The Experts below are selected from a list of 339 Experts worldwide ranked by ideXlab platform
Georgi V. Petkov - One of the best experts on this subject based on the ideXlab platform.
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Regulation of Guinea Pig Detrusor Smooth Muscle Excitability by 17β-Estradiol: The Role of the Large Conductance Voltage- and Ca2+-Activated K+ Channels.
PLOS ONE, 2015Co-Authors: Aaron Provence, Shankar P. Parajuli, Kiril Hristov, Georgi V. PetkovAbstract:Estrogen replacement therapies have been suggested to be beneficial in alleviating symptoms of overactive bladder. However, the precise regulatory mechanisms of estrogen in urinary bladder smooth Muscle (UBSM) at the cellular level remain unknown. Large conductance voltage- and Ca2+-activated K+ (BK) channels, which are key regulators of UBSM function, are suggested to be non-genomic targets of estrogens. This study provides an electrophysiological investigation into the role of UBSM BK channels as direct targets for 17β-estradiol, the principle estrogen in human circulation. Single BK channel recordings on inside-out excised membrane patches and perforated whole cell patch-clamp were applied in combination with the BK channel selective inhibitor paxilline to elucidate the mechanism of regulation of BK channel activity by 17β-estradiol in freshly-isolated guinea pig UBSM cells. 17β-Estradiol (100 nM) significantly increased the amplitude of depolarization-induced whole cell steady-state BK currents and the frequency of spontaneous transient BK currents in freshly-isolated UBSM cells. The increase in whole cell BK currents by 17β-estradiol was eliminated upon blocking BK channels with paxilline. 17β-Estradiol (100 nM) significantly increased (~3-fold) the single BK channel open probability, indicating direct 17β-estradiol-BK channel interactions. 17β-Estradiol (100 nM) caused a significant hyperpolarization of the membrane potential of UBSM cells, and this hyperpolarization was reversed by blocking the BK channels with paxilline. 17β-Estradiol (100 nM) had no effects on L-type voltage-gated Ca2+ channel currents recorded under perforated patch-clamp conditions. This study reveals a new regulatory mechanism in the urinary bladder whereby BK channels are directly activated by 17β-estradiol to reduce UBSM cell Excitability.
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suppression of human detrusor smooth Muscle Excitability and contractility via pharmacological activation of large conductance ca2 activated k channels
American Journal of Physiology-cell Physiology, 2012Co-Authors: Kiril L Hristov, Georgi V. Petkov, Shankar P. Parajuli, Rupal P Soder, Qiuping Cheng, Eric S RovnerAbstract:Overactive bladder syndrome is frequently associated with increased detrusor smooth Muscle (DSM) contractility. We tested the hypothesis that pharmacological activation of the large-conductance voltage- and Ca2+-activated K+ (BK) channel with NS-1619, a selective BK channel opener, reduces the Excitability and contractility of human DSM. We used the amphotericin-perforated whole cell patch-clamp technique on freshly isolated human DSM cells, live-cell Ca2+ imaging, and isometric DSM tension recordings of human DSM strips obtained from open bladder surgeries. NS-1619 (30 μM) significantly increased the amplitude of the voltage step-induced whole cell BK currents, and this effect was abolished by pretreatment with 200 nM iberiotoxin (IBTX), a selective BK channel inhibitor. In current-clamp mode, NS-1619 (30 μM) significantly hyperpolarized the resting membrane potential, and the hyperpolarization was reversed by IBTX (200 nM). NS-1619 (30 μM) significantly decreased the intracellular Ca2+ level in isolated human DSM cells. BK channel activation with NS-1619 (30 μM) significantly inhibited the amplitude, Muscle force, frequency, duration, and tone of the spontaneous phasic and pharmacologically induced DSM contractions from human DSM isolated strips. IBTX (200 nM) suppressed the inhibitory effects of NS-1619 on spontaneous contractions. The amplitude of electrical field stimulation (0.5–50 Hz)-induced contractions was significantly reduced by NS-1619 (30 μM). Our data suggest that pharmacological activation of BK channels could represent a novel treatment option to control bladder dysfunction in humans.
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pharmacological activation of small conductance calcium activated potassium channels with naphtho 1 2 d thiazol 2 ylamine decreases guinea pig detrusor smooth Muscle Excitability and contractility
Journal of Pharmacology and Experimental Therapeutics, 2012Co-Authors: Shankar P. Parajuli, Kiril L Hristov, Rupal P Soder, Georgi V. PetkovAbstract:Small conductance Ca2+-activated K+ (SK) and intermediate conductance Ca2+-activated K+ (IK) channels are thought to be involved in detrusor smooth Muscle (DSM) Excitability and contractility. Using naphtho[1,2-d]thiazol-2-ylamine (SKA-31), a novel and highly specific SK/IK channel activator, we investigated whether pharmacological activation of SK/IK channels reduced guinea pig DSM Excitability and contractility. We detected the expression of all known isoforms of SK (SK1-SK3) and IK channels at mRNA and protein levels in DSM by single-cell reverse transcription-polymerase chain reaction and Western blot. Using the perforated patch-clamp technique on freshly isolated DSM cells, we observed that SKA-31 (10 μM) increased SK currents, which were blocked by apamin (1 μM), a selective SK channel inhibitor. In current-clamp mode, SKA-31 (10 μM) hyperpolarized the cell resting membrane potential, which was blocked by apamin (1 μM) but not by 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34) (1 μM), a selective IK channel inhibitor. SKA-31 (10 nM-10 μM) significantly inhibited the spontaneous phasic contraction amplitude, frequency, duration, and Muscle force in DSM isolated strips. The SKA-31 inhibitory effects on DSM contractility were blocked by apamin (1 μM) but not by TRAM-34 (1 μM), which did not per se significantly affect DSM spontaneous contractility. SK channel activation with SKA-31 reduced contractions evoked by electrical field stimulation. SKA-31 effects were reversible upon washout. In conclusion, SK channels, but not IK channels, mediate SKA-31 effects in guinea pig DSM. Pharmacological activation of SK channels reduces DSM Excitability and contractility and therefore may provide a novel therapeutic approach for controlling bladder dysfunction.
Thomas Holm Pedersen - One of the best experts on this subject based on the ideXlab platform.
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depletion of atp limits membrane Excitability of skeletal Muscle by increasing both clc1 open probability and membrane conductance
Frontiers in Neurology, 2020Co-Authors: Pieter Arnold Leermakers, Frank Vincenzo De Paoli, Kamilla Lohde Tordrup Dybdahl, Kristian Soborg Husted, Anders Riisager, Tomas Pinos, John Vissing, Thomas Krag, Thomas Holm PedersenAbstract:Activation of skeletal Muscle contractions require that action potentials can be excited and propagated along the Muscle fibers. Recent studies have revealed that Muscle fiber Excitability is regulated during repeated firing of action potentials by cellular signaling systems that control the function of ion channel that determine the resting membrane conductance (G (m) ). In fast-twitch Muscle, prolonged firing of action potentials triggers a marked increase in G (m) , reducing Muscle fiber Excitability and causing action potential failure. Both ClC-1 and K(ATP) ion channels contribute to this G (m) rise, but the exact molecular regulation underlying their activation remains unclear. Studies in expression systems have revealed that ClC-1 is able to bind adenosine nucleotides, and that low adenosine nucleotide levels result in ClC-1 activation. In three series of experiments, this study aimed to explore whether ClC-1 is also regulated by adenosine nucleotides in native skeletal Muscle fibers, and whether the adenosine nucleotide sensitivity of ClC-1 could explain the rise in G (m) Muscle fibers during prolonged action potential firing. First, whole cell patch clamping of mouse Muscle fibers demonstrated that ClC-1 activation shifted in the hyperpolarized direction when clamping pipette solution contained 0 mM ATP compared with 5 mM ATP. Second, three-electrode G (m) measurement during Muscle fiber stimulation showed that glycolysis inhibition, with 2-deoxy-glucose or iodoacetate, resulted in an accelerated and rapid >400% G (m) rise during short periods of repeated action potential firing in both fast-twitch and slow-twitch rat, and in human Muscle fibers. Moreover, ClC-1 inhibition with 9-anthracenecarboxylic acid resulted in either an absence or blunted G (m) rise during action potential firing in human Muscle fibers. Third, G (m) measurement during repeated action potential firing in Muscle fibers from a murine McArdle disease model suggest that the rise in G (m) was accelerated in a subset of fibers. Together, these results are compatible with ClC-1 function being regulated by the level of adenosine nucleotides in native tissue, and that the channel operates as a sensor of skeletal Muscle metabolic state, limiting Muscle Excitability when energy status is low.
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chloride channels take center stage in acute regulation of Excitability in skeletal Muscle implications for fatigue
Physiology, 2017Co-Authors: Ole Baekgaard Nielsen, Frank Vincenzo De Paoli, Anders Riisager, Thomas Holm PedersenAbstract:Initiation and propagation of action potentials in Muscle fibers is a key element in the transmission of activating motor input from the central nervous system to their contractile apparatus, and maintenance of Excitability is therefore paramount for their endurance during work. Here, we review current knowledge about the acute regulation of ClC-1 channels in active Muscles and its importance for Muscle Excitability, function, and fatigue.
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an analysis of the relationships between subthreshold electrical properties and Excitability in skeletal Muscle
The Journal of General Physiology, 2011Co-Authors: Thomas Holm Pedersen, Christopher L H Huang, James A FraserAbstract:Skeletal Muscle activation requires action potential (AP) initiation followed by its sarcolemmal propagation and tubular excitation to trigger Ca2+ release and contraction. Recent studies demonstrate that ion channels underlying the resting membrane conductance (GM) of fast-twitch mammalian Muscle fibers are highly regulated during Muscle activity. Thus, onset of activity reduces GM, whereas prolonged activity can markedly elevate GM. Although these observations implicate GM regulation in control of Muscle Excitability, classical theoretical studies in un-myelinated axons predict little influence of GM on membrane Excitability. However, surface membrane morphologies differ markedly between un-myelinated axons and Muscle fibers, predominantly because of the tubular (t)-system of Muscle fibers. This study develops a linear circuit model of mammalian Muscle fiber and uses this to assess the role of subthreshold electrical properties, including GM changes during Muscle activity, for AP initiation, AP propagation, and t-system excitation. Experimental observations of frequency-dependent length constant and membrane-phase properties in fast-twitch rat fibers could only be replicated by models that included t-system luminal resistances. Having quantified these resistances, the resulting models showed enhanced conduction velocity of passive current flow also implicating elevated AP propagation velocity. Furthermore, the resistances filter passive currents such that higher frequency current components would determine sarcolemma AP conduction velocity, whereas lower frequency components excite t-system APs. Because GM modulation affects only the low-frequency membrane impedance, the GM changes in active Muscle would predominantly affect neuromuscular transmission and low-frequency t-system excitation while exerting little influence on the high-frequency process of sarcolemmal AP propagation. This physiological role of GM regulation was increased by high Cl− permeability, as in Muscle endplate regions, and by increased extracellular [K+], as observed in working Muscle. Thus, reduced GM at the onset of exercise would enhance t-system excitation and neuromuscular transmission, whereas elevated GM after sustained activity would inhibit these processes and thereby accentuate Muscle fatigue.
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lactate per se improves the Excitability of depolarized rat skeletal Muscle by reducing the cl conductance
The Journal of Physiology, 2010Co-Authors: Frank Vincenzo De Paoli, Thomas Holm Pedersen, Niels Ortenblad, Rasmus Nyholm Jorgensen, Ole Baekgaard NielsenAbstract:Studies on rats have shown that lactic acid can improve Excitability and function of depolarized Muscles. The effect has been related to the ensuing reduction in intracellular pH causing inhibition of Muscle fibre Cl− channels. However, since several carboxylic acids with structural similarities to lactate can inhibit Muscle Cl− channels it is possible that lactate per se can increase Muscle Excitability by exerting a direct effect on these channels. We therefore examined the effects of lactate on the function of intact Muscles and skinned fibres together with effects on pH and Cl− conductance (Gcl). In Muscles where extracellular compound action potentials (M-waves) and tetanic force response to excitation were reduced by (mean ±s.e.m.) 82 ± 4% and 83 ± 2%, respectively, by depolarization with 11 mm extracellular K+, both M-waves and force exhibited an up to 4-fold increase when 20 mm lactate was added. This effect was present already at 5 mm and saturated at 15 mm lactate, and was associated with a 31% reduction in GCl. The effects of lactate were completely blocked by Cl− channel inhibition or use of Cl−-free solutions. Finally, both experiments where effects of lactate on intracellular pH in intact Muscles were mimicked by increased CO2 tension and experiments with skinned fibres showed that the effects of lactate could not be related to reduced intracellular pH. It is concluded that addition of lactate can inhibit ClC-1 Cl− channels and increase the Excitability and contractile function of depolarized rat Muscles via mechanisms not related to a reduction in intracellular pH.
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comparison of regulated passive membrane conductance in action potential firing fast and slow twitch Muscle
The Journal of General Physiology, 2009Co-Authors: Thomas Holm Pedersen, William Macdonald, Frank Vincenzo De Paoli, Iman S Gurung, Ole Baekgaard NielsenAbstract:In several pathological and experimental conditions, the passive membrane conductance of Muscle fibers (Gm) and their Excitability are inversely related. Despite this capacity of Gm to determine Muscle Excitability, its regulation in active Muscle fibers is largely unexplored. In this issue, our previous study (Pedersen et al. 2009. J. Gen. Physiol. doi:10.1085/jgp.200910291) established a technique with which biphasic regulation of Gm in action potential (AP)-firing fast-twitch fibers of rat extensor digitorum longus Muscles was identified and characterized with temporal resolution of seconds. This showed that AP firing initially reduced Gm via ClC-1 channel inhibition but after ∼1,800 APs, Gm rose substantially, causing AP excitation failure. This late increase of Gm reflected activation of ClC-1 and KATP channels. The present study has explored regulation of Gm in AP-firing slow-twitch fibers of soleus Muscle and compared it to Gm dynamics in fast-twitch fibers. It further explored aspects of the cellular signaling that conveyed regulation of Gm in AP-firing fibers. Thus, in both fiber types, AP firing first triggered protein kinase C (PKC)-dependent ClC-1 channel inhibition that reduced Gm by ∼50%. Experiments with dantrolene showed that AP-triggered SR Ca2+ release activated this PKC-mediated ClC-1 channel inhibition that was associated with reduced rheobase current and improved function of depolarized Muscles, indicating that the reduced Gm enhanced Muscle fiber Excitability. In fast-twitch fibers, the late rise in Gm was accelerated by glucose-free conditions, whereas it was postponed when intermittent resting periods were introduced during AP firing. Remarkably, elevation of Gm was never encountered in AP-firing slow-twitch fibers, even after 15,000 APs. These observations implicate metabolic depression in the elevation of Gm in AP-firing fast-twitch fibers. It is concluded that regulation of Gm is a general phenomenon in AP-firing Muscle, and that differences in Gm regulation may contribute to the different phenotypes of fast- and slow-twitch Muscle.
Ole Baekgaard Nielsen - One of the best experts on this subject based on the ideXlab platform.
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chloride channels take center stage in acute regulation of Excitability in skeletal Muscle implications for fatigue
Physiology, 2017Co-Authors: Ole Baekgaard Nielsen, Frank Vincenzo De Paoli, Anders Riisager, Thomas Holm PedersenAbstract:Initiation and propagation of action potentials in Muscle fibers is a key element in the transmission of activating motor input from the central nervous system to their contractile apparatus, and maintenance of Excitability is therefore paramount for their endurance during work. Here, we review current knowledge about the acute regulation of ClC-1 channels in active Muscles and its importance for Muscle Excitability, function, and fatigue.
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lactate per se improves the Excitability of depolarized rat skeletal Muscle by reducing the cl conductance
The Journal of Physiology, 2010Co-Authors: Frank Vincenzo De Paoli, Thomas Holm Pedersen, Niels Ortenblad, Rasmus Nyholm Jorgensen, Ole Baekgaard NielsenAbstract:Studies on rats have shown that lactic acid can improve Excitability and function of depolarized Muscles. The effect has been related to the ensuing reduction in intracellular pH causing inhibition of Muscle fibre Cl− channels. However, since several carboxylic acids with structural similarities to lactate can inhibit Muscle Cl− channels it is possible that lactate per se can increase Muscle Excitability by exerting a direct effect on these channels. We therefore examined the effects of lactate on the function of intact Muscles and skinned fibres together with effects on pH and Cl− conductance (Gcl). In Muscles where extracellular compound action potentials (M-waves) and tetanic force response to excitation were reduced by (mean ±s.e.m.) 82 ± 4% and 83 ± 2%, respectively, by depolarization with 11 mm extracellular K+, both M-waves and force exhibited an up to 4-fold increase when 20 mm lactate was added. This effect was present already at 5 mm and saturated at 15 mm lactate, and was associated with a 31% reduction in GCl. The effects of lactate were completely blocked by Cl− channel inhibition or use of Cl−-free solutions. Finally, both experiments where effects of lactate on intracellular pH in intact Muscles were mimicked by increased CO2 tension and experiments with skinned fibres showed that the effects of lactate could not be related to reduced intracellular pH. It is concluded that addition of lactate can inhibit ClC-1 Cl− channels and increase the Excitability and contractile function of depolarized rat Muscles via mechanisms not related to a reduction in intracellular pH.
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comparison of regulated passive membrane conductance in action potential firing fast and slow twitch Muscle
The Journal of General Physiology, 2009Co-Authors: Thomas Holm Pedersen, William Macdonald, Frank Vincenzo De Paoli, Iman S Gurung, Ole Baekgaard NielsenAbstract:In several pathological and experimental conditions, the passive membrane conductance of Muscle fibers (Gm) and their Excitability are inversely related. Despite this capacity of Gm to determine Muscle Excitability, its regulation in active Muscle fibers is largely unexplored. In this issue, our previous study (Pedersen et al. 2009. J. Gen. Physiol. doi:10.1085/jgp.200910291) established a technique with which biphasic regulation of Gm in action potential (AP)-firing fast-twitch fibers of rat extensor digitorum longus Muscles was identified and characterized with temporal resolution of seconds. This showed that AP firing initially reduced Gm via ClC-1 channel inhibition but after ∼1,800 APs, Gm rose substantially, causing AP excitation failure. This late increase of Gm reflected activation of ClC-1 and KATP channels. The present study has explored regulation of Gm in AP-firing slow-twitch fibers of soleus Muscle and compared it to Gm dynamics in fast-twitch fibers. It further explored aspects of the cellular signaling that conveyed regulation of Gm in AP-firing fibers. Thus, in both fiber types, AP firing first triggered protein kinase C (PKC)-dependent ClC-1 channel inhibition that reduced Gm by ∼50%. Experiments with dantrolene showed that AP-triggered SR Ca2+ release activated this PKC-mediated ClC-1 channel inhibition that was associated with reduced rheobase current and improved function of depolarized Muscles, indicating that the reduced Gm enhanced Muscle fiber Excitability. In fast-twitch fibers, the late rise in Gm was accelerated by glucose-free conditions, whereas it was postponed when intermittent resting periods were introduced during AP firing. Remarkably, elevation of Gm was never encountered in AP-firing slow-twitch fibers, even after 15,000 APs. These observations implicate metabolic depression in the elevation of Gm in AP-firing fast-twitch fibers. It is concluded that regulation of Gm is a general phenomenon in AP-firing Muscle, and that differences in Gm regulation may contribute to the different phenotypes of fast- and slow-twitch Muscle.
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regulation of clc 1 and katp channels in action potential firing fast twitch Muscle fibers
The Journal of General Physiology, 2009Co-Authors: Thomas Holm Pedersen, Frank Vincenzo De Paoli, J A Flatman, Ole Baekgaard NielsenAbstract:Action potential (AP) excitation requires a transient dominance of depolarizing membrane currents over the repolarizing membrane currents that stabilize the resting membrane potential. Such stabilizing currents, in turn, depend on passive membrane conductance (Gm), which in skeletal Muscle fibers covers membrane conductances for K+ (GK) and Cl− (GCl). Myotonic disorders and studies with metabolically poisoned Muscle have revealed capacities of GK and GCl to inversely interfere with Muscle Excitability. However, whether regulation of GK and GCl occur in AP-firing Muscle under normal physiological conditions is unknown. This study establishes a technique that allows the determination of GCl and GK with a temporal resolution of seconds in AP-firing Muscle fibers. With this approach, we have identified and quantified a biphasic regulation of Gm in active fast-twitch extensor digitorum longus fibers of the rat. Thus, at the onset of AP firing, a reduction in GCl of ∼70% caused Gm to decline by ∼55% in a manner that is well described by a single exponential function characterized by a time constant of ∼200 APs (phase 1). When stimulation was continued beyond ∼1,800 APs, synchronized elevations in GK (∼14-fold) and GCl (∼3-fold) caused Gm to rise sigmoidally to ∼400% of its level before AP firing (phase 2). Phase 2 was often associated with a failure to excite APs. When AP firing was ceased during phase 2, Gm recovered to its level before AP firing in ∼1 min. Experiments with glibenclamide (KATP channel inhibitor) and 9-anthracene carboxylic acid (ClC-1 Cl− channel inhibitor) revealed that the decreased Gm during phase 1 reflected ClC-1 channel inhibition, whereas the massively elevated Gm during phase 2 reflected synchronized openings of ClC-1 and KATP channels. In conclusion, GCl and GK are acutely regulated in AP-firing fast-twitch Muscle fibers. Such regulation may contribute to the physiological control of Excitability in active Muscle.
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additive protective effects of the addition of lactic acid and adrenaline on Excitability and force in isolated rat skeletal Muscle depressed by elevated extracellular k
The Journal of Physiology, 2007Co-Authors: Frank Vincenzo De Paoli, Thomas Holm Pedersen, Kristian Overgaard, Ole Baekgaard NielsenAbstract:During strenuous exercise, extracellular K+ ([K+]o) is increased, which potentially can reduce Muscle Excitability and force production. In addition, exercise leads to accumulation of lactate and H+ and increased levels of circulating catecholamines. Individually, reduced pH and increased catecholamines have been shown to counteract the depressing effect of elevated K+. This study examines (i) whether the effects of addition of lactic acid and adrenaline on the Excitability of isolated Muscles are caused by separate mechanisms and are additive and (ii) whether the effect of adding lactic acid or increasing CO2 is related to a reduction of intra- or extracellular pH. Rat soleus Muscles were incubated at a [K+]o of 15 mm, which reduced tetanic force by 85%. Subsequent addition of 20 mm lactic acid or 10−5m adrenaline led to a small recovery of force, but when added together induced an almost complete force recovery. Compound action potentials showed that the force recovery was associated with recovery of Muscle Excitability. The improved Excitability after addition of adrenaline was associated with increased Na+–K+ pump activity resulting in hyperpolarization and an increase in the chemical Na+ gradient. In contrast, addition of lactic acid had no effect on the membrane potential or the Na+–K+ pump activity, but most likely increased Excitability via a reduction in intracellular pH. It is concluded that the protective effects of acidosis and adrenaline on Muscle Excitability and force took place via different mechanisms and were additive. The results suggest that circulating catecholamines and development of acidosis during exercise may improve the tolerance of Muscles to elevated [K+]o.
Frank Vincenzo De Paoli - One of the best experts on this subject based on the ideXlab platform.
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depletion of atp limits membrane Excitability of skeletal Muscle by increasing both clc1 open probability and membrane conductance
Frontiers in Neurology, 2020Co-Authors: Pieter Arnold Leermakers, Frank Vincenzo De Paoli, Kamilla Lohde Tordrup Dybdahl, Kristian Soborg Husted, Anders Riisager, Tomas Pinos, John Vissing, Thomas Krag, Thomas Holm PedersenAbstract:Activation of skeletal Muscle contractions require that action potentials can be excited and propagated along the Muscle fibers. Recent studies have revealed that Muscle fiber Excitability is regulated during repeated firing of action potentials by cellular signaling systems that control the function of ion channel that determine the resting membrane conductance (G (m) ). In fast-twitch Muscle, prolonged firing of action potentials triggers a marked increase in G (m) , reducing Muscle fiber Excitability and causing action potential failure. Both ClC-1 and K(ATP) ion channels contribute to this G (m) rise, but the exact molecular regulation underlying their activation remains unclear. Studies in expression systems have revealed that ClC-1 is able to bind adenosine nucleotides, and that low adenosine nucleotide levels result in ClC-1 activation. In three series of experiments, this study aimed to explore whether ClC-1 is also regulated by adenosine nucleotides in native skeletal Muscle fibers, and whether the adenosine nucleotide sensitivity of ClC-1 could explain the rise in G (m) Muscle fibers during prolonged action potential firing. First, whole cell patch clamping of mouse Muscle fibers demonstrated that ClC-1 activation shifted in the hyperpolarized direction when clamping pipette solution contained 0 mM ATP compared with 5 mM ATP. Second, three-electrode G (m) measurement during Muscle fiber stimulation showed that glycolysis inhibition, with 2-deoxy-glucose or iodoacetate, resulted in an accelerated and rapid >400% G (m) rise during short periods of repeated action potential firing in both fast-twitch and slow-twitch rat, and in human Muscle fibers. Moreover, ClC-1 inhibition with 9-anthracenecarboxylic acid resulted in either an absence or blunted G (m) rise during action potential firing in human Muscle fibers. Third, G (m) measurement during repeated action potential firing in Muscle fibers from a murine McArdle disease model suggest that the rise in G (m) was accelerated in a subset of fibers. Together, these results are compatible with ClC-1 function being regulated by the level of adenosine nucleotides in native tissue, and that the channel operates as a sensor of skeletal Muscle metabolic state, limiting Muscle Excitability when energy status is low.
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chloride channels take center stage in acute regulation of Excitability in skeletal Muscle implications for fatigue
Physiology, 2017Co-Authors: Ole Baekgaard Nielsen, Frank Vincenzo De Paoli, Anders Riisager, Thomas Holm PedersenAbstract:Initiation and propagation of action potentials in Muscle fibers is a key element in the transmission of activating motor input from the central nervous system to their contractile apparatus, and maintenance of Excitability is therefore paramount for their endurance during work. Here, we review current knowledge about the acute regulation of ClC-1 channels in active Muscles and its importance for Muscle Excitability, function, and fatigue.
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lactate per se improves the Excitability of depolarized rat skeletal Muscle by reducing the cl conductance
The Journal of Physiology, 2010Co-Authors: Frank Vincenzo De Paoli, Thomas Holm Pedersen, Niels Ortenblad, Rasmus Nyholm Jorgensen, Ole Baekgaard NielsenAbstract:Studies on rats have shown that lactic acid can improve Excitability and function of depolarized Muscles. The effect has been related to the ensuing reduction in intracellular pH causing inhibition of Muscle fibre Cl− channels. However, since several carboxylic acids with structural similarities to lactate can inhibit Muscle Cl− channels it is possible that lactate per se can increase Muscle Excitability by exerting a direct effect on these channels. We therefore examined the effects of lactate on the function of intact Muscles and skinned fibres together with effects on pH and Cl− conductance (Gcl). In Muscles where extracellular compound action potentials (M-waves) and tetanic force response to excitation were reduced by (mean ±s.e.m.) 82 ± 4% and 83 ± 2%, respectively, by depolarization with 11 mm extracellular K+, both M-waves and force exhibited an up to 4-fold increase when 20 mm lactate was added. This effect was present already at 5 mm and saturated at 15 mm lactate, and was associated with a 31% reduction in GCl. The effects of lactate were completely blocked by Cl− channel inhibition or use of Cl−-free solutions. Finally, both experiments where effects of lactate on intracellular pH in intact Muscles were mimicked by increased CO2 tension and experiments with skinned fibres showed that the effects of lactate could not be related to reduced intracellular pH. It is concluded that addition of lactate can inhibit ClC-1 Cl− channels and increase the Excitability and contractile function of depolarized rat Muscles via mechanisms not related to a reduction in intracellular pH.
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comparison of regulated passive membrane conductance in action potential firing fast and slow twitch Muscle
The Journal of General Physiology, 2009Co-Authors: Thomas Holm Pedersen, William Macdonald, Frank Vincenzo De Paoli, Iman S Gurung, Ole Baekgaard NielsenAbstract:In several pathological and experimental conditions, the passive membrane conductance of Muscle fibers (Gm) and their Excitability are inversely related. Despite this capacity of Gm to determine Muscle Excitability, its regulation in active Muscle fibers is largely unexplored. In this issue, our previous study (Pedersen et al. 2009. J. Gen. Physiol. doi:10.1085/jgp.200910291) established a technique with which biphasic regulation of Gm in action potential (AP)-firing fast-twitch fibers of rat extensor digitorum longus Muscles was identified and characterized with temporal resolution of seconds. This showed that AP firing initially reduced Gm via ClC-1 channel inhibition but after ∼1,800 APs, Gm rose substantially, causing AP excitation failure. This late increase of Gm reflected activation of ClC-1 and KATP channels. The present study has explored regulation of Gm in AP-firing slow-twitch fibers of soleus Muscle and compared it to Gm dynamics in fast-twitch fibers. It further explored aspects of the cellular signaling that conveyed regulation of Gm in AP-firing fibers. Thus, in both fiber types, AP firing first triggered protein kinase C (PKC)-dependent ClC-1 channel inhibition that reduced Gm by ∼50%. Experiments with dantrolene showed that AP-triggered SR Ca2+ release activated this PKC-mediated ClC-1 channel inhibition that was associated with reduced rheobase current and improved function of depolarized Muscles, indicating that the reduced Gm enhanced Muscle fiber Excitability. In fast-twitch fibers, the late rise in Gm was accelerated by glucose-free conditions, whereas it was postponed when intermittent resting periods were introduced during AP firing. Remarkably, elevation of Gm was never encountered in AP-firing slow-twitch fibers, even after 15,000 APs. These observations implicate metabolic depression in the elevation of Gm in AP-firing fast-twitch fibers. It is concluded that regulation of Gm is a general phenomenon in AP-firing Muscle, and that differences in Gm regulation may contribute to the different phenotypes of fast- and slow-twitch Muscle.
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regulation of clc 1 and katp channels in action potential firing fast twitch Muscle fibers
The Journal of General Physiology, 2009Co-Authors: Thomas Holm Pedersen, Frank Vincenzo De Paoli, J A Flatman, Ole Baekgaard NielsenAbstract:Action potential (AP) excitation requires a transient dominance of depolarizing membrane currents over the repolarizing membrane currents that stabilize the resting membrane potential. Such stabilizing currents, in turn, depend on passive membrane conductance (Gm), which in skeletal Muscle fibers covers membrane conductances for K+ (GK) and Cl− (GCl). Myotonic disorders and studies with metabolically poisoned Muscle have revealed capacities of GK and GCl to inversely interfere with Muscle Excitability. However, whether regulation of GK and GCl occur in AP-firing Muscle under normal physiological conditions is unknown. This study establishes a technique that allows the determination of GCl and GK with a temporal resolution of seconds in AP-firing Muscle fibers. With this approach, we have identified and quantified a biphasic regulation of Gm in active fast-twitch extensor digitorum longus fibers of the rat. Thus, at the onset of AP firing, a reduction in GCl of ∼70% caused Gm to decline by ∼55% in a manner that is well described by a single exponential function characterized by a time constant of ∼200 APs (phase 1). When stimulation was continued beyond ∼1,800 APs, synchronized elevations in GK (∼14-fold) and GCl (∼3-fold) caused Gm to rise sigmoidally to ∼400% of its level before AP firing (phase 2). Phase 2 was often associated with a failure to excite APs. When AP firing was ceased during phase 2, Gm recovered to its level before AP firing in ∼1 min. Experiments with glibenclamide (KATP channel inhibitor) and 9-anthracene carboxylic acid (ClC-1 Cl− channel inhibitor) revealed that the decreased Gm during phase 1 reflected ClC-1 channel inhibition, whereas the massively elevated Gm during phase 2 reflected synchronized openings of ClC-1 and KATP channels. In conclusion, GCl and GK are acutely regulated in AP-firing fast-twitch Muscle fibers. Such regulation may contribute to the physiological control of Excitability in active Muscle.
Shankar P. Parajuli - One of the best experts on this subject based on the ideXlab platform.
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Regulation of Guinea Pig Detrusor Smooth Muscle Excitability by 17β-Estradiol: The Role of the Large Conductance Voltage- and Ca2+-Activated K+ Channels.
PLOS ONE, 2015Co-Authors: Aaron Provence, Shankar P. Parajuli, Kiril Hristov, Georgi V. PetkovAbstract:Estrogen replacement therapies have been suggested to be beneficial in alleviating symptoms of overactive bladder. However, the precise regulatory mechanisms of estrogen in urinary bladder smooth Muscle (UBSM) at the cellular level remain unknown. Large conductance voltage- and Ca2+-activated K+ (BK) channels, which are key regulators of UBSM function, are suggested to be non-genomic targets of estrogens. This study provides an electrophysiological investigation into the role of UBSM BK channels as direct targets for 17β-estradiol, the principle estrogen in human circulation. Single BK channel recordings on inside-out excised membrane patches and perforated whole cell patch-clamp were applied in combination with the BK channel selective inhibitor paxilline to elucidate the mechanism of regulation of BK channel activity by 17β-estradiol in freshly-isolated guinea pig UBSM cells. 17β-Estradiol (100 nM) significantly increased the amplitude of depolarization-induced whole cell steady-state BK currents and the frequency of spontaneous transient BK currents in freshly-isolated UBSM cells. The increase in whole cell BK currents by 17β-estradiol was eliminated upon blocking BK channels with paxilline. 17β-Estradiol (100 nM) significantly increased (~3-fold) the single BK channel open probability, indicating direct 17β-estradiol-BK channel interactions. 17β-Estradiol (100 nM) caused a significant hyperpolarization of the membrane potential of UBSM cells, and this hyperpolarization was reversed by blocking the BK channels with paxilline. 17β-Estradiol (100 nM) had no effects on L-type voltage-gated Ca2+ channel currents recorded under perforated patch-clamp conditions. This study reveals a new regulatory mechanism in the urinary bladder whereby BK channels are directly activated by 17β-estradiol to reduce UBSM cell Excitability.
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suppression of human detrusor smooth Muscle Excitability and contractility via pharmacological activation of large conductance ca2 activated k channels
American Journal of Physiology-cell Physiology, 2012Co-Authors: Kiril L Hristov, Georgi V. Petkov, Shankar P. Parajuli, Rupal P Soder, Qiuping Cheng, Eric S RovnerAbstract:Overactive bladder syndrome is frequently associated with increased detrusor smooth Muscle (DSM) contractility. We tested the hypothesis that pharmacological activation of the large-conductance voltage- and Ca2+-activated K+ (BK) channel with NS-1619, a selective BK channel opener, reduces the Excitability and contractility of human DSM. We used the amphotericin-perforated whole cell patch-clamp technique on freshly isolated human DSM cells, live-cell Ca2+ imaging, and isometric DSM tension recordings of human DSM strips obtained from open bladder surgeries. NS-1619 (30 μM) significantly increased the amplitude of the voltage step-induced whole cell BK currents, and this effect was abolished by pretreatment with 200 nM iberiotoxin (IBTX), a selective BK channel inhibitor. In current-clamp mode, NS-1619 (30 μM) significantly hyperpolarized the resting membrane potential, and the hyperpolarization was reversed by IBTX (200 nM). NS-1619 (30 μM) significantly decreased the intracellular Ca2+ level in isolated human DSM cells. BK channel activation with NS-1619 (30 μM) significantly inhibited the amplitude, Muscle force, frequency, duration, and tone of the spontaneous phasic and pharmacologically induced DSM contractions from human DSM isolated strips. IBTX (200 nM) suppressed the inhibitory effects of NS-1619 on spontaneous contractions. The amplitude of electrical field stimulation (0.5–50 Hz)-induced contractions was significantly reduced by NS-1619 (30 μM). Our data suggest that pharmacological activation of BK channels could represent a novel treatment option to control bladder dysfunction in humans.
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pharmacological activation of small conductance calcium activated potassium channels with naphtho 1 2 d thiazol 2 ylamine decreases guinea pig detrusor smooth Muscle Excitability and contractility
Journal of Pharmacology and Experimental Therapeutics, 2012Co-Authors: Shankar P. Parajuli, Kiril L Hristov, Rupal P Soder, Georgi V. PetkovAbstract:Small conductance Ca2+-activated K+ (SK) and intermediate conductance Ca2+-activated K+ (IK) channels are thought to be involved in detrusor smooth Muscle (DSM) Excitability and contractility. Using naphtho[1,2-d]thiazol-2-ylamine (SKA-31), a novel and highly specific SK/IK channel activator, we investigated whether pharmacological activation of SK/IK channels reduced guinea pig DSM Excitability and contractility. We detected the expression of all known isoforms of SK (SK1-SK3) and IK channels at mRNA and protein levels in DSM by single-cell reverse transcription-polymerase chain reaction and Western blot. Using the perforated patch-clamp technique on freshly isolated DSM cells, we observed that SKA-31 (10 μM) increased SK currents, which were blocked by apamin (1 μM), a selective SK channel inhibitor. In current-clamp mode, SKA-31 (10 μM) hyperpolarized the cell resting membrane potential, which was blocked by apamin (1 μM) but not by 1-[(2-chlorophenyl)diphenylmethyl]-1H-pyrazole (TRAM-34) (1 μM), a selective IK channel inhibitor. SKA-31 (10 nM-10 μM) significantly inhibited the spontaneous phasic contraction amplitude, frequency, duration, and Muscle force in DSM isolated strips. The SKA-31 inhibitory effects on DSM contractility were blocked by apamin (1 μM) but not by TRAM-34 (1 μM), which did not per se significantly affect DSM spontaneous contractility. SK channel activation with SKA-31 reduced contractions evoked by electrical field stimulation. SKA-31 effects were reversible upon washout. In conclusion, SK channels, but not IK channels, mediate SKA-31 effects in guinea pig DSM. Pharmacological activation of SK channels reduces DSM Excitability and contractility and therefore may provide a novel therapeutic approach for controlling bladder dysfunction.
