scholarly journals Extremely Potent Block of Bacterial Voltage-Gated Sodium Channels by µ-Conotoxin PIIIA

Marine Drugs ◽  
2019 ◽  
Vol 17 (9) ◽  
pp. 510 ◽  
Author(s):  
Rocio K. Finol-Urdaneta ◽  
Jeffrey R. McArthur ◽  
Vyacheslav S. Korkosh ◽  
Sun Huang ◽  
Denis McMaster ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Sodium Channel ◽  
Sodium Channels ◽  
Cell Voltage ◽  
Hill Coefficient ◽  
Dependent Manner ◽  
Binding Modes ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels ◽  
Conducting Pathway

µ-Conotoxin PIIIA, in the sub-picomolar, range inhibits the archetypal bacterial sodium channel NaChBac (NavBh) in a voltage- and use-dependent manner. Peptide µ-conotoxins were first recognized as potent components of the venoms of fish-hunting cone snails that selectively inhibit voltage-gated skeletal muscle sodium channels, thus preventing muscle contraction. Intriguingly, computer simulations predicted that PIIIA binds to prokaryotic channel NavAb with much higher affinity than to fish (and other vertebrates) skeletal muscle sodium channel (Nav 1.4). Here, using whole-cell voltage clamp, we demonstrate that PIIIA inhibits NavBac mediated currents even more potently than predicted. From concentration-response data, with [PIIIA] varying more than 6 orders of magnitude (10−12 to 10−5 M), we estimated an IC50 = ~5 pM, maximal block of 0.95 and a Hill coefficient of 0.81 for the inhibition of peak currents. Inhibition was stronger at depolarized holding potentials and was modulated by the frequency and duration of the stimulation pulses. An important feature of the PIIIA action was acceleration of macroscopic inactivation. Docking of PIIIA in a NaChBac (NavBh) model revealed two interconvertible binding modes. In one mode, PIIIA sterically and electrostatically blocks the permeation pathway. In a second mode, apparent stabilization of the inactivated state was achieved by PIIIA binding between P2 helices and trans-membrane S5s from adjacent channel subunits, partially occluding the outer pore. Together, our experimental and computational results suggest that, besides blocking the channel-mediated currents by directly occluding the conducting pathway, PIIIA may also change the relative populations of conducting (activated) and non-conducting (inactivated) states.

scholarly journals Sodium Channel Nav1.3 Is Expressed by Polymorphonuclear Neutrophils during Mouse Heart and Kidney Ischemia InVivo and Regulates Adhesion, Transmigration, and Chemotaxis of Human and Mouse Neutrophils In Vitro

2018 ◽  
Vol 128 (6) ◽  
pp. 1151-1166 ◽  
Author(s):  
Marit Poffers ◽  
Nathalie Bühne ◽  
Christine Herzog ◽  
Anja Thorenz ◽  
Rongjun Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Sodium Channel ◽  
Sodium Channels ◽  
Polymorphonuclear Neutrophils ◽  
Human Neutrophils ◽  
Mouse Heart ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels

Abstract Background Voltage-gated sodium channels generate action potentials in excitable cells, but they have also been attributed noncanonical roles in nonexcitable cells. We hypothesize that voltage-gated sodium channels play a functional role during extravasation of neutrophils. Methods Expression of voltage-gated sodium channels was analyzed by polymerase chain reaction. Distribution of Nav1.3 was determined by immunofluorescence and flow cytometry in mouse models of ischemic heart and kidney injury. Adhesion, transmigration, and chemotaxis of neutrophils to endothelial cells and collagen were investigated with voltage-gated sodium channel inhibitors and lidocaine in vitro. Sodium currents were examined with a whole cell patch clamp. Results Mouse and human neutrophils express multiple voltage-gated sodium channels. Only Nav1.3 was detected in neutrophils recruited to ischemic mouse heart (25 ± 7%, n = 14) and kidney (19 ± 2%, n = 6) in vivo. Endothelial adhesion of mouse neutrophils was reduced by tetrodotoxin (56 ± 9%, unselective Nav-inhibitor), ICA121431 (53 ± 10%), and Pterinotoxin-2 (55 ± 9%; preferential inhibitors of Nav1.3, n = 10). Tetrodotoxin (56 ± 19%), ICA121431 (62 ± 22%), and Pterinotoxin-2 (59 ± 22%) reduced transmigration of human neutrophils through endothelial cells, and also prevented chemotactic migration (n = 60, 3 × 20 cells). Lidocaine reduced neutrophil adhesion to 60 ± 9% (n = 10) and transmigration to 54 ± 8% (n = 9). The effect of lidocaine was not increased by ICA121431 or Pterinotoxin-2. Conclusions Nav1.3 is expressed in neutrophils in vivo; regulates attachment, transmigration, and chemotaxis in vitro; and may serve as a relevant target for antiinflammatory effects of lidocaine.

scholarly journals Axons provide the secretory machinery for trafficking of voltage-gated sodium channels in peripheral nerve

2016 ◽  
Vol 113 (7) ◽  
pp. 1823-1828 ◽  
Author(s):  
Carolina González ◽  
José Cánovas ◽  
Javiera Fresno ◽  
Eduardo Couve ◽  
Felipe A. Court ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Sodium Channel ◽  
Sodium Channels ◽  
Neural Function ◽  
Axonal Membrane ◽  
Local Synthesis ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels

The regulation of the axonal proteome is key to generate and maintain neural function. Fast and slow axoplasmic waves have been known for decades, but alternative mechanisms to control the abundance of axonal proteins based on local synthesis have also been identified. The presence of the endoplasmic reticulum has been documented in peripheral axons, but it is still unknown whether this localized organelle participates in the delivery of axonal membrane proteins. Voltage-gated sodium channels are responsible for action potentials and are mostly concentrated in the axon initial segment and nodes of Ranvier. Despite their fundamental role, little is known about the intracellular trafficking mechanisms that govern their availability in mature axons. Here we describe the secretory machinery in axons and its contribution to plasma membrane delivery of sodium channels. The distribution of axonal secretory components was evaluated in axons of the sciatic nerve and in spinal nerve axons after in vivo electroporation. Intracellular protein trafficking was pharmacologically blocked in vivo and in vitro. Axonal voltage-gated sodium channel mRNA and local trafficking were examined by RT-PCR and a retention-release methodology. We demonstrate that mature axons contain components of the endoplasmic reticulum and other biosynthetic organelles. Axonal organelles and sodium channel localization are sensitive to local blockade of the endoplasmic reticulum to Golgi transport. More importantly, secretory organelles are capable of delivering sodium channels to the plasma membrane in isolated axons, demonstrating an intrinsic capacity of the axonal biosynthetic route in regulating the axonal proteome in mammalian axons.

scholarly journals Tracking S4 movement by gating pore currents in the bacterial sodium channel NaChBac

2014 ◽  
Vol 144 (2) ◽  
pp. 147-157 ◽  
Author(s):  
Tamer M. Gamal El-Din ◽  
Todd Scheuer ◽  
William A. Catterall
Keyword(s):  
Membrane Potential ◽  
Sodium Channel ◽  
Sodium Channels ◽  
Voltage Dependence ◽  
Periodic Paralysis ◽  
Membrane Potentials ◽  
Structural Basis ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels ◽  
Gating Charges

Voltage-gated sodium channels mediate the initiation and propagation of action potentials in excitable cells. Transmembrane segment S4 of voltage-gated sodium channels resides in a gating pore where it senses the membrane potential and controls channel gating. Substitution of individual S4 arginine gating charges (R1–R3) with smaller amino acids allows ionic currents to flow through the mutant gating pore, and these gating pore currents are pathogenic in some skeletal muscle periodic paralysis syndromes. The voltage dependence of gating pore currents provides information about the transmembrane position of the gating charges as S4 moves in response to membrane potential. Here we studied gating pore current in mutants of the homotetrameric bacterial sodium channel NaChBac in which individual arginine gating charges were replaced by cysteine. Gating pore current was observed for each mutant channel, but with different voltage-dependent properties. Mutating the first (R1C) or second (R2C) arginine to cysteine resulted in gating pore current at hyperpolarized membrane potentials, where the channels are in resting states, but not at depolarized potentials, where the channels are activated. Conversely, the R3C gating pore is closed at hyperpolarized membrane potentials and opens with channel activation. Negative conditioning pulses revealed time-dependent deactivation of the R3C gating pore at the most hyperpolarized potentials. Our results show sequential voltage dependence of activation of gating pore current from R1 to R3 and support stepwise outward movement of the substituted cysteines through the narrow portion of the gating pore that is sealed by the arginine side chains in the wild-type channel. This pattern of voltage dependence of gating pore current is consistent with a sliding movement of the S4 helix through the gating pore. Through comparison with high-resolution models of the voltage sensor of bacterial sodium channels, these results shed light on the structural basis for pathogenic gating pore currents in periodic paralysis syndromes.

scholarly journals Block of Voltage-Gated Sodium Channels by Atomoxetine in a State- and Use-dependent Manner

2021 ◽  
Vol 12 ◽  
Author(s):  
Karl Josef Föhr ◽  
Ariadni Nastos ◽  
Michael Fauler ◽  
Thomas Zimmer ◽  
Bettina Jungwirth ◽  
...  
Keyword(s):  
Sodium Channels ◽  
Heart Muscle ◽  
Brain Regions ◽  
Clinical Effects ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Human Embryonic Kidney Cells ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels ◽  
Slow Kinetics

Atomoxetine, a neuroactive drug, is approved for the treatment of attention-deficit/hyperactivity disorder (ADHD). It is primarily known as a high affinity blocker of the noradrenaline transporter, whereby its application leads to an increased level of the corresponding neurotransmitter in different brain regions. However, the concentrations used to obtain clinical effects are much higher than those which are required to block the transporter system. Thus, off-target effects are likely to occur. In this way, we previously identified atomoxetine as blocker of NMDA receptors. As many psychotropic drugs give rise to sudden death of cardiac origin, we now tested the hypothesis whether atomoxetine also interacts with voltage-gated sodium channels of heart muscle type in clinically relevant concentrations. Electrophysiological experiments were performed by means of the patch-clamp technique at human heart muscle sodium channels (hNav1.5) heterogeneously expressed in human embryonic kidney cells. Atomoxetine inhibited sodium channels in a state- and use-dependent manner. Atomoxetine had only a weak affinity for the resting state of the hNav1.5 (Kr: ∼ 120 µM). The efficacy of atomoxetine strongly increased with membrane depolarization, indicating that the inactivated state is an important target. A hallmark of this drug was its slow interaction. By use of different experimental settings, we concluded that the interaction occurs with the slow inactivated state as well as by slow kinetics with the fast-inactivated state. Half-maximal effective concentrations (2–3 µM) were well within the concentration range found in plasma of treated patients. Atomoxetine also interacted with the open channel. However, the interaction was not fast enough to accelerate the time constant of fast inactivation. Nevertheless, when using the inactivation-deficient hNav1.5_I408W_L409C_A410W mutant, we found that the persistent late current was blocked half maximal at about 3 µM atomoxetine. The interaction most probably occurred via the local anesthetic binding site. Atomoxetine inhibited sodium channels at a similar concentration as it is used for the treatment of ADHD. Due to its slow interaction and by inhibiting the late current, it potentially exerts antiarrhythmic properties.

scholarly journals Point-Mutations in Skeletal Muscle Voltage-Gated Sodium Channels Confer Resistance to Tetrodotoxin: But at a Cost?

2016 ◽  
Vol 110 (3) ◽  
pp. 436a-437a ◽  
Author(s):  
Robert E. del Carlo ◽  
Normand Leblanc ◽  
Edmund D. Brodie ◽  
Chis R. Feldman
Keyword(s):  
Skeletal Muscle ◽  
Sodium Channels ◽  
Point Mutations ◽  
Voltage Gated ◽  
Voltage Gated Sodium Channels

Voltage-Gated Sodium Channel β1 Gene: An Overview

2021 ◽  
pp. 1-9
Author(s):  
Hisham Al-Ward ◽  
Chun-Yang Liu ◽  
Ning Liu ◽  
Fahmi Shaher ◽  
Murad Al-Nusaif ◽  
...  
Keyword(s):  
Sodium Channel ◽  
Sodium Channels ◽  
Brugada Syndrome ◽  
Protein Complexes ◽  
Sodium Ion ◽  
Dravet Syndrome ◽  
Β Subunit ◽  
Voltage Gated ◽  
Physiological Processes ◽  
Voltage Gated Sodium Channels

<b><i>Background:</i></b> Voltage-gated sodium channels are protein complexes composed of 2 subunits, namely, pore-forming α- and regulatory β-subunits. A β-subunit consists of 5 proteins encoded by 4 genes (i.e., <i>SCN1B–SCN4B</i>). <b><i>Summary:</i></b> β<sub>1</sub>-Subunits regulate sodium ion channel functions, including gating properties, subcellular localization, and kinetics. <b><i>Key Message:</i></b> Sodium channel β<sub>1</sub>- and its variant β<sub>1B</sub>-subunits are encoded by <i>SCN1B</i>. These variants are associated with many human diseases, such as epilepsy, Brugada syndrome, Dravet syndrome, and cancers. On the basis of previous research, we aimed to provide an overview of the structure, expression, and involvement of <i>SCN1B</i> in physiological processes and focused on its role in diseases.

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