[Effect of insulin on characteristics of contractile responses of fast and slow skeletal muscles of rats with streptozotocin-induced overt diabetes].

Comparison of amplitude-time characteristics of fast (m. EDL) muscles in rats with acute model of streptozotocin diabetes (SD) (12 and 30 days after treatment with streptozotocin) did not reveal significant changes in strength of single normalized contractile responses as compared with control. In slow (m. Soleus) muscles of rats with the 30-day SD there were observed essential changes in amplitude-time characteristics of such contractile responses: a decrease of their amplitude and an increase of duration. In diabetic rats treated with a course of insulin, resistance of skeletal muscles to both types of exogenous insulin is developing. Both in control and in diabetic animals the fatiguing stimulation of m. EDL by firing from 5 impulses did not reveal significant differences at early (up to 3 min) terms of development of fatigue. Under similar conditions, fatigue of m. Soleus in rats of both diabetic groups developed significantly faster as compared with control (as early as 30 s after the beginning of stimulation). Insulin at a concentration of 5-1 nM produced a dose-dependent decrease of amplitude of single contractile responses in fast and slow muscles of rats with the acute SD model (the negative inotropic action). The same effect of insulin, but at the higher concentrations, we demonstrated earlier in health rats. Insulin at a concentration of 10 nM did not produce essential effect on the time course of depression of responses in the course of development of fatigue at tetanic stimulation of m. EDL and m. Soleus both in control and in diabetic rats, but affected essentially the time course of change in duration of the half-decline (T(hd)) of their tetanic responses. The presence of insulin in the bath solution led to stabilization of the muscle relaxation period in the course of development fatigue in all studied animal groups.

[Action of insulin on contraction and electrical responses of rat skeletal muscle].

The effect of insulin on contractility of directly stimulated skeletal muscles was studied in experiments in isolated preparations of rat fast, extensor digitorum longus (m. EDL), slow, soleus (m. SOL) and mixed, diaphragm muscles. In addition (diaphragm only) characteristics of extracellularly recorded muscle fiber action potentials (APs) were evaluated before and after addition of insulin to a bath solution. Insulin (0.5-10 nM) decreased muscle twitch force. This negative inotropic effect of insulin was dose-dependent, with m. SOL appearing to be more sensitive to insulin than either m. EDL or diaphragm. Insulin did not affect strength of isotonic KCL- or caffeine-induced muscle contractures, but decreased second and increased first and third phases of extracellularly recorded muscle fiber APs. The analysis of the data obtained in this study and the data from the literature suggests changes in electrogenesis of the muscle fiber's t-tubular plasma membrane as a key element of the negative inotropic effect of the hormone on contractility of mammalian skeletal musculature. Possible mechanisms of such putative changes are discussed.

[Effect of barium and ouabain on electrogenesis in various sites of intact and detubulated skeletal muscle fibers of the frog R. temporaria].

In loose patch clamp experiments on intact sartorious muscle fibers of the frog Rana temporaria there were revealed two types of wave forms of the extracellularly recorded action potentials (AP). Responses of the first type (T1AP) consisted of the initial positive phase with subsequent phase of strong negativity. Responses of the second type (T2AP) had an additional positive phase concluding their waveform. In the detubulated fibers, only T1AP were recorded. Application Ba2+ (10 microM) to the muscle led to a significant increase in the amplitude of the third T2AP phase, whereas the T1AP characteristics of both intact and detubulated muscle preparations remained unchanged. After the higher applied Ba2+ concentration (50 microM), in some of the studied intact fibers the late positive signal phase was replaced by the late negative phase. The amplitude of the late negative phase was increased markedly by the high-frequency stimulation. At the combined action of ouabain and Ba2+ there was summation of effects. These results can tentatively be explained by that the locally recorded T1AP and T2AP represent responses of the muscle fiber surface and T-tubular membranes and that caracteristics of the late phase of T2AP are determined by activities of T-tubular potassium channels and of Na, K-ATPase.

[Ouabain-induced blockade of alpha2 isoform of the Na,K-ATPase on electrophysiological and contractile characteristics of the rat diaphragm].

In experiments with isolated neuromuscular preparation of the rat diaphragm, selective blockade of alpha2 isoform of the Na,K-ATPase with ouabain (1 mcmol/L) induced steady depolarization of muscle fibers that reached a maximum of 4 mV, a decrease in amplitude of muscle fiber action potential, and prolonged raising and decline phases of the action potential. At the same time, the force, time to peak, and half relaxation time of the isometric muscle twitch were increased, as well as the area under the contraction curve. During continuous fatiguing stimulation (2/s), a more pronounced decline of contraction speed was observed in presence of ouabain; dynamics of the half-relaxation time remaining unchanged. It is suggested that blockade of alpha2 isoform of the Na,K-ATPase impairs excitation-contraction coupling resulting in a delay of Ca2+ release from sarcoplasmic reticulum. The increase in contraction force seems to result from a mechanism similar to that of positive inotropic effect of cardiac glycosides in heart muscle. Physiological significance of the skeletal muscle alpha2 isoform of the Na,K-ATPase in regulation of Ca2+ and Na+ concentrations near triadic junctions and in regulatory processes involving the Na,K-ATPase endogenous modulators or transmitter acetylcholine is discussed.

[Functional interaction between nicotinic cholinergic receptors and Na, K-ATPase in the skeletal muscles]. / Funktsional'naia sviaz' nikotinovykh kholinoretseptorov i Na, K-ATFazy v skeletnoi myshtse.

Acetylcholine (ACh) hyperpolarized the rat diaphragm muscle fibers by 4.5 +/- 0.8 mV (K0.5 = = 36 +/- 6 nmol/l). The AC-induced hyperpolarization was blocked by d-tubocurarine and ouabain in nanomolar concentrations. This effect of ACh was not observed in cultured C2C12 muscle cells and in Xenopus oocytes with expressed embryonic mouse muscle nicotinic acetylcholine receptors (nAChR) or with neuronal alpha 4 beta 2 nAChR. In membrane preparations from the Torpedo californica electric organ, containing both nAChR and Na, K-ATPase, 10 nmol/l ouabain modulated the binding kinetics of the cholinergic ligand dansyl-C6-choline to the nAChR. These results suggest that in-sensitive alpha 2 isoform) and nAChR in a state with high affinity to Ach and d-tubocurarine may form a functional complex in which binding of ACh to nAchR is coupled to activation of the Na, K-ATPase.

[Porcine kidney extract contains a specific inhibitor of the ouabain-sensitive alpha2-isoform of Na, K-ATPase present in rat diaphragm fibres]. / Ekstrakt pochek svin'i soderzhit spetsificheskii ingibitor ouabain-chuvstvitel'noi alfa2 izoformy Na, K-ATFazy myshechnykh volokon diafragmy krysy.

In experiments on isolated rat diaphragm muscle, acetylcholine (100 nmol/l) hyperpolarized muscle fibres due to activation of the alpha 2 isoform of Na,K-ATPase. This hyperpolarization was blocked in a dose-dependent manner by ouabain (K0.5 = 8 +/- 4 nmol/l) as well as by a solution of porcine kidney extract (10 kDa cut-off filtration), with the K0.5 approximately equal to a 1:20,000-fold dilution. The inhibitory activity of the developed slowly over a period of 3 hours and, in contrast to ouabain, was still present after 1 hour of washing. Ouabain, but not the extract, inhibits Rb+ uptake in human erythrocytes that only express the alpha = 1 isoform of Na, K-ATPase. Our data suggest that in rat skeletal muscle the alpha 1 isoform of Na,K-ATPase is primarily responsible for ionic homeostasis, while the alpha 2 isoform provides a "regulatable" function and may be controlled by cholinergic stimulation and/or endogenous digitalis-like factors (EDLFs). Porcine kidney extract contains a factor (M. W. < 10 kDa) that selectively inhibits the rat alpha 2 isoform and differs from ouabain. Our experimental protocol can be used as a highly sensitive physiological assay for factors that selectively inhibit the alpha 2 isoform of Na,K-ATPase.

A possible function of Schwann cells in neuromuscular transmission in the frog.

The total treatment of the cutaneous pectoris muscle of the frog with rabbit antiserum to galactocerebrosides leads to the slowing of the development of the depression of end-plate currents when the transected preparations are stimulated in normal Ringer's solution at a frequency of 100 imp/sec. Changes are not observed in the quantal composition and the binomial characteristics of neuromuscular transmission when nontransected preparations are stimulated at 1 imp/sec in solutions containing magnesium. The action of the antiserum to galactocerebrosides elicits an increase in the relative number of typical miniature end-plate potentials that are characterized by a prolonged time course. The electron microscopic investigation of the preparations demonstrates the absence of lysis of the schwann cells in the preparations subjected to treatment with antiserum. It is possible that the effects obtained reflect changes in the functional state of the Schwann cells of the synaptic region that arise as the result of the fixation on their membrane of antibodies to galactocerebrosides.

[Atypical endplate miniature potentials in the frog neuromuscular junction after modification of the intercellular matrix and osmotic exposures]. / Atipichnye miniatiurnye potentsialy kontsevoi plastinki v nervno-myshechnom soedinenii liagushki posle modifikatsii mezhkletochnogo matriksa i osmoticheskikh vozdeistvii.

In frog cutaneous-pectoris muscles the frequency of slowly rising atypical miniature endplate potentials (MEPPs) was significantly enhanced after collagenase (0.1%) treatment. Treatment with trypsin, hyaluronidase, hyper- and hypoosmotic solutions caused no changes in slowly rising MEPP (frequency in muscle fibers with intact acetylcholinesterase (AChE). Inhibition of AChE caused appearance of giant MEPPs. Acceleration of acetylcholine diffusion from synaptic cleft after treatment with hyaluronidase decreased giant MEPP frequency demonstrating their dependence upon nonhydrolyzed acetylcholine in synaptic cleft. The relation between slowly rising MEPPs and activity of synaptic Schwann cells in discussed.

[The possible function of Schwann cells in neuromuscular transmission in the frog]. / O vozmozhnoi funktsii shvannovskikh kletok v nervno-myshechnoi peredache u liagushki.

Anti-galactoside rabbit antisera increased the frequency of slow (atypical) MEPPs. No lysis of the Schwann cells was found in synaptic regions. The data obtained suggest that fixation of antibodies to galactocerebroside on the Schwann cell surface membrane alters the cell's activity thus affecting some characteristics of neuromuscular transmission.

[Psychosine-induced changes in neuromuscular transmission and in the structure of the neuromuscular junction in the frog]. / Vyzyvaemye psikhozinom izmeneniia nervno-myshechnoi peredachi i struktury nervno-myshechnogo soedineniia u liagushki.

A decrease in the amplitude of miniature and evoked end-plate potentials (MEPPs and EPPs) as well as changes in EPP facilitation and depression during frequency stimulation were observed in psychosine-treated m. cutaneous-pectoris of the frog. Electron-microscopic investigation demonstrated severe changes in synaptic Schwann cells embracing motor terminals and disturbances in the inner mesaxon structure of the myelinated parts of motor terminals.

[The formation of nerve endings in the phasic muscles of the frog]. / Formirovanie nervnykh okonchanii v faznykh myshtsakh liagushki.

Synapses with the complex nerve endings consisting of several terminal branchings of a single axon divided by myelin segments were studied at the cutaneous-pectoris muscles of a frog by histological and electrophysiological methods. It has been revealed that the number of the synapses with complex nerve endings rises during the postnatal development. The relation between the nerve ending complexity, total terminal length and size of the muscle fibres is described. Some of branches in complex nerve endings arise from the Ranvier nodes. They are characterized by a low level of quantal secretion and a peculiar fall of the sodium current along nerve terminals. Mechanisms that induce growth of nerve endings, branching and peculiarities of the transmitter release are discussed. It is supposed that the processes of growth and myelinization of nerve endings take place simultaneously.

[Seasonal changes in neuromuscular transmission in experiments on dissected or curarized frog muscles]. / Sezonnye izmeneniia nervno-myshechnoi peredachi v opytakh na rassechennykh ili kurarizovannykh myshtsakh liagushki.

The end-plate currents and potentials (EPCs and EPPs) in cut or curarized muscles were investigated in frogs maintained at different temperatures (5 and 20-25 degrees C) before experiments in summer and in winter. EPCs and EPPs were elicited by nerve stimulation with different frequencies (1 and 100/s). Seasonal changes in amplitude, facilitation and depression of the EPPs and EPCs observed in curarized and cut (not curarized) muscles differed quantitatively and qualitatively. These differences may be explained assuming that sensitivity of the neuromuscular junction to the curare action (probably, a presynaptic one) is season-variable.

[Myelinization within the limits of the motor nerve ending as a possible factor in increasing synaptic transmission efficiency]. / Mielinizatsiia v predelakh motornogo nervnogo okonchaniia kak vozmozhnyi faktor uvelicheniia éffektivnosti sinapticheskoi peredachi.

The end-plate potentials (EPPs) were investigated in frog neuromuscular junctions with different nerve endings structure (cutaneous pectoris muscle). When the muscle fibres input resistance and nerve endings length in selected pairs of synapses were similar, larger EPPs amplitudes were noted in synapses with myelin segments within nerve ending. It is suggested, that myelinization within nerve ending is one of the factors of synaptic efficacy.

[Effect of antibodies to glial glycolipid antigens on miniature end-plate potentials]. / Vliianie antitel k glikolipidnym antigenam glii na miniatiurnye potentsial'y kontsevoi plastinki.

Less than 4% of miniature end-plate potentials (MEPPs) in frog cutaneous-pectoral muscles, normal or exposed to nonimmune rabbit serum were atypical. These MEPPs were of normal amplitude, but their time course was nearly twice as slow as that of normal MEPPs. The exposure of muscles to rabbit anti-galactocerebroside serum induced more than three-fold increase in the fraction of atypically slow MEPPs. Inhibition of acetylcholinesterase caused an appearance of giant MEPPs, the fraction of slow MEPPs being unchanged. Possible mechanisms of increase in fraction of atypical slow MEPPs based on modification of the synaptic Schwann cell functioning are discussed.

[Variations in the sensitivity of the neuromuscular junctions of the frog to the blocking action of hyperkalemic solutions]. / Variiatsii chuvstvitel'nosti nervno-myshechnykh soedinenii liagushek k blokiruiushchemu deistviiu giperkalievykh rastvorov.

The depression of evoked transmitter release with solutions with enhanced (up to 8 mM) potassium concentration was studied in cut and voltage-clamped muscle fibers of frog m. cutaneous pectoris. As potassium blockade proceeds, a reduction of the second end-plate current to paired stimuli occurs before a change in the first end-plate current, the reduction of end-plate currents being simultaneous with the latency increase. The existence of variable sensitivity of different end-plates to potassium blocking action depends on year season and ionic environment. Possible mechanisms of potassium blocking action are discussed.

[Effect of etimizol on the neuromuscular synapse of the frog]. / Deistvie étimizola na nervno-myshechnyi sinaps liagushki.

The effect of aethimizol, a substance activating the memory processes, was studied by intracellular recording at the frog cutaneous pectoris neuro-muscular preparation. Presynaptic action of aethimizol (0.5-2 mM) was found. This action consisted in the increase of miniature end-plate potentials (mEPP's) frequency and the modulation of quantal content of evoked end-plate potentials (EPP's) due to changes in binomial quantal release. parameters p and n. These effects depended on the initial level of quantal release. Aethimizol at the concentrations of 5-10 mM strongly decreased the amplitudes of mEPP's and EPP's while at the concentrations of 0.5-10 mM aethimizol increased the rising- and half-decay time of mEPP's and EPP's and half decay time of evoked end-plate currents presumably affecting the postsynaptic ionic channels properties. Aethimizol increased the time integral of EPP's. Effects of aethimizol are discussed in relation to its possible modes of action in the central neural system as a presumed unspecific connector.