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Telezhkin V. S. Tsvilovskii V. V. Dyskina Yu. B. Davydovskaya N. V. Shuba M. F. 《Neurophysiology》2001,33(5):281-288
Using a sucrose-bridge technique, we studied electrical and mechanical responses of smooth muscle ring strips of the rabbit main pulmonary artery to applications of blockers of voltage-operated (including Ca2+-dependent) K+ channels, tetraethylammonium (TEA) and 4-aminopyridine (4-AP), as well to application of nitric oxide (NO); nitroglycerin (NG) was used as a donor of the latter. All experiments were carried out under conditions of blockade of the adreno- and cholinoreceptors in the preparation. Both TEA and 4-AP evoked dose-dependent effects: depolarization of smooth muscle cells (SMC) and their contraction. Simultaneous addition of TEA and 4-AP to the normal superfusate (Krebs solution) resulted in intensification of depolarization and initiated generation of action potentials (AP); contractions became rather intensive and possessed a tetanic pattern. Addition of NG to TEA- and 4-AP-containing Krebs solution effectively suppressed AP generation and contractions, whereas the depolarization level underwent only mild modifications. These findings show that Ca2+-dependent high-conductance K+ channels (KCa channels) and 4-AP-sensitive voltage-operated K+ channels (KV channels) are involved in the formation of the resting membrane potential (RMP) in SMC of the rabbit main pulmonary artery. The impact of the KCa channels is greater than that of the KV channels. We suppose that the effects of NO on SMC are related to inhibition of the activity of high-threshold voltage-operated L-type Ca2+ channels and, probably, to lowering of the sensitivity of the contractile SMC apparatus to Ca2+. 相似文献
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A. V. Zholos T. B. Bolton A. V. Dresvyannikov M. V. Kustov V. V. Tsvilovskii M. F. Shuba 《Neurophysiology》2004,36(5-6):398-406
Acetylcholine, the main neurotransmitter of the parasympathetic nervous system, depolarizes various smooth muscles and initiates their contraction via activating muscarinic cholinergic receptors. In most visceral smooth muscle tissues, such as the gastrointestinal tract, airways, and the urinary system, muscarinic receptors are comprised of predominant M2 (about 80%)and minor M3 (about 20%) subtypes. Cholinergic excitation is generally mediated by the opening of ion channels selective for monovalent cations (under physiological conditions, Na+ and K+); among them the cationic channel of an about 60 pS unitary conductance has been recently identified as the main target for acetylcholine action. The signal transduction leading to channel opening is very complex and involves activation of Go protein (an M2 effect), activation of phospholipase C (an M3 effect), and [Ca2+]i and voltage dependence of channel opening. These multiple signaling pathways were difficult to reconcile with the channel gating mechanisms since only a simplified two-state channel mechanism (e.g., one open and one shut state) was until recently available. However, our recent studies of channel gating in isolated outside-out membrane patches revealed a greater complexity. Thus, this cationic channel shows transitions between at least eight states, four open and four shut, with strong connections between adjacent shut and open states. Therefore, four pairs of connected states have been identified, which showed voltage-dependent transitions in each pair of shut/open states. Since the membrane potential did not affect the relative proportions between the pairs, we have assumed that these effects are controlled by ligands that bind to the channel and, thus, stabilize its various open conformations. In this work, direct tests of the above hypothesis have been performed, and their results showed that spontaneous brief channel gating exists in the absence of receptor or G-protein activation, which is strongly voltage-dependent (increasing at depolarized potentials). Furthermore, this activity was potentiated at a low agonist concentration, while channel openings generally remained brief. An increasing receptor occupancy by the agonist produced long channel openings, indicating a shift of gating towards a long open/brief shut pair of the channel states. These findings are interpreted in the context of the established signal transduction pathways;certain predictions for the whole-cell current are also examined.Neirofiziologiya/Neurophysiology, Vol. 36, Nos. 5/6, pp. 446–454, September–December, 2004.This revised version was published online in April 2005 with a corrected cover date and copyright year. 相似文献
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M. V. Kustov V. V. Tsvilovskii A. V. Zholos M. F. Shuba T. B. Bolton 《Neurophysiology》2006,38(1):1-8
Using a two-wave fluorescence probe, Fura-2, we studied changes in the intracellular concentration of calcium ions ([Ca2+]i) resulting from activation of muscarinic and purine receptors in single myocytes of the guinea-pig small intestine. Applications
of the respective agonists added to the normal Krebs solution (1.0, 10.0, and 100.0 μM carbachol, CCh, as well as 10.0 and
100.0 μM ATP) induced a rise in the [Ca2+]i. Carbachol evoked an increase in the [Ca2+]i, including two components (a rapid and a plateaulike), while ATP under analogous conditions led only to a short-lasting rise
in the [Ca2+]i. Transients induced by CCh or ATP applied in different concentrations, which exceeded a certain level, did not significantly
differ from each other in their amplitudes, i.e., they were generated according to an all-or-none principle. In the nominally
Ca-and Mg-free solution, CCh and ATP induced only rapid increases in the [Ca2+]i in myocytes. The absence of the slow component in the [Ca2+]i elevation upon the action of CCh under such conditions indicates that the effect of ATP, as compared with that of CCh, is
not related to activation of the entry of Ca2+ ions into cells through voltage-operated calcium channels. After the addition of CCh, repeated application of CCh or ATP
induced no effect, while application of CCh after the addition of ATP initiated a rise in the [Ca2+]i. These data show that intracellular calcium stores are depleted completely upon the action of CCh, while they are depleted
only partially after the action of ATP. An inhibitor of phospholipase C (PLC), U-73122 (5.0 μM), completely blocked rises
in the [Ca2+]i induced by both CCh and ATP; therefore, the release of Ca2+ ions from the intracellular calcium stores after application of these agonists is mediated by PLC. We hypothesize that the
difference in the release of Ca2+ ions from the intracellular stores observed in our experiments upon activation of choline and purine receptors (partial and
complete depletion of the stores upon the action of ATP and CCh, respectively) is responsible for the opposite functional
effects of the above-mentioned neurotransmitters on smooth muscles.
Neirofiziologiya/Neurophysiology, Vol. 38, No. 1, pp. 3–10, January–February, 2006. 相似文献
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Using a patch-clamp technique in the whole-cell configuration, we studied the effect of a nitric oxide (NO) donor, nitroglycerin (NG), on outward transmembrane ion current in isolated smooth muscle cells (SMC) of the main pulmonary artery of the rabbit. We also studied the characteristics of unitary high-conductance Ca2+-dependent K+ channels (KCa channels) in the SMC membrane in the cell-attached and outside-out configurations. Nitroglycerin in a 10 M concentration increased the amplitude and intensified oscillations of outward transmembrane current induced by step depolarization. In this case, the threshold of activation of the current (–40 mV) did not change. If the potential was +70 mV, the transmembrane current in the presence of NG increased, as compared with the control, by 32.6 ± 19.4% (n = 6), on average. Simultaneous addition of 10 M NG and 1 mM tetraethylammonium chloride (TEA), a blocker of KCa channels, to the external solution at the potential of +70 mV decreased the amplitude of outward transmembrane current with respect to the control by 25.2 ± 11% (n = 6) and suppressed oscillations of this current. In the series of experiments carried out in the outside-out configuration (concentration of K+ ions in the external solution was 5.9 mM), we calculated the conductance of a single KCa channel, which was approximately 150 pS. In the case where the potential was equal to +40 mV, 1 mM TEA suppressed completely the current through unitary KCa channels. In the series of experiments performed in the cell-attached configuration, 100 M NG to a considerable extent intensified the activity of unitary high-conductance KCa channels by increasing the probability of the channel open state (P
0), on average, by 80 ± 1%, as compared with the control. In this case, NG did not influence the conductance of single KCa channels. We concluded that the NO donor NG increases the amplitude of outward transmembrane current in SMC of the rabbit main pulmonary artery by stimulation of the activity of TEA-sensitive high-conductance KCa channels. Our experiments carried out on single KCa channels demonstrated that the activating effect of NG on KCa channels is realized at the expense of an increase in the P
0 of these channels, but not of a change in the conductance of single channels. 相似文献
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