adrenoreceptors.

[Role of acetylcholine in the Ca2+-dependent regulation of functional activity of myocardium of frog Rana temporaria].

To study role of ACh in the Ca2+-dependent regulation of rhythm and strength of cardiac contractions in frog Rana temporaria, the ACh chrono- and inotropic effects have been studied in parallel experiments on the background of blockers of potential-controlled Ca2+-channels, ryanodine and muscarine receptors. The obtained results indicate participation of acetylcholine in the Ca2+-dependent regulation of rhythm and strength of frog cardiac contractions.

[Effect of ethanol and polyethylene glycol-400 on frequency dependence of the inotropic effect of verapamil on the frog myocardium]. / Vliianie étanola i poliétilenglikolia-400 na chastotozavisimost' inotropnogo deistviia verapamila na serdechnuiu myshtsu liagushki.

The PEG-400 was found to be neutral in respect to the frequency-dependent V negative inotropic action whereas ethanol enhanced this dependence. The findings suggest a possibility for V inotropic activity to be modified by lipophilic agents, provided they sterically complementary to hydrophobic domains of the Ca2(+)-channel structure.

[Hydrophobic interactions as the basis of the inotropic effect of verapamil]. / Gidrofobnye vzaimodeistviia kak osnova inotropnogo éffekta verapamila.

The role of hydrophobic interactions (HI) of verapamil (V) in the mechanism of its blocking action on voltage-dependent Ca(2+)-channels was considered. For this purpose a comparative study of V inotropic effects and general anesthetics (GA) modeling V action on rhythmic switches of frog heart muscle was made. Concentration thresholds of the agents of inotropic action (Cth) were determined at two frequencies of stimulation, 4 and 30 switches per minute, resp. According to features of the inotropic action the studied agents were divided into two groups: Cth values of the former group were dependent on stimulation frequency (V, methanol, ethanol, isopropanol, chloral hydrate and acetone); Cth values of agents of the latter group were independent on stimulation frequency (n-butanol, n-pentanol, n-hexanol, and chloroform). In both groups Cth values were linearly dependent on agent hydrophobicities (in logarithmic scale). It indicates that HI play the main role in interaction of the studied agents with biostructures and primarily with protein structures of Ca(2+)-channels, which are targets for V-like ligand action. The results obtained make an experimental support of our earlier proposed "hydrophobic" model of blocking action of V on plasmalemmal L-type Ca(2+)-channels.

[Effect of verapamil and anesthetics on the level of sodium and potassium ions in frog muscles. Role of hydrophobic interactions]. / Vliianie verapamila i anestetikov na soderzhanie ionov natriia i kaliia v myshtsakh liagushki. Rol' gidrofobnykh vzaimodeistvii.

Verapamil, general and local anesthetics influences on ionic contents of frog sartorius and cardiac muscles in respect with these drugs hydrophobicity were studied to clarify the role of verapamil hydrophobic interactions in its physiological activity. It was found that concentration thresholds of the agents toxicities were linearly linked with their hydrophobicity (in logarithmic scale). This relationship provides support for our conclusion that verapamil, general and some local anesthetics are the members of a single drug family. It is a reason to believe that a hydrophobic mechanism of general anesthetic action on cellular structure is the same as a mechanism of verapamil activity in muscles. It is possible that verapamil is accepted as medical drug exactly due to optimum combination of high verapamil hydrophobicity with its structural complementarity to receptor site responsible for use-dependent channel blocking.