Free energy dissipation of the spontaneous gating of a single voltage-gated potassium channel

NASA Astrophysics Data System (ADS)

Wang, Jia-Zeng; Wang, Rui-Zhen

2018-02-01

Potassium channels mainly contribute to the resting potential and re-polarizations, with the potassium electrochemical gradient being maintained by the pump Na+/K+-ATPase. In this paper, we construct a stochastic model mimicking the kinetics of a potassium channel, which integrates temporal evolving of the membrane voltage and the spontaneous gating of the channel. Its stationary probability density functions (PDFs) are found to be singular at the boundaries, which result from the fact that the evolving rates of voltage are greater than the gating rates of the channel. We apply PDFs to calculate the power dissipations of the potassium current, the leakage, and the gating currents. On a physical perspective, the essential role of the system is the K+-battery charging the leakage (L-)battery. A part of power will inevitably be dissipated among the process. So, the efficiency of energy transference is calculated.

Cardiac Delayed Rectifier Potassium Channels in Health and Disease

PubMed Central

Chen, Lei; Sampson, Kevin J.; Kass, Robert S.

2016-01-01

Cardiac delayed rectifier potassium channels conduct outward potassium currents during the plateau phase of action potentials and play pivotal roles in cardiac repolarization. These include IKs, IKr and the atrial specific IKur channels. In this chapter, we will review the molecular identities and biophysical properties of these channels. Mutations in the genes encoding delayed rectifiers lead to loss- or gain-of-function phenotypes, disrupt normal cardiac repolarization and result in various cardiac rhythm disorders, including congenital Long QT Syndrome, Short QT Syndrome and familial atrial fibrillation. We will also discuss the possibility and prospect of using delayed rectifier channels as therapeutic targets to manage cardiac arrhythmia. PMID:27261823

Carbamazepine as a novel small molecule corrector of trafficking-impaired ATP-sensitive potassium channels identified in congenital hyperinsulinism.

PubMed

Chen, Pei-Chun; Olson, Erik M; Zhou, Qing; Kryukova, Yelena; Sampson, Heidi M; Thomas, David Y; Shyng, Show-Ling

2013-07-19

ATP-sensitive potassium (KATP) channels consisting of sulfonylurea receptor 1 (SUR1) and the potassium channel Kir6.2 play a key role in insulin secretion by coupling metabolic signals to β-cell membrane potential. Mutations in SUR1 and Kir6.2 that impair channel trafficking to the cell surface lead to loss of channel function and congenital hyperinsulinism. We report that carbamazepine, an anticonvulsant, corrects the trafficking defects of mutant KATP channels previously identified in congenital hyperinsulinism. Strikingly, of the 19 SUR1 mutations examined, only those located in the first transmembrane domain of SUR1 responded to the drug. We show that unlike that reported for several other protein misfolding diseases, carbamazepine did not correct KATP channel trafficking defects by activating autophagy; rather, it directly improved the biogenesis efficiency of mutant channels along the secretory pathway. In addition to its effect on channel trafficking, carbamazepine also inhibited KATP channel activity. Upon subsequent removal of carbamazepine, however, the function of rescued channels was recovered. Importantly, combination of the KATP channel opener diazoxide and carbamazepine led to enhanced mutant channel function without carbamazepine washout. The corrector effect of carbamazepine on mutant KATP channels was also demonstrated in rat and human β-cells with an accompanying increase in channel activity. Our findings identify carbamazepine as a novel small molecule corrector that may be used to restore KATP channel expression and function in a subset of congenital hyperinsulinism patients.

International Union of Basic and Clinical Pharmacology. C. Nomenclature and Properties of Calcium-Activated and Sodium-Activated Potassium Channels.

PubMed

Kaczmarek, Leonard K; Aldrich, Richard W; Chandy, K George; Grissmer, Stephan; Wei, Aguan D; Wulff, Heike

2017-01-01

A subset of potassium channels is regulated primarily by changes in the cytoplasmic concentration of ions, including calcium, sodium, chloride, and protons. The eight members of this subfamily were originally all designated as calcium-activated channels. More recent studies have clarified the gating mechanisms for these channels and have documented that not all members are sensitive to calcium. This article describes the molecular relationships between these channels and provides an introduction to their functional properties. It also introduces a new nomenclature that differentiates between calcium- and sodium-activated potassium channels. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

Slack, Slick, and Sodium-Activated Potassium Channels

PubMed Central

Kaczmarek, Leonard K.

2013-01-01

The Slack and Slick genes encode potassium channels that are very widely expressed in the central nervous system. These channels are activated by elevations in intracellular sodium, such as those that occur during trains of one or more action potentials, or following activation of nonselective cationic neurotransmitter receptors such as AMPA receptors. This review covers the cellular and molecular properties of Slack and Slick channels and compares them with findings on the properties of sodium-activated potassium currents (termed KNa currents) in native neurons. Human mutations in Slack channels produce extremely severe defects in learning and development, suggesting that KNa channels play a central role in neuronal plasticity and intellectual function. PMID:24319675

Plant Ion Channels: Gene Families, Physiology, and Functional Genomics Analyses

PubMed Central

Ward, John M.; Mäser, Pascal; Schroeder, Julian I.

2016-01-01

Distinct potassium, anion, and calcium channels in the plasma membrane and vacuolar membrane of plant cells have been identified and characterized by patch clamping. Primarily owing to advances in Arabidopsis genetics and genomics, and yeast functional complementation, many of the corresponding genes have been identified. Recent advances in our understanding of ion channel genes that mediate signal transduction and ion transport are discussed here. Some plant ion channels, for example, ALMT and SLAC anion channel subunits, are unique. The majority of plant ion channel families exhibit homology to animal genes; such families include both hyperpolarization-and depolarization-activated Shaker-type potassium channels, CLC chloride transporters/channels, cyclic nucleotide–gated channels, and ionotropic glutamate receptor homologs. These plant ion channels offer unique opportunities to analyze the structural mechanisms and functions of ion channels. Here we review gene families of selected plant ion channel classes and discuss unique structure-function aspects and their physiological roles in plant cell signaling and transport. PMID:18842100

Plant ion channels: gene families, physiology, and functional genomics analyses.

PubMed

Ward, John M; Mäser, Pascal; Schroeder, Julian I

2009-01-01

Distinct potassium, anion, and calcium channels in the plasma membrane and vacuolar membrane of plant cells have been identified and characterized by patch clamping. Primarily owing to advances in Arabidopsis genetics and genomics, and yeast functional complementation, many of the corresponding genes have been identified. Recent advances in our understanding of ion channel genes that mediate signal transduction and ion transport are discussed here. Some plant ion channels, for example, ALMT and SLAC anion channel subunits, are unique. The majority of plant ion channel families exhibit homology to animal genes; such families include both hyperpolarization- and depolarization-activated Shaker-type potassium channels, CLC chloride transporters/channels, cyclic nucleotide-gated channels, and ionotropic glutamate receptor homologs. These plant ion channels offer unique opportunities to analyze the structural mechanisms and functions of ion channels. Here we review gene families of selected plant ion channel classes and discuss unique structure-function aspects and their physiological roles in plant cell signaling and transport.

Pharmacological Correction of Trafficking Defects in ATP-sensitive Potassium Channels Caused by Sulfonylurea Receptor 1 Mutations*

PubMed Central

Martin, Gregory M.; Rex, Emily A.; Devaraneni, Prasanna; Denton, Jerod S.; Boodhansingh, Kara E.; DeLeon, Diva D.; Stanley, Charles A.; Shyng, Show-Ling

2016-01-01

ATP-sensitive potassium (KATP) channels play a key role in mediating glucose-stimulated insulin secretion by coupling metabolic signals to β-cell membrane potential. Loss of KATP channel function due to mutations in ABCC8 or KCNJ11, genes encoding the sulfonylurea receptor 1 (SUR1) or the inwardly rectifying potassium channel Kir6.2, respectively, results in congenital hyperinsulinism. Many SUR1 mutations prevent trafficking of channel proteins from the endoplasmic reticulum to the cell surface. Channel inhibitors, including sulfonylureas and carbamazepine, have been shown to correct channel trafficking defects. In the present study, we identified 13 novel SUR1 mutations that cause channel trafficking defects, the majority of which are amenable to pharmacological rescue by glibenclamide and carbamazepine. By contrast, none of the mutant channels were rescued by KATP channel openers. Cross-linking experiments showed that KATP channel inhibitors promoted interactions between the N terminus of Kir6.2 and SUR1, whereas channel openers did not, suggesting the inhibitors enhance intersubunit interactions to overcome channel biogenesis and trafficking defects. Functional studies of rescued mutant channels indicate that most mutants rescued to the cell surface exhibited WT-like sensitivity to ATP, MgADP, and diazoxide. In intact cells, recovery of channel function upon trafficking rescue by reversible sulfonylureas or carbamazepine was facilitated by the KATP channel opener diazoxide. Our study expands the list of KATP channel trafficking mutations whose function can be recovered by pharmacological ligands and provides further insight into the structural mechanism by which channel inhibitors correct channel biogenesis and trafficking defects. PMID:27573238

Pharmacological rescue of trafficking-impaired ATP-sensitive potassium channels

PubMed Central

Martin, Gregory M.; Chen, Pei-Chun; Devaraneni, Prasanna; Shyng, Show-Ling

2013-01-01

ATP-sensitive potassium (KATP) channels link cell metabolism to membrane excitability and are involved in a wide range of physiological processes including hormone secretion, control of vascular tone, and protection of cardiac and neuronal cells against ischemic injuries. In pancreatic β-cells, KATP channels play a key role in glucose-stimulated insulin secretion, and gain or loss of channel function results in neonatal diabetes or congenital hyperinsulinism, respectively. The β-cell KATP channel is formed by co-assembly of four Kir6.2 inwardly rectifying potassium channel subunits encoded by KCNJ11 and four sulfonylurea receptor 1 subunits encoded by ABCC8. Many mutations in ABCC8 or KCNJ11 cause loss of channel function, thus, congenital hyperinsulinism by hampering channel biogenesis and hence trafficking to the cell surface. The trafficking defects caused by a subset of these mutations can be corrected by sulfonylureas, KATP channel antagonists that have long been used to treat type 2 diabetes. More recently, carbamazepine, an anticonvulsant that is thought to target primarily voltage-gated sodium channels has been shown to correct KATP channel trafficking defects. This article reviews studies to date aimed at understanding the mechanisms by which mutations impair channel biogenesis and trafficking and the mechanisms by which pharmacological ligands overcome channel trafficking defects. Insight into channel structure-function relationships and therapeutic implications from these studies are discussed. PMID:24399968

G-protein-coupled inward rectifier potassium channels involved in corticostriatal presynaptic modulation.

PubMed

Meneses, David; Mateos, Verónica; Islas, Gustavo; Barral, Jaime

2015-09-01

Presynaptic modulation has been associated mainly with calcium channels but recent data suggests that inward rectifier potassium channels (K(IR)) also play a role. In this work we set to characterize the role of presynaptic K(IR) channels in corticostriatal synaptic transmission. We elicited synaptic potentials in striatum by stimulating cortical areas and then determined the synaptic responses of corticostriatal synapsis by using paired pulse ratio (PPR) in the presence and absence of several potassium channel blockers. Unspecific potassium channels blockers Ba(2+) and Cs(+) reduced the PPR, suggesting that these channels are presynaptically located. Further pharmacological characterization showed that application of tertiapin-Q, a specific K(IR)3 channel family blocker, also induced a reduction of PPR, suggesting that K(IR)3 channels are present at corticostriatal terminals. In contrast, exposure to Lq2, a specific K(IR)1.1 inward rectifier potassium channel, did not induce any change in PPR suggesting the absence of these channels in the presynaptic corticostriatal terminals. Our results indicate that K(IR)3 channels are functionally expressed at the corticostriatal synapses, since blockage of these channels result in PPR decrease. Our results also help to explain how synaptic activity may become sensitive to extracellular signals mediated by G-protein coupled receptors. A vast repertoire of receptors may influence neurotransmitter release in an indirect manner through regulation of K(IR)3 channels. © 2015 Wiley Periodicals, Inc.

Thylakoid potassium channel is required for efficient photosynthesis in cyanobacteria.

PubMed

Checchetto, Vanessa; Segalla, Anna; Allorent, Guillaume; La Rocca, Nicoletta; Leanza, Luigi; Giacometti, Giorgio Mario; Uozumi, Nobuyuki; Finazzi, Giovanni; Bergantino, Elisabetta; Szabò, Ildikò

2012-07-03

A potassium channel (SynK) of the cyanobacterium Synechocystis sp. PCC 6803, a photoheterotrophic model organism for the study of photosynthesis, has been recently identified and demonstrated to function as a potassium selective channel when expressed in a heterologous system and to be located predominantly to the thylakoid membrane in cyanobacteria. To study its physiological role, a SynK-less knockout mutant was generated and characterized. Fluorimetric experiments indicated that SynK-less cyanobacteria cannot build up a proton gradient as efficiently as WT organisms, suggesting that SynK might be involved in the regulation of the electric component of the proton motive force. Accordingly, measurements of flash-induced cytochrome b(6)f turnover and respiration pointed to a reduced generation of ΔpH and to an altered linear electron transport in mutant cells. The lack of the channel did not cause an altered membrane organization, but decreased growth and modified the photosystem II/photosystem I ratio at high light intensities because of enhanced photosensitivity. These data shed light on the function of a prokaryotic potassium channel and reports evidence, by means of a genetic approach, on the requirement of a thylakoid ion channel for optimal photosynthesis.

Oxidative Stress and Maxi Calcium-Activated Potassium (BK) Channels

PubMed Central

Hermann, Anton; Sitdikova, Guzel F.; Weiger, Thomas M.

2015-01-01

All cells contain ion channels in their outer (plasma) and inner (organelle) membranes. Ion channels, similar to other proteins, are targets of oxidative impact, which modulates ion fluxes across membranes. Subsequently, these ion currents affect electrical excitability, such as action potential discharge (in neurons, muscle, and receptor cells), alteration of the membrane resting potential, synaptic transmission, hormone secretion, muscle contraction or coordination of the cell cycle. In this chapter we summarize effects of oxidative stress and redox mechanisms on some ion channels, in particular on maxi calcium-activated potassium (BK) channels which play an outstanding role in a plethora of physiological and pathophysiological functions in almost all cells and tissues. We first elaborate on some general features of ion channel structure and function and then summarize effects of oxidative alterations of ion channels and their functional consequences. PMID:26287261

Role of ER Export Signals in Controlling Surface Potassium Channel Numbers

NASA Astrophysics Data System (ADS)

Ma, Dzwokai; Zerangue, Noa; Lin, Yu-Fung; Collins, Anthony; Yu, Mei; Jan, Yuh Nung; Yeh Jan, Lily

2001-01-01

Little is known about the identity of endoplasmic reticulum (ER) export signals and how they are used to regulate the number of proteins on the cell surface. Here, we describe two ER export signals that profoundly altered the steady-state distribution of potassium channels and were required for channel localization to the plasma membrane. When transferred to other potassium channels or a G protein-coupled receptor, these ER export signals increased the number of functional proteins on the cell surface. Thus, ER export of membrane proteins is not necessarily limited by folding or assembly, but may be under the control of specific export signals.

Cardiac Delayed Rectifier Potassium Channels in Health and Disease.

PubMed

Chen, Lei; Sampson, Kevin J; Kass, Robert S

2016-06-01

Cardiac delayed rectifier potassium channels conduct outward potassium currents during the plateau phase of action potentials and play pivotal roles in cardiac repolarization. These include IKs, IKr and the atrial specific IKur channels. In this article, we will review their molecular identities and biophysical properties. Mutations in the genes encoding delayed rectifiers lead to loss- or gain-of-function phenotypes, disrupt normal cardiac repolarization and result in various cardiac rhythm disorders, including congenital Long QT Syndrome, Short QT Syndrome and familial atrial fibrillation. We will also discuss the prospect of using delayed rectifier channels as therapeutic targets to manage cardiac arrhythmia. Copyright © 2016 Elsevier Inc. All rights reserved.

Adenosine triphosphate-sensitive potassium channel blocking agent ameliorates, but the opening agent aggravates, ischemia/reperfusion-induced injury. Heart function studies in nonfibrillating isolated hearts.

PubMed

Tosaki, A; Hellegouarch, A

1994-02-01

This study was conducted to elucidate the role of the adenosine triphosphate (ATP)-sensitive potassium channel blocking agent glibenclamide and the opener cromakalim in the mechanism of reperfusion-induced injury. Recently, ATP-sensitive potassium channel openers have been proposed to reduce ischemia/reperfusion-induced injury, including arrhythmias and heart function. Thus, one might hypothesize that pharmacologic agents that enhance the loss of potassium ions in the myocardium through ATP-sensitive potassium channels would be arrhythmogenic, and agents that interfere with tissue potassium ion loss would be antiarrhythmic. Isolated "working" guinea pig hearts and phosphorus-31 nuclear magnetic resonance spectroscopy were used to study the recovery of myocardial function and phosphorus compounds after 30, 40 and 50 min of normothermic global ischemia followed by reperfusion in untreated control and glibenclamide- and cromakalim-treated groups. After 30 min of ischemia, 1, 3, 10 and 30 mumol/liter of glibenclamide dose-dependently reduced the incidence of reperfusion-induced ventricular fibrillation (total) from its control value of 92% to 75%, 33% (p < 0.05), 33% (p < 0.05) and 42% (p < 0.05), respectively. The incidence of ventricular tachycardia followed the same pattern. A reduction of arrhythmias was also observed after 40 and 50 min of ischemia followed by reperfusion in the glibenclamide-treated hearts. Cromakalim, at the same concentrations, did not reduce the incidence of reperfusion-induced arrhythmias. During reperfusion, glibenclamide (3 and 10 mumol/liter) improved the recovery of coronary blood flow, aortic flow, myocardial contractility and tissue ATP and creatine phosphate content, but cromakalim failed to ameliorate the recovery of postischemic myocardium compared with that in the drug-free control hearts. The preservation of myocardial potassium ions and phosphorus compounds by glibenclamide can improve the recovery of postischemic function, but the use of ATP-sensitive potassium channel openers as antihypertensive or antiarrhythmic agents may be of particular concern in those postinfarction patients who are known to be at high risk for sudden cardiac death.

Pharmacological Correction of Trafficking Defects in ATP-sensitive Potassium Channels Caused by Sulfonylurea Receptor 1 Mutations.

PubMed

Martin, Gregory M; Rex, Emily A; Devaraneni, Prasanna; Denton, Jerod S; Boodhansingh, Kara E; DeLeon, Diva D; Stanley, Charles A; Shyng, Show-Ling

2016-10-14

ATP-sensitive potassium (K ATP ) channels play a key role in mediating glucose-stimulated insulin secretion by coupling metabolic signals to β-cell membrane potential. Loss of K ATP channel function due to mutations in ABCC8 or KCNJ11, genes encoding the sulfonylurea receptor 1 (SUR1) or the inwardly rectifying potassium channel Kir6.2, respectively, results in congenital hyperinsulinism. Many SUR1 mutations prevent trafficking of channel proteins from the endoplasmic reticulum to the cell surface. Channel inhibitors, including sulfonylureas and carbamazepine, have been shown to correct channel trafficking defects. In the present study, we identified 13 novel SUR1 mutations that cause channel trafficking defects, the majority of which are amenable to pharmacological rescue by glibenclamide and carbamazepine. By contrast, none of the mutant channels were rescued by K ATP channel openers. Cross-linking experiments showed that K ATP channel inhibitors promoted interactions between the N terminus of Kir6.2 and SUR1, whereas channel openers did not, suggesting the inhibitors enhance intersubunit interactions to overcome channel biogenesis and trafficking defects. Functional studies of rescued mutant channels indicate that most mutants rescued to the cell surface exhibited WT-like sensitivity to ATP, MgADP, and diazoxide. In intact cells, recovery of channel function upon trafficking rescue by reversible sulfonylureas or carbamazepine was facilitated by the K ATP channel opener diazoxide. Our study expands the list of K ATP channel trafficking mutations whose function can be recovered by pharmacological ligands and provides further insight into the structural mechanism by which channel inhibitors correct channel biogenesis and trafficking defects. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Specific residues of the cytoplasmic domains of cardiac inward rectifier potassium channels are effective antifibrillatory targets

PubMed Central

Noujaim, Sami F.; Stuckey, Jeanne A.; Ponce-Balbuena, Daniela; Ferrer-Villada, Tania; López-Izquierdo, Angelica; Pandit, Sandeep; Calvo, Conrado J.; Grzeda, Krzysztof R.; Berenfeld, Omer; Sánchez Chapula, José A.; Jalife, José

2010-01-01

Atrial and ventricular tachyarrhythmias can be perpetuated by up-regulation of inward rectifier potassium channels. Thus, it may be beneficial to block inward rectifier channels under conditions in which their function becomes arrhythmogenic (e.g., inherited gain-of-function mutation channelopathies, ischemia, and chronic and vagally mediated atrial fibrillation). We hypothesize that the antimalarial quinoline chloroquine exerts potent antiarrhythmic effects by interacting with the cytoplasmic domains of Kir2.1 (IK1), Kir3.1 (IKACh), or Kir6.2 (IKATP) and reducing inward rectifier potassium currents. In isolated hearts of three different mammalian species, intracoronary chloroquine perfusion reduced fibrillatory frequency (atrial or ventricular), and effectively terminated the arrhythmia with resumption of sinus rhythm. In patch-clamp experiments chloroquine blocked IK1, IKACh, and IKATP. Comparative molecular modeling and ligand docking of chloroquine in the intracellular domains of Kir2.1, Kir3.1, and Kir6.2 suggested that chloroquine blocks or reduces potassium flow by interacting with negatively charged amino acids facing the ion permeation vestibule of the channel in question. These results open a novel path toward discovering antiarrhythmic pharmacophores that target specific residues of the cytoplasmic domain of inward rectifier potassium channels.—Noujaim, S. F., Stuckey, J. A., Ponce-Balbuena, D., Ferrer-Villada, T., López-Izquierdo, A., Pandit, S., Calvo, C. J., Grzeda, K. R., Berenfeld, O., Sánchez Chapula, J. A., Jalife, J. Specific residues of the cytoplasmic domains of cardiac inward rectifier potassium channels are effective antifibrillatory targets. PMID:20585026

Potassium Channels in Epilepsy

PubMed Central

Köhling, Rüdiger; Wolfart, Jakob

2016-01-01

This review attempts to give a concise and up-to-date overview on the role of potassium channels in epilepsies. Their role can be defined from a genetic perspective, focusing on variants and de novo mutations identified in genetic studies or animal models with targeted, specific mutations in genes coding for a member of the large potassium channel family. In these genetic studies, a demonstrated functional link to hyperexcitability often remains elusive. However, their role can also be defined from a functional perspective, based on dynamic, aggravating, or adaptive transcriptional and posttranslational alterations. In these cases, it often remains elusive whether the alteration is causal or merely incidental. With ∼80 potassium channel types, of which ∼10% are known to be associated with epilepsies (in humans) or a seizure phenotype (in animals), if genetically mutated, a comprehensive review is a challenging endeavor. This goal may seem all the more ambitious once the data on posttranslational alterations, found both in human tissue from epilepsy patients and in chronic or acute animal models, are included. We therefore summarize the literature, and expand only on key findings, particularly regarding functional alterations found in patient brain tissue and chronic animal models. PMID:27141079

The Ketogenic Diet and Potassium Channel Function

DTIC Science & Technology

2014-10-01

1 AWARD NUMBER: W81XWH-13-1-0463 TITLE: The Ketogenic Diet and Potassium Channel Function...Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT The overall objective of this Discovery Award is to explore the hypothesis the ketogenic diet ...have examining the impact of the ketogenic diet on mice in which the gene that encodes Kvβ2 has been deleted (Kvβ2 KO mice) using an in vitro model of

In vivo Expression of a Light-activatable Potassium Channel Using Unnatural Amino Acids

PubMed Central

Kang, Ji-Yong; Kawaguchi, Daichi; Coin, Irene; Xiang, Zheng; O’Leary, Dennis D. M.; Slesinger, Paul A.; Wang, Lei

2013-01-01

SUMMARY Optical control of protein function provides excellent spatial-temporal resolution for studying proteins in situ. Although light-sensitive exogenous proteins and ligands have been employed to manipulate neuronal activity, a method for optical control of neuronal proteins using unnatural amino acids (Uaa) in vivo is lacking. Here, we describe the genetic incorporation of a photoreactive Uaa into the pore of an inwardly-rectifying potassium channel Kir2.1. The Uaa occluded the pore, rendering the channel non-conducting, and upon brief light illumination, was released to permit outward K+ current. Expression of this photo-inducible inwardly rectifying potassium (PIRK) channel in rat hippocampal neurons created a light-activatable PIRK switch for suppressing neuronal firing. We also expressed PIRK channels in embryonic mouse neocortex in vivo and demonstrated a light-activated PIRK current in cortical neurons. The principles applied here to a potassium channel could be generally expanded to other proteins expressed in the brain to enable optical regulation. PMID:24139041

Students' Understanding of External Representations of the Potassium Ion Channel Protein Part II: Structure-Function Relationships and Fragmented Knowledge

ERIC Educational Resources Information Center

Harle, Marissa; Towns, Marcy H.

2012-01-01

Research that has focused on external representations in biochemistry has uncovered student difficulties in comprehending and interpreting external representations. This study focuses on students' understanding of three external representations (ribbon diagram, wireframe, and hydrophobic/hydrophilic) of the potassium ion channel protein. Analysis…

Calcium-Activated Potassium Channels at Nodes of Ranvier Secure Axonal Spike Propagation

PubMed Central

Gründemann, Jan; Clark, Beverley A.

2015-01-01

Summary Functional connectivity between brain regions relies on long-range signaling by myelinated axons. This is secured by saltatory action potential propagation that depends fundamentally on sodium channel availability at nodes of Ranvier. Although various potassium channel types have been anatomically localized to myelinated axons in the brain, direct evidence for their functional recruitment in maintaining node excitability is scarce. Cerebellar Purkinje cells provide continuous input to their targets in the cerebellar nuclei, reliably transmitting axonal spikes over a wide range of rates, requiring a constantly available pool of nodal sodium channels. We show that the recruitment of calcium-activated potassium channels (IK, KCa3.1) by local, activity-dependent calcium (Ca2+) influx at nodes of Ranvier via a T-type voltage-gated Ca2+ current provides a powerful mechanism that likely opposes depolarizing block at the nodes and is thus pivotal to securing continuous axonal spike propagation in spontaneously firing Purkinje cells. PMID:26344775

The Segregated Expression of Voltage-Gated Potassium and Sodium Channels in Neuronal Membranes: Functional Implications and Regulatory Mechanisms

PubMed Central

Duménieu, Maël; Oulé, Marie; Kreutz, Michael R.; Lopez-Rojas, Jeffrey

2017-01-01

Neurons are highly polarized cells with apparent functional and morphological differences between dendrites and axon. A critical determinant for the molecular and functional identity of axonal and dendritic segments is the restricted expression of voltage-gated ion channels (VGCs). Several studies show an uneven distribution of ion channels and their differential regulation within dendrites and axons, which is a prerequisite for an appropriate integration of synaptic inputs and the generation of adequate action potential (AP) firing patterns. This review article will focus on the signaling pathways leading to segmented expression of voltage-gated potassium and sodium ion channels at the neuronal plasma membrane and the regulatory mechanisms ensuring segregated functions. We will also discuss the relevance of proper ion channel targeting for neuronal physiology and how alterations in polarized distribution contribute to neuronal pathology. PMID:28484374

Big Potassium (BK) ion channels in biology, disease and possible targets for cancer immunotherapy

PubMed Central

Ge, Lisheng; Hoa, Neil T.; Wilson, Zechariah; Arismendi-Morillo, Gabriel; Kong, Xia-Tang; Tajhya, Rajeev B.; Beeton, Christine; Jadus, Martin R.

2017-01-01

The Big Potassium (BK) ion channel is commonly known by a variety of names (Maxi-K, KCNMA1, slo, Stretch-activated potassium channels, KCa1.1). Each name reflects a different physical property displayed by this single ion channel. This transmembrane channel is found on nearly every cell type of the body and has its own distinctive roles for that tissue type. The BKα channel contains the pore that releases potassium ions from intracellular stores. This ion channel is found on the cell membrane, endoplasmic reticulum, Golgi and mitochondria. Complex splicing pathways produce different isoforms. The BKα channels can be phosphorylated, palmitoylated and myristylated. BK is composed of a homo-tetramer that interacts with β and γ chains. These accessory proteins provide a further modulating effect on the functions of BKα channels. BK channels play important roles in cell division and migration. In this review, we will focus on the biology of BK channels, especially its role, and that it has in the immune response towards cancer. Recent proteomic studies have linked BK channels with various proteins. Some of these interactions offer further insight into the role that BK channels have with cancers, especially with brain tumors. This review shows that BK channels have a complex interplay with intracellular components of cancer cells and still have plenty of secrets to be discovered. PMID:25027630

Effects of channel blocking on information transmission and energy efficiency in squid giant axons.

PubMed

Liu, Yujiang; Yue, Yuan; Yu, Yuguo; Liu, Liwei; Yu, Lianchun

2018-04-01

Action potentials are the information carriers of neural systems. The generation of action potentials involves the cooperative opening and closing of sodium and potassium channels. This process is metabolically expensive because the ions flowing through open channels need to be restored to maintain concentration gradients of these ions. Toxins like tetraethylammonium can block working ion channels, thus affecting the function and energy cost of neurons. In this paper, by computer simulation of the Hodgkin-Huxley neuron model, we studied the effects of channel blocking with toxins on the information transmission and energy efficiency in squid giant axons. We found that gradually blocking sodium channels will sequentially maximize the information transmission and energy efficiency of the axons, whereas moderate blocking of potassium channels will have little impact on the information transmission and will decrease the energy efficiency. Heavy blocking of potassium channels will cause self-sustained oscillation of membrane potentials. Simultaneously blocking sodium and potassium channels with the same ratio increases both information transmission and energy efficiency. Our results are in line with previous studies suggesting that information processing capacity and energy efficiency can be maximized by regulating the number of active ion channels, and this indicates a viable avenue for future experimentation.

Mechanism of resveratrol-induced relaxation of the guinea pig fundus.

PubMed

Tsai, Ching-Chung; Tey, Shu-Leei; Lee, Ming-Che; Liu, Ching-Wen; Su, Yu-Tsun; Huang, Shih-Che

2018-04-01

Resveratrol is a polyphenolic compound that can be isolated from plants and also is a constituent of red wine. Resveratrol induces relaxation of vascular smooth muscle and may prevent cardiovascular diseases. Impaired gastric accommodation plays an important role in functional dyspepsia and fundic relaxation and is a therapeutic target of functional dyspepsia. Although drugs for fundic relaxation have been developed, these types of drugs are still rare. The purpose of this study was to investigate the relaxant effects of resveratrol in the guinea pig fundus. We studied the relaxant effects of resveratrol in the guinea pig fundus. In addition, we investigated the mechanism of resveratrol-induced relaxation on the guinea pig fundus by using tetraethylammonium (a non-selective potassium channel blocker), apamine (a selective inhibitor of the small conductance calcium-activated potassium channel), iberiotoxin (an inhibitor of large conductance calcium-activated potassium channels), glibenclamide (an ATP-sensitive potassium channel blocker), KT 5720 (a cAMP-dependent protein kinase A inhibitor), KT 5823 (a cGMP-dependent protein kinase G inhibitor), NG-nitro-L-arginine (a competitive inhibitor of nitric oxide synthase), tetrodotoxin (a selective neuronal Na + channel blocker), ω-conotoxin GVIA (a selective neuronal Ca 2+ channel blocker) and G-15 (a G-protein coupled estrogen receptor antagonist). The results of this study showed that resveratrol has potent and dose-dependent relaxant effects on the guinea pig fundic muscle. In addition, the results showed that resveratrol-induced relaxation of the guinea pig fundus occurs through nitric oxide and ATP-sensitive potassium channels. This study provides the first evidence concerning the relaxant effects of resveratrol in the guinea pig fundic muscle strips. Furthermore, resveratrol may be a potential drug to relieve gastrointestinal dyspepsia. Copyright © 2018 Elsevier GmbH. All rights reserved.

Altered potassium ATP channel signaling in mesenteric arteries of old high salt-fed rats

PubMed Central

Whidden, Melissa A.; Basgut, Bilgen; Kirichenko, Nataliya; Erdos, Benedek; Tümer, Nihal

2016-01-01

[Purpose] Both aging and the consumption of a high salt diet are associated with clear changes in the vascular system that can lead to the development of cardiovascular disease; however the mechanisms are not clearly understood. Therefore, we examined whether aging and the consumption of excess salt alters the function of potassium ATP-dependent channel signaling in mesenteric arteries [Methods] Young (7 months) and old (29 months) Fischer 344 x Brown Norway rats were fed a control or a high salt diet (8% NaCl) for 12 days and mesenteric arteries were utilized for vascular reactivity measurements. [Results] Acetylcholine-induced endothelium relaxation was significantly reduced in old arteries (81 ± 4%) when compared with young arteries (92 ± 2%). Pretreatment with the potassium-ATP channel blocker glibenclamide reduced relaxation to acetylcholine in young arteries but did not alter dilation in old arteries. On a high salt diet, endothelium dilation to acetylcholine was significantly reduced in old salt arteries (60 ± 3%) when compared with old control arteries (81 ± 4%). Glibenclamide reduced acetylcholine-induced dilation in young salt arteries but had no effect on old salt arteries. Dilation to cromakalim, a potassium-ATP channel opener, was reduced in old salt arteries when compared with old control arteries. [Conclusion] These findings demonstrate that aging impairs endothelium-dependent relaxation in mesenteric arteries. Furthermore, a high salt diet alters the function of potassium-ATP-dependent channel signaling in old isolated mesenteric arteries and affects the mediation of relaxation stimuli. PMID:27508155

Slick (Kcnt2) Sodium-Activated Potassium Channels Limit Peptidergic Nociceptor Excitability and Hyperalgesia.

PubMed

Tomasello, Danielle L; Hurley, Edward; Wrabetz, Lawrence; Bhattacharjee, Arin

2017-01-01

The Slick (Kcnt2) sodium-activated potassium (K Na ) channel is a rapidly gating and weakly voltage-dependent and sodium-dependent potassium channel with no clearly defined physiological function. Within the dorsal root ganglia (DRGs), we show Slick channels are exclusively expressed in small-sized and medium-sized calcitonin gene-related peptide (CGRP)-containing DRG neurons, and a pool of channels are localized to large dense-core vesicles (LDCV)-containing CGRP. We stimulated DRG neurons for CGRP release and found Slick channels contained within CGRP-positive LDCV translocated to the neuronal membrane. Behavioral studies in Slick knockout (KO) mice indicated increased basal heat detection and exacerbated thermal hyperalgesia compared with wild-type littermate controls during neuropathic and chronic inflammatory pain. Electrophysiologic recordings of DRG neurons from Slick KO mice revealed that Slick channels contribute to outward current, propensity to fire action potentials (APs), and to AP properties. Our data suggest that Slick channels restrain the excitability of CGRP-containing neurons, diminishing pain behavior after inflammation and injury.

Disruption of the potassium channel regulatory subunit KCNE2 causes iron-deficient anemia

PubMed Central

Salsbury, Grace; Cambridge, Emma L.; McIntyre, Zoe; Arends, Mark J.; Karp, Natasha A.; Isherwood, Christopher; Shannon, Carl; Hooks, Yvette; Ramirez-Solis, Ramiro; Adams, David J.; White, Jacqueline K.; Speak, Anneliese O.

2014-01-01

Iron homeostasis is a dynamic process that is tightly controlled to balance iron uptake, storage, and export. Reduction of dietary iron from the ferric to the ferrous form is required for uptake by solute carrier family 11 (proton-coupled divalent metal ion transporters), member 2 (Slc11a2) into the enterocytes. Both processes are proton dependent and have led to the suggestion of the importance of acidic gastric pH for the absorption of dietary iron. Potassium voltage-gated channel subfamily E, member 2 (KCNE2), in combination with potassium voltage-gated channel, KQT-like subfamily, member 1 (KCNQ1), form a gastric potassium channel essential for gastric acidification. Deficiency of either Kcne2 or Kcnq1 results in achlorhydia, gastric hyperplasia, and neoplasia, but the impact on iron absorption has not, to our knowledge, been investigated. Here we report that Kcne2-deficient mice, in addition to the previously reported phenotypes, also present with iron-deficient anemia. Interestingly, impaired function of KCNQ1 results in iron-deficient anemia in Jervell and Lange-Nielsen syndrome patients. We speculate that impaired function of KCNE2 could result in the same clinical phenotype. PMID:25127743

Physical and functional interaction between integrins and hERG potassium channels.

PubMed

Arcangeli, A; Becchetti, A; Cherubini, A; Crociani, O; Defilippi, P; Guasti, L; Hofmann, G; Pillozzi, S; Olivotto, M; Wanke, E

2004-11-01

Integrins are adhesion receptors capable of transmitting intracellular signals that regulate many different cellular functions. Among integrin-mediated signals, the activation of ion channels can be included. We demonstrated that a long-lasting activation of hERG (human ether-a-go-go-related gene) potassium channels occurs in both human neuroblastoma and leukaemia cells after the activation of the beta1 integrin subunit. This activation is apparently a determining factor inducing neurite extension and osteoclastic differentiation in both the cell types. More recently, we provided evidences that beta1 integrins and hERG channels co-precipitate in both the cell types. Preliminary results suggest that a macromolecular signalling complex indeed occurs between integrins and the hERG1 protein and that hERG channel activity can modulate integrin downstream signalling.

Altered profile of mRNA expression in atrioventricular node of streptozotocin-induced diabetic rats

PubMed Central

Howarth, Frank Christopher; Parekh, Khatija; Jayaprakash, Petrilla; Inbaraj, Edward Samuel; Oz, Murat; Dobrzynski, Halina; Adrian, Thomas Edward

2017-01-01

Prolonged action potential duration, reduced action potential firing rate, upstroke velocity and rate of diastolic depolarization have been demonstrated in atrioventricular node (AVN) cells from streptozotocin (STZ)-induced diabetic rats. To further clarify the molecular basis of these electrical disturbances, the mRNA profiles encoding a variety of proteins associated with the generation and conduction of electrical activity in the AVN, were evaluated in the STZ-induced diabetic rat heart. Expression of mRNA was measured in AVN biopsies using reverse transcription-quantitative polymerase chain reaction techniques. Notable differences in mRNA expression included upregulation of genes encoding membrane and intracellular Ca2+ transport, including solute carrier family 8 member A1, transient receptor potential channel 1, ryanodine receptor 2/3, hyperpolarization-activated cyclic-nucleotide 2 and 3, calcium channel voltage-dependent, β2 subunit and sodium channels 3a, 4a, 7a and 3b. In addition to this, potassium channels potassium voltage-gated channel subfamily A member 4, potassium channel calcium activated intermediate/small conductance subfamily N α member 2, potassium voltage-gated channel subfamily J members 3, 5, and 11, potassium channel subfamily K members 1, 2, 3 and natriuretic peptide B (BNP) were upregulated in AVN of STZ heart, compared with controls. Alterations in gene expression were associated with upregulation of various proteins including the inwardly rectifying, potassium channel Kir3.4, NCX1 and BNP. The present study demonstrated notable differences in the profile of mRNA encoding proteins associated with the generation, conduction and regulation of electrical signals in the AVN of the STZ-induced diabetic rat heart. These data will provide a basis for a substantial range of future studies to investigate whether variations in mRNA translate into alterations in electrophysiological function. PMID:28731153

The role of ATP-sensitive potassium channels in cellular function and protection in the cardiovascular system.

PubMed

Tinker, Andrew; Aziz, Qadeer; Thomas, Alison

2014-01-01

ATP-sensitive potassium channels (K(ATP)) are widely distributed and present in a number of tissues including muscle, pancreatic beta cells and the brain. Their activity is regulated by adenine nucleotides, characteristically being activated by falling ATP and rising ADP levels. Thus, they link cellular metabolism with membrane excitability. Recent studies using genetically modified mice and genomic studies in patients have implicated K(ATP) channels in a number of physiological and pathological processes. In this review, we focus on their role in cellular function and protection particularly in the cardiovascular system. © 2013 The British Pharmacological Society.

Inwardly rectifying potassium channels influence Drosophila wing morphogenesis by regulating Dpp release.

PubMed

Dahal, Giri Raj; Pradhan, Sarala Joshi; Bates, Emily Anne

2017-08-01

Loss of embryonic ion channel function leads to morphological defects, but the underlying reason for these defects remains elusive. Here, we show that inwardly rectifying potassium (Irk) channels regulate release of the Drosophila bone morphogenetic protein Dpp in the developing fly wing and that this is necessary for developmental signaling. Inhibition of Irk channels decreases the incidence of distinct Dpp-GFP release events above baseline fluorescence while leading to a broader distribution of Dpp-GFP. Work by others in different cell types has shown that Irk channels regulate peptide release by modulating membrane potential and calcium levels. We found calcium transients in the developing wing, and inhibition of Irk channels reduces the duration and amplitude of calcium transients. Depolarization with high extracellular potassium evokes Dpp release. Taken together, our data implicate Irk channels as a requirement for regulated release of Dpp, highlighting the importance of the temporal pattern of Dpp presentation for morphogenesis of the wing. © 2017. Published by The Company of Biologists Ltd.

Differential Regulation of Action Potential Shape and Burst-Frequency Firing by BK and Kv2 Channels in Substantia Nigra Dopaminergic Neurons

PubMed Central

Kimm, Tilia; Khaliq, Zayd M.

2015-01-01

Little is known about the voltage-dependent potassium currents underlying spike repolarization in midbrain dopaminergic neurons. Studying mouse substantia nigra pars compacta dopaminergic neurons both in brain slice and after acute dissociation, we found that BK calcium-activated potassium channels and Kv2 channels both make major contributions to the depolarization-activated potassium current. Inhibiting Kv2 or BK channels had very different effects on spike shape and evoked firing. Inhibiting Kv2 channels increased spike width and decreased the afterhyperpolarization, as expected for loss of an action potential-activated potassium conductance. BK inhibition also increased spike width but paradoxically increased the afterhyperpolarization. Kv2 channel inhibition steeply increased the slope of the frequency–current (f–I) relationship, whereas BK channel inhibition had little effect on the f–I slope or decreased it, sometimes resulting in slowed firing. Action potential clamp experiments showed that both BK and Kv2 current flow during spike repolarization but with very different kinetics, with Kv2 current activating later and deactivating more slowly. Further experiments revealed that inhibiting either BK or Kv2 alone leads to recruitment of additional current through the other channel type during the action potential as a consequence of changes in spike shape. Enhancement of slowly deactivating Kv2 current can account for the increased afterhyperpolarization produced by BK inhibition and likely underlies the very different effects on the f–I relationship. The cross-regulation of BK and Kv2 activation illustrates that the functional role of a channel cannot be defined in isolation but depends critically on the context of the other conductances in the cell. SIGNIFICANCE STATEMENT This work shows that BK calcium-activated potassium channels and Kv2 voltage-activated potassium channels both regulate action potentials in dopamine neurons of the substantia nigra pars compacta. Although both channel types participate in action potential repolarization about equally, they have contrasting and partially opposite effects in regulating neuronal firing at frequencies typical of bursting. Our analysis shows that this results from their different kinetic properties, with fast-activating BK channels serving to short-circuit activation of Kv2 channels, which tend to slow firing by producing a deep afterhyperpolarization. The cross-regulation of BK and Kv2 activation illustrates that the functional role of a channel cannot be defined in isolation but depends critically on the context of the other conductances in the cell. PMID:26674866

Differential Regulation of Action Potential Shape and Burst-Frequency Firing by BK and Kv2 Channels in Substantia Nigra Dopaminergic Neurons.

PubMed

Kimm, Tilia; Khaliq, Zayd M; Bean, Bruce P

2015-12-16

Little is known about the voltage-dependent potassium currents underlying spike repolarization in midbrain dopaminergic neurons. Studying mouse substantia nigra pars compacta dopaminergic neurons both in brain slice and after acute dissociation, we found that BK calcium-activated potassium channels and Kv2 channels both make major contributions to the depolarization-activated potassium current. Inhibiting Kv2 or BK channels had very different effects on spike shape and evoked firing. Inhibiting Kv2 channels increased spike width and decreased the afterhyperpolarization, as expected for loss of an action potential-activated potassium conductance. BK inhibition also increased spike width but paradoxically increased the afterhyperpolarization. Kv2 channel inhibition steeply increased the slope of the frequency-current (f-I) relationship, whereas BK channel inhibition had little effect on the f-I slope or decreased it, sometimes resulting in slowed firing. Action potential clamp experiments showed that both BK and Kv2 current flow during spike repolarization but with very different kinetics, with Kv2 current activating later and deactivating more slowly. Further experiments revealed that inhibiting either BK or Kv2 alone leads to recruitment of additional current through the other channel type during the action potential as a consequence of changes in spike shape. Enhancement of slowly deactivating Kv2 current can account for the increased afterhyperpolarization produced by BK inhibition and likely underlies the very different effects on the f-I relationship. The cross-regulation of BK and Kv2 activation illustrates that the functional role of a channel cannot be defined in isolation but depends critically on the context of the other conductances in the cell. This work shows that BK calcium-activated potassium channels and Kv2 voltage-activated potassium channels both regulate action potentials in dopamine neurons of the substantia nigra pars compacta. Although both channel types participate in action potential repolarization about equally, they have contrasting and partially opposite effects in regulating neuronal firing at frequencies typical of bursting. Our analysis shows that this results from their different kinetic properties, with fast-activating BK channels serving to short-circuit activation of Kv2 channels, which tend to slow firing by producing a deep afterhyperpolarization. The cross-regulation of BK and Kv2 activation illustrates that the functional role of a channel cannot be defined in isolation but depends critically on the context of the other conductances in the cell. Copyright © 2015 the authors 0270-6474/15/3516404-14$15.00/0.

Study of the interaction of potassium ion channel protein with micelle by molecular dynamics simulation

NASA Astrophysics Data System (ADS)

Shantappa, Anil; Talukdar, Keka

2018-04-01

Ion channels are proteins forming pore inside the body of all living organisms. This potassium ion channel known as KcsA channel and it is found in the each cell and nervous system. Flow of various ions is regulated by the function of the ion channels. The nerve ion channel protein with protein data bank entry 1BL8, which is basically an ion channel protein in Streptomyces Lividans and which is taken up to form micelle-protein system and the system is analyzed by using molecular dynamics simulation. Firstly, ion channel pore is engineered by CHARMM potential and then Micelle-protein system is subjected to molecular dynamics simulation. For some specific micelle concentration, the protein unfolding is observed.

Endoplasmic reticulum membrane potassium channel dysfunction in high fat diet induced stress in rat hepatocytes

PubMed Central

Khodaee, Naser; Ghasemi, Maedeh; Saghiri, Reza; Eliassi, Afsaneh

2014-01-01

In a previous study we reported the presence of a large conductance K+ channel in the membrane of endoplasmic reticulum (ER) from rat hepatocytes. The channel open probability (Po) appeared voltage dependent and reached to a minimum 0.2 at +50 mV. Channel activity in this case was found to be totally inhibited at ATP concentration 2.5 mM, glibenclamide 100 µM and tolbutamide 400 µM. Existing evidence indicates an impairment of endoplasmic reticulum functions in ER stress condition. Because ER potassium channels have been involved in several ER functions including cytoprotection, apoptosis and calcium homeostasis, a study was carried out to consider whether the ER potassium channel function is altered in a high fat diet model of ER stress. Male Wistar rats were made ER stress for 2 weeks with a high fat diet. Ion channel incorporation of ER stress model into the bilayer lipid membrane allowed the characterization of K+ channel. Our results indicate that the channel Po was significantly increased at voltages above +30 mV. Interestingly, addition of ATP 7.5 mM, glibenclamide 400 µM and tolbutamide 2400 µM totally inhibited the channel activities, 3-fold, 4-fold and 6-fold higher than that in the control groups, respectively. Our results thus demonstrate a modification in the ER K+ channel gating properties and decreased sensitivity to drugs in membrane preparations coming from ER high fat model of ER stress, an effect potentially linked to a change in ER K+ channel subunits in ER stress condition. Our results may provide new insights into the cellular mechanisms underlying ER dysfunctions in ER stress. PMID:26417322

Alterations of sodium and potassium channels of RGCs in RCS rat with the development of retinal degeneration.

PubMed

Chen, Zhongshan; Song, Yanping; Yao, Junping; Weng, Chuanhuang; Yin, Zheng Qin

2013-11-01

All know that retinitis pigmentosa (RP) is a group of hereditary retinal degenerative diseases characterized by progressive dysfunction of photoreceptors and associated with progressive cells loss; nevertheless, little is known about how rods and cones loss affects the surviving inner retinal neurons and networks. Retinal ganglion cells (RGCs) process and convey visual information from retina to visual centers in the brain. The healthy various ion channels determine the normal reception and projection of visual signals from RGCs. Previous work on the Royal College of Surgeons (RCS) rat, as a kind of classical RP animal model, indicated that, at late stages of retinal degeneration in RCS rat, RGCs were also morphologically and functionally affected. Here, retrograde labeling for RGCs with Fluorogold was performed to investigate the distribution, density, and morphological changes of RGCs during retinal degeneration. Then, patch clamp recording, western blot, and immunofluorescence staining were performed to study the channels of sodium and potassium properties of RGCs, so as to explore the molecular and proteinic basis for understanding the alterations of RGCs membrane properties and firing functions. We found that the resting membrane potential, input resistance, and capacitance of RGCs changed significantly at the late stage of retinal degeneration. Action potential could not be evoked in a part of RGCs. Inward sodium current and outward potassium current recording showed that sodium current was impaired severely but only slightly in potassium current. Expressions of sodium channel protein were impaired dramatically at the late stage of retinal degeneration. The results suggested that the density of RGCs decreased, process ramification impaired, and sodium ion channel proteins destructed, which led to the impairment of electrophysiological functions of RGCs and eventually resulted in the loss of visual function.

Preliminary Studies of Acute Cadmium Administration Effects on the Calcium-Activated Potassium (SKCa and BKCa) Channels and Na+/K+-ATPase Activity in Isolated Aortic Rings of Rats.

PubMed

Vassallo, Dalton V; Almenara, Camila C P; Broseghini-Filho, Gilson Brás; Teixeira, Ariane Calazans; da Silva, David Chaves F; Angeli, Jhuli K; Padilha, Alessandra S

2018-06-01

Cadmium is an environmental pollutant closely linked with cardiovascular diseases that seems to involve endothelium dysfunction and reduced nitric oxide (NO) bioavailability. Knowing that NO causes dilatation through the activation of potassium channels and Na + /K + -ATPase, we aimed to determine whether acute cadmium administration (10 μM) alters the participation of K + channels, voltage-activated calcium channel, and Na + /K + -ATPase activity in vascular function of isolated aortic rings of rats. Cadmium did not modify the acetylcholine-induced relaxation. After L-NAME addition, the relaxation induced by acetylcholine was abolished in presence or absence of cadmium, suggesting that acutely, this metal did not change NO release. However, tetraethylammonium (a nonselective K + channels blocker) reduced acetylcholine-induced relaxation but this effect was lower in the preparations with cadmium, suggesting a decrease of K + channels function in acetylcholine response after cadmium incubation. Apamin (a selective blocker of small Ca 2+ -activated K + channels-SK Ca ), iberiotoxin (a selective blocker of large-conductance Ca 2+ -activated K + channels-BK Ca ), and verapamil (a blocker of calcium channel) reduced the endothelium-dependent relaxation only in the absence of cadmium. Finally, cadmium decreases Na + /K + -ATPase activity. Our results provide evidence that the cadmium acute incubation unaffected the calcium-activated potassium channels (SK Ca and BK Ca ) and voltage-calcium channels on the acetylcholine vasodilatation. In addition, acute cadmium incubation seems to reduce the Na + /K + -ATPase activity.

Membrane transporters for nitrogen, phosphate and potassium uptake in plants.

PubMed

Chen, Yi-Fang; Wang, Yi; Wu, Wei-Hua

2008-07-01

Nitrogen, phosphorous and potassium are essential nutrients for plant growth and development. However, their contents in soils are limited so that crop production needs to invest a lot for fertilizer supply. To explore the genetic potentialities of crops (or plants) for their nutrient utilization efficiency has been an important research task for many years. In fact, a number of evidences have revealed that plants, during their evolution, have developed many morphological, physiological, biochemical and molecular adaptation mechanisms for acquiring nitrate, phosphate and potassium under stress conditions. Recent discoveries of many transporters and channels for nitrate, phosphate and potassium uptake have opened up opportunities to study the molecular regulatory mechanisms for acquisition of these nutrients. This review aims to briefly discuss the genes and gene families for these transporters and channels. In addition, the functions and regulation of some important transporters and channels are particularly emphasized.

Potassium Channels in Regulation of Vascular Smooth Muscle Contraction and Growth

PubMed Central

Jackson, William F.

2017-01-01

Potassium channels importantly contribute to the regulation of vascular smooth muscle (VSM) contraction and growth. They are the dominant ion conductance of the VSM cell membrane and importantly determine and regulate membrane potential. Membrane potential, in turn, regulates the open-state probability of voltage-gated Ca2+ channels (VGCC), Ca2+ influx through VGCC, intracellular Ca2+ and VSM contraction. Membrane potential also affects release of Ca2+ from internal stores and the Ca2+ sensitivity of the contractile machinery such that K+ channels participate in all aspects of regulation of VSM contraction. Potassium channels also regulate proliferation of VSM cells through membrane potential-dependent and membrane potential-independent mechanisms. Vascular smooth muscle cells express multiple isoforms of at least five classes of K+ channels contribute to the regulation of contraction and cell proliferation (growth). This review will examine the structure, expression and function of large-conductance, Ca2+-activated K+ (BKCa) channels, intermediate-conductance Ca2+-activated K+ (KCa3.1) channels, multiple isoforms of voltage-gated K+ (KV) channels, ATP-sensitive K+ (KATP) channels, and inward-rectifier K+ (KIR) channels in both contractile and proliferating VSM cells. PMID:28212804

A heteromeric potassium channel involved in the modulation of the plasma membrane potential is essential for the survival of African trypanosomes.

PubMed

Steinmann, Michael E; González-Salgado, Amaia; Bütikofer, Peter; Mäser, Pascal; Sigel, Erwin

2015-08-01

Discovery of novel drug targets may lead to improved treatment of trypanosomiasis. We characterize here 2 gene products of Trypanosoma brucei that are essential for the growth of bloodstream form (BSF) parasites, as shown by RNA interference (RNAi)-mediated down-regulation of the individual mRNAs. The primary sequences of the 2 proteins--protein encoded by gene Tb927.1.4450 (TbK1) and protein encoded by gene Tb927.9.4820 (TbK2)--indicate that both belong to the family of putative, Ca(2+)-activated potassium channels. The proteins were expressed in Xenopus laevis oocytes and their functions investigated by use of electrophysiological techniques. Only combined expression of TbK1 and TbK2 results in the formation of sizeable currents, indicating that these proteins probably assemble into a heteromeric ion channel. The current mediated by this channel shows little time and voltage dependence and displays a permeability ratio of K(+)/Na(+) of >20. The known potassium channel blocker barium inhibits this channel with a half-maximal inhibitory concentration (IC50) of 98 ± 15 μM. The membrane potential of trypanosomes was measured with a fluorescent dye. Individual RNAi-mediated down-regulation of TbK1 or TbK2 eliminates a potassium conductance in the plasma membrane of BSF. Thus, this heteromeric potassium channel is involved in the modulation of the plasma membrane potential and represents a novel drug target in T. brucei. © FASEB.

A new pH-sensitive rectifying potassium channel in mitochondria from the embryonic rat hippocampus.

PubMed

Kajma, Anna; Szewczyk, Adam

2012-10-01

Patch-clamp single-channel studies on mitochondria isolated from embryonic rat hippocampus revealed the presence of two different potassium ion channels: a large-conductance (288±4pS) calcium-activated potassium channel and second potassium channel with outwardly rectifying activity under symmetric conditions (150/150mM KCl). At positive voltages, this channel displayed a conductance of 67.84pS and a strong voltage dependence at holding potentials from -80mV to +80mV. The open probability was higher at positive than at negative voltages. Patch-clamp studies at the mitoplast-attached mode showed that the channel was not sensitive to activators and inhibitors of mitochondrial potassium channels but was regulated by pH. Moreover, we demonstrated that the channel activity was not affected by the application of lidocaine, an inhibitor of two-pore domain potassium channels, or by tertiapin, an inhibitor of inwardly rectifying potassium channels. In summary, based on the single-channel recordings, we characterised for the first time mitochondrial pH-sensitive ion channel that is selective for cations, permeable to potassium ions, displays voltage sensitivity and does not correspond to any previously described potassium ion channels in the inner mitochondrial membrane. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012). Copyright © 2012 Elsevier B.V. All rights reserved.

Connexin 43 and ATP-sensitive potassium channels crosstalk: a missing link in hypoxia/ischemia stress.

PubMed

Ahmad Waza, Ajaz; Ahmad Bhat, Shabir; Ul Hussain, Mahboob; Ganai, Bashir A

2018-02-01

Connexin 43 (Cx43) is a gap junction protein expressed in various tissues and organs of vertebrates. Besides functioning as a gap junction, Cx43 also regulates diverse cellular processes like cell growth and differentiation, cell migration, cell survival, etc. Cx43 is critical for normal cardiac functioning and is therefore abundantly expressed in cardiomyocytes. On the other hand, ATP-sensitive potassium (K ATP ) channels are metabolic sensors converting metabolic changes into electrical activity. These channels are important in maintaining the neurotransmitter release, smooth muscle relaxation, cardiac action potential repolarization, normal physiology of cellular repolarization, insulin secretion and immune function. Cx43 and K ATP channels are part of the same signaling pathway, regulating cell survival during stress conditions and ischemia/hypoxia preconditioning. However, the underlying molecular mechanism for their combined role in ischemia/hypoxia preconditioning is largely unknown. The current review focuses on understanding the molecular mechanism responsible for the coordinated role of Cx43 and K ATP channel protein in protecting cardiomyocytes against ischemia/hypoxia stress.

High temperature sensitivity is intrinsic to voltage-gated potassium channels

PubMed Central

Yang, Fan; Zheng, Jie

2014-01-01

Temperature-sensitive transient receptor potential (TRP) ion channels are members of the large tetrameric cation channels superfamily but are considered to be uniquely sensitive to heat, which has been presumed to be due to the existence of an unidentified temperature-sensing domain. Here we report that the homologous voltage-gated potassium (Kv) channels also exhibit high temperature sensitivity comparable to that of TRPV1, which is detectable under specific conditions when the voltage sensor is functionally decoupled from the activation gate through either intrinsic mechanisms or mutations. Interestingly, mutations could tune Shaker channel to be either heat-activated or heat-deactivated. Therefore, high temperature sensitivity is intrinsic to both TRP and Kv channels. Our findings suggest important physiological roles of heat-induced variation in Kv channel activities. Mechanistically our findings indicate that temperature-sensing TRP channels may not contain a specialized heat-sensor domain; instead, non-obligatory allosteric gating permits the intrinsic heat sensitivity to drive channel activation, allowing temperature-sensitive TRP channels to function as polymodal nociceptors. DOI: http://dx.doi.org/10.7554/eLife.03255.001 PMID:25030910

Ras-Association Domain of Sorting Nexin 27 Is Critical for Regulating Expression of GIRK Potassium Channels

PubMed Central

Bodhinathan, Karthik; Taura, Jaume J.; Taylor, Natalie M.; Nettleton, Margaret Y.; Ciruela, Francisco; Slesinger, Paul A.

2013-01-01

G protein-gated inwardly rectifying potassium (GIRK) channels play an important role in regulating neuronal excitability. Sorting nexin 27b (SNX27b), which reduces surface expression of GIRK channels through a PDZ domain interaction, contains a putative Ras-association (RA) domain with unknown function. Deleting the RA domain in SNX27b (SNX27b-ΔRA) prevents the down-regulation of GIRK2c/GIRK3 channels. Similarly, a point mutation (K305A) in the RA domain disrupts regulation of GIRK2c/GIRK3 channels and reduces H-Ras binding in vitro. Finally, the dominant-negative H-Ras (S17N) occludes the SNX27b-dependent decrease in surface expression of GIRK2c/GIRK3 channels. Thus, the presence of a functional RA domain and the interaction with Ras-like G proteins comprise a novel mechanism for modulating SNX27b control of GIRK channel surface expression and cellular excitability. PMID:23536889

Transportation behavior of alkali ions through a cell membrane ion channel. A quantum chemical description of a simplified isolated model.

PubMed

Billes, Ferenc; Mohammed-Ziegler, Ildikó; Mikosch, Hans

2012-08-01

Quantum chemical model calculations were carried out for modeling the ion transport through an isolated ion channel of a cell membrane. An isolated part of a natural ion channel was modeled. The model channel was a calixarene derivative, hydrated sodium and potassium ions were the models of the transported ion. The electrostatic potential of the channel and the energy of the channel-ion system were calculated as a function of the alkali ion position. Both attractive and repulsive ion-channel interactions were found. The calculations - namely the dependence of the system energy and the atomic charges of the water molecules with respect to the position of the alkali ion in the channel - revealed the molecular-structural background of the potassium selectivity of this artificial ion channel. It was concluded that the studied ion channel mimics real biological ion channel quite well.

Expression and distribution of Kv4 potassium channel subunits and potassium channel interacting proteins in subpopulations of interneurons in the basolateral amygdala.

PubMed

Dabrowska, J; Rainnie, D G

2010-12-15

The Kv4 potassium channel α subunits, Kv4.1, Kv4.2, and Kv4.3, determine some of the fundamental physiological properties of neurons in the CNS. Kv4 subunits are associated with auxiliary β-subunits, such as the potassium channel interacting proteins (KChIP1 - 4), which are thought to regulate the trafficking and gating of native Kv4 potassium channels. Intriguingly, KChIP1 is thought to show cell type-selective expression in GABA-ergic inhibitory interneurons, while other β-subunits (KChIP2-4) are associated with principal glutamatergic neurons. However, nothing is known about the expression of Kv4 family α- and β-subunits in specific interneurons populations in the BLA. Here, we have used immunofluorescence, co-immunoprecipitation, and Western Blotting to determine the relative expression of KChIP1 in the different interneuron subtypes within the BLA, and its co-localization with one or more of the Kv4 α subunits. We show that all three α-subunits of Kv4 potassium channel are found in rat BLA neurons, and that the immunoreactivity of KChIP1 closely resembles that of Kv4.3. Indeed, Kv4.3 showed almost complete co-localization with KChIP1 in the soma and dendrites of a distinct subpopulation of BLA neurons. Dual-immunofluorescence studies revealed this to be in BLA interneurons immunoreactive for parvalbumin, cholecystokin-8, and somatostatin. Finally, co-immunoprecipitation studies showed that KChIP1 was associated with all three Kv4 α subunits. Together our results suggest that KChIP1 is selectively expressed in BLA interneurons where it may function to regulate the activity of A-type potassium channels. Hence, KChIP1 might be considered as a cell type-specific regulator of GABAergic inhibitory circuits in the BLA. Copyright © 2010 IBRO. Published by Elsevier Ltd. All rights reserved.

Potassium Channels in Peripheral Pain Pathways: Expression, Function and Therapeutic Potential

PubMed Central

Du, Xiaona; Gamper, Nikita

2013-01-01

Electrical excitation of peripheral somatosensory nerves is a first step in generation of most pain signals in mammalian nervous system. Such excitation is controlled by an intricate set of ion channels that are coordinated to produce a degree of excitation that is proportional to the strength of the external stimulation. However, in many disease states this coordination is disrupted resulting in deregulated peripheral excitability which, in turn, may underpin pathological pain states (i.e. migraine, neuralgia, neuropathic and inflammatory pains). One of the major groups of ion channels that are essential for controlling neuronal excitability is potassium channel family and, hereby, the focus of this review is on the K+ channels in peripheral pain pathways. The aim of the review is threefold. First, we will discuss current evidence for the expression and functional role of various K+ channels in peripheral nociceptive fibres. Second, we will consider a hypothesis suggesting that reduced functional activity of K+ channels within peripheral nociceptive pathways is a general feature of many types of pain. Third, we will evaluate the perspectives of pharmacological enhancement of K+ channels in nociceptive pathways as a strategy for new analgesic drug design. PMID:24396338

Vertebrate rod photoreceptors express both BK and IK calcium-activated potassium channels, but only BK channels are involved in receptor potential regulation.

PubMed

Pelucchi, Bruna; Grimaldi, Annalisa; Moriondo, Andrea

2008-01-01

In salamander rods, Ca(2+)-activated K(+) current (I(KCa)) provides an effective "clamp" of the dark membrane potential to its normal resting level. By a combination of electrophysiological, pharmacological, and immunohistochemical approaches, we show that salamander rods functionally express large-conductance Ca(2+)- and voltage-dependent potassium (BK) channel and intermediate-conductance Ca(2+)-dependent potassium (IK) channel, but not small-conductance Ca(2+)-dependent potassium channel (SK) subtypes. Application of 100 nM iberiotoxin and 100 nM clotrimazole reduced net I(KCa) to 36% and 63%, respectively, whereas the current was unaffected by application of 1 microM apamin. Consistently, anti- SK1, -SK2, and -SK3 antibodies were unable to stain rod photoreceptors, whereas both anti-BK and -SK4/ IK1 antibodies heavily stained the ellipsoid region of the inner segments of the rods. Moreover, by using current-clamp experiments, it was clearly seen that the strong clamping effect of the total I(KCa) was lost when IbTx, but not CLTZ, was applied to the bath. This behavior strongly suggests that of BK and IK channels, only the former are responsible for the clamping effect on the photoreceptor membrane potential.

VOLTAGE-GATED POTASSIUM CHANNELS AT THE CROSSROADS OF NEURONAL FUNCTION, ISCHEMIC TOLERANCE, AND NEURODEGENERATION

PubMed Central

Shah, Niyathi Hegde; Aizenman, Elias

2013-01-01

Voltage-gated potassium (Kv) channels are widely expressed in the central and peripheral nervous system, and are crucial mediators of neuronal excitability. Importantly, these channels also actively participate in cellular and molecular signaling pathways that regulate the life and death of neurons. Injury-mediated increased K+ efflux through Kv2.1 channels promotes neuronal apoptosis, contributing to widespread neuronal loss in neurodegenerative disorders such as Alzheimer’s disease and stroke. In contrast, some forms of neuronal activity can dramatically alter Kv2.1 channel phosphorylation levels and influence their localization. These changes are normally accompanied by modifications in channel voltage-dependence, which may be neuroprotective within the context of ischemic injury. Kv1 and Kv7 channel dysfunction leads to neuronal hyperexcitability that critically contributes to the pathophysiology of human clinical disorders such as episodic ataxia and epilepsy. This review summarizes the neurotoxic, neuroprotective, and neuroregulatory roles of Kv channels, and highlights the consequences of Kv channel dysfunction on neuronal physiology. The studies described in this review thus underscore the importance of normal Kv channel function in neurons, and emphasize the therapeutic potential of targeting Kv channels in the treatment of a wide range of neurological diseases. PMID:24323720

United in Diversity: Mechanosensitive Ion Channels in Plants

PubMed Central

Hamilton, Eric S.; Schlegel, Angela M.; Haswell, Elizabeth S.

2015-01-01

Mechanosensitive (MS) ion channels are a common mechanism for perceiving and responding to mechanical force. This class of mechanoreceptors is capable of transducing membrane tension directly into ion flux. In plant systems, MS ion channels have been proposed to play a wide array of roles, from the perception of touch and gravity to the osmotic homeostasis of intracellular organelles. Three families of plant MS ion channels have been identified: the MscS-like (MSL), Mid1-complementing activity (MCA), and two-pore potassium (TPK) families. Channels from these families vary widely in structure and function, localize to multiple cellular compartments, and conduct chloride, calcium, and/or potassium ions. However, they are still likely to represent only a fraction of the MS ion channel diversity in plant systems. PMID:25494462

Distribution of the messenger RNA for the small conductance calcium-activated potassium channel SK3 in the adult rat brain and correlation with immunoreactivity.

PubMed

Tacconi, S; Carletti, R; Bunnemann, B; Plumpton, C; Merlo Pich, E; Terstappen, G C

2001-01-01

Small conductance calcium-activated potassium channels are voltage independent potassium channels which modulate the firing patterns of neurons by activating the slow component of the afterhyperpolarization. The genes encoding a family of small conductance calcium-activated potassium channels have been cloned and up to now three known members have been described and named small conductance calcium-activated potassium channel type 1, small conductance calcium-activated potassium channel type 2 and small conductance calcium-activated potassium channel type 3; the distribution of their messenger RNA in the rat CNS has already been performed but only in a limited detail. The present study represents the first detailed analysis of small conductance calcium-activated potassium channel type 3 mRNA distribution in the adult rat brain and resulted in a strong to moderate expression of signal in medial habenular nucleus, substantia nigra compact part, suprachiasmatic nucleus, ventral tegmental area, lateral septum, dorsal raphe and locus coeruleus. Immunohistological experiments were also performed and confirmed the presence of small conductance calcium-activated potassium channel type 3 protein in medial habenular nucleus, locus coeruleus and dorsal raphe. Given the importance of dorsal raphe, locus coeruleus and substantia nigra/ventral tegmental area for serotonergic, noradrenergic and dopaminergic transmission respectively, our results pose the morphological basis for further studies on the action of small conductance calcium-activated potassium channel type 3 in serotonergic, noradrenergic and dopaminergic transmission.

Diverse Roles for Auxiliary Subunits in Phosphorylation-Dependent Regulation of Mammalian Brain Voltage-Gated Potassium Channels

PubMed Central

Vacher, Helene; Trimmer, James S.

2012-01-01

Voltage-gated ion channels are a diverse family of signaling proteins that mediate rapid electrical signaling events. Among these, voltage-gated potassium or Kv channels are the most diverse, in part due to the large number of principal (or α) subunits and auxiliary subunits that can assemble in different combinations to generate Kv channel complexes with distinct structures and functions. The diversity of Kv channels underlies much of the variability in the active properties between different mammalian central neurons, and the dynamic changes that lead to experience-dependent plasticity in intrinsic excitability. Recent studies have revealed that Kv channel α subunits and auxiliary subunits are extensively phosphorylated, contributing to additional structural and functional diversity. Here we highlight recent studies that show that auxiliary subunits exert some of their profound effects on dendritic Kv4 and axonal Kv1 channels through phosphorylation-dependent mechanisms, either due to phosphorylation on the auxiliary subunit itself, or by influencing the extent and/or impact of α subunit phosphorylation. The complex effects of auxiliary subunits and phosphorylation provide a potent mechanism to generate additional diversity in the structure and function of Kv4 and Kv1 channels, as well as allowing for dynamic reversible regulation of these important ion channels. PMID:21822597

Elimination of fast inactivation in Kv4 A-type potassium channels by an auxiliary subunit domain.

PubMed

Holmqvist, Mats H; Cao, Jie; Hernandez-Pineda, Ricardo; Jacobson, Michael D; Carroll, Karen I; Sung, M Amy; Betty, Maria; Ge, Pei; Gilbride, Kevin J; Brown, Melissa E; Jurman, Mark E; Lawson, Deborah; Silos-Santiago, Inmaculada; Xie, Yu; Covarrubias, Manuel; Rhodes, Kenneth J; Distefano, Peter S; An, W Frank

2002-01-22

The Kv4 A-type potassium currents contribute to controlling the frequency of slow repetitive firing and back-propagation of action potentials in neurons and shape the action potential in heart. Kv4 currents exhibit rapid activation and inactivation and are specifically modulated by K-channel interacting proteins (KChIPs). Here we report the discovery and functional characterization of a modular K-channel inactivation suppressor (KIS) domain located in the first 34 aa of an additional KChIP (KChIP4a). Coexpression of KChIP4a with Kv4 alpha-subunits abolishes fast inactivation of the Kv4 currents in various cell types, including cerebellar granule neurons. Kinetic analysis shows that the KIS domain delays Kv4.3 opening, but once the channel is open, it disrupts rapid inactivation and slows Kv4.3 closing. Accordingly, KChIP4a increases the open probability of single Kv4.3 channels. The net effects of KChIP4a and KChIP1-3 on Kv4 gating are quite different. When both KChIP4a and KChIP1 are present, the Kv4.3 current shows mixed inactivation profiles dependent on KChIP4a/KChIP1 ratios. The KIS domain effectively converts the A-type Kv4 current to a slowly inactivating delayed rectifier-type potassium current. This conversion is opposite to that mediated by the Kv1-specific "ball" domain of the Kv beta 1 subunit. Together, these results demonstrate that specific auxiliary subunits with distinct functions actively modulate gating of potassium channels that govern membrane excitability.

Molecular basis of potassium channels in pancreatic duct epithelial cells

PubMed Central

Hayashi, Mikio; Novak, Ivana

2013-01-01

Potassium channels regulate excitability, epithelial ion transport, proliferation, and apoptosis. In pancreatic ducts, K+ channels hyperpolarize the membrane potential and provide the driving force for anion secretion. This review focuses on the molecular candidates of functional K+ channels in pancreatic duct cells, including KCNN4 (KCa3.1), KCNMA1 (KCa1.1), KCNQ1 (Kv7.1), KCNH2 (Kv11.1), KCNH5 (Kv10.2), KCNT1 (KCa4.1), KCNT2 (KCa4.2), and KCNK5 (K2P5.1). We will give an overview of K+ channels with respect to their electrophysiological and pharmacological characteristics and regulation, which we know from other cell types, preferably in epithelia, and, where known, their identification and functions in pancreatic ducts and in adenocarcinoma cells. We conclude by pointing out some outstanding questions and future directions in pancreatic K+ channel research with respect to the physiology of secretion and pancreatic pathologies, including pancreatitis, cystic fibrosis, and cancer, in which the dysregulation or altered expression of K+ channels may be of importance. PMID:23962792

Functional significance of M-type potassium channels in nociceptive cutaneous sensory endings

PubMed Central

Passmore, Gayle M.; Reilly, Joanne M.; Thakur, Matthew; Keasberry, Vanessa N.; Marsh, Stephen J.; Dickenson, Anthony H.; Brown, David A.

2012-01-01

M-channels carry slowly activating potassium currents that regulate excitability in a variety of central and peripheral neurons. Functional M-channels and their Kv7 channel correlates are expressed throughout the somatosensory nervous system where they may play an important role in controlling sensory nerve activity. Here we show that Kv7.2 immunoreactivity is expressed in the peripheral terminals of nociceptive primary afferents. Electrophysiological recordings from single afferents in vitro showed that block of M-channels by 3 μM XE991 sensitized Aδ- but not C-fibers to noxious heat stimulation and induced spontaneous, ongoing activity at 32°C in many Aδ-fibers. These observations were extended in vivo: intraplantar injection of XE991 selectively enhanced the response of deep dorsal horn (DH) neurons to peripheral mid-range mechanical and higher range thermal stimuli, consistent with a selective effect on Aδ-fiber peripheral terminals. These results demonstrate an important physiological role of M-channels in controlling nociceptive Aδ-fiber responses and provide a rationale for the nocifensive behaviors that arise following intraplantar injection of the M-channel blocker XE991. PMID:22593734

Tyrosine Phosphatases ε and α Perform Specific and Overlapping Functions in Regulation of Voltage-gated Potassium Channels in Schwann Cells

PubMed Central

Tiran, Zohar; Peretz, Asher; Sines, Tal; Shinder, Vera; Sap, Jan; Attali, Bernard

2006-01-01

Tyrosine phosphatases (PTPs) ε and α are closely related and share several molecular functions, such as regulation of Src family kinases and voltage-gated potassium (Kv) channels. Functional interrelationships between PTPε and PTPα and the mechanisms by which they regulate K+ channels and Src were analyzed in vivo in mice lacking either or both PTPs. Lack of either PTP increases Kv channel activity and phosphorylation in Schwann cells, indicating these PTPs inhibit Kv current amplitude in vivo. Open probability and unitary conductance of Kv channels are unchanged, suggesting an effect on channel number or organization. PTPα inhibits Kv channels more strongly than PTPε; this correlates with constitutive association of PTPα with Kv2.1, driven by membranal localization of PTPα. PTPα, but not PTPε, activates Src in sciatic nerve extracts, suggesting Src deregulation is not responsible exclusively for the observed phenotypes and highlighting an unexpected difference between both PTPs. Developmentally, sciatic nerve myelination is reduced transiently in mice lacking either PTP and more so in mice lacking both PTPs, suggesting both PTPs support myelination but are not fully redundant. We conclude that PTPε and PTPα differ significantly in their regulation of Kv channels and Src in the system examined and that similarity between PTPs does not necessarily result in full functional redundancy in vivo. PMID:16870705

The renal TRPV4 channel is essential for adaptation to increased dietary potassium

PubMed Central

Mamenko, Mykola; Boukelmoune, Nabila; Tomilin, Viktor; Zaika, Oleg; Jensen, V. Behrana; O’Neil, Roger G.; Pochynyuk, Oleh

2016-01-01

To maintain potassium homeostasis, kidneys exert flow-dependent potassium secretion to facilitate kaliuresis in response to elevated dietary potassium intake. This process involves stimulation of calcium-activated large conductance maxi-K (BK) channels in the distal nephron, namely the connecting tubule and the collecting duct. Recent evidence suggests that the TRPV4 channel is a critical determinant of flow-dependent intracellular calcium elevations in these segments of the renal tubule. Here, we demonstrate that elevated dietary potassium intake (five percent potassium) increases renal TRPV4 mRNA and protein levels in an aldosterone-dependent manner and causes redistribution of the channel to the apical plasma membrane in native collecting duct cells. This, in turn, leads to augmented TRPV4-mediated flow-dependent calcium ion responses in freshly isolated split-opened collecting ducts from mice fed the high potassium diet. Genetic TRPV4 ablation greatly diminished BK channel activity in collecting duct cells pointing to a reduced capacity to excrete potassium. Consistently, elevated potassium intake induced hyperkalemia in TRPV4 knockout mice due to deficient renal potassium excretion. Thus, regulation of TRPV4 activity in the distal nephron by dietary potassium is an indispensable component of whole body potassium balance. PMID:28187982

Coupling of activation and inactivation gate in a K+-channel: potassium and ligand sensitivity

PubMed Central

Ader, Christian; Schneider, Robert; Hornig, Sönke; Velisetty, Phanindra; Vardanyan, Vitya; Giller, Karin; Ohmert, Iris; Becker, Stefan; Pongs, Olaf; Baldus, Marc

2009-01-01

Potassium (K+)-channel gating is choreographed by a complex interplay between external stimuli, K+ concentration and lipidic environment. We combined solid-state NMR and electrophysiological experiments on a chimeric KcsA–Kv1.3 channel to delineate K+, pH and blocker effects on channel structure and function in a membrane setting. Our data show that pH-induced activation is correlated with protonation of glutamate residues at or near the activation gate. Moreover, K+ and channel blockers distinctly affect the open probability of both the inactivation gate comprising the selectivity filter of the channel and the activation gate. The results indicate that the two gates are coupled and that effects of the permeant K+ ion on the inactivation gate modulate activation-gate opening. Our data suggest a mechanism for controlling coordinated and sequential opening and closing of activation and inactivation gates in the K+-channel pore. PMID:19661921

Neuroprotective role of ATP-sensitive potassium channels in cerebral ischemia

PubMed Central

Sun, Hong-shuo; Feng, Zhong-ping

2013-01-01

ATP-sensitive potassium (KATP) channels are weak, inward rectifiers that couple metabolic status to cell membrane electrical activity, thus modulating many cellular functions. An increase in the ADP/ATP ratio opens KATP channels, leading to membrane hyperpolarization. KATP channels are ubiquitously expressed in neurons located in different regions of the brain, including the hippocampus and cortex. Brief hypoxia triggers membrane hyperpolarization in these central neurons. In vivo animal studies confirmed that knocking out the Kir6.2 subunit of the KATP channels increases ischemic infarction, and overexpression of the Kir6.2 subunit reduces neuronal injury from ischemic insults. These findings provide the basis for a practical strategy whereby activation of endogenous KATP channels reduces cellular damage resulting from cerebral ischemic stroke. KATP channel modulators may prove to be clinically useful as part of a combination therapy for stroke management in the future. PMID:23123646

Mechanism underlying selective regulation of G protein-gated inwardly rectifying potassium channels by the psychostimulant-sensitive sorting nexin 27

PubMed Central

Balana, Bartosz; Maslennikov, Innokentiy; Kwiatkowski, Witek; Stern, Kalyn M.; Bahima, Laia; Choe, Senyon; Slesinger, Paul A.

2011-01-01

G protein-gated inwardly rectifying potassium (GIRK) channels are important gatekeepers of neuronal excitability. The surface expression of neuronal GIRK channels is regulated by the psychostimulant-sensitive sorting nexin 27 (SNX27) protein through a class I (-X-Ser/Thr-X-Φ, where X is any residue and Φ is a hydrophobic amino acid) PDZ-binding interaction. The G protein-insensitive inward rectifier channel (IRK1) contains the same class I PDZ-binding motif but associates with a different synaptic PDZ protein, postsynaptic density protein 95 (PSD95). The mechanism by which SNX27 and PSD95 discriminate these channels was previously unclear. Using high-resolution structures coupled with biochemical and functional analyses, we identified key amino acids upstream of the channel's canonical PDZ-binding motif that associate electrostatically with a unique structural pocket in the SNX27-PDZ domain. Changing specific charged residues in the channel's carboxyl terminus or in the PDZ domain converts the selective association and functional regulation by SNX27. Elucidation of this unique interaction site between ion channels and PDZ-containing proteins could provide a therapeutic target for treating brain diseases. PMID:21422294

Functional characterization of Kv11.1 (hERG) potassium channels split in the voltage-sensing domain.

PubMed

de la Peña, Pilar; Domínguez, Pedro; Barros, Francisco

2018-03-23

Voltage-dependent KCNH family potassium channel functionality can be reconstructed using non-covalently linked voltage-sensing domain (VSD) and pore modules (split channels). However, the necessity of a covalent continuity for channel function has not been evaluated at other points within the two functionally independent channel modules. We find here that by cutting Kv11.1 (hERG, KCNH2) channels at the different loops linking the transmembrane spans of the channel core, not only channels split at the S4-S5 linker level, but also those split at the intracellular S2-S3 and the extracellular S3-S4 loops, yield fully functional channel proteins. Our data indicate that albeit less markedly, channels split after residue 482 in the S2-S3 linker resemble the uncoupled gating phenotype of those split at the C-terminal end of the VSD S4 transmembrane segment. Channels split after residues 514 and 518 in the S3-S4 linker show gating characteristics similar to those of the continuous wild-type channel. However, breaking the covalent link at this level strongly accelerates the voltage-dependent accessibility of a membrane impermeable methanethiosulfonate reagent to an engineered cysteine at the N-terminal region of the S4 transmembrane helix. Thus, besides that of the S4-S5 linker, structural integrity of the intracellular S2-S3 linker seems to constitute an important factor for proper transduction of VSD rearrangements to opening and closing the cytoplasmic gate. Furthermore, our data suggest that the short and probably rigid characteristics of the extracellular S3-S4 linker are not an essential component of the Kv11.1 voltage sensing machinery.

Conformational heterogeneity in closed and open states of the KcsA potassium channel in lipid bicelles

PubMed Central

Kim, Dorothy M.; Dikiy, Igor; Upadhyay, Vikrant; Posson, David J.

2016-01-01

The process of ion channel gating—opening and closing—involves local and global structural changes in the channel in response to external stimuli. Conformational changes depend on the energetic landscape that underlies the transition between closed and open states, which plays a key role in ion channel gating. For the prokaryotic, pH-gated potassium channel KcsA, closed and open states have been extensively studied using structural and functional methods, but the dynamics within each of these functional states as well as the transition between them is not as well understood. In this study, we used solution nuclear magnetic resonance (NMR) spectroscopy to investigate the conformational transitions within specific functional states of KcsA. We incorporated KcsA channels into lipid bicelles and stabilized them into a closed state by using either phosphatidylcholine lipids, known to favor the closed channel, or mutations designed to trap the channel shut by disulfide cross-linking. A distinct state, consistent with an open channel, was uncovered by the addition of cardiolipin lipids. Using selective amino acid labeling at locations within the channel that are known to move during gating, we observed at least two different slowly interconverting conformational states for both closed and open channels. The pH dependence of these conformations and the predictable disruptions to this dependence observed in mutant channels with altered pH sensing highlight the importance of conformational heterogeneity for KcsA gating. PMID:27432996

The sodium-activated potassium channel Slack is required for optimal cognitive flexibility in mice.

PubMed

Bausch, Anne E; Dieter, Rebekka; Nann, Yvette; Hausmann, Mario; Meyerdierks, Nora; Kaczmarek, Leonard K; Ruth, Peter; Lukowski, Robert

2015-07-01

Kcnt1 encoded sodium-activated potassium channels (Slack channels) are highly expressed throughout the brain where they modulate the firing patterns and general excitability of many types of neurons. Increasing evidence suggests that Slack channels may be important for higher brain functions such as cognition and normal intellectual development. In particular, recent findings have shown that human Slack mutations produce very severe intellectual disability and that Slack channels interact directly with the Fragile X mental retardation protein (FMRP), a protein that when missing or mutated results in Fragile X syndrome (FXS), the most common form of inherited intellectual disability and autism in humans. We have now analyzed a recently developed Kcnt1 null mouse model in several behavioral tasks to assess which aspects of memory and learning are dependent on Slack. We demonstrate that Slack deficiency results in mildly altered general locomotor activity, but normal working memory, reference memory, as well as cerebellar control of motor functions. In contrast, we find that Slack channels are required for cognitive flexibility, including reversal learning processes and the ability to adapt quickly to unfamiliar situations and environments. Our data reveal that hippocampal-dependent spatial learning capabilities require the proper function of Slack channels. © 2015 Bausch et al.; Published by Cold Spring Harbor Laboratory Press.

The sodium-activated potassium channel Slack is required for optimal cognitive flexibility in mice

PubMed Central

Bausch, Anne E.; Dieter, Rebekka; Nann, Yvette; Hausmann, Mario; Meyerdierks, Nora; Kaczmarek, Leonard K.

2015-01-01

Kcnt1 encoded sodium-activated potassium channels (Slack channels) are highly expressed throughout the brain where they modulate the firing patterns and general excitability of many types of neurons. Increasing evidence suggests that Slack channels may be important for higher brain functions such as cognition and normal intellectual development. In particular, recent findings have shown that human Slack mutations produce very severe intellectual disability and that Slack channels interact directly with the Fragile X mental retardation protein (FMRP), a protein that when missing or mutated results in Fragile X syndrome (FXS), the most common form of inherited intellectual disability and autism in humans. We have now analyzed a recently developed Kcnt1 null mouse model in several behavioral tasks to assess which aspects of memory and learning are dependent on Slack. We demonstrate that Slack deficiency results in mildly altered general locomotor activity, but normal working memory, reference memory, as well as cerebellar control of motor functions. In contrast, we find that Slack channels are required for cognitive flexibility, including reversal learning processes and the ability to adapt quickly to unfamiliar situations and environments. Our data reveal that hippocampal-dependent spatial learning capabilities require the proper function of Slack channels. PMID:26077685

The renal TRPV4 channel is essential for adaptation to increased dietary potassium.

PubMed

Mamenko, Mykola V; Boukelmoune, Nabila; Tomilin, Viktor N; Zaika, Oleg L; Jensen, V Behrana; O'Neil, Roger G; Pochynyuk, Oleh M

2017-06-01

To maintain potassium homeostasis, kidneys exert flow-dependent potassium secretion to facilitate kaliuresis in response to elevated dietary potassium intake. This process involves stimulation of calcium-activated large conductance maxi-K (BK) channels in the distal nephron, namely the connecting tubule and the collecting duct. Recent evidence suggests that the TRPV4 channel is a critical determinant of flow-dependent intracellular calcium elevations in these segments of the renal tubule. Here, we demonstrate that elevated dietary potassium intake (five percent potassium) increases renal TRPV4 mRNA and protein levels in an aldosterone-dependent manner and causes redistribution of the channel to the apical plasma membrane in native collecting duct cells. This, in turn, leads to augmented TRPV4-mediated flow-dependent calcium ion responses in freshly isolated split-opened collecting ducts from mice fed the high potassium diet. Genetic TRPV4 ablation greatly diminished BK channel activity in collecting duct cells pointing to a reduced capacity to excrete potassium. Consistently, elevated potassium intake induced hyperkalemia in TRPV4 knockout mice due to deficient renal potassium excretion. Thus, regulation of TRPV4 activity in the distal nephron by dietary potassium is an indispensable component of whole body potassium balance. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Free RCK arrangement in Kch, a putative escherichia coli potassium channel, as suggested by electron crystallography.

PubMed

Kuang, Qie; Purhonen, Pasi; Jegerschöld, Caroline; Koeck, Philip J B; Hebert, Hans

2015-01-06

The ligand-gated potassium channels are stimulated by various kinds of messengers. Previous studies showed that ligand-gated potassium channels containing RCK domains (the regulator of the conductance of potassium ion) form a dimer of tetramer structure through the RCK octameric gating ring in the presence of detergent. Here, we have analyzed the structure of Kch, a channel of this type from Escherichia coli, in a lipid environment using electron crystallography. By combining information from the 3D map of the transmembrane part of the protein and docking of an atomic model of a potassium channel, we conclude that the RCK domains face the solution and that an RCK octameric gating ring arrangement does not form under our crystallization condition. Our findings may be applied to other potassium channels that have an RCK gating ring arrangement. Copyright © 2015 Elsevier Ltd. All rights reserved.

Delayed rectifier potassium channels are involved in SO2 derivative-induced hippocampal neuronal injury.

PubMed

Li, Guangke; Sang, Nan

2009-01-01

Recent studies implicate the possible neurotoxicity of SO(2), however, its mechanisms remain unclear. In the present study, we investigated SO(2) derivative-induced effect on delayed rectifier potassium channels (I(K)) and cellular death/apoptosis in primary cultured hippocampal neurons. The results demonstrate that SO(2) derivatives (NaHSO(3) and Na(2)SO(3), 3:1M/M) effectively augmented I(K) and promoted the activation of delayed rectifier potassium channels. Also, SO(2) derivatives increased neuronal death percentage and contributed to the formation of DNA ladder in concentration-dependent manners. Interestingly, the neuronal death and DNA ladder formation, caused by SO(2) derivatives, could be attenuated by the delayed rectifier potassium channel blocker (tetraethylammonium, TEA), but not by the transient outward potassium channel blocker (4-aminopyridine, 4-AP). It implies that stimulating delayed rectifier potassium channels were involved in SO(2) derivative-caused hippocampal neuronal insults, and blocking these channels might be one of the possibly clinical treatment for SO(2)-caused neuronal dysfunction.

Intractable hyperkalemia due to nicorandil induced potassium channel syndrome.

PubMed

Chowdhry, Vivek; Mohanty, B B

2015-01-01

Nicorandil is a commonly used antianginal agent, which has both nitrate-like and ATP-sensitive potassium (K ATP ) channel activator properties. Activation of potassium channels by nicorandil causes expulsion of potassium ions into the extracellular space leading to membrane hyperpolarization, closure of voltage-gated calcium channels and finally vasodilatation. However, on the other hand, being an activator of K ATP channel, it can expel K + ions out of the cells and can cause hyperkalemia. Here, we report a case of nicorandil induced hyperkalemia unresponsive to medical treatment in a patient with diabetic nephropathy.

Roderick MacKinnon and Ion Channels - Potassium Channels and Sodium

Science.gov Websites

very first potassium channel structure, which revealed the way that positively charged potassium ions explain how a fundamental feedback loop worked. Now, with the structure of the voltage-dependent ion channel, based on research carried out at The Rockefeller University, the National Synchrotron Light

[Architecture of receptor-operated ionic channels of biological membranes].

PubMed

Bregestovski, P D

2011-01-01

Ion channels of biological membranes are the key proteins, which provide bioelectric functioning of living systems. These proteins are homo- or heterooligomers assembled from several identical or different subunits. Understanding the architectural organization and functioning of ion channels has been significantly extended due to resolving the crystal structure of several types of voltage-gated and receptor-operated channels. This review summarizes the information obtained from crystal structures of potassium, nicotinic acetylcholine receptor, P2X, and other ligand-gated ion channels. Despite the differences in the function, topology, ionic selectivity, and the subunit stoichiometry, a high similarity in the principles of organization of these macromolecular complexes has been revealed.

An interview with Arthur M. "Buzz" Brown, M.D., Ph.D. by Vicki Glaser.

PubMed

Brown, Arthur M

2008-12-01

Dr. Arthur M. "Buzz" Brown is the founder and CEO of ChanTest Corporation, an ion channel company specializing in drug discovery and safety services. He is Adjunct Professor of Physiology and Biophysics, School of Medicine, Case Western Reserve University. Dr. Brown has more than 30 years of experience in ion channel structure-function relationships and their associations with human health. He established world-leading ion channel departments at University of Texas Medical Branch, Baylor College of Medicine, and Case Western Reserve University. His lab first applied liquid ion exchanger ion-selective microelectrodes to single cells, introduced the concept of membrane delimited action of G proteins on ion channels, identified the ion channel conduction pathway or pore of voltage-gated channels and inwardly rectifying potassium channels, showed that the human ether-à-go-go-related gene potassium channel was the molecular target for lethal arrhythmias associated with noncardiac drugs, and established that noncardiac drugs could also produce lethal arrhythmias by inhibiting ion channel trafficking. Dr. Brown holds eight patents on ion channel methodology and application of ion channel pharmacology to therapeutics.

Molecular and functional characterization of Anopheles gambiae inward rectifier potassium (Kir1) channels: a novel role in egg production.

PubMed

Raphemot, Rene; Estévez-Lao, Tania Y; Rouhier, Matthew F; Piermarini, Peter M; Denton, Jerod S; Hillyer, Julián F

2014-08-01

Inward rectifier potassium (Kir) channels play essential roles in regulating diverse physiological processes. Although Kir channels are encoded in mosquito genomes, their functions remain largely unknown. In this study, we identified the members of the Anopheles gambiae Kir gene family and began to investigate their function. Notably, we sequenced the A. gambiae Kir1 (AgKir1) gene and showed that it encodes all the canonical features of a Kir channel: an ion pore that is composed of a pore helix and a selectivity filter, two transmembrane domains that flank the ion pore, and the so-called G-loop. Heterologous expression of AgKir1 in Xenopus oocytes revealed that this gene encodes a functional, barium-sensitive Kir channel. Quantitative RT-PCR experiments then showed that relative AgKir1 mRNA levels are highest in the pupal stage, and that AgKir1 mRNA is enriched in the adult ovaries. Gene silencing of AgKir1 by RNA interference did not affect the survival of female mosquitoes following a blood meal, but decreased their egg output. These data provide evidence for a new role of Kir channels in mosquito fecundity, and further validates them as promising molecular targets for the development of a new class of mosquitocides to be used in vector control. Copyright © 2014 Elsevier Ltd. All rights reserved.

Hydrogen Sulfide Targets the Cys320/Cys529 Motif in Kv4.2 to Inhibit the Ito Potassium Channels in Cardiomyocytes and Regularizes Fatal Arrhythmia in Myocardial Infarction

PubMed Central

Ma, Shan-Feng; Luo, Yan; Ding, Ying-Jiong; Chen, Ying; Pu, Shi-Xin; Wu, Hang-Jing; Wang, Zhong-Feng; Tao, Bei-Bei; Wang, Wen-Wei

2015-01-01

Abstract Aims: The mechanisms underlying numerous biological roles of hydrogen sulfide (H2S) remain largely unknown. We have previously reported an inhibitory role of H2S in the L-type calcium channels in cardiomyocytes. This prompts us to examine the mechanisms underlying the potential regulation of H2S on the ion channels. Results: H2S showed a novel inhibitory effect on Ito potassium channels, and this effect was blocked by mutation at the Cys320 and/or Cys529 residues of the Kv4.2 subunit. H2S broke the disulfide bridge between a pair of oxidized cysteine residues; however, it did not modify single cysteine residues. H2S extended action potential duration in epicardial myocytes and regularized fatal arrhythmia in a rat model of myocardial infarction. H2S treatment significantly increased survival by ∼1.4-fold in the critical 2-h time window after myocardial infarction with a protection against ventricular premature beats and fatal arrhythmia. However, H2S did not change the function of other ion channels, including IK1 and INa. Innovation and Conclusion: H2S targets the Cys320/Cys529 motif in Kv4.2 to regulate the Ito potassium channels. H2S also shows a potent regularizing effect against fatal arrhythmia in a rat model of myocardial infarction. The study provides the first piece of evidence for the role of H2S in regulating Ito potassium channels and also the specific motif in an ion channel labile for H2S regulation. Antioxid. Redox Signal. 23, 129–147. PMID:25756524

The receptor-like pseudokinase MRH1 interacts with the voltage-gated potassium channel AKT2.

PubMed

Sklodowski, Kamil; Riedelsberger, Janin; Raddatz, Natalia; Riadi, Gonzalo; Caballero, Julio; Chérel, Isabelle; Schulze, Waltraud; Graf, Alexander; Dreyer, Ingo

2017-03-16

The potassium channel AKT2 plays important roles in phloem loading and unloading. It can operate as inward-rectifying channel that allows H + -ATPase-energized K + uptake. Moreover, through reversible post-translational modifications it can also function as an open, K + -selective channel, which taps a 'potassium battery', providing additional energy for transmembrane transport processes. Knowledge about proteins involved in the regulation of the operational mode of AKT2 is very limited. Here, we employed a large-scale yeast two-hybrid screen in combination with fluorescence tagging and null-allele mutant phenotype analysis and identified the plasma membrane localized receptor-like kinase MRH1/MDIS2 (AT4G18640) as interaction partner of AKT2. The phenotype of the mrh1-1 knockout plant mirrors that of akt2 knockout plants in energy limiting conditions. Electrophysiological analyses showed that MRH1/MDIS2 failed to exert any functional regulation on AKT2. Using structural protein modeling approaches, we instead gathered evidence that the putative kinase domain of MRH1/MDIS2 lacks essential sites that are indispensable for a functional kinase suggesting that MRH1/MDIS2 is a pseudokinase. We propose that MRH1/MDIS2 and AKT2 are likely parts of a bigger protein complex. MRH1 might help to recruit other, so far unknown partners, which post-translationally regulate AKT2. Additionally, MRH1 might be involved in the recognition of chemical signals.

The receptor-like pseudokinase MRH1 interacts with the voltage-gated potassium channel AKT2

NASA Astrophysics Data System (ADS)

Sklodowski, Kamil; Riedelsberger, Janin; Raddatz, Natalia; Riadi, Gonzalo; Caballero, Julio; Chérel, Isabelle; Schulze, Waltraud; Graf, Alexander; Dreyer, Ingo

2017-03-01

The potassium channel AKT2 plays important roles in phloem loading and unloading. It can operate as inward-rectifying channel that allows H+-ATPase-energized K+ uptake. Moreover, through reversible post-translational modifications it can also function as an open, K+-selective channel, which taps a ‘potassium battery’, providing additional energy for transmembrane transport processes. Knowledge about proteins involved in the regulation of the operational mode of AKT2 is very limited. Here, we employed a large-scale yeast two-hybrid screen in combination with fluorescence tagging and null-allele mutant phenotype analysis and identified the plasma membrane localized receptor-like kinase MRH1/MDIS2 (AT4G18640) as interaction partner of AKT2. The phenotype of the mrh1-1 knockout plant mirrors that of akt2 knockout plants in energy limiting conditions. Electrophysiological analyses showed that MRH1/MDIS2 failed to exert any functional regulation on AKT2. Using structural protein modeling approaches, we instead gathered evidence that the putative kinase domain of MRH1/MDIS2 lacks essential sites that are indispensable for a functional kinase suggesting that MRH1/MDIS2 is a pseudokinase. We propose that MRH1/MDIS2 and AKT2 are likely parts of a bigger protein complex. MRH1 might help to recruit other, so far unknown partners, which post-translationally regulate AKT2. Additionally, MRH1 might be involved in the recognition of chemical signals.

Role of potassium channels in chlorogenic acid-induced apoptotic volume decrease and cell cycle arrest in Candida albicans.

PubMed

Yun, JiEun; Lee, Dong Gun

2017-03-01

Chlorogenic acid (CRA) is an abundant phenolic compound in the human diet. CRA has a potent antifungal effect, inducing cell death in Candida albicans. However, there are no further studies to investigate the antifungal mechanism of CRA, associated with ion channels. To evaluate the inhibitory effects on CRA-induced cell death, C. albicans cells were pretreated with potassium and chloride channel blockers, separately. Flow cytometry was carried out to detect several hallmarks of apoptosis, such as cell cycle arrest, caspase activation, and DNA fragmentation, after staining of the cells with SYTOX green, FITC-VAD-FMK, and TUNEL. CRA caused excessive potassium efflux, and an apoptotic volume decrease (AVD) was observed. This change, in turn, induced cytosolic calcium uptake and cell cycle arrest in C. albicans. Moreover, CRA induced caspase activation and DNA fragmentation, which are considered apoptotic markers. In contrast, the potassium efflux and proapoptotic changes were inhibited when potassium channels were blocked, whereas there was no inhibitory effect when chloride channels were blocked. CRA induces potassium efflux, leading to AVD and G2/M cell cycle arrest in C. albicans. Therefore, potassium efflux via potassium channels regulates the CRA-induced apoptosis, stimulating several apoptotic processes. This study improves the understanding of the antifungal mechanism of CRA and its association with ion homeostasis, thereby pointing to a role of potassium channels in CRA-induced apoptosis. Copyright © 2016. Published by Elsevier B.V.

Distribution, expression and functional effects of small conductance Ca-activated potassium (SK) channels in rat myometrium.

PubMed

Noble, Karen; Floyd, Rachel; Shmygol, Andre; Shmygol, Anatoly; Mobasheri, A; Wray, Susan

2010-01-01

Calcium-activated potassium channels are important in a variety of smooth muscles, contributing to excitability and contractility. In the myometrium previous work has focussed on the large conductance channels (BK), and the role of small conductance channels (SK) has received scant attention, despite the finding that over-expression of an SK channel isoform (SK3) results in uterine dysfunction and delayed parturition. This study therefore characterises the expression of the three SK channel isoforms (SK1-3) in rat myometrium throughout pregnancy and investigates their effect on cytosolic [Ca] and force and compares this with that of BK channels. Consistent expression of all SK isoform transcripts and clear immunostaining of SK1-3 was found. Inhibition of SK1-3 channels (apamin, scyllatoxin) significantly inhibited outward current, caused membrane depolarisation and elicited action potentials in previously quiescent cells. Apamin or scyllatoxin increased the amplitude of [Ca] and force in spontaneously contracting myometrial strips throughout gestation. The functional effect of SK inhibition was larger than that of BK channel inhibition. Thus we show for the first time that SK1-3 channels are expressed and translated throughout pregnancy and contribute to outward current, regulate membrane potential and hence Ca signals in pregnant rat myometrium. They contribute more to quiescence that BK channels. 2009 Elsevier Ltd. All rights reserved.

Ion channel profile of TRPM8 cold receptors reveals a novel role of TASK-3 potassium channels in thermosensation

PubMed Central

Morenilla-Palao, Cruz; Luis, Enoch; Fernández-Peña, Carlos; Quintero, Eva; Weaver, Janelle L.; Bayliss, Douglas A.; Viana, Félix

2017-01-01

Summary Animals sense cold ambient temperatures through the activation of peripheral thermoreceptors that express TRPM8, a cold- and menthol-activated ion channel. These receptors can discriminate a very wide range of temperatures from innocuous to noxious. The molecular mechanism responsible for the variable sensitivity of individual cold receptors to temperature is unclear. To address this question, we performed a detailed ion channel expression analysis of cold sensitive neurons, combining BAC transgenesis with a molecular profiling approach in FACS purified TRPM8 neurons. We found that TASK-3 leak potassium channels are highly enriched in a subpopulation of these sensory neurons. The thermal threshold of TRPM8 cold neurons is decreased during TASK-3 blockade and in mice lacking TASK-3 and, most importantly, these mice display hypersensitivity to cold. Our results demonstrate a novel role of TASK-3 channels in thermosensation, showing that a channel-based combinatorial strategy in TRPM8 cold thermoreceptors leads to molecular specialization and functional diversity. PMID:25199828

Role of the activation gate in determining the extracellular potassium dependency of block of HERG by trapped drugs.

PubMed

Pareja, Kristeen; Chu, Elaine; Dodyk, Katrina; Richter, Kristofer; Miller, Alan

2013-01-01

Drug induced long QT syndrome (diLQTS) results primarily from block of the cardiac potassium channel HERG (human-ether-a-go-go related gene). In some cases long QT syndrome can result in the lethal arrhythmia torsade de pointes, an arrhythmia characterized by a rapid heart rate and severely compromised cardiac output. Many patients requiring medication present with serum potassium abnormalities due to a variety of conditions including gastrointestinal dysfunction, renal and endocrine disorders, diuretic use, and aging. Extracellular potassium influences HERG channel inactivation and can alter block of HERG by some drugs. However, block of HERG by a number of drugs is not sensitive to extracellular potassium. In this study, we show that block of WT HERG by bepridil and terfenadine, two drugs previously shown to be trapped inside the HERG channel after the channel closes, is insensitive to extracellular potassium over the range of 0 mM to 20 mM. We also show that bepridil block of the HERG mutant D540K, a mutant channel that is unable to trap drugs, is dependent on extracellular potassium, correlates with the permeant ion, and is independent of HERG inactivation. These results suggest that the lack of extracellular potassium dependency of block of HERG by some drugs may in part be related to the ability of these drugs to be trapped inside the channel after the channel closes.

Rearrangement of potassium ions and Kv1.1/Kv1.2 potassium channels in regenerating axons following end-to-end neurorrhaphy: ionic images from TOF-SIMS.

PubMed

Liu, Chiung-Hui; Chang, Hung-Ming; Wu, Tsung-Huan; Chen, Li-You; Yang, Yin-Shuo; Tseng, To-Jung; Liao, Wen-Chieh

2017-10-01

The voltage-gated potassium channels Kv1.1 and Kv1.2 that cluster at juxtaparanodal (JXP) regions are essential in the regulation of nerve excitability and play a critical role in axonal conduction. When demyelination occurs, Kv1.1/Kv1.2 activity increases, suppressing the membrane potential nearly to the equilibrium potential of K + , which results in an axonal conduction blockade. The recovery of K + -dependent communication signals and proper clustering of Kv1.1/Kv1.2 channels at JXP regions may directly reflect nerve regeneration following peripheral nerve injury. However, little is known about potassium channel expression and its relationship with the dynamic potassium ion distribution at the node of Ranvier during the regenerative process of peripheral nerve injury (PNI). In the present study, end-to-end neurorrhaphy (EEN) was performed using an in vivo model of PNI. The distribution of K + at regenerating axons following EEN was detected by time-of-flight secondary-ion mass spectrometry. The specific localization and expression of Kv1.1/Kv1.2 channels were examined by confocal microscopy and western blotting. Our data showed that the re-establishment of K + distribution and intensity was correlated with the functional recovery of compound muscle action potential morphology in EEN rats. Furthermore, the re-clustering of Kv1.1/1.2 channels 1 and 3 months after EEN at the nodal region of the regenerating nerve corresponded to changes in the K + distribution. This study provided direct evidence of K + distribution in regenerating axons for the first time. We proposed that the Kv1.1/Kv1.2 channels re-clustered at the JXP regions of regenerating axons are essential for modulating the proper patterns of K + distribution in axons for maintaining membrane potential stability after EEN.

Nuclear BK Channels Regulate Gene Expression via the Control of Nuclear Calcium Signaling

PubMed Central

Li, Boxing; Jie, Wei; Huang, Lianyan; Wei, Peng; Li, Shuji; Luo, Zhengyi; Friedman, Allyson K.; Meredith, Andrea L.; Han, Ming-Hu; Zhu, Xin-Hong; Gao, Tian-Ming

2014-01-01

Ion channels are essential for the regulation of neuronal functions. The significance of plasma membrane, mitochondrial, endoplasmic reticulum, and lysosomal ion channels in the regulation of Ca2+ is well established. In contrast, surprisingly less is known about the function of ion channels on the nuclear envelope (NE). Here we demonstrate the presence of functional large-conductance, calcium-activated potassium channels (BK channels) on the NE of rodent hippocampal neurons. Functionally blockade of nuclear BK channels (nBK channels) induces NE-derived Ca2+ release, nucleoplasmic Ca2+ elevation, and cAMP response element binding protein (CREB)-dependent transcription. More importantly, blockade of nBK channels regulates nuclear Ca2+-sensitive gene expression and promotes dendritic arborization in a nuclear Ca2+-dependent manner. These results suggest that nBK channel functions as a molecular linker between neuronal activity and nuclear Ca2+ to convey the signals from synapse to nucleus and is a new modulator for synaptic activity-dependent neuronal functions at the NE level. PMID:24952642

Mechanism of resveratrol-induced relaxation in the human gallbladder.

PubMed

Tsai, Ching-Chung; Lee, Ming-Che; Tey, Shu-Leei; Liu, Ching-Wen; Huang, Shih-Che

2017-05-08

Resveratrol is a polyphenolic compound extracted from plants and is also a constituent of red wine. Resveratrol produces relaxation of vascular smooth muscle and may prevent cardiovascular diseases. Although resveratrol has been reported to cause relaxation of the guinea pig gallbladder, limited data are available about the effect of resveratrol on the gallbladder smooth muscle in humans. The purpose of this study was to investigate the relaxation effects of resveratrol in human gallbladder muscle strips. We studied the relaxant effects of resveratrol in human gallbladder. In addition, we also investigated mechanism of resveratrol-induced relaxation in human gallbladder by tetraethylammonium (a non-selective potassium channels blocker), iberiotoxin (an inhibitor of large conductance calcium-activated potassium channel), glibenclamide (an ATP-sensitive potassium channel blocker), charybdotoxin (an inhibitor of large conductance calcium-activated potassium channels and slowly inactivating voltage-gated potassium channels), apamine (a selective inhibitor of the small conductance calcium-activated potassium channel), KT 5720 (a cAMP-dependent protein kinase A inhibitor), KT 5823 (a cGMP-dependent protein kinase G inhibitor), NG-Nitro-L-arginine (a competitive inhibitor of nitric oxide synthase), tetrodotoxin (a selective neuronal Na + channel blocker), and ω-conotoxin GVIA (a selective neuronal Ca 2+ channel blocker). The present study showed that resveratrol has relaxant effects in human gallbladder muscle strips. In addition, we found that resveratrol-induced relaxation in human gallbladder is associated with nitric oxide, ATP-sensitive potassium channel, and large conductance calcium-activated potassium channel pathways. This study provides the first evidence concerning the relaxant effects of resveratrol in human gallbladder muscle strips. Furthermore, these results demonstrate that resveratrol is a potential new drug or health supplement in the treatment of biliary colic.

Cantú Syndrome Resulting from Activating Mutation in the KCNJ8 Gene

PubMed Central

Cooper, Paige E.; Reutter, Heiko; Woelfle, Joachim; Engels, Hartmut; Grange, Dorothy K.; van Haaften, Gijs; van Bon, Bregje W.; Hoischen, Alexander; Nichols, Colin G.

2014-01-01

ATP-sensitive potassium (KATP) channels, composed of inward-rectifying potassium channel subunits (Kir6.1 and Kir6.2, encoded by KCNJ8 and KCNJ11, respectively) and regulatory sulfonylurea receptor (SUR1 and SUR2, encoded by ABCC8 and ABCC9, respectively), couple metabolism to excitability in multiple tissues. Mutations in ABCC9 cause Cantú syndrome, a distinct multi-organ disease, potentially via enhanced KATP channel activity. We screened KCNJ8 in an ABCC9 mutation-negative patient who also exhibited clinical hallmarks of Cantú syndrome (hypertrichosis, macrosomia, macrocephaly, coarse facial appearance, cardiomegaly, and skeletal abnormalities). We identified a de novo missense mutation encoding Kir6.1[p.Cys176Ser] in the patient. Kir6.1[p.Cys176Ser] channels exhibited markedly higher activity than wild-type channels, as a result of reduced ATP sensitivity, whether co-expressed with SUR1 or SUR2A subunits. Our results identify a novel causal gene in Cantú syndrome, but also demonstrate that the cardinal features of the disease result from gain of KATP channel function, not from Kir6-independent SUR2 function. PMID:24700710

Cantú syndrome resulting from activating mutation in the KCNJ8 gene.

PubMed

Cooper, Paige E; Reutter, Heiko; Woelfle, Joachim; Engels, Hartmut; Grange, Dorothy K; van Haaften, Gijs; van Bon, Bregje W; Hoischen, Alexander; Nichols, Colin G

2014-07-01

ATP-sensitive potassium (KATP ) channels, composed of inward-rectifying potassium channel subunits (Kir6.1 and Kir6.2, encoded by KCNJ8 and KCNJ11, respectively) and regulatory sulfonylurea receptor (SUR1 and SUR2, encoded by ABCC8 and ABCC9, respectively), couple metabolism to excitability in multiple tissues. Mutations in ABCC9 cause Cantú syndrome (CS), a distinct multiorgan disease, potentially via enhanced KATP channel activity. We screened KCNJ8 in an ABCC9 mutation-negative patient who also exhibited clinical hallmarks of CS (hypertrichosis, macrosomia, macrocephaly, coarse facial appearance, cardiomegaly, and skeletal abnormalities). We identified a de novo missense mutation encoding Kir6.1[p.Cys176Ser] in the patient. Kir6.1[p.Cys176Ser] channels exhibited markedly higher activity than wild-type channels, as a result of reduced ATP sensitivity, whether coexpressed with SUR1 or SUR2A subunits. Our results identify a novel causal gene in CS, but also demonstrate that the cardinal features of the disease result from gain of KATP channel function, not from a Kir6-independent SUR2 function. © 2014 WILEY PERIODICALS, INC.

Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus

PubMed Central

Bukiya, Anna N.; Noskov, Sergei; Rosenhouse-Dantsker, Avia

2017-01-01

Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. PMID:28213520

Persistent anterograde amnesia following limbic encephalitis associated with antibodies to the voltage-gated potassium channel complex.

PubMed

Butler, Christopher R; Miller, Thomas D; Kaur, Manveer S; Baker, Ian W; Boothroyd, Georgie D; Illman, Nathan A; Rosenthal, Clive R; Vincent, Angela; Buckley, Camilla J

2014-04-01

Limbic encephalitis (LE) associated with antibodies to the voltage-gated potassium channel complex (VGKC) is a potentially reversible cause of cognitive impairment. Despite the prominence of cognitive dysfunction in this syndrome, little is known about patients' neuropsychological profile at presentation or their long-term cognitive outcome. We used a comprehensive neuropsychological test battery to evaluate cognitive function longitudinally in 19 patients with VGKC-LE. Before immunotherapy, the group had significant impairment of memory, processing speed and executive function, whereas language and perceptual organisation were intact. At follow-up, cognitive impairment was restricted to the memory domain, with processing speed and executive function having returned to the normal range. Residual memory function was predicted by the antibody titre at presentation. The results show that, despite broad cognitive dysfunction in the acute phase, patients with VGKC-LE often make a substantial recovery with immunotherapy but may be left with permanent anterograde amnesia.

Potassium channel gene associations with joint processing speed and white matter impairments in schizophrenia.

PubMed

Bruce, H A; Kochunov, P; Paciga, S A; Hyde, C L; Chen, X; Xie, Z; Zhang, B; Xi, H S; O'Donnell, P; Whelan, C; Schubert, C R; Bellon, A; Ament, S A; Shukla, D K; Du, X; Rowland, L M; O'Neill, H; Hong, L E

2017-06-01

Patients with schizophrenia show decreased processing speed on neuropsychological testing and decreased white matter integrity as measured by diffusion tensor imaging, two traits shown to be both heritable and genetically associated indicating that there may be genes that influence both traits as well as schizophrenia disease risk. The potassium channel gene family is a reasonable candidate to harbor such a gene given the prominent role potassium channels play in the central nervous system in signal transduction, particularly in myelinated axons. We genotyped members of the large potassium channel gene family focusing on putatively functional single nucleotide polymorphisms (SNPs) in a population of 363 controls, 194 patients with schizophrenia spectrum disorder (SSD) and 28 patients with affective disorders with psychotic features who completed imaging and neuropsychological testing. We then performed three association analyses using three phenotypes - processing speed, whole-brain white matter fractional anisotropy (FA) and schizophrenia spectrum diagnosis. We extracted SNPs showing an association at a nominal P value of <0.05 with all three phenotypes in the expected direction: decreased processing speed, decreased FA and increased risk of SSD. A single SNP, rs8234, in the 3' untranslated region of voltage-gated potassium channel subfamily Q member 1 (KCNQ1) was identified. Rs8234 has been shown to affect KCNQ1 expression levels, and KCNQ1 levels have been shown to affect neuronal action potentials. This exploratory analysis provides preliminary data suggesting that KCNQ1 may contribute to the shared risk for diminished processing speed, diminished white mater integrity and increased risk of schizophrenia. © 2017 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

Ion channels in plants.

PubMed

Hedrich, Rainer

2012-10-01

Since the first recordings of single potassium channel activities in the plasma membrane of guard cells more than 25 years ago, patch-clamp studies discovered a variety of ion channels in all cell types and plant species under inspection. Their properties differed in a cell type- and cell membrane-dependent manner. Guard cells, for which the existence of plant potassium channels was initially documented, advanced to a versatile model system for studying plant ion channel structure, function, and physiology. Interestingly, one of the first identified potassium-channel genes encoding the Shaker-type channel KAT1 was shown to be highly expressed in guard cells. KAT1-type channels from Arabidopsis thaliana and its homologs from other species were found to encode the K(+)-selective inward rectifiers that had already been recorded in early patch-clamp studies with guard cells. Within the genome era, additional Arabidopsis Shaker-type channels appeared. All nine members of the Arabidopsis Shaker family are localized at the plasma membrane, where they either operate as inward rectifiers, outward rectifiers, weak voltage-dependent channels, or electrically silent, but modulatory subunits. The vacuole membrane, in contrast, harbors a set of two-pore K(+) channels. Just very recently, two plant anion channel families of the SLAC/SLAH and ALMT/QUAC type were identified. SLAC1/SLAH3 and QUAC1 are expressed in guard cells and mediate Slow- and Rapid-type anion currents, respectively, that are involved in volume and turgor regulation. Anion channels in guard cells and other plant cells are key targets within often complex signaling networks. Here, the present knowledge is reviewed for the plant ion channel biology. Special emphasis is drawn to the molecular mechanisms of channel regulation, in the context of model systems and in the light of evolution.

Pharmacological enhancement of calcium-activated potassium channel function reduces the effects of repeated stress on fear memory

PubMed Central

Atchley, Derek; Hankosky, Emily R.; Gasparotto, Kaylyn; Rosenkranz, J. Amiel

2012-01-01

Repeated stress impacts emotion, and can induce mood and anxiety disorders. These disorders are characterized by imbalance of emotional responses. The amygdala is fundamental in expression of emotion, and is hyperactive in many patients with mood or anxiety disorders. Stress also leads to hyperactivity of the amygdala in humans. In rodent studies, repeated stress causes hyperactivity of the amygdala, and increases fear conditioning behavior that is mediated by the basolateral amygdala (BLA). Calcium-activated potassium (KCa) channels regulate BLA neuronal activity, and evidence suggests reduced small conductance KCa (SK) channel function in male rats exposed to repeated stress. Pharmacological enhancement of SK channels reverses the BLA neuronal hyperexcitability caused by repeated stress. However, it is not known if pharmacological targeting of SK channels can repair the effects of repeated stress on amygdala-dependent behaviors. The purpose of this study was to test whether enhancement of SK channel function reverses the effects of repeated restraint on BLA-dependent auditory fear conditioning. We found that repeated restraint stress increased the expression of cued conditioned fear in male rats. However, 1-EBIO (1 or 10 mg/kg) or CyPPA (5 mg/kg) administered 30 minutes prior to testing of fear expression brought conditioned freezing to control levels, with little impact on fear expression in control handled rats. These results demonstrate that enhancement of SK channel function can reduce the abnormalities of BLA-dependent fear memory caused by repeated stress. Furthermore, this indicates that pharmacological targeting of SK channels may provide a novel target for alleviation of psychiatric symptoms associated with amygdala hyperactivity. PMID:22487247

S-acylation dependent post-translational cross-talk regulates large conductance calcium- and voltage- activated potassium (BK) channels

PubMed Central

Shipston, Michael J.

2014-01-01

Mechanisms that control surface expression and/or activity of large conductance calcium-activated potassium (BK) channels are important determinants of their (patho)physiological function. Indeed, BK channel dysfunction is associated with major human disorders ranging from epilepsy to hypertension and obesity. S-acylation (S-palmitoylation) represents a major reversible, post-translational modification controlling the properties and function of many proteins including ion channels. Recent evidence reveals that both pore-forming and regulatory subunits of BK channels are S-acylated and control channel trafficking and regulation by AGC-family protein kinases. The pore-forming α-subunit is S-acylated at two distinct sites within the N- and C-terminus, each site being regulated by different palmitoyl acyl transferases (zDHHCs) and acyl thioesterases (APTs). S-acylation of the N-terminus controls channel trafficking and surface expression whereas S-acylation of the C-terminal domain determines regulation of channel activity by AGC-family protein kinases. S-acylation of the regulatory β4-subunit controls ER exit and surface expression of BK channels but does not affect ion channel kinetics at the plasma membrane. Furthermore, a significant number of previously identified BK-channel interacting proteins have been shown, or are predicted to be, S-acylated. Thus, the BK channel multi-molecular signaling complex may be dynamically regulated by this fundamental post-translational modification and thus S-acylation likely represents an important determinant of BK channel physiology in health and disease. PMID:25140154

Quasi-specific access of the potassium channel inactivation gate

PubMed Central

Venkataraman, Gaurav; Srikumar, Deepa; Holmgren, Miguel

2014-01-01

Many voltage-gated potassium channels open in response to membrane depolarization and then inactivate within milliseconds. Neurons use these channels to tune their excitability. In Shaker K+ channels, inactivation is caused by the cytoplasmic amino terminus, termed the inactivation gate. Despite having four such gates, inactivation is caused by the movement of a single gate into a position that occludes ion permeation. The pathway that this single inactivation gate takes into its inactivating position remains unknown. Here we show that a single gate threads through the intracellular entryway of its own subunit, but the tip of the gate has sufficient freedom to interact with all four subunits deep in the pore, and does so with equal probability. This pathway demonstrates that flexibility afforded by the inactivation peptide segment at the tip of the N-terminus is used to mediate function. PMID:24909510

Terbinafine is a novel and selective activator of the two-pore domain potassium channel TASK3.

PubMed

Wright, Paul D; Veale, Emma L; McCoull, David; Tickle, David C; Large, Jonathan M; Ococks, Emma; Gothard, Gemma; Kettleborough, Catherine; Mathie, Alistair; Jerman, Jeffrey

2017-11-04

Two-pore domain potassium channels (K2Ps) are characterized by their four transmembrane domain and two-pore topology. They carry background (or leak) potassium current in a variety of cell types. Despite a number of important roles there is currently a lack of pharmacological tools with which to further probe K2P function. We have developed a cell-based thallium flux assay, using baculovirus delivered TASK3 (TWIK-related acid-sensitive K + channel 3, KCNK9, K2P9.1) with the aim of identifying novel, selective TASK3 activators. After screening a library of 1000 compounds, including drug-like and FDA approved molecules, we identified Terbinafine as an activator of TASK3. In a thallium flux assay a pEC50 of 6.2 ( ±0.12) was observed. When Terbinafine was screened against TASK2, TREK2, THIK1, TWIK1 and TRESK no activation was observed in thallium flux assays. Several analogues of Terbinafine were also purchased and structure activity relationships examined. To confirm Terbinafine's activation of TASK3 whole cell patch clamp electrophysiology was carried out and clear potentiation observed in both the wild type channel and the pathophysiological, Birk-Barel syndrome associated, G236R TASK3 mutant. No activity at TASK1 was observed in electrophysiology studies. In conclusion, we have identified the first selective activator of the two-pore domain potassium channel TASK3. Copyright © 2017 Elsevier Inc. All rights reserved.

Functional coupling between sodium-activated potassium channels and voltage-dependent persistent sodium currents in cricket Kenyon cells.

PubMed

Takahashi, Izumi; Yoshino, Masami

2015-10-01

In this study, we examined the functional coupling between Na(+)-activated potassium (KNa) channels and Na(+) influx through voltage-dependent Na(+) channels in Kenyon cells isolated from the mushroom body of the cricket Gryllus bimaculatus. Single-channel activity of KNa channels was recorded with the cell-attached patch configuration. The open probability (Po) of KNa channels increased with increasing Na(+) concentration in a bath solution, whereas it decreased by the substitution of Na(+) with an equimolar concentration of Li(+). The Po of KNa channels was also found to be reduced by bath application of a high concentration of TTX (1 μM) and riluzole (100 μM), which inhibits both fast (INaf) and persistent (INaP) Na(+) currents, whereas it was unaffected by a low concentration of TTX (10 nM), which selectively blocks INaf. Bath application of Cd(2+) at a low concentration (50 μM), as an inhibitor of INaP, also decreased the Po of KNa channels. Conversely, bath application of the inorganic Ca(2+)-channel blockers Co(2+) and Ni(2+) at high concentrations (500 μM) had little effect on the Po of KNa channels, although Cd(2+) (500 μM) reduced the Po of KNa channels. Perforated whole cell clamp analysis further indicated the presence of sustained outward currents for which amplitude was dependent on the amount of Na(+) influx. Taken together, these results indicate that KNa channels could be activated by Na(+) influx passing through voltage-dependent persistent Na(+) channels. The functional significance of this coupling mechanism was discussed in relation to the membrane excitability of Kenyon cells and its possible role in the formation of long-term memory. Copyright © 2015 the American Physiological Society.

A human intermediate conductance calcium-activated potassium channel.

PubMed

Ishii, T M; Silvia, C; Hirschberg, B; Bond, C T; Adelman, J P; Maylie, J

1997-10-14

An intermediate conductance calcium-activated potassium channel, hIK1, was cloned from human pancreas. The predicted amino acid sequence is related to, but distinct from, the small conductance calcium-activated potassium channel subfamily, which is approximately 50% conserved. hIK1 mRNA was detected in peripheral tissues but not in brain. Expression of hIK1 in Xenopus oocytes gave rise to inwardly rectifying potassium currents, which were activated by submicromolar concentrations of intracellular calcium (K0.5 = 0.3 microM). Although the K0.5 for calcium was similar to that of small conductance calcium-activated potassium channels, the slope factor derived from the Hill equation was significantly reduced (1.7 vs. 3. 5). Single-channel current amplitudes reflected the macroscopic inward rectification and revealed a conductance level of 39 pS in the inward direction. hIK1 currents were reversibly blocked by charybdotoxin (Ki = 2.5 nM) and clotrimazole (Ki = 24.8 nM) but were minimally affected by apamin (100 nM), iberiotoxin (50 nM), or ketoconazole (10 microM). These biophysical and pharmacological properties are consistent with native intermediate conductance calcium-activated potassium channels, including the erythrocyte Gardos channel.

Identification of the functional binding pocket for compounds targeting small-conductance Ca2+-activated potassium channels

PubMed Central

Zhang, Miao; Pascal, John M.; Schumann, Marcel; Armen, Roger S.; Zhang, Ji-fang

2012-01-01

Small- and intermediate-conductance Ca2+-activated potassium channels, activated by Ca2+-bound calmodulin, play an important role in regulating membrane excitability. These channels are also linked to clinical abnormalities. A tremendous amount of effort has been devoted to developing small molecule compounds targeting these channels. However, these compounds often suffer from low potency and lack of selectivity, hindering their potentials for clinical use. A key contributing factor is the lack of knowledge of the binding site(s) for these compounds. Here we demonstrate by X-ray crystallography that the binding pocket for the compounds of the 1-EBIO class is located at the calmodulin-channel interface. We show that, based on structure data and molecular docking, mutations of the channel can effectively change the potency of these compounds. Our results provide insight into the molecular nature of the binding pocket and its contribution to the potency and selectivity of the compounds of the 1-EBIO class. PMID:22929778

Identification of the functional binding pocket for compounds targeting small-conductance Ca²⁺-activated potassium channels.

PubMed

Zhang, Miao; Pascal, John M; Schumann, Marcel; Armen, Roger S; Zhang, Ji-Fang

2012-01-01

Small- and intermediate-conductance Ca(2+)-activated potassium channels, activated by Ca(2+)-bound calmodulin, have an important role in regulating membrane excitability. These channels are also linked to clinical abnormalities. A tremendous amount of effort has been devoted to developing small molecule compounds targeting these channels. However, these compounds often suffer from low potency and lack of selectivity, hindering their potential for clinical use. A key contributing factor is the lack of knowledge of the binding site(s) for these compounds. Here we demonstrate by X-ray crystallography that the binding pocket for the compounds of the 1-ethyl-2-benzimidazolinone (1-EBIO) class is located at the calmodulin-channel interface. We show that, based on structure data and molecular docking, mutations of the channel can effectively change the potency of these compounds. Our results provide insight into the molecular nature of the binding pocket and its contribution to the potency and selectivity of the compounds of the 1-EBIO class.

Planar patch clamp: advances in electrophysiology.

PubMed

Brüggemann, Andrea; Farre, Cecilia; Haarmann, Claudia; Haythornthwaite, Ali; Kreir, Mohamed; Stoelzle, Sonja; George, Michael; Fertig, Niels

2008-01-01

Ion channels have gained increased interest as therapeutic targets over recent years, since a growing number of human and animal diseases have been attributed to defects in ion channel function. Potassium channels are the largest and most diverse family of ion channels. Pharmaceutical agents such as Glibenclamide, an inhibitor of K(ATP) channel activity which promotes insulin release, have been successfully sold on the market for many years. So far, only a small group of the known ion channels have been addressed as potential drug targets. The functional testing of drugs on these ion channels has always been the bottleneck in the development of these types of pharmaceutical compounds.New generations of automated patch clamp screening platforms allow a higher throughput for drug testing and widen this bottleneck. Due to their planar chip design not only is a higher throughput achieved, but new applications have also become possible. One of the advantages of planar patch clamp is the possibility of perfusing the intracellular side of the membrane during a patch clamp experiment in the whole-cell configuration. Furthermore, the extracellular membrane remains accessible for compound application during the experiment.Internal perfusion can be used not only for patch clamp experiments with cell membranes, but also for those with artificial lipid bilayers. In this chapter we describe how internal perfusion can be applied to potassium channels expressed in Jurkat cells, and to Gramicidin channels reconstituted in a lipid bilayer.

Crystal Structure of the Potassium Importing KdpFABC Membrane Complex

PubMed Central

Huang, Ching-Shin; Pedersen, Bjørn Panyella; Stokes, David Lloyd

2017-01-01

Cellular potassium import systems play a fundamental role in osmoregulation, pH homeostasis and membrane potential in all domains of life. In bacteria, the kdp operon encodes a four subunit potassium pump that maintains intracellular homeostasis as well as cell shape and turgor under conditions where potassium is limiting1. This membrane complex, called KdpFABC, has one channel-like subunit (KdpA) belonging to the Superfamily of Potassium Transporters and another pump-like subunit (KdpB) belonging to the Superfamily of P-type ATPases. Although there is considerable structural and functional information about members from both superfamilies, the mechanism by which uphill potassium transport through KdpA is coupled with ATP hydrolysis by KdpB remains poorly understood. Here we report the 2.9 Å X-ray structure of the complete Escherichia coli KdpFABC complex with a potassium ion within the selectivity filter of KdpA as well as a water molecule at a canonical cation site in the transmembrane domain of KdpB. The structure also reveals two structural elements that appear to mediate the coupling between these two subunits. Specifically, a protein-embedded tunnel runs between these potassium and water sites and a helix controlling the cytoplasmic gate of KdpA is linked to the phosphorylation domain of KdpB. Based on these observations, we propose an unprecedented mechanism that repurposes protein channel architecture for active transport across biomembranes. PMID:28636601

Kv7 potassium channel subunits and M currents in cultured hippocampal interneurons.

PubMed

Grigorov, Alexej; Moskalyuk, Anastasia; Kravchenko, Mykola; Veselovsky, Nikolai; Verkhratsky, Alexei; Fedulova, Svetlana

2014-09-01

Potassium channels of the Kv7 family that mediate the non-inactivating M current regulate the excitability of many types of neurons in the central nervous system, including some in the hippocampus. We report here that individual interneurons from newborn rat hippocampi in long-term culture strongly express messenger RNA specific for Kv7.2 and Kv7.3 and, to a lesser extent, Kv7.5 channel subunits but not for the Kv7.4 subunit. An M-like current was electrophysiologically identified in two subpopulations of interneurons distinct in their spiking behaviour (regular or fast spiking). The M-channel enhancer retigabine reduced interneuronal excitability by constraining the number of action potentials generated during imposed depolarisations; this effect was inhibited by specific the M-channel blocking drugs. In paired synaptically connected interneuron-target cell recordings, anatomically localised applications of retigabine indicated that M channels were present in both the interneuron soma and its GABA-ergic inhibitory axon. We conclude that M-channel subunits and functional M channels are broadly expressed in hippocampal interneurons and their axons and are potentially capable of strongly regulating their firing properties.

[Water regulation in the cochlea : Do molecular water channels facilitate potassium-dependent sound transduction?].

PubMed

Eckhard, A; Löwenheim, H

2014-06-01

Sound transduction in the cochlea critically depends on the circulation of potassium ions (K(+)) along so-called "K(+) recycling routes" between the endolymph and perilymph. These K(+) currents generate high ionic and osmotic gradients, which potentially impair the excitability of sensory hair cells and threaten cell survival in the entire cochlear duct. Molecular water channels-aquaporins (AQP)-are expressed in all cochlear supporting cells along the K(+) recycling routes; however, their significance for osmotic equilibration in cochlear duct cells is unknown. The diffusive and osmotic water permeabilies of Reissner's membrane, the organ of Corti and the entire cochlear duct epithelium were determined. Expression of the potassium channel Kir4.1 and the water channel AQP4 in the cochlear duct was investigated by immunohistochemistry. The calculated water permeability values indicate the extent of AQP-facilitated water flux across the cochlear duct epithelium. Immunohistochemically, Kir4.1 and AQP4 were found to colocalize in distinct membrane domains of supporting cells along the K(+)-recycling routes. These observations suggest the presence of a rapid AQP-mediated water exchange between the endolymph, the cells of the cochlear duct and the perilymph. The subcellular colocalization of Kir4.1 and AQP4 in epithelial supporting cells indicates functional coupling of potassium and water flow in the cochlea. Finally, this offers an explanation for the hearing impairment observed in individuals with mutations in the AQP4 gene.

Effect of tyrphostin AG879 on Kv4.2 and Kv4.3 potassium channels

PubMed Central

Yu, Haibo; Zou, Beiyan; Wang, Xiaoliang; Li, Min

2015-01-01

Background and Purpose A-type potassium channels (IA) are important proteins for modulating neuronal membrane excitability. The expression and activity of Kv4.2 channels are critical for neurological functions and pharmacological inhibitors of Kv4.2 channels may have therapeutic potential for Fragile X syndrome. While screening various compounds, we identified tyrphostin AG879, a tyrosine kinase inhibitor, as a Kv4.2 inhibitor from. In the present study we characterized the effect of AG879 on cloned Kv4.2/Kv channel-interacting protein 2 (KChIP2) channels. Experimental Approach To screen the library of pharmacologically active compounds, the thallium flux assay was performed on HEK-293 cells transiently-transfected with Kv4.2 cDNA using the Maxcyte transfection system. The effects of AG879 were further examined on CHO-K1 cells expressing Kv4.2/KChIP2 channels using a whole-cell patch-clamp technique. Key Results Tyrphostin AG879 selectively and dose-dependently inhibited Kv4.2 and Kv4.3 channels. In Kv4.2/KChIP2 channels, AG879 induced prominent acceleration of the inactivation rate, use-dependent block and slowed the recovery from inactivation. AG879 induced a hyperpolarizing shift in the voltage-dependence of the steady-state inactivation of Kv4.2 channels without apparent effect on the V1/2 of the voltage-dependent activation. The blocking effect of AG879 was enhanced as channel inactivation increased. Furthermore, AG879 significantly inhibited the A-type potassium currents in the cultured hippocampus neurons. Conclusion and Implications AG879 was identified as a selective and potent inhibitor the Kv4.2 channel. AG879 inhibited Kv4.2 channels by preferentially interacting with the open state and further accelerating their inactivation. PMID:25752739

Effect of tyrphostin AG879 on Kv 4.2 and Kv 4.3 potassium channels.

PubMed

Yu, Haibo; Zou, Beiyan; Wang, Xiaoliang; Li, Min

2015-07-01

A-type potassium channels (IA) are important proteins for modulating neuronal membrane excitability. The expression and activity of Kv 4.2 channels are critical for neurological functions and pharmacological inhibitors of Kv 4.2 channels may have therapeutic potential for Fragile X syndrome. While screening various compounds, we identified tyrphostin AG879, a tyrosine kinase inhibitor, as a Kv 4.2 inhibitor from. In the present study we characterized the effect of AG879 on cloned Kv 4.2/Kv channel-interacting protein 2 (KChIP2) channels. To screen the library of pharmacologically active compounds, the thallium flux assay was performed on HEK-293 cells transiently-transfected with Kv 4.2 cDNA using the Maxcyte transfection system. The effects of AG879 were further examined on CHO-K1 cells expressing Kv 4.2/KChIP2 channels using a whole-cell patch-clamp technique. Tyrphostin AG879 selectively and dose-dependently inhibited Kv 4.2 and Kv 4.3 channels. In Kv 4.2/KChIP2 channels, AG879 induced prominent acceleration of the inactivation rate, use-dependent block and slowed the recovery from inactivation. AG879 induced a hyperpolarizing shift in the voltage-dependence of the steady-state inactivation of Kv 4.2 channels without apparent effect on the V1/2 of the voltage-dependent activation. The blocking effect of AG879 was enhanced as channel inactivation increased. Furthermore, AG879 significantly inhibited the A-type potassium currents in the cultured hippocampus neurons. AG879 was identified as a selective and potent inhibitor the Kv 4.2 channel. AG879 inhibited Kv 4.2 channels by preferentially interacting with the open state and further accelerating their inactivation. © 2015 The British Pharmacological Society.

Sodium and potassium competition in potassium-selective and non-selective channels

NASA Astrophysics Data System (ADS)

Sauer, David B.; Zeng, Weizhong; Canty, John; Lam, Yeeling; Jiang, Youxing

2013-11-01

Potassium channels selectively conduct K+, primarily to the exclusion of Na+, despite the fact that both ions can bind within the selectivity filter. Here we perform crystallographic titration and single-channel electrophysiology to examine the competition of Na+ and K+ binding within the filter of two NaK channel mutants; one is the potassium-selective NaK2K mutant and the other is the non-selective NaK2CNG, a CNG channel pore mimic. With high-resolution structures of these engineered NaK channel constructs, we explicitly describe the changes in K+ occupancy within the filter upon Na+ competition by anomalous diffraction. Our results demonstrate that the non-selective NaK2CNG still retains a K+-selective site at equilibrium, whereas the NaK2K channel filter maintains two high-affinity K+ sites. A double-barrier mechanism is proposed to explain K+ channel selectivity at low K+ concentrations.

Complementary functions of SK and Kv7/M potassium channels in excitability control and synaptic integration in rat hippocampal dentate granule cells

PubMed Central

Mateos-Aparicio, Pedro; Murphy, Ricardo; Storm, Johan F

2014-01-01

The dentate granule cells (DGCs) form the most numerous neuron population of the hippocampal memory system, and its gateway for cortical input. Yet, we have only limited knowledge of the intrinsic membrane properties that shape their responses. Since SK and Kv7/M potassium channels are key mechanisms of neuronal spiking and excitability control, afterhyperpolarizations (AHPs) and synaptic integration, we studied their functions in DGCs. The specific SK channel blockers apamin or scyllatoxin increased spike frequency (excitability), reduced early spike frequency adaptation, fully blocked the medium-duration AHP (mAHP) after a single spike or spike train, and increased postsynaptic EPSP summation after spiking, but had no effect on input resistance (Rinput) or spike threshold. In contrast, blockade of Kv7/M channels by XE991 increased Rinput, lowered the spike threshold, and increased excitability, postsynaptic EPSP summation, and EPSP–spike coupling, but only slightly reduced mAHP after spike trains (and not after single spikes). The SK and Kv7/M channel openers 1-EBIO and retigabine, respectively, had effects opposite to the blockers. Computational modelling reproduced many of these effects. We conclude that SK and Kv7/M channels have complementary roles in DGCs. These mechanisms may be important for the dentate network function, as CA3 neurons can be activated or inhibition recruited depending on DGC firing rate. PMID:24366266

Phosphatidylinositol-4,5-bisphosphate is required for KCNQ1/KCNE1 channel function but not anterograde trafficking.

PubMed

Royal, Alice A; Tinker, Andrew; Harmer, Stephen C

2017-01-01

The slow delayed-rectifier potassium current (IKs) is crucial for human cardiac action potential repolarization. The formation of IKs requires co-assembly of the KCNQ1 α-subunit and KCNE1 β-subunit, and mutations in either of these subunits can lead to hereditary long QT syndrome types 1 and 5, respectively. It is widely recognised that the KCNQ1/KCNE1 (Q1/E1) channel requires phosphatidylinositol-4,5-bisphosphate (PIP2) binding for function. We previously identified a cluster of basic residues in the proximal C-terminus of KCNQ1 that form a PIP2/phosphoinositide binding site. Upon charge neutralisation of these residues we found that the channel became more retained in the endoplasmic reticulum, which raised the possibility that channel-phosphoinositide interactions could play a role in channel trafficking. To explore this further we used a chemically induced dimerization (CID) system to selectively deplete PIP2 and/or phosphatidylinositol-4-phosphate (PI(4)P) at the plasma membrane (PM) or Golgi, and we subsequently monitored the effects on both channel trafficking and function. The depletion of PIP2 and/or PI(4)P at either the PM or Golgi did not alter channel cell-surface expression levels. However, channel function was extremely sensitive to the depletion of PIP2 at the PM, which is in contrast to the response of other cardiac potassium channels tested (Kir2.1 and Kv11.1). Surprisingly, when using the CID system IKs was dramatically reduced even before dimerization was induced, highlighting limitations regarding the utility of this system when studying processes highly sensitive to PIP2 depletion. In conclusion, we identify that the Q1/E1 channel does not require PIP2 or PI(4)P for anterograde trafficking, but is heavily reliant on PIP2 for channel function once at the PM.

Sodium channel selectivity and conduction: Prokaryotes have devised their own molecular strategy

PubMed Central

Finol-Urdaneta, Rocio K.; Wang, Yibo; Al-Sabi, Ahmed; Zhao, Chunfeng

2014-01-01

Striking structural differences between voltage-gated sodium (Nav) channels from prokaryotes (homotetramers) and eukaryotes (asymmetric, four-domain proteins) suggest the likelihood of different molecular mechanisms for common functions. For these two channel families, our data show similar selectivity sequences among alkali cations (relative permeability, Pion/PNa) and asymmetric, bi-ionic reversal potentials when the Na/K gradient is reversed. We performed coordinated experimental and computational studies, respectively, on the prokaryotic Nav channels NaChBac and NavAb. NaChBac shows an “anomalous,” nonmonotonic mole-fraction dependence in the presence of certain sodium–potassium mixtures; to our knowledge, no comparable observation has been reported for eukaryotic Nav channels. NaChBac’s preferential selectivity for sodium is reduced either by partial titration of its highly charged selectivity filter, when extracellular pH is lowered from 7.4 to 5.8, or by perturbation—likely steric—associated with a nominally electro-neutral substitution in the selectivity filter (E191D). Although no single molecular feature or energetic parameter appears to dominate, our atomistic simulations, based on the published NavAb crystal structure, revealed factors that may contribute to the normally observed selectivity for Na over K. These include: (a) a thermodynamic penalty to exchange one K+ for one Na+ in the wild-type (WT) channel, increasing the relative likelihood of Na+ occupying the binding site; (b) a small tendency toward weaker ion binding to the selectivity filter in Na–K mixtures, consistent with the higher conductance observed with both sodium and potassium present; and (c) integrated 1-D potentials of mean force for sodium or potassium movement that show less separation for the less selective E/D mutant than for WT. Overall, tight binding of a single favored ion to the selectivity filter, together with crucial inter-ion interactions within the pore, suggests that prokaryotic Nav channels use a selective strategy more akin to those of eukaryotic calcium and potassium channels than that of eukaryotic Nav channels. PMID:24420772

Overexpression of calcium-activated potassium channels underlies cortical dysfunction in a model of PTEN-associated autism.

PubMed

Garcia-Junco-Clemente, Pablo; Chow, David K; Tring, Elaine; Lazaro, Maria T; Trachtenberg, Joshua T; Golshani, Peyman

2013-11-05

De novo phosphatase and tensin homolog on chromosome ten (PTEN) mutations are a cause of sporadic autism. How single-copy loss of PTEN alters neural function is not understood. Here we report that Pten haploinsufficiency increases the expression of small-conductance calcium-activated potassium channels. The resultant augmentation of this conductance increases the amplitude of the afterspike hyperpolarization, causing a decrease in intrinsic excitability. In vivo, this change in intrinsic excitability reduces evoked firing rates of cortical pyramidal neurons but does not alter receptive field tuning. The decreased in vivo firing rate is not associated with deficits in the dendritic integration of synaptic input or with changes in dendritic complexity. These findings identify calcium-activated potassium channelopathy as a cause of cortical dysfunction in the PTEN model of autism and provide potential molecular therapeutic targets.

Overcoming HERG affinity in the discovery of the CCR5 antagonist maraviroc.

PubMed

Price, David A; Armour, Duncan; de Groot, Marcel; Leishman, Derek; Napier, Carolyn; Perros, Manos; Stammen, Blanda L; Wood, Anthony

2006-09-01

The discovery of maraviroc 17 is described with particular reference to the generation of high selectivity over affinity for the HERG potassium channel. This was achieved through the use of a high throughput binding assay for the HERG channel that is known to show an excellent correlation with functional effects.

Reversible dementia: two nursing home patients with voltage-gated potassium channel antibody-associated limbic encephalitis.

PubMed

Reintjes, Wesley; Romijn, Marloes D M; Hollander, Daan; Ter Bruggen, Jan P; van Marum, Rob J

2015-09-01

Voltage-gated potassium channel antibody-associated limbic encephalitis (VGKC-LE) is a rare disease that is a diagnostic and therapeutic challenge for medical practitioners. Two patients with VGKC-LE, both developing dementia are presented. Following treatment, both patients showed remarkable cognitive and functional improvement enabling them to leave the psychogeriatric nursing homes they both were admitted to. Patients with VGKC-LE can have a major cognitive and functional improvement even after a diagnostic delay of more than 1 year. Medical practitioners who treat patients with unexplained cognitive decline, epileptic seizures, or psychiatric symptoms should be aware of LE as an underlying rare cause. Copyright © 2015 AMDA – The Society for Post-Acute and Long-Term Care Medicine. Published by Elsevier Inc. All rights reserved.

Chronic Manganese Toxicity Associated with Voltage-Gated Potassium Channel Complex Antibodies in a Relapsing Neuropsychiatric Disorder

PubMed Central

Ho, Cyrus S.H.; Quek, Amy M.L.

2018-01-01

Heavy metal poisoning is a rare but important cause of encephalopathy. Manganese (Mn) toxicity is especially rare in the modern world, and clinicians’ lack of recognition of its neuropsychiatric manifestations can lead to misdiagnosis and mismanagement. We describe the case of a man who presented with recurrent episodes of confusion, psychosis, dystonic limb movement and cognitive impairment and was initially diagnosed with anti-voltage-gated potassium channel (VGKC) complex limbic encephalitis in view of previous positive autoantibodies. His failure to respond to immunotherapy prompted testing for heavy metal poisoning, which was positive for Mn. This is the first report to examine an association between Mn and VGKC antibodies and the effects of Mn on functional brain activity using functional near-infrared spectroscopy (fNIRS). PMID:29669989

Chronic Manganese Toxicity Associated with Voltage-Gated Potassium Channel Complex Antibodies in a Relapsing Neuropsychiatric Disorder.

PubMed

Ho, Cyrus S H; Ho, Roger C M; Quek, Amy M L

2018-04-18

Heavy metal poisoning is a rare but important cause of encephalopathy. Manganese (Mn) toxicity is especially rare in the modern world, and clinicians’ lack of recognition of its neuropsychiatric manifestations can lead to misdiagnosis and mismanagement. We describe the case of a man who presented with recurrent episodes of confusion, psychosis, dystonic limb movement and cognitive impairment and was initially diagnosed with anti-voltage-gated potassium channel (VGKC) complex limbic encephalitis in view of previous positive autoantibodies. His failure to respond to immunotherapy prompted testing for heavy metal poisoning, which was positive for Mn. This is the first report to examine an association between Mn and VGKC antibodies and the effects of Mn on functional brain activity using functional near-infrared spectroscopy (fNIRS).

Automated Patch-Clamp Methods for the hERG Cardiac Potassium Channel.

PubMed

Houtmann, Sylvie; Schombert, Brigitte; Sanson, Camille; Partiseti, Michel; Bohme, G Andrees

2017-01-01

The human Ether-a-go-go Related Gene (hERG) product has been identified as a central ion channel underlying both familial forms of elongated QT interval on the electrocardiogram and drug-induced elongation of the same QT segment. Indeed, reduced function of this potassium channel involved in the repolarization of the cardiac action potential can produce a type of life-threatening cardiac ventricular arrhythmias called Torsades de Pointes (TdP). Therefore, hERG inhibitory activity of newly synthetized molecules is a relevant structure-activity metric for compound prioritization and optimization in medicinal chemistry phases of drug discovery. Electrophysiology remains the gold standard for the functional assessment of ion channel pharmacology. The recent years have witnessed automatization and parallelization of the manual patch-clamp technique, allowing higher throughput screening on recombinant hERG channels. However, the multi-well plate format of automatized patch-clamp does not allow visual detection of potential micro-precipitation of poorly soluble compounds. In this chapter we describe bench procedures for the culture and preparation of hERG-expressing CHO cells for recording on an automated patch-clamp workstation. We also show that the sensitivity of the assay can be improved by adding a surfactant to the extracellular medium.

Fragile X mental retardation protein controls ion channel expression and activity.

PubMed

Ferron, Laurent

2016-10-15

Fragile X-associated disorders are a family of genetic conditions resulting from the partial or complete loss of fragile X mental retardation protein (FMRP). Among these disorders is fragile X syndrome, the most common cause of inherited intellectual disability and autism. FMRP is an RNA-binding protein involved in the control of local translation, which has pleiotropic effects, in particular on synaptic function. Analysis of the brain FMRP transcriptome has revealed hundreds of potential mRNA targets encoding postsynaptic and presynaptic proteins, including a number of ion channels. FMRP has been confirmed to bind voltage-gated potassium channels (K v 3.1 and K v 4.2) mRNAs and regulates their expression in somatodendritic compartments of neurons. Recent studies have uncovered a number of additional roles for FMRP besides RNA regulation. FMRP was shown to directly interact with, and modulate, a number of ion channel complexes. The sodium-activated potassium (Slack) channel was the first ion channel shown to directly interact with FMRP; this interaction alters the single-channel properties of the Slack channel. FMRP was also shown to interact with the auxiliary β4 subunit of the calcium-activated potassium (BK) channel; this interaction increases calcium-dependent activation of the BK channel. More recently, FMRP was shown to directly interact with the voltage-gated calcium channel, Ca v 2.2, and reduce its trafficking to the plasma membrane. Studies performed on animal models of fragile X syndrome have revealed links between modifications of ion channel activity and changes in neuronal excitability, suggesting that these modifications could contribute to the phenotypes observed in patients with fragile X-associated disorders. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

A novel crystallization method for visualizing the membrane localization of potassium channels.

PubMed Central

Lopatin, A N; Makhina, E N; Nichols, C G

1998-01-01

The high permeability of K+ channels to monovalent thallium (Tl+) ions and the low solubility of thallium bromide salt were used to develop a simple yet very sensitive approach to the study of membrane localization of potassium channels. K+ channels (Kir1.1, Kir2.1, Kir2.3, Kv2.1), were expressed in Xenopus oocytes and loaded with Br ions by microinjection. Oocytes were then exposed to extracellular thallium. Under conditions favoring influx of Tl+ ions (negative membrane potential under voltage clamp, or high concentration of extracellular Tl+), crystals of TlBr, visible under low-power microscopy, formed under the membrane in places of high density of K+ channels. Crystals were not formed in uninjected oocytes, but were formed in oocytes expressing as little as 5 microS K+ conductance. The number of observed crystals was much lower than the estimated number of functional channels. Based on the pattern of crystal formation, K+ channels appear to be expressed mostly around the point of cRNA injection when injected either into the animal or vegetal hemisphere. In addition to this pseudopolarized distribution of K+ channels due to localized microinjection of cRNA, a naturally polarized (animal/vegetal side) distribution of K+ channels was also frequently observed when K+ channel cRNA was injected at the equator. A second novel "agarose-hemiclamp" technique was developed to permit direct measurements of K+ currents from different hemispheres of oocytes under two-microelectrode voltage clamp. This technique, together with direct patch-clamping of patches of membrane in regions of high crystal density, confirmed that the localization of TlBr crystals corresponded to the localization of functional K+ channels and suggested a clustered organization of functional channels. With appropriate permeant ion/counterion pairs, this approach may be applicable to the visualization of the membrane distribution of any functional ion channel. PMID:9591643

A novel crystallization method for visualizing the membrane localization of potassium channels.

PubMed

Lopatin, A N; Makhina, E N; Nichols, C G

1998-05-01

The high permeability of K+ channels to monovalent thallium (Tl+) ions and the low solubility of thallium bromide salt were used to develop a simple yet very sensitive approach to the study of membrane localization of potassium channels. K+ channels (Kir1.1, Kir2.1, Kir2.3, Kv2.1), were expressed in Xenopus oocytes and loaded with Br ions by microinjection. Oocytes were then exposed to extracellular thallium. Under conditions favoring influx of Tl+ ions (negative membrane potential under voltage clamp, or high concentration of extracellular Tl+), crystals of TlBr, visible under low-power microscopy, formed under the membrane in places of high density of K+ channels. Crystals were not formed in uninjected oocytes, but were formed in oocytes expressing as little as 5 microS K+ conductance. The number of observed crystals was much lower than the estimated number of functional channels. Based on the pattern of crystal formation, K+ channels appear to be expressed mostly around the point of cRNA injection when injected either into the animal or vegetal hemisphere. In addition to this pseudopolarized distribution of K+ channels due to localized microinjection of cRNA, a naturally polarized (animal/vegetal side) distribution of K+ channels was also frequently observed when K+ channel cRNA was injected at the equator. A second novel "agarose-hemiclamp" technique was developed to permit direct measurements of K+ currents from different hemispheres of oocytes under two-microelectrode voltage clamp. This technique, together with direct patch-clamping of patches of membrane in regions of high crystal density, confirmed that the localization of TlBr crystals corresponded to the localization of functional K+ channels and suggested a clustered organization of functional channels. With appropriate permeant ion/counterion pairs, this approach may be applicable to the visualization of the membrane distribution of any functional ion channel.

Poplar potassium transporters capable of controlling K+ homeostasis and K+-dependent xylogenesis.

PubMed

Langer, Katharina; Ache, Peter; Geiger, Dietmar; Stinzing, Andrea; Arend, Matthias; Wind, Christa; Regan, Sharon; Fromm, Jörg; Hedrich, Rainer

2002-12-01

The cambial K+ content of poplar increases during the growth period in a K+ supply dependent manner. Upon K+ starvation or application of tetraethylammoniumchloride (TEA+), a K+ channel blocker, the average vessel lumen and expansion zone area were significantly reduced. In search for the molecular basis of potassium-dependent xylogenesis in poplar, K+ transporters homologous to those of known function in Arabidopis phloem- and xylem-physiology were isolated from a poplar wood EST library. The expression profile of three distinct K+ channel types and one K+ transporter, Populus tremula K+ uptake transporter 1 (PtKUP1), was analysed by quantitative RT-PCR. Thereby, we found P. tremula outward rectifying K+ channel (PTORK) and P. tremula K+ channel 2 (PTK2) correlated with the seasonal wood production. K+ transporter P. tremula 1 (KPT1) was predominantly found in guard cells. Following the heterologous expression in Xenopus oocytes the biophysical properties of the different channels were determined. PTORK, upon membrane de-polarization mediates potassium release. PTK2 is almost voltage independent, carrying inward K+ flux at hyperpolarized potential and K+ release upon de-polarization. PtKUP1 was expressed in a K+ uptake-deficient Escherichia coli strain, where this K+ transporter rescued K+-dependent growth. In order to link the different K+ transporters to the cambial activity and wood production, we compared the expression profiles to seasonal changes in the K+ content of the bark as well as xylem vessel diameter. Thereby, we found PTORK and PTK2 transcripts to follow the annual K+ variations in poplar branches. PtKUP1 was expressed at a low level throughout the year, suggesting a housekeeping function. From these data, we conclude that K+ channels are involved in the regulation of K+-dependent wood production.

International Union of Basic and Clinical Pharmacology. LXXVI. Current Progress in the Mammalian TRP Ion Channel Family

PubMed Central

Wu, Long-Jun; Sweet, Tara-Beth

2010-01-01

Transient receptor potential (TRP) channels are a large family of ion channel proteins, surpassed in number in mammals only by voltage-gated potassium channels. TRP channels are activated and regulated through strikingly diverse mechanisms, making them suitable candidates for cellular sensors. They respond to environmental stimuli such as temperature, pH, osmolarity, pheromones, taste, and plant compounds, and intracellular stimuli such as Ca2+ and phosphatidylinositol signal transduction pathways. However, it is still largely unknown how TRP channels are activated in vivo. Despite the uncertainties, emerging evidence using TRP channel knockout mice indicates that these channels have broad function in physiology. Here we review the recent progress on the physiology, pharmacology and pathophysiological function of mammalian TRP channels. PMID:20716668

Cholesterol up-regulates neuronal G protein-gated inwardly rectifying potassium (GIRK) channel activity in the hippocampus.

PubMed

Bukiya, Anna N; Durdagi, Serdar; Noskov, Sergei; Rosenhouse-Dantsker, Avia

2017-04-14

Hypercholesterolemia is a well known risk factor for the development of neurodegenerative disease. However, the underlying mechanisms are mostly unknown. In recent years, it has become increasingly evident that cholesterol-driven effects on physiology and pathophysiology derive from its ability to alter the function of a variety of membrane proteins including ion channels. Yet, the effect of cholesterol on G protein-gated inwardly rectifying potassium (GIRK) channels expressed in the brain is unknown. GIRK channels mediate the actions of inhibitory brain neurotransmitters. As a result, loss of GIRK function can enhance neuron excitability, whereas gain of GIRK function can reduce neuronal activity. Here we show that in rats on a high-cholesterol diet, cholesterol levels in hippocampal neurons are increased. We also demonstrate that cholesterol plays a critical role in modulating neuronal GIRK currents. Specifically, cholesterol enrichment of rat hippocampal neurons resulted in enhanced channel activity. In accordance, elevated currents upon cholesterol enrichment were also observed in Xenopus oocytes expressing GIRK2 channels, the primary GIRK subunit expressed in the brain. Furthermore, using planar lipid bilayers, we show that although cholesterol did not affect the unitary conductance of GIRK2, it significantly enhanced the frequency of channel openings. Last, combining computational and functional approaches, we identified two putative cholesterol-binding sites in the transmembrane domain of GIRK2. These findings establish that cholesterol plays a critical role in modulating GIRK activity in the brain. Because up-regulation of GIRK function can reduce neuronal activity, our findings may lead to novel approaches for prevention and therapy of cholesterol-driven neurodegenerative disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Functional conversion between A-type and delayed rectifier K+ channels by membrane lipids.

PubMed

Oliver, Dominik; Lien, Cheng-Chang; Soom, Malle; Baukrowitz, Thomas; Jonas, Peter; Fakler, Bernd

2004-04-09

Voltage-gated potassium (Kv) channels control action potential repolarization, interspike membrane potential, and action potential frequency in excitable cells. It is thought that the combinatorial association between distinct alpha and beta subunits determines whether Kv channels function as non-inactivating delayed rectifiers or as rapidly inactivating A-type channels. We show that membrane lipids can convert A-type channels into delayed rectifiers and vice versa. Phosphoinositides remove N-type inactivation from A-type channels by immobilizing the inactivation domains. Conversely, arachidonic acid and its amide anandamide endow delayed rectifiers with rapid voltage-dependent inactivation. The bidirectional control of Kv channel gating by lipids may provide a mechanism for the dynamic regulation of electrical signaling in the nervous system.

Potassium Inhibits Dietary Salt-Induced Transforming Growth Factor-β Production

PubMed Central

Ying, Wei-Zhong; Aaron, Kristal; Wang, Pei-Xuan; Sanders, Paul W.

2009-01-01

Human and animal studies demonstrate an untoward effect of excess dietary NaCl (salt) intake on cardiovascular function and life span. The endothelium in particular augments the production of transforming growth factor (TGF)-β, a fibrogenic growth factor, in response to excess dietary salt intake. This study explored the initiating mechanism that regulates salt-induced endothelial cell production of TGF-β. Male Sprague-Dawley rats were given diets containing different amounts of NaCl and potassium for 4 days. A bioassay for TGF-β demonstrated increased (35.2%) amounts of active TGF-β in the medium of aortic ring segments from rats on the high-salt diet compared with rats maintained on a 0.3% NaCl diet. Inhibition of the large-conductance, calcium-activated potassium channel inhibited dietary salt-induced vascular production of TGF-β but did not affect production of TGF-β by ring segments from rats on the low-salt diet. Immunohistochemical and Western analyses demonstrated the α subunit of the calcium-activated potassium channel in endothelial cells. Increasing medium [K+] inhibited production of dietary salt-induced vascular production levels of total and active TGF-β but did not alter TGF-β production by aortic rings from rats on the 0.3% NaCl diet. Increasing dietary potassium content decreased urinary active TGF-β in animals receiving the high-salt diet but did not change urinary active TGF-β in animals receiving the low-salt diet. The findings demonstrated an interesting interaction between the dietary intake of potassium and excess NaCl and further showed the fundamental role of the endothelial calcium-activated potassium channel in the vascular response to excess salt intake. PMID:19738156

Decreased inward rectifier potassium current IK1 in dystrophin-deficient ventricular cardiomyocytes.

PubMed

Rubi, Lena; Koenig, Xaver; Kubista, Helmut; Todt, Hannes; Hilber, Karlheinz

2017-03-04

Kir2.x channels in ventricular cardiomyocytes (most prominently Kir2.1) account for the inward rectifier potassium current I K1 , which controls the resting membrane potential and the final phase of action potential repolarization. Recently it was hypothesized that the dystrophin-associated protein complex (DAPC) is important in the regulation of Kir2.x channels. To test this hypothesis, we investigated potential I K1 abnormalities in dystrophin-deficient ventricular cardiomyocytes derived from the hearts of Duchenne muscular dystrophy mouse models. We found that I K1 was substantially diminished in dystrophin-deficient cardiomyocytes when compared to wild type myocytes. This finding represents the first functional evidence for a significant role of the DAPC in the regulation of Kir2.x channels.

Glucose recruits K(ATP) channels via non-insulin-containing dense-core granules.

PubMed

Yang, Shao-Nian; Wenna, Nancy Dekki; Yu, Jia; Yang, Guang; Qiu, Hua; Yu, Lina; Juntti-Berggren, Lisa; Köhler, Martin; Berggren, Per-Olof

2007-09-01

beta cells rely on adenosine triphosphate-sensitive potassium (K(ATP)) channels to initiate and end glucose-stimulated insulin secretion through changes in membrane potential. These channels may also act as a constituent of the exocytotic machinery to mediate insulin release independent of their electrical function. However, the molecular mechanisms whereby the beta cell plasma membrane maintains an appropriate number of K(ATP) channels are not known. We now show that glucose increases K(ATP) current amplitude by increasing the number of K(ATP) channels in the beta cell plasma membrane. The effect was blocked by inhibition of protein kinase A (PKA) as well as by depletion of extracellular or intracellular Ca(2+). Furthermore, glucose promoted recruitment of the potassium inward rectifier 6.2 to the plasma membrane, and intracellular K(ATP) channels localized in chromogranin-positive/insulin-negative dense-core granules. Our data suggest that glucose can recruit K(ATP) channels to the beta cell plasma membrane via non-insulin-containing dense-core granules in a Ca(2+)- and PKA-dependent manner.

Phosphatidylinositol-4,5-bisphosphate is required for KCNQ1/KCNE1 channel function but not anterograde trafficking

PubMed Central

Royal, Alice A.

2017-01-01

The slow delayed-rectifier potassium current (IKs) is crucial for human cardiac action potential repolarization. The formation of IKs requires co-assembly of the KCNQ1 α-subunit and KCNE1 β-subunit, and mutations in either of these subunits can lead to hereditary long QT syndrome types 1 and 5, respectively. It is widely recognised that the KCNQ1/KCNE1 (Q1/E1) channel requires phosphatidylinositol-4,5-bisphosphate (PIP2) binding for function. We previously identified a cluster of basic residues in the proximal C-terminus of KCNQ1 that form a PIP2/phosphoinositide binding site. Upon charge neutralisation of these residues we found that the channel became more retained in the endoplasmic reticulum, which raised the possibility that channel–phosphoinositide interactions could play a role in channel trafficking. To explore this further we used a chemically induced dimerization (CID) system to selectively deplete PIP2 and/or phosphatidylinositol-4-phosphate (PI(4)P) at the plasma membrane (PM) or Golgi, and we subsequently monitored the effects on both channel trafficking and function. The depletion of PIP2 and/or PI(4)P at either the PM or Golgi did not alter channel cell-surface expression levels. However, channel function was extremely sensitive to the depletion of PIP2 at the PM, which is in contrast to the response of other cardiac potassium channels tested (Kir2.1 and Kv11.1). Surprisingly, when using the CID system IKs was dramatically reduced even before dimerization was induced, highlighting limitations regarding the utility of this system when studying processes highly sensitive to PIP2 depletion. In conclusion, we identify that the Q1/E1 channel does not require PIP2 or PI(4)P for anterograde trafficking, but is heavily reliant on PIP2 for channel function once at the PM. PMID:29020060

Neuronal Atrophy Early in Degenerative Ataxia Is a Compensatory Mechanism to Regulate Membrane Excitability

PubMed Central

Dell'Orco, James M.; Wasserman, Aaron H.; Chopra, Ravi; Ingram, Melissa A. C.; Hu, Yuan-Shih; Singh, Vikrant; Wulff, Heike; Opal, Puneet; Orr, Harry T.

2015-01-01

Neuronal atrophy in neurodegenerative diseases is commonly viewed as an early event in a continuum that ultimately results in neuronal loss. In a mouse model of the polyglutamine disorder spinocerebellar ataxia type 1 (SCA1), we tested the hypothesis that cerebellar Purkinje neuron atrophy serves an adaptive role rather than being simply a nonspecific response to injury. In acute cerebellar slices from SCA1 mice, we find that Purkinje neuron pacemaker firing is initially normal but, with the onset of motor dysfunction, becomes disrupted, accompanied by abnormal depolarization. Remarkably, subsequent Purkinje cell atrophy is associated with a restoration of pacemaker firing. The early inability of Purkinje neurons to support repetitive spiking is due to unopposed calcium currents resulting from a reduction in large-conductance calcium-activated potassium (BK) and subthreshold-activated potassium channels. The subsequent restoration of SCA1 Purkinje neuron firing correlates with the recovery of the density of these potassium channels that accompanies cell atrophy. Supporting a critical role for BK channels, viral-mediated increases in BK channel expression in SCA1 Purkinje neurons improves motor dysfunction and partially restores Purkinje neuron morphology. Cerebellar perfusion of flufenamic acid, an agent that restores the depolarized membrane potential of SCA1 Purkinje neurons by activating potassium channels, prevents Purkinje neuron dendritic atrophy. These results suggest that Purkinje neuron dendritic remodeling in ataxia is an adaptive response to increases in intrinsic membrane excitability. Similar adaptive remodeling could apply to other vulnerable neuronal populations in neurodegenerative disease. SIGNIFICANCE STATEMENT In neurodegenerative disease, neuronal atrophy has long been assumed to be an early nonspecific event preceding neuronal loss. However, in a mouse model of spinocerebellar ataxia type 1 (SCA1), we identify a previously unappreciated compensatory role for neuronal shrinkage. Purkinje neuron firing in these mice is initially normal, but is followed by abnormal membrane depolarization resulting from a reduction in potassium channels. Subsequently, these electrophysiological effects are counteracted by cell atrophy, which by restoring normal potassium channel membrane density, re-establishes pacemaker firing. Reversing the initial membrane depolarization improved motor function and Purkinje neuron morphology in the SCA1 mice. These results suggest that Purkinje neuron remodeling in ataxia is an active compensatory response that serves to normalize intrinsic membrane excitability. PMID:26269637

Mechanism of inhibition of mouse Slo3 (KCa 5.1) potassium channels by quinine, quinidine and barium.

PubMed

Wrighton, David C; Muench, Stephen P; Lippiat, Jonathan D

2015-09-01

The Slo3 (KCa 5.1) channel is a major component of mammalian KSper (sperm potassium conductance) channels and inhibition of these channels by quinine and barium alters sperm motility. The aim of this investigation was to determine the mechanism by which these drugs inhibit Slo3 channels. Mouse (m) Slo3 (KCa 5.1) channels or mutant forms were expressed in Xenopus oocytes and currents recorded with 2-electrode voltage-clamp. Gain-of-function mSlo3 mutations were used to explore the state-dependence of the inhibition. The interaction between quinidine and mSlo3 channels was modelled by in silico docking. Several drugs known to block KSper also affected mSlo3 channels with similar levels of inhibition. The inhibition induced by extracellular barium was prevented by increasing the extracellular potassium concentration. R196Q and F304Y mutations in the mSlo3 voltage sensor and pore, respectively, both increased channel activity. The F304Y mutation did not alter the effects of barium, but increased the potency of inhibition by both quinine and quinidine approximately 10-fold; this effect was not observed with the R196Q mutation. Block of mSlo3 channels by quinine, quinidine and barium is not state-dependent. Barium inhibits mSlo3 outside the cell by interacting with the selectivity filter, whereas quinine and quinidine act from the inside, by binding in a hydrophobic pocket formed by the S6 segment of each subunit. Furthermore, we propose that the Slo3 channel activation gate lies deep within the pore between F304 in the S6 segment and the selectivity filter. © 2015 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of The British Pharmacological Society.

BK potassium channels facilitate high-frequency firing and cause early spike frequency adaptation in rat CA1 hippocampal pyramidal cells

PubMed Central

Gu, Ning; Vervaeke, Koen; Storm, Johan F

2007-01-01

Neuronal potassium (K+) channels are usually regarded as largely inhibitory, i.e. reducing excitability. Here we show that BK-type calcium-activated K+ channels enhance high-frequency firing and cause early spike frequency adaptation in neurons. By combining slice electrophysiology and computational modelling, we investigated functions of BK channels in regulation of high-frequency firing in rat CA1 pyramidal cells. Blockade of BK channels by iberiotoxin (IbTX) selectively reduced the initial discharge frequency in response to strong depolarizing current injections, thus reducing the early spike frequency adaptation. IbTX also blocked the fast afterhyperpolarization (fAHP), slowed spike rise and decay, and elevated the spike threshold. Simulations with a computational model of a CA1 pyramidal cell confirmed that the BK channel-mediated rapid spike repolarization and fAHP limits activation of slower K+ channels (in particular the delayed rectifier potassium current (IDR)) and Na+ channel inactivation, whereas M-, sAHP- or SK-channels seem not to be important for the early facilitating effect. Since the BK current rapidly inactivates, its facilitating effect diminishes during the initial discharge, thus producing early spike frequency adaptation by an unconventional mechanism. This mechanism is highly frequency dependent. Thus, IbTX had virtually no effect at spike frequencies < 40 Hz. Furthermore, extracellular field recordings demonstrated (and model simulations supported) that BK channels contribute importantly to high-frequency burst firing in response to excitatory synaptic input to distal dendrites. These results strongly support the idea that BK channels play an important role for early high-frequency, rapidly adapting firing in hippocampal pyramidal neurons, thus promoting the type of bursting that is characteristic of these cells in vivo, during behaviour. PMID:17303637

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Ching-Shin; Pedersen, Bjørn Panyella; Stokes, David L.

Cellular potassium import systems play a fundamental role in osmoregulation, pH homeostasis and membrane potential in all domains of life. In bacteria, the kdp operon encodes a four-subunit potassium pump that maintains intracellular homeostasis, cell shape and turgor under conditions in which potassium is limiting1. This membrane complex, called KdpFABC, has one channel-like subunit (KdpA) belonging to the superfamily of potassium transporters and another pump-like subunit (KdpB) belonging to the superfamily of P-type ATPases. Although there is considerable structural and functional information about members of both superfamilies, the mechanism by which uphill potassium transport through KdpA is coupled with ATPmore » hydrolysis by KdpB remains poorly understood. Here we report the 2.9 Å X-ray structure of the complete Escherichia coli KdpFABC complex with a potassium ion within the selectivity filter of KdpA and a water molecule at a canonical cation site in the transmembrane domain of KdpB. The structure also reveals two structural elements that appear to mediate the coupling between these two subunits. Specifically, a protein-embedded tunnel runs between these potassium and water sites and a helix controlling the cytoplasmic gate of KdpA is linked to the phosphorylation domain of KdpB. On the basis of these observations, we propose a mechanism that repurposes protein channel architecture for active transport across biomembranes.« less

Overexpression of the rice AKT1 potassium channel affects potassium nutrition and rice drought tolerance

PubMed Central

Ahmad, Izhar; Mian, Afaq; Maathuis, Frans J. M.

2016-01-01

Potassium (K+) is the most important cationic nutrient for all living organisms and has roles in most aspects of plant physiology. To assess the impact of one of the main K+ uptake components, the K+ inward rectifying channel AKT1, we characterized both loss of function and overexpression of OsAKT1 in rice. In many conditions, AKT1 expression correlated with K+ uptake and tissue K+ levels. No salinity-related growth phenotype was observed for either loss or gain of function mutants. However, a correlation between AKT1 expression and root Na+ when the external Na/K ratio was high suggests that there may be a role for AKT1 in Na+ uptake in such conditions. In contrast to findings with Arabidopsis thaliana, we did not detect any change in growth of AKT1 loss of function mutants in the presence of NH4 +. Nevertheless, NH4 +-dependent inhibition was detected during K+ uptake assays in loss of function and wild type plants, depending on pre-growth conditions. The most prominent result of OsAKT1 overexpression was a reduction in sensitivity to osmotic/drought stress in transgenic plants: the data suggest that AKT1 overexpression improved rice osmotic and drought stress tolerance by increasing tissue levels of K+, especially in the root. PMID:26969743

Biophysics of BK Channel Gating.

PubMed

Pantazis, A; Olcese, R

2016-01-01

BK channels are universal regulators of cell excitability, given their exceptional unitary conductance selective for K(+), joint activation mechanism by membrane depolarization and intracellular [Ca(2+)] elevation, and broad expression pattern. In this chapter, we discuss the structural basis and operational principles of their activation, or gating, by membrane potential and calcium. We also discuss how the two activation mechanisms interact to culminate in channel opening. As members of the voltage-gated potassium channel superfamily, BK channels are discussed in the context of archetypal family members, in terms of similarities that help us understand their function, but also seminal structural and biophysical differences that confer unique functional properties. © 2016 Elsevier Inc. All rights reserved.

Genome-wide screening identifies a KCNIP1 copy number variant as a genetic predictor for atrial fibrillation

PubMed Central

Tsai, Chia-Ti; Hsieh, Chia-Shan; Chang, Sheng-Nan; Chuang, Eric Y.; Ueng, Kwo-Chang; Tsai, Chin-Feng; Lin, Tsung-Hsien; Wu, Cho-Kai; Lee, Jen-Kuang; Lin, Lian-Yu; Wang, Yi-Chih; Yu, Chih-Chieh; Lai, Ling-Ping; Tseng, Chuen-Den; Hwang, Juey-Jen; Chiang, Fu-Tien; Lin, Jiunn-Lee

2016-01-01

Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia. Previous genome-wide association studies had identified single-nucleotide polymorphisms in several genomic regions to be associated with AF. In human genome, copy number variations (CNVs) are known to contribute to disease susceptibility. Using a genome-wide multistage approach to identify AF susceptibility CNVs, we here show a common 4,470-bp diallelic CNV in the first intron of potassium interacting channel 1 gene (KCNIP1) is strongly associated with AF in Taiwanese populations (odds ratio=2.27 for insertion allele; P=6.23 × 10−24). KCNIP1 insertion is associated with higher KCNIP1 mRNA expression. KCNIP1-encoded protein potassium interacting channel 1 (KCHIP1) is physically associated with potassium Kv channels and modulates atrial transient outward current in cardiac myocytes. Overexpression of KCNIP1 results in inducible AF in zebrafish. In conclusions, a common CNV in KCNIP1 gene is a genetic predictor of AF risk possibly pointing to a functional pathway. PMID:26831368

Actions of bis(7)-tacrine and tacrine on transient potassium current in rat DRG neurons and potassium current mediated by K(V)4.2 expressed in Xenopus oocyte.

PubMed

Li, Xiang-Yuan; Zhang, Jian; Dai, Jia-Pei; Liu, Xiang-Ming; Li, Zhi-Wang

2010-03-08

Bis(7)-tacrine [bis(7)-tetrahydroaminacrine] is a dimeric AChE inhibitor derived from tacrine with a potential to treat Alzheimer's disease. Actions of bis(7)-tacrine on ligand-gated ion channels and voltage-gated cation channels have been identified on neurons of both central and peripheral nervous systems. In the present study, the effect of bis(7)-tacrine was investigated on the K(V)4.2 encoded potassium currents expressed in Xenopus oocytes and the transient A-type potassium current (I(K(A))) on rat DRG neurons. Bis(7)-tacrine suppressed recombinant Kv4.2 potassium channels in a concentration-dependent manner, with IC(50) value of 0.53+/-0.13 muM. Tacrine also inhibited Kv4.2 channels, but with a much lower potency (IC(50) 74+/-15 muM).The possible mechanisms underlying the inhibition on potassium currents by bis(7)-tacrine/tacrine could be that inactivation of the transient potassium currents was accelerated and recovery of the native or Kv4.2 expressed potassium currents was suppressed by bis(7)-tacrine/tacrine. Copyright 2010 Elsevier B.V. All rights reserved.

Antibodies to Kv1 potassium channel-complex proteins leucine-rich, glioma inactivated 1 protein and contactin-associated protein-2 in limbic encephalitis, Morvan’s syndrome and acquired neuromyotonia

PubMed Central

Irani, Sarosh R.; Alexander, Sian; Waters, Patrick; Kleopa, Kleopas A.; Pettingill, Philippa; Zuliani, Luigi; Peles, Elior; Buckley, Camilla; Lang, Bethan

2010-01-01

Antibodies that immunoprecipitate 125I-α-dendrotoxin-labelled voltage-gated potassium channels extracted from mammalian brain tissue have been identified in patients with neuromyotonia, Morvan’s syndrome, limbic encephalitis and a few cases of adult-onset epilepsy. These conditions often improve following immunomodulatory therapies. However, the proportions of the different syndromes, the numbers with associated tumours and the relationships with potassium channel subunit antibody specificities have been unclear. We documented the clinical phenotype and tumour associations in 96 potassium channel antibody positive patients (titres >400 pM). Five had thymomas and one had an endometrial adenocarcinoma. To define the antibody specificities, we looked for binding of serum antibodies and their effects on potassium channel currents using human embryonic kidney cells expressing the potassium channel subunits. Surprisingly, only three of the patients had antibodies directed against the potassium channel subunits. By contrast, we found antibodies to three proteins that are complexed with 125I-α-dendrotoxin-labelled potassium channels in brain extracts: (i) contactin-associated protein-2 that is localized at the juxtaparanodes in myelinated axons; (ii) leucine-rich, glioma inactivated 1 protein that is most strongly expressed in the hippocampus; and (iii) Tag-1/contactin-2 that associates with contactin-associated protein-2. Antibodies to Kv1 subunits were found in three sera, to contactin-associated protein-2 in 19 sera, to leucine-rich, glioma inactivated 1 protein in 55 sera and to contactin-2 in five sera, four of which were also positive for the other antibodies. The remaining 18 sera were negative for potassium channel subunits and associated proteins by the methods employed. Of the 19 patients with contactin-associated protein-antibody-2, 10 had neuromyotonia or Morvan’s syndrome, compared with only 3 of the 55 leucine-rich, glioma inactivated 1 protein-antibody positive patients (P < 0.0001), who predominantly had limbic encephalitis. The responses to immunomodulatory therapies, defined by changes in modified Rankin scores, were good except in the patients with tumours, who all had contactin-associated-2 protein antibodies. This study confirms that the majority of patients with high potassium channel antibodies have limbic encephalitis without tumours. The identification of leucine-rich, glioma inactivated 1 protein and contactin-associated protein-2 as the major targets of potassium channel antibodies, and their associations with different clinical features, begins to explain the diversity of these syndromes; furthermore, detection of contactin-associated protein-2 antibodies should help identify the risk of an underlying tumour and a poor prognosis in future patients. PMID:20663977

Selection of Inhibitor-Resistant Viral Potassium Channels Identifies a Selectivity Filter Site that Affects Barium and Amantadine Block

PubMed Central

Fujiwara, Yuichiro; Arrigoni, Cristina; Domigan, Courtney; Ferrara, Giuseppina; Pantoja, Carlos; Thiel, Gerhard; Moroni, Anna; Minor, Daniel L.

2009-01-01

Background Understanding the interactions between ion channels and blockers remains an important goal that has implications for delineating the basic mechanisms of ion channel function and for the discovery and development of ion channel directed drugs. Methodology/Principal Findings We used genetic selection methods to probe the interaction of two ion channel blockers, barium and amantadine, with the miniature viral potassium channel Kcv. Selection for Kcv mutants that were resistant to either blocker identified a mutant bearing multiple changes that was resistant to both. Implementation of a PCR shuffling and backcrossing procedure uncovered that the blocker resistance could be attributed to a single change, T63S, at a position that is likely to form the binding site for the inner ion in the selectivity filter (site 4). A combination of electrophysiological and biochemical assays revealed a distinct difference in the ability of the mutant channel to interact with the blockers. Studies of the analogous mutation in the mammalian inward rectifier Kir2.1 show that the T→S mutation affects barium block as well as the stability of the conductive state. Comparison of the effects of similar barium resistant mutations in Kcv and Kir2.1 shows that neighboring amino acids in the Kcv selectivity filter affect blocker binding. Conclusions/Significance The data support the idea that permeant ions have an integral role in stabilizing potassium channel structure, suggest that both barium and amantadine act at a similar site, and demonstrate how genetic selections can be used to map blocker binding sites and reveal mechanistic features. PMID:19834614

Movement of Potassium Ions inside KcsA in the High Concentration Regime using a Molecular Dynamics Simulation

NASA Astrophysics Data System (ADS)

Kim, Myojeong; Jo, Byeong Chul; Yoon, Hyun Jung; Wu, Sangwook; Thangappan, Jayaraman; Eun, Changsun

2018-05-01

The selectivity and conduction specificity of the KcsA channel toward potassium ions is crucial to the activity of this protein and this channel is intricately associated with several osmotic regulation and neuronal signaling processes. Despite multi-ion characteristics, the selective conduction behavior of KcsA is controlled by the size and distance specific electrostatic interaction between the selected residues and the potassium ions. The mechanism describing the movement of potassium ions in the channel and the conformational changes to KcsA that facilitate ion movement were investigated by a molecular dynamics (MD) simulation. In this study, we analyze the movement of potassium ions and water molecules at various time intervals during a 90 ns molecular dynamics simulation in the high potassium ion concentration regime and in the absence of the voltage. Examination of specific (3.6, 17.3, 43.38 and 43.44 ns) simulation periods revealed that key residues in the selectivity filter of KcsA influence the movement of potassium ions in the channel.

Sodium-dependent potassium channels of a Slack-like subtype contribute to the slow afterhyperpolarization in lamprey spinal neurons

PubMed Central

Wallén, Peter; Robertson, Brita; Cangiano, Lorenzo; Löw, Peter; Bhattacharjee, Arin; Kaczmarek, Leonard K; Grillner, Sten

2007-01-01

The slow afterhyperpolarization (sAHP) following the action potential is the main determinant of spike frequency regulation. The sAHP after single action potentials in neurons of the lamprey locomotor network is largely due to calcium-dependent K+ channels (80%), activated by calcium entering the cell during the spike. The residual (20%) component becomes prominent during high level activity (50% of the sAHP). It is not Ca2+ dependent, has a reversal potential like that of potassium, and is not affected by chloride injection. It is not due to rapid activation of Na+/K+-ATPase. This non-KCa-sAHP is reduced markedly in amplitude when sodium ions are replaced by lithium ions, and is thus sodium dependent. Quinidine also blocks this sAHP component, further indicating an involvement of sodium-dependent potassium channels (KNa). Modulators tested do not influence the KNa-sAHP amplitude. Immunofluorescence labelling with an anti-Slack antibody revealed distinct immunoreactivity of medium-sized and large neurons in the grey matter of the lamprey spinal cord, suggesting the presence of a Slack-like subtype of KNa channel. The results strongly indicate that a KNa potassium current contributes importantly to the sAHP and thereby to neuronal frequency regulation during high level burst activity as during locomotion. This is, to our knowledge, the first demonstration of a functional role for the Slack gene in contributing to the slow AHP. PMID:17884929

A human intermediate conductance calcium-activated potassium channel

PubMed Central

Ishii, Takahiro M.; Silvia, Christopher; Hirschberg, Birgit; Bond, Chris T.; Adelman, John P.; Maylie, James

1997-01-01

An intermediate conductance calcium-activated potassium channel, hIK1, was cloned from human pancreas. The predicted amino acid sequence is related to, but distinct from, the small conductance calcium-activated potassium channel subfamily, which is ≈50% conserved. hIK1 mRNA was detected in peripheral tissues but not in brain. Expression of hIK1 in Xenopus oocytes gave rise to inwardly rectifying potassium currents, which were activated by submicromolar concentrations of intracellular calcium (K0.5 = 0.3 μM). Although the K0.5 for calcium was similar to that of small conductance calcium-activated potassium channels, the slope factor derived from the Hill equation was significantly reduced (1.7 vs. 3.5). Single-channel current amplitudes reflected the macroscopic inward rectification and revealed a conductance level of 39 pS in the inward direction. hIK1 currents were reversibly blocked by charybdotoxin (Ki = 2.5 nM) and clotrimazole (Ki = 24.8 nM) but were minimally affected by apamin (100 nM), iberiotoxin (50 nM), or ketoconazole (10 μM). These biophysical and pharmacological properties are consistent with native intermediate conductance calcium-activated potassium channels, including the erythrocyte Gardos channel. PMID:9326665

Down-regulation of A-type potassium channel in gastric-specific DRG neurons in a rat model of functional dyspepsia.

PubMed

Li, S; Chen, J D Z

2014-07-01

Although without evidence of organic structural abnormalities, pain or discomfort is a prominent symptom of functional dyspepsia and considered to reflect visceral hypersensitivity whose underlying mechanism is poorly understood. Here, we studied electrophysiological properties and expression of voltage-gated potassium channels in dorsal root ganglion (DRG) neurons in a rat model of functional dyspepsia induced by neonatal gastric irritation. Male Sprague-Dawley rat pups at 10-day old received 0.1% iodoacetamide (IA) or vehicle by oral gavage for 6 days and studied at adulthood. Retrograde tracer-labeled gastric-specific T8 -T12 DRG neurons were harvested for the patch-clamp study in voltage and current-clamp modes and protein expression of K(+) channel in T8 -T12 DRGs was examined by western blotting. (1) Gastric specific but not non-gastric DRG neurons showed an enhanced excitability in neonatal IA-treated rats compared to the control: depolarized resting membrane potentials, a lower current threshold for action potential (AP) activation, and an increase in the number of APs in response to current stimulation. (2) The current density of tetraethylammonium insensitive (transiently inactivating A-type current), but not the tetraethylammonium sensitive (slow-inactivating delayed rectifier K(+) currents), was significantly smaller in IA-treated rats (65.4 ± 6.9 pA/pF), compared to that of control (93.1 ± 8.3 pA/pF). (3) Protein expression of KV 4.3 was down-regulated in IA-treated rats. A-type potassium channels are significantly down-regulated in the gastric-specific DRG neurons in adult rats with mild neonatal gastric irritation, which in part contribute to the enhanced DRG neuron excitabilities that leads to the development of gastric hypersensitivity. © 2014 John Wiley & Sons Ltd.

Molecular basis for the toxin insensitivity of scorpion voltage-gated potassium channel MmKv1.

PubMed

Zhang, Chuangeng; Xie, Zili; Li, Xinxin; Chen, Jing; Feng, Jing; Lang, Yange; Yang, Weishan; Li, Wenxin; Chen, Zongyun; Yao, Jing; Cao, Zhijian; Wu, Yingliang

2016-05-01

Scorpions are insensitive to their own venoms, which contain various neurotoxins specific for mammalian or insect ion channels, whose molecular mechanism remains unsolved. Using MmKv1, a potassium channel identified from the genome of the scorpion Mesobuthus martensii, channel kinetic experiments showed that MmKv1 was a classical voltage-gated potassium channel with a voltage-dependent fast activation and slow inactivation. Compared with the human Kv1.3 channel (hKv1.3), the MmKv1 channel exhibited a remarkable insensitivity to both scorpion venom and toxin. The chimaeric channels of MmKv1 and hKv1.3 revealed that both turret and filter regions of the MmKv1 channel were critical for the toxin insensitivity of MmKv1. Furthermore, mutagenesis of the chimaeric channel indicated that two basic residues (Arg(399) and Lys(403)) in the MmKv1 turret region and Arg(425) in the MmKv1 filter region significantly affected its toxin insensitivity. Moreover, when these three basic residues of MmKv1 were simultaneously substituted with the corresponding residues from hKv1.3, the MmKv1-R399T/K403S/R425H mutant channels exhibited similar sensitivity to both scorpion venom and toxin to hKv1.3, which revealed the determining role of these three basic residues in the toxin insensitivity of the MmKv1 channel. More strikingly, a similar triad sequence structure is present in all Shaker-like channels from venomous invertebrates, which suggested a possible convergent functional evolution of these channels to enable them to resist their own venoms. Together, these findings first illustrate the mechanism by which scorpions are insensitive to their own venoms at the ion channel receptor level and enrich our knowledge of the insensitivity of scorpions and other venomous animals to their own venoms. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

A Nanoscale Interface Promoting Molecular and Functional Differentiation of Neural Cells

NASA Astrophysics Data System (ADS)

Posati, Tamara; Pistone, Assunta; Saracino, Emanuela; Formaggio, Francesco; Mola, Maria Grazia; Troni, Elisabetta; Sagnella, Anna; Nocchetti, Morena; Barbalinardo, Marianna; Valle, Francesco; Bonetti, Simone; Caprini, Marco; Nicchia, Grazia Paola; Zamboni, Roberto; Muccini, Michele; Benfenati, Valentina

2016-08-01

Potassium channels and aquaporins expressed by astrocytes are key players in the maintenance of cerebral homeostasis and in brain pathophysiologies. One major challenge in the study of astrocyte membrane channels in vitro, is that their expression pattern does not resemble the one observed in vivo. Nanostructured interfaces represent a significant resource to control the cellular behaviour and functionalities at micro and nanoscale as well as to generate novel and more reliable models to study astrocytes in vitro. However, the potential of nanotechnologies in the manipulation of astrocytes ion channels and aquaporins has never been previously reported. Hydrotalcite-like compounds (HTlc) are layered materials with increasing potential as biocompatible nanoscale interface. Here, we evaluate the effect of the interaction of HTlc nanoparticles films with primary rat neocortical astrocytes. We show that HTlc films are biocompatible and do not promote gliotic reaction, while favouring astrocytes differentiation by induction of F-actin fibre alignment and vinculin polarization. Western Blot, Immunofluorescence and patch-clamp revealed that differentiation was accompanied by molecular and functional up-regulation of both inward rectifying potassium channel Kir 4.1 and aquaporin 4, AQP4. The reported results pave the way to engineering novel in vitro models to study astrocytes in a in vivo like condition.

Potassium channels: the importance of transport signals.

PubMed

Griffith, L C

2001-03-20

The number, type and distribution of ion channels on a neuron's surface determine its electrical response to stimulation. One way that a cell determines how many molecules of each channel type are sent to the surface has been eludicated in a recent study of intrinsic protein transport signals within potassium channels.

Effects of channel noise on firing coherence of small-world Hodgkin-Huxley neuronal networks

NASA Astrophysics Data System (ADS)

Sun, X. J.; Lei, J. Z.; Perc, M.; Lu, Q. S.; Lv, S. J.

2011-01-01

We investigate the effects of channel noise on firing coherence of Watts-Strogatz small-world networks consisting of biophysically realistic HH neurons having a fraction of blocked voltage-gated sodium and potassium ion channels embedded in their neuronal membranes. The intensity of channel noise is determined by the number of non-blocked ion channels, which depends on the fraction of working ion channels and the membrane patch size with the assumption of homogeneous ion channel density. We find that firing coherence of the neuronal network can be either enhanced or reduced depending on the source of channel noise. As shown in this paper, sodium channel noise reduces firing coherence of neuronal networks; in contrast, potassium channel noise enhances it. Furthermore, compared with potassium channel noise, sodium channel noise plays a dominant role in affecting firing coherence of the neuronal network. Moreover, we declare that the observed phenomena are independent of the rewiring probability.

Arrhythmogenic KCNE gene variants: current knowledge and future challenges

PubMed Central

Crump, Shawn M.; Abbott, Geoffrey W.

2014-01-01

There are twenty-five known inherited cardiac arrhythmia susceptibility genes, all of which encode either ion channel pore-forming subunits or proteins that regulate aspects of ion channel biology such as function, trafficking, and localization. The human KCNE gene family comprises five potassium channel regulatory subunits, sequence variants in each of which are associated with cardiac arrhythmias. KCNE gene products exhibit promiscuous partnering and in some cases ubiquitous expression, hampering efforts to unequivocally correlate each gene to specific native potassium currents. Likewise, deducing the molecular etiology of cardiac arrhythmias in individuals harboring rare KCNE gene variants, or more common KCNE polymorphisms, can be challenging. In this review we provide an update on putative arrhythmia-causing KCNE gene variants, and discuss current thinking and future challenges in the study of molecular mechanisms of KCNE-associated cardiac rhythm disturbances. PMID:24478792

The inward rectifier potassium channel Kir2.1 is expressed in mouse neutrophils from bone marrow and liver.

PubMed

Masia, Ricard; Krause, Daniela S; Yellen, Gary

2015-02-01

Neutrophils are phagocytic cells that play a critical role in innate immunity by destroying bacterial pathogens. Channels belonging to the inward rectifier potassium channel subfamily 2 (Kir2 channels) have been described in other phagocytes (monocytes/macrophages and eosinophils) and in hematopoietic precursors of phagocytes. Their physiological function in these cells remains unclear, but some evidence suggests a role in growth factor-dependent proliferation and development. Expression of functional Kir2 channels has not been definitively demonstrated in mammalian neutrophils. Here, we show by RT-PCR that neutrophils from mouse bone marrow and liver express mRNA for the Kir2 subunit Kir2.1 but not for other subunits (Kir2.2, Kir2.3, and Kir2.4). In electrophysiological experiments, resting (unstimulated) neutrophils from mouse bone marrow and liver exhibit a constitutively active, external K(+)-dependent, strong inwardly rectifying current that constitutes the dominant current. The reversal potential is dependent on the external K(+) concentration in a Nernstian fashion, as expected for a K(+)-selective current. The current is not altered by changes in external or internal pH, and it is blocked by Ba(2+), Cs(+), and the Kir2-selective inhibitor ML133. The single-channel conductance is in agreement with previously reported values for Kir2.1 channels. These properties are characteristic of homomeric Kir2.1 channels. Current density in short-term cultures of bone marrow neutrophils is decreased in the absence of growth factors that are important for neutrophil proliferation [granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF)]. These results demonstrate that mouse neutrophils express functional Kir2.1 channels and suggest that these channels may be important for neutrophil function, possibly in a growth factor-dependent manner. Copyright © 2015 the American Physiological Society.

Regulation of human cardiac potassium channels by full-length KCNE3 and KCNE4.

PubMed

Abbott, Geoffrey W

2016-12-06

Voltage-gated potassium (Kv) channels comprise pore-forming α subunits and a multiplicity of regulatory proteins, including the cardiac-expressed and cardiac arrhythmia-linked transmembrane KCNE subunits. After recently uncovering novel, N-terminally extended (L) KCNE3 and KCNE4 isoforms and detecting their transcripts in human atrium, reported here are their functional effects on human cardiac Kv channel α subunits expressed in Xenopus laevis oocytes. As previously reported for short isoforms KCNE3S and KCNE4S, KCNE3L inhibited hERG; KCNE4L inhibited Kv1.1; neither form regulated the HCN1 pacemaker channel. Unlike KCNE4S, KCNE4L was a potent inhibitor of Kv4.2 and Kv4.3; co-expression of cytosolic β subunit KChIP2, which regulates Kv4 channels in cardiac myocytes, partially relieved Kv4.3 but not Kv4.2 inhibition. Inhibition of Kv4.2 and Kv4.3 by KCNE3L was weaker, and its inhibition of Kv4.2 abolished by KChIP2. KCNE3L and KCNE4L also exhibited subunit-specific effects on Kv4 channel complex inactivation kinetics, voltage dependence and recovery. Further supporting the potential physiological significance of the robust functional effects of KCNE4L on Kv4 channels, KCNE4L protein was detected in human atrium, where it co-localized with Kv4.3. The findings establish functional effects of novel human cardiac-expressed KCNE isoforms and further contribute to our understanding of the potential mechanisms influencing cardiomyocyte repolarization.

A potent potassium channel blocker from Mesobuthus eupeus scorpion venom.

PubMed

Gao, Bin; Peigneur, Steve; Tytgat, Jan; Zhu, Shunyi

2010-12-01

Scorpion venom-derived peptidyl toxins are valuable pharmacological tools for investigating the structure-function relationship of ion channels. Here, we report the purification, sequencing and functional characterization of a new K(+) channel blocker (MeuKTX) from the venom of the scorpion Mesobuthus eupeus. Effects of MeuKTX on ten cloned potassium channels in Xenopus oocytes were evaluated using two-electrode voltage-clamp recordings. MeuKTX is the orthologue of BmKTX (α-KTx3.6), a known Kv1.3 blocker from the scorpion Mesobuthus martensii, and classified as α-KTx3.13. MeuKTX potently blocks rKv1.1, rKv1.2 and hKv1.3 channels with 50% inhibitory concentration (IC(50)) of 203.15 ± 4.06 pM, 8.92 ± 2.3 nM and 171 ± 8.56 pM, respectively, but does not affect rKv1.4, rKv1.5, hKv3.1, rKv4.3, and hERG channels even at 2 μM concentration. At this high concentration, MeuKTX is also active on rKv1.6 and Shaker IR. Our results also demonstrate that MeuKTX and BmKTX have the same channel spectrum and similar pharmacological potency. Analysis of the structure-function relationships of α-KTx3 subfamily toxins allows us to recognize several key sites which may be useful for designing toxins with improved activity on hKv1.3, an attractive target for T-cell mediated autoimmune diseases. Copyright © 2010 Elsevier Masson SAS. All rights reserved.

Overexpression of Large-Conductance Calcium-Activated Potassium Channels in Human Glioblastoma Stem-Like Cells and Their Role in Cell Migration.

PubMed

Rosa, Paolo; Sforna, Luigi; Carlomagno, Silvia; Mangino, Giorgio; Miscusi, Massimo; Pessia, Mauro; Franciolini, Fabio; Calogero, Antonella; Catacuzzeno, Luigi

2017-09-01

Glioblastomas (GBMs) are brain tumors characterized by diffuse invasion of cancer cells into the healthy brain parenchyma, and establishment of secondary foci. GBM cells abundantly express large-conductance, calcium-activated potassium (BK) channels that are thought to promote cell invasion. Recent evidence suggests that the GBM high invasive potential mainly originates from a pool of stem-like cells, but the expression and function of BK channels in this cell subpopulation have not been studied. We investigated the expression of BK channels in GBM stem-like cells using electrophysiological and immunochemical techniques, and assessed their involvement in the migratory process of this important cell subpopulation. In U87-MG cells, BK channel expression and function were markedly upregulated by growth conditions that enriched the culture in GBM stem-like cells (U87-NS). Cytofluorimetric analysis further confirmed the appearance of a cell subpopulation that co-expressed high levels of BK channels and CD133, as well as other stem cell markers. A similar association was also found in cells derived from freshly resected GBM biopsies. Finally, transwell migration tests showed that U87-NS cells migration was much more sensitive to BK channel block than U87-MG cells. Our data show that BK channels are highly expressed in GBM stem-like cells, and participate to their high migratory activity. J. Cell. Physiol. 232: 2478-2488, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Genetics Home Reference: Cantú syndrome

MedlinePlus

... syndrome is a rare condition characterized by excess hair growth (hypertrichosis), a distinctive facial appearance, heart defects, and ... problem with potassium channel function leads to excess hair growth, heart defects, and the other features of Cantú ...

Broadening roles for FMRP: big news for big potassium (BK) channels.

PubMed

Contractor, Anis

2013-02-20

FMRP is an RNA-binding protein that negatively regulates translation and which is lost in fragile X syndrome. In this issue of Neuron, Deng et al. (2013) demonstrate a novel translation-independent function for FMRP as a regulator of presynaptic BK channels that modulate the dynamics of neurotransmitter release. Copyright © 2013 Elsevier Inc. All rights reserved.

Structural Analysis and Deletion Mutagenesis Define Regions of QUIVER/SLEEPLESS that Are Responsible for Interactions with Shaker-Type Potassium Channels and Nicotinic Acetylcholine Receptors

PubMed Central

Wu, Meilin; Liu, Clifford Z.; Joiner, William J.

2016-01-01

Ly6 proteins are endogenous prototoxins found in most animals. They show striking structural and functional parallels to snake α-neurotoxins, including regulation of ion channels and cholinergic signaling. However, the structural contributions of Ly6 proteins to regulation of effector molecules is poorly understood. This question is particularly relevant to the Ly6 protein QUIVER/SLEEPLESS (QVR/SSS), which has previously been shown to suppress excitability and synaptic transmission by upregulating potassium (K) channels and downregulating nicotinic acetylcholine receptors (nAChRs) in wake-promoting neurons to facilitate sleep in Drosophila. Using deletion mutagenesis, co-immunoprecipitations, ion flux assays, surface labeling and confocal microscopy, we demonstrate that only loop 2 is required for many of the previously described properties of SSS in transfected cells, including interactions with K channels and nAChRs. Collectively our data suggest that QVR/SSS, and by extension perhaps other Ly6 proteins, target effector molecules using limited protein motifs. Mapping these motifs may be useful in rational design of drugs that mimic or suppress Ly6-effector interactions to modulate nervous system function. PMID:26828958

Role of potassium ion channels in detrusor smooth muscle function and dysfunction

PubMed Central

Petkov, Georgi V.

2013-01-01

Contraction and relaxation of the detrusor smooth muscle (DSM), which makes up the wall of the urinary bladder, facilitates the storage and voiding of urine. Several families of K+ channels, including voltage-gated K+ (KV) channels, Ca2+-activated K+ (KCa) channels, inward-rectifying ATP-sensitive K+ (Kir, KATP) channels, and two-pore-domain K+ (K2P) channels, are expressed and functional in DSM. They control DSM excitability and contractility by maintaining the resting membrane potential and shaping the action potentials that determine the phasic nature of contractility in this tissue. Defects in DSM K+ channel proteins or in the molecules involved in their regulatory pathways may underlie certain forms of bladder dysfunction, such as overactive bladder. K+ channels represent an opportunity for novel pharmacological manipulation and therapeutic intervention in human DSM. Modulation of DSM K+ channels directly or indirectly by targeting their regulatory mechanisms has the potential to control urinary bladder function. This Review summarizes our current state of knowledge of the functional role of K+ channels in DSM in health and disease, with special emphasis on current advancements in the field. PMID:22158596

Remote and reversible inhibition of neurons and circuits by small molecule induced potassium channel stabilization

PubMed Central

Auffenberg, Eva; Jurik, Angela; Mattusch, Corinna; Stoffel, Rainer; Genewsky, Andreas; Namendorf, Christian; Schmid, Roland M.; Rammes, Gerhard; Biel, Martin; Uhr, Manfred; Moosmang, Sven; Michalakis, Stylianos; Wotjak, Carsten T.; Thoeringer, Christoph K.

2016-01-01

Manipulating the function of neurons and circuits that translate electrical and chemical signals into behavior represents a major challenges in neuroscience. In addition to optogenetic methods using light-activatable channels, pharmacogenetic methods with ligand induced modulation of cell signaling and excitability have been developed. However, they are largely based on ectopic expression of exogenous or chimera proteins. Now, we describe the remote and reversible expression of a Kir2.1 type potassium channel using the chemogenetic technique of small molecule induced protein stabilization. Based on shield1-mediated shedding of a destabilizing domain fused to a protein of interest and inhibition of protein degradation, this principle has been adopted for biomedicine, but not in neuroscience so far. Here, we apply this chemogenetic approach in brain research for the first time in order to control a potassium channel in a remote and reversible manner. We could show that shield1-mediated ectopic Kir2.1 stabilization induces neuronal silencing in vitro and in vivo in the mouse brain. We also validated this novel pharmacogenetic method in different neurobehavioral paradigms.The DD-Kir2.1 may complement the existing portfolio of pharmaco- and optogenetic techniques for specific neuron manipulation, but it may also provide an example for future applications of this principle in neuroscience research. PMID:26757616

Interaction of the scorpion toxin discrepin with Kv4.3 channels and A-type K(+) channels in cerebellum granular cells.

PubMed

Picco, Cristiana; Corzo, Gerardo; Possani, Lourival D; Prestipino, Gianfranco

2014-09-01

The peptide discrepin from the α-KTx15 subfamily of scorpion toxins preferentially affects transient A-type potassium currents, which regulate many aspects of neuronal function in the central nervous system. However, the specific Kv channel targeted by discrepin and the molecular mechanism of interaction are still unknown. Different variant peptides of discrepin were chemically synthesized and their effects were studied using patch clamp technique on rat cerebellum granular cells (CGC) and HEK cells transiently expressing Kv4.3 channels. Functional analysis indicated that nanomolar concentrations of native discrepin blocked Kv4.3 expressed channels, as previously observed in CGC. Similarly, the apparent affinities of all mutated peptides for Kv4.3 expressed channels were analogous to those found in CGC. In particular, in the double variant [V6K, D20K] the apparent affinity increased about 10-fold, whereas in variants carrying a deletion (ΔK13) or substitution (K13A) at position K13, the blockage was removed and the apparent affinity decreased more than 20-fold. These results indicate that Kv4.3 is likely the target of discrepin and highlight the importance of the basic residue K13, located in the α-helix of the toxin, for current blockage. We report the first example of a Kv4 subfamily potassium channel blocked by discrepin and identify the amino acid residues responsible for the blockage. The availability of discrepin variant peptides stimulates further research on the functions and pharmacology of neuronal Kv4 channels and on their possible roles in neurodegenerative disorders. Copyright © 2014 Elsevier B.V. All rights reserved.

Toxin-induced conformational changes in a potassium channel revealed by solid-state NMR

NASA Astrophysics Data System (ADS)

Lange, Adam; Giller, Karin; Hornig, Sönke; Martin-Eauclaire, Marie-France; Pongs, Olaf; Becker, Stefan; Baldus, Marc

2006-04-01

The active site of potassium (K+) channels catalyses the transport of K+ ions across the plasma membrane-similar to the catalytic function of the active site of an enzyme-and is inhibited by toxins from scorpion venom. On the basis of the conserved structures of K+ pore regions and scorpion toxins, detailed structures for the K+ channel-scorpion toxin binding interface have been proposed. In these models and in previous solution-state nuclear magnetic resonance (NMR) studies using detergent-solubilized membrane proteins, scorpion toxins were docked to the extracellular entrance of the K+ channel pore assuming rigid, preformed binding sites. Using high-resolution solid-state NMR spectroscopy, here we show that high-affinity binding of the scorpion toxin kaliotoxin to a chimaeric K+ channel (KcsA-Kv1.3) is associated with significant structural rearrangements in both molecules. Our approach involves a combined analysis of chemical shifts and proton-proton distances and demonstrates that solid-state NMR is a sensitive method for analysing the structure of a membrane protein-inhibitor complex. We propose that structural flexibility of the K+ channel and the toxin represents an important determinant for the high specificity of toxin-K+ channel interactions.

Scorpion Toxins Specific for Potassium (K+) Channels: A Historical Overview of Peptide Bioengineering

PubMed Central

Bergeron, Zachary L.; Bingham, Jon-Paul

2012-01-01

Scorpion toxins have been central to the investigation and understanding of the physiological role of potassium (K+) channels and their expansive function in membrane biophysics. As highly specific probes, toxins have revealed a great deal about channel structure and the correlation between mutations, altered regulation and a number of human pathologies. Radio- and fluorescently-labeled toxin isoforms have contributed to localization studies of channel subtypes in expressing cells, and have been further used in competitive displacement assays for the identification of additional novel ligands for use in research and medicine. Chimeric toxins have been designed from multiple peptide scaffolds to probe channel isoform specificity, while advanced epitope chimerization has aided in the development of novel molecular therapeutics. Peptide backbone cyclization has been utilized to enhance therapeutic efficiency by augmenting serum stability and toxin half-life in vivo as a number of K+-channel isoforms have been identified with essential roles in disease states ranging from HIV, T-cell mediated autoimmune disease and hypertension to various cardiac arrhythmias and Malaria. Bioengineered scorpion toxins have been monumental to the evolution of channel science, and are now serving as templates for the development of invaluable experimental molecular therapeutics. PMID:23202307

Lower KV7.5 Potassium Channel Subunit Expression in an Animal Model of Paroxysmal Dystonia.

PubMed

Sander, Svenja E; Diwan, Mustansir; Raymond, Roger; Nobrega, José N; Richter, Angelika

2016-01-01

Dystonia is a hyperkinetic disabling movement disorder. In the dt(sz) hamster, a model of paroxysmal dystonia, pronounced antidystonic effects of the KV7.2-5 potassium channel opener retigabine and aggravation of dystonia by a selective KV7.2-5 blocker indicated a pathophysiological role of an abnormal expression of KV7 channels. We therefore investigated the expression of KV7 subunits in brains of dystonic hamsters. While KV7.2 and KV7.3 subunits were unaltered, lower KV7.5 mRNA levels became evident in motor areas and in limbic structures of dystonic hamsters. The KV7.2/3 subunit-preferring channel opener N-(6-chloropyridin-3-yl)-3,4- difluorobenzamide (ICA 27243; 10-30 mg/kg i.p.) failed to reduce the severity of dystonia in mutant hamsters, suggesting that the previously observed antidystonic action of retigabine is mediated by the activation of KV7.5 channels. The experiments indicate a functional relevance for KV7.5 channels in paroxysmal dystonia. We suggest that compounds highly selective for subtypes of KV7 channels, i.e. for KV7.5, may provide new therapeutic approaches.

The Role of Potassium Channels in Arabidopsis thaliana Long Distance Electrical Signalling: AKT2 Modulates Tissue Excitability While GORK Shapes Action Potentials.

PubMed

Cuin, Tracey Ann; Dreyer, Ingo; Michard, Erwan

2018-03-21

Fast responses to an external threat depend on the rapid transmission of signals through a plant. Action potentials (APs) are proposed as such signals. Plant APs share similarities with their animal counterparts; they are proposed to depend on the activity of voltage-gated ion channels. Nonetheless, despite their demonstrated role in (a)biotic stress responses, the identities of the associated voltage-gated channels and transporters remain undefined in higher plants. By demonstrating the role of two potassium-selective channels in Arabidopsis thaliana in AP generation and shaping, we show that the plant AP does depend on similar Kv -like transport systems to those of the animal signal. We demonstrate that the outward-rectifying potassium-selective channel GORK limits the AP amplitude and duration, while the weakly-rectifying channel AKT2 affects membrane excitability. By computational modelling of plant APs, we reveal that the GORK activity not only determines the length of an AP but also the steepness of its rise and the maximal amplitude. Thus, outward-rectifying potassium channels contribute to both the repolarisation phase and the initial depolarisation phase of the signal. Additionally, from modelling considerations we provide indications that plant APs might be accompanied by potassium waves, which prime the excitability of the green cable.

Inward rectifier potassium (Kir2.1) channels as end-stage boosters of endothelium-dependent vasodilators.

PubMed

Sonkusare, Swapnil K; Dalsgaard, Thomas; Bonev, Adrian D; Nelson, Mark T

2016-06-15

Increase in endothelial cell (EC) calcium activates calcium-sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium-dependent vasodilatation. Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium-dependent vasodilatation is not clear. In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform. Endothelium-dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown. These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Endothelium-dependent vasodilators, such as acetylcholine, increase intracellular Ca(2+) through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca(2+) -sensitive intermediate and small conductance K(+) (IK and SK, respectively) channels. Although strong inward rectifier K(+) (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC-dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC-dependent vasodilatation of resistance-sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC-specific Kir2.1 knockdown (EC-Kir2.1(-/-) ) mice. Elevation of extracellular K(+) to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial denudation and Kir channel inhibitors (Ba(2+) , ML-133) or in the arteries from EC-Kir2.1(-/-) mice. Potassium-induced dilatations were unaffected by inhibitors of TRPV4, IK and SK channels. The Kir channel blocker, Ba(2+) , did not affect currents through TRPV4, IK or SK channels. Endothelial cell-dependent vasodilatations in response to activation of muscarinic receptors, TRPV4 channels or IK/SK channels were reduced, but not eliminated, by Kir channel inhibitors or EC-Kir2.1(-/-) . In angiotensin II-induced hypertension, the Kir channel function was not altered, although the endothelium-dependent vasodilatation was severely impaired. Our results support the concept that EC Kir2 channels boost vasodilatory signals that are generated by Ca(2+) -dependent activation of IK and SK channels. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

Dynamics of internal pore opening in KV channels probed by a fluorescent unnatural amino acid

PubMed Central

Kalstrup, Tanja; Blunck, Rikard

2013-01-01

Atomic-scale models on the gating mechanism of voltage-gated potassium channels (Kv) are based on linear interpolations between static structures of their initial and final state derived from crystallography and molecular dynamics simulations, and, thus, lack dynamic structural information. The lack of information on dynamics and intermediate states makes it difficult to associate the structural with the dynamic functional data obtained with electrophysiology. Although voltage-clamp fluorometry fills this gap, it is limited to sites extracellularly accessible, when the key region for gating is located at the cytosolic side of the channels. Here, we solved this problem by performing voltage-clamp fluorometry with a fluorescent unnatural amino acid. By using an orthogonal tRNA-synthetase pair, the fluorescent unnatural amino acid was incorporated in the Shaker voltage-gated potassium channel at key regions that were previously inaccessible. Thus, we defined which parts act independently and which parts act cooperatively and found pore opening to occur in two sequential transitions. PMID:23630265

Dendritic Kv3.3 potassium channels in cerebellar purkinje cells regulate generation and spatial dynamics of dendritic Ca2+ spikes.

PubMed

Zagha, Edward; Manita, Satoshi; Ross, William N; Rudy, Bernardo

2010-06-01

Purkinje cell dendrites are excitable structures with intrinsic and synaptic conductances contributing to the generation and propagation of electrical activity. Voltage-gated potassium channel subunit Kv3.3 is expressed in the distal dendrites of Purkinje cells. However, the functional relevance of this dendritic distribution is not understood. Moreover, mutations in Kv3.3 cause movement disorders in mice and cerebellar atrophy and ataxia in humans, emphasizing the importance of understanding the role of these channels. In this study, we explore functional implications of this dendritic channel expression and compare Purkinje cell dendritic excitability in wild-type and Kv3.3 knockout mice. We demonstrate enhanced excitability of Purkinje cell dendrites in Kv3.3 knockout mice, despite normal resting membrane properties. Combined data from local application pharmacology, voltage clamp analysis of ionic currents, and assessment of dendritic Ca(2+) spike threshold in Purkinje cells suggest a role for Kv3.3 channels in opposing Ca(2+) spike initiation. To study the physiological relevance of altered dendritic excitability, we measured [Ca(2+)](i) changes throughout the dendritic tree in response to climbing fiber activation. Ca(2+) signals were specifically enhanced in distal dendrites of Kv3.3 knockout Purkinje cells, suggesting a role for dendritic Kv3.3 channels in regulating propagation of electrical activity and Ca(2+) influx in distal dendrites. These findings characterize unique roles of Kv3.3 channels in dendrites, with implications for synaptic integration, plasticity, and human disease.

[Mechanisms of action of voltage-gated sodium channel ligands].

PubMed

Tikhonov, D B

2007-05-01

The voltage-gated sodium channels play a key role in the generation of action potential in excitable cells. Sodium channels are targeted by a number of modulating ligands. Despite numerous studies, the mechanisms of action of many ligands are still unknown. The main cause of the problem is the absence of the channel structure. Sodium channels belong to the superfamily of P-loop channels that also the data abowt includes potassium and calcium channels and the channels of ionotropic glutamate receptors. Crystallization of several potassium channels has opened a possibility to analyze the structure of other members of the superfamily using the homology modeling approach. The present study summarizes the results of several recent modelling studies of such sodium channel ligands as tetrodotoxin, batrachotoxin and local anesthetics. Comparison of available experimental data with X-ray structures of potassium channels has provided a new level of understanding of the mechanisms of action of sodium channel ligands and has allowed proposing several testable hypotheses.

New Molecular Targets for Antiepileptic Drugs: α2δ, SV2A, and Kv7/KCNQ/M Potassium Channels

PubMed Central

Rogawski, Michael A.; Bazil, Carl W.

2008-01-01

Many currently prescribed antiepileptic drugs (AEDs) act via voltage-gated sodium channels, through effects on γ-aminobutyric acid–mediated inhibition, or via voltage-gated calcium channels. Some newer AEDs do not act via these traditional mechanisms. The molecular targets for several of these nontraditional AEDs have been defined using cellular electrophysiology and molecular approaches. Here, we describe three of these targets: α2δ, auxiliary subunits of voltage-gated calcium channels through which the gabapentinoids gabapentin and pregabalin exert their anticonvulsant and analgesic actions; SV2A, a ubiquitous synaptic vesicle glycoprotein that may prepare vesicles for fusion and serves as the target for levetiracetam and its analog brivaracetam (which is currently in late-stage clinical development); and Kv7/KCNQ/M potassium channels that mediate the M-current, which acts a brake on repetitive firing and burst generation and serves as the target for the investigational AEDs retigabine and ICA-105665. Functionally, all of the new targets modulate neurotransmitter output at synapses, focusing attention on presynaptic terminals as critical sites of action for AEDs. PMID:18590620

New molecular targets for antiepileptic drugs: alpha(2)delta, SV2A, and K(v)7/KCNQ/M potassium channels.

PubMed

Rogawski, Michael A; Bazil, Carl W

2008-07-01

Many currently prescribed antiepileptic drugs (AEDs) act via voltage-gated sodium channels, through effects on gamma-aminobutyric acid-mediated inhibition, or via voltage-gated calcium channels. Some newer AEDs do not act via these traditional mechanisms. The molecular targets for several of these nontraditional AEDs have been defined using cellular electrophysiology and molecular approaches. Here, we describe three of these targets: alpha(2)delta, auxiliary subunits of voltage-gated calcium channels through which the gabapentinoids gabapentin and pregabalin exert their anticonvulsant and analgesic actions; SV2A, a ubiquitous synaptic vesicle glycoprotein that may prepare vesicles for fusion and serves as the target for levetiracetam and its analog brivaracetam (which is currently in late-stage clinical development); and K(v)7/KCNQ/M potassium channels that mediate the M-current, which acts a brake on repetitive firing and burst generation and serves as the target for the investigational AEDs retigabine and ICA-105665. Functionally, all of the new targets modulate neurotransmitter output at synapses, focusing attention on presynaptic terminals as critical sites of action for AEDs.

Involvement of a Gardos-type potassium channel in head activator-induced mitosis of BON cells.

PubMed

Kayser, S T; Ulrich, H; Schaller, H C

1998-06-01

The human neuroendocrine cell line BON was used to study second messengers involved in signal transduction for entry into mitosis. BON cells produce the neuropeptide head activator (HA) and use it as autocrine growth factor. HA stimulates BON cell proliferation by triggering entry into mitosis. HA-induced mitosis is mediated by an inhibitory G protein, the action of which is blocked by pertussis toxin. HA signaling requires inhibition of the cAMP pathway, calcium influx, and hyperpolarization of cells. The latter is a very important and sensitive step involving a calcium-activated potassium channel. Cell cycle progression and proliferation of BON cells are most efficiently inhibited with specific inhibitors of this potassium channel. Pharmacology and RNA analysis suggest identity with the recently cloned Gardos-type potassium channel.

Identification of a pharmacological target for genioglossus reactivation throughout sleep.

PubMed

Grace, Kevin P; Hughes, Stuart W; Horner, Richard L

2014-01-01

Obstructive sleep apnea (OSA) is a significant public health problem caused by repeated episodes of upper airway closure that occur only during sleep. Attempts to treat OSA pharmacologically have been unsuccessful because there has not been identification of a target operating at cranial motor nuclei, blockade of which can reactivate pharyngeal muscle activity throughout sleep. Increasing potassium conductance is a common mechanism by which state-dependent neuromodulators reduce motoneuron excitability. Therefore, we aimed to determine if potassium channel blockade is an effective strategy to reactivate the pharyngeal musculature throughout sleep. In rats chronically instrumented for recording sleep-wake states and respiratory motor activities, we locally microperfused pharmacological agents into the hypoglossal motor pool to modulate potassium channels of three major classes: inwardly rectifying, two-pore domain, and voltage-gated. Microperfusion of the inwardly rectifying potassium channel blocker, barium, as well as the voltage-gated potassium channel blockers, tetraethylammonium and 4-aminopyridine, increased tonic and respiratory-related genioglossus activities throughout nonrapid eye movement (non-REM) and rapid eye movement (REM) sleep to 133-300% of levels present during baseline wakefulness. In contrast, microperfusion of methanandamide (TWIK-related acid-sensitive potassium [TASK] channel blocker/cannabinoid receptor agonist) activated genioglossus in wakefulness but not in sleep. These findings establish proof-of-principle that targeted blockade of certain potassium channels at the hypoglossal motor pool is an effective strategy for reversing upper airway hypotonia and causing sustained reactivation of genioglossus throughout nonrapid eye movement and rapid eye movement sleep. These findings identify an important new direction for translational approaches to the pharmacological treatment of obstructive sleep apnea.

The temperature dependence of the BK channel activity - kinetics, thermodynamics, and long-range correlations.

PubMed

Wawrzkiewicz-Jałowiecka, Agata; Dworakowska, Beata; Grzywna, Zbigniew J

2017-10-01

Large-conductance, voltage dependent, Ca 2+ -activated potassium channels (BK) are transmembrane proteins that regulate many biological processes by controlling potassium flow across cell membranes. Here, we investigate to what extent temperature (in the range of 17-37°C with ΔT=5°C step) is a regulating parameter of kinetic properties of the channel gating and memory effect in the series of dwell-time series of subsequent channel's states, at membrane depolarization and hyperpolarization. The obtained results indicate that temperature affects strongly the BK channels' gating, but, counterintuitively, it exerts no effect on the long-range correlations, as measured by the Hurst coefficient. Quantitative differences between dependencies of appropriate channel's characteristics on temperature are evident for different regimes of voltage. Examining the characteristics of BK channel activity as a function of temperature allows to estimate the net activation energy (E act ) and changes of thermodynamic parameters (ΔH, ΔS, ΔG) by channel opening. Larger E act corresponds to the channel activity at membrane hyperpolarization. The analysis of entropy and enthalpy changes of closed to open channel's transition suggest the entropy-driven nature of the increase of open state probability during voltage activation and supports the hypothesis about the voltage-dependent geometry of the channel vestibule. Copyright © 2017 Elsevier B.V. All rights reserved.

Effect of urinary trypsin inhibitor on potassium currents: fetus modulates membrane excitability by production of UTI.

PubMed

Takeuchi, Kinya; Fukuda, Atsuo; Kanayama, Naohiro

2004-01-01

Amniotic fluid contains a significant level of urinary trypsin inhibitor (UTI). Previously, we reported that UTI inhibits calcium influx of myometrium and it is effective in preventing uterine contraction. This study examined the effects of UTI upon potassium channels, which is important for membrane excitability. Whole-cell patch-clamp recordings were performed in fibroblasts derived from human fetal skin. Potassium currents were recorded and the effects of exogenous UTI and/or cadmium determined. Tetraethylammonium sensitive potassium currents were elicited by step or ramp stimulations at depolarized membrane potentials (over +30 mV). Administration of 1 micro M UTI significantly increased these potassium currents by 16.9%. When calcium channels were blocked by the administration of cadmium, UTI increased the rest of the potassium currents by 4.8%. This indicates that UTI increased calcium-dependent potassium currents by 94.8% but only increased voltage-dependent potassium currents by 4.8%. Urinary trypsin inhibitor is a physiological substance of fetal origin that modulates calcium-dependent and voltage-dependent potassium channels. These data suggest that UTI is capable of regulating the membrane properties of the fetal and myometrial cells in contact with amniotic fluid.

Angiotensin II Inhibits the ROMK-like Small Conductance K Channel in Renal Cortical Collecting Duct during Dietary Potassium Restriction*

PubMed Central

Wei, Yuan; Zavilowitz, Beth; Satlin, Lisa M.; Wang, Wen-Hui

2010-01-01

Base-line urinary potassium secretion in the distal nephron is mediated by small conductance rat outer medullary K (ROMK)-like channels. We used the patch clamp technique applied to split-open cortical collecting ducts (CCDs) isolated from rats fed a normal potassium (NK) or low potassium (LK) diet to test the hypothesis that AngII directly inhibits ROMK channel activity. We found that AngII inhibited ROMK channel activity in LK but not NK rats in a dose-dependent manner. The AngII-induced reduction in channel activity was mediated by AT1 receptor (AT1R) binding, because pretreatment of CCDs with losartan but not PD123319 AT1 and AT2 receptor antagonists, respectively, blocked the response. Pretreatment of CCDs with U73122 and calphostin C, inhibitors of phospholipase C (PLC) and protein kinase C (PKC), respectively, abolished the AngII-induced decrease in ROMK channel activity, confirming a role of the PLC-PKC pathway in this response. Studies by others suggest that AngII stimulates an Src family protein-tyrosine kinase (PTK) via PKC-NADPH oxidase. PTK has been shown to regulate the ROMK channel. Inhibition of NADPH oxidase with diphenyliodonium abolished the inhibitory effect of AngII or the PKC activator phorbol 12-myristate 13-acetate on ROMK channels. Suppression of PTK by herbimycin A significantly attenuated the inhibitory effect of AngII on ROMK channel activity. We conclude that AngII inhibits ROMK channel activity through PKC-, NADPH oxidase-, and PTK-dependent pathways under conditions of dietary potassium restriction. PMID:17194699

Angiotensin II inhibits the ROMK-like small conductance K channel in renal cortical collecting duct during dietary potassium restriction.

PubMed

Wei, Yuan; Zavilowitz, Beth; Satlin, Lisa M; Wang, Wen-Hui

2007-03-02

Base-line urinary potassium secretion in the distal nephron is mediated by small conductance rat outer medullary K (ROMK)-like channels. We used the patch clamp technique applied to split-open cortical collecting ducts (CCDs) isolated from rats fed a normal potassium (NK) or low potassium (LK) diet to test the hypothesis that AngII directly inhibits ROMK channel activity. We found that AngII inhibited ROMK channel activity in LK but not NK rats in a dose-dependent manner. The AngII-induced reduction in channel activity was mediated by AT1 receptor (AT1R) binding, because pretreatment of CCDs with losartan but not PD123319 AT1 and AT2 receptor antagonists, respectively, blocked the response. Pretreatment of CCDs with U73122 and calphostin C, inhibitors of phospholipase C (PLC) and protein kinase C (PKC), respectively, abolished the AngII-induced decrease in ROMK channel activity, confirming a role of the PLC-PKC pathway in this response. Studies by others suggest that AngII stimulates an Src family protein-tyrosine kinase (PTK) via PKC-NADPH oxidase. PTK has been shown to regulate the ROMK channel. Inhibition of NADPH oxidase with diphenyliodonium abolished the inhibitory effect of AngII or the PKC activator phorbol 12-myristate 13-acetate on ROMK channels. Suppression of PTK by herbimycin A significantly attenuated the inhibitory effect of AngII on ROMK channel activity. We conclude that AngII inhibits ROMK channel activity through PKC-, NADPH oxidase-, and PTK-dependent pathways under conditions of dietary potassium restriction.

Depolarization of sperm membrane potential is a common feature of men with subfertility and is associated with low fertilization rate at IVF.

PubMed

Brown, Sean G; Publicover, Stephen J; Mansell, Steven A; Lishko, Polina V; Williams, Hannah L; Ramalingam, Mythili; Wilson, Stuart M; Barratt, Christopher L R; Sutton, Keith A; Da Silva, Sarah Martins

2016-06-01

Are significant abnormalities in outward (K(+)) conductance and resting membrane potential (Vm) present in the spermatozoa of patients undertaking IVF and ICSI and if so, what is their functional effect on fertilization success? Negligible outward conductance (≈5% of patients) or an enhanced inward conductance (≈4% of patients), both of which caused depolarization of Vm, were associated with a low rate of fertilization following IVF. Sperm-specific potassium channel knockout mice are infertile with defects in sperm function, suggesting that these channels are essential for fertility. These observations suggest that malfunction of K(+) channels in human spermatozoa might contribute significantly to the occurrence of subfertility in men. However, remarkably little is known of the nature of K(+) channels in human spermatozoa or the incidence and functional consequences of K(+) channel defects. Spermatozoa were obtained from healthy volunteer research donors and subfertile IVF and ICSI patients attending a hospital assisted reproductive techniques clinic between May 2013 and December 2015. In total, 40 IVF patients, 41 ICSI patients and 26 normozoospermic donors took part in the study. Samples were examined using electrophysiology (whole-cell patch clamping). Where abnormal electrophysiological characteristics were identified, spermatozoa were further examined for Ca(2+) influx induced by progesterone and penetration into viscous media if sufficient sample was available. Full exome sequencing was performed to specifically evaluate potassium calcium-activated channel subfamily M α 1 (KCNMA1), potassium calcium-activated channel subfamily U member 1 (KCNU1) and leucine-rich repeat containing 52 (LRRC52) genes and others associated with K(+) signalling. In IVF patients, comparison with fertilization rates was done to assess the functional significance of the electrophysiological abnormalities. Patch clamp electrophysiology was used to assess outward (K(+)) conductance and resting membrane potential (Vm) and signalling/motility assays were used to assess functional characteristics of sperm from IVF and ICSI patient samples. The mean Vm and outward membrane conductance in sperm from IVF and ICSI patients were not significantly different from those of control (donor) sperm prepared under the same conditions, but variation between individuals was significantly greater (P< 0.02) with a large number of outliers (>25%). In particular, in ≈10% of patients (7/81), we observed either a negligible outward conductance (4 patients) or an enhanced inward current (3 patients), both of which caused depolarization of Vm. Analysis of clinical data from the IVF patients showed significant association of depolarized Vm (≥0 mV) with low fertilization rate (P= 0.012). Spermatozoa with electrophysiological abnormities (conductance and Vm) responded normally to progesterone with elevation of [Ca(2+)]i and penetration of viscous medium, indicating retention of cation channel of sperm (CatSper) channel function. For practical, technical, ethical and logistical reasons, we could not obtain sufficient additional semen samples from men with conductance abnormalities to establish the cause of the conductance defects. Full exome sequencing was only available in two men with conductance defects. These data add significantly to the understanding of the role of ion channels in human sperm function and its impact on male fertility. Impaired potassium channel conductance (Gm) and/or Vm regulation is both common and complex in human spermatozoa and importantly is associated with impaired fertilization capacity when the Vm of cells is completely depolarized. The majority of the data were obtained using funding from MRC project grants (#MR/K013343/1, MR/012492/1). Additional funding was provided by NHS Tayside, TENOVUS, Chief Scientist Office NRS Fellowship and University of Abertay. The authors declare that there is no conflict of interest. Not applicable. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology.

Depolarization of sperm membrane potential is a common feature of men with subfertility and is associated with low fertilization rate at IVF

PubMed Central

Brown, Sean G.; Publicover, Stephen J.; Mansell, Steven A.; Lishko, Polina V.; Williams, Hannah L.; Ramalingam, Mythili; Wilson, Stuart M.; Barratt, Christopher L.R.; Sutton, Keith A.; Da Silva, Sarah Martins

2016-01-01

STUDY QUESTION Are significant abnormalities in outward (K+) conductance and resting membrane potential (Vm) present in the spermatozoa of patients undertaking IVF and ICSI and if so, what is their functional effect on fertilization success? SUMMARY ANSWER Negligible outward conductance (≈5% of patients) or an enhanced inward conductance (≈4% of patients), both of which caused depolarization of Vm, were associated with a low rate of fertilization following IVF. WHAT IS KNOWN ALREADY Sperm-specific potassium channel knockout mice are infertile with defects in sperm function, suggesting that these channels are essential for fertility. These observations suggest that malfunction of K+ channels in human spermatozoa might contribute significantly to the occurrence of subfertility in men. However, remarkably little is known of the nature of K+ channels in human spermatozoa or the incidence and functional consequences of K+ channel defects. STUDY DESIGN, SIZE AND DURATION Spermatozoa were obtained from healthy volunteer research donors and subfertile IVF and ICSI patients attending a hospital assisted reproductive techniques clinic between May 2013 and December 2015. In total, 40 IVF patients, 41 ICSI patients and 26 normozoospermic donors took part in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS Samples were examined using electrophysiology (whole-cell patch clamping). Where abnormal electrophysiological characteristics were identified, spermatozoa were further examined for Ca2+ influx induced by progesterone and penetration into viscous media if sufficient sample was available. Full exome sequencing was performed to specifically evaluate potassium calcium-activated channel subfamily M α 1 (KCNMA1), potassium calcium-activated channel subfamily U member 1 (KCNU1) and leucine-rich repeat containing 52 (LRRC52) genes and others associated with K+ signalling. In IVF patients, comparison with fertilization rates was done to assess the functional significance of the electrophysiological abnormalities. MAIN RESULTS AND THE ROLE OF CHANCE Patch clamp electrophysiology was used to assess outward (K+) conductance and resting membrane potential (Vm) and signalling/motility assays were used to assess functional characteristics of sperm from IVF and ICSI patient samples. The mean Vm and outward membrane conductance in sperm from IVF and ICSI patients were not significantly different from those of control (donor) sperm prepared under the same conditions, but variation between individuals was significantly greater (P< 0.02) with a large number of outliers (>25%). In particular, in ≈10% of patients (7/81), we observed either a negligible outward conductance (4 patients) or an enhanced inward current (3 patients), both of which caused depolarization of Vm. Analysis of clinical data from the IVF patients showed significant association of depolarized Vm (≥0 mV) with low fertilization rate (P= 0.012). Spermatozoa with electrophysiological abnormities (conductance and Vm) responded normally to progesterone with elevation of [Ca2+]i and penetration of viscous medium, indicating retention of cation channel of sperm (CatSper) channel function. LIMITATIONS, REASONS FOR CAUTION For practical, technical, ethical and logistical reasons, we could not obtain sufficient additional semen samples from men with conductance abnormalities to establish the cause of the conductance defects. Full exome sequencing was only available in two men with conductance defects. WIDER IMPLICATIONS OF THE FINDINGS These data add significantly to the understanding of the role of ion channels in human sperm function and its impact on male fertility. Impaired potassium channel conductance (Gm) and/or Vm regulation is both common and complex in human spermatozoa and importantly is associated with impaired fertilization capacity when the Vm of cells is completely depolarized. STUDY FUNDING/COMPETING INTEREST(S) The majority of the data were obtained using funding from MRC project grants (#MR/K013343/1, MR/012492/1). Additional funding was provided by NHS Tayside, TENOVUS, Chief Scientist Office NRS Fellowship and University of Abertay. The authors declare that there is no conflict of interest. TRIAL REGISTRATION NUMBER Not applicable. PMID:27052499

A Conserved Bicycle Model for Circadian Clock Control of Membrane Excitability

PubMed Central

Flourakis, Matthieu; Kula-Eversole, Elzbieta; Hutchison, Alan L.; Han, Tae Hee; Aranda, Kimberly; Moose, Devon L.; White, Kevin P.; Dinner, Aaron R.; Lear, Bridget C.; Ren, Dejian; Diekman, Casey O.; Raman, Indira M.; Allada, Ravi

2015-01-01

Summary Circadian clocks regulate membrane excitability in master pacemaker neurons to control daily rhythms of sleep and wake. Here we find that two distinctly timed electrical drives collaborate to impose rhythmicity on Drosophila clock neurons. In the morning, a voltage-independent sodium conductance via the NA/NALCN ion channel depolarizes these neurons. This current is driven by the rhythmic expression of NCA localization factor-1, linking the molecular clock to ion channel function. In the evening, basal potassium currents peak to silence clock neurons. Remarkably, daily antiphase cycles of sodium and potassium currents also drive mouse clock neuron rhythms. Thus, we reveal an evolutionarily ancient strategy for the neural mechanisms that govern daily sleep and wake. PMID:26276633

Regulation of potassium transport and signaling in plants.

PubMed

Wang, Yi; Wu, Wei-Hua

2017-10-01

As an essential macronutrient, potassium (K + ) plays crucial roles in diverse physiological processes during plant growth and development. The K + concentration in soils is relatively low and fluctuating. Plants are able to perceive external K + changes and generate chemical and physical signals in plant cells. The signals can be transducted across the plasma membrane and into the cytosol, and eventually regulates the downstream targets, particularly K + channels and transporters. As a result, K + homeostasis in plant cells is modulated, which facilitates plant adaptation to K + deficient conditions. This minireview focuses on the latest research progress in the diverse functions of K + channels and transporters as well as their regulatory mechanisms in plant response to low-K + stress. Copyright © 2017 Elsevier Ltd. All rights reserved.

Novel ion channel targets in atrial fibrillation.

PubMed

Hancox, Jules C; James, Andrew F; Marrion, Neil V; Zhang, Henggui; Thomas, Dierk

2016-08-01

Atrial fibrillation (AF) is the most common arrhythmia in humans. It is progressive and the development of electrical and structural remodeling makes early intervention desirable. Existing antiarrhythmic pharmacological approaches are not always effective and can produce unwanted side effects. Additional atrial-selective antiarrhythmic strategies are therefore desirable. Evidence for three novel ion channel atrial-selective therapeutic targets is evaluated: atrial-selective fast sodium channel current (INa) inhibition; small conductance calcium-activated potassium (SK) channels; and two-pore (K2P) potassium channels. Data from animal models support atrial-ventricular differences in INa kinetics and also suggest atrial-ventricular differences in sodium channel β subunit expression. Further work is required to determine whether intrinsic atrial-ventricular differences in human INa exist or whether functional differences occur due to distinct atrial and ventricular action and resting potentials. SK and K2P channels (particularly K2P 3.1) offer potentially attractive atrial-selective targets. Work is needed to identify the underlying basis of SK current that contributes to (patho)physiological atrial repolarization and settings in which SK inhibition is anti- versus pro-arrhythmic. Although K2P3.1 appears to be a promising target with comparatively selective drugs for experimental use, a lack of selective pharmacology hinders evaluation of other K2P channels as potential atrial-selective targets.

Disruption of ATP-sensitive potassium channel function in skeletal muscles promotes production and secretion of musclin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sierra, Ana, E-mail: ana-sierra@uiowa.edu; Subbotina, Ekaterina, E-mail: ekaterina-subbotina@uiowa.edu; Zhu, Zhiyong, E-mail: zhiyong-zhu@uiowa.edu

Sarcolemmal ATP-sensitive potassium (K{sub ATP}) channels control skeletal muscle energy use through their ability to adjust membrane excitability and related cell functions in accordance with cellular metabolic status. Mice with disrupted skeletal muscle K{sub ATP} channels exhibit reduced adipocyte size and increased fatty acid release into the circulation. As yet, the molecular mechanisms underlying this link between skeletal muscle K{sub ATP} channel function and adipose mobilization have not been established. Here, we demonstrate that skeletal muscle-specific disruption of K{sub ATP} channel function in transgenic (TG) mice promotes production and secretion of musclin. Musclin is a myokine with high homology tomore » atrial natriuretic peptide (ANP) that enhances ANP signaling by competing for elimination. Augmented musclin production in TG mice is driven by a molecular cascade resulting in enhanced acetylation and nuclear exclusion of the transcription factor forkhead box O1 (FOXO1) – an inhibitor of transcription of the musclin encoding gene. Musclin production/secretion in TG is paired with increased mobilization of fatty acids and a clear trend toward increased circulating ANP, an activator of lipolysis. These data establish K{sub ATP} channel-dependent musclin production as a potential mechanistic link coupling “local” skeletal muscle energy consumption with mobilization of bodily resources from fat. Understanding such mechanisms is an important step toward designing interventions to manage metabolic disorders including those related to excess body fat and associated co-morbidities. - Highlights: • ATP-sensitive K{sup +} channels regulate musclin production by skeletal muscles. • Lipolytic ANP signaling is promoted by augmented skeletal muscle musclin production. • Skeletal muscle musclin transcription is promoted by a CaMKII/HDAC/FOXO1 pathway. • Musclin links adipose mobilization to energy use in K{sub ATP} channel deficient skeletal muscle.« less

Regulation of voltage-gated potassium channels attenuates resistance of side-population cells to gefitinib in the human lung cancer cell line NCI-H460.

PubMed

Choi, Seon Young; Kim, Hang-Rae; Ryu, Pan Dong; Lee, So Yeong

2017-02-21

Side-population (SP) cells that exclude anti-cancer drugs have been found in various tumor cell lines. Moreover, SP cells have a higher proliferative potential and drug resistance than main population cells (Non-SP cells). Also, several ion channels are responsible for the drug resistance and proliferation of SP cells in cancer. To confirm the expression and function of voltage-gated potassium (Kv) channels of SP cells, these cells, as well as highly expressed ATP-binding cassette (ABC) transporters and stemness genes, were isolated from a gefitinib-resistant human lung adenocarcinoma cell line (NCI-H460), using Hoechst 33342 efflux. In the present study, we found that mRNA expression of Kv channels in SP cells was different compared to Non-SP cells, and the resistance of SP cells to gefitinib was weakened with a combination treatment of gefitinib and Kv channel blockers or a Kv7 opener, compared to single-treatment gefitinib, through inhibition of the Ras-Raf signaling pathway. The findings indicate that Kv channels in SP cells could be new targets for reducing the resistance to gefitinib.

Ion channels to inactivate neurons in Drosophila.

PubMed

Hodge, James J L

2009-01-01

Ion channels are the determinants of excitability; therefore, manipulation of their levels and properties provides an opportunity for the investigator to modulate neuronal and circuit function. There are a number of ways to suppress electrical activity in Drosophila neurons, for instance, over-expression of potassium channels (i.e. Shaker Kv1, Shaw Kv3, Kir2.1 and DORK) that are open at resting membrane potential. This will result in increased potassium efflux and membrane hyperpolarisation setting resting membrane potential below the threshold required to fire action potentials. Alternatively over-expression of other channels, pumps or co-transporters that result in a hyperpolarised membrane potential will also prevent firing. Lastly, neurons can be inactivated by, disrupting or reducing the level of functional voltage-gated sodium (Nav1 paralytic) or calcium (Cav2 cacophony) channels that mediate the depolarisation phase of action potentials. Similarly, strategies involving the opposite channel manipulation should allow net depolarisation and hyperexcitation in a given neuron. These changes in ion channel expression can be brought about by the versatile transgenic (i.e. Gal4/UAS based) systems available in Drosophila allowing fine temporal and spatial control of (channel) transgene expression. These systems are making it possible to electrically inactivate (or hyperexcite) any neuron or neural circuit in the fly brain, and much like an exquisite lesion experiment, potentially elucidate whatever interesting behaviour or phenotype each network mediates. These techniques are now being used in Drosophila to reprogram electrical activity of well-defined circuits and bring about robust and easily quantifiable changes in behaviour, allowing different models and hypotheses to be rapidly tested.

Chronic Inhibition of Renal Outer Medullary Potassium Channel Not Only Prevented but Also Reversed Development of Hypertension and End-Organ Damage in Dahl Salt-Sensitive Rats.

PubMed

Zhou, Xiaoyan; Forrest, Michael J; Sharif-Rodriguez, Wanda; Forrest, Gail; Szeto, Daphne; Urosevic-Price, Olga; Zhu, Yonghua; Stevenson, Andra S; Zhou, Yuchen; Stribling, Sloan; Dajee, Maya; Walsh, Shawn P; Pasternak, Alexander; Sullivan, Kathleen A

2017-02-01

The renal outer medullary potassium (ROMK) channel mediates potassium recycling and facilitates sodium reabsorption through the Na + /K + /2Cl - cotransporter in the loop of Henle and potassium secretion at the cortical collecting duct. Evidence from the phenotype of humans and rodents with functional ROMK deficiency supports the contention that selective ROMK inhibitors (ROMKi) will represent a novel diuretic with potential of therapeutic benefit for hypertension. ROMKi have recently been synthesized by Merck & Co, Inc. The present studies were designed to examine the effects of ROMKi B on systemic hemodynamics, renal function and structure, and vascular function in Dahl salt-sensitive rats. Four experimental groups-control, high-salt diet alone; ROMKi B 3 mg·kg - 1 ·d - 1 ; ROMKi B 10 mg·kg - 1 ·d - 1 ; and hydrochlorothiazide 25 mg·kg - 1 ·d - 1 -were included in prophylactic (from week 1 to week 9 on high-salt diet) and therapeutic studies (from week 5 to week 9 on high-salt diet), respectively. ROMKi B produced sustained blood pressure reduction and improved renal and vascular function and histological alterations induced by a high-salt diet. ROMKi B was superior to hydrochlorothiazide at reducing blood pressure. Furthermore, ROMKi B provided beneficial effects on both the plasma lipid profile and bone mineral density. Chronic ROMK inhibition not only prevented but also reversed the development of hypertension and end-organ damage in Dahl salt-sensitive rats. Our findings suggest a potential utility of ROMKi B as a novel antihypertensive agent, particularly for the treatment of the salt-sensitive hypertension patient population. © 2016 American Heart Association, Inc.

Neonatal Diabetes Caused by Mutations in Sulfonylurea Receptor 1: Interplay between Expression and Mg-Nucleotide Gating Defects of ATP-Sensitive Potassium Channels

PubMed Central

Zhou, Qing; Garin, Intza; Castaño, Luis; Argente, Jesús; Muñoz-Calvo, Ma. Teresa; Perez de Nanclares, Guiomar; Shyng, Show-Ling

2010-01-01

Context: ATP-sensitive potassium (KATP) channels regulate insulin secretion by coupling glucose metabolism to β-cell membrane potential. Gain-of-function mutations in the sulfonylurea receptor 1 (SUR1) or Kir6.2 channel subunit underlie neonatal diabetes. Objective: The objective of the study was to determine the mechanisms by which two SUR1 mutations, E208K and V324M, associated with transient neonatal diabetes affect KATP channel function. Design: E208K or V324M mutant SUR1 was coexpressed with Kir6.2 in COS cells, and expression and gating properties of the resulting channels were assessed biochemically and electrophysiologically. Results: Both E208K and V324M augment channel response to MgADP stimulation without altering sensitivity to ATP4− or sulfonylureas. Surprisingly, whereas E208K causes only a small increase in MgADP response consistent with the mild transient diabetes phenotype, V324M causes a severe activating gating defect. Unlike E208K, V324M also impairs channel expression at the cell surface, which is expected to dampen its functional impact on β-cells. When either mutation was combined with a mutation in the second nucleotide binding domain of SUR1 previously shown to abolish Mg-nucleotide response, the activating effect of E208K and V324M was also abolished. Moreover, combination of E208K and V324M results in channels with Mg-nucleotide sensitivity greater than that seen in individual mutations alone. Conclusion: The results demonstrate that E208K and V324M, located in distinct domains of SUR1, enhance transduction of Mg-nucleotide stimulation from the SUR1 nucleotide binding folds to Kir6.2. Furthermore, they suggest that diabetes severity is determined by interplay between effects of a mutation on channel expression and channel gating. PMID:20810569

Kir6.2 activation by sulfonylurea receptors: a different mechanism of action for SUR1 and SUR2A subunits via the same residues

PubMed Central

Principalli, Maria A; Dupuis, Julien P; Moreau, Christophe J; Vivaudou, Michel; Revilloud, Jean

2015-01-01

ATP-sensitive potassium channels (K-ATP channels) play a key role in adjusting the membrane potential to the metabolic state of cells. They result from the unique combination of two proteins: the sulfonylurea receptor (SUR), an ATP-binding cassette (ABC) protein, and the inward rectifier K+ channel Kir6.2. Both subunits associate to form a heterooctamer (4 SUR/4 Kir6.2). SUR modulates channel gating in response to the binding of nucleotides or drugs and Kir6.2 conducts potassium ions. The activity of K-ATP channels varies with their localization. In pancreatic β-cells, SUR1/Kir6.2 channels are partly active at rest while in cardiomyocytes SUR2A/Kir6.2 channels are mostly closed. This divergence of function could be related to differences in the interaction of SUR1 and SUR2A with Kir6.2. Three residues (E1305, I1310, L1313) located in the linker region between transmembrane domain 2 and nucleotide-binding domain 2 of SUR2A were previously found to be involved in the activation pathway linking binding of openers onto SUR2A and channel opening. To determine the role of the equivalent residues in the SUR1 isoform, we designed chimeras between SUR1 and the ABC transporter multidrug resistance-associated protein 1 (MRP1), and used patch clamp recordings on Xenopus oocytes to assess the functionality of SUR1/MRP1 chimeric K-ATP channels. Our results reveal that the same residues in SUR1 and SUR2A are involved in the functional association with Kir6.2, but they display unexpected side-chain specificities which could account for the contrasted properties of pancreatic and cardiac K-ATP channels. PMID:26416970

The large-conductance calcium-activated potassium channel holds the key to the conundrum of familial hypokalemic periodic paralysis

PubMed Central

Kim, Sung-Jo; Kang, Sun-Yang; Yi, Jin Woong; Kim, Seung-Min

2014-01-01

Purpose Familial hypokalemic periodic paralysis (HOKPP) is an autosomal dominant channelopathy characterized by episodic attacks of muscle weakness and hypokalemia. Mutations in the calcium channel gene, CACNA1S, or the sodium channel gene, SCN4A, have been found to be responsible for HOKPP; however, the mechanism that causes hypokalemia remains to be determined. The aim of this study was to improve the understanding of this mechanism by investigating the expression of calcium-activated potassium (KCa) channel genes in HOKPP patients. Methods We measured the intracellular calcium concentration with fura-2-acetoxymethyl ester in skeletal muscle cells of HOKPP patients and healthy individuals. We examined the mRNA and protein expression of KCa channel genes (KCNMA1, KCNN1, KCNN2, KCNN3, and KCNN4) in both cell types. Results Patient cells exhibited higher cytosolic calcium levels than normal cells. Quantitative reverse transcription polymerase chain reaction analysis showed that the mRNA levels of the KCa channel genes did not significantly differ between patient and normal cells. However, western blot analysis showed that protein levels of the KCNMA1 gene, which encodes KCa1.1 channels (also called big potassium channels), were significantly lower in the membrane fraction and higher in the cytosolic fraction of patient cells than normal cells. When patient cells were exposed to 50 mM potassium buffer, which was used to induce depolarization, the altered subcellular distribution of BK channels remained unchanged. Conclusion These findings suggest a novel mechanism for the development of hypokalemia and paralysis in HOKPP and demonstrate a connection between disease-associated mutations in calcium/sodium channels and pathogenic changes in nonmutant potassium channels. PMID:25379045

Molecular characterization of voltage-gated potassium channel (Kv) and its importance in functional dynamics in bull spermatozoa.

PubMed

Gupta, Rishi Kumar; Swain, Dilip Kumar; Singh, Vijay; Anand, Mukul; Choudhury, Soumen; Yadav, Sarvajeet; Saxena, Atul; Garg, Satish Kumar

2018-07-01

Present study was undertaken to characterize the voltage gated potassium channel (K v 1.1) in bull spermatozoa using sixty four ejaculates collected from four Hariana bulls. Functional characterization was undertaken using a selective blocker of Kv channel, 4-Aminopyridine (4-AP) while molecular presence of Kv on bull spermatozoa by immunoblotting and indirect immunofluorescence. Three sets of 100 μL diluted sperm samples namely-negative control (100 μL of sperm dilution medium (SDM) containing 10 × 10 6  cells), vehicle control (99 μL of SDM containing 10 × 10 6  cells, and DMSO- 1  μL) and 4-AP treatment group (99 μL of SDM containing 10 × 10 6  cells, and drug 1 μL 4-AP) were used in the study. Immunoblotting identified a single band of 56 kDa corresponding to Kv1.1 in Hariana bull spermatozoa. Immunolocalization showed the positive immunoreactivity at head, middle piece and principal piece of the spermatozoa for Kv 1.1. Blocking of Kv using 4-AP resulted in significant (p < 0.05) reduction in sperm progressive motility, per cent capacitated spermatozoa (B-pattern) and acrosome reacted (AR-pattern) spermatozoa, while significant (P < 0.05) increase in per cent swollen spermatozoa. Blocking of Kv channels resulted in significantly (P < 0.05) increased percentage of spermatozoa with lower mitochondrial transmembrane potential. Computer assisted semen analysis (CASA) of motion and kinematic parameters in 4-AP treated spermatozoa indicated reduction in sperm motion parameters like LIN, STR, VSL and VAP and higher ALH, VCL, and BCF indicating hyperactivity of spermatozoa. Based on our findings, it may be concluded that voltage-gated potassium channel (Kv) are present on bull spermatozoa and these are associated with functional dynamics of spermatozoa. However, based on our limited study, it is not possible to deduce that how these channels are associated with induction of hyperactivity. Therefore, further studies are warranted to unravel the mechanistic signaling pathways involved in Kv-mediated alterations in functional dynamics of spermatozoa. Copyright © 2018 Elsevier Inc. All rights reserved.

Kv2 Channel Regulation of Action Potential Repolarization and Firing Patterns in Superior Cervical Ganglion Neurons and Hippocampal CA1 Pyramidal Neurons

PubMed Central

Liu, Pin W.

2014-01-01

Kv2 family “delayed-rectifier” potassium channels are widely expressed in mammalian neurons. Kv2 channels activate relatively slowly and their contribution to action potential repolarization under physiological conditions has been unclear. We explored the function of Kv2 channels using a Kv2-selective blocker, Guangxitoxin-1E (GxTX-1E). Using acutely isolated neurons, mixed voltage-clamp and current-clamp experiments were done at 37°C to study the physiological kinetics of channel gating and action potentials. In both rat superior cervical ganglion (SCG) neurons and mouse hippocampal CA1 pyramidal neurons, 100 nm GxTX-1E produced near-saturating block of a component of current typically constituting ∼60–80% of the total delayed-rectifier current. GxTX-1E also reduced A-type potassium current (IA), but much more weakly. In SCG neurons, 100 nm GxTX-1E broadened spikes and voltage clamp experiments using action potential waveforms showed that Kv2 channels carry ∼55% of the total outward current during action potential repolarization despite activating relatively late in the spike. In CA1 neurons, 100 nm GxTX-1E broadened spikes evoked from −70 mV, but not −80 mV, likely reflecting a greater role of Kv2 when other potassium channels were partially inactivated at −70 mV. In both CA1 and SCG neurons, inhibition of Kv2 channels produced dramatic depolarization of interspike voltages during repetitive firing. In CA1 neurons and some SCG neurons, this was associated with increased initial firing frequency. In all neurons, inhibition of Kv2 channels depressed maintained firing because neurons entered depolarization block more readily. Therefore, Kv2 channels can either decrease or increase neuronal excitability depending on the time scale of excitation. PMID:24695716

Cardioprotective effect of diadenosine tetraphosphate (AP4A) preservation in hypothermic storage and its relation with mitochondrial ATP-sensitive potassium channels.

PubMed

Ahmet, I; Sawa, Y; Nishimura, M; Kitakaze, M; Matsuda, H

2000-01-15

The preconditioning effect of diadenosine tetraphosphate (AP4A) was reported in ischemia/reperfused hearts, but its effect in heart preservation was unknown. According to the possible role of mitochondrial ATP-sensitive potassium channel (mK(ATP) channel) in the effect of ischemic preconditioning, the contribution of mK(ATP) channel to the effect of AP4A was tested. Isolated rat hearts were arrested and preserved by Eurocollin's (EC) solution at 4 degrees C for 8 hr. AP4A (80 microM) or AP4A with the 5-hydroxydecanoic acid (100 microM), a selective inhibitor of the mK(ATP) channel, was added into the EC solution. The preischemic and postischemic cardiac functions were evaluated on a buffer-perfused Langendorff apparatus before storage and after 20 min of reperfusion. AP4A administration improved the recovery of poststorage cardiac functions (the rate-pressure production, left ventricular systolic pressure, heart rate, coronary flow rate, and derivative of left ventricular systolic pressure; P<0.05) and reduced the leakage of lactate dehydrate and creatine kinase during reperfusion, compared with EC alone. Those effects of AP4A were completely reversed by 5-hydroxydecanoic acid administration in combination subjects. AP4A administration protects the heart through opening of the mK(ATP) channel during hypothermic preservation. Thus, addition of AP4A into cardioplegia may be a novel method of ischemic preconditioning in the transplantation context.

Functional characterization of a prokaryotic Kir channel.

PubMed

Enkvetchakul, Decha; Bhattacharyya, Jaya; Jeliazkova, Iana; Groesbeck, Darcy K; Cukras, Catherine A; Nichols, Colin G

2004-11-05

The Kir gene family encodes inward rectifying K+ (Kir) channels that are widespread and critical regulators of excitability in eukaryotic cells. A related gene family (KirBac) has recently been identified in prokaryotes. While a crystal structure of one member, Kir-Bac1.1, has been solved, there has been no functional characterization of any KirBac gene products. Here we present functional characterization of KirBac1.1 reconstituted in liposomes. Utilizing a 86Rb+ uptake assay, we demonstrate that KirBac1.1 generates a K+ -selective permeation path that is inhibited by extraliposomal Ba2+ and Ca2+ ions. In contrast to KcsA (an acid-activated bacterial potassium channel), KirBac1.1 is inhibited by extraliposomal acid (pKa approximately 6). This characterization of KirBac1.1 activity now paves the way for further correlation of structure and function in this model Kir channel.

SLO-2 potassium channel is an important regulator of neurotransmitter release in Caenorhabditis elegans

PubMed Central

Liu, Ping; Chen, Bojun; Wang, Zhao-Wen

2014-01-01

Slo2 channels are prominent K+ channels in mammalian neurons but their physiological functions are not well understood. Here we investigate physiological functions and regulation of the C. elegans homologue SLO-2 in motor neurons through electrophysiological analyses of wild-type and mutant worms. We find that SLO-2 is the primary K+ channel conducting delayed outward current in cholinergic motor neurons, and one of two K+ channels with this function in GABAergic motor neurons. Loss-of-function mutation of slo-2 increases the duration and charge transfer rate of spontaneous postsynaptic current bursts at the neuromuscular junction, which are physiological signals used by motor neurons to control muscle cells, without altering postsynaptic receptor sensitivity. SLO-2 activity in motor neurons depends on Ca2+ entry through EGL-19, an L-type voltage-gated Ca2+ channel (CaV1), but not on other proteins implicated in either Ca2+ entry or intracellular Ca2+ release. Thus, SLO-2 is functionally coupled with CaV1 and regulates neurotransmitter release. PMID:25300429

Tuning the allosteric regulation of artificial muscarinic and dopaminergic ligand-gated potassium channels by protein engineering of G protein-coupled receptors

PubMed Central

Moreau, Christophe J.; Revilloud, Jean; Caro, Lydia N.; Dupuis, Julien P.; Trouchet, Amandine; Estrada-Mondragón, Argel; Nieścierowicz, Katarzyna; Sapay, Nicolas; Crouzy, Serge; Vivaudou, Michel

2017-01-01

Ligand-gated ion channels enable intercellular transmission of action potential through synapses by transducing biochemical messengers into electrical signal. We designed artificial ligand-gated ion channels by coupling G protein-coupled receptors to the Kir6.2 potassium channel. These artificial channels called ion channel-coupled receptors offer complementary properties to natural channels by extending the repertoire of ligands to those recognized by the fused receptors, by generating more sustained signals and by conferring potassium selectivity. The first artificial channels based on the muscarinic M2 and the dopaminergic D2L receptors were opened and closed by acetylcholine and dopamine, respectively. We find here that this opposite regulation of the gating is linked to the length of the receptor C-termini, and that C-terminus engineering can precisely control the extent and direction of ligand gating. These findings establish the design rules to produce customized ligand-gated channels for synthetic biology applications. PMID:28145461

The changes of potassium currents in RCS rat Müller cell during retinal degeneration.

PubMed

Zhao, TongTao; Li, YaoChen; Weng, ChuanHuang; Yin, ZhengQin

2012-01-03

Müller cells are the principal glial cells expressing membrane-bound potassium channel and predominantly mediating the homeostatic regulation of extracellular K+ produced by neuronal activity in retina. It's well known that Müller cells can be activated in many pathological conditions, but little is known about the change of potassium currents of Müller cells during the progression of retinitis pigmentosa. Herein, the Royal College of Surgeons rats (RCS rat) were employed to investigate some phenotypic and functional changes of Müller cells during retinal degeneration such as the expression of Kir4.1, membrane properties and K+ channel currents by using immunohistochemistry, RT-PCR, western blot and whole-cell patch clamping respectively. Compared with Müller cells in control retina, increased glutamine synthetase (GS) mRNA levels were seen at P30 and P60, and then decreased gradually in RCS rat retina. Morphologically, Müller cells showed significant hypertrophy and proliferation after p60. The increased expression of intermediate filament, glial fibrillary acidic protein (GFAP) and vimentin began at P30 and reached a peak at p60. Kir4.1 channels presented a peak expression at P30. Concomitantly, K(+) currents of Müller cells increased at P30 and decreased at P90 significantly. We concluded that retinal Müller cells of RCS rats underwent an activation initiated by the onset of retinal degeneration before p60 and then an obvious reactive gliosis, which led the basic membrane properties to suffer marked changes, and caused the Kir4.1 channels of Müller cells to occur a clear functional shift, even lose their normal electrophysiological properties. This process aggravates the impairment caused by the initial photoreceptor degeneration. Copyright © 2011 Elsevier B.V. All rights reserved.

Prostatic relaxation induced by agmatine is decreased in spontaneously hypertensive rats.

PubMed

Lee, Liang-Ming; Tsai, Tsung-Chin; Chung, Hsien-Hui; Tong, Yat-Ching; Cheng, Juei-Tang

2012-09-01

What's known on the subject? and What does the study add? Neurotransmitters are known to control prostate contractility. Agmatine is one of them and induces relaxation through imidazoline receptors. The paper shows that the action of agmatine is reduced in hypertensive rats, and that this change is related to the decrease of ATP-sensitive potassium channels in the prostate. The findings can increase our understanding of the possible underlying mechanism for the development of clinical benign prostatic hyperplasia. To compare agmatine-induced prostatic relaxation in hypertensive and control rats. To investigate the responsible mechanism(s) and the role of the ATP-sensitive potassium channel. Prostate strips were isolated from male spontaneously hypertensive (SH) rats and normal Wistar-Kyoto (WKY) rats for measurement of isometric tension. The strips were precontracted with 1 µmol/L phenylephrine or 50 mmol/L KCl. Dose-dependent relaxation of the prostatic strips was studied by cumulative administration of agmatine, 1 to 100 µmol/L, into the organ bath. Effects of specific antagonists on agmatine-induced relaxation were studied. Western blotting analysis was used to measure the gene expression of the ATP-sensitive potassium channel in the rat prostate. Prostatic relaxation induced by agmatine was markedly reduced in SH rats compared with WKY rats. The relaxation caused by agmatine was abolished by BU224, a selective imidazoline I(2)-receptor antagonist, but was not modified by efaroxan at a dose sufficient to block imidazoline I(1)-receptors. The relaxation induced by diazoxide at a concentration sufficient to activate ATP-sensitive potassium channels was markedly reduced in the SH rat prostate. Expressions of ATP-sensitive potassium channel sulphonylurea receptor and inwardly rectifying potassium channel (Kir) 6.2 subunits were both decreased in the prostate of SH rats. The decrease of agmatine-induced prostatic relaxation in SH rats is related to the change in ATP-sensitive potassium channels. © 2012 THE AUTHORS. BJU INTERNATIONAL © 2012 BJU INTERNATIONAL.

Extracellular Potassium Homeostasis: Insights from Hypokalemic Periodic Paralysis

PubMed Central

Cheng, Chih-Jen; Kuo, Elizabeth; Huang, Chou-Long

2014-01-01

The extracellular potassium makes up only about 2% of the total body potassium store. The majority of the body potassium is distributed in the intracellular space, and of which about 80% is in skeletal muscle. Movement of potassium in and out of skeletal muscle thus plays a pivotal role in extracellular potassium homeostasis. The exchange of potassium between the extracellular space and skeletal muscle is mediated by specific membrane transporters. These include potassium uptake by Na+, K+-ATPase and release by inward rectifier K+ channels. These processes are regulated by circulating hormones, peptides, ions, and by physical activity of muscle as well as dietary potassium intake. Pharmaceutical agents, poisons and disease conditions also affect the exchange and alter extracellular potassium concentration. Here, we review extracellular potassium homeostasis focusing on factors and conditions that influence the balance of potassium movement in skeletal muscle. Recent findings that mutations of a skeletal muscle-specific inward rectifier K+ channel cause hypokalemic periodic paralysis provide interesting insights into the role of skeletal muscle in extracellular potassium homeostasis. These recent findings will be reviewed. PMID:23953801

C-Terminal residues in small potassium channel blockers OdK1 and OSK3 from scorpion venom fine-tune the selectivity.

PubMed

Kuzmenkov, Alexey I; Peigneur, Steve; Chugunov, Anton O; Tabakmakher, Valentin M; Efremov, Roman G; Tytgat, Jan; Grishin, Eugene V; Vassilevski, Alexander A

2017-05-01

We report isolation, sequencing, and electrophysiological characterization of OSK3 (α-KTx 8.8 in Kalium and Uniprot databases), a potassium channel blocker from the scorpion Orthochirus scrobiculosus venom. Using the voltage clamp technique, OSK3 was tested on a wide panel of 11 voltage-gated potassium channels expressed in Xenopus oocytes, and was found to potently inhibit Kv1.2 and Kv1.3 with IC 50 values of ~331nM and ~503nM, respectively. OdK1 produced by the scorpion Odontobuthus doriae differs by just two C-terminal residues from OSK3, but shows marked preference to Kv1.2. Based on the charybdotoxin-potassium channel complex crystal structure, a model was built to explain the role of the variable residues in OdK1 and OSK3 selectivity. Copyright © 2017 Elsevier B.V. All rights reserved.

The episodic ataxia type 1 mutation I262T alters voltage-dependent gating and disrupts protein biosynthesis of human Kv1.1 potassium channels.

PubMed

Chen, Szu-Han; Fu, Ssu-Ju; Huang, Jing-Jia; Tang, Chih-Yung

2016-01-18

Voltage-gated potassium (Kv) channels are essential for setting neuronal membrane excitability. Mutations in human Kv1.1 channels are linked to episodic ataxia type 1 (EA1). The EA1-associated mutation I262T was identified from a patient with atypical phenotypes. Although a previous report has characterized its suppression effect, several key questions regarding the impact of the I262T mutation on Kv1.1 as well as other members of the Kv1 subfamily remain unanswered. Herein we show that the dominant-negative effect of I262T on Kv1.1 current expression is not reversed by co-expression with Kvβ1.1 or Kvβ2 subunits. Biochemical examinations indicate that I262T displays enhanced protein degradation and impedes membrane trafficking of Kv1.1 wild-type subunits. I262T appears to be the first EA1 mutation directly associated with impaired protein stability. Further functional analyses demonstrate that I262T changes the voltage-dependent activation and Kvβ1.1-mediated inactivation, uncouples inactivation from activation gating, and decelerates the kinetics of cumulative inactivation of Kv1.1 channels. I262T also exerts similar dominant effects on the gating of Kv1.2 and Kv1.4 channels. Together our data suggest that I262T confers altered channel gating and reduced functional expression of Kv1 channels, which may account for some of the phenotypes of the EA1 patient.

The amiodarone derivative KB130015 activates hERG1 potassium channels via a novel mechanism

PubMed Central

Gessner, Guido; Macianskiene, Regina; Starkus, John G.; Schönherr, Roland; Heinemann, Stefan H.

2010-01-01

Human ether à go-go related gene (hERG1) potassium channels underlie the repolarizing IKr current in the heart. Since they are targets of various drugs with cardiac side effects we tested whether the amiodarone derivative 2-methyl-3-(3,5-diiodo-4-carboxymethoxybenzyl)benzofuran (KB130015) blocks hERG1 channels like its parent compound. Using patch-clamp and two-electrode voltage-clamp techniques we found that KB130015 blocks native and recombinant hERG1 channels at high voltages, but it activates them at low voltages. The activating effect has an apparent EC50 value of 12 μM and is brought about by an about 4-fold acceleration of activation kinetics and a shift in voltage-dependent activation by −16 mV. Channel activation was not use-dependent and was independent of inactivation gating. KB130015 presumably binds to the hERG1 pore from the cytosolic side and functionally competes with hERG1 block by amiodarone, E4031 (N-[4-[[1-[2-(6-methyl-2-pyridinyl)ethyl] -4-piperidinyl] carbonyl] phenyl] methanesulfonamide dihydrochloride), and sertindole. Vice versa, amiodarone attenuates hERG1 activation by KB130015. Based on synergic channel activation by mallotoxin and KB130015 we conclude that the hERG1 pore contains at least two sites for activators that are functionally coupled among each other and to the cavity-blocker site. KB130015 and amiodarone may serve as lead structures for the identification of hERG1 pore-interacting drugs favoring channel activation vs. block. PMID:20097192

Robustness, Death of Spiral Wave in the Network of Neurons under Partial Ion Channel Block

NASA Astrophysics Data System (ADS)

Ma, Jun; Huang, Long; Wang, Chun-Ni; Pu, Zhong-Sheng

2013-02-01

The development of spiral wave in a two-dimensional square array due to partial ion channel block (Potassium, Sodium) is investigated, the dynamics of the node is described by Hodgkin—Huxley neuron and these neurons are coupled with nearest neighbor connection. The parameter ratio xNa (and xK), which defines the ratio of working ion channel number of sodium (potassium) to the total ion channel number of sodium (and potassium), is used to measure the shift conductance induced by channel block. The distribution of statistical variable R in the two-parameter phase space (parameter ratio vs. poisoning area) is extensively calculated to mark the parameter region for transition of spiral wave induced by partial ion channel block, the area with smaller factors of synchronization R is associated the parameter region that spiral wave keeps alive and robust to the channel poisoning. Spiral wave keeps alive when the poisoned area (potassium or sodium) and degree of intoxication are small, distinct transition (death, several spiral waves coexist or multi-arm spiral wave emergence) occurs under moderate ratio xNa (and xK) when the size of blocked area exceeds certain thresholds. Breakup of spiral wave occurs and multi-arm of spiral waves are observed when the channel noise is considered.

Kcnh1 Voltage-gated Potassium Channels Are Essential for Early Zebrafish Development*

PubMed Central

Stengel, Rayk; Rivera-Milla, Eric; Sahoo, Nirakar; Ebert, Christina; Bollig, Frank; Heinemann, Stefan H.; Schönherr, Roland; Englert, Christoph

2012-01-01

The Kcnh1 gene encodes a voltage-gated potassium channel highly expressed in neurons and involved in tumor cell proliferation, yet its physiological roles remain unclear. We have used the zebrafish as a model to analyze Kcnh1 function in vitro and in vivo. We found that the kcnh1 gene is duplicated in teleost fish (i.e. kcnh1a and kcnh1b) and that both genes are maternally expressed during early development. In adult zebrafish, kcnh1a and kcnh1b have distinct expression patterns but share expression in brain and testis. Heterologous expression of both genes in Xenopus oocytes revealed a strong conservation of characteristic functional properties between human and fish channels, including a unique sensitivity to intracellular Ca2+/calmodulin and modulation of voltage-dependent gating by extracellular Mg2+. Using a morpholino antisense approach, we demonstrate a strong kcnh1 loss-of-function phenotype in developing zebrafish, characterized by growth retardation, delayed hindbrain formation, and embryonic lethality. This late phenotype was preceded by transcriptional up-regulation of known cell-cycle inhibitors (p21, p27, cdh2) and down-regulation of pro-proliferative factors, including cyclin D1, at 70% epiboly. These results reveal an unanticipated basic activity of kcnh1 that is crucial for early embryonic development and patterning. PMID:22927438

Clinical Aspects of Type-1 Long-QT Syndrome by Location, Coding Type, and Biophysical Function of Mutations Involving the KCNQ1 Gene

PubMed Central

Moss, Arthur J.; Shimizu, Wataru; Wilde, Arthur A.M.; Towbin, Jeffrey A.; Zareba, Wojciech; Robinson, Jennifer L.; Qi, Ming; Vincent, G. Michael; Ackerman, Michael J.; Kaufman, Elizabeth S.; Hofman, Nynke; Seth, Rahul; Kamakura, Shiro; Miyamoto, Yoshihiro; Goldenberg, Ilan; Andrews, Mark L.; McNitt, Scott

2012-01-01

Background Type-1 long-QT syndrome (LQTS) is caused by loss-of-function mutations in the KCNQ1-encoded IKs cardiac potassium channel. We evaluated the effect of location, coding type, and biophysical function of KCNQ1 mutations on the clinical phenotype of this disorder. Methods and Results We investigated the clinical course in 600 patients with 77 different KCNQ1 mutations in 101 proband-identified families derived from the US portion of the International LQTS Registry (n=425), the Netherlands’ LQTS Registry (n=93), and the Japanese LQTS Registry (n=82). The Cox proportional hazards survivorship model was used to evaluate the independent contribution of clinical and genetic factors to the first occurrence of time-dependent cardiac events from birth through age 40 years. The clinical characteristics, distribution of mutations, and overall outcome event rates were similar in patients enrolled from the 3 geographic regions. Biophysical function of the mutations was categorized according to dominant-negative (>50%) or haploinsufficiency (≤50%) reduction in cardiac repolarizing IKs potassium channel current. Patients with transmembrane versus C-terminus mutations (hazard ratio, 2.06; P<0.001) and those with mutations having dominant-negative versus haploinsufficiency ion channel effects (hazard ratio, 2.26; P<0.001) were at increased risk for cardiac events, and these genetic risks were independent of traditional clinical risk factors. Conclusions This genotype–phenotype study indicates that in type-1 LQTS, mutations located in the transmembrane portion of the ion channel protein and the degree of ion channel dysfunction caused by the mutations are important independent risk factors influencing the clinical course of this disorder. PMID:17470695

Alternative splice isoforms of small conductance calcium-activated SK2 channels differ in molecular interactions and surface levels

PubMed Central

Scholl, Elizabeth Storer; Pirone, Antonella; Cox, Daniel H; Duncan, R Keith; Jacob, Michele H

2014-01-01

Small conductance Ca2+-sensitive potassium (SK2) channels are voltage-independent, Ca2+-activated ion channels that conduct potassium cations and thereby modulate the intrinsic excitability and synaptic transmission of neurons and sensory hair cells. In the cochlea, SK2 channels are functionally coupled to the highly Ca2+ permeant α9/10-nicotinic acetylcholine receptors (nAChRs) at olivocochlear postsynaptic sites. SK2 activation leads to outer hair cell hyperpolarization and frequency-selective suppression of afferent sound transmission. These inhibitory responses are essential for normal regulation of sound sensitivity, frequency selectivity, and suppression of background noise. However, little is known about the molecular interactions of these key functional channels. Here we show that SK2 channels co-precipitate with α9/10-nAChRs and with the actin-binding protein α-actinin-1. SK2 alternative splicing, resulting in a 3 amino acid insertion in the intracellular 3′ terminus, modulates these interactions. Further, relative abundance of the SK2 splice variants changes during developmental stages of synapse maturation in both the avian cochlea and the mammalian forebrain. Using heterologous cell expression to separately study the 2 distinct isoforms, we show that the variants differ in protein interactions and surface expression levels, and that Ca2+ and Ca2+-bound calmodulin differentially regulate their protein interactions. Our findings suggest that the SK2 isoforms may be distinctly modulated by activity-induced Ca2+ influx. Alternative splicing of SK2 may serve as a novel mechanism to differentially regulate the maturation and function of olivocochlear and neuronal synapses. PMID:24394769

Clofilium inhibits Slick and Slack potassium channels.

PubMed

de Los Angeles Tejada, Maria; Stolpe, Kathleen; Meinild, Anne-Kristine; Klaerke, Dan A

2012-01-01

Slick and Slack high-conductance potassium channels have been recently discovered, and are found in the central nervous system and in the heart. Both channels are activated by Na(+) and Cl(-), and Slick channels are also inhibited by adenosine triphospate (ATP). An important role of setting the resting membrane potential and controlling the basal excitability of neurons has been suggested for these channels. In addition, no specific blockers for these channels are known up to the present. With the purpose of studying the pharmacological characteristics of Slick and Slack channels, the effects of exposure to the antiarrhythmic compound clofilium were evaluated. Clofilium was able to modulate the activity of Slick and Slack channels effectively, with a stronger effect on Slack than Slick channels. In order to evaluate the pharmacological behavior of Slick and Slack channels further, 38 commonly used potassium channel blockers were tested. Screening of these compounds did not reveal any modulators of Slick and Slack channels, except for clofilium. The present study provides a first approach towards elucidating the pharmacological characteristics of Slick and Slack channels and could be the basis for future studies aimed at developing potent and specific blockers and activators for these channels.

[Lead compound optimization strategy(5) – reducing the hERG cardiac toxicity in drug development].

PubMed

Zhou, Sheng-bin; Wang, Jiang; Liu, Hong

2016-10-01

The potassium channel encoded by the human ether-a-go-go related gene(hERG) plays a very important role in the physiological and pathological processes in human. hERG potassium channel determines the outward currents which facilitate the repolarization of the myocardial cells. Some drugs were withdrawn from the market for the serious side effect of long QT interval and arrhythmia due to blockade of hERG channel. The strategies for lead compound optimization are to reduce inhibitory activity of hERG potassium channel and decrease cardiac toxicity. These methods include reduction of lipophilicity and basicity of amines, introduction of hydroxyl and acidic groups, and restricting conformation.

Potassium dependent rescue of a myopathy with core-like structures in mouse

PubMed Central

Hanson, M Gartz; Wilde, Jonathan J; Moreno, Rosa L; Minic, Angela D; Niswander, Lee

2015-01-01

Myopathies decrease muscle functionality. Mutations in ryanodine receptor 1 (RyR1) are often associated with myopathies with microscopic core-like structures in the muscle fiber. In this study, we identify a mouse RyR1 model in which heterozygous animals display clinical and pathological hallmarks of myopathy with core-like structures. The RyR1 mutation decreases sensitivity to activated calcium release and myoplasmic calcium levels, subsequently affecting mitochondrial calcium and ATP production. Mutant muscle shows a persistent potassium leak and disrupted expression of regulators of potassium homeostasis. Inhibition of KATP channels or increasing interstitial potassium by diet or FDA-approved drugs can reverse the muscle weakness, fatigue-like physiology and pathology. We identify regulators of potassium homeostasis as biomarkers of disease that may reveal therapeutic targets in human patients with myopathy of central core disease (CCD). Altogether, our results suggest that amelioration of potassium leaks through potassium homeostasis mechanisms may minimize muscle damage of myopathies due to certain RyR1 mutations. DOI: http://dx.doi.org/10.7554/eLife.02923.001 PMID:25564733

Incorporating Born solvation energy into the three-dimensional Poisson-Nernst-Planck model to study ion selectivity in KcsA K+ channels

NASA Astrophysics Data System (ADS)

Liu, Xuejiao; Lu, Benzhuo

2017-12-01

Potassium channels are much more permeable to potassium than sodium ions, although potassium ions are larger and both carry the same positive charge. This puzzle cannot be solved based on the traditional Poisson-Nernst-Planck (PNP) theory of electrodiffusion because the PNP model treats all ions as point charges, does not incorporate ion size information, and therefore cannot discriminate potassium from sodium ions. The PNP model can qualitatively capture some macroscopic properties of certain channel systems such as current-voltage characteristics, conductance rectification, and inverse membrane potential. However, the traditional PNP model is a continuum mean-field model and has no or underestimates the discrete ion effects, in particular the ion solvation or self-energy (which can be described by Born model). It is known that the dehydration effect (closely related to ion size) is crucial to selective permeation in potassium channels. Therefore, we incorporated Born solvation energy into the PNP model to account for ion hydration and dehydration effects when passing through inhomogeneous dielectric channel environments. A variational approach was adopted to derive a Born-energy-modified PNP (BPNP) model. The model was applied to study a cylindrical nanopore and a realistic KcsA channel, and three-dimensional finite element simulations were performed. The BPNP model can distinguish different ion species by ion radius and predict selectivity for K+ over Na+ in KcsA channels. Furthermore, ion current rectification in the KcsA channel was observed by both the PNP and BPNP models. The I -V curve of the BPNP model for the KcsA channel indicated an inward rectifier effect for K+ (rectification ratio of ˜3 /2 ) but indicated an outward rectifier effect for Na+ (rectification ratio of ˜1 /6 ) .

Accumulation of K+ in the synaptic cleft modulates activity by influencing both vestibular hair cell and calyx afferent in the turtle

PubMed Central

Contini, Donatella; Price, Steven D.

2016-01-01

Key points In the synaptic cleft between type I hair cells and calyceal afferents, K+ ions accumulate as a function of activity, dynamically altering the driving force and permeation through ion channels facing the synaptic cleft.High‐fidelity synaptic transmission is possible due to large conductances that minimize hair cell and afferent time constants in the presence of significant membrane capacitance.Elevated potassium maintains hair cells near a potential where transduction currents are sufficient to depolarize them to voltages necessary for calcium influx and synaptic vesicle fusion.Elevated potassium depolarizes the postsynaptic afferent by altering ion permeation through hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels, and contributes to depolarizing the afferent to potentials where a single EPSP (quantum) can generate an action potential.With increased stimulation, hair cell depolarization increases the frequency of quanta released, elevates [K+]cleft and depolarizes the afferent to potentials at which smaller and smaller EPSPs would be sufficient to trigger APs. Abstract Fast neurotransmitters act in conjunction with slower modulatory effectors that accumulate in restricted synaptic spaces found at giant synapses such as the calyceal endings in the auditory and vestibular systems. Here, we used dual patch‐clamp recordings from turtle vestibular hair cells and their afferent neurons to show that potassium ions accumulating in the synaptic cleft modulated membrane potentials and extended the range of information transfer. High‐fidelity synaptic transmission was possible due to large conductances that minimized hair cell and afferent time constants in the presence of significant membrane capacitance. Increased potassium concentration in the cleft maintained the hair cell near potentials that promoted the influx of calcium necessary for synaptic vesicle fusion. The elevated potassium concentration also depolarized the postsynaptic neuron by altering ion permeation through hyperpolarization‐activated cyclic nucleotide‐gated (HCN) channels. This depolarization enabled the afferent to reliably generate action potentials evoked by single AMPA‐dependent EPSPs. Depolarization of the postsynaptic afferent could also elevate potassium in the synaptic cleft, and would depolarize other hair cells enveloped by the same neuritic process increasing the fidelity of neurotransmission at those synapses as well. Collectively, these data demonstrate that neuronal activity gives rise to potassium accumulation, and suggest that potassium ion action on HCN channels can modulate neurotransmission, preserving the fidelity of high‐speed synaptic transmission by dynamically shifting the resting potentials of both presynaptic and postsynaptic cells. PMID:27633787

A Disease Mutation Causing Episodic Ataxia Type I in the S1 Links Directly to the Voltage Sensor and the Selectivity Filter in Kv Channels.

PubMed

Petitjean, Dimitri; Kalstrup, Tanja; Zhao, Juan; Blunck, Rikard

2015-09-02

The mutation F184C in Kv1.1 leads to development of episodic ataxia type I (EA1). Although the mutation has been said to alter activation kinetics and to lower expression, we show here that the underlying molecular mechanisms may be more complex. Although F184 is positioned in the "peripheral" S1 helix, it occupies a central position in the 3D fold. We show in cut-open oocyte voltage-clamp recordings of gating and ionic currents of the Shaker Kv channel expressed in Xenopus oocytes that F184 not only interacts directly with the gating charges of the S4, but also creates a functional link to the selectivity filter of the neighboring subunit. This link leads to impaired fast and slow inactivation. The effect on fast inactivation is of an allosteric nature considering that fast inactivation is caused by a linked cytosolic ball peptide. The extensive effects of F184C provide a new mechanism underlying EA. Episodic ataxia (EA) is an inherited disease that leads to occasional loss of motor control in combination with variable other symptoms such as vertigo or migraine. EA type I (EA1), studied here, is caused by mutations in a voltage-gated potassium channel that contributes to the generation of electrical signals in the brain. The mechanism by which mutations in voltage-gated potassium channels lead to EA is still unknown and there is no consistent pharmacological treatment. By studying in detail one disease-causing mutation in Kv1.1, we describe a novel molecular mechanism distinct from mechanisms described previously. This mechanism contributes to the understanding of potassium channel function in general and might lead to a better understanding of how EA develops. Copyright © 2015 the authors 0270-6474/15/3512198-09$15.00/0.

Functional analysis of potassium channels in Kv7.2 G271V mutant causing early onset familial epilepsy.

PubMed

Wang, Juanjuan; Li, Yuan; Hui, Zhiyan; Cao, Min; Shi, Ruiming; Zhang, Wei; Geng, Limeng; Zhou, Xihui

2015-08-07

Kv7 (KCNQ) channels underlying a class of voltage-gated K+ current are best known for regulating neuronal excitability. The first glycine (G) residue in the pore helix of Kv7.2 (KCNQ2) subunit is highly conserved among different classes of Kv7 channel family. A missense mutation causing the replacement of the corresponding G residues with a valine (p.G271V) in Kv7.2 was found in a large, four-generation pedigree. Here, we set out to examine the molecular pathomechanism of G271V mutants using patch clamp technology combined with biochemical and immunocytochemical techniques in transiently transfected human embryonic kidney (HEK) 293 cells. The expression of Kv7.2 protein had the same intensity for both wild type (WT) and G271V. In transfected HEK cells, G271V mutants induced large depolarizing shifts of the conductance-voltage relationships and marked slowing of current activation kinetics compared to WT. In addition, G271V mutants abolished currents in homomeric channels, and resulted in about 50% reduction of current in Kv7.2/G271V/Kv7.3 heteromultimeric condition, indicating a more severe functional defect. To test for G271V mutant channel expression in surface membrane, we performed fluorescence confocal microscopy imaging, which revealed no differences between the mutant and WT, suggesting that G271V channels fail to open in response to depolarization even though they are present in the membrane. Furthermore, pharmacologic intervention experiments revealed that upon specific incubation of transfected HEK 293 cells expressing G271V heteromultimeric channels in presence of Kv7 channel enhancer retigabine (ezogabine), the potassium currents increased significantly, suggesting the potential of retigabine as gene-specific therapy. Copyright © 2015 Elsevier B.V. All rights reserved.

Structural and functional analysis of the putative pH sensor in the Kir1.1 (ROMK) potassium channel.

PubMed

Rapedius, Markus; Haider, Shozeb; Browne, Katharine F; Shang, Lijun; Sansom, Mark S P; Baukrowitz, Thomas; Tucker, Stephen J

2006-06-01

The pH-sensitive renal potassium channel Kir1.1 is important for K+ homeostasis. Disruption of the pH-sensing mechanism causes type II Bartter syndrome. The pH sensor is thought to be an anomalously titrated lysine residue (K80) that interacts with two arginine residues as part of an 'RKR triad'. We show that a Kir1.1 orthologue from Fugu rubripes lacks this lysine and yet is still highly pH sensitive, indicating that K80 is not the H+ sensor. Instead, K80 functionally interacts with A177 on transmembrane domain 2 at the 'helix-bundle crossing' and controls the ability of pH-dependent conformational changes to induce pore closure. Although not required for pH inhibition, K80 is indispensable for the coupling of pH gating to the extracellular K+ concentration, explaining its conservation in most Kir1.1 orthologues. Furthermore, we demonstrate that instead of interacting with K80, the RKR arginine residues form highly conserved inter- and intra-subunit interactions that are important for Kir channel gating and influence pH sensitivity indirectly.

Killing K channels with TEA+.

PubMed

Khodakhah, K; Melishchuk, A; Armstrong, C M

1997-11-25

Tetraethylammonium (TEA+) is widely used for reversible blockade of K channels in many preparations. We noticed that intracellular perfusion of voltage-clamped squid giant axons with a solution containing K+ and TEA+ irreversibly decreased the potassium current when there was no K+ outside. Five minutes of perfusion with 20 mM TEA+, followed by removal of TEA+, reduced potassium current to < 5% of its initial value. The irreversible disappearance of K channels with TEA+ could be prevented by addition of > or = 10 mM K+ to the extracellular solution. The rate of disappearance of K channels followed first-order kinetics and was slowed by reducing the concentration of TEA+. Killing is much less evident when an axon is held at -110 mV to tightly close all of the channels. The longer-chain TEA+ derivative decyltriethylammonium (C10+) had irreversible effects similar to TEA+. External K+ also protected K channels against the irreversible action of C10+. It has been reported that removal of all K+ internally and externally (dekalification) can result in the disappearance of K channels, suggesting that binding of K+ within the pore is required to maintain function. Our evidence further suggests that the crucial location for K+ binding is external to the (internal) TEA+ site and that TEA+ prevents refilling of this location by intracellular K+. Thus in the absence of extracellular K+, application of TEA+ (or C10+) has effects resembling dekalification and kills the K channels.

Novel inhibitors of the Gardos channel for the treatment of sickle cell disease.

PubMed

McNaughton-Smith, Grant A; Burns, J Ford; Stocker, Jonathan W; Rigdon, Gregory C; Creech, Christopher; Arrington, Susan; Shelton, Tara; de Franceschi, Lucia

2008-02-28

Sickle cell disease (SCD) is a hereditary condition characterized by deformation of red blood cells (RBCs). This phenomenon is due to the presence of abnormal hemoglobin that polymerizes upon deoxygenation. This effect is exacerbated when dehydrated RBCs experience a loss of both water and potassium salts. One critical pathway for the regulation of potassium efflux from RBCs is the Gardos channel, a calcium-activated potassium channel. This paper describes the synthesis and biological evaluation of a series of potent inhibitors of the Gardos channel. The goal was to identify compounds that were potent and selective inhibitors of the channel but had improved pharmacokinetic properties compared to 1, Clotrimazole. Several triarylamides such as 10 and 21 were potent inhibitors of the Gardos channel (IC50 of <10 nM) and active in a mouse model of SCD. Compound 21 (ICA-17043) was advanced into phase 3 clinical trials for SCD.

Neuro-Modulation of Immuno-Endocrine Response Induced by Kaliotoxin of Androctonus Scorpion Venom.

PubMed

Ladjel-Mendil, Amina; Martin-Eauclaire, Marie-France; Laraba-Djebari, Fatima

2016-12-01

Kaliotoxin (KTX), a specific blocker of potassium channels, exerts various toxic effects due to its action on the central nervous system. Its use in experimental model could help the understanding of the cellular and molecular mechanisms involved in the neuropathological processes related to potassium channel dysfunctions. In this study, the ability of KTX to stimulate neuro-immuno-endocrine axis was investigated. As results, the intracerebroventricular injection of KTX leads to severe structural-functional alterations of both hypothalamus and thyroid. These alterations were characterized by a massive release of hormones' markers of thyroid function associated with damaged tissue which was infiltrated by inflammatory cell and an imbalanced redox status. Taken together, these data highlight that KTX is able to modulate the neuro-endocrine response after binding to its targets leading to the hypothalamus and the thyroid stimulation, probably by inflammatory response activation and the installation of oxidative stress in these organs. © 2016 Wiley Periodicals, Inc.

Kcna1-mutant rats dominantly display myokymia, neuromyotonia and spontaneous epileptic seizures.

PubMed

Ishida, Saeko; Sakamoto, Yu; Nishio, Takeshi; Baulac, Stéphanie; Kuwamura, Mitsuru; Ohno, Yukihiro; Takizawa, Akiko; Kaneko, Shuji; Serikawa, Tadao; Mashimo, Tomoji

2012-01-30

Mutations in the KCNA1 gene, which encodes for the α subunit of the voltage-gated potassium channel Kv1.1, cause episodic ataxia type 1 (EA1). EA1 is a dominant human neurological disorder characterized by variable phenotypes of brief episodes of ataxia, myokymia, neuromyotonia, and associated epilepsy. Animal models for EA1 include Kcna1-deficient mice, which recessively display severe seizures and die prematurely, and V408A-knock-in mice, which dominantly exhibit stress-induced loss of motor coordination. In the present study, we have identified an N-ethyl-N-nitrosourea-mutagenized rat, named autosomal dominant myokymia and seizures (ADMS), with a missense mutation (S309T) in the voltage-sensor domain, S4, of the Kcna1 gene. ADMS rats dominantly exhibited myokymia, neuromyotonia and generalized tonic-clonic seizures. They also showed cold stress-induced tremor, neuromyotonia, and motor incoordination. Expression studies of homomeric and heteromeric Kv1.1 channels in HEK cells and Xenopus oocytes, showed that, although S309T channels are transferred to the cell membrane surface, they remained non-functional in terms of their biophysical properties, suggesting a dominant-negative effect of the S309T mutation on potassium channel function. ADMS rats provide a new model, distinct from previously reported mouse models, for studying the diverse functions of Kv1.1 in vivo, as well as for understanding the pathology of EA1. Copyright © 2011 Elsevier B.V. All rights reserved.

Prolonged action potential duration in cardiac ablation of PDK1 mice.

PubMed

Han, Zhonglin; Jiang, Yu; Yang, Zhongzhou; Cao, Kejiang; Wang, Dao W

2015-01-01

The involvement of the AGC protein kinase family in regulating arrhythmia has drawn considerable attention, but the underlying mechanisms are still not clear. The aim of this study is to explore the role of 3-phosphoinositide-dependent protein kinase-1 (PDK1), one of upstream protein kinases of the AGC protein kinase family, in the pathogenesis of dysregulated electrophysiological basis. PDK1(F/F) αMHC-Cre mice and PDK1(F/F) mice were divided into experiment group and control group. Using patch clamping technology, we explored action potential duration in both groups, and investigated the functions of transient outward potassium channel and L-type Ca(2+) channel to explain the abnormal action potential duration. Significant prolongation action potential duration was found in mice with PDK1 deletion. Further, the peak current of transient outward potassium current and L-type Ca(2+) current were decreased by 84% and 49% respectively. In addition, dysregulation of channel kinetics lead to action potential duration prolongation further. In conclusion, we have demonstrated that PDK1 participates in action potential prolongation in cardiac ablation of PDK1 mice. This effect is likely to be mediated largely through downregulation of transient outward potassium current. These findings indicate the modulation of the PDK1 pathway could provide a new mechanism for abnormal electrophysiological basis.

Involvement of WNK1-mediated potassium channels in the sexual dimorphism of blood pressure.

PubMed

Yu, Guofeng; Cheng, Mengting; Wang, Wei; Zhao, Rong; Liu, Zhen

2017-04-01

Potassium homeostasis plays an essential role in the control of blood pressure. It is unknown, however, whether potassium balance is involved in the gender-associated blood pressure differences. We therefore investigated the possible mechanism of sexual dimorphism in blood pressure regulation by measuring the blood pressure, plasma potassium, renal actions of potassium channels and upstream regulator in male and female mice. Here we found that female mice exhibited lower blood pressure and higher plasma K + level as compared to male littermates. Western blot analyses of mouse kidney extract revealed a significant decrease in renal outer medullary potassium (ROMK) channel expression, while large-conductance Ca 2+ -activated K + (BK) channel and Na-K-2Cl cotransporter (NKCC2) as well as the upstream regulator with-no-lysine kinase 1 (WNK1) enhanced in female mice under normal condition. Surprisingly, both dietary K + loading and K + depletion eliminated the differences in plasma K + and blood pressure between females and males, and the differences of renal K + channels and WNK1 also attenuated in both groups of mice. These findings indicated the existence of a close correlation between K + homeostasis and sex-associated blood pressure. Moreover, the differential regulation of ROMK, BK-α and NKCC2 between female and male mice, at least, were partly mediated via WNK1 pathway, which may contribute to the sexual dimorphism of plasma K + and blood pressure control. Copyright © 2017 Elsevier Inc. All rights reserved.

KCNJ11: Genetic Polymorphisms and Risk of Diabetes Mellitus

PubMed Central

Mohamed, Zahurin; Abdullah, Nor Azizan; Haghvirdizadeh, Pantea; Haerian, Monir Sadat

2015-01-01

Diabetes mellitus (DM) is a major worldwide health problem and its prevalence has been rapidly increasing in the last century. It is caused by defects in insulin secretion or insulin action or both, leading to hyperglycemia. Of the various types of DM, type 2 occurs most frequently. Multiple genes and their interactions are involved in the insulin secretion pathway. Insulin secretion is mediated through the ATP-sensitive potassium (KATP) channel in pancreatic beta cells. This channel is a heteromeric protein, composed of four inward-rectifier potassium ion channel (Kir6.2) tetramers, which form the pore of the KATP channel, as well as sulfonylurea receptor 1 subunits surrounding the pore. Kir6.2 is encoded by the potassium inwardly rectifying channel, subfamily J, member 11 (KCNJ11) gene, a member of the potassium channel genes. Numerous studies have reported the involvement of single nucleotide polymorphisms of the KCNJ11 gene and their interactions in the susceptibility to DM. This review discusses the current evidence for the contribution of common KCNJ11 genetic variants to the development of DM. Future studies should concentrate on understanding the exact role played by these risk variants in the development of DM. PMID:26448950

Comparison of the effects of the K(+)-channel openers cromakalim and minoxidil sulphate on vascular smooth muscle.

PubMed Central

Wickenden, A. D.; Grimwood, S.; Grant, T. L.; Todd, M. H.

1991-01-01

1 The actions of the potassium channel openers, cromakalim and minoxidil sulphate, were compared in a range of isolated blood vessel preparations. 2 Cromakalim and minoxidil sulphate inhibited spontaneous mechanical activity of the guinea-pig portal vein and relaxed the noradrenaline precontracted rat aorta with similar potency. In contrast, minoxidil sulphate was less potent than cromakalim in inhibiting spontaneous activity in the rat portal vein and was essentially inactive in the noradrenaline precontracted rat mesenteric artery and rabbit aorta. 3 Minoxidil sulphate did not antagonize the effects of cromakalim in the rabbit aorta indicating it was not acting as a partial 'agonist'. 4 Charybdotoxin, noxiustoxin and rubidium failed to discriminate between cromakalim and minoxidil sulphate indicating that the apparently selective effects of minoxidil sulphate were not mediated by either Ca(2+)-activated potassium channels, delayed rectifiers or rubidium impermeable potassium channels. 5 Glibenclamide antagonized the effects of cromakalim in an apparently competitive manner whereas the effects of minoxidil sulphate were antagonized in a non-competitive manner. The involvement of subtypes of ATP-sensitive potassium channels is discussed. PMID:1878752

Cellular Mechanism of Inner Ear Genetic Disease, roles of Kv7.1 (KCNQ1) Channel

NASA Astrophysics Data System (ADS)

Mousavi Nik, Atefeh

Potassium channels are the most diverse and widely distributed membrane protein in all living organisms. They have various roles in the body such as controlling membrane potential, cell volume, and cell migration. Many studies have shown that mutation in these channels is associated with different diseases for example: Hearing Defect, Cardiac Arrhythmia, Episodic Ataxia, Seizure and Neuromyotonia. One of the most important diseases associated with K+ channel mutations is called Jervell and Lange-Nielsen syndrome (JLNS). This disease causes bilateral congenital deafness and the patients also suffer from Long QT and they usually experience syncopal episodes in their life and eventually die as a result of cardiac arrest. The gene KCNQ1 encodes the Kv7.1 voltage gated potassium channel. This channel expresses in apical membrane of marginal cell in stria vasularis of cochlea and secret K+ ion to endolymp to keep the endocochlear potential stable, which is necessary for the inner ear to function properly. Kv7.1 channel also expresses in cardiac myocytes and mutation in this gene is associated with another syndrome called Romano-Ward syndrome (RWS). Although Romano-Ward patients have mutation in KCNQ1, similar to Jervell and Lange-Nielsen patients, they only suffer from cardiac defect, and their hearing is completely normal. Several studies identified that mutations in Kv7.1 gene is associated with JLNS and RWS, but the biophysical and cellular mechanisms of these mutations are still unknown. To determine the cellular mechanisms of JLNS and RWS, and to provide mechanistic insight on the functional outputs of JLNS versus RWS mutations, we generated several mutant forms of the human Kv7.1 ( KCNQ1) clone, using site-directed mutagenesis to define their sub-cellular localization and examined their electrophysiological properties. We identified JLNS and RWS mutations at the S4-S5-linker, the pore loop (P-loop) and the C-terminus of hKv7.1 which have been found to control channel gating, permeation and modulation, respectively. The result showed that for JLNS, all P-loop and C-terminal mutations (seven mutations) yielded non-functional channels when expressed alone. Moreover, the W248F at the end of the S4 domain yielded a functional current, but it became inactivated at positive step potentials, therefore the channel essentially is non-functional. All the JLNS mutant channels are non-functional, and have impaired membrane trafficking. In contrast, the RWS mutants showed wide-ranging functional phenotypes consisting of channels with large current, channel with no measurable current when expressed alone, but significant current upon addition of the WT subunit and finally channel with no measurable current even in presence of WT subunit. The RWS mutants, however, produced dominant negative effect. These findings provide integrated cellular and molecular mechanisms of hKv7.1 functions and the consequent diseased phenotype in JLNS and RWS may come from the tissue-specific function of the channel.

Mechanism of Electromechanical Coupling in Voltage-Gated Potassium Channels

PubMed Central

Blunck, Rikard; Batulan, Zarah

2012-01-01

Voltage-gated ion channels play a central role in the generation of action potentials in the nervous system. They are selective for one type of ion – sodium, calcium, or potassium. Voltage-gated ion channels are composed of a central pore that allows ions to pass through the membrane and four peripheral voltage sensing domains that respond to changes in the membrane potential. Upon depolarization, voltage sensors in voltage-gated potassium channels (Kv) undergo conformational changes driven by positive charges in the S4 segment and aided by pairwise electrostatic interactions with the surrounding voltage sensor. Structure-function relations of Kv channels have been investigated in detail, and the resulting models on the movement of the voltage sensors now converge to a consensus; the S4 segment undergoes a combined movement of rotation, tilt, and vertical displacement in order to bring 3–4e+ each through the electric field focused in this region. Nevertheless, the mechanism by which the voltage sensor movement leads to pore opening, the electromechanical coupling, is still not fully understood. Thus, recently, electromechanical coupling in different Kv channels has been investigated with a multitude of techniques including electrophysiology, 3D crystal structures, fluorescence spectroscopy, and molecular dynamics simulations. Evidently, the S4–S5 linker, the covalent link between the voltage sensor and pore, plays a crucial role. The linker transfers the energy from the voltage sensor movement to the pore domain via an interaction with the S6 C-termini, which are pulled open during gating. In addition, other contact regions have been proposed. This review aims to provide (i) an in-depth comparison of the molecular mechanisms of electromechanical coupling in different Kv channels; (ii) insight as to how the voltage sensor and pore domain influence one another; and (iii) theoretical predictions on the movement of the cytosolic face of the Kv channels during gating. PMID:22988442

Compartmentalized beta subunit distribution determines characteristics and ethanol sensitivity of somatic, dendritic, and terminal large-conductance calcium-activated potassium channels in the rat central nervous system.

PubMed

Wynne, P M; Puig, S I; Martin, G E; Treistman, S N

2009-06-01

Neurons are highly differentiated and polarized cells, whose various functions depend upon the compartmentalization of ion channels. The rat hypothalamic-neurohypophysial system (HNS), in which cell bodies and dendrites reside in the hypothalamus, physically separated from their nerve terminals in the neurohypophysis, provides a particularly powerful preparation in which to study the distribution and regional properties of ion channel proteins. Using electrophysiological and immunohistochemical techniques, we characterized the large-conductance calcium-activated potassium (BK) channel in each of the three primary compartments (soma, dendrite, and terminal) of HNS neurons. We found that dendritic BK channels, in common with somatic channels but in contrast to nerve terminal channels, are insensitive to iberiotoxin. Furthermore, analysis of dendritic BK channel gating kinetics indicates that they, like somatic channels, have fast activation kinetics, in contrast to the slow gating of terminal channels. Dendritic and somatic channels are also more sensitive to calcium and have a greater conductance than terminal channels. Finally, although terminal BK channels are highly potentiated by ethanol, somatic and dendritic channels are insensitive to the drug. The biophysical and pharmacological properties of somatic and dendritic versus nerve terminal channels are consistent with the characteristics of exogenously expressed alphabeta1 versus alphabeta4 channels, respectively. Therefore, one possible explanation for our findings is a selective distribution of auxiliary beta1 subunits to the somatic and dendritic compartments and beta4 to the terminal compartment. This hypothesis is supported immunohistochemically by the appearance of distinct punctate beta1 or beta4 channel clusters in the membrane of somatic and dendritic or nerve terminal compartments, respectively.

Synergistic action of cyclic adenosine monophosphate- and calcium-mediated chloride secretion in a colonic epithelial cell line.

PubMed Central

Cartwright, C A; McRoberts, J A; Mandel, K G; Dharmsathaphorn, K

1985-01-01

Vasoactive intestinal polypeptide (VIP) and the calcium ionophore A23187 caused dose-dependent changes in the potential difference and the short circuit current (Isc) across confluent T84 cell monolayers mounted in modified Ussing chambers. Both VIP and A23187 stimulated net chloride secretion without altering sodium transport. Net chloride secretion accounted for the increase in Isc. When A23187 was tested in combination with VIP, net chloride secretion was significantly greater than predicted from the calculated sum of their individual responses indicating a synergistic effect. VIP increased cellular cyclic AMP (cAMP) production in a dose-dependent manner, whereas A23187 had no effect on cellular cAMP. We then determined whether VIP and A23187 activated different transport pathways. Earlier studies suggest that VIP activates a basolaterally localized, barium-sensitive potassium channel as well as an apically localized chloride conductance pathway. In this study, stimulation of basolateral membrane potassium efflux by A23187 was documented by preloading the monolayers with 86Rb+. Stimulation of potassium efflux by A23187 was additive to the VIP-stimulated potassium efflux. By itself, 0.3 microM A23187 did not alter transepithelial chloride permeability, and its stimulation of basolateral membrane potassium efflux caused only a relatively small amount of chloride secretion. However, in the presence of an increased transepithelial chloride permeability induced by VIP, the effectiveness of A23187 on chloride secretion was greatly augmented. Our studies suggest that cAMP and calcium each activate basolateral potassium channels, but cAMP also activates an apically localized chloride channel. Synergism results from cooperative interaction of potassium channels and the chloride channel. PMID:2997291

Oxidative Modulation of Voltage-Gated Potassium Channels

PubMed Central

Sahoo, Nirakar; Hoshi, Toshinori

2014-01-01

Abstract Significance: Voltage-gated K+ channels are a large family of K+-selective ion channel protein complexes that open on membrane depolarization. These K+ channels are expressed in diverse tissues and their function is vital for numerous physiological processes, in particular of neurons and muscle cells. Potentially reversible oxidative regulation of voltage-gated K+ channels by reactive species such as reactive oxygen species (ROS) represents a contributing mechanism of normal cellular plasticity and may play important roles in diverse pathologies including neurodegenerative diseases. Recent Advances: Studies using various protocols of oxidative modification, site-directed mutagenesis, and structural and kinetic modeling provide a broader phenomenology and emerging mechanistic insights. Critical Issues: Physicochemical mechanisms of the functional consequences of oxidative modifications of voltage-gated K+ channels are only beginning to be revealed. In vivo documentation of oxidative modifications of specific amino-acid residues of various voltage-gated K+ channel proteins, including the target specificity issue, is largely absent. Future Directions: High-resolution chemical and proteomic analysis of ion channel proteins with respect to oxidative modification combined with ongoing studies on channel structure and function will provide a better understanding of how the function of voltage-gated K+ channels is tuned by ROS and the corresponding reducing enzymes to meet cellular needs. Antioxid. Redox Signal. 21, 933–952. PMID:24040918

Co-expression of the voltage-gated potassium channel Kv1.4 with transient receptor potential channels (TRPV1 and TRPV2) and the cannabinoid receptor CB1 in rat dorsal root ganglion neurons.

PubMed

Binzen, U; Greffrath, W; Hennessy, S; Bausen, M; Saaler-Reinhardt, S; Treede, R-D

2006-10-13

Potassium channels contribute to basic neuronal excitability and modulation. Here, we examined expression patterns of the voltage-gated potassium channel Kv1.4, the nociceptive transduction channels TRPV1 and TRPV2 as well as the putative anti-nociceptive cannabinoid receptor CB1 by immunofluorescence double-labelings in sections of rat dorsal root ganglia (DRGs). Kv1.4, TRPV1 and CB1 were each detected in about one third of neurons (35.7+/-0.5%, 29.4+/-1.1% and 36.4+/-0.5%, respectively, mean diameter 19.1+/-0.3 microm). TRPV2 was present in 4.4+/-0.4% of all neurons that were significantly larger in diameter (27.4+/-0.7 microm; P < 0.001). Antibody double-labeling revealed that the majority of Kv1.4-positive neurons co-expressed TRPV1 (73.9+/-1.5%) whereas none expressed TRPV2. The largest overlap was found with CB1 (93.1+/-0.1%). CB1 expression resembled that seen for Kv1.4 since the majority of neurons expressing CB1-protein also expressed TRPV1 (69.4+/-6.5%) but not TRPV2 (0.6+/-0.3%). When CB1-mRNA was detected using in situ hybridizations an additional subset of larger neurons was labeled including 82.4+/-17.7% of the TRPV2 expressing neurons. However, co-localization of Kv1.4 with CB1-mRNA (92%, mean diameter: 18.5 microm) was essentially the same as with CB1-protein. The almost complete overlap of CB1 and Kv1.4 in nociceptive DRG neurons suggests a functional synergistic action between Kv1.4 and CB1. The potassium channel may have two important roles in nociception. As the molecular basis of A-type current it could be involved in the control of repetitive discharges at peripheral terminals and as a downstream signal transduction site of CB1 in the control of presynaptic transmitter release at central terminals.

Modulation of Potassium Channels Inhibits Bunyavirus Infection.

PubMed

Hover, Samantha; King, Barnabas; Hall, Bradley; Loundras, Eleni-Anna; Taqi, Hussah; Daly, Janet; Dallas, Mark; Peers, Chris; Schnettler, Esther; McKimmie, Clive; Kohl, Alain; Barr, John N; Mankouri, Jamel

2016-02-12

Bunyaviruses are considered to be emerging pathogens facilitated by the segmented nature of their genome that allows reassortment between different species to generate novel viruses with altered pathogenicity. Bunyaviruses are transmitted via a diverse range of arthropod vectors, as well as rodents, and have established a global disease range with massive importance in healthcare, animal welfare, and economics. There are no vaccines or anti-viral therapies available to treat human bunyavirus infections and so development of new anti-viral strategies is urgently required. Bunyamwera virus (BUNV; genus Orthobunyavirus) is the model bunyavirus, sharing aspects of its molecular and cellular biology with all Bunyaviridae family members. Here, we show for the first time that BUNV activates and requires cellular potassium (K(+)) channels to infect cells. Time of addition assays using K(+) channel modulating agents demonstrated that K(+) channel function is critical to events shortly after virus entry but prior to viral RNA synthesis/replication. A similar K(+) channel dependence was identified for other bunyaviruses namely Schmallenberg virus (Orthobunyavirus) as well as the more distantly related Hazara virus (Nairovirus). Using a rational pharmacological screening regimen, two-pore domain K(+) channels (K2P) were identified as the K(+) channel family mediating BUNV K(+) channel dependence. As several K2P channel modulators are currently in clinical use, our work suggests they may represent a new and safe drug class for the treatment of potentially lethal bunyavirus disease. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Chloride and potassium channels in cystic fibrosis airway epithelia

NASA Astrophysics Data System (ADS)

Welsh, Michael J.; Liedtke, Carole M.

1986-07-01

Cystic fibrosis, the most common lethal genetic disease in Caucasians, is characterized by a decreased permeability in sweat gland duct and airway epithelia. In sweat duct epithelium, a decreased Cl- permeability accounts for the abnormally increased salt content of sweat1. In airway epithelia a decreased Cl- permeability, and possibly increased sodium absorption, may account for the abnormal respiratory tract fluid2,3. The Cl- impermeability has been localized to the apical membrane of cystic fibrosis airway epithelial cells4. The finding that hormonally regulated Cl- channels make the apical membrane Cl- permeable in normal airway epithelial cells5 suggested abnormal Cl- channel function in cystic fibrosis. Here we report that excised, cell-free patches of membrane from cystic fibrosis epithelial cells contain Cl- channels that have the same conductive properties as Cl- channels from normal cells. However, Cl- channels from cystic fibrosis cells did not open when they were attached to the cell. These findings suggest defective regulation of Cl- channels in cystic fibrosis epithelia; to begin to address this issue, we performed two studies. First, we found that isoprenaline, which stimulates Cl- secretion, increases cellular levels of cyclic AMP in a similar manner in cystic fibrosis and non-cystic fibrosis epithelial cells. Second, we show that adrenergic agonists open calcium-activated potassium channels, indirectly suggesting that calcium-dependent stimulus-response coupling is intact in cystic fibrosis. These data suggest defective regulation of Cl- channels at a site distal to cAMP accumulation.

Potassium Intake, Bioavailability, Hypertension, and Glucose Control

PubMed Central

Stone, Michael S.; Martyn, Lisa; Weaver, Connie M.

2016-01-01

Potassium is an essential nutrient. It is the most abundant cation in intracellular fluid where it plays a key role in maintaining cell function. The gradient of potassium across the cell membrane determines cellular membrane potential, which is maintained in large part by the ubiquitous ion channel the sodium-potassium (Na+-K+) ATPase pump. Approximately 90% of potassium consumed (60–100 mEq) is lost in the urine, with the other 10% excreted in the stool, and a very small amount lost in sweat. Little is known about the bioavailability of potassium, especially from dietary sources. Less is understood on how bioavailability may affect health outcomes. Hypertension (HTN) is the leading cause of cardiovascular disease (CVD) and a major financial burden ($50.6 billion) to the US public health system, and has a significant impact on all-cause morbidity and mortality worldwide. The relationship between increased potassium supplementation and a decrease in HTN is relatively well understood, but the effect of increased potassium intake from dietary sources on blood pressure overall is less clear. In addition, treatment options for hypertensive individuals (e.g., thiazide diuretics) may further compound chronic disease risk via impairments in potassium utilization and glucose control. Understanding potassium bioavailability from various sources may help to reveal how specific compounds and tissues influence potassium movement, and further the understanding of its role in health. PMID:27455317

Potassium Intake, Bioavailability, Hypertension, and Glucose Control.

PubMed

Stone, Michael S; Martyn, Lisa; Weaver, Connie M

2016-07-22

Potassium is an essential nutrient. It is the most abundant cation in intracellular fluid where it plays a key role in maintaining cell function. The gradient of potassium across the cell membrane determines cellular membrane potential, which is maintained in large part by the ubiquitous ion channel the sodium-potassium (Na+-K+) ATPase pump. Approximately 90% of potassium consumed (60-100 mEq) is lost in the urine, with the other 10% excreted in the stool, and a very small amount lost in sweat. Little is known about the bioavailability of potassium, especially from dietary sources. Less is understood on how bioavailability may affect health outcomes. Hypertension (HTN) is the leading cause of cardiovascular disease (CVD) and a major financial burden ($50.6 billion) to the US public health system, and has a significant impact on all-cause morbidity and mortality worldwide. The relationship between increased potassium supplementation and a decrease in HTN is relatively well understood, but the effect of increased potassium intake from dietary sources on blood pressure overall is less clear. In addition, treatment options for hypertensive individuals (e.g., thiazide diuretics) may further compound chronic disease risk via impairments in potassium utilization and glucose control. Understanding potassium bioavailability from various sources may help to reveal how specific compounds and tissues influence potassium movement, and further the understanding of its role in health.

Long-pore Electrostatics in Inward-rectifier Potassium Channels

PubMed Central

Robertson, Janice L.; Palmer, Lawrence G.; Roux, Benoît

2008-01-01

Inward-rectifier potassium (Kir) channels differ from the canonical K+ channel structure in that they possess a long extended pore (∼85 Å) for ion conduction that reaches deeply into the cytoplasm. This unique structural feature is presumably involved in regulating functional properties specific to Kir channels, such as conductance, rectification block, and ligand-dependent gating. To elucidate the underpinnings of these functional roles, we examine the electrostatics of an ion along this extended pore. Homology models are constructed based on the open-state model of KirBac1.1 for four mammalian Kir channels: Kir1.1/ROMK, Kir2.1/IRK, Kir3.1/GIRK, and Kir6.2/KATP. By solving the Poisson-Boltzmann equation, the electrostatic free energy of a K+ ion is determined along each pore, revealing that mammalian Kir channels provide a favorable environment for cations and suggesting the existence of high-density regions in the cytoplasmic domain and cavity. The contribution from the reaction field (the self-energy arising from the dielectric polarization induced by the ion's charge in the complex geometry of the pore) is unfavorable inside the long pore. However, this is well compensated by the electrostatic interaction with the static field arising from the protein charges and shielded by the dielectric surrounding. Decomposition of the static field provides a list of residues that display remarkable correspondence with existing mutagenesis data identifying amino acids that affect conduction and rectification. Many of these residues demonstrate interactions with the ion over long distances, up to 40 Å, suggesting that mutations potentially affect ion or blocker energetics over the entire pore. These results provide a foundation for understanding ion interactions in Kir channels and extend to the study of ion permeation, block, and gating in long, cation-specific pores. PMID:19001143

Functional interactions at the interface between voltage-sensing and pore domains in the Shaker K(v) channel.

PubMed

Soler-Llavina, Gilberto J; Chang, Tsg-Hui; Swartz, Kenton J

2006-11-22

Voltage-activated potassium (K(v)) channels contain a central pore domain that is partially surrounded by four voltage-sensing domains. Recent X-ray structures suggest that the two domains lack extensive protein-protein contacts within presumed transmembrane regions, but whether this is the case for functional channels embedded in lipid membranes remains to be tested. We investigated domain interactions in the Shaker K(v) channel by systematically mutating the pore domain and assessing tolerance by examining channel maturation, S4 gating charge movement, and channel opening. When mapped onto the X-ray structure of the K(v)1.2 channel the large number of permissive mutations support the notion of relatively independent domains, consistent with crystallographic studies. Inspection of the maps also identifies portions of the interface where residues are sensitive to mutation, an external cluster where mutations hinder voltage sensor activation, and an internal cluster where domain interactions between S4 and S5 helices from adjacent subunits appear crucial for the concerted opening transition.

Chondrocyte channel transcriptomics

PubMed Central

Lewis, Rebecca; May, Hannah; Mobasheri, Ali; Barrett-Jolley, Richard

2013-01-01

To date, a range of ion channels have been identified in chondrocytes using a number of different techniques, predominantly electrophysiological and/or biomolecular; each of these has its advantages and disadvantages. Here we aim to compare and contrast the data available from biophysical and microarray experiments. This letter analyses recent transcriptomics datasets from chondrocytes, accessible from the European Bioinformatics Institute (EBI). We discuss whether such bioinformatic analysis of microarray datasets can potentially accelerate identification and discovery of ion channels in chondrocytes. The ion channels which appear most frequently across these microarray datasets are discussed, along with their possible functions. We discuss whether functional or protein data exist which support the microarray data. A microarray experiment comparing gene expression in osteoarthritis and healthy cartilage is also discussed and we verify the differential expression of 2 of these genes, namely the genes encoding large calcium-activated potassium (BK) and aquaporin channels. PMID:23995703

Comparative Study of the Energetics of Ion Permeation in Kv1.2 and KcsA Potassium Channels

PubMed Central

Baştuğ, Turgut; Kuyucak, Serdar

2011-01-01

Biological ion channels rely on a multi-ion transport mechanism for fast yet selective permeation of ions. The crystal structure of the KcsA potassium channel provided the first microscopic picture of this process. A similar mechanism is assumed to operate in all potassium channels, but the validity of this assumption has not been well investigated. Here, we examine the energetics of ion permeation in Shaker Kv1.2 and KcsA channels, which exemplify the six-transmembrane voltage-gated and two-transmembrane inward-rectifier channels. We study the feasibility of binding a third ion to the filter and the concerted motion of ions in the channel by constructing the potential of mean force for K+ ions in various configurations. For both channels, we find that a pair of K+ ions can move almost freely within the filter, but a relatively large free-energy barrier hinders the K+ ion from stepping outside the filter. We discuss the effect of the CMAP dihedral energy correction that was recently incorporated into the CHARMM force field on ion permeation dynamics. PMID:21281577

Competition of calcified calmodulin N lobe and PIP2 to an LQT mutation site in Kv7.1 channel

PubMed Central

Tobelaim, William Sam; Dvir, Meidan; Lebel, Guy; Cui, Meng; Buki, Tal; Peretz, Asher; Marom, Milit; Haitin, Yoni; Logothetis, Diomedes E.; Hirsch, Joel Alan; Attali, Bernard

2017-01-01

Voltage-gated potassium 7.1 (Kv7.1) channel and KCNE1 protein coassembly forms the slow potassium current IKS that repolarizes the cardiac action potential. The physiological importance of the IKS channel is underscored by the existence of mutations in human Kv7.1 and KCNE1 genes, which cause cardiac arrhythmias, such as the long-QT syndrome (LQT) and atrial fibrillation. The proximal Kv7.1 C terminus (CT) binds calmodulin (CaM) and phosphatidylinositol-4,5-bisphosphate (PIP2), but the role of CaM in channel function is still unclear, and its possible interaction with PIP2 is unknown. Our recent crystallographic study showed that CaM embraces helices A and B with the apo C lobe and calcified N lobe, respectively. Here, we reveal the competition of PIP2 and the calcified CaM N lobe to a previously unidentified site in Kv7.1 helix B, also known to harbor an LQT mutation. Protein pulldown, molecular docking, molecular dynamics simulations, and patch-clamp recordings indicate that residues K526 and K527 in Kv7.1 helix B form a critical site where CaM competes with PIP2 to stabilize the channel open state. Data indicate that both PIP2 and Ca2+-CaM perform the same function on IKS channel gating by producing a left shift in the voltage dependence of activation. The LQT mutant K526E revealed a severely impaired channel function with a right shift in the voltage dependence of activation, a reduced current density, and insensitivity to gating modulation by Ca2+-CaM. The results suggest that, after receptor-mediated PIP2 depletion and increased cytosolic Ca2+, calcified CaM N lobe interacts with helix B in place of PIP2 to limit excessive IKS current inhibition. PMID:28096388

Competition of calcified calmodulin N lobe and PIP2 to an LQT mutation site in Kv7.1 channel.

PubMed

Tobelaim, William Sam; Dvir, Meidan; Lebel, Guy; Cui, Meng; Buki, Tal; Peretz, Asher; Marom, Milit; Haitin, Yoni; Logothetis, Diomedes E; Hirsch, Joel Alan; Attali, Bernard

2017-01-31

Voltage-gated potassium 7.1 (Kv7.1) channel and KCNE1 protein coassembly forms the slow potassium current I KS that repolarizes the cardiac action potential. The physiological importance of the I KS channel is underscored by the existence of mutations in human Kv7.1 and KCNE1 genes, which cause cardiac arrhythmias, such as the long-QT syndrome (LQT) and atrial fibrillation. The proximal Kv7.1 C terminus (CT) binds calmodulin (CaM) and phosphatidylinositol-4,5-bisphosphate (PIP 2 ), but the role of CaM in channel function is still unclear, and its possible interaction with PIP 2 is unknown. Our recent crystallographic study showed that CaM embraces helices A and B with the apo C lobe and calcified N lobe, respectively. Here, we reveal the competition of PIP 2 and the calcified CaM N lobe to a previously unidentified site in Kv7.1 helix B, also known to harbor an LQT mutation. Protein pulldown, molecular docking, molecular dynamics simulations, and patch-clamp recordings indicate that residues K526 and K527 in Kv7.1 helix B form a critical site where CaM competes with PIP 2 to stabilize the channel open state. Data indicate that both PIP 2 and Ca 2+ -CaM perform the same function on I KS channel gating by producing a left shift in the voltage dependence of activation. The LQT mutant K526E revealed a severely impaired channel function with a right shift in the voltage dependence of activation, a reduced current density, and insensitivity to gating modulation by Ca 2+ -CaM. The results suggest that, after receptor-mediated PIP 2 depletion and increased cytosolic Ca 2+ , calcified CaM N lobe interacts with helix B in place of PIP 2 to limit excessive I KS current inhibition.

Synthesis, folding, structure and activity of a predicted peptide from the sea anemone Oulactis sp. with an ShKT fold.

PubMed

Krishnarjuna, Bankala; Villegas-Moreno, Jessica; Mitchell, Michela L; Csoti, Agota; Peigneur, Steve; Amero, Carlos; Pennington, Michael W; Tytgat, Jan; Panyi, Gyorgy; Norton, Raymond S

2018-05-19

Sea anemone venom is rich in bioactive compounds, including peptides containing multiple disulfide bridges. In a transcriptomic study on Oulactis sp., we identified the putative 36-residue peptide, OspTx2b, which is an isoform of the K V channel blocker OspTx2a (Sunanda P et al. [2018] Identification, chemical synthesis, structure and function of a new K V 1 channel blocking peptide from Oulactis sp. Peptide Science, in press). As OspTx2b contains a ShK/BgK-like cysteine framework, with high amino acid sequence similarity to BgK, we were interested to investigate its structure and function. The solution structure of OspTx2b was determined using nuclear magnetic resonance spectroscopy. OspTx2b does indeed possess a BgK-like scaffold, with the same disulfide bond connectivities. The orientation of the Lys-Tyr dyad in OspTx2b is more similar to that in ShK than in BgK. However, it failed to show against a range of voltage-gated potassium channels in Xenopus oocytes and human T lymphocytes. OspTx2b also showed no growth inhibitory activity against several strains of bacteria and fungi. Having a BgK-like fold with the Lys-Tyr dyad but no BgK-like activity highlights the importance of key amino acid residues in BgK that are missing in OspTx2b. The lack of activity against the K V channels assessed in this study emphasises that the ShK/BgK scaffold is capable of supporting functional activity beyond potassium channel blockade. Copyright © 2018. Published by Elsevier Ltd.

Coulomb interaction rules timescales in potassium ion channel tunneling

NASA Astrophysics Data System (ADS)

De March, N.; Prado, S. D.; Brunnet, L. G.

2018-06-01

Assuming the selectivity filter of KcsA potassium ion channel may exhibit quantum coherence, we extend a previous model by Vaziri and Plenio (2010 New J. Phys. 12 085001) to take into account Coulomb repulsion between potassium ions. We show that typical ion transit timescales are determined by this interaction, which imposes optimal input/output parameter ranges. Also, as observed in other examples of quantum tunneling in biological systems, the addition of moderate noise helps coherent ion transport.

SLO BK Potassium Channels Couple Gap Junctions to Inhibition of Calcium Signaling in Olfactory Neuron Diversification.

PubMed

Alqadah, Amel; Hsieh, Yi-Wen; Schumacher, Jennifer A; Wang, Xiaohong; Merrill, Sean A; Millington, Grethel; Bayne, Brittany; Jorgensen, Erik M; Chuang, Chiou-Fen

2016-01-01

The C. elegans AWC olfactory neuron pair communicates to specify asymmetric subtypes AWCOFF and AWCON in a stochastic manner. Intercellular communication between AWC and other neurons in a transient NSY-5 gap junction network antagonizes voltage-activated calcium channels, UNC-2 (CaV2) and EGL-19 (CaV1), in the AWCON cell, but how calcium signaling is downregulated by NSY-5 is only partly understood. Here, we show that voltage- and calcium-activated SLO BK potassium channels mediate gap junction signaling to inhibit calcium pathways for asymmetric AWC differentiation. Activation of vertebrate SLO-1 channels causes transient membrane hyperpolarization, which makes it an important negative feedback system for calcium entry through voltage-activated calcium channels. Consistent with the physiological roles of SLO-1, our genetic results suggest that slo-1 BK channels act downstream of NSY-5 gap junctions to inhibit calcium channel-mediated signaling in the specification of AWCON. We also show for the first time that slo-2 BK channels are important for AWC asymmetry and act redundantly with slo-1 to inhibit calcium signaling. In addition, nsy-5-dependent asymmetric expression of slo-1 and slo-2 in the AWCON neuron is necessary and sufficient for AWC asymmetry. SLO-1 and SLO-2 localize close to UNC-2 and EGL-19 in AWC, suggesting a role of possible functional coupling between SLO BK channels and voltage-activated calcium channels in AWC asymmetry. Furthermore, slo-1 and slo-2 regulate the localization of synaptic markers, UNC-2 and RAB-3, in AWC neurons to control AWC asymmetry. We also identify the requirement of bkip-1, which encodes a previously identified auxiliary subunit of SLO-1, for slo-1 and slo-2 function in AWC asymmetry. Together, these results provide an unprecedented molecular link between gap junctions and calcium pathways for terminal differentiation of olfactory neurons.

Effects of calcium antagonists on isolated bovine cerebral arteries: inhibition of constriction and calcium-45 uptake induced by potassium or serotonin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wendling, W.W.; Harakal, C.

1987-05-01

The purpose of this study was to determine the mechanisms by which organic calcium channel blockers inhibit cerebral vasoconstriction. Isolated bovine middle cerebral arteries were cut into rings to measure contractility or into strips to measure radioactive calcium (/sup 45/Ca) influx and efflux. Calcium channel blockers (10(-5) M verapamil or 3.3 X 10(-7) M nifedipine) and calcium-deficient solutions all produced near-maximal inhibition of both potassium- and serotonin-induced constriction. In calcium-deficient solutions containing potassium or serotonin, verapamil and nifedipine each blocked subsequent calcium-induced constriction in a competitive manner. Potassium and serotonin significantly increased /sup 45/Ca uptake into cerebral artery strips duringmore » 5 minutes of /sup 45/Ca loading; for potassium /sup 45/Ca uptake increased from 62 to 188 nmol/g, and for serotonin from 65 to 102 nmol/g. Verapamil or nifedipine had no effect on basal /sup 45/Ca uptake but significantly blocked the increase in /sup 45/Ca uptake induced by potassium or serotonin. Potassium, and to a lesser extent serotonin, each induced a brief increase in the rate of /sup 45/Ca efflux into calcium-deficient solutions. Verapamil or nifedipine had no effect on basal or potassium-stimulated /sup 45/Ca efflux. The results demonstrate that verapamil and nifedipine block /sup 45/Ca uptake through both potential-operated (potassium) and receptor-operated (serotonin) channels in bovine middle cerebral arteries.« less

Inactivation properties of voltage-gated K+ channels altered by presence of beta-subunit.

PubMed

Rettig, J; Heinemann, S H; Wunder, F; Lorra, C; Parcej, D N; Dolly, J O; Pongs, O

1994-05-26

Structural and functional diversity of voltage-gated Kv1-type potassium channels in rat brain is enhanced by the association of two different types of subunits, the membrane-bound, poreforming alpha-subunits and a peripheral beta-subunit. We have cloned a beta-subunit (Kv beta 1) that is specifically expressed in the rat nervous system. Association of Kv beta 1 with alpha-subunits confers rapid A-type inactivation on non-inactivating Kv1 channels (delayed rectifiers) in expression systems in vitro. This effect is mediated by an inactivating ball domain in the Kv beta 1 amino terminus.

Differential Expression of Renal Outer Medullary K+ Channel and Voltage-gated K+ Channel 7.1 in Bladder Urothelium of Patients With Interstitial Cystitis/Painful Bladder Syndrome.

PubMed

Lee, Jane-Dar; Lee, Ming-Huei; Yang, Wen-Kai; Wang, Kuan-Lin; Lee, Tsung-Han

2017-03-01

To investigate the changes including expression and localization of 2 potassium channels, renal outer medullary K + channel (ROMK) and voltage-gated K + channel 7.1 (KCNQ1), after increased urinary potassium leakage in patients with interstitial cystitis/painful bladder syndrome (IC/PBS). The study group included 24 patients with IC/PBS and a control group consisting of 12 volunteers without any IC/PBS symptoms. Bladder biopsies were taken from both groups. We determined the protein expression and distribution of potassium channels using immunoblotting, immunohistochemistry, and immunofluorescent staining under confocal laser microscopy. The results revealed that ROMK was predominantly expressed in apical cells of the bladder urothelium at significantly higher levels (3.3-fold) in the study group than in the control group. In contrast, KCNQ1 was expressed in the basolateral membrane according to confocal microscopy results and did not significantly differ between groups. Our data showed that the abundance of ROMK protein in apical cells was increased in the IC/PBS group, whereas KCNQ1, which was distributed in the basolateral membrane of the bladder urothelium, showed similar abundance between groups. These results suggest that upregulation of the ROMK channel in apical cells might permit avid potassium flux into the bladder lumen to maintain intracellular K + homeostasis in the dysfunctional urothelium. Copyright © 2016 Elsevier Inc. All rights reserved.

Recent Advances in the Pathogenesis and Drug Action in Periodic Paralyses and Related Channelopathies

PubMed Central

Tricarico, Domenico; Camerino, Diana Conte

2011-01-01

The periodic paralysis (PP) are rare autosomal-dominant disorders associated to mutations in the skeletal muscle sodium, calcium, and potassium channel genes characterized by muscle fiber depolarization with un-excitability, episodes of weakness with variations in serum potassium concentrations. Recent advances in thyrotoxic PP and hypokalemic PP (hypoPP) confirm the involvement of the muscle potassium channels in the pathogenesis of the diseases and their role as target of action for drugs of therapeutic interest. The novelty in the gating pore currents theory help to explain the disease symptoms, and open the possibility to more specifically target the disease. It is now known that the fiber depolarization in the hypoPP is due to an unbalance between the novel identified depolarizing gating pore currents (Igp) carried by protons or Na+ ions flowing through aberrant alternative pathways of the mutant subunits and repolarizing inwardly rectifying potassium channel (Kir) currents which also includes the ATP-sensitive subtype. Abnormal activation of the Igp or deficiency in the Kir channels predispose to fiber depolarization. One pharmacological strategy is based on blocking the Igp without affecting normal channel gating. It remains safe and effective the proposal of targeting the KATP, Kir channels, or BK channels by drugs capable to specifically open at nanomolar concentrations the skeletal muscle subtypes with less side effects. PMID:21687503

Evaluation of potassium permanganate against an experimental subacute infection of Flavobacterium columnare in channel catfish, Icatlurus punctatus

USDA-ARS?s Scientific Manuscript database

The efficacy of potassium permanganate (KMnO4) as a prophylactic and therapeutic treatment for subacute infection of Flavobacterium columnare was demonstrated in experimentally infected channel catfish, Ictalurus punctatus. Catfish experimentally infected with F. columnare to mimic a subacute infec...

Effectiveness of copper sulfate and potassium permanganate on channel catfish infected with Flavobacterium columnare

USDA-ARS?s Scientific Manuscript database

Copper sulfate (CuSO4) and potassium permanganate (KMnO4) were evaluated for their effectiveness to curtail mortality and decrease bacterial load in fish tissues and water in channel catfish Ictalurus punctatus naturally infected with Flavobacterium columnare, the causative agent of columnaris. Fis...

The effect of copper sulfate, potassium permanganate, and peracetic acid on Ichthyobodo necator in channel catfish

USDA-ARS?s Scientific Manuscript database

Ichthyobodo necator is a single celled biflagellate that can cause significant mortalities in fish, particularly young, tank-reared fish. Copper sulfate (CuSO4), potassium permanganate (KMnO4) and peracetic acid (PAA) were evaluated for effectiveness against Ichthybodosis in juvenile channel catfis...

Simultaneous optical and electrical recording of a single ion-channel.

PubMed

Ide, Toru; Takeuchi, Yuko; Aoki, Takaaki; Yanagida, Toshio

2002-10-01

In recent years, the single-molecule imaging technique has proven to be a valuable tool in solving many basic problems in biophysics. The technique used to measure single-molecule functions was initially developed to study electrophysiological properties of channel proteins. However, the technology to visualize single channels at work has not received as much attention. In this study, we have for the first time, simultaneously measured the optical and electrical properties of single-channel proteins. The large conductance calcium-activated potassium channel (BK-channel) labeled with fluorescent dye molecules was incorporated into a planar bilayer membrane and the fluorescent image captured with a total internal reflection fluorescence microscope simultaneously with single-channel current recording. This innovative technology will greatly advance the study of channel proteins as well as signal transduction processes that involve ion permeation processes.

Protective effects of phosphodiesterase-1 (PDE1) and ATP sensitive potassium (KATP) channel modulators against 3-nitropropionic acid induced behavioral and biochemical toxicities in experimental Huntington׳s disease.

PubMed

Gupta, Surbhi; Sharma, Bhupesh

2014-06-05

Huntington׳s disease (HD), a devastating neurodegenerative disorder, is characterized by weight loss, impairment of motor function, cognitive dysfunction, neuropsychiatric disturbances and striatal damage. Phosphodiesterase-1 (PDE1) has been implicated in various neurological diseases. Mitochondrial potassium channels in the brain take part in neuroprotection. This study has been structured to investigate the role of vinpocetine, a selective PDE1 inhibitor as well as nicorandil, selective ATP sensitive potassium (KATP) channel opener in 3-nitropropionic acid (3-NP) induced HD symptoms in rats. Systemic administration of 3-NP significantly, reduced body weight, impaired locomotion, grip strength and impaired cognition. 3-NP elicited marked oxidative stress in the brain (enhanced malondialdehyde-MDA, reduced glutathione-GSH content, superoxide dismutase-SOD and catalase-CAT), elevated brain acetylcholinesterase activity and inflammation (myeloperoxidase-MPO), with marked nitrosative stress (nitrite/nitrate) in the brain. 3-NP has also induced mitochondrial dysfunction (impaired mitochondrial NADH dehydrogenase-complex I, succinate dehydrogenase-complex II and cytochrome oxidase-complex IV) activities in the striatum of the rat. Tetrabenazine was used as a positive control. Treatment with vinpocetine, nicorandil and tetrabenazine ameliorated 3-NP induced reduction in body weight, impaired locomotion, grip strength and impaired cognition. Treatment with these drugs reduced brain striatum oxidative (MDA, GSH, SOD and CAT) and nitrosative (nitrite/nitrate) stress, acetylcholinesterase activity, inflammation and mitochondrial dysfunctions. These results indicate that vinpocetine, a selective PDE1 inhibitor and nicorandil, a KATP channel opener have attenuated 3-NP induced experimental HD. Hence, pharmacological modulation of PDE1 as well as KATP channels may be considered as potential research targets for mitigation of HD. Copyright © 2014 Elsevier B.V. All rights reserved.

Kir2 potassium channels in rat striatum are strategically localized to control basal ganglia function.

PubMed

Prüss, Harald; Wenzel, Mareike; Eulitz, Dirk; Thomzig, Achim; Karschin, Andreas; Veh, Rüdiger W

2003-02-20

Parkinson's disease is the most frequent movement disorder caused by loss of dopaminergic neurons in the midbrain. Intentions to avoid side effects of the conventional therapy should aim to identify additional targets for potential pharmacological intervention. In principle, every step of a signal transduction cascade such as presynaptic transmitter release, type and occupation of postsynaptic receptors, G protein-mediated effector mechanisms, and the alterations of pre- or postsynaptic potentials as determined by the local ion channel composition, have to be considered. Due to their diversity and their widespread but distinct localizations, potassium channels represent interesting candidates for new therapeutic strategies. As a first step, the present report aimed to study in the striatum the cellular and subcellular distribution of the individual members of the Kir2 family, a group of proteins forming inwardly rectifying potassium channels. For this purpose polyclonal monospecific affinity-purified antibodies against the less conserved carboxyterminal sequences from the Kir2.1, Kir2.2, Kir2.3, and Kir2.4 proteins were prepared. All subunits of the Kir2 family were detected on somata and dendrites of most striatal neurons. However, the distribution of two of them was not homogeneous. Striatal patch areas were largely devoid of the Kir2.3 protein, and the Kir2.4 subunit was most prominently expressed on the tonically active, giant cholinergic interneurons of the striatum. These two structures are among the key players in regulating dopaminergic and cholinergic neurotransmission within the striatum, and therefore are of major importance for the output of the basal ganglia. The heterogeneous localization of the Kir2.3 and the Kir2.4 subunits with respect to these strategic structures pinpoints to these channel proteins as promising targets for future pharmacological efforts.

Cardiac transient outward potassium current: a pulse chemistry model of frequency-dependent properties.

PubMed

Liu, L; Krinsky, V I; Grant, A O; Starmer, C F

1996-01-01

Recent voltage-clamp studies of isolated myocytes have demonstrated widespread occurrence of a transient outward current (I(to)) carried by potassium ions. In the canine ventricle, this current is well developed in epicardial cells but not in endocardial cells. The resultant spatial dispersion of refractoriness is potentially proarrhythmic and may be amplified by channel blockade. The inactivation and recovery time constants of this channel are in excess of several hundred milliseconds, and consequently channel availability is frequency dependent at physiological stimulation rates. When the time constants associated with transitions between different channel conformations are rapid relative to drug binding kinetics, the interactions between drugs and an ion channel can be approximated by a sequence of first-order reactions, in which binding occurs in pulses in response to pulse train stimulation (pulse chemistry). When channel conformation transition time constants do not meet this constraint, analytical characterizations of the drug-channel interaction must then be modified to reflect the channel time-dependent properties. Here we report that the rate and steady-state amount of frequency-dependent inactivation of I(to) are consistent with a generalization of the channel blockade model: channel availability is reduced in a pulsatile exponential pattern as the stimulation frequency is increased, and the rate of reduction is a linear function of the pulse train depolarizing and recovery intervals. I(to) was reduced in the presence of quinidine. After accounting for the use-dependent availability of I(to) channels, we found little evidence of an additional use-dependent component of block after exposure to quinidine, suggesting that quinidine reacts with both open and closed I(to) channels as though the binding site is continuously accessible. The model provides a useful tool for assessing drug-channel interactions when the reaction cannot be continuously monitored.

BK channel β1 subunits regulate airway contraction secondary to M2 muscarinic acetylcholine receptor mediated depolarization.

PubMed

Semenov, Iurii; Wang, Bin; Herlihy, Jeremiah T; Brenner, Robert

2011-04-01

The large conductance calcium- and voltage-activated potassium channel (BK channel) and its smooth muscle-specific β1 subunit regulate excitation–contraction coupling in many types of smooth muscle cells. However, the relative contribution of BK channels to control of M2- or M3-muscarinic acetylcholine receptor mediated airway smooth muscle contraction is poorly understood. Previously, we showed that knockout of the BK channel β1 subunit enhances cholinergic-evoked trachea contractions. Here, we demonstrate that the enhanced contraction of the BK β1 knockout can be ascribed to a defect in BK channel opposition of M2 receptor-mediated contractions. Indeed, the enhanced contraction of β1 knockout is eliminated by specific M2 receptor antagonism. The role of BK β1 to oppose M2 signalling is evidenced by a greater than fourfold increase in the contribution of L-type voltage-dependent calcium channels to contraction that otherwise does not occur with M2 antagonist or with β1 containing BK channels. The mechanism through which BK channels oppose M2-mediated recruitment of calcium channels is through a negative shift in resting voltage that offsets, rather than directly opposes, M2-mediated depolarization. The negative shift in resting voltage is reduced to similar extents by BK β1 knockout or by paxilline block of BK channels. Normalization of β1 knockout baseline voltage with low external potassium eliminated the enhanced M2-receptor mediated contraction. In summary, these findings indicate that an important function of BK/β1 channels is to oppose cholinergic M2 receptor-mediated depolarization and activation of calcium channels by restricting excitation–contraction coupling to more negative voltage ranges.

BK channel β1 subunits regulate airway contraction secondary to M2 muscarinic acetylcholine receptor mediated depolarization

PubMed Central

Semenov, Iurii; Wang, Bin; Herlihy, Jeremiah T; Brenner, Robert

2011-01-01

Abstract The large conductance calcium- and voltage-activated potassium channel (BK channel) and its smooth muscle-specific β1 subunit regulate excitation–contraction coupling in many types of smooth muscle cells. However, the relative contribution of BK channels to control of M2- or M3-muscarinic acetylcholine receptor mediated airway smooth muscle contraction is poorly understood. Previously, we showed that knockout of the BK channel β1 subunit enhances cholinergic-evoked trachea contractions. Here, we demonstrate that the enhanced contraction of the BK β1 knockout can be ascribed to a defect in BK channel opposition of M2 receptor-mediated contractions. Indeed, the enhanced contraction of β1 knockout is eliminated by specific M2 receptor antagonism. The role of BK β1 to oppose M2 signalling is evidenced by a greater than fourfold increase in the contribution of L-type voltage-dependent calcium channels to contraction that otherwise does not occur with M2 antagonist or with β1 containing BK channels. The mechanism through which BK channels oppose M2-mediated recruitment of calcium channels is through a negative shift in resting voltage that offsets, rather than directly opposes, M2-mediated depolarization. The negative shift in resting voltage is reduced to similar extents by BK β1 knockout or by paxilline block of BK channels. Normalization of β1 knockout baseline voltage with low external potassium eliminated the enhanced M2-receptor mediated contraction. In summary, these findings indicate that an important function of BK/β1 channels is to oppose cholinergic M2 receptor-mediated depolarization and activation of calcium channels by restricting excitation–contraction coupling to more negative voltage ranges. PMID:21300746

Identification of quaternary ammonium compounds as potent inhibitors of hERG potassium channels

PubMed Central

Xia, Menghang; Shahane, Sampada; Huang, Ruili; Titus, Steven A.; Shum, Enoch; Zhao, Yong; Southall, Noel; Zheng, Wei; Witt, Kristine L.; Tice, Raymond R.; Austin, Christopher P.

2011-01-01

The human ether-a-go-go-related gene (hERG) channel, a member of a family of voltage-gated potassium (K+) channels, plays a critical role in the repolarization of the cardiac action potential. The reduction of hERG channel activity as a result of adverse drug effects or genetic mutations may cause QT interval prolongation and potentially lead to acquired long QT syndrome. Thus, screening for hERG channel activity is important in drug development. Cardiotoxicity associated with the inhibition of hERG channels by environmental chemicals is also a public health concern. To assess the inhibitory effects of environmental chemicals on hERG channel function, we screened the National Toxicology Program (NTP) collection of 1408 compounds by measuring thallium influx into cells through hERG channels. Seventeen compounds with hERG channel inhibition were identified with IC50 potencies ranging from 0.26 to 22 μM. Twelve of these compounds were confirmed as hERG channel blockers in an automated whole cell patch clamp experiment. In addition, we investigated the structure-activity relationship of seven compounds belonging to the quaternary ammonium compound (QAC) series on hERG channel inhibition. Among four active QAC compounds, tetra-n-octylammonium bromide was the most potent with an IC50 value of 260 nM in the thallium influx assay and 80 nM in the patch clamp assay. The potency of this class of hERG channel inhibitors appears to depend on the number and length of their aliphatic side-chains surrounding the charged nitrogen. Profiling environmental compound libraries for hERG channel inhibition provides information useful in prioritizing these compounds for cardiotoxicity assessment in vivo. PMID:21362439

Updating In Vivo and In Vitro Phosphorylation and Methylation Sites of Voltage-Gated Kv7.2 Potassium Channels.

PubMed

Erdem, Fatma Asli; Salzer, Isabella; Heo, Seok; Chen, Wei-Qiang; Jung, Gangsoo; Lubec, Gert; Boehm, Stefan; Yang, Jae-Won

2017-10-01

Voltage-gated Kv7.2 potassium channels regulate neuronal excitability. The gating of these channels is tightly controlled by various mediators and neurotransmitters acting via G protein-coupled receptors; the underlying signaling cascades involve phosphatidylinositol-4,5-bisphosphate (PIP 2 ), Ca 2+ /calmodulin, and phosphorylation. Recent studies found that the PIP 2 sensitivity of Kv7.2 channels is affected by two posttranslational modifications, phosphorylation and methylation, harboured within putative PIP 2 -binding domains. In this study, we updated phosphorylation and methylation sites in Kv7.2 either heterologously expressed in mammalian cells or as GST-fusion proteins exposed to recombinant protein kinases by using LC-MS/MS. In vitro kinase assays revealed that CDK5, protein kinase C (PKC) alpha, PKA, p38 MAPK, CamKIIα, and GSK3β could mediate phosphorylation. Taken together, we provided a comprehensive map of phosphorylation and methylation in Kv7.2 within protein-protein and protein-lipid interaction domains. This may help to interpret the functional roles of individual PTM sites in Kv7.2 channels. All MS data are available via ProteomeXchange with the identifier PXD005567. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biophysical Properties of ATP-sensitive Potassium Channels in CA3 Hippocampal Neurons

NASA Astrophysics Data System (ADS)

Obregón-Herrera, Armando; Márquez-Gamiño, Sergio; Onetti, Carlos G.

2004-09-01

Single-channel activity of glucose-sensitive channels from CA3 neurons of the rat hippocampus, was studied in cell-attached membrane patches. Single-channel activity was totally abolished at 20 mM external glucose. Glucose-sensitive channels were selective to K+ ions; the unitary conductance was 170 pS in 140 mM K+, and the K+ permeability was 3.86×10-13 cmṡs-1. The open-state probability (PO) increased with membrane depolarization as a result of mean open time enhancement and shortening of the closure periods. The activation midpoint was -79 mV. Glucose-sensitive K+ channel of CA3 neurons could be considered as an ATP-sensitive potassium channel.

Potassium dynamics and seizures: Why is potassium ictogenic?

PubMed

de Curtis, Marco; Uva, Laura; Gnatkovsky, Vadym; Librizzi, Laura

2018-07-01

Potassium channels dysfunction and altered genes encoding for molecules involved in potassium homeostasis have been associated with human epilepsy. These observations are in agreement with a control role of extracellular potassium on neuronal excitability and seizure generation. Epileptiform activity, in turn, regulates potassium homeostasis through mechanisms that are still not well established. We review here how potassium-associated processes are regulated in the brain and examine the mechanisms that support the role of potassium in triggering epileptiform activities. Copyright © 2018 Elsevier B.V. All rights reserved.

How does KCNE1 regulate the Kv7.1 potassium channel? Model-structure, mutations, and dynamics of the Kv7.1-KCNE1 complex.

PubMed

Gofman, Yana; Shats, Simona; Attali, Bernard; Haliloglu, Turkan; Ben-Tal, Nir

2012-08-08

The voltage-gated potassium channel Kv7.1 and its auxiliary subunit KCNE1 are expressed in the heart and give rise to the major repolarization current. The interaction of Kv7.1 with the single transmembrane helix of KCNE1 considerably slows channel activation and deactivation, raises single-channel conductance, and prevents slow voltage-dependent inactivation. We built a Kv7.1-KCNE1 model-structure. The model-structure agrees with previous disulfide mapping studies and enables us to derive molecular interpretations of electrophysiological recordings that we obtained for two KCNE1 mutations. An elastic network analysis of Kv7.1 fluctuations in the presence and absence of KCNE1 suggests a mechanistic perspective on the known effects of KCNE1 on Kv7.1 function: slow deactivation is attributed to the low mobility of the voltage-sensor domains upon KCNE1 binding, abolishment of voltage-dependent inactivation could result from decreased fluctuations in the external vestibule, and amalgamation of the fluctuations in the pore region is associated with enhanced ion conductivity. Copyright © 2012 Elsevier Ltd. All rights reserved.

The polarization of the G-protein activated potassium channel GIRK5 to the vegetal pole of Xenopus laevis oocytes is driven by a di-leucine motif.

PubMed

Díaz-Bello, Beatriz; Rangel-García, Claudia I; Salvador, Carolina; Carrisoza-Gaytán, Rolando; Escobar, Laura I

2013-01-01

The G protein-coupled inwardly-rectifying potassium channels (known as GIRK or Kir3) form functional heterotetramers gated by G-βγ subunits. GIRK channels participate in heart rate modulation and neuronal postsynaptic inhibition in mammals. In Xenopus laevis oocytes, GIRK5 is a functional homomultimer. Previously, we found that phosphorylation of a tyrosine (Y16) at its N-terminus downregulates the surface expression of GIRK5. In this work, we elucidated the subcellular localization and trafficking of GIRK5 in oocytes. Several EGFP-GIRK5 chimeras were produced and an ECFP construct was used to identify the endoplasmic reticulum (ER). Whereas GIRK5-WT was retained in the ER at the animal pole, the phospho-null GIRK5-Y16A was localized to the vegetal pole. Interestingly, a construct with an N-terminal Δ25 deletion produced an even distribution of the channel in the whole oocyte. Through an alanine-scan, we identified an acidic cluster/di-leucine sorting-signal recognition motif between E17 and I22. We quantified the effect of each amino acid residue within this di-leucine motif in determining the distribution of GIRK5 to the animal and vegetal poles. We found that Y16 and I22 contributed to functional expression and were dominant in the polarization of GIRK5. We thus conclude that the N-terminal acidic di-leucine motif of GIRK5 determines its retention and polarized trafficking within Xl oocytes.

The Polarization of the G-Protein Activated Potassium Channel GIRK5 to the Vegetal Pole of Xenopus laevis Oocytes Is Driven by a Di-Leucine Motif

PubMed Central

Díaz-Bello, Beatriz; Rangel-García, Claudia I.; Salvador, Carolina; Carrisoza-Gaytán, Rolando; Escobar, Laura I.

2013-01-01

The G protein-coupled inwardly-rectifying potassium channels (known as GIRK or Kir3) form functional heterotetramers gated by G-βγ subunits. GIRK channels participate in heart rate modulation and neuronal postsynaptic inhibition in mammals. In Xenopus laevis oocytes, GIRK5 is a functional homomultimer. Previously, we found that phosphorylation of a tyrosine (Y16) at its N-terminus downregulates the surface expression of GIRK5. In this work, we elucidated the subcellular localization and trafficking of GIRK5 in oocytes. Several EGFP-GIRK5 chimeras were produced and an ECFP construct was used to identify the endoplasmic reticulum (ER). Whereas GIRK5-WT was retained in the ER at the animal pole, the phospho-null GIRK5-Y16A was localized to the vegetal pole. Interestingly, a construct with an N-terminal Δ25 deletion produced an even distribution of the channel in the whole oocyte. Through an alanine-scan, we identified an acidic cluster/di-leucine sorting-signal recognition motif between E17 and I22. We quantified the effect of each amino acid residue within this di-leucine motif in determining the distribution of GIRK5 to the animal and vegetal poles. We found that Y16 and I22 contributed to functional expression and were dominant in the polarization of GIRK5. We thus conclude that the N-terminal acidic di-leucine motif of GIRK5 determines its retention and polarized trafficking within Xl oocytes. PMID:23717539

Identification of potential novel interaction partners of the sodium-activated potassium channels Slick and Slack in mouse brain.

PubMed

Rizzi, Sandra; Schwarzer, Christoph; Kremser, Leopold; Lindner, Herbert H; Knaus, Hans-Günther

2015-12-01

The sodium-activated potassium channels Slick (Slo2.1, KCNT2) and Slack (Slo2.2, KCNT1) are paralogous channels of the Slo family of high-conductance potassium channels. Slick and Slack channels are widely distributed in the mammalian CNS and they play a role in slow afterhyperpolarization, generation of depolarizing afterpotentials and in setting and stabilizing the resting potential. In the present study we used a combined approach of (co)-immunoprecipitation studies, Western blot analysis, double immunofluorescence and mass spectrometric sequencing in order to investigate protein-protein interactions of the Slick and Slack channels. The data strongly suggest that Slick and Slack channels co-assemble into identical cellular complexes. Double immunofluorescence experiments revealed that Slick and Slack channels co-localize in distinct mouse brain regions. Moreover, we identified the small cytoplasmic protein beta-synuclein and the transmembrane protein 263 (TMEM 263) as novel interaction partners of both, native Slick and Slack channels. In addition, the inactive dipeptidyl-peptidase (DPP 10) and the synapse associated protein 102 (SAP 102) were identified as constituents of the native Slick and Slack channel complexes in the mouse brain. This study presents new insights into protein-protein interactions of native Slick and Slack channels in the mouse brain.

Disruption of Inhibitory Function in the Ts65Dn Mouse Hippocampus Through Overexpression of GIRK2

DTIC Science & Technology

2007-10-24

are prominent (Galdzicki and Siarey, 2003). We found that GIRK2 mRNA and protein subunits are highly overexpressed in multiple CNS structures ... STRUCTURE GIRK channels are members of the large family of potassium inward rectifiers (Kir). The seven subfamilies of Kir channels (Kir1-7) differ as...This ability to discriminate against the smaller Na+ (atomic radius: 0.95 Å) was elucidated by examining the pore structure of the bacterial KcsA

A conserved threonine in the S1-S2 loop of KV7.2 and K V7.3 channels regulates voltage-dependent activation.

PubMed

Füll, Yvonne; Seebohm, Guiscard; Lerche, Holger; Maljevic, Snezana

2013-06-01

The voltage-gated potassium channels KV7.2 and KV7.3 (KCNQ2/3 genes) play an important role in regulating neuronal excitability. More than 50 KCNQ2/3 mutations have been identified to cause an inherited form of epilepsy in newborns. For two of those (E119G and S122L) found in the S1-S2 region of KV7.2, we previously showed a decreased channel availability mainly at action potential subthreshold voltages caused by a slight depolarizing shift of the activation curve. Interestingly, recent studies revealed that a threonine residue within the S1-S2 loop, highly conserved among different classes of KV channels, is crucial for both their function and surface expression. To investigate the functional role of the homologous threonine residues in KV7.2 (T114) and KV7.3 (T144) channels, we replaced them with alanine and examined the electrophysiological properties using heterologous expression in CHO cells and whole cell patch clamping. Channels comprising mutant subunits yielded decreased potassium currents with slowed activation and accelerated deactivation kinetics. However, the most striking effect was a depolarizing shift in the voltage dependence of activation reaching +30 mV upon co-expression of both mutant subunits. Potential interactions of T114 within the channel were analyzed by creating a 3D homology model of KV7.2 in an open state suggesting that this residue plays a central role in the formation of a stable interface between the S1-S2 and the S5 segment helices. This could be the explanation why substitution of the conserved threonine in KV7.2 and KV7.3 channels destabilizes the open and favors the closed state of these channels.

Loss of Sodium-Activated Potassium Channel Slack and FMRP Differentially Affect Social Behavior in Mice.

PubMed

Bausch, Anne E; Ehinger, Rebekka; Straubinger, Julia; Zerfass, Patrick; Nann, Yvette; Lukowski, Robert

2018-05-31

The sodium-activated potassium channel Slack (Slo2.2) is widely expressed in central and peripheral neurons where it is supposed to shape firing properties important for neuronal excitability. Slack activity is enhanced by interaction with the Fragile-X-Mental-Retardation-Protein (FMRP) and loss of FMRP leads to decreased sodium-activated potassium currents in medial nucleus of the trapezoid body neurons of the Fmr1-knockout (KO) mouse representing a mouse model of the human Fragile-X-Syndrome (FXS) and autism. Autism is a frequent comorbidity of FXS, but it is unclear whether Slack is involved in autistic or related conditions of FXS in vivo. By applying a wide range of behavioral tests, we compared social and autism-related behaviors in Slack- and FMRP-deficient mice. In our hands, as expected, FMRP-deficiency causes autism-related behavioral changes in nesting and in a marble-burying test. In contrast, Slack-deficient males exhibited specific abnormalities in sociability in direct and indirect social interaction tests. Hence, we show for the first time that a proper Slack channel function is mandatory for normal social behavior in mice. Nevertheless, as deficits in social behaviors seem to occur independently from each other in FMRP and Slack null mutants, we conclude that Slack is not involved in the autistic phenotype of FMRP KO mice. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Ion Conduction through the hERG Potassium Channel

PubMed Central

Cavalli, Andrea; Recanatini, Maurizio

2012-01-01

The inward rectifier voltage-gated potassium channel hERG is of primary importance for the regulation of the membrane potential of cardiomyocytes. Unlike most voltage-gated K+-channels, hERG shows a low elementary conductance at physiological voltage and potassium concentration. To investigate the molecular features underlying this unusual behavior, we simulated the ion conduction through the selectivity filter at a fully atomistic level by means of molecular dynamics-based methods, using a homology-derived model. According to our calculations, permeation of potassium ions can occur along two pathways, one involving site vacancies inside the filter (showing an energy barrier of about 6 kcal mol−1), and the other characterized by the presence of a knock-on intermediate (about 8 kcal mol−1). These barriers are indeed in accordance with a low conductance behavior, and can be explained in terms of a series of distinctive structural features displayed by the hERG ion permeation pathway. PMID:23133669

Localization and function of ATP-sensitive potassium channels in human skeletal muscle.

PubMed

Nielsen, Jens Jung; Kristensen, Michael; Hellsten, Ylva; Bangsbo, Jens; Juel, Carsten

2003-02-01

The present study investigated the localization of ATP-sensitive K+ (KATP) channels in human skeletal muscle and the functional importance of these channels for human muscle K+ distribution at rest and during muscle activity. Membrane fractionation based on the giant vesicle technique or the sucrose-gradient technique in combination with Western blotting demonstrated that the KATP channels are mainly located in the sarcolemma. This localization was confirmed by immunohistochemical measurements. With the microdialysis technique, it was demonstrated that local application of the KATP channel inhibitor glibenclamide reduced (P < 0.05) interstitial K+ at rest from approximately 4.5 to 4.0 mM, whereas the concentration in the control leg remained constant. Glibenclamide had no effect on the interstitial K+ accumulation during knee-extensor exercise at a power output of 60 W. In contrast to in vitro conditions, the present study demonstrated that under in vivo conditions the KATP channels are active at rest and contribute to the accumulation of interstitial K+.

hERG1/Kv11.1 activation stimulates transcription of p21waf/cip in breast cancer cells via a calcineurin-dependent mechanism.

PubMed

Perez-Neut, Mathew; Rao, Vidhya R; Gentile, Saverio

2016-09-13

The function of Kv11.1 is emerging in breast cancer biology, as a growing body of evidence indicates that the hERG1/Kv11.1 potassium channel is aberrantly expressed in several cancer types including breast cancers.The biological effects of Kv11.1 channel blockers and their associated side effects are very well known but the potential use of Kv11.1 activators as an anticancer strategy are still unexplored. In our previous work, we have established that stimulation of the Kv11.1 potassium channel activates a senescent-like program that is characterized by a significant increase in tumor suppressor protein levels, such as p21waf/cip and p16INK4A. In this study we investigated the mechanism linking Kv11.1 stimulation to augmentation of p21waf/cip protein level. We have demonstrated that the Kv11.1 channel activator NS1643 activates a calcineurin-dependent transcription of p21waf/cip and that this event is fundamental for the inhibitory effect of NS1643 on cell proliferation. Our results reveal a novel mechanism by which stimulation of Kv11.1 channel leads to transcription of a potent tumor suppressor and suggest a potential therapeutic use for Kv11.1 channel activators.

hERG1/Kv11.1 activation stimulates transcription of p21waf/cip in breast cancer cells via a calcineurin-dependent mechanism

PubMed Central

Perez-Neut, Mathew; Rao, Vidhya R.; Gentile, Saverio

2016-01-01

The function of Kv11.1 is emerging in breast cancer biology, as a growing body of evidence indicates that the hERG1/Kv11.1 potassium channel is aberrantly expressed in several cancer types including breast cancers. The biological effects of Kv11.1 channel blockers and their associated side effects are very well known but the potential use of Kv11.1 activators as an anticancer strategy are still unexplored. In our previous work, we have established that stimulation of the Kv11.1 potassium channel activates a senescent-like program that is characterized by a significant increase in tumor suppressor protein levels, such as p21waf/cip and p16INK4A. In this study we investigated the mechanism linking Kv11.1 stimulation to augmentation of p21waf/cip protein level. We have demonstrated that the Kv11.1 channel activator NS1643 activates a calcineurin-dependent transcription of p21waf/cip and that this event is fundamental for the inhibitory effect of NS1643 on cell proliferation. Our results reveal a novel mechanism by which stimulation of Kv11.1 channel leads to transcription of a potent tumor suppressor and suggest a potential therapeutic use for Kv11.1 channel activators. PMID:25945833

Dissection of K+ currents in Caenorhabditis elegans muscle cells by genetics and RNA interference

PubMed Central

Santi, C. M.; Yuan, A.; Fawcett, G.; Wang, Z.-W.; Butler, A.; Nonet, M. L.; Wei, A.; Rojas, P.; Salkoff, L.

2003-01-01

GFP-promoter experiments have previously shown that at least nine genes encoding potassium channel subunits are expressed in Caenorhabditis elegans muscle. By using genetic, RNA interference, and physiological techniques we revealed the molecular identity of the major components of the outward K+ currents in body wall muscle cells in culture. We found that under physiological conditions, outward current is dominated by the products of only two genes, Shaker (Kv1) and Shal (Kv4), both expressing voltage-dependent potassium channels. Other channels may be held in reserve to respond to particular circumstances. Because GFP-promoter experiments indicated that slo-2 expression is prominent, we created a deletion mutant to identify the SLO-2 current in vivo. In both whole-cell and single-channel modes, in vivo SLO-2 channels were active only when intracellular Ca2+ and Cl- were raised above normal physiological conditions, as occurs during hypoxia. Under such conditions, SLO-2 is the largest outward current, contributing up to 87% of the total current. Other channels are present in muscle, but our results suggest that they are unlikely to contribute a large outward component under physiological conditions. However, they, too, may contribute currents conditional on other factors. Hence, the picture that emerges is of a complex membrane with a small number of household conductances functioning under normal circumstances, but with additional conductances that are activated during unusual circumstances. PMID:14612577

Synergistic activation of G protein-gated inwardly rectifying potassium channels by cholesterol and PI(4,5)P2.

PubMed

Bukiya, Anna N; Rosenhouse-Dantsker, Avia

2017-07-01

G-protein gated inwardly rectifying potassium (GIRK or Kir3) channels play a major role in the control of the heart rate, and require the membrane phospholipid phosphatidylinositol-bis-phosphate (PI(4,5)P 2 ) for activation. Recently, we have shown that the activity of the heterotetrameric Kir3.1/Kir3.4 channel that underlies atrial K ACh currents was enhanced by cholesterol. Similarly, the activities of both the Kir3.4 homomer and its active pore mutant Kir3.4* (Kir3.4_S143T) were also enhanced by cholesterol. Here we employ planar lipid bilayers to investigate the crosstalk between PI(4,5)P 2 and cholesterol, and demonstrate that these two lipids act synergistically to activate Kir3.4* currents. Further studies using the Xenopus oocytes heterologous expression system suggest that PI(4,5)P 2 and cholesterol act via distinct binding sites. Whereas PI(4,5)P 2 binds to the cytosolic domain of the channel, the putative binding region of cholesterol is located at the center of the transmembrane domain overlapping the central glycine hinge region of the channel. Together, our data suggest that changes in the levels of two key membrane lipids - cholesterol and PI(4,5)P 2 - could act in concert to provide fine-tuning of Kir3 channel function. Copyright © 2017 Elsevier B.V. All rights reserved.

Deletion of the Kv2.1 delayed rectifier potassium channel leads to neuronal and behavioral hyperexcitability

PubMed Central

Speca, David J.; Ogata, Genki; Mandikian, Danielle; Bishop, Hannah I.; Wiler, Steve W.; Eum, Kenneth; Wenzel, H. Jürgen; Doisy, Emily T.; Matt, Lucas; Campi, Katharine L.; Golub, Mari S.; Nerbonne, Jeanne M.; Hell, Johannes W.; Trainor, Brian C.; Sack, Jon T.; Schwartzkroin, Philip A.; Trimmer, James S.

2014-01-01

The Kv2.1 delayed rectifier potassium channel exhibits high-level expression in both principal and inhibitory neurons throughout the central nervous system, including prominent expression in hippocampal neurons. Studies of in vitro preparations suggest that Kv2.1 is a key yet conditional regulator of intrinsic neuronal excitability, mediated by changes in Kv2.1 expression, localization and function via activity-dependent regulation of Kv2.1 phosphorylation. Here we identify neurological and behavioral deficits in mutant (Kv2.1−/−) mice lacking this channel. Kv2.1−/− mice have grossly normal characteristics. No impairment in vision or motor coordination was apparent, although Kv2.1−/− mice exhibit reduced body weight. The anatomic structure and expression of related Kv channels in the brains of Kv2.1−/− mice appears unchanged. Delayed rectifier potassium current is diminished in hippocampal neurons cultured from Kv2.1−/− animals. Field recordings from hippocampal slices of Kv2.1−/− mice reveal hyperexcitability in response to the convulsant bicuculline, and epileptiform activity in response to stimulation. In Kv2.1−/− mice, long-term potentiation at the Schaffer collateral – CA1 synapse is decreased. Kv2.1−/− mice are strikingly hyperactive, and exhibit defects in spatial learning, failing to improve performance in a Morris Water Maze task. Kv2.1−/− mice are hypersensitive to the effects of the convulsants flurothyl and pilocarpine, consistent with a role for Kv2.1 as a conditional suppressor of neuronal activity. Although not prone to spontaneous seizures, Kv2.1−/− mice exhibit accelerated seizure progression. Together, these findings suggest homeostatic suppression of elevated neuronal activity by Kv2.1 plays a central role in regulating neuronal network function. PMID:24494598

Three C-terminal residues from the sulphonylurea receptor contribute to the functional coupling between the KATP channel subunits SUR2A and Kir6.2

PubMed Central

Dupuis, Julien P; Revilloud, Jean; Moreau, Christophe J; Vivaudou, Michel

2008-01-01

Cardiac ATP-sensitive potassium (KATP) channels are metabolic sensors formed by the association of the inward rectifier potassium channel Kir6.2 and the sulphonylurea receptor SUR2A. SUR2A adjusts channel gating as a function of intracellular ATP and ADP and is the target of pharmaceutical openers and blockers which, respectively, up- and down-regulate Kir6.2. In an effort to understand how effector binding to SUR2A translates into Kir6.2 gating modulation, we examined the role of a 65-residue SUR2A fragment linking transmembrane domain TMD2 and nucleotide-binding domain NBD2 that has been shown to interact with Kir6.2. This fragment of SUR2A was replaced by the equivalent residues of its close homologue, the multidrug resistance protein MRP1. The chimeric construct was expressed in Xenopus oocytes and characterized using the patch-clamp technique. We found that activation by MgADP and synthetic openers was greatly attenuated although apparent affinities were unchanged. Further chimeragenetic and mutagenetic studies showed that mutation of three residues, E1305, I1310 and L1313 (rat numbering), was sufficient to confer this defective phenotype. The same mutations had no effects on channel block by the sulphonylurea glibenclamide or by ATP, suggesting a role for these residues in activatory – but not inhibitory – transduction processes. These results indicate that, within the KATP channel complex, the proximal C-terminal of SUR2A is a critical link between ligand binding to SUR2A and Kir6.2 up-regulation. PMID:18450778

Comparative effects of copper sulfate or potassium permanganate on channel catfish concurrently infected with Flavobacterium columnare and Ichthyobodo necator

USDA-ARS?s Scientific Manuscript database

An opportunistic study was conducted to determine the effects of two chemical therapeutants on channel catfish (CCF) Ictalurus punctatus concurrently infected Flavobacterium columnare and Ichthyobodo necator. Copper sulfate (CuSO4) and potassium permanganate (KMnO4) were investigated for their abil...

Treating a natural outbreak of columnaris in channel catfish with copper sulfate and potassium permanganate

USDA-ARS?s Scientific Manuscript database

An F. Columnare-exclusive epizootic occurred in fingerling channel catfish (Ictalurus punctatus) during normal tank culture practices at SNARC. Fish were transferred to the ultra low-flow system and 2.1 mg/L copper sulfate or 3 mg/L potassium permanganate was administered; an untreated control was ...

Loss of Female Sex Hormones Exacerbates Cerebrovascular and Cognitive Dysfunction in Aortic Banded Miniswine Through a Neuropeptide Y-Ca2+-Activated Potassium Channel-Nitric Oxide Mediated Mechanism.

PubMed

Olver, T Dylan; Hiemstra, Jessica A; Edwards, Jenna C; Schachtman, Todd R; Heesch, Cheryl M; Fadel, Paul J; Laughlin, M Harold; Emter, Craig A

2017-10-31

Postmenopausal women represent the largest cohort of patients with heart failure with preserved ejection fraction, and vascular dementia represents the most common form of dementia in patients with heart failure with preserved ejection fraction. Therefore, we tested the hypotheses that the combination of cardiac pressure overload (aortic banding [AB]) and the loss of female sex hormones (ovariectomy [OVX]) impairs cerebrovascular control and spatial memory. Female Yucatan miniswine were separated into 4 groups (n=7 per group): (1) control, (2) AB, (3) OVX, and (4) AB-OVX. Pigs underwent OVX and AB at 7 and 8 months of age, respectively. At 14 months, cerebral blood flow velocity and spatial memory (spatial hole-board task) were lower in the OVX groups ( P <0.05), with significant impairments in the AB-OVX group ( P <0.05). Resting carotid artery β stiffness and vascular resistance during central hypovolemia were increased in the AB-OVX group ( P <0.05), and blood flow recovery after central hypovolemia was reduced in both OVX groups ( P <0.05). Isolated pial artery (pressure myography) vasoconstriction to neuropeptide Y was greatest in the AB-OVX group ( P <0.05), and vasodilation to the Ca 2+ -activated potassium channel α-subunit agonist NS-1619 was impaired in both AB groups ( P <0.05). The ratio of phosphorylated endothelial nitric oxide synthase:total endothelial nitric oxide synthase was depressed and Ca 2+ -activated potassium channel α-subunit protein was increased in AB groups ( P <0.05). Mechanistically, impaired cerebral blood flow control in experimental heart failure may be the result of heightened neuropeptide Y-induced vasoconstriction along with reduced vasodilation associated with decreased Ca 2+ -activated potassium channel function and impaired nitric oxide signaling, the effects of which are exacerbated in the absence of female sex hormones. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

The calcium-activated potassium channel KCa3.1 plays a central role in the chemotactic response of mammalian neutrophils

PubMed Central

Henríquez, C.; Riquelme, T. T.; Vera, D.; Julio-Kalajzić, F.; Ehrenfeld, P.; Melvin, J. E.; Figueroa, C. D.; Sarmiento, J.; Flores, C. A.

2017-01-01

Aim Neutrophils are the first cells to arrive at sites of injury. Nevertheless, many inflammatory diseases are characterized by an uncontrolled infiltration and action of these cells. Cell migration depends on volume changes that are governed by ion channel activity, but potassium channels in neutrophil have not been clearly identified. We aim to test whether KCa3.1 participates in neutrophil migration and other relevant functions of the cell. Methods Cytometer and confocal measurements to determine changes in cell volume were used. Cells isolated from human, mouse and horse were tested for KCa3.1-dependent chemotaxis. Chemokinetics, calcium handling and release of reactive oxygen species were measured to determine the role of KCa3.1 in those processes. A mouse model was used to test for neutrophil recruitment after acute lung injury in vivo. Results We show for the first time that KCa3.1 is expressed in mammalian neutrophils. When the channel is inhibited by a pharmacological blocker or by genetic silencing, it profoundly affects cell volume regulation, and chemotactic and chemokinetic properties of the cells. We also demonstrated that pharmacological inhibition of KCa3.1 did not affect calcium entry or reactive oxygen species production in neutrophils. Using a mouse model of acute lung injury, we observed that Kca3.1−/− mice are significantly less effective at recruiting neutrophils into the site of inflammation. Conclusions These results demonstrate that KCa3.1 channels are key actors in the migration capacity of neutrophils, and its inhibition did not affect other relevant cellular functions. PMID:26138196

The mechanism of gentisic acid-induced relaxation of the guinea pig isolated trachea: the role of potassium channels and vasoactive intestinal peptide receptors.

PubMed

Cunha, J F; Campestrini, F D; Calixto, J B; Scremin, A; Paulino, N

2001-03-01

We examined some of the mechanisms by which the aspirin metabolite and the naturally occurring metabolite gentisic acid induced relaxation of the guinea pig trachea in vitro. In preparations with or without epithelium and contracted by histamine, gentisic acid caused concentration-dependent and reproducible relaxation, with mean EC(50) values of 18 microM and E(max) of 100% (N = 10) or 20 microM and E(max) of 92% (N = 10), respectively. The relaxation caused by gentisic acid was of slow onset in comparison to that caused by norepinephrine, theophylline or vasoactive intestinal peptide (VIP). The relative rank order of potency was: salbutamol 7.9 > VIP 7.0 > gentisic acid 4.7 > theophylline 3.7. Gentisic acid-induced relaxation was markedly reduced (24 +/- 7.0, 43 +/- 3.9 and 78 +/- 5.6%) in preparations with elevated potassium concentration in the medium (20, 40 or 80 mM, respectively). Tetraethylammonium (100 microM), a nonselective blocker of the potassium channels, partially inhibited the relaxation response to gentisic acid, while 4-AP (10 microM), a blocker of the voltage potassium channel, inhibited gentisic acid-induced relaxation by 41 +/- 12%. Glibenclamide (1 or 3 microM), at a concentration which markedly inhibited the relaxation induced by the opener of ATP-sensitive K(+) channels, levcromakalim, had no effect on the relaxation induced by gentisic acid. Charybdotoxin (0.1 or 0.3 microM), a selective blocker of the large-conductance Ca(2+)-activated K(+) channels, caused rightward shifts (6- and 7-fold) of the gentisic acid concentration-relaxation curve. L-N(G)-nitroarginine (100 microM), a NO synthase inhibitor, had no effect on the relaxant effect of gentisic acid, and caused a slight displacement to the right in the relaxant effect of the gentisic acid curve at 300 microM, while methylene blue (10 or 30 microM) or ODQ (1 microM), the inhibitors of soluble guanylate cyclase, all failed to affect gentisic acid-induced relaxation. D-(P)-Cl-Phe(6),Leu(17)[VIP] (0.1 microM), a VIP receptor antagonist, significantly inhibited (37 +/- 7%) relaxation induced by gentisic acid, whereas CGRP (8-37) (0.1 microM), a CGRP antagonist, only slightly enhanced the action of gentisic acid. Taken together, these results provide functional evidence for the direct activation of voltage and large-conductance Ca(+2)-activated K(+) channels, or indirect modulation of potassium channels induced by VIP receptors and accounts for the predominant relaxation response caused by gentisic acid in the guinea pig trachea.

Adenosine Monophosphate Binding Stabilizes the KTN Domain of the Shewanella denitrificans Kef Potassium Efflux System.

PubMed

Pliotas, Christos; Grayer, Samuel C; Ekkerman, Silvia; Chan, Anthony K N; Healy, Jess; Marius, Phedra; Bartlett, Wendy; Khan, Amjad; Cortopassi, Wilian A; Chandler, Shane A; Rasmussen, Tim; Benesch, Justin L P; Paton, Robert S; Claridge, Timothy D W; Miller, Samantha; Booth, Ian R; Naismith, James H; Conway, Stuart J

2017-08-15

Ligand binding is one of the most fundamental properties of proteins. Ligand functions fall into three basic types: substrates, regulatory molecules, and cofactors essential to protein stability, reactivity, or enzyme-substrate complex formation. The regulation of potassium ion movement in bacteria is predominantly under the control of regulatory ligands that gate the relevant channels and transporters, which possess subunits or domains that contain Rossmann folds (RFs). Here we demonstrate that adenosine monophosphate (AMP) is bound to both RFs of the dimeric bacterial Kef potassium efflux system (Kef), where it plays a structural role. We conclude that AMP binds with high affinity, ensuring that the site is fully occupied at all times in the cell. Loss of the ability to bind AMP, we demonstrate, causes protein, and likely dimer, instability and consequent loss of function. Kef system function is regulated via the reversible binding of comparatively low-affinity glutathione-based ligands at the interface between the dimer subunits. We propose this interfacial binding site is itself stabilized, at least in part, by AMP binding.

Adenosine Monophosphate Binding Stabilizes the KTN Domain of the Shewanella denitrificans Kef Potassium Efflux System

PubMed Central

2017-01-01

Ligand binding is one of the most fundamental properties of proteins. Ligand functions fall into three basic types: substrates, regulatory molecules, and cofactors essential to protein stability, reactivity, or enzyme–substrate complex formation. The regulation of potassium ion movement in bacteria is predominantly under the control of regulatory ligands that gate the relevant channels and transporters, which possess subunits or domains that contain Rossmann folds (RFs). Here we demonstrate that adenosine monophosphate (AMP) is bound to both RFs of the dimeric bacterial Kef potassium efflux system (Kef), where it plays a structural role. We conclude that AMP binds with high affinity, ensuring that the site is fully occupied at all times in the cell. Loss of the ability to bind AMP, we demonstrate, causes protein, and likely dimer, instability and consequent loss of function. Kef system function is regulated via the reversible binding of comparatively low-affinity glutathione-based ligands at the interface between the dimer subunits. We propose this interfacial binding site is itself stabilized, at least in part, by AMP binding. PMID:28656748

[Role of ATP-sensitive potassium channel activators in liver mitochondrial function in rats with different resistance to hypoxia].

PubMed

Tkachenko, H M; Kurhaliuk, N M; Vovkanych, L S

2003-01-01

Effects of ATP-sensitive potassium (KATP) channels opener pinacidil (0.06 mg/kg) and inhibitor glibenclamide (1 mg/kg) in rats with different resistance to hypoxia on indices of ADP-stimulation of mitochondrial respiration by Chance, calcium capacity and processes of lipid peroxidation in liver has been investigated. We used next substrates of oxidation: 0.35 mM succinate, 1 mM alpha-ketoglutarate. Additional analyses contain the next inhibitors: mitochondrial fermentative complex I-10 mkM rotenone, succinate dehydrogenase 2 mM malonic acid. It was shown that effects of pinacidil induced the increasing of oxidative phosporylation efficacy and ATP synthesis together with lowering of calcium capacity in rats with low resistance to hypoxia. Effects of pinacidil were leveled by glibenclamide. These changes are connected with the increasing of respiratory rate, calcium overload and intensification of lipid peroxidation processes. A conclusion was made about protective effect of pinacidil on mitochondrial functioning by economization of oxygen-dependent processes, adaptive potentialities of organisms with low resistance to hypoxia being increased.

CNG and HCN channels: two peas, one pod.

PubMed

Craven, Kimberley B; Zagotta, William N

2006-01-01

Cyclic nucleotide-activated ion channels play a fundamental role in a variety of physiological processes. By opening in response to intracellular cyclic nucleotides, they translate changes in concentrations of signaling molecules to changes in membrane potential. These channels belong to two families: the cyclic nucleotide-gated (CNG) channels and the hyperpolarization-activated cyclic nucleotide-modulated (HCN) channels. The two families exhibit high sequence similarity and belong to the superfamily of voltage-gated potassium channels. Whereas HCN channels are activated by voltage and CNG channels are virtually voltage independent, both channels are activated by cyclic nucleotide binding. Furthermore, the channels are thought to have similar channel structures, leading to similar mechanisms of activation by cyclic nucleotides. However, although these channels are structurally and behaviorally similar, they have evolved to perform distinct physiological functions. This review describes the physiological roles and biophysical behavior of CNG and HCN channels. We focus on how similarities in structure and activation mechanisms result in common biophysical models, allowing CNG and HCN channels to be viewed as a single genre.

Magnolol and honokiol regulate the calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli-induced diarrhea mice.

PubMed

Deng, Yanli; Han, Xuefeng; Tang, Shaoxun; Xiao, Wenjun; Tan, Zhiliang; Zhou, Chuanshe; Wang, Min; Kang, Jinghe

2015-05-15

To explore the regulatory mechanisms of magnolol and honokiol on calcium-activated potassium channels signaling pathway in Enterotoxigenic Escherichia coli (ETEC)-induced diarrhea mice, the concentrations of serum chloride ion (Cl(-)), sodium ion (Na(+)), potassium ion (K(+)) and calcium ion (Ca(2+)) were measured. Additionally, the mRNA expressions of calmodulin 1 (CaM), calcium/calmodulin-dependent protein kinase II alpha subunit (CaMKIIα) and beta subunit (CaMKIIβ), ryanodine receptor 1, inositol 1,4,5-trisphosphate receptors (IP3 receptors), protein kinases C (PKC), potassium intermediate/small conductance calcium-activated channels (SK) and potassium large conductance calcium-activated channels(BK)were determined. A diarrhea mouse model was established using ETEC suspensions (3.29×10(9)CFU/ml) at a dosage of 0.02ml/g live body weight (BW). Magnolol or honokiol was intragastrically administered at dosages of 100 (M100 or H100), 300 (M300 or H300) and 500 (M500 or H500) mg/kg BW according to a 3×3 factorial arrangement. Magnolol and honokiol increased the Cl(-) and K(+) concentrations, further, upregulated the CaM, BKα1 and BKβ3 mRNA levels but downregulated the IP3 receptors 1, PKC, SK1, SK2, SK3, SK4 and BKβ4 mRNA expressions. Magnolol and honokiol did not alter the CaMKIIα, CaMKIIβ, ryanodine receptor 1, IP3 receptor 2, IP3 receptor 3, BKβ1 and BKβ2 mRNA expressions. These results clarify that magnolol and honokiol, acting through Ca(2+) channel blockade, inhibit the activation of IP3 receptor 1 to regulate the IP3-Ca(2+) store release, activate CaM to inhibit SK channels, and effectively suppress PKC kinases to promote BKα1 and BKβ3 channels opening and BKβ4 channel closing, which modulates the intestinal ion secretion. Copyright © 2015 Elsevier B.V. All rights reserved.

Computer simulations of transport through membranes: passive diffusion, pores, channels and transporters.

PubMed

Tieleman, D Peter

2006-10-01

A key function of biological membranes is to provide mechanisms for the controlled transport of ions, nutrients, metabolites, peptides and proteins between a cell and its environment. We are using computer simulations to study several processes involved in transport. In model membranes, the distribution of small molecules can be accurately calculated; we are making progress towards understanding the factors that determine the partitioning behaviour in the inhomogeneous lipid environment, with implications for drug distribution, membrane protein folding and the energetics of voltage gating. Lipid bilayers can be simulated at a scale that is sufficiently large to study significant defects, such as those caused by electroporation. Computer simulations of complex membrane proteins, such as potassium channels and ATP-binding cassette (ABC) transporters, can give detailed information about the atomistic dynamics that form the basis of ion transport, selectivity, conformational change and the molecular mechanism of ATP-driven transport. This is illustrated in the present review with recent simulation studies of the voltage-gated potassium channel KvAP and the ABC transporter BtuCD.

Suppression of KV7/KCNQ potassium channel enhances neuronal differentiation of PC12 cells.

PubMed

Zhou, Najing; Huang, Sha; Li, Li; Huang, Dongyang; Yan, Yunli; Du, Xiaona; Zhang, Hailin

2016-10-01

Membrane potential shift driven by electrical activity is critical in determining the cell fate of proliferation or differentiation. As such, the ion channels that underlie the membrane electrical activity play an important role in cell proliferation/differentiation. KV7/KCNQ potassium channels are critical in determining the resting membrane potentials in many neuronal cells. However, the role of these channels in cell differentiation is not well studied. In the present study, we used PC12 cells as well as primary cultured rat cortical neurons to study the role and mechanism of KV7/KCNQ in neuronal differentiation. NGF induced PC12 cell differentiation into neuron-like cells with growth of neurites showing typical growth cone-like extensions. The Kv7/KCNQ blocker XE991 promoted NGF-induced neurite outgrowth, whereas Kv7/KCNQ opener retigabine (RTG) inhibited outgrowth. M-type Kv7 channels are likely involved in regulating neurite growth because overexpression of KCNQ2/Q3 inhibited neurite growth whereas suppression of KCNQ2/Q3 with shRNA promoted neurite growth. Membrane depolarization possibly underpins enhanced neurite growth induced by the suppression of Kv7/KCNQ. Additionally, high extracellular K(+) likely induced membrane depolarization and also promoted neurite growth. Finally, T-type Ca(2+) channels may be involved in membrane-depolarization-induced neurite growth. This study provides a new perspective for understanding neuronal differentiation as well as KV7/KCNQ channel function. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

Inward rectifier potassium (Kir2.1) channels as end‐stage boosters of endothelium‐dependent vasodilators

PubMed Central

Dalsgaard, Thomas; Bonev, Adrian D.; Nelson, Mark T.

2016-01-01

Key points Increase in endothelial cell (EC) calcium activates calcium‐sensitive intermediate and small conductance potassium (IK and SK) channels, thereby causing hyperpolarization and endothelium‐dependent vasodilatation.Endothelial cells express inward rectifier potassium (Kir) channels, but their role in endothelium‐dependent vasodilatation is not clear.In the mesenteric arteries, only ECs, but not smooth muscle cells, displayed Kir currents that were predominantly mediated by the Kir2.1 isoform.Endothelium‐dependent vasodilatations in response to muscarinic receptor, TRPV4 (transient receptor potential vanilloid 4) channel and IK/SK channel agonists were highly attenuated by Kir channel inhibitors and by Kir2.1 channel knockdown.These results point to EC Kir channels as amplifiers of vasodilatation in response to increases in EC calcium and IK/SK channel activation and suggest that EC Kir channels could be targeted to treat endothelial dysfunction, which is a hallmark of vascular disorders. Abstract Endothelium‐dependent vasodilators, such as acetylcholine, increase intracellular Ca2+ through activation of transient receptor potential vanilloid 4 (TRPV4) channels in the plasma membrane and inositol trisphosphate receptors in the endoplasmic reticulum, leading to stimulation of Ca2+‐sensitive intermediate and small conductance K+ (IK and SK, respectively) channels. Although strong inward rectifier K+ (Kir) channels have been reported in the native endothelial cells (ECs) their role in EC‐dependent vasodilatation is not clear. Here, we test the idea that Kir channels boost the EC‐dependent vasodilatation of resistance‐sized arteries. We show that ECs, but not smooth muscle cells, of small mesenteric arteries have Kir currents, which are substantially reduced in EC‐specific Kir2.1 knockdown (EC‐Kir2.1 −/−) mice. Elevation of extracellular K+ to 14 mm caused vasodilatation of pressurized arteries, which was prevented by endothelial denudation and Kir channel inhibitors (Ba2+, ML‐133) or in the arteries from EC‐Kir2.1 −/− mice. Potassium‐induced dilatations were unaffected by inhibitors of TRPV4, IK and SK channels. The Kir channel blocker, Ba2+, did not affect currents through TRPV4, IK or SK channels. Endothelial cell‐dependent vasodilatations in response to activation of muscarinic receptors, TRPV4 channels or IK/SK channels were reduced, but not eliminated, by Kir channel inhibitors or EC‐Kir2.1 −/−. In angiotensin II‐induced hypertension, the Kir channel function was not altered, although the endothelium‐dependent vasodilatation was severely impaired. Our results support the concept that EC Kir2 channels boost vasodilatory signals that are generated by Ca2+‐dependent activation of IK and SK channels. PMID:26840527

The dipole moment of membrane proteins: potassium channel protein and beta-subunit.

PubMed

Takashima, S

2001-12-25

The mechanism of ion channel opening is one of the most fascinating problems in membrane biology. Based on phenomenological studies, early researchers suggested that the elementary process of ion channel opening may be the intramembrane charge movement or the orientation of dipolar proteins in the channel. In spite of the far reaching significance of these hypotheses, it has not been possible to formulate a comprehensive molecular theory for the mechanism of channel opening. This is because of the lack of the detailed knowledge on the structure of channel proteins. In recent years, however, the research on the structure of channel proteins made marked advances and, at present, we are beginning to have sufficient information on the structure of some of the channel proteins, e.g. potassium-channel protein and beta-subunits. With these new information, we are now ready to have another look at the old hypothesis, in particular, the dipole moment of channel proteins being the voltage sensor for the opening and closing of ion channels. In this paper, the dipole moments of potassium channel protein and beta-subunit, are calculated using X-ray diffraction data. A large dipole moment was found for beta-subunits while the dipole moment of K-channel protein was found to be considerably smaller than that of beta-subunits. These calculations were conducted as a preliminary study of the comprehensive research on the dipolar structure of channel proteins in excitable membranes, above all, sodium channel proteins.

Conopeptide Vt3.1 preferentially inhibits BK potassium channels containing β4 subunits via electrostatic interactions.

PubMed

Li, Min; Chang, Shan; Yang, Longjin; Shi, Jingyi; McFarland, Kelli; Yang, Xiao; Moller, Alyssa; Wang, Chunguang; Zou, Xiaoqin; Chi, Chengwu; Cui, Jianmin

2014-02-21

BK channel β subunits (β1-β4) modulate the function of channels formed by slo1 subunits to produce tissue-specific phenotypes. The molecular mechanism of how the homologous β subunits differentially alter BK channel functions and the role of different BK channel functions in various physiologic processes remain unclear. By studying channels expressed in Xenopus laevis oocytes, we show a novel disulfide-cross-linked dimer conopeptide, Vt3.1 that preferentially inhibits BK channels containing the β4 subunit, which is most abundantly expressed in brain and important for neuronal functions. Vt3.1 inhibits the currents by a maximum of 71%, shifts the G-V relation by 45 mV approximately half-saturation concentrations, and alters both open and closed time of single channel activities, indicating that the toxin alters voltage dependence of the channel. Vt3.1 contains basic residues and inhibits voltage-dependent activation by electrostatic interactions with acidic residues in the extracellular loops of the slo1 and β4 subunits. These results suggest a large interaction surface between the slo1 subunit of BK channels and the β4 subunit, providing structural insight into the molecular interactions between slo1 and β4 subunits. The results also suggest that Vt3.1 is an excellent tool for studying β subunit modulation of BK channels and for understanding the physiological roles of BK channels in neurophysiology.

A unique alkaline pH-regulated and fatty acid-activated tandem pore domain potassium channel (K2P) from a marine sponge

PubMed Central

Wells, Gregory D.; Tang, Qiong-Yao; Heler, Robert; Tompkins-MacDonald, Gabrielle J.; Pritchard, Erica N.; Leys, Sally P.; Logothetis, Diomedes E.; Boland, Linda M.

2012-01-01

SUMMARY A cDNA encoding a potassium channel of the two-pore domain family (K2P, KCNK) of leak channels was cloned from the marine sponge Amphimedon queenslandica. Phylogenetic analysis indicated that AquK2P cannot be placed into any of the established functional groups of mammalian K2P channels. We used the Xenopus oocyte expression system, a two-electrode voltage clamp and inside-out patch clamp electrophysiology to determine the physiological properties of AquK2P. In whole cells, non-inactivating, voltage-independent, outwardly rectifying K+ currents were generated by external application of micromolar concentrations of arachidonic acid (AA; EC50 ∼30 μmol l–1), when applied in an alkaline solution (≥pH 8.0). Prior activation of channels facilitated the pH-regulated, AA-dependent activation of AquK2P but external pH changes alone did not activate the channels. Unlike certain mammalian fatty-acid-activated K2P channels, the sponge K2P channel was not activated by temperature and was insensitive to osmotically induced membrane distortion. In inside-out patch recordings, alkalinization of the internal pH (pKa 8.18) activated the AquK2P channels independently of AA and also facilitated activation by internally applied AA. The gating of the sponge K2P channel suggests that voltage-independent outward rectification and sensitivity to pH and AA are ancient and fundamental properties of animal K2P channels. In addition, the membrane potential of some poriferan cells may be dynamically regulated by pH and AA. PMID:22723483

A unique alkaline pH-regulated and fatty acid-activated tandem pore domain potassium channel (K₂P) from a marine sponge.

PubMed

Wells, Gregory D; Tang, Qiong-Yao; Heler, Robert; Tompkins-MacDonald, Gabrielle J; Pritchard, Erica N; Leys, Sally P; Logothetis, Diomedes E; Boland, Linda M

2012-07-15

A cDNA encoding a potassium channel of the two-pore domain family (K(2P), KCNK) of leak channels was cloned from the marine sponge Amphimedon queenslandica. Phylogenetic analysis indicated that AquK(2P) cannot be placed into any of the established functional groups of mammalian K(2P) channels. We used the Xenopus oocyte expression system, a two-electrode voltage clamp and inside-out patch clamp electrophysiology to determine the physiological properties of AquK(2P). In whole cells, non-inactivating, voltage-independent, outwardly rectifying K(+) currents were generated by external application of micromolar concentrations of arachidonic acid (AA; EC(50) ∼30 μmol l(-1)), when applied in an alkaline solution (≥pH 8.0). Prior activation of channels facilitated the pH-regulated, AA-dependent activation of AquK(2P) but external pH changes alone did not activate the channels. Unlike certain mammalian fatty-acid-activated K(2P) channels, the sponge K(2P) channel was not activated by temperature and was insensitive to osmotically induced membrane distortion. In inside-out patch recordings, alkalinization of the internal pH (pK(a) 8.18) activated the AquK(2P) channels independently of AA and also facilitated activation by internally applied AA. The gating of the sponge K(2P) channel suggests that voltage-independent outward rectification and sensitivity to pH and AA are ancient and fundamental properties of animal K(2P) channels. In addition, the membrane potential of some poriferan cells may be dynamically regulated by pH and AA.

Molecular basis of ancestral vertebrate electroreception

PubMed Central

Bellono, Nicholas W.; Leitch, Duncan B.; Julius, David

2017-01-01

Elasmobranch fishes, including sharks, rays, and skates, use specialized electrosensory organs called Ampullae of Lorenzini to detect extremely small changes in environmental electric fields. Electrosensory cells within these ampullae are able to discriminate and respond to minute changes in environmental voltage gradients through an as-yet unknown mechanism. Here we show that the voltage-gated calcium channel CaV1.3 and big conductance calcium-activated potassium (BK) channel are preferentially expressed by electrosensory cells in little skate (Leucoraja erinacea) and functionally couple to mediate electrosensory cell membrane voltage oscillations, which are important in the detection of specific, weak electrical signals. Both channels exhibit unique properties compared with their mammalian orthologues to support electrosensory functions: structural adaptations in CaV1.3 mediate a low voltage threshold for activation, while alterations in BK support specifically tuned voltage oscillations. These findings reveal a molecular basis of electroreception and demonstrate how discrete evolutionary changes in ion channel structure facilitate sensory adaptation. PMID:28264196

KCNE Regulation of K+ Channel Trafficking – a Sisyphean Task?

PubMed Central

Kanda, Vikram A.; Abbott, Geoffrey W.

2012-01-01

Voltage-gated potassium (Kv) channels shape the action potentials of excitable cells and regulate membrane potential and ion homeostasis in excitable and non-excitable cells. With 40 known members in the human genome and a variety of homomeric and heteromeric pore-forming α subunit interactions, post-translational modifications, cellular locations, and expression patterns, the functional repertoire of the Kv α subunit family is monumental. This versatility is amplified by a host of interacting proteins, including the single membrane-spanning KCNE ancillary subunits. Here, examining both the secretory and the endocytic pathways, we review recent findings illustrating the surprising virtuosity of the KCNE proteins in orchestrating not just the function, but also the composition, diaspora and retrieval of channels formed by their Kv α subunit partners. PMID:22754540

Structure of the voltage-gated K⁺ channel Eag1 reveals an alternative voltage sensing mechanism.

PubMed

Whicher, Jonathan R; MacKinnon, Roderick

2016-08-12

Voltage-gated potassium (K(v)) channels are gated by the movement of the transmembrane voltage sensor, which is coupled, through the helical S4-S5 linker, to the potassium pore. We determined the single-particle cryo-electron microscopy structure of mammalian K(v)10.1, or Eag1, bound to the channel inhibitor calmodulin, at 3.78 angstrom resolution. Unlike previous K(v) structures, the S4-S5 linker of Eag1 is a five-residue loop and the transmembrane segments are not domain swapped, which suggest an alternative mechanism of voltage-dependent gating. Additionally, the structure and position of the S4-S5 linker allow calmodulin to bind to the intracellular domains and to close the potassium pore, independent of voltage-sensor position. The structure reveals an alternative gating mechanism for K(v) channels and provides a template to further understand the gating properties of Eag1 and related channels. Copyright © 2016, American Association for the Advancement of Science.

CaV channels and cancer: canonical functions indicate benefits of repurposed drugs as cancer therapeutics.

PubMed

Buchanan, Paul J; McCloskey, Karen D

2016-10-01

The importance of ion channels in the hallmarks of many cancers is increasingly recognised. This article reviews current knowledge of the expression of members of the voltage-gated calcium channel family (Ca V ) in cancer at the gene and protein level and discusses their potential functional roles. The ten members of the Ca V channel family are classified according to expression of their pore-forming α-subunit; moreover, co-expression of accessory α2δ, β and γ confers a spectrum of biophysical characteristics including voltage dependence of activation and inactivation, current amplitude and activation/inactivation kinetics. Ca V channels have traditionally been studied in excitable cells including neurones, smooth muscle, skeletal muscle and cardiac cells, and drugs targeting the channels are used in the treatment of hypertension and epilepsy. There is emerging evidence that several Ca V channels are differentially expressed in cancer cells compared to their normal counterparts. Interestingly, a number of Ca V channels also have non-canonical functions and are involved in transcriptional regulation of the expression of other proteins including potassium channels. Pharmacological studies show that Ca V canonical function contributes to the fundamental biology of proliferation, cell-cycle progression and apoptosis. This raises the intriguing possibility that calcium channel blockers, approved for the treatment of other conditions, could be repurposed to treat particular cancers. Further research will reveal the full extent of both the canonical and non-canonical functions of Ca V channels in cancer and whether calcium channel blockers are beneficial in cancer treatment.

Intracellular segment between transmembrane helices S0 and S1 of BK channel α subunit contains two amphipathic helices connected by a flexible loop

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shi, Pan; High Magnetic Field Laboratory, Chinese Academy of Sciences, Hefei, Anhui, 230031; Li, Dong

2013-08-02

Highlights: •The loop between S0 and S1 of BK channel was overexpressed and purified in DPC. •NMR studies indicated BK-IS1 contained two helices connected by a flexible loop. •Mg{sup 2+} titration of BK-IS1 indicated two possible binding sites of divalent ions. -- Abstract: The BK channel, a tetrameric potassium channel with very high conductance, has a central role in numerous physiological functions. The BK channel can be activated by intracellular Ca{sup 2+} and Mg{sup 2+}, as well as by membrane depolarization. Unlike other tetrameric potassium channels, the BK channel has seven transmembrane helices (S0–S6) including an extra helix S0. Themore » intracellular segment between S0 and S1 (BK-IS1) is essential to BK channel functions and Asp99 in BK-IS1 is reported to be responsible for Mg{sup 2+} coordination. In this study, BK-IS1 (44–113) was over-expressed using a bacterial system and purified in the presence of detergent micelles for multidimensional heteronuclear nuclear magnetic resonance (NMR) structural studies. Backbone resonance assignment and secondary structure analysis showed that BK-IS1 contains two amphipathic helices connected by a 36-residue loop. Amide {sup 1}H–{sup 15}N heteronuclear NOE analysis indicated that the loop is very flexible, while the two amphipathic helices are possibly stabilized through interaction with the membrane. A solution NMR-based titration assay of BK-IS1 was performed with various concentrations of Mg{sup 2+}. Two residues (Thr45 and Leu46) with chemical shift changes were observed but no, or very minor, chemical shift difference was observed for Asp99, indicating a possible site for binding divalent ions or other modulation partners.« less

Potassium intake modulates the thiazide-sensitive sodium-chloride cotransporter (NCC) activity via the Kir4.1 potassium channel.

PubMed

Wang, Ming-Xiao; Cuevas, Catherina A; Su, Xiao-Tong; Wu, Peng; Gao, Zhong-Xiuzi; Lin, Dao-Hong; McCormick, James A; Yang, Chao-Ling; Wang, Wen-Hui; Ellison, David H

2018-04-01

Kir4.1 in the distal convoluted tubule plays a key role in sensing plasma potassium and in modulating the thiazide-sensitive sodium-chloride cotransporter (NCC). Here we tested whether dietary potassium intake modulates Kir4.1 and whether this is essential for mediating the effect of potassium diet on NCC. High potassium intake inhibited the basolateral 40 pS potassium channel (a Kir4.1/5.1 heterotetramer) in the distal convoluted tubule, decreased basolateral potassium conductance, and depolarized the distal convoluted tubule membrane in Kcnj10flox/flox mice, herein referred to as control mice. In contrast, low potassium intake activated Kir4.1, increased potassium currents, and hyperpolarized the distal convoluted tubule membrane. These effects of dietary potassium intake on the basolateral potassium conductance and membrane potential in the distal convoluted tubule were completely absent in inducible kidney-specific Kir4.1 knockout mice. Furthermore, high potassium intake decreased, whereas low potassium intake increased the abundance of NCC expression only in the control but not in kidney-specific Kir4.1 knockout mice. Renal clearance studies demonstrated that low potassium augmented, while high potassium diminished, hydrochlorothiazide-induced natriuresis in control mice. Disruption of Kir4.1 significantly increased basal urinary sodium excretion but it abolished the natriuretic effect of hydrochlorothiazide. Finally, hypokalemia and metabolic alkalosis in kidney-specific Kir4.1 knockout mice were exacerbated by potassium restriction and only partially corrected by a high-potassium diet. Thus, Kir4.1 plays an essential role in mediating the effect of dietary potassium intake on NCC activity and potassium homeostasis. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Thermal Mechanisms of Millimeter Wave Stimulation of Excitable Cells

PubMed Central

Shapiro, Mikhail G.; Priest, Michael F.; Siegel, Peter H.; Bezanilla, Francisco

2013-01-01

Interactions between millimeter waves (MMWs) and biological systems have received increasing attention due to the growing use of MMW radiation in technologies ranging from experimental medical devices to telecommunications and airport security. Studies have shown that MMW exposure alters cellular function, especially in neurons and muscles. However, the biophysical mechanisms underlying such effects are still poorly understood. Due to the high aqueous absorbance of MMW, thermal mechanisms are likely. However, nonthermal mechanisms based on resonance effects have also been postulated. We studied MMW stimulation in a simplified preparation comprising Xenopus laevis oocytes expressing proteins that underlie membrane excitability. Using electrophysiological recordings simultaneously with 60 GHz stimulation, we observed changes in the kinetics and activity levels of voltage-gated potassium and sodium channels and a sodium-potassium pump that are consistent with a thermal mechanism. Furthermore, we showed that MMW stimulation significantly increased the action potential firing rate in oocytes coexpressing voltage-gated sodium and potassium channels, as predicted by thermal terms in the Hodgkin-Huxley model of neurons. Our results suggest that MMW stimulation produces significant thermally mediated effects on excitable cells via basic thermodynamic mechanisms that must be taken into account in the study and use of MMW radiation in biological systems. PMID:23790370

Evaluation of the therapeutic effect of potassium permanganate at early stages of an experimental acute infection of Flavobacterium columnare in channel catfish (Ictalurus punctatus)

USDA-ARS?s Scientific Manuscript database

The efficacy of potassium permanganate (KMnO4) against early stages of an experimental acute infection of Flavobacterium columnare in channel catfish (Ictalurus punctatus) was evaluated. Fish were experimentally challenged, by waterborne exposure for 2 h to F. columnare after cutaneous abrasion, an...

Investigation of MHD flow structure and fluctuations by potassium lineshape fitting

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bauman, L.E.

1993-12-31

Multiple Potassium D-line emission absorption spectra from a high temperature, coal-fired flow have been fit to a radiative transfer, boundary layer flow model. The results of fitting spectra from the aerodynamic duct of the Department of Energy Coal-Fired Flow Facility provide information about the thickness and shape of the thermal boundary layer and the bulk potassium seed atom density in a simulated magnetohydrodynamic channel flow. Probability distribution functions for the entire set of more than six thousand spectra clearly indicate the typical values and magnitude of fluctuations for the flow: core temperature of 2538 {plus_minus} 20 K, near wall temperaturemore » of 1945 {plus_minus} 135 K, boundary layer width of about 1 cm, and potassium seed atom density of (5.1 {plus_minus} 0.8)x 10{sup 22}/m{sup 3}. Probability distribution functions for selected times during the eight hours of measurements indicate occasional periods of unstable combustion. In addition, broadband particle parameters during the unstable start of the test may be related to differing particle and gas temperatures. The results clearly demonstrate the ability of lineshape fitting to provide valuable data for diagnosing the high speed turbulent flow.« less

Ion channels in inflammation.

PubMed

Eisenhut, Michael; Wallace, Helen

2011-04-01

Most physical illness in vertebrates involves inflammation. Inflammation causes disease by fluid shifts across cell membranes and cell layers, changes in muscle function and generation of pain. These disease processes can be explained by changes in numbers or function of ion channels. Changes in ion channels have been detected in diarrhoeal illnesses, pyelonephritis, allergy, acute lung injury and systemic inflammatory response syndromes involving septic shock. The key role played by changes in ion transport is directly evident in inflammation-induced pain. Expression or function of all major categories of ion channels like sodium, chloride, calcium, potassium, transient receptor potential, purinergic receptor and acid-sensing ion channels can be influenced by cyto- and chemokines, prostaglandins, leukotrienes, histamine, ATP, reactive oxygen species and protons released in inflammation. Key pathways in this interaction are cyclic nucleotide, phosphoinositide and mitogen-activated protein kinase-mediated signalling, direct modification by reactive oxygen species like nitric oxide, ATP or protons and disruption of the cytoskeleton. Therapeutic interventions to modulate the adverse and overlapping effects of the numerous different inflammatory mediators on each ion transport system need to target adversely affected ion transport systems directly and locally.

Computational models of O-LM cells are recruited by low or high theta frequency inputs depending on h-channel distributions

PubMed Central

Sekulić, Vladislav; Skinner, Frances K

2017-01-01

Although biophysical details of inhibitory neurons are becoming known, it is challenging to map these details onto function. Oriens-lacunosum/moleculare (O-LM) cells are inhibitory cells in the hippocampus that gate information flow, firing while phase-locked to theta rhythms. We build on our existing computational model database of O-LM cells to link model with function. We place our models in high-conductance states and modulate inhibitory inputs at a wide range of frequencies. We find preferred spiking recruitment of models at high (4–9 Hz) or low (2–5 Hz) theta depending on, respectively, the presence or absence of h-channels on their dendrites. This also depends on slow delayed-rectifier potassium channels, and preferred theta ranges shift when h-channels are potentiated by cyclic AMP. Our results suggest that O-LM cells can be differentially recruited by frequency-modulated inputs depending on specific channel types and distributions. This work exposes a strategy for understanding how biophysical characteristics contribute to function. DOI: http://dx.doi.org/10.7554/eLife.22962.001 PMID:28318488

[G-protein potentiates the activation of TNF-alpha on calcium-activated potassium channel in ECV304].

PubMed

Lin, L; Zheng, Y; Qu, J; Bao, G

2000-06-01

Observe the effect of tumor necrosis factor-alpha (TNF-alpha) on calcium-activated potassium channel in ECV304 and the possible involvement of G-protein mediation in the action of TNF-alpha. Using the cell-attached configuration of patch clamp technique. (1) the activity of high-conductance calcium-activated potassium channel (BKca) was recorded. Its conductance is (202.54 +/- 16.62) pS; (2) the activity of BKca was potentiated by 200 U/ml TNF-alpha; (3) G-protein would intensify this TNF-alpha activation. TNF-alpha acted on vascular endothelial cell ECV304 could rapidly activate the activity of BKca. Opening of BKca resulted in membrane hyper-polarization which could increase electro-chemical gradient for the resting Ca2+ influx and open leakage calcium channel, thus resting cytoplasmic free Ca2+ concentration could be elevated. G-protein may exert an important regulation in this process.

Interleukin-4 activates large-conductance, calciumactivated potassium (BKCa) channels in human airway smooth muscle cells

PubMed Central

Martin, Gilles; O’Connell, Robert J.; Pietrzykowski, Andrzej Z.; Treistman, Steven N.; Ethier, Michael F.; Madison, J. Mark

2014-01-01

Large-conductance, calcium-activated potassium (BKCa) channels are regulated by voltage and near-membrane calcium concentrations and are determinants of membrane potential and excitability in airway smooth muscle cells. Since the T helper–2 (Th2) cytokine, interleukin (IL)-4, is an important mediator of airway inflammation, we investigated whether IL-4 rapidly regulated BKCa activity in normal airway smooth muscle cells. On-cell voltage clamp recordings were made on subconfluent, cultured human bronchial smooth muscle cells (HBSMC). Interleukin-4 (50 ng ml−1), IL-13 (50 ng ml−1) or histamine (10 μm) was added to the bath during the recordings. Immunofluorescence studies with selective antibodies against the α and β1 subunits of BKCa were also performed. Both approaches demonstrated that HBSMC membranes contained large-conductance channels (>200 pS) with both calcium and voltage sensitivity, all of which is characteristic of the BKCa channel. Histamine caused a rapid increase in channel activity, as expected. A new finding was that perfusion with IL-4 stimulated rapid, large increases in BKCa channel activity (77.2 ± 63.3-fold increase, P < 0.05, n = 18). This large potentiation depended on the presence of external calcium. In contrast, IL-13 (50 ng ml−1) had little effect on BKCa channel activity, but inhibited the effect of IL-4. Thus, HBSMC contain functional BKCa channels whose activity is rapidly potentiated by the cytokine, IL-4, but not by IL-13.These findings are consistent with a model in which IL-4 rapidly increases near-membrane calcium concentrations to regulate BKCa activity. PMID:18403443

Actions and Mechanisms of Polyunsaturated Fatty Acids on Voltage-Gated Ion Channels.

PubMed

Elinder, Fredrik; Liin, Sara I

2017-01-01

Polyunsaturated fatty acids (PUFAs) act on most ion channels, thereby having significant physiological and pharmacological effects. In this review we summarize data from numerous PUFAs on voltage-gated ion channels containing one or several voltage-sensor domains, such as voltage-gated sodium (Na V ), potassium (K V ), calcium (Ca V ), and proton (H V ) channels, as well as calcium-activated potassium (K Ca ), and transient receptor potential (TRP) channels. Some effects of fatty acids appear to be channel specific, whereas others seem to be more general. Common features for the fatty acids to act on the ion channels are at least two double bonds in cis geometry and a charged carboxyl group. In total we identify and label five different sites for the PUFAs. PUFA site 1 : The intracellular cavity. Binding of PUFA reduces the current, sometimes as a time-dependent block, inducing an apparent inactivation. PUFA site 2 : The extracellular entrance to the pore. Binding leads to a block of the channel. PUFA site 3 : The intracellular gate. Binding to this site can bend the gate open and increase the current. PUFA site 4 : The interface between the extracellular leaflet of the lipid bilayer and the voltage-sensor domain. Binding to this site leads to an opening of the channel via an electrostatic attraction between the negatively charged PUFA and the positively charged voltage sensor. PUFA site 5 : The interface between the extracellular leaflet of the lipid bilayer and the pore domain. Binding to this site affects slow inactivation. This mapping of functional PUFA sites can form the basis for physiological and pharmacological modifications of voltage-gated ion channels.

Actions and Mechanisms of Polyunsaturated Fatty Acids on Voltage-Gated Ion Channels

PubMed Central

Elinder, Fredrik; Liin, Sara I.

2017-01-01

Polyunsaturated fatty acids (PUFAs) act on most ion channels, thereby having significant physiological and pharmacological effects. In this review we summarize data from numerous PUFAs on voltage-gated ion channels containing one or several voltage-sensor domains, such as voltage-gated sodium (NaV), potassium (KV), calcium (CaV), and proton (HV) channels, as well as calcium-activated potassium (KCa), and transient receptor potential (TRP) channels. Some effects of fatty acids appear to be channel specific, whereas others seem to be more general. Common features for the fatty acids to act on the ion channels are at least two double bonds in cis geometry and a charged carboxyl group. In total we identify and label five different sites for the PUFAs. PUFA site 1: The intracellular cavity. Binding of PUFA reduces the current, sometimes as a time-dependent block, inducing an apparent inactivation. PUFA site 2: The extracellular entrance to the pore. Binding leads to a block of the channel. PUFA site 3: The intracellular gate. Binding to this site can bend the gate open and increase the current. PUFA site 4: The interface between the extracellular leaflet of the lipid bilayer and the voltage-sensor domain. Binding to this site leads to an opening of the channel via an electrostatic attraction between the negatively charged PUFA and the positively charged voltage sensor. PUFA site 5: The interface between the extracellular leaflet of the lipid bilayer and the pore domain. Binding to this site affects slow inactivation. This mapping of functional PUFA sites can form the basis for physiological and pharmacological modifications of voltage-gated ion channels. PMID:28220076

Analysis of inter-residue contacts reveals folding stabilizers in P-loops of potassium, sodium, and TRPV channels.

PubMed

Korkosh, V S; Zhorov, B S; Tikhonov, D B

2016-05-01

The family of P-loop channels includes potassium, sodium, calcium, cyclic nucleotide-gated and TRPV channels, as well as ionotropic glutamate receptors. Despite vastly different physiological and pharmacological properties, the channels have structurally conserved folding of the pore domain. Furthermore, crystallographic data demonstrate surprisingly similar mutual disposition of transmembrane and membrane-diving helices. To understand determinants of this conservation, here we have compared available high-resolution structures of sodium, potassium, and TRPV1 channels. We found that some residues, which are in matching positions of the sequence alignment, occur in different positions in the 3D alignment. Surprisingly, we found 3D mismatches in well-packed P-helices. Analysis of energetics of individual residues in Monte Carlo minimized structures revealed cyclic patterns of energetically favorable inter- and intra-subunit contacts of P-helices with S6 helices. The inter-subunit contacts are rather conserved in all the channels, whereas the intra-subunit contacts are specific for particular types of the channels. Our results suggest that these residue-residue contacts contribute to the folding stabilization. Analysis of such contacts is important for structural and phylogenetic studies of homologous proteins.

Surface dynamics of voltage-gated ion channels.

PubMed

Heine, Martin; Ciuraszkiewicz, Anna; Voigt, Andreas; Heck, Jennifer; Bikbaev, Arthur

2016-07-03

Neurons encode information in fast changes of the membrane potential, and thus electrical membrane properties are critically important for the integration and processing of synaptic inputs by a neuron. These electrical properties are largely determined by ion channels embedded in the membrane. The distribution of most ion channels in the membrane is not spatially uniform: they undergo activity-driven changes in the range of minutes to days. Even in the range of milliseconds, the composition and topology of ion channels are not static but engage in highly dynamic processes including stochastic or activity-dependent transient association of the pore-forming and auxiliary subunits, lateral diffusion, as well as clustering of different channels. In this review we briefly discuss the potential impact of mobile sodium, calcium and potassium ion channels and the functional significance of this for individual neurons and neuronal networks.

Surface dynamics of voltage-gated ion channels

PubMed Central

Heine, Martin; Ciuraszkiewicz, Anna; Voigt, Andreas; Heck, Jennifer; Bikbaev, Arthur

2016-01-01

ABSTRACT Neurons encode information in fast changes of the membrane potential, and thus electrical membrane properties are critically important for the integration and processing of synaptic inputs by a neuron. These electrical properties are largely determined by ion channels embedded in the membrane. The distribution of most ion channels in the membrane is not spatially uniform: they undergo activity-driven changes in the range of minutes to days. Even in the range of milliseconds, the composition and topology of ion channels are not static but engage in highly dynamic processes including stochastic or activity-dependent transient association of the pore-forming and auxiliary subunits, lateral diffusion, as well as clustering of different channels. In this review we briefly discuss the potential impact of mobile sodium, calcium and potassium ion channels and the functional significance of this for individual neurons and neuronal networks. PMID:26891382

Molecular identity of cardiac mitochondrial chloride intracellular channel proteins.

PubMed

Ponnalagu, Devasena; Gururaja Rao, Shubha; Farber, Jason; Xin, Wenyu; Hussain, Ahmed Tafsirul; Shah, Kajol; Tanda, Soichi; Berryman, Mark; Edwards, John C; Singh, Harpreet

2016-03-01

Emerging evidences demonstrate significance of chloride channels in cardiac function and cardioprotection from ischemia-reperfusion (IR) injury. Unlike mitochondrial potassium channels sensitive to calcium (BKCa) and ATP (KATP), molecular identity of majority of cardiac mitochondrial chloride channels located at the inner membrane is not known. In this study, we report the presence of unique dimorphic chloride intracellular channel (CLIC) proteins namely CLIC1, CLIC4 and CLIC5 as abundant CLICs in the rodent heart. Further, CLIC4, CLIC5, and an ortholog present in Drosophila (DmCLIC) localize to adult cardiac mitochondria. We found that CLIC4 is enriched in the outer mitochondrial membrane, whereas CLIC5 is present in the inner mitochondrial membrane. Also, CLIC5 plays a direct role in regulating mitochondrial reactive oxygen species (ROS) generation. Our study highlights that CLIC5 is localized to the cardiac mitochondria and directly modulates mitochondrial function. Copyright © 2016 Elsevier B.V. and Mitochondria Research Society. All rights reserved.

Profiling neuronal ion channelopathies with non-invasive brain imaging and dynamic causal models: Case studies of single gene mutations

PubMed Central

Gilbert, Jessica R.; Symmonds, Mkael; Hanna, Michael G.; Dolan, Raymond J.; Friston, Karl J.; Moran, Rosalyn J.

2016-01-01

Clinical assessments of brain function rely upon visual inspection of electroencephalographic waveform abnormalities in tandem with functional magnetic resonance imaging. However, no current technology proffers in vivo assessments of activity at synapses, receptors and ion-channels, the basis of neuronal communication. Using dynamic causal modeling we compared electrophysiological responses from two patients with distinct monogenic ion channelopathies and a large cohort of healthy controls to demonstrate the feasibility of assaying synaptic-level channel communication non-invasively. Synaptic channel abnormality was identified in both patients (100% sensitivity) with assay specificity above 89%, furnishing estimates of neurotransmitter and voltage-gated ion throughput of sodium, calcium, chloride and potassium. This performance indicates a potential novel application as an adjunct for clinical assessments in neurological and psychiatric settings. More broadly, these findings indicate that biophysical models of synaptic channels can be estimated non-invasively, having important implications for advancing human neuroimaging to the level of non-invasive ion channel assays. PMID:26342528

Solution structure and function of the "tandem inactivation domain" of the neuronal A-type potassium channel Kv1.4.

PubMed

Wissmann, Ralph; Bildl, Wolfgang; Oliver, Dominik; Beyermann, Michael; Kalbitzer, Hans-Robert; Bentrop, Detlef; Fakler, Bernd

2003-05-02

Cumulative inactivation of voltage-gated (Kv) K(+) channels shapes the presynaptic action potential and determines timing and strength of synaptic transmission. Kv1.4 channels exhibit rapid "ball-and-chain"-type inactivation gating. Different from all other Kvalpha subunits, Kv1.4 harbors two inactivation domains at its N terminus. Here we report the solution structure and function of this "tandem inactivation domain" using NMR spectroscopy and patch clamp recordings. Inactivation domain 1 (ID1, residues 1-38) consists of a flexible N terminus anchored at a 5-turn helix, whereas ID2 (residues 40-50) is a 2.5-turn helix made up of small hydrophobic amino acids. Functional analysis suggests that only ID1 may work as a pore-occluding ball domain, whereas ID2 most likely acts as a "docking domain" that attaches ID1 to the cytoplasmic face of the channel. Deletion of ID2 slows inactivation considerably and largely impairs cumulative inactivation. Together, the concerted action of ID1 and ID2 may promote rapid inactivation of Kv1.4 that is crucial for the channel function in short term plasticity.

Computational Models for Understanding of Structure, Function and Pharmacology of the Cardiac Potassium Channel Kv11.1 (hERG).

PubMed

Wacker, Soren; Noskov, Sergei Yu; Perissinotti, Laura L

2017-01-01

The rapid delayed rectifier current IKr is one of the major K+ currents involved in repolarization of the human cardiac action potential. Various inherited or drug-induced forms of the long QT syndrome (LQTS) in humans are linked to functional and structural modifications in the IKr conducting channels. IKr is carried by the potassium channel Kv11.1 encoded by the gene KCNH2 (commonly referred to as human ether-a-go-go-related gene or hERG) [1, 2]. The first necessary step for predicting emergent drug effects on the heart is determining and modeling the binding thermodynamics and kinetics of primary and major off-target drug interactions with subcellular targets. The bulk of drugs that target hERG channels are known to have complex interactions at the atomic scale. Accordingly, one of the goals for this review is to provide comprehensive guide in the universe of computational models aiming to refine our understanding of structure-function relations in Kv11.1 and its isoforms. The special emphasis is placed on the mapping of drug binding sites and tentative mechanisms of channel inhibition and activation by drugs. An overview over recent structural models and mapping of binding sites for blockers and activators of IKr current along with the discussion on agreements and discrepancies among different models is presented. There is an apparent reciprocity or feedback loop between drug binding and action potential of the cardiac myocytes. Thus one has to connect drug binding to a particular receptor so that its functional consequences impact on the action potential duration. The natural pathway is to develop multi-scale models that connect between receptor and cellular scales. The potential for such multi-scale model development is discussed through the lens of common gating models. Accordingly, the second part of this review covers an ongoing development of the kinetic models of gating transitions and cardiac ion currents carried by hERG channels with and without drug bound. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Modulation of Kv7 channels and excitability in the brain.

PubMed

Greene, Derek L; Hoshi, Naoto

2017-02-01

Neuronal Kv7 channels underlie a voltage-gated non-inactivating potassium current known as the M-current. Due to its particular characteristics, Kv7 channels show pronounced control over the excitability of neurons. We will discuss various factors that have been shown to drastically alter the activity of this channel such as protein and phospholipid interactions, phosphorylation, calcium, and numerous neurotransmitters. Kv7 channels locate to key areas for the control of action potential initiation and propagation. Moreover, we will explore the dynamic surface expression of the channel modulated by neurotransmitters and neural activity. We will also focus on known principle functions of neural Kv7 channels: control of resting membrane potential and spiking threshold, setting the firing frequency, afterhyperpolarization after burst firing, theta resonance, and transient hyperexcitability from neurotransmitter-induced suppression of the M-current. Finally, we will discuss the contribution of altered Kv7 activity to pathologies such as epilepsy and cognitive deficits.

Modulation of Kv7 channels and excitability in the brain

PubMed Central

Greene, Derek L; Hoshi, Naoto

2016-01-01

Neuronal Kv7 channels underlie a voltage-gated non-inactivating potassium current known as the M-current. Due to its particular characteristics, Kv7 channels show pronounced control over the excitability of neurons. We will discuss various factors that have been shown to drastically alter the activity of this channel such as protein and phospholipid interactions, phosphorylation, calcium, and numerous neurotransmitters. Kv7 channels locate to key areas for the control of action potential initiation and propagation. Moreover, we will explore the dynamic surface expression of the channel modulated by neurotransmitters and neural activity. We will also focus on known principle functions of neural Kv7 channels: control of resting membrane potential and spiking threshold, setting the firing frequency, afterhyperpolarization after burst firing, theta resonance, and transient hyperexcitability from neurotransmitter-induced suppression of the M-current. Finally, we will discuss the contribution of altered Kv7 activity to pathologies such as epilepsy and cognitive deficits. PMID:27645822

CNS Schwann cells display oligodendrocyte precursor-like potassium channel activation and antigenic expression in vitro.

PubMed

Kegler, Kristel; Imbschweiler, Ilka; Ulrich, Reiner; Kovermann, Peter; Fahlke, Christoph; Deschl, Ulrich; Kalkuhl, Arno; Baumgärnter, Wolfgang; Wewetzer, Konstantin

2014-06-01

Central nervous system (CNS) injury triggers production of myelinating Schwann cells from endogenous oligodendrocyte precursors (OLPs). These CNS Schwann cells may be attractive candidates for novel therapeutic strategies aiming to promote endogenous CNS repair. However, CNS Schwann cells have been so far mainly characterized in situ regarding morphology and marker expression, and it has remained enigmatic whether they display functional properties distinct from peripheral nervous system (PNS) Schwann cells. Potassium channels (K+) have been implicated in progenitor and glial cell proliferation after injury and may, therefore, represent a suitable pharmacological target. In the present study, we focused on the function and expression of voltage-gated K+ channels Kv(1-12) and accessory β-subunits in purified adult canine CNS and PNS Schwann cell cultures using electrophysiology and microarray analysis and characterized their antigenic phenotype. We show here that K+ channels differed significantly in both cell types. While CNS Schwann cells displayed prominent K D-mediated K+ currents, PNS Schwann cells elicited K(D-) and K(A-type) K+ currents. Inhibition of K+ currents by TEA and Ba2+ was more effective in CNS Schwann cells. These functional differences were not paralleled by differential mRNA expression of Kv(1-12) and accessory β-subunits. However, O4/A2B5 and GFAP expressions were significantly higher and lower, respectively, in CNS than in PNS Schwann cells. Taken together, this is the first evidence that CNS Schwann cells display specific properties not shared by their peripheral counterpart. Both Kv currents and increased O4/A2B5 expression were reminiscent of OLPs suggesting that CNS Schwann cells retain OLP features during maturation.

The role of entropic potential in voltage activation and K+ transport through Kv 1.2 channels

NASA Astrophysics Data System (ADS)

Wawrzkiewicz-Jałowiecka, Agata; Grzywna, Zbigniew J.

2018-03-01

We analyze the entropic effects of inner pore geometry changes of Kv 1.2 channel during membrane depolarization and their implications for the rate of transmembrane transport of potassium ions. We base this on the idea that spatial confinements within the channel pore give rise to entropic barriers which can both effectively affect the stability of open macroconformation and influence channel's ability to conduct the potassium ions through the membrane. First, we calculate the differences in entropy between voltage-activated and resting states of the channel. As a template, we take a set of structures of channel pore in an open state at different membrane potentials generated in our previous research. The obtained results indicate that tendency to occupy open states at membrane depolarization is entropy facilitated. Second, we describe the differences in rates of K+ transport through the channel pore at different voltages based on the results of appropriate random walk simulations in entropic and electric potentials. The simulated single channel currents (I) suggest that the geometry changes during membrane depolarization are an important factor contributing to the observed flow of potassium ions through the channel. Nevertheless, the charge distribution within the channel pore (especially at the extracellular entrance) seems most prominent for the observed I/Imax relation at a qualitative level at analyzed voltages.

More than Memory Impairment in Voltage-Gated Potassium Channel Complex Encephalopathy

PubMed Central

Bettcher, Brianne M.; Gelfand, Jeffrey M.; Irani, Sarosh R.; Neuhaus, John; Forner, Sven; Hess, Christopher P.; Geschwind, Michael D.

2014-01-01

Objective Autoimmune encephalopathies (AE) are a heterogeneous group of neurological disorders that affect cognition. Although memory difficulties are commonly endorsed, few reports of AE inclusively assess all cognitive domains in detail. Our aim was to perform an unbiased cognitive evaluation of AE patients with voltage-gated potassium channel complex antibodies (VGKCC-Abs) in order to delineate cognitive strengths and weaknesses. Methods We assessed serial VGKCC-Abs AE subjects (n=12) with a comprehensive evaluation of memory, executive functions, visuospatial skills, and language. Clinical MRI (n=10/12) was evaluated. Five subjects had serial cognitive testing available, permitting descriptive analysis of change. Results Subjects demonstrated mild to moderate impairment in memory (mean Z=−1.9) and executive functions (mean Z=−1.5), with variable impairments in language and sparing of visuospatial skills. MRI findings showed T2 hyperintensities in medial temporal lobe (10/10) and basal ganglia (2/10). Serial cognitive examination revealed heterogeneity in cognitive function; whereas most patients improved in one or more domains, residual impairments were observed in some patients. Conclusions This study augments prior neuropsychological analyses in VGKCC-Ab AE by identifying not only memory and executive function deficits, but also language impairments, with preservation of visuospatial functioning. This study further highlights the importance of domain-specific testing to parse out the complex cognitive phenotypes of VGKCC-Ab AE. PMID:24981998

Neuronal and Cardiovascular Potassium Channels as Therapeutic Drug Targets

PubMed Central

Humphries, Edward S. A.

2015-01-01

Potassium (K+) channels, with their diversity, often tissue-defined distribution, and critical role in controlling cellular excitability, have long held promise of being important drug targets for the treatment of dysrhythmias in the heart and abnormal neuronal activity within the brain. With the exception of drugs that target one particular class, ATP-sensitive K+ (KATP) channels, very few selective K+ channel activators or inhibitors are currently licensed for clinical use in cardiovascular and neurological disease. Here we review what a range of human genetic disorders have told us about the role of specific K+ channel subunits, explore the potential of activators and inhibitors of specific channel populations as a therapeutic strategy, and discuss possible reasons for the difficulty in designing clinically relevant K+ channel modulators. PMID:26303307

Artificial phosphorylation sites modulate the activity of a voltage-gated potassium channel

NASA Astrophysics Data System (ADS)

Ariyaratne, Amila; Zocchi, Giovanni

2015-03-01

The KvAP potassium channel is representative of a family of voltage-gated ion channels where the membrane potential is sensed by a transmembrane helix containing several positively charged arginines. Previous work by Wang and Zocchi [A. Wang and G. Zocchi, PLoS ONE 6, e18598 (2011), 10.1371/journal.pone.0018598] showed how a negatively charged polyelectrolyte attached in proximity to the voltage sensing element can bias the opening probability of the channel. Here we introduce three phosphorylation sites at the same location and show that the response curve of the channel shifts by about 20 mV upon phosphorylation, while other characteristics such as the single-channel conductance are unaffected. In summary, we construct an artificial phosphorylation site which confers allosteric regulation to the channel.

Pharmacologic study of calcium influx pathways in rabbit aortic smooth muscle

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lukeman, D.S.

1987-01-01

Functional characteristics and pharmacologic domains of receptor-operated and potential-sensitive calcium (Ca/sup 2 +/) channels (ROCs and PSCs, respectively) were derived via measurements of /sup 45/Ca/sup 2 +/ influx (M/sup Ca/) during activation by the neurotransmitters norepinephrine (NE), histamine (HS), and serotonin (5-HT) and by elevated extracellular potassium (K/sup +/) in the individual or combined presence of organic Ca/sup 2 +/ channel antagonists (CAts), calmodulin antagonists (Calm-ants), lanthanum (La/sup 3 +/), and agents that increase intracellular levels of cyclic AMP.

Students' Understanding of External Representations of the Potassium Ion Channel Protein, Part I: Affordances and Limitations of Ribbon Diagrams, Vines, and Hydrophobic/Polar Representations

ERIC Educational Resources Information Center

Harle, Marissa; Towns, Marcy H.

2012-01-01

Research on external representations in biochemistry has uncovered student difficulties in comprehending and interpreting external representations. This project focuses on students' understanding of three external representations of the potassium ion channel protein. This is part I of a two-part study, which focuses on the affordances and…

Intracellular spermine blocks TRPC4 channel via electrostatic interaction with C-terminal negative amino acids.

PubMed

Kim, Jinsung; Moon, Sang Hui; Shin, Young-Cheul; Jeon, Ju-Hong; Park, Kyu Joo; Lee, Kyu Pil; So, Insuk

2016-04-01

Transient receptor potential canonical (TRPC) 4 channels are calcium-permeable, nonselective cation channels and are widely expressed in mammalian tissue, especially in the GI tract and brain. TRPC4 channels are known to be involved in neurogenic contraction of ileal smooth muscle cells via generating cationic current after muscarinic stimulation (muscarinic cationic current (mIcat)). Polyamines exist in numerous tissues and are believed to be involved in cell proliferation, differentiation, scar formation, wound healing, and carcinogenesis. Besides, physiological polyamines are essential to maintain inward rectification of cardiac potassium channels (Kir2.1). At membrane potentials more positive than equilibrium potential, intracellular polyamines plug the cytosolic surface of the Kir2.1 so that potassium ions cannot pass through the pore. Recently, it was reported that polyamines inhibit not only cardiac potassium channels but also nonselective cation channels that mediate the generation of mIcat. Here, we report that TRPC4, a definite mIcat mediator, is inhibited by intracellular spermine with great extent. The inhibition was specific to TRPC4 and TRPC5 channels but was not effective to TRPC1/4, TRPC1/5, and TRPC3 channels. For this inhibition to occur, we found that glutamates at 728th and 729th position of TRPC4 channels are essential whereby we conclude that spermine blocks the TRPC4 channel with electrostatic interaction between negative amino acids at the C-terminus of the channel.

Protons stabilize the closed conformation of gain-of-function mutants of the TRPV1 channel.

PubMed

Boukalova, Stepana; Teisinger, Jan; Vlachova, Viktorie

2013-03-01

The vanilloid transient receptor potential channel TRPV1 is a molecular integrator of noxious stimuli, including capsaicin, heat and protons. Despite clear similarities between the overall architecture of TRPV1 and voltage-dependent potassium (Kv) channels, the extent of conservation in the molecular logic for gating is unknown. In Kv channels, a small contact surface between S1 and the pore-helix is required for channel functioning. To explore the function of S1 in TRPV1, we used tryptophan-scanning mutagenesis and characterized the responses to capsaicin and protons. Wild-type-like currents were generated in 9 out of 17 mutants; three mutants (M445W, A452W, R455W) were non-functional. The conservative mutation R455K in the extracellular extent of S1 slowed down capsaicin-induced activation and prevented normal channel closure. This mutant was neither activated nor potentiated by protons, on the contrary, protons promoted a rapid deactivation of its currents. Similar phenotypes were found in two other gain-of-function mutants and also in the pore-helix mutant T633A, known to uncouple proton activation. We propose that the S1 domain contains a functionally important region that may be specifically involved in TRPV1 channel gating, and thus be important for the energetic coupling between S1-S4 sensor activation and gate opening. Analogous to Kv channels, the S1-pore interface might serve to stabilize conformations associated with TRPV1 channel gating. Copyright © 2012 Elsevier B.V. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lolicato, Marco; Arrigoni, Cristina; Mori, Takahiro

Polymodal thermo- and mechanosensitive two-pore domain potassium (K2P) channels of the TREK1 subfamily generate ‘leak’ currents that regulate neuronal excitability, respond to lipids, temperature and mechanical stretch, and influence pain, temperature perception and anaesthetic responses1, 2, 3. These dimeric voltage-gated ion channel (VGIC) superfamily members have a unique topology comprising two pore-forming regions per subunit4, 5, 6. In contrast to other potassium channels, K2P channels use a selectivity filter ‘C-type’ gate7, 8, 9, 10 as the principal gating site. Despite recent advances3, 11, 12, poor pharmacological profiles of K2P channels limit mechanistic and biological studies. Here we describe a classmore » of small-molecule TREK activators that directly stimulate the C-type gate by acting as molecular wedges that restrict interdomain interface movement behind the selectivity filter. Structures of K2P2.1 (also known as TREK-1) alone and with two selective K2P2.1 (TREK-1) and K2P10.1 (TREK-2) activators—an N-aryl-sulfonamide, ML335, and a thiophene-carboxamide, ML402—define a cryptic binding pocket unlike other ion channel small-molecule binding sites and, together with functional studies, identify a cation–π interaction that controls selectivity. Together, our data reveal a druggable K2P site that stabilizes the C-type gate ‘leak mode’ and provide direct evidence for K2P selectivity filter gating.« less

Postsynaptic ERG potassium channels limit muscle excitability to allow distinct egg-laying behavior states in C. elegans

PubMed Central

Collins, Kevin M.; Koelle, Michael R.

2013-01-01

C. elegans regulates egg laying by alternating between an inactive phase and a serotonin-triggered active phase. We found that the conserved ERG potassium channel UNC-103 enables this two-state behavior by limiting excitability of the egg-laying muscles. Using both high-speed video recording and calcium imaging of egg-laying muscles in behaving animals, we found that the muscles appear to be excited at a particular phase of each locomotor body bend. During the inactive phase, this rhythmic excitation infrequently evokes calcium transients or contraction of the egg-laying muscles. During the serotonin-triggered active phase, however, these muscles are more excitable and each body bend is accompanied by a calcium transient that drives twitching or full contraction of the egg-laying muscles. We found that ERG null mutants lay eggs too frequently, and that ERG function is necessary and sufficient in the egg-laying muscles to limit egg laying. ERG K+ channels localize to postsynaptic sites in the egg-laying muscle, and mutants lacking ERG have more frequent calcium transients and contractions of the egg-laying muscles even during the inactive phase. Thus ERG channels set postsynaptic excitability at a threshold so that further adjustments of excitability by serotonin generate two distinct behavioral states. PMID:23303953

Structural basis for KCNE3 modulation of potassium recycling in epithelia.

PubMed

Kroncke, Brett M; Van Horn, Wade D; Smith, Jarrod; Kang, CongBao; Welch, Richard C; Song, Yuanli; Nannemann, David P; Taylor, Keenan C; Sisco, Nicholas J; George, Alfred L; Meiler, Jens; Vanoye, Carlos G; Sanders, Charles R

2016-09-01

The single-span membrane protein KCNE3 modulates a variety of voltage-gated ion channels in diverse biological contexts. In epithelial cells, KCNE3 regulates the function of the KCNQ1 potassium ion (K(+)) channel to enable K(+) recycling coupled to transepithelial chloride ion (Cl(-)) secretion, a physiologically critical cellular transport process in various organs and whose malfunction causes diseases, such as cystic fibrosis (CF), cholera, and pulmonary edema. Structural, computational, biochemical, and electrophysiological studies lead to an atomically explicit integrative structural model of the KCNE3-KCNQ1 complex that explains how KCNE3 induces the constitutive activation of KCNQ1 channel activity, a crucial component in K(+) recycling. Central to this mechanism are direct interactions of KCNE3 residues at both ends of its transmembrane domain with residues on the intra- and extracellular ends of the KCNQ1 voltage-sensing domain S4 helix. These interactions appear to stabilize the activated "up" state configuration of S4, a prerequisite for full opening of the KCNQ1 channel gate. In addition, the integrative structural model was used to guide electrophysiological studies that illuminate the molecular basis for how estrogen exacerbates CF lung disease in female patients, a phenomenon known as the "CF gender gap."

Developmental Expression of Kv Potassium Channels at the Axon Initial Segment of Cultured Hippocampal Neurons

PubMed Central

Sánchez-Ponce, Diana; DeFelipe, Javier; Garrido, Juan José; Muñoz, Alberto

2012-01-01

Axonal outgrowth and the formation of the axon initial segment (AIS) are early events in the acquisition of neuronal polarity. The AIS is characterized by a high concentration of voltage-dependent sodium and potassium channels. However, the specific ion channel subunits present and their precise localization in this axonal subdomain vary both during development and among the types of neurons, probably determining their firing characteristics in response to stimulation. Here, we characterize the developmental expression of different subfamilies of voltage-gated potassium channels in the AISs of cultured mouse hippocampal neurons, including subunits Kv1.2, Kv2.2 and Kv7.2. In contrast to the early appearance of voltage-gated sodium channels and the Kv7.2 subunit at the AIS, Kv1.2 and Kv2.2 subunits were tethered at the AIS only after 10 days in vitro. Interestingly, we observed different patterns of Kv1.2 and Kv2.2 subunit expression, with each confined to distinct neuronal populations. The accumulation of Kv1.2 and Kv2.2 subunits at the AIS was dependent on ankyrin G tethering, it was not affected by disruption of the actin cytoskeleton and it was resistant to detergent extraction, as described previously for other AIS proteins. This distribution of potassium channels in the AIS further emphasizes the heterogeneity of this structure in different neuronal populations, as proposed previously, and suggests corresponding differences in action potential regulation. PMID:23119056

Differential effect of brief electrical stimulation on voltage-gated potassium channels

PubMed Central

Al Abed, Amr; Buskila, Yossi; Dokos, Socrates; Lovell, Nigel H.; Morley, John W.

2017-01-01

Electrical stimulation of neuronal tissue is a promising strategy to treat a variety of neurological disorders. The mechanism of neuronal activation by external electrical stimulation is governed by voltage-gated ion channels. This stimulus, typically brief in nature, leads to membrane potential depolarization, which increases ion flow across the membrane by increasing the open probability of these voltage-gated channels. In spiking neurons, it is activation of voltage-gated sodium channels (NaV channels) that leads to action potential generation. However, several other types of voltage-gated channels are expressed that also respond to electrical stimulation. In this study, we examine the response of voltage-gated potassium channels (KV channels) to brief electrical stimulation by whole cell patch-clamp electrophysiology and computational modeling. We show that nonspiking amacrine neurons of the retina exhibit a large variety of responses to stimulation, driven by different KV-channel subtypes. Computational modeling reveals substantial differences in the response of specific KV-channel subtypes that is dependent on channel kinetics. This suggests that the expression levels of different KV-channel subtypes in retinal neurons are a crucial predictor of the response that can be obtained. These data expand our knowledge of the mechanisms of neuronal activation and suggest that KV-channel expression is an important determinant of the sensitivity of neurons to electrical stimulation. NEW & NOTEWORTHY This paper describes the response of various voltage-gated potassium channels (KV channels) to brief electrical stimulation, such as is applied during prosthetic electrical stimulation. We show that the pattern of response greatly varies between KV channel subtypes depending on activation and inactivation kinetics of each channel. Our data suggest that problems encountered when artificially stimulating neurons such as cessation in firing at high frequencies, or “fading,” may be attributed to KV-channel activation. PMID:28202576

Voltage-dependent gating and gating charge measurements in the Kv1.2 potassium channel

PubMed Central

Ishida, Itzel G.; Rangel-Yescas, Gisela E.; Carrasco-Zanini, Julia

2015-01-01

Much has been learned about the voltage sensors of ion channels since the x-ray structure of the mammalian voltage-gated potassium channel Kv1.2 was published in 2005. High resolution structural data of a Kv channel enabled the structural interpretation of numerous electrophysiological findings collected in various ion channels, most notably Shaker, and permitted the development of meticulous computational simulations of the activation mechanism. The fundamental premise for the structural interpretation of functional measurements from Shaker is that this channel and Kv1.2 have the same characteristics, such that correlation of data from both channels would be a trivial task. We tested these assumptions by measuring Kv1.2 voltage-dependent gating and charge per channel. We found that the Kv1.2 gating charge is near 10 elementary charges (eo), ∼25% less than the well-established 13–14 eo in Shaker. Next, we neutralized positive residues in the Kv1.2 S4 transmembrane segment to investigate the cause of the reduction of the gating charge and found that, whereas replacing R1 with glutamine decreased voltage sensitivity to ∼50% of the wild-type channel value, mutation of the subsequent arginines had a much smaller effect. These data are in marked contrast to the effects of charge neutralization in Shaker, where removal of the first four basic residues reduces the gating charge by roughly the same amount. In light of these differences, we propose that the voltage-sensing domains (VSDs) of Kv1.2 and Shaker might undergo the same physical movement, but the septum that separates the aqueous crevices in the VSD of Kv1.2 might be thicker than Shaker’s, accounting for the smaller Kv1.2 gating charge. PMID:25779871

Voltage-Dependent Gating of hERG Potassium Channels

PubMed Central

Cheng, Yen May; Claydon, Tom W.

2012-01-01

The mechanisms by which voltage-gated channels sense changes in membrane voltage and energetically couple this with opening of the ion conducting pore has been the source of significant interest. In voltage-gated potassium (Kv) channels, much of our knowledge in this area comes from Shaker-type channels, for which voltage-dependent gating is quite rapid. In these channels, activation and deactivation are associated with rapid reconfiguration of the voltage-sensing domain unit that is electromechanically coupled, via the S4–S5 linker helix, to the rate-limiting opening of an intracellular pore gate. However, fast voltage-dependent gating kinetics are not typical of all Kv channels, such as Kv11.1 (human ether-à-go-go related gene, hERG), which activates and deactivates very slowly. Compared to Shaker channels, our understanding of the mechanisms underlying slow hERG gating is much poorer. Here, we present a comparative review of the structure–function relationships underlying activation and deactivation gating in Shaker and hERG channels, with a focus on the roles of the voltage-sensing domain and the S4–S5 linker that couples voltage sensor movements to the pore. Measurements of gating current kinetics and fluorimetric analysis of voltage sensor movement are consistent with models suggesting that the hERG activation pathway contains a voltage independent step, which limits voltage sensor transitions. Constraints upon hERG voltage sensor movement may result from loose packing of the S4 helices and additional intra-voltage sensor counter-charge interactions. More recent data suggest that key amino acid differences in the hERG voltage-sensing unit and S4–S5 linker, relative to fast activating Shaker-type Kv channels, may also contribute to the increased stability of the resting state of the voltage sensor. PMID:22586397

Ionic mechanisms of action of prion protein fragment PrP(106-126) in rat basal forebrain neurons.

PubMed

Alier, Kwai; Li, Zongming; Mactavish, David; Westaway, David; Jhamandas, Jack H

2010-08-01

Prion diseases are neurodegenerative disorders that are characterized by the presence of the misfolded prion protein (PrP). Neurotoxicity in these diseases may result from prion-induced modulation of ion channel function, changes in neuronal excitability, and consequent disruption of cellular homeostasis. We therefore examined PrP effects on a suite of potassium (K(+)) conductances that govern excitability of basal forebrain neurons. Our study examined the effects of a PrP fragment [PrP(106-126), 50 nM] on rat neurons using the patch clamp technique. In this paradigm, PrP(106-126) peptide, but not the "scrambled" sequence of PrP(106-126), evoked a reduction of whole-cell outward currents in a voltage range between -30 and +30 mV. Reduction of whole-cell outward currents was significantly attenuated in Ca(2+)-free external media and also in the presence of iberiotoxin, a blocker of calcium-activated potassium conductance. PrP(106-126) application also evoked a depression of the delayed rectifier (I(K)) and transient outward (I(A)) potassium currents. By using single cell RT-PCR, we identified the presence of two neuronal chemical phenotypes, GABAergic and cholinergic, in cells from which we recorded. Furthermore, cholinergic and GABAergic neurons were shown to express K(v)4.2 channels. Our data establish that the central region of PrP, defined by the PrP(106-126) peptide used at nanomolar concentrations, induces a reduction of specific K(+) channel conductances in basal forebrain neurons. These findings suggest novel links between PrP signalling partners inferred from genetic experiments, K(+) channels, and PrP-mediated neurotoxicity.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bean, Bruce Palmer

The effects of ether and halothane on membrane currents in the voltage clamped crayfish giant axon membrane were investigated. Concentrations of ether up to 300 mM and of halothane up to 32 mM had no effect on resting potential or leakage conductance. Ether and halothane reduced the size of sodium currents without changing the voltage dependence of the peak currents or their reversal potential. Ether and halothane also produced a reversible, dose-dependent speeding of sodium current decay at all membrane potentials. Ether reduced the time constants for inactivation, and also shifted the midpoint of the steady-state inactivation curve in themore » hyperpolarizing direction. Potassium currents were smaller with ether present, with no change in the voltage dependence of steady-state currents. The activation of potassium channels was faster with ether present. There was no apparent change in the capacitance of the crayfish giant axon membrane with ether concentrations of up to 100 mM. Experiments on sodium channel inactivation kinetics were performed using 4-aminopyridine to block potassium currents. Sodium currents decayed with a time course generally fit well by a single exponential. The time constant of decay was a steep function of voltage, especially in the negative resistance region of the peak current vs voltage relation.The time course of inactivation was very similar to that of the decay of the current at the same potential. The measurement of steady-state inactivation curves with different test pulses showed no shifts along the voltage asix. The voltage-dependence of the integral of sodium conductance was measured to test models of sodium channel inactivation in which channels must open before inactivating; the results appear inconsistent with some of the simplest cases of such models.« less

The calcium-activated potassium channel KCa3.1 plays a central role in the chemotactic response of mammalian neutrophils.

PubMed

Henríquez, C; Riquelme, T T; Vera, D; Julio-Kalajzić, F; Ehrenfeld, P; Melvin, J E; Figueroa, C D; Sarmiento, J; Flores, C A

2016-01-01

Neutrophils are the first cells to arrive at sites of injury. Nevertheless, many inflammatory diseases are characterized by an uncontrolled infiltration and action of these cells. Cell migration depends on volume changes that are governed by ion channel activity, but potassium channels in neutrophil have not been clearly identified. We aim to test whether KCa3.1 participates in neutrophil migration and other relevant functions of the cell. Cytometer and confocal measurements to determine changes in cell volume were used. Cells isolated from human, mouse and horse were tested for KCa3.1-dependent chemotaxis. Chemokinetics, calcium handling and release of reactive oxygen species were measured to determine the role of KCa3.1 in those processes. A mouse model was used to test for neutrophil recruitment after acute lung injury in vivo. We show for the first time that KCa3.1 is expressed in mammalian neutrophils. When the channel is inhibited by a pharmacological blocker or by genetic silencing, it profoundly affects cell volume regulation, and chemotactic and chemokinetic properties of the cells. We also demonstrated that pharmacological inhibition of KCa3.1 did not affect calcium entry or reactive oxygen species production in neutrophils. Using a mouse model of acute lung injury, we observed that Kca3.1(-/-) mice are significantly less effective at recruiting neutrophils into the site of inflammation. These results demonstrate that KCa3.1 channels are key actors in the migration capacity of neutrophils, and its inhibition did not affect other relevant cellular functions. © 2015 Scandinavian Physiological Society. Published by John Wiley & Sons Ltd.

Biochemical and Electrophysiological Characterization of Two Sea Anemone Type 1 Potassium Toxins from a Geographically Distant Population of Bunodosoma caissarum

PubMed Central

Orts, Diego J. B.; Peigneur, Steve; Madio, Bruno; Cassoli, Juliana S.; Montandon, Gabriela G.; Pimenta, Adriano M. C.; Bicudo, José E. P. W.; Freitas, José C.; Zaharenko, André J.; Tytgat, Jan

2013-01-01

Sea anemone (Cnidaria, Anthozoa) venom is an important source of bioactive compounds used as tools to study the pharmacology and structure-function of voltage-gated K+ channels (KV). These neurotoxins can be divided into four different types, according to their structure and mode of action. In this work, for the first time, two toxins were purified from the venom of Bunodosoma caissarum population from Saint Peter and Saint Paul Archipelago, Brazil. Sequence alignment and phylogenetic analysis reveals that BcsTx1 and BcsTx2 are the newest members of the sea anemone type 1 potassium channel toxins. Their functional characterization was performed by means of a wide electrophysiological screening on 12 different subtypes of KV channels (KV1.1–KV1.6; KV2.1; KV3.1; KV4.2; KV4.3; hERG and Shaker IR). BcsTx1 shows a high affinity for rKv1.2 over rKv1.6, hKv1.3, Shaker IR and rKv1.1, while Bcstx2 potently blocked rKv1.6 over hKv1.3, rKv1.1, Shaker IR and rKv1.2. Furthermore, we also report for the first time a venom composition and biological activity comparison between two geographically distant populations of sea anemones. PMID:23466933

A novel frequency analysis method for assessing K(ir)2.1 and Na (v)1.5 currents.

PubMed

Rigby, J R; Poelzing, S

2012-04-01

Voltage clamping is an important tool for measuring individual currents from an electrically active cell. However, it is difficult to isolate individual currents without pharmacological or voltage inhibition. Herein, we present a technique that involves inserting a noise function into a standard voltage step protocol, which allows one to characterize the unique frequency response of an ion channel at different step potentials. Specifically, we compute the fast Fourier transform for a family of current traces at different step potentials for the inward rectifying potassium channel, K(ir)2.1, and the channel encoding the cardiac fast sodium current, Na(v)1.5. Each individual frequency magnitude, as a function of voltage step, is correlated to the peak current produced by each channel. The correlation coefficient vs. frequency relationship reveals that these two channels are associated with some unique frequencies with high absolute correlation. The individual IV relationship can then be recreated using only the unique frequencies with magnitudes of high absolute correlation. Thus, this study demonstrates that ion channels may exhibit unique frequency responses.

TMEM150C/Tentonin3 Is a Regulator of Mechano-gated Ion Channels.

PubMed

Anderson, Evan O; Schneider, Eve R; Matson, Jon D; Gracheva, Elena O; Bagriantsev, Sviatoslav N

2018-04-17

Neuronal mechano-sensitivity relies on mechano-gated ion channels, but pathways regulating their activity remain poorly understood. TMEM150C was proposed to mediate mechano-activated current in proprioceptive neurons. Here, we studied functional interaction of TMEM150C with mechano-gated ion channels from different classes (Piezo2, Piezo1, and the potassium channel TREK-1) using two independent methods of mechanical stimulation. We found that TMEM150C significantly prolongs the duration of the mechano-current produced by all three channels, decreases apparent activation threshold in Piezo2, and induces persistent current in Piezo1. We also show that TMEM150C is co-expressed with Piezo2 in trigeminal neurons, expanding its role beyond proprioceptors. Finally, we cloned TMEM150C from the trigeminal neurons of the tactile-foraging domestic duck and showed that it functions similarly to the mouse ortholog, demonstrating evolutionary conservation among vertebrates. Our studies reveal TMEM150C as a general regulator of mechano-gated ion channels from different classes. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

Optical control of insulin release using a photoswitchable sulfonylurea.

PubMed

Broichhagen, Johannes; Schönberger, Matthias; Cork, Simon C; Frank, James A; Marchetti, Piero; Bugliani, Marco; Shapiro, A M James; Trapp, Stefan; Rutter, Guy A; Hodson, David J; Trauner, Dirk

2014-10-14

Sulfonylureas are widely prescribed for the treatment of type 2 diabetes mellitus (T2DM). Through their actions on ATP-sensitive potassium (KATP) channels, sulfonylureas boost insulin release from the pancreatic beta cell mass to restore glucose homeostasis. A limitation of these compounds is the elevated risk of developing hypoglycemia and cardiovascular disease, both potentially fatal complications. Here, we describe the design and development of a photoswitchable sulfonylurea, JB253, which reversibly and repeatedly blocks KATP channel activity following exposure to violet-blue light. Using in situ imaging and hormone assays, we further show that JB253 bestows light sensitivity upon rodent and human pancreatic beta cell function. Thus, JB253 enables the optical control of insulin release and may offer a valuable research tool for the interrogation of KATP channel function in health and T2DM.

Mechanosensitive Piezo Channels in the Gastrointestinal Tract

PubMed Central

Alcaino, C.; Farrugia, G.; Beyder, A.

2017-01-01

Sensation of mechanical forces is critical for normal function of the gastrointestinal (GI) tract and abnormalities in mechanosensation are linked to GI pathologies. In the GI tract there are several mechanosensitive cell types—epithelial enterochromaffin cells, intrinsic and extrinsic enteric neurons, smooth muscle cells and interstitial cells of Cajal. These cells use mechanosensitive ion channels that respond to mechanical forces by altering transmembrane ionic currents in a process called mechanoelectrical coupling. Several mechanosensitive ionic conductances have been identified in the mechano-sensory GI cells, ranging from mechanosensitive voltage-gated sodium and calcium channels to the mechanogated ion channels, such as the two-pore domain potassium channels K2P (TREK-1) and nonselective cation channels from the transient receptor potential family. The recently discovered Piezo channels are increasingly recognized as significant contributors to cellular mechanosensitivity. Piezo1 and Piezo2 are nonselective cationic ion channels that are directly activated by mechanical forces and have well-defined biophysical and pharmacologic properties. The role of Piezo channels in the GI epithelium is currently under investigation and their role in the smooth muscle syncytium and enteric neurons is still not known. In this review, we outline the current state of knowledge on mechanosensitive ion channels in the GI tract, with a focus on the known and potential functions of the Piezo channels. PMID:28728818

Potassium channels and prostacyclin contribute to vasorelaxant activities of Tridax procumbens crude aqueous leaf extract in rat superior mesenteric arteries.

PubMed

Salahdeen, H M; Adebari, A O; Murtala, B A; Alada, A R A

2015-03-01

Previous studies have shown that aqueous extract of the leaf of Tridax procuinbens is capable of lowering blood pressure through its vasodilatory effects. In the present study attempt was made to examine the biological active components of T procuinbens leaf using GC-MS methods. We further investigated the role of K+ channels in the vasorelaxation effects of Tridax procumbens using rat isolated mesenteric artery. The superior mesenteric artery isolated from healthy, young adult Wistar rats (250-300 g) were precontracted with phenylephrine (PE) (10(-7) M) and potassium chloride (KCl) (60 mM) and were treated with Various concentrations of aqueous extract ofT procumbens (0.9.0 mg/ml). The changes in arterial tension were recorded using a force-displacement transducer (Model 7004; Ugo Basil Varese, Italy) coupled to data capsule acquisition system. The results of GG-MS revealed the presence of linoleic acid. The T. procumbens extract (TPE) ranging from 0.5-9.0 mg/mI significantly (p<0.05) reduced the, contraction induced by (PE) and (KCl) in a concentration-dependent manner. The extract also antagonised the calcium-induced vasoconstriction (1(-9) - 10(-5)) in calcium-free with high concentration of potassium as well as. in calcium- and potassium free physiological solutions. The vasorelaxing effect caused by TPE was significantly (p<0.05) attenuated with preincubation of potassium channels blockers (Barium chloride and apamin), NO synthaseinhibitor (L-NAME), prostacyclin inhibitor (indomethacin), atropine; propranolol, and methylene blue while it was not affected by preincubation with glibenclamide and tetra ethyl ammonium, 4-aminopyridine (4-AP) and oxadiazolo quinoxalin (ODQ). The results of this study demonstrate that T procumbens extract causes vasodilatory effects by blocking calcium channels and the vasodilatory effect of the extract may also be due to stimulation of prostacyclin production and opening of small-conductance Ga2+ activated potassium channels. The observed effect of this extract may be probably due to the presence of linoleic acid in this extract.

Involvement of Potassium and Cation Channels in Hippocampal Abnormalities of Embryonic Ts65Dn and Tc1 Trisomic Mice

PubMed Central

Stern, Shani; Segal, Menahem; Moses, Elisha

2015-01-01

Down syndrome (DS) mouse models exhibit cognitive deficits, and are used for studying the neuronal basis of DS pathology. To understand the differences in the physiology of DS model neurons, we used dissociated neuronal cultures from the hippocampi of Ts65Dn and Tc1 DS mice. Imaging of [Ca2+]i and whole cell patch clamp recordings were used to analyze network activity and single neuron properties, respectively. We found a decrease of ~ 30% in both fast (A-type) and slow (delayed rectifier) outward potassium currents. Depolarization of Ts65Dn and Tc1 cells produced fewer spikes than diploid cells. Their network bursts were smaller and slower than diploids, displaying a 40% reduction in Δf / f0 of the calcium signals, and a 30% reduction in propagation velocity. Additionally, Ts65Dn and Tc1 neurons exhibited changes in the action potential shape compared to diploid neurons, with an increase in the amplitude of the action potential, a lower threshold for spiking, and a sharp decrease of about 65% in the after-hyperpolarization amplitude. Numerical simulations reproduced the DS measured phenotype by variations in the conductance of the delayed rectifier and A-type, but necessitated also changes in inward rectifying and M-type potassium channels and in the hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. We therefore conducted whole cell patch clamp measurements of M-type potassium currents, which showed a ~ 90% decrease in Ts65Dn neurons, while HCN measurements displayed an increase of ~ 65% in Ts65Dn cells. Quantitative real-time PCR analysis indicates overexpression of 40% of KCNJ15, an inward rectifying potassium channel, contributing to the increased inhibition. We thus find that changes in several types of potassium channels dominate the observed DS model phenotype. PMID:26501103

Histidine phosphorylation relieves copper inhibition in the mammalian potassium channel KCa3.1

PubMed Central

Srivastava, Shekhar; Panda, Saswati; Li, Zhai; Fuhs, Stephen R; Hunter, Tony; Thiele, Dennis J; Hubbard, Stevan R; Skolnik, Edward Y

2016-01-01

KCa2.1, KCa2.2, KCa2.3 and KCa3.1 constitute a family of mammalian small- to intermediate-conductance potassium channels that are activated by calcium-calmodulin. KCa3.1 is unique among these four channels in that activation requires, in addition to calcium, phosphorylation of a single histidine residue (His358) in the cytoplasmic region, by nucleoside diphosphate kinase-B (NDPK-B). The mechanism by which KCa3.1 is activated by histidine phosphorylation is unknown. Histidine phosphorylation is well characterized in prokaryotes but poorly understood in eukaryotes. Here, we demonstrate that phosphorylation of His358 activates KCa3.1 by antagonizing copper-mediated inhibition of the channel. Furthermore, we show that activated CD4+ T cells deficient in intracellular copper exhibit increased KCa3.1 histidine phosphorylation and channel activity, leading to increased calcium flux and cytokine production. These findings reveal a novel regulatory mechanism for a mammalian potassium channel and for T-cell activation, and highlight a unique feature of histidine versus serine/threonine and tyrosine as a regulatory phosphorylation site. DOI: http://dx.doi.org/10.7554/eLife.16093.001 PMID:27542194

Microelectrode array measurement of potassium ion channel remodeling on the field action potential duration in rapid atrial pacing rabbits model.

PubMed

Sun, Juan; Yan, Huang; Wugeti, Najina; Guo, Yujun; Zhang, Ling; Ma, Mei; Guo, Xingui; Jiao, Changan; Xu, Wenli; Li, Tianqi

2015-01-01

Atrial fibrillation (AF) arises from abnormalities in atrial structure and electrical activity. Microelectrode arrays (MEA) is a real-time, nondestructive measurement of the resting and action potential signal, from myocardial cells, to the peripheral circuit of electrophysiological activity. This study examined the field action potential duration (fAPD) of the right atrial appendage (RAA) by MEA in rapid atrial pacing (RAP) in the right atrium of rabbits. In addition, this study also investigated the effect of potassium ion channel blockers on fAPD. 40 New Zealand white rabbits of either sex were randomly divided into 3 groups: 1) the control, 2) potassium ion channel blocker (TEA, 4-Ap and BaCl2), and 3) amiodarone groups. The hearts were quickly removed and right atrial appendage sectioned (slice thickness 500 μm). Each slice was perfused with Tyrode's solution and continuously stimulated for 30 minutes. Sections from the control group were superfused with Tyrode's solution for 10 minutes, while the blocker groups and amiodarone were both treated with their respective compounds for 10 minutes each. The fAPD of RAA and action field action potential morphology were measured using MEA. In non-pace (control) groups, fAPD was 188.33 ± 18.29 ms after Tyrode's solution superfusion, and 173.91 ± 6.83 ms after RAP. In pace/potassium ion channel groups, TEA and BaCl2 superfusion prolonged atrial field action potential (fAPD) (control vs blocker: 176.67 ± 8.66 ms vs 196.11 ± 10.76 ms, 182.22 ± 12.87 ms vs 191.11 ± 13.09 ms with TEA and BaCl2 superfusion, respectively, P < 0.05). 4-AP superfusion significantly prolonged FAPD. In pace/amiodarone groups, 4-Ap superfusion extended fAPD. MEA was a sensitive and stable reporter for the measurement of the tissue action potential in animal heart slices. After superfusing potassium ion channel blockers, fAPD was prolonged. These results suggest that Ito, IKur and IK1 remodel and mediate RAP-induced atrial electrical remodeling. Amiodarone alter potassium ion channel activity (Ito, IKur, IK1 and IKs), shortening fAPD.

Activation of mTORC1 in Collecting Ducts Causes Hyperkalemia

PubMed Central

Chen, Zhenguo; Dong, Heling; Jia, Chunhong; Song, Qiancheng; Chen, Juan; Zhang, Yue; Lai, Pinglin; Fan, Xiaorong; Zhou, Xuan; Liu, Miao; Lin, Jun; Yang, Cuilan; Li, Ming; Gao, Tianming

2014-01-01

Mutation of TSC (encoding tuberous sclerosis complex protein) and activation of mammalian target of rapamycin (mTOR) have been implicated in the pathogenesis of several renal diseases, such as diabetic nephropathy and polycystic kidney disease. However, the role of mTOR in renal potassium excretion and hyperkalemia is not known. We showed that mice with collecting-duct (CD)–specific ablation of TSC1 (CDTsc1KO) had greater mTOR complex 1 (mTORC1) activation in the CD and demonstrated features of pseudohypoaldosteronism, including hyperkalemia, hyperaldosteronism, and metabolic acidosis. mTORC1 activation caused endoplasmic reticulum stress, columnar cell lesions, and dedifferentiation of CD cells with loss of aquaporin-2 and epithelial-mesenchymal transition-like phenotypes. Of note, mTORC1 activation also reduced the expression of serum- and glucocorticoid-inducible kinase 1, a crucial regulator of potassium homeostasis in the kidney, and decreased the expression and/or activity of epithelial sodium channel-α, renal outer medullary potassium channel, and Na+, K+-ATPase in the CD, which probably contributed to the aldosterone resistance and hyperkalemia in these mice. Rapamycin restored these phenotypic changes. Overall, this study identifies a novel function of mTORC1 in regulating potassium homeostasis and demonstrates that loss of TSC1 and activation of mTORC1 results in dedifferentiation and dysfunction of the CD and causes hyperkalemia. The CDTsc1KO mice provide a novel model for hyperkalemia induced exclusively by dysfunction of the CD. PMID:24203997

Activation of mTORC1 in collecting ducts causes hyperkalemia.

PubMed

Chen, Zhenguo; Dong, Heling; Jia, Chunhong; Song, Qiancheng; Chen, Juan; Zhang, Yue; Lai, Pinglin; Fan, Xiaorong; Zhou, Xuan; Liu, Miao; Lin, Jun; Yang, Cuilan; Li, Ming; Gao, Tianming; Bai, Xiaochun

2014-03-01

Mutation of TSC (encoding tuberous sclerosis complex protein) and activation of mammalian target of rapamycin (mTOR) have been implicated in the pathogenesis of several renal diseases, such as diabetic nephropathy and polycystic kidney disease. However, the role of mTOR in renal potassium excretion and hyperkalemia is not known. We showed that mice with collecting-duct (CD)-specific ablation of TSC1 (CDTsc1KO) had greater mTOR complex 1 (mTORC1) activation in the CD and demonstrated features of pseudohypoaldosteronism, including hyperkalemia, hyperaldosteronism, and metabolic acidosis. mTORC1 activation caused endoplasmic reticulum stress, columnar cell lesions, and dedifferentiation of CD cells with loss of aquaporin-2 and epithelial-mesenchymal transition-like phenotypes. Of note, mTORC1 activation also reduced the expression of serum- and glucocorticoid-inducible kinase 1, a crucial regulator of potassium homeostasis in the kidney, and decreased the expression and/or activity of epithelial sodium channel-α, renal outer medullary potassium channel, and Na(+), K(+)-ATPase in the CD, which probably contributed to the aldosterone resistance and hyperkalemia in these mice. Rapamycin restored these phenotypic changes. Overall, this study identifies a novel function of mTORC1 in regulating potassium homeostasis and demonstrates that loss of TSC1 and activation of mTORC1 results in dedifferentiation and dysfunction of the CD and causes hyperkalemia. The CDTsc1KO mice provide a novel model for hyperkalemia induced exclusively by dysfunction of the CD.

Ion conduction in the KcsA potassium channel analyzed with a minimal kinetic model.

PubMed

Mafé, Salvador; Pellicer, Julio

2005-02-01

We use a model by Nelson to study the current-voltage and conductance-concentration curves of bacterial potassium channel KcsA without assuming rapid ion translocation. Ion association to the channel filter is rate controlling at low concentrations, but dissociation and transport in the filter can limit conduction at high concentration for ions other than K+. The absolute values of the effective rate constants are tentative but the relative changes in these constants needed to qualitatively explain the experiments should be of significance.

[Channels: a new way to revisit pathology].

PubMed

Fournier, Emmanuel

2014-02-01

Many "essential" diseases that manifest themselves in the form of crises or fits (epilepsies, episodic ataxia, periodic paralyses, myotonia, heart rhythm disorders, etc.) are due to ionic channel dysfunction and are thus referred to as "channelopathies". Some of these disorders are congenital, due to mutations of genes encoding channel subunits, while others result from toxic, immune or hormonal disturbances affecting channelfunction. Channelopathies take on a wide variety of clinical forms, depending on the type of channel (sodium, potassium, calcium, chloride...) and the type of dysfunction (loss or gain of function). Some apparently unrelated diseases affecting distinct organs are due to a similar dysfunction of the same channel, revealing unsuspected relationships between organs and between medical specialties. In addition, a given syndrome can be caused by distinct channel dysfunctions. This provides new opportunities for diferential diagnosis and specific correction of the causal defects, although some treatments find applications across multiple medical specialties.

Ion Channels in Innate and Adaptive Immunity

PubMed Central

Feske, Stefan; Wulff, Heike; Skolnik, Edward Y.

2016-01-01

Ion channels and transporters mediate the transport of charged ions across hydrophobic lipid membranes. In immune cells, divalent cations such as calcium, magnesium, and zinc have important roles as second messengers to regulate intracellular signaling pathways. By contrast, monovalent cations such as sodium and potassium mainly regulate the membrane potential, which indirectly controls the influx of calcium and immune cell signaling. Studies investigating human patients with mutations in ion channels and transporters, analysis of gene-targeted mice, or pharmacological experiments with ion channel inhibitors have revealed important roles of ionic signals in lymphocyte development and in innate and adaptive immune responses. We here review the mechanisms underlying the function of ion channels and transporters in lymphocytes and innate immune cells and discuss their roles in lymphocyte development, adaptive and innate immune responses, and autoimmunity, as well as recent efforts to develop pharmacological inhibitors of ion channels for immunomodulatory therapy. PMID:25861976

Heme impairs the ball-and-chain inactivation of potassium channels.

PubMed

Sahoo, Nirakar; Goradia, Nishit; Ohlenschläger, Oliver; Schönherr, Roland; Friedrich, Manfred; Plass, Winfried; Kappl, Reinhard; Hoshi, Toshinori; Heinemann, Stefan H

2013-10-15

Fine-tuned regulation of K(+) channel inactivation enables excitable cells to adjust action potential firing. Fast inactivation present in some K(+) channels is mediated by the distal N-terminal structure (ball) occluding the ion permeation pathway. Here we show that Kv1.4 K(+) channels are potently regulated by intracellular free heme; heme binds to the N-terminal inactivation domain and thereby impairs the inactivation process, thus enhancing the K(+) current with an apparent EC50 value of ∼20 nM. Functional studies on channel mutants and structural investigations on recombinant inactivation ball domain peptides encompassing the first 61 residues of Kv1.4 revealed a heme-responsive binding motif involving Cys13:His16 and a secondary histidine at position 35. Heme binding to the N-terminal inactivation domain induces a conformational constraint that prevents it from reaching its receptor site at the vestibule of the channel pore.

Quantitative autoradiography of the binding sites for ( sup 125 I) iodoglyburide, a novel high-affinity ligand for ATP-sensitive potassium channels in rat brain

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gehlert, D.R.; Gackenheimer, S.L.; Mais, D.E.

1991-05-01

We have developed a high specific activity ligand for localization of ATP-sensitive potassium channels in the brain. When brain sections were incubated with ({sup 125}I)iodoglyburide (N-(2-((((cyclohexylamino)carbonyl)amino)sulfonyl)ethyl)-5-{sup 125}I-2- methoxybenzamide), the ligand bound to a single site with a KD of 495 pM and a maximum binding site density of 176 fmol/mg of tissue. Glyburide was the most potent inhibitor of specific ({sup 125}I)iodoglyburide binding to rat forebrain sections whereas iodoglyburide and glipizide were slightly less potent. The binding was also sensitive to ATP which completely inhibited binding at concentrations of 10 mM. Autoradiographic localization of ({sup 125}I)iodoglyburide binding indicated a broadmore » distribution of the ATP-sensitive potassium channel in the brain. The highest levels of binding were seen in the globus pallidus and ventral pallidum followed by the septohippocampal nucleus, anterior pituitary, the CA2 and CA3 region of the hippocampus, ventral pallidum, the molecular layer of the cerebellum and substantia nigra zona reticulata. The hilus and dorsal subiculum of the hippocampus, molecular layer of the dentate gyrus, cerebral cortex, lateral olfactory tract nucleus, olfactory tubercle and the zona incerta contained relatively high levels of binding. A lower level of binding (approximately 3- to 4-fold) was found throughout the remainder of the brain. These results indicate that the ATP-sensitive potassium channel has a broad presence in the rat brain and that a few select brain regions are enriched in this subtype of neuronal potassium channels.« less

Local calcium signalling is mediated by mechanosensitive ion channels in mesenchymal stem cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chubinskiy-Nadezhdin, Vladislav I., E-mail: vchubinskiy@gmail.com; Vasileva, Valeria Y.; Pugovkina, Natalia A.

Mechanical forces are implicated in key physiological processes in stem cells, including proliferation, differentiation and lineage switching. To date, there is an evident lack of understanding of how external mechanical cues are coupled with calcium signalling in stem cells. Mechanical reactions are of particular interest in adult mesenchymal stem cells because of their promising potential for use in tissue remodelling and clinical therapy. Here, single channel patch-clamp technique was employed to search for cation channels involved in mechanosensitivity in mesenchymal endometrial-derived stem cells (hMESCs). Functional expression of native mechanosensitive stretch-activated channels (SACs) and calcium-sensitive potassium channels of different conductances inmore » hMESCs was shown. Single current analysis of stretch-induced channel activity revealed functional coupling of SACs and BK channels in plasma membrane. The combination of cell-attached and inside-out experiments have indicated that highly localized Ca{sup 2+} entry via SACs triggers BK channel activity. At the same time, SK channels are not coupled with SACs despite of high calcium sensitivity as compared to BK. Our data demonstrate novel mechanism controlling BK channel activity in native cells. We conclude that SACs and BK channels are clusterized in functional mechanosensitive domains in the plasma membrane of hMESCs. Co-clustering of ion channels may significantly contribute to mechano-dependent calcium signalling in stem cells. - Highlights: • Stretch-induced channel activity in human mesenchymal stem cells was analyzed. • Functional expression of SACs and Ca{sup 2+}-sensitive BK and SK channels was shown. • Local Ca{sup 2+} influx via stretch-activated channels triggers BK channel activity. • SK channels are not coupled with SACs despite higher sensitivity to [Ca{sup 2+}]{sub i}. • Functional clustering of SACs and BK channels in stem cell membrane is proposed.« less

Differential distribution of the sodium‐activated potassium channels slick and slack in mouse brain

PubMed Central

Knaus, Hans‐Günther; Schwarzer, Christoph

2015-01-01

ABSTRACT The sodium‐activated potassium channels Slick (Slo2.1, KCNT2) and Slack (Slo2.2, KCNT1) are high‐conductance potassium channels of the Slo family. In neurons, Slick and Slack channels are involved in the generation of slow afterhyperpolarization, in the regulation of firing patterns, and in setting and stabilizing the resting membrane potential. The distribution and subcellular localization of Slick and Slack channels in the mouse brain have not yet been established in detail. The present study addresses this issue through in situ hybridization and immunohistochemistry. Both channels were widely distributed and exhibited distinct distribution patterns. However, in some brain regions, their expression overlapped. Intense Slick channel immunoreactivity was observed in processes, varicosities, and neuronal cell bodies of the olfactory bulb, granular zones of cortical regions, hippocampus, amygdala, lateral septal nuclei, certain hypothalamic and midbrain nuclei, and several regions of the brainstem. The Slack channel showed primarily a diffuse immunostaining pattern, and labeling of cell somata and processes was observed only occasionally. The highest Slack channel expression was detected in the olfactory bulb, lateral septal nuclei, basal ganglia, and distinct areas of the midbrain, brainstem, and cerebellar cortex. In addition, comparing our data obtained from mouse brain with a previously published study on rat brain revealed some differences in the expression and distribution of Slick and Slack channels in these species. J. Comp. Neurol. 524:2093–2116, 2016. © 2015 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc. PMID:26587966

Reciprocal voltage sensor-to-pore coupling leads to potassium channel C-type inactivation

PubMed Central

Conti, Luca; Renhorn, Jakob; Gabrielsson, Anders; Turesson, Fredrik; Liin, Sara I; Lindahl, Erik; Elinder, Fredrik

2016-01-01

Voltage-gated potassium channels open at depolarized membrane voltages. A prolonged depolarization causes a rearrangement of the selectivity filter which terminates the conduction of ions – a process called slow or C-type inactivation. How structural rearrangements in the voltage-sensor domain (VSD) cause alteration in the selectivity filter, and vice versa, are not fully understood. We show that pulling the pore domain of the Shaker potassium channel towards the VSD by a Cd2+ bridge accelerates C-type inactivation. Molecular dynamics simulations show that such pulling widens the selectivity filter and disrupts the K+ coordination, a hallmark for C-type inactivation. An engineered Cd2+ bridge within the VSD also affect C-type inactivation. Conversely, a pore domain mutation affects VSD gating-charge movement. Finally, C-type inactivation is caused by the concerted action of distant amino acid residues in the pore domain. All together, these data suggest a reciprocal communication between the pore domain and the VSD in the extracellular portion of the channel. PMID:27278891

Reciprocal voltage sensor-to-pore coupling leads to potassium channel C-type inactivation

NASA Astrophysics Data System (ADS)

Conti, Luca; Renhorn, Jakob; Gabrielsson, Anders; Turesson, Fredrik; Liin, Sara I.; Lindahl, Erik; Elinder, Fredrik

2016-06-01

Voltage-gated potassium channels open at depolarized membrane voltages. A prolonged depolarization causes a rearrangement of the selectivity filter which terminates the conduction of ions - a process called slow or C-type inactivation. How structural rearrangements in the voltage-sensor domain (VSD) cause alteration in the selectivity filter, and vice versa, are not fully understood. We show that pulling the pore domain of the Shaker potassium channel towards the VSD by a Cd2+ bridge accelerates C-type inactivation. Molecular dynamics simulations show that such pulling widens the selectivity filter and disrupts the K+ coordination, a hallmark for C-type inactivation. An engineered Cd2+ bridge within the VSD also affect C-type inactivation. Conversely, a pore domain mutation affects VSD gating-charge movement. Finally, C-type inactivation is caused by the concerted action of distant amino acid residues in the pore domain. All together, these data suggest a reciprocal communication between the pore domain and the VSD in the extracellular portion of the channel.

Antibodies to AChR, MuSK and VGKC in a patient with myasthenia gravis and Morvan's syndrome.

PubMed

Díaz-Manera, Jordi; Rojas-García, Ricard; Gallardo, Eduard; Juárez, Cándido; Martínez-Domeño, Alejandro; Martínez-Ramírez, Sergi; Dalmau, Josep; Blesa, Rafael; Illa, Isabel

2007-07-01

A 46-year-old woman presented to a local hospital with acute respiratory failure and a 2-year progressive history of fatigue, personality changes, increased sweating, dysphagia with substantial weight loss, dysarthria, and intermittent ptosis and diplopia. Neurological examination showed facial weakness, lingual atrophy and bulbar palsy, which necessitated the use of a feeding tube and ventilatory support. Mild limb weakness with severe muscle atrophy and diffuse muscle twitches were observed. The patient had also developed visual hallucinations and persecutory delusions. Her personal and family medical histories were unremarkable. Sensory and motor nerve conduction studies, repetitive nerve stimulation, electromyogram, blood-cell counts, general chemistry and metabolic function tests, a CT scan, an [(18)F]fluorodeoxyglucose-PET scan, and tests for serum antibodies to acetylcholine receptors, muscle-specific tyrosine kinase, voltage-gated potassium channels, P/Q-type voltage-gated calcium channels, and paraneoplastic antigens, were carried out. Myasthenia gravis associated with antibodies to acetylcholine receptor and muscle-specific tyrosine kinase, and Morvan's syndrome associated with antibodies to voltage-gated potassium channels in the absence of thymoma. Combined treatment with prednisone, intravenous immunoglobulin, ciclosporin, and rituximab.

Inactivation of the first nucleotide-binding fold of the sulfonylurea receptor, and familial persistent hyperinsulinemic hypoglycemia of infancy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thomas, P.M.; Wohllk, N.; Huang, E.

1996-09-01

Familial persistent hyperinsulinemic hypoglycemia of infancy is a disorder of glucose homeostasis and is characterized by unregulated insulin secretion and profound hypoglycemia. Loss-of-function mutations in the second nucleotide-binding fold of the sulfonylurea receptor, a subunit of the pancreatic-islet {beta}-cell ATP-dependent potassium channel, has been demonstrated to be causative for persistent hyperinsulinemic hypoglycemia of infancy. We now describe three additional mutations in the first nucleotide-binding fold of the sulfonylurea-receptor gene. One point mutation disrupts the highly conserved Walker A motif of the first nucleotide-binding-fold region. The other two mutations occur in noncoding sequences required for RNA processing and are predicted tomore » disrupt the normal splicing pathway of the sulfonylurea-receptor mRNA precursor. These data suggest that both nucleotide-binding-fold regions of the sulfortylurea receptor are required for normal regulation of {beta}-cell ATP-dependent potassium channel activity and insulin secretion. 32 refs., 4 figs., 1 tab.« less

Dynamic redistribution of calcium sensitive potassium channels (hK(Ca)3.1) in migrating cells.

PubMed

Schwab, Albrecht; Nechyporuk-Zloy, Volodymyr; Gassner, Birgit; Schulz, Christoph; Kessler, Wolfram; Mally, Sabine; Römer, Michael; Stock, Christian

2012-02-01

Calcium-sensitive potassium channels (K(Ca)3.1) are expressed in virtually all migrating cells. Their activity is required for optimal cell migration so that their blockade leads to slowing down. K(Ca)3.1 channels must be inserted into the plasma membrane in order to elicit their physiological function. However, the plasma membrane of migrating cells is subject to rapid recycling by means of endo- and exocytosis. Here, we focussed on the endocytic internalization and the intracellular transport of the human isoform hK(Ca)3.1. A hK(Ca)3.1 channel construct with an HA-tag in the extracellularly located S3-S4 linker was transfected into migrating transformed renal epithelial MDCK-F cells. Channel internalization was visualized and quantified with immunofluorescence and a cell-based ELISA. Movement of hK(Ca)3.1 channel containing vesicles as well as migration of MDCK-F cells were monitored by means of time lapse video microscopy. hK(Ca)3.1 channels are endocytosed during migration. Most of the hK(Ca)3.1 channel containing vesicles are moving at a speed of up to 2 µm/sec in a microtubule-dependent manner towards the front of MDCK-F cells. Our experiments indicate that endocytosis of hK(Ca)3.1 channels is clathrin-dependent since they colocalize with clathrin adaptor proteins and since it is impaired when a C-terminal dileucine motif is mutated. The C-terminal dileucine motif is also important for the subcellular localization of hK(Ca)3.1 channels in migrating cells. Mutated channels are no longer concentrated at the leading edge. We therefore propose that recycling of hK(Ca)3.1 channels contributes to their characteristic subcellular distribution in migrating cells. Copyright © 2011 Wiley Periodicals, Inc.

‘And then there were three’: highly efficient uptake of potassium by foliar trichomes of epiphytic bromeliads

PubMed Central

Winkler, Uwe; Zotz, Gerhard

2010-01-01

Background and Aims Vascular epiphytes have to acquire nutrients from atmospheric wash out, stem-flow, canopy soils and trapped litter. Physiological studies on the adaptations to nutrient acquisition and plant utilization of nutrients have focused on phosphorus and nitrogen; potassium, as a third highly abundant nutrient element, has received minor attention. In the present study, potassium uptake kinetics by leaves, within-plant distribution and nutrient accumulation were analysed to gain an improved understanding of physiological adaptations to non-terrestrial nutrient supply of plants. Methods Radioactively labelled 86RbCl was used as an analogue to study uptake kinetics of potassium absorbed from tanks of epiphytes, its plant distribution and the correlation between uptake efficiency and abundance of trichomes, functioning as uptake organs of leaves. Potassium in leaves was additionally analysed by atomic absorption spectroscopy to assess plant responses to potassium deficiency. Key Results Labelled rubidium was taken up from tanks over a wide range of concentrations, 0·01–90 mm, which was achieved by two uptake systems. In four tank epiphytes, the high-affinity transporters had average Km values of 41·2 µm, and the low-affinity transporters average Km values of 44·8 mm. Further analysis in Vriesea splenriet showed that high-affinity uptake of rubidium was an ATP-dependent process, while low-affinity uptake was mediated by a K+-channel. The kinetic properties of both types of transporters are comparable with those of potassium transporters in roots of terrestrial plants. Specific differences in uptake velocities of epiphytes are correlated with the abundance of trichomes on their leaf surfaces. The main sinks for potassium were fully grown leaves. These leaves thus function as internal potassium sources, which allow growth to be maintained during periods of low external potassium availability. Conclusions Vascular epiphytes possess effective mechanisms to take up potassium from both highly diluted and highly concentrated solutions, enabling the plant to incorporate this nutrient element quickly and almost quantitatively from tank solutions. A surplus not needed for current metabolism is stored, i.e. plants show luxury consumption. PMID:20542886

Cytoplasmic Domains and Voltage-Dependent Potassium Channel Gating

PubMed Central

Barros, Francisco; Domínguez, Pedro; de la Peña, Pilar

2012-01-01

The basic architecture of the voltage-dependent K+ channels (Kv channels) corresponds to a transmembrane protein core in which the permeation pore, the voltage-sensing components and the gating machinery (cytoplasmic facing gate and sensor–gate coupler) reside. Usually, large protein tails are attached to this core, hanging toward the inside of the cell. These cytoplasmic regions are essential for normal channel function and, due to their accessibility to the cytoplasmic environment, constitute obvious targets for cell-physiological control of channel behavior. Here we review the present knowledge about the molecular organization of these intracellular channel regions and their role in both setting and controlling Kv voltage-dependent gating properties. This includes the influence that they exert on Kv rapid/N-type inactivation and on activation/deactivation gating of Shaker-like and eag-type Kv channels. Some illustrative examples about the relevance of these cytoplasmic domains determining the possibilities for modulation of Kv channel gating by cellular components are also considered. PMID:22470342

Select α-arrestins control cell-surface abundance of the mammalian Kir2.1 potassium channel in a yeast model.

PubMed

Hager, Natalie A; Krasowski, Collin J; Mackie, Timothy D; Kolb, Alexander R; Needham, Patrick G; Augustine, Andrew A; Dempsey, Alison; Szent-Gyorgyi, Christopher; Bruchez, Marcel P; Bain, Daniel J; Kwiatkowski, Adam V; O'Donnell, Allyson F; Brodsky, Jeffrey L

2018-05-21

Protein composition at the plasma membrane is tightly regulated, with rapid protein internalization and selective targeting to the cell surface occurring in response to environmental changes. For example, ion channels are dynamically relocalized to or from the plasma membrane in response to physiological alterations, allowing cells and organisms to maintain osmotic and salt homeostasis. To identify additional factors that regulate the selective trafficking of a specific ion channel, we used a yeast model for a mammalian potassium channel, the K+ inwardly rectifying channel Kir2.1. Kir2.1 maintains potassium homeostasis in heart muscle cells, and Kir2.1 defects lead to human disease. By examining the ability of Kir2.1 to rescue the growth of yeast cells lacking endogenous potassium channels, we discovered that specific α-arrestins regulate Kir2.1 localization. Specifically, we found that the Ldb19/Art1, Aly1/Art6, and Aly2/Art3 α-arrestin adaptor proteins promote Kir2.1 trafficking to the cell surface, increase Kir2.1 activity at the plasma membrane, and raise intracellular potassium levels. To better quantify the intracellular and cell-surface populations of Kir2.1, we created fluorescence-activating protein fusions and for the first time used this technique to measure the cell-surface residency of a plasma membrane protein in yeast. Our experiments revealed that two α-arrestin effectors also control Kir2.1 localization. In particular, both the Rsp5 ubiquitin ligase and the protein phosphatase calcineurin facilitated the α-arrestin-mediated trafficking of Kir2.1. Together, our findings implicate α-arrestins in regulating an additional class of plasma membrane proteins and establish a new tool for dissecting the trafficking itinerary of any membrane protein in yeast. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Molecular mechanism of a COOH-terminal gating determinant in the ROMK channel revealed by a Bartter's disease mutation

PubMed Central

Flagg, Thomas P; Yoo, Dana; Sciortino, Christopher M; Tate, Margaret; Romero, Michael F; Welling, Paul A

2002-01-01

The ROMK subtypes of inward-rectifier K+ channels mediate potassium secretion and regulate NaCl reabsorption in the kidney. Loss-of-function mutations in this pH-sensitive K+ channel cause Bartter's disease, a familial salt wasting nephropathy. One disease-causing mutation truncates the extreme COOH-terminus and induces a closed gating conformation. Here we identify a region within the deleted domain that plays an important role in pH-dependent gating. The domain contains a structural element that functionally interacts with the pH sensor in the cytoplasmic NH2-terminus to set a physiological range of pH sensitivity. Removal of the domain shifts the pKa towards alkaline pH values, causing channel inactivation under physiological conditions. Suppressor mutations within the pH sensor rescued channel gating and trans addition of the cognate peptide restored pH sensitivity. A specific interdomain interaction was revealed in an in vitro protein-protein binding assay between the NH2- and COOH-terminal cytoplasmic domains expressed as bacterial fusion proteins. These results provide new insights into the molecular mechanisms underlying Kir channel regulation and channel gating defects that are associated with Bartter's disease. PMID:12381810

The Caenorhabditis elegans Iodotyrosine Deiodinase Ortholog SUP-18 Functions through a Conserved Channel SC-Box to Regulate the Muscle Two-Pore Domain Potassium Channel SUP-9

PubMed Central

de la Cruz, Ignacio Perez; Ma, Long; Horvitz, H. Robert

2014-01-01

Loss-of-function mutations in the Caenorhabditis elegans gene sup-18 suppress the defects in muscle contraction conferred by a gain-of-function mutation in SUP-10, a presumptive regulatory subunit of the SUP-9 two-pore domain K+ channel associated with muscle membranes. We cloned sup-18 and found that it encodes the C. elegans ortholog of mammalian iodotyrosine deiodinase (IYD), an NADH oxidase/flavin reductase that functions in iodine recycling and is important for the biosynthesis of thyroid hormones that regulate metabolism. The FMN-binding site of mammalian IYD is conserved in SUP-18, which appears to require catalytic activity to function. Genetic analyses suggest that SUP-10 can function with SUP-18 to activate SUP-9 through a pathway that is independent of the presumptive SUP-9 regulatory subunit UNC-93. We identified a novel evolutionarily conserved serine-cysteine-rich region in the C-terminal cytoplasmic domain of SUP-9 required for its specific activation by SUP-10 and SUP-18 but not by UNC-93. Since two-pore domain K+ channels regulate the resting membrane potentials of numerous cell types, we suggest that the SUP-18 IYD regulates the activity of the SUP-9 channel using NADH as a coenzyme and thus couples the metabolic state of muscle cells to muscle membrane excitability. PMID:24586202

Negative inotropic effect of carbachol and interaction between acetylcholine receptor-operated potassium channel (K.ACh channel) and GTP binding protein in mouse isolated atrium--a novel methodological trial.

PubMed

Okada, Muneyoshi; Noma, Chihiro; Yamawaki, Hideyuki; Hara, Yukio

2013-01-01

Interaction between acetylcholine receptor-operated potassium channel (K.ACh channel) and GTP binding protein was examined by an immunoprecipitation-Western blotting system in mouse isolated atrium. The carbachol-induced negative inotropic action in indomethacin-pretreated mouse atrium was significantly inhibited by a K.ACh channel blocker, tertiapin or atropine. Kir3.1 K.ACh channel (Kir3.1) was immunoprecipitated with a mouse anti-Kir3.1 antibody. Coprecipitating Gβ with Kir3.1, detected by Western blotting, was significantly augmented by carbachol. Atropine, but not tertiapin, significantly inhibited the carbachol-induced coprecipitating Gβ with Kir3.1. The data indicate that immunoprecipitation with Kir3.1 and Western blotting of Gβ system is a useful method for assessing interaction between K.ACh channel and GTP binding protein in mouse atrium.

Insights in KIR2.1 channel structure and function by an evolutionary approach; cloning and functional characterization of the first reptilian inward rectifier channel KIR2.1, derived from the California kingsnake (Lampropeltis getula californiae).

PubMed

Houtman, Marien J C; Korte, Sanne M; Ji, Yuan; Kok, Bart; Vos, Marc A; Stary-Weinzinger, Anna; van der Heyden, Marcel A G

2014-10-03

Potassium inward rectifier KIR2.1 channels contribute to the stable resting membrane potential in a variety of muscle and neuronal cell-types. Mutations in the KIR2.1 gene KCNJ2 have been associated with human disease, such as cardiac arrhythmias and periodic paralysis. Crystal structure and homology modelling of KIR2.1 channels combined with functional current measurements provided valuable insights in mechanisms underlying channel function. KIR2.1 channels have been cloned and analyzed from all main vertebrate phyla, except reptilians. To address this lacuna, we set out to clone reptilian KIR2.1 channels. Using a degenerated primer set we cloned the KCNJ2 coding regions from muscle tissue of turtle, snake, bear, quail and bream, and compared their deduced amino acid sequences with those of KIR2.1 sequences from 26 different animal species obtained from Genbank. Furthermore, expression constructs were prepared for functional electrophysiological studies of ectopically expressed KIR2.1 ion channels. In general, KCNJ2 gene evolution followed normal phylogenetic patterns, however turtle KIR2.1 ion channel sequence is more homologues to avians than to snake. Alignment of all 31 KIR2.1 sequences showed that all disease causing KIR2.1 mutations, except V93I, V123G and N318S, are fully conserved. Homology models were built to provide structural insights into species specific amino acid substitutions. Snake KIR2.1 channels became expressed at the plasmamembrane and produced typical barium sensitive (IC50 ∼6μM) inward rectifier currents. Copyright © 2014 Elsevier Inc. All rights reserved.

ATP-sensitive potassium-channel inhibitor glibenclamide attenuates HPA axis hyperactivity, depression- and anxiety-related symptoms in a rat model of Alzheimer's disease.

PubMed

Esmaeili, Mohammad Hossein; Bahari, Behnam; Salari, Ali-Akbar

2018-03-01

Affective disorders including depression and anxiety are among the most prevalent behavioral abnormalities in patients with Alzheimer's disease (AD), which affect the quality of life and progression of the disease. Dysregulation of the hypothalamic-pituitary-adrenal-(HPA) axis has been reported in affective disorders and AD. Recent studies revealed that current antidepressant drugs are not completely effective for treating anxiety- and depression-related disorders in people with dementia. ATP-sensitive-potassium-(K ATP ) channels are well-known to be involved in AD pathophysiology, HPA axis function and the pathogenesis of depression and anxiety-related behaviors. Thus, targeting of K ATP channel may be a potential therapeutic strategy in AD. Hence, we investigated the effects of intracerebroventricular injection of Aβ25-35 alone or in combination with glibenclamide, K ATP channel inhibitor on depression- and anxiety-related behaviors as well as HPA axis response to stress in rats. To do this, non-Aβ25-35- and Aβ25-35-treated rats were orally treated with glibenclamide, then the behavioral consequences were assessed using sucrose preference, forced swim, light-dark box and plus maze tests. Stress-induced corticosterone levels following forced swim and plus maze tests were also evaluated as indicative of abnormal HPA-axis-function. Aβ25-35 induced HPA axis hyperreactivity and increased depression- and anxiety-related symptoms in rats. Our results showed that blockade of K ATP channels with glibenclamide decreased depression- and anxiety-related behaviors by normalizing HPA axis activity in Aβ25-35-treated rats. This study provides additional evidence that Aβ administration can induce depression- and anxiety-like symptoms in rodents, and suggests that K ATP channel inhibitors may be a plausible therapeutic strategy for treating affective disorders in AD patients. Copyright © 2018 Elsevier Inc. All rights reserved.

The polar T1 interface is linked to conformational changes that open the voltage-gated potassium channel.

PubMed

Minor, D L; Lin, Y F; Mobley, B C; Avelar, A; Jan, Y N; Jan, L Y; Berger, J M

2000-09-01

Kv voltage-gated potassium channels share a cytoplasmic assembly domain, T1. Recent mutagenesis of two T1 C-terminal loop residues implicates T1 in channel gating. However, structural alterations of these mutants leave open the question concerning direct involvement of T1 in gating. We find in mammalian Kv1.2 that gating depends critically on residues at complementary T1 surfaces in an unusually polar interface. An isosteric mutation in this interface causes surprisingly little structural alteration while stabilizing the closed channel and increasing the stability of T1 tetramers. Replacing T1 with a tetrameric coiled-coil destabilizes the closed channel. Together, these data suggest that structural changes involving the buried polar T1 surfaces play a key role in the conformational changes leading to channel opening.

Kv1 channels and neural processing in vestibular calyx afferents.

PubMed

Meredith, Frances L; Kirk, Matthew E; Rennie, Katherine J

2015-01-01

Potassium-selective ion channels are important for accurate transmission of signals from auditory and vestibular sensory end organs to their targets in the central nervous system. During different gravity conditions, astronauts experience altered input signals from the peripheral vestibular system resulting in sensorimotor dysfunction. Adaptation to altered sensory input occurs, but it is not explicitly known whether this involves synaptic modifications within the vestibular epithelia. Future investigations of such potential plasticity require a better understanding of the electrophysiological mechanisms underlying the known heterogeneity of afferent discharge under normal conditions. This study advances this understanding by examining the role of the Kv1 potassium channel family in mediating action potentials in specialized vestibular afferent calyx endings in the gerbil crista and utricle. Pharmacological agents selective for different sub-types of Kv1 channels were tested on membrane responses in whole cell recordings in the crista. Kv1 channels sensitive to α-dendrotoxin and dendrotoxin-K were found to prevail in the central regions, whereas K(+) channels sensitive to margatoxin, which blocks Kv1.3 and 1.6 channels, were more prominent in peripheral regions. Margatoxin-sensitive currents showed voltage-dependent inactivation. Dendrotoxin-sensitive currents showed no inactivation and dampened excitability in calyces in central neuroepithelial regions. The differential distribution of Kv1 potassium channels in vestibular afferents supports their importance in accurately relaying gravitational and head movement signals through specialized lines to the central nervous system. Pharmacological modulation of specific groups of K(+) channels could help alleviate vestibular dysfunction on earth and in space.

Involvement of ATP-sensitive potassium channels and the opioid system in the anticonvulsive effect of zolpidem in mice.

PubMed

Sheikhi, Mehdi; Shirzadian, Armin; Dehdashtian, Amir; Amiri, Shayan; Ostadhadi, Sattar; Ghasemi, Mehdi; Dehpour, Ahmad Reza

2016-09-01

Zolpidem is a hypnotic medication that mainly exerts its function through activating γ-aminobutyric acid (GABA)A receptors. There is some evidence that zolpidem may have anticonvulsive effects. However, the mechanisms underlying this effect have not been elucidated yet. In the present study, we used the pentylentetrazole (PTZ)-induced generalized seizure model in mice to investigate whether zolpidem can affect seizure threshold. We also further evaluated the roles of ATP-sensitive potassium (KATP) channels as well as μ-opioid receptors in the effects of zolpidem on seizure threshold. Our data showed that zolpidem in a dose-dependent manner increased the PTZ-induced seizure threshold. The noneffective (i.e., did not significantly alter the PTZ-induced seizure threshold by itself) doses of KATP channel blocker (glibenclamide) and nonselective opioid receptor antagonist (naloxone) were able to inhibit the anticonvulsive effect of zolpidem. Additionally, noneffective doses of either KATP channel opener (cromakalim) or nonselective μ-opioid receptor agonist (morphine) in combination with a noneffective dose of zolpidem exerted a significant anticonvulsive effect on PTZ-induced seizures in mice. A combination of noneffective doses of naloxone and glibenclamide, which separately did not affect zolpidem effect on seizure threshold, inhibited the anticonvulsive effects of zolpidem. These results suggest a role for KATP channels and the opioid system, alone or in combination, in the anticonvulsive effects of zolpidem. Copyright © 2016 Elsevier Inc. All rights reserved.

Amino acid residues involved in membrane insertion and pore formation of Clostridium botulinum C2 toxin.

PubMed

Lang, Alexander E; Neumeyer, Tobias; Sun, Jianjun; Collier, R John; Benz, Roland; Aktories, Klaus

2008-08-12

The actin-ADP-ribosylating Clostridium botulinum C2 toxin consists of the enzymatic component C2I and the binding component C2II. C2II forms heptameric channels involved in translocation of the enzymatic component into the target cell. On the basis of the heptameric toxin channel, we studied functional consequences of mutagenesis of amino acid residues probably lining the lumen of the toxin channel. Substitution of glutamate-399 of C2II with alanine blocked channel formation and cytotoxicity of the holotoxin. Although cytotoxicity and rounding up of cells by C2I were completely blocked by exchange of phenylalanine-428 with alanine, the mutation increased potassium conductance caused by C2II in artificial membranes by about 2-3-fold over that of wild-type toxin. In contrast to its effects on single-channel potassium conductance in artificial membranes, the F428A mutation delayed the kinetics of pore formation in lipid vesicles and inhibited the activity of C2II in promoting (86)Rb (+) release from preloaded intact cells after pH shift of the medium. Moreover, F428A C2II exhibited delayed and diminished formation of C2II aggregates at low pH, indicating major changes of the biophysical properties of the toxin. The data indicate that phenylalanine-428 of C2II plays a major role in conformational changes occurring during pore formation of the binding component of C2II.

Osteogenic and chondrogenic master genes expression is dependent on the Kir2.1 potassium channel through the bone morphogenetic protein pathway.

PubMed

Pini, Jonathan; Giuliano, Serena; Matonti, Julia; Simkin, Dina; Rouleau, Matthieu; Bendahhou, Saïd

2018-05-29

Andersen's syndrome is a rare disorder affecting muscle, heart, and bone, that is associated with mutations leading to a loss of function of the inwardly rectifying K + channel Kir2.1. While the Kir2.1 function can be anticipated in excitable cells by controlling the electrical activity, its role in non-excitable cells remains to be investigated. Using Andersen's syndrome induced Pluripotent Stem cells, we investigated the cellular and molecular events during the osteoblastic and chondrogenic differentiation that are affected by the loss of the Ik1 current. We show that loss of Kir2.1 channel function impairs both osteoblastic and chondrogenic processes through the down regulation master gene expression. This down regulation is due to an impairment of the bone morphogenetic proteins signaling pathway through de-phosphorylation of the Smad proteins. Restoring Kir2.1 channel function in Andersen's syndrome cells rescued master genes expression, and restored normal osteoblasts and chondrocytes behavior. Our results show that Kir2.1-mediated activity controls endochondral and intramembranous ossification signaling pathways. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

KCNE4 and KCNE5: K+ channel regulation and cardiac arrhythmogenesis

PubMed Central

Abbott, Geoffrey W.

2016-01-01

KCNE proteins are single transmembrane-segment voltage-gated potassium (Kv) channel ancillary subunits that exhibit a diverse range of physiological functions. Human KCNE gene mutations are associated with various pathophysiological states, most notably cardiac arrhythmias. Of the five isoforms in the human KCNE gene family, KCNE4 and the X-linked KCNE5 are, to date, the least-studied. Recently, however, interest in these neglected genes has been stoked by their putative association with debilitating or lethal cardiac arrhythmias. The sometimes-overlapping functional effects of KCNE4 and KCNE5 vary depending on both their Kv α subunit partner and on other ancillary subunits within the channel complex, but mostly fall into two contrasting categories either inhibition, or fine-tuning of gating kinetics. This review covers current knowledge regarding the molecular mechanisms of KCNE4 and KCNE5 function, human disease associations, and findings from very recent studies of cardiovascular pathophysiology in Kcne4−/− mice. PMID:27484720

KCNE4 and KCNE5: K(+) channel regulation and cardiac arrhythmogenesis.

PubMed

Abbott, Geoffrey W

2016-11-30

KCNE proteins are single transmembrane-segment voltage-gated potassium (Kv) channel ancillary subunits that exhibit a diverse range of physiological functions. Human KCNE gene mutations are associated with various pathophysiological states, most notably cardiac arrhythmias. Of the five isoforms in the human KCNE gene family, KCNE4 and the X-linked KCNE5 are, to date, the least-studied. Recently, however, interest in these neglected genes has been stoked by their putative association with debilitating or lethal cardiac arrhythmias. The sometimes-overlapping functional effects of KCNE4 and KCNE5 vary depending on both their Kv α subunit partner and on other ancillary subunits within the channel complex, but mostly fall into two contrasting categories - either inhibition, or fine-tuning of gating kinetics. This review covers current knowledge regarding the molecular mechanisms of KCNE4 and KCNE5 function, human disease associations, and findings from very recent studies of cardiovascular pathophysiology in Kcne4(-/-) mice. Copyright © 2016 Elsevier B.V. All rights reserved.

Physiological roles of Kv2 channels in entorhinal cortex layer II stellate cells revealed by Guangxitoxin‐1E

PubMed Central

Hönigsperger, Christoph; Nigro, Maximiliano J.

2016-01-01

Key points Kv2 channels underlie delayed‐rectifier potassium currents in various neurons, although their physiological roles often remain elusive. Almost nothing is known about Kv2 channel functions in medial entorhinal cortex (mEC) neurons, which are involved in representing space, memory formation, epilepsy and dementia.Stellate cells in layer II of the mEC project to the hippocampus and are considered to be space‐representing grid cells. We used the new Kv2 blocker Guangxitoxin‐1E (GTx) to study Kv2 functions in these neurons.Voltage clamp recordings from mEC stellate cells in rat brain slices showed that GTx inhibited delayed‐rectifier K+ current but not transient A‐type current.In current clamp, GTx had multiple effects: (i) increasing excitability and bursting at moderate spike rates but reducing firing at high rates; (ii) enhancing after‐depolarizations; (iii) reducing the fast and medium after‐hyperpolarizations; (iv) broadening action potentials; and (v) reducing spike clustering.GTx is a useful tool for studying Kv2 channels and their functions in neurons. Abstract The medial entorhinal cortex (mEC) is strongly involved in spatial navigation, memory, dementia and epilepsy. Although potassium channels shape neuronal activity, their roles in mEC are largely unknown. We used the new Kv2 blocker Guangxitoxin‐1E (GTx; 10–100 nm) in rat brain slices to investigate Kv2 channel functions in mEC layer II stellate cells (SCs). These neurons project to the hippocampus and are considered to be grid cells representing space. Voltage clamp recordings from SCs nucleated patches showed that GTx inhibited a delayed rectifier K+ current activating beyond –30 mV but not transient A‐type current. In current clamp, GTx (i) had almost no effect on the first action potential but markedly slowed repolarization of late spikes during repetitive firing; (ii) enhanced the after‐depolarization (ADP); (iii) reduced fast and medium after‐hyperpolarizations (AHPs); (iv) strongly enhanced burst firing and increased excitability at moderate spike rates but reduced spiking at high rates; and (v) reduced spike clustering and rebound potentials. The changes in bursting and excitability were related to the altered ADPs and AHPs. Kv2 channels strongly shape the activity of mEC SCs by affecting spike repolarization, after‐potentials, excitability and spike patterns. GTx is a useful tool and may serve to further clarify Kv2 channel functions in neurons. We conclude that Kv2 channels in mEC SCs are important determinants of intrinsic properties that allow these neurons to produce spatial representation. The results of the present study may also be important for the accurate modelling of grid cells. PMID:27562026

Mechanisms of action of ligands of potential-dependent sodium channels.

PubMed

Tikhonov, D B

2008-06-01

Potential-dependent sodium channels play a leading role in generating action potentials in excitable cells. Sodium channels are the site of action of a variety of modulator ligands. Despite numerous studies, the mechanisms of action of many modulators remain incompletely understood. The main reason that many important questions cannot be resolved is that there is a lack of precise data on the structures of the channels themselves. Structurally, potential-dependent sodium channels are members of the P-loop channel superfamily, which also include potassium and calcium channels and glutamate receptor channels. Crystallization of a series of potassium channels showed that it was possible to analyze the structures of different members of the superfamily using the "homologous modeling" method. The present study addresses model investigations of the actions of ligands of sodium channels, including tetrodotoxin and batrachotoxin, as well as local anesthetics. Comparison of experimental data on sodium channel ligands with x-ray analysis data allowed us to reach a new level of understanding of the mechanisms of channel modulation and to propose a series of experimentally verifiable hypotheses.

Diabetes attenuates urothelial modulation of detrusor contractility and spontaneous activity.

PubMed

Wang, Yi; Tar, Moses T; Fu, Shibo; Melman, Arnold; Davies, Kelvin P

2014-10-01

To investigate the effect of diabetes on urothelial modulation of bladder contractility. Bladder strips (urothelium intact or denuded) were prepared from 8-week-old streptozotocin-induced diabetic (n = 19) and non-diabetic control rats (n = 10). The effect of modulators of MaxiK (iberiotoxin and tetraethylammonium) and Kv7 (XE991 and retigabine) potassium channel activity were investigated for their effects on both carbachol-induced force generation and spontaneous contractile activity. In bladder strips from non-diabetic animals, the presence of the urothelium resulted in marked sensitivity to carbachol-induced force generation by modulators of MaxiK and Kv7 channel activity, whereas in the diabetic animal urothelial sensitivity to these agents was significantly diminished. Urothelial-intact bladder strips from non-diabetic animals were more sensitive to modulators of Kv7 activity in reducing the amplitude of spontaneous phasic contractions than urothelial-denuded bladder strips, whereas in diabetic animals the presence or absence of the urothelium did not alter the sensitivity to modulators of Kv7 activity. Spontaneous activity in the presence of tetraethylammonium was not affected by the urothelium in bladder strips from either diabetic or non-diabetic animals. The presence of the urothelium in bladders from non-diabetic animals modulates the activity of potassium blockers to affect bladder contractility, whereas in the diabetic bladder this effect is attenuated. These findings could help to explain the lack of success of pharmaceutical treatments targeting potassium channels to treat bladder pathology in patients with diseases imparing urothelial function. © 2014 The Japanese Urological Association.

Genomics analysis of potassium channel genes in songbirds reveals molecular specializations of brain circuits for the maintenance and production of learned vocalizations

PubMed Central

2013-01-01

Background A fundamental question in molecular neurobiology is how genes that determine basic neuronal properties shape the functional organization of brain circuits underlying complex learned behaviors. Given the growing availability of complete vertebrate genomes, comparative genomics represents a promising approach to address this question. Here we used genomics and molecular approaches to study how ion channel genes influence the properties of the brain circuitry that regulates birdsong, a learned vocal behavior with important similarities to human speech acquisition. We focused on potassium (K-)Channels, which are major determinants of neuronal cell excitability. Starting with the human gene set of K-Channels, we used cross-species mRNA/protein alignments, and syntenic analysis to define the full complement of orthologs, paralogs, allelic variants, as well as novel loci not previously predicted in the genome of zebra finch (Taeniopygia guttata). We also compared protein coding domains in chicken and zebra finch orthologs to identify genes under positive selective pressure, and those that contained lineage-specific insertions/deletions in functional domains. Finally, we conducted comprehensive in situ hybridizations to determine the extent of brain expression, and identify K-Channel gene enrichments in nuclei of the avian song system. Results We identified 107 K-Channel finch genes, including 6 novel genes common to non-mammalian vertebrate lineages. Twenty human genes are absent in songbirds, birds, or sauropsids, or unique to mammals, suggesting K-Channel properties may be lineage-specific. We also identified specific family members with insertions/deletions and/or high dN/dS ratios compared to chicken, a non-vocal learner. In situ hybridization revealed that while most K-Channel genes are broadly expressed in the brain, a subset is selectively expressed in song nuclei, representing molecular specializations of the vocal circuitry. Conclusions Together, these findings shed new light on genes that may regulate biophysical and excitable properties of the song circuitry, identify potential targets for the manipulation of the song system, and reveal genomic specializations that may relate to the emergence of vocal learning and associated brain areas in birds. PMID:23845108

Opening of the inward rectifier potassium channel alleviates maladaptive tissue repair following myocardial infarction.

PubMed

Liu, Chengfang; Liu, Enli; Luo, Tiane; Zhang, Weifang; He, Rongli

2016-08-01

Activation of the inward rectifier potassium current (IK1) channel has been reported to be associated with suppression of ventricular arrhythmias. In this study, we tested the hypothesis that opening of the IK1 channel with zacopride (ZAC) was involved in the modulation of tissue repair after myocardial infarction. Sprague-Dawley rats were subject to coronary artery ligation and ZAC was administered intraperitoneally (15 µg/kg/day) for 28 days. Compared with the ischemia group, treatment with ZAC significantly reduced the ratio of heart/body weight and the cross-sectional area of cardiomyocytes, suggesting less cardiac hypertrophy. ZAC reduced the accumulation of collagen types I and III, accompanied with decrease of collagen area, which were associated with a reduction of collagen deposition in the fibrotic myocardium. Echocardiography showed improved cardiac function, evidenced by the reduced left ventricular end-diastolic dimension and left ventricular end-systolic dimension, and the increased ejection fraction and fractional shortening in ZAC-treated animals (all P < 0.05 vs. ischemia group). In coincidence with these changes, ZAC up-regulated the protein level of the IK1 channel and down-regulated the phosphorylation of mammalian target of rapamycin (mTOR) and 70-kDa ribosomal protein S6 (p70S6) kinase. Administration of chloroquine alone, an IK1 channel antagonist, had no effect on all the parameters measured, but significantly blocked the beneficial effects of ZAC on cardiac repair. In conclusion, opening of the IK1 channel with ZAC inhibits maladaptive tissue repair and improves cardiac function, potentially mediated by the inhibition of ischemia-activated mTOR-p70S6 signaling pathway via the IK1 channel. So the development of pharmacological agents specifically targeting the activation of the IK1 channel may protect the heart against myocardial ischemia-induced cardiac dysfunction. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Interaction with caveolin-1 modulates vascular ATP-sensitive potassium (KATP) channel activity

PubMed Central

Davies, Lowri M; Purves, Gregor I; Barrett-Jolley, Richard; Dart, Caroline

2010-01-01

ATP-sensitive potassium channels (KATP channels) of arterial smooth muscle are important regulators of arterial tone, and hence blood flow, in response to vasoactive transmitters. Recent biochemical and electron microscopic evidence suggests that these channels localise to small vesicular invaginations of the plasma membrane, known as caveolae, and interact with the caveolae-associated protein, caveolin. Here we report that interaction with caveolin functionally regulates the activity of the vascular subtype of KATP channel, Kir6.1/SUR2B. Pinacidil-evoked recombinant whole-cell Kir6.1/SUR2B currents recorded in HEK293 cells stably expressing caveolin-1 (69.6 ± 8.3 pA pF−1, n= 8) were found to be significantly smaller than currents recorded in caveolin-null cells (179.7 ± 35.9 pA pF−1, n= 6; P < 0.05) indicating that interaction with caveolin may inhibit channel activity. Inclusion in the pipette-filling solution of a peptide corresponding to the scaffolding domain of caveolin-1 had a similar inhibitory effect on whole-cell Kir6.1/SUR2B currents as co-expression with full-length caveolin-1, while a scrambled version of the same peptide had no effect. Interestingly, intracellular dialysis of vascular smooth muscle cells with the caveolin-1 scaffolding domain peptide (SDP) also caused inhibition of pinacidil-evoked native whole-cell KATP currents, indicating that a significant proportion of vascular KATP channels are susceptible to block by exogenously applied SDP. In cell-attached recordings of Kir6.1/SUR2B single channel activity, the presence of caveolin-1 significantly reduced channel open probability (from 0.05 ± 0.01 to 0.005 ± 0.001; P < 0.05) and the amount of time spent in a relatively long-lived open state. These changes in kinetic behaviour can be explained by a caveolin-induced shift in the channel's sensitivity to its physiological regulator MgADP. Our findings thus suggest that interaction with caveolin-1 suppresses vascular-type KATP channel activity. Since caveolin expression is regulated by cellular free cholesterol and plasma levels of low-density lipoprotein (LDL), this interaction may have implications in both the physiological and pathophysiological control of vascular function. PMID:20624795

The potent insulin secretagogue effect of betulinic acid is mediated by potassium and chloride channels.

PubMed

Gomes Castro, Allisson Jhonatan; Cazarolli, Luisa Helena; Bretanha, Lizandra C; Sulis, Paola Miranda; Rey Padilla, Diana Patricia; Aragón Novoa, Diana Marcela; Dambrós, Betina Fernanda; Pizzolatti, Moacir G; Mena Barreto Silva, Fátima Regina

2018-06-15

Betulinic acid (BA) has been described as an insulin secretagogue which may explain its potent antihyperglycemic effect; however, the exact role of BA as an insulinogenic agent is not clear. The aim of this study was to investigate the mechanism of BA on calcium influx and static insulin secretion in pancreatic islets isolated from euglycemic rats. We found that BA triggers calcium influx by a mechanism dependent on ATP-dependent potassium channels and L-type voltage-dependent calcium channels. Additionally, the voltage-dependent and calcium-dependent chloride channels are also involved in the mechanism of BA, probably due to an indirect stimulation of calcium entry and increased intracellular calcium. Additionally, the downstream activation of PKC, which is necessary for the effect of BA on calcium influx, is involved in the full stimulatory response of the triterpene. BA stimulated the static secretion of insulin in pancreatic islets, indicating that the abrupt calcium influx may be a key step in its secretagogue effect. As such, BA stimulates insulin secretion through the activation of electrophysiological mechanisms, such as the closure of potassium channels and opening of calcium and chloride channels, inducing cellular depolarization associated with metabolic-biochemical effects, in turn activating PKC and ensuring the secretion of insulin. Copyright © 2018 Elsevier Inc. All rights reserved.

The voltage-sensing domain of a phosphatase gates the pore of a potassium channel.

PubMed

Arrigoni, Cristina; Schroeder, Indra; Romani, Giulia; Van Etten, James L; Thiel, Gerhard; Moroni, Anna

2013-03-01

The modular architecture of voltage-gated K(+) (Kv) channels suggests that they resulted from the fusion of a voltage-sensing domain (VSD) to a pore module. Here, we show that the VSD of Ciona intestinalis phosphatase (Ci-VSP) fused to the viral channel Kcv creates Kv(Synth1), a functional voltage-gated, outwardly rectifying K(+) channel. Kv(Synth1) displays the summed features of its individual components: pore properties of Kcv (selectivity and filter gating) and voltage dependence of Ci-VSP (V(1/2) = +56 mV; z of ~1), including the depolarization-induced mode shift. The degree of outward rectification of the channel is critically dependent on the length of the linker more than on its amino acid composition. This highlights a mechanistic role of the linker in transmitting the movement of the sensor to the pore and shows that electromechanical coupling can occur without coevolution of the two domains.

Behavioral and Genetic Evidence for GIRK Channels in the CNS: Role in Physiology, Pathophysiology, and Drug Addiction.

PubMed

Mayfield, Jody; Blednov, Yuri A; Harris, R Adron

2015-01-01

G protein-coupled inwardly rectifying potassium (GIRK) channels are widely expressed throughout the brain and mediate the inhibitory effects of many neurotransmitters. As a result, these channels are important for normal CNS function and have also been implicated in Down syndrome, Parkinson's disease, psychiatric disorders, epilepsy, and drug addiction. Knockout mouse models have provided extensive insight into the significance of GIRK channels under these conditions. This review examines the behavioral and genetic evidence from animal models and genetic association studies in humans linking GIRK channels with CNS disorders. We further explore the possibility that subunit-selective modulators and other advanced research tools will be instrumental in establishing the role of individual GIRK subunits in drug addiction and other relevant CNS diseases and in potentially advancing treatment options for these disorders. © 2015 Elsevier Inc. All rights reserved.

Atomic basis for therapeutic activation of neuronal potassium channels

NASA Astrophysics Data System (ADS)

Kim, Robin Y.; Yau, Michael C.; Galpin, Jason D.; Seebohm, Guiscard; Ahern, Christopher A.; Pless, Stephan A.; Kurata, Harley T.

2015-09-01

Retigabine is a recently approved anticonvulsant that acts by potentiating neuronal M-current generated by KCNQ2-5 channels, interacting with a conserved Trp residue in the channel pore domain. Using unnatural amino-acid mutagenesis, we subtly altered the properties of this Trp to reveal specific chemical interactions required for retigabine action. Introduction of a non-natural isosteric H-bond-deficient Trp analogue abolishes channel potentiation, indicating that retigabine effects rely strongly on formation of a H-bond with the conserved pore Trp. Supporting this model, substitution with fluorinated Trp analogues, with increased H-bonding propensity, strengthens retigabine potency. In addition, potency of numerous retigabine analogues correlates with the negative electrostatic surface potential of a carbonyl/carbamate oxygen atom present in most KCNQ activators. These findings functionally pinpoint an atomic-scale interaction essential for effects of retigabine and provide stringent constraints that may guide rational improvement of the emerging drug class of KCNQ channel activators.

[Modulation of Kv4 channels by KChIPs clamping].

PubMed

Cui, Yuan-Yuan; Wang, Ke-Wei

2009-01-01

The rapidly inactivating (A-type) potassium channels regulate membrane excitability that defines the fundamental mechanism of neuronal functions such as pain signaling. Cytosolic Kv channel-interacting proteins KChIPs co-assemble with Kv4 (Shal) alpha subunits to form a native complex. The specific binding of auxiliary KChIPs to the Kv4 N-terminus results in modulation of gating properties, surface expression and subunit assembly of Kv4 channels. Based on recent structural efforts, here we attempt to emphasize the interaction between KChIPs and Kv4 channel complex in which a single KChIP1 molecule laterally clamps two neighboring Kv4.3 N-termini in a 4:4 manner. Greater insights into molecular mechanism between KChIPs and Kv4 interaction may provide therapeutic potentials by structure-based design of chemical compounds aimed at disrupting the protein-protein interaction for treatment of membrane excitability-related disorders.

BAD and KATP channels regulate neuron excitability and epileptiform activity

PubMed Central

Fernández-Agüera, María Carmen; Nathwani, Nidhi; Lahmann, Carolina; Burnham, Veronica L

2018-01-01

Brain metabolism can profoundly influence neuronal excitability. Mice with genetic deletion or alteration of Bad (BCL-2 agonist of cell death) exhibit altered brain-cell fuel metabolism, accompanied by resistance to acutely induced epileptic seizures; this seizure protection is mediated by ATP-sensitive potassium (KATP) channels. Here we investigated the effect of BAD manipulation on KATP channel activity and excitability in acute brain slices. We found that BAD’s influence on neuronal KATP channels was cell-autonomous and directly affected dentate granule neuron (DGN) excitability. To investigate the role of neuronal KATP channels in the anticonvulsant effects of BAD, we imaged calcium during picrotoxin-induced epileptiform activity in entorhinal-hippocampal slices. BAD knockout reduced epileptiform activity, and this effect was lost upon knockout or pharmacological inhibition of KATP channels. Targeted BAD knockout in DGNs alone was sufficient for the antiseizure effect in slices, consistent with a ‘dentate gate’ function that is reinforced by increased KATP channel activity. PMID:29368690

Differential effect of brief electrical stimulation on voltage-gated potassium channels.

PubMed

Cameron, Morven A; Al Abed, Amr; Buskila, Yossi; Dokos, Socrates; Lovell, Nigel H; Morley, John W

2017-05-01

Electrical stimulation of neuronal tissue is a promising strategy to treat a variety of neurological disorders. The mechanism of neuronal activation by external electrical stimulation is governed by voltage-gated ion channels. This stimulus, typically brief in nature, leads to membrane potential depolarization, which increases ion flow across the membrane by increasing the open probability of these voltage-gated channels. In spiking neurons, it is activation of voltage-gated sodium channels (Na V channels) that leads to action potential generation. However, several other types of voltage-gated channels are expressed that also respond to electrical stimulation. In this study, we examine the response of voltage-gated potassium channels (K V channels) to brief electrical stimulation by whole cell patch-clamp electrophysiology and computational modeling. We show that nonspiking amacrine neurons of the retina exhibit a large variety of responses to stimulation, driven by different K V -channel subtypes. Computational modeling reveals substantial differences in the response of specific K V -channel subtypes that is dependent on channel kinetics. This suggests that the expression levels of different K V -channel subtypes in retinal neurons are a crucial predictor of the response that can be obtained. These data expand our knowledge of the mechanisms of neuronal activation and suggest that K V -channel expression is an important determinant of the sensitivity of neurons to electrical stimulation. NEW & NOTEWORTHY This paper describes the response of various voltage-gated potassium channels (K V channels) to brief electrical stimulation, such as is applied during prosthetic electrical stimulation. We show that the pattern of response greatly varies between K V channel subtypes depending on activation and inactivation kinetics of each channel. Our data suggest that problems encountered when artificially stimulating neurons such as cessation in firing at high frequencies, or "fading," may be attributed to K V -channel activation. Copyright © 2017 the American Physiological Society.

Differential regulation of ROMK (Kir1.1) in distal nephron segments by dietary potassium.

PubMed

Wade, James B; Fang, Liang; Coleman, Richard A; Liu, Jie; Grimm, P Richard; Wang, Tong; Welling, Paul A

2011-06-01

ROMK channels are well-known to play a central role in renal K secretion, but the absence of highly specific and avid-ROMK antibodies has presented significant roadblocks toward mapping the extent of expression along the entire distal nephron and determining whether surface density of these channels is regulated in response to physiological stimuli. Here, we prepared new ROMK antibodies verified to be highly specific, using ROMK knockout mice as a control. Characterization with segmental markers revealed a more extensive pattern of ROMK expression along the entire distal nephron than previously thought, localizing to distal convoluted tubule regions, DCT1 and DCT2; the connecting tubule (CNT); and cortical collecting duct (CD). ROMK was diffusely distributed in intracellular compartments and at the apical membrane of each tubular region. Apical labeling was significantly increased by high-K diet in DCT2, CNT1, CNT2, and CD (P < 0.05) but not in DCT1. Consistent with the large increase in apical ROMK, dramatically increased mature glycosylation was observed following dietary potassium augmentation. We conclude 1) our new antibody provides a unique tool to characterize ROMK channel localization and expression and 2) high-K diet causes a large increase in apical expression of ROMK in DCT2, CNT, and CD but not in DCT1, indicating that different regulatory mechanisms are involved in K diet-regulated ROMK channel functions in the distal nephron.

Electric organ discharge diversification in mormyrid weakly electric fish is associated with differential expression of voltage-gated ion channel genes.

PubMed

Nagel, Rebecca; Kirschbaum, Frank; Tiedemann, Ralph

2017-03-01

In mormyrid weakly electric fish, the electric organ discharge (EOD) is used for species recognition, orientation and prey localization. Produced in the muscle-derived adult electric organ, the EOD exhibits a wide diversity across species in both waveform and duration. While certain defining EOD characteristics can be linked to anatomical features of the electric organ, many factors underlying EOD differentiation are yet unknown. Here, we report the differential expression of 13 Kv1 voltage-gated potassium channel genes, two inwardly rectifying potassium channel genes, two previously studied sodium channel genes and an ATPase pump in two sympatric species of the genus Campylomormyrus in both the adult electric organ and skeletal muscle. Campylomormyrus compressirostris displays a basal EOD, largely unchanged during development, while C. tshokwe has an elongated, putatively derived discharge. We report an upregulation in all Kv1 genes in the electric organ of Campylomormyrus tshokwe when compared to both skeletal muscle and C. compressirostris electric organ. This pattern of upregulation in a species with a derived EOD form suggests that voltage-gated potassium channels are potentially involved in the diversification of the EOD signal among mormyrid weakly electric fish.

Differential distribution of the sodium-activated potassium channels slick and slack in mouse brain.

PubMed

Rizzi, Sandra; Knaus, Hans-Günther; Schwarzer, Christoph

2016-07-01

The sodium-activated potassium channels Slick (Slo2.1, KCNT2) and Slack (Slo2.2, KCNT1) are high-conductance potassium channels of the Slo family. In neurons, Slick and Slack channels are involved in the generation of slow afterhyperpolarization, in the regulation of firing patterns, and in setting and stabilizing the resting membrane potential. The distribution and subcellular localization of Slick and Slack channels in the mouse brain have not yet been established in detail. The present study addresses this issue through in situ hybridization and immunohistochemistry. Both channels were widely distributed and exhibited distinct distribution patterns. However, in some brain regions, their expression overlapped. Intense Slick channel immunoreactivity was observed in processes, varicosities, and neuronal cell bodies of the olfactory bulb, granular zones of cortical regions, hippocampus, amygdala, lateral septal nuclei, certain hypothalamic and midbrain nuclei, and several regions of the brainstem. The Slack channel showed primarily a diffuse immunostaining pattern, and labeling of cell somata and processes was observed only occasionally. The highest Slack channel expression was detected in the olfactory bulb, lateral septal nuclei, basal ganglia, and distinct areas of the midbrain, brainstem, and cerebellar cortex. In addition, comparing our data obtained from mouse brain with a previously published study on rat brain revealed some differences in the expression and distribution of Slick and Slack channels in these species. J. Comp. Neurol. 524:2093-2116, 2016. © 2015 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc. © 2015 The Authors The Journal of Comparative Neurology Published by Wiley Periodicals, Inc.

Evolving regulatory paradigm for proarrhythmic risk assessment for new drugs.

PubMed

Vicente, Jose; Stockbridge, Norman; Strauss, David G

Fourteen drugs were removed from the market worldwide because their potential to cause torsade de pointes (torsade), a potentially fatal ventricular arrhythmia. The observation that most drugs that cause torsade block the potassium channel encoded by the human ether-à-go-go related gene (hERG) and prolong the heart rate corrected QT interval (QTc) on the ECG, led to a focus on screening new drugs for their potential to block the hERG potassium channel and prolong QTc. This has been a successful strategy keeping torsadogenic drugs off the market, but has resulted in drugs being dropped from development, sometimes inappropriately. This is because not all drugs that block the hERG potassium channel and prolong QTc cause torsade, sometimes because they block other channels. The regulatory paradigm is evolving to improve proarrhythmic risk prediction. ECG studies can now use exposure-response modeling for assessing the effect of a drug on the QTc in small sample size first-in-human studies. Furthermore, the Comprehensive in vitro Proarrhythmia Assay (CiPA) initiative is developing and validating a new in vitro paradigm for cardiac safety evaluation of new drugs that provides a more accurate and comprehensive mechanistic-based assessment of proarrhythmic potential. Under CiPA, the prediction of proarrhythmic potential will come from in vitro ion channel assessments coupled with an in silico model of the human ventricular myocyte. The preclinical assessment will be checked with an assessment of human phase 1 ECG data to determine if there are unexpected ion channel effects in humans compared to preclinical ion channel data. While there is ongoing validation work, the heart rate corrected J-T peak interval is likely to be assessed under CiPA to detect inward current block in presence of hERG potassium channel block. Copyright © 2016 Elsevier Inc. All rights reserved.

A bursting potassium channel in isolated cholinergic synaptosomes of Torpedo electric organ.

PubMed Central

Edry-Schiller, J; Ginsburg, S; Rahamimoff, R

1991-01-01

1. Pinched-off cholinergic nerve terminals (synaptosomes) prepared from the electric organ of Torpedo ocelata were fused into large structures (greater than 20 microns) using dimethyl sulphoxide and polyethylene glycol 1500, as previously described for synaptic vesicles from the same organ. 2. The giant fused synaptosomes were easily amenable to the patch clamp technique and 293 seals with a resistance greater than 4 G omega were obtained in the 'cell-attached' configuration. In a large fraction of the experiments, an 'inside-out' patch configuration was achieved. 3. Several types of unitary ionic currents were observed. This study describes the most frequently observed single-channel activity which was found in 247 out of the 293 membrane patches (84.3%). 4. The single-channel current-voltage relation was linear between -60 and 20 mV and showed a slope conductance of 23.8 +/- 1.3 pS when the pipette contained 350-390 mM-Na+ and the bath facing the inside of the synaptosomal membrane contained 390 mM-K+. 5. From extrapolated reversal potential measurements, it was concluded that this channel has a large selectivity for K+ over Na+ (70.4 +/- 11.5, mean +/- S.E.M.). Chloride ions are not transported significantly through this potassium channel. 6. This potassium channel has a low probability of opening. The probability of being in the open state increases upon depolarization and reaches about 1% when the inside of the patch is 20 mV positive compared to the pipette side. 7. The mean channel open time increases with depolarization; thus the product current x time (= charge) also increases upon depolarization, showing properties of an outward rectifier. 8. The potassium channel in the giant synaptosome membrane has a bursting behaviour. Open-time distribution, closed-time distribution and a Poisson analysis indicate that the minimal kinetic scheme requires one open state and three closed states. PMID:1654418

Mechanosensitive Piezo Channels in the Gastrointestinal Tract.

PubMed

Alcaino, C; Farrugia, G; Beyder, A

2017-01-01

Sensation of mechanical forces is critical for normal function of the gastrointestinal (GI) tract and abnormalities in mechanosensation are linked to GI pathologies. In the GI tract there are several mechanosensitive cell types-epithelial enterochromaffin cells, intrinsic and extrinsic enteric neurons, smooth muscle cells and interstitial cells of Cajal. These cells use mechanosensitive ion channels that respond to mechanical forces by altering transmembrane ionic currents in a process called mechanoelectrical coupling. Several mechanosensitive ionic conductances have been identified in the mechanosensory GI cells, ranging from mechanosensitive voltage-gated sodium and calcium channels to the mechanogated ion channels, such as the two-pore domain potassium channels K2P (TREK-1) and nonselective cation channels from the transient receptor potential family. The recently discovered Piezo channels are increasingly recognized as significant contributors to cellular mechanosensitivity. Piezo1 and Piezo2 are nonselective cationic ion channels that are directly activated by mechanical forces and have well-defined biophysical and pharmacologic properties. The role of Piezo channels in the GI epithelium is currently under investigation and their role in the smooth muscle syncytium and enteric neurons is still not known. In this review, we outline the current state of knowledge on mechanosensitive ion channels in the GI tract, with a focus on the known and potential functions of the Piezo channels. Copyright © 2017 Elsevier Inc. All rights reserved.

Functional and molecular identification of a TASK-1 potassium channel regulating chloride secretion through CFTR channels in the shark rectal gland: implications for cystic fibrosis

PubMed Central

Telles, Connor J.; Decker, Sarah E.; Motley, William W.; Peters, Alexander W.; Mehr, Ali Poyan; Frizzell, Raymond A.

2016-01-01

In the shark rectal gland (SRG), apical chloride secretion through CFTR channels is electrically coupled to a basolateral K+ conductance whose type and molecular identity are unknown. We performed studies in the perfused SRG with 17 K+ channel inhibitors to begin this search. Maximal chloride secretion was markedly inhibited by low-perfusate pH, bupivicaine, anandamide, zinc, quinidine, and quinine, consistent with the properties of an acid-sensitive, four-transmembrane, two-pore-domain K+ channel (4TM-K2P). Using PCR with degenerate primers to this family, we identified a TASK-1 fragment in shark rectal gland, brain, gill, and kidney. Using 5′ and 3′ rapid amplification of cDNA ends PCR and genomic walking, we cloned the full-length shark gene (1,282 bp), whose open reading frame encodes a protein of 375 amino acids that was 80% identical to the human TASK-1 protein. We expressed shark and human TASK-1 cRNA in Xenopus oocytes and characterized these channels using two-electrode voltage clamping. Both channels had identical current-voltage relationships (outward rectifying) and a reversal potential of −90 mV. Both were inhibited by quinine, bupivicaine, and acidic pH. The pKa for current inhibition was 7.75 for shark TASK-1 vs. 7.37 for human TASK-1, values similar to the arterial pH for each species. We identified this protein in SRG by Western blot and confocal immunofluorescent microscopy and detected the protein in SRG and human airway cells. Shark TASK-1 is the major K+ channel coupled to chloride secretion in the SRG, is the oldest 4TM 2P family member identified, and is the first TASK-1 channel identified to play a role in setting the driving force for chloride secretion in epithelia. The detection of this potassium channel in mammalian lung tissue has implications for human biology and disease. PMID:27653983

KATP channel inhibition blunts electromechanical decline during hypoxia in left ventricular working rabbit hearts

PubMed Central

Garrott, Kara; Kuzmiak‐Glancy, Sarah; Wengrowski, Anastasia; Zhang, Hanyu; Rogers, Jack

2017-01-01

Key points Heart function is critically dependent upon the balance of energy production and utilization. Sarcolemmal ATP‐sensitive potassium channels (KATP channels) in cardiac myocytes adjust contractile function to compensate for the level of available energy.Understanding the activation of KATP channels in working myocardium during high‐stress situations is crucial to the treatment of cardiovascular disease, especially ischaemic heart disease.Using a new optical mapping approach, we measured action potentials from the surface of excised contracting rabbit hearts to assess when sarcolemmal KATP channels were activated during physiologically relevant workloads and during gradual reductions in myocardial oxygenation.We demonstrate that left ventricular pressure is closely linked to KATP channel activation and that KATP channel inhibition with a low concentration of tolbutamide prevents electromechanical decline when oxygen availability is reduced. As a result, KATP channel inhibition probably exacerbates a mismatch between energy demand and energy production when myocardial oxygenation is low. Abstract Sarcolemmal ATP‐sensitive potassium channel (KATP channel) activation in isolated cells is generally understood, although the relationship between myocardial oxygenation and KATP activation in excised working rabbit hearts remains unknown. We optically mapped action potentials (APs) in excised rabbit hearts to test the hypothesis that hypoxic changes would be more severe in left ventricular (LV) working hearts (LWHs) than Langendorff (LANG) perfused hearts. We further hypothesized that KATP inhibition would prevent those changes. Optical APs were mapped when measuring LV developed pressure (LVDP), coronary flow rate and oxygen consumption in LANG and LWHs. Hearts were paced to increase workload and perfusate was deoxygenated to study the effects of myocardial hypoxia. A subset of hearts was perfused with 1 μm tolbutamide (TOLB) to identify the level of AP duration (APD) shortening attributed to KATP channel activation. During sinus rhythm, APD was shorter in LWHs compared to LANG hearts. APD in both LWHs and LANG hearts dropped steadily during deoxygenation. With TOLB, APDs in LWHs were longer at all workloads and APD reductions during deoxygenation were blunted in both LWHs and LANG hearts. At 50% perfusate oxygenation, APD and LVDP were significantly higher in LWHs perfused with TOLB (199 ± 16 ms; 92 ± 5.3 mmHg) than in LWHs without TOLB (109 ± 14 ms, P = 0.005; 65 ± 6.5 mmHg, P = 0.01). Our results indicate that KATP channels are activated to a greater extent in perfused hearts when the LV performs pressure–volume work. The results of the present study demonstrate the critical role of KATP channels in modulating myocardial function over a wide range of physiological conditions. PMID:28177123

The N-terminal domain of Slack determines the formation and trafficking of Slick/Slack heteromeric sodium-activated potassium channels.

PubMed

Chen, Haijun; Kronengold, Jack; Yan, Yangyang; Gazula, Valeswara-Rao; Brown, Maile R; Ma, Liqun; Ferreira, Gonzalo; Yang, Youshan; Bhattacharjee, Arin; Sigworth, Fred J; Salkoff, Larry; Kaczmarek, Leonard K

2009-04-29

Potassium channels activated by intracellular Na(+) ions (K(Na)) play several distinct roles in regulating the firing patterns of neurons, and, at the single channel level, their properties are quite diverse. Two known genes, Slick and Slack, encode K(Na) channels. We have now found that Slick and Slack subunits coassemble to form heteromeric channels that differ from the homomers in their unitary conductance, kinetic behavior, subcellular localization, and response to activation of protein kinase C. Heteromer formation requires the N-terminal domain of Slack-B, one of the alternative splice variants of the Slack channel. This cytoplasmic N-terminal domain of Slack-B also facilitates the localization of heteromeric K(Na) channels to the plasma membrane. Immunocytochemical studies indicate that Slick and Slack-B subunits are coexpressed in many central neurons. Our findings provide a molecular explanation for some of the diversity in reported properties of neuronal K(Na) channels.

Modulatory mechanisms and multiple functions of somatodendritic A-type K+ channel auxiliary subunits

PubMed Central

Jerng, Henry H.; Pfaffinger, Paul J.

2014-01-01

Auxiliary subunits are non-conducting, modulatory components of the multi-protein ion channel complexes that underlie normal neuronal signaling. They interact with the pore-forming α-subunits to modulate surface distribution, ion conductance, and channel gating properties. For the somatodendritic subthreshold A-type potassium (ISA) channel based on Kv4 α-subunits, two types of auxiliary subunits have been extensively studied: Kv channel-interacting proteins (KChIPs) and dipeptidyl peptidase-like proteins (DPLPs). KChIPs are cytoplasmic calcium-binding proteins that interact with intracellular portions of the Kv4 subunits, whereas DPLPs are type II transmembrane proteins that associate with the Kv4 channel core. Both KChIPs and DPLPs genes contain multiple start sites that are used by various neuronal populations to drive the differential expression of functionally distinct N-terminal variants. In turn, these N-terminal variants generate tremendous functional diversity across the nervous system. Here, we focus our review on (1) the molecular mechanism underlying the unique properties of different N-terminal variants, (2) the shaping of native ISA properties by the concerted actions of KChIPs and DPLP variants, and (3) the surprising ways that KChIPs and DPLPs coordinate the activity of multiple channels to fine-tune neuronal excitability. Unlocking the unique contributions of different auxiliary subunit N-terminal variants may provide an important opportunity to develop novel targeted therapeutics to treat numerous neurological disorders. PMID:24723849

Modulatory mechanisms and multiple functions of somatodendritic A-type K (+) channel auxiliary subunits.

PubMed

Jerng, Henry H; Pfaffinger, Paul J

2014-01-01

Auxiliary subunits are non-conducting, modulatory components of the multi-protein ion channel complexes that underlie normal neuronal signaling. They interact with the pore-forming α-subunits to modulate surface distribution, ion conductance, and channel gating properties. For the somatodendritic subthreshold A-type potassium (ISA) channel based on Kv4 α-subunits, two types of auxiliary subunits have been extensively studied: Kv channel-interacting proteins (KChIPs) and dipeptidyl peptidase-like proteins (DPLPs). KChIPs are cytoplasmic calcium-binding proteins that interact with intracellular portions of the Kv4 subunits, whereas DPLPs are type II transmembrane proteins that associate with the Kv4 channel core. Both KChIPs and DPLPs genes contain multiple start sites that are used by various neuronal populations to drive the differential expression of functionally distinct N-terminal variants. In turn, these N-terminal variants generate tremendous functional diversity across the nervous system. Here, we focus our review on (1) the molecular mechanism underlying the unique properties of different N-terminal variants, (2) the shaping of native ISA properties by the concerted actions of KChIPs and DPLP variants, and (3) the surprising ways that KChIPs and DPLPs coordinate the activity of multiple channels to fine-tune neuronal excitability. Unlocking the unique contributions of different auxiliary subunit N-terminal variants may provide an important opportunity to develop novel targeted therapeutics to treat numerous neurological disorders.

Origin of heterogeneous spiking patterns from continuously distributed ion channel densities: a computational study in spinal dorsal horn neurons.

PubMed

Balachandar, Arjun; Prescott, Steven A

2018-05-01

Distinct spiking patterns may arise from qualitative differences in ion channel expression (i.e. when different neurons express distinct ion channels) and/or when quantitative differences in expression levels qualitatively alter the spike generation process. We hypothesized that spiking patterns in neurons of the superficial dorsal horn (SDH) of spinal cord reflect both mechanisms. We reproduced SDH neuron spiking patterns by varying densities of K V 1- and A-type potassium conductances. Plotting the spiking patterns that emerge from different density combinations revealed spiking-pattern regions separated by boundaries (bifurcations). This map suggests that certain spiking pattern combinations occur when the distribution of potassium channel densities straddle boundaries, whereas other spiking patterns reflect distinct patterns of ion channel expression. The former mechanism may explain why certain spiking patterns co-occur in genetically identified neuron types. We also present algorithms to predict spiking pattern proportions from ion channel density distributions, and vice versa. Neurons are often classified by spiking pattern. Yet, some neurons exhibit distinct patterns under subtly different test conditions, which suggests that they operate near an abrupt transition, or bifurcation. A set of such neurons may exhibit heterogeneous spiking patterns not because of qualitative differences in which ion channels they express, but rather because quantitative differences in expression levels cause neurons to operate on opposite sides of a bifurcation. Neurons in the spinal dorsal horn, for example, respond to somatic current injection with patterns that include tonic, single, gap, delayed and reluctant spiking. It is unclear whether these patterns reflect five cell populations (defined by distinct ion channel expression patterns), heterogeneity within a single population, or some combination thereof. We reproduced all five spiking patterns in a computational model by varying the densities of a low-threshold (K V 1-type) potassium conductance and an inactivating (A-type) potassium conductance and found that single, gap, delayed and reluctant spiking arise when the joint probability distribution of those channel densities spans two intersecting bifurcations that divide the parameter space into quadrants, each associated with a different spiking pattern. Tonic spiking likely arises from a separate distribution of potassium channel densities. These results argue in favour of two cell populations, one characterized by tonic spiking and the other by heterogeneous spiking patterns. We present algorithms to predict spiking pattern proportions based on ion channel density distributions and, conversely, to estimate ion channel density distributions based on spiking pattern proportions. The implications for classifying cells based on spiking pattern are discussed. © 2018 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.

Two inwardly rectifying potassium channels, Irk1 and Irk2, play redundant roles in Drosophila renal tubule function

PubMed Central

Wu, Yipin; Baum, Michel; Huang, Chou-Long

2015-01-01

Inwardly rectifying potassium channels play essential roles in renal physiology across phyla. Barium-sensitive K+ conductances are found on the basolateral membrane of a variety of insect Malpighian (renal) tubules, including Drosophila melanogaster. We found that barium decreases the lumen-positive transepithelial potential difference in isolated perfused Drosophila tubules and decreases fluid secretion and transepithelial K+ flux. In those insect species in which it has been studied, transcripts from multiple genes encoding inwardly rectifying K+ channels are expressed in the renal (Malpighian) tubule. In Drosophila melanogaster, this includes transcripts of the Irk1, Irk2, and Irk3 genes. The role of each of these gene products in renal tubule function is unknown. We found that simultaneous knockdown of Irk1 and Irk2 in the principal cell of the fly tubule decreases transepithelial K+ flux, with no additive effect of Irk3 knockdown, and decreases barium sensitivity of transepithelial K+ flux by ∼50%. Knockdown of any of the three inwardly rectifying K+ channels individually has no effect, nor does knocking down Irk3 simultaneously with Irk1 or Irk2. Irk1/Irk2 principal cell double-knockdown tubules remain sensitive to the kaliuretic effect of cAMP. Inhibition of the Na+/K+-ATPase with ouabain and Irk1/Irk2 double knockdown have additive effects on K+ flux, and 75% of transepithelial K+ transport is due to Irk1/Irk2 or ouabain-sensitive pathways. In conclusion, Irk1 and Irk2 play redundant roles in transepithelial ion transport in the Drosophila melanogaster renal tubule and are additive to Na+/K+-ATPase-dependent pathways. PMID:26224687

The effect of hypercholesterolemia on carbachol-induced contractions of the detrusor smooth muscle in rats: increased role of L-type Ca2+ channels.

PubMed

Balkanci, Zeynep Dicle; Pehlivanoğlu, Bilge; Bayrak, Sibel; Karabulut, Ismail; Karaismailoğlu, Serkan; Erdem, Ayşen

2012-11-01

To investigate a possible relation between hypercholesterolemia and detrusor smooth muscle function, we studied the contractile response to potassium challenge, carbachol (CCh), and the components of CCh-induced contractile mechanism in high-cholesterol diet-fed rats. Adult male Sprague-Dawley rats were fed with standard (control group, N = 17) or 4 % cholesterol diet (hypercholesterolemia group (HC), N = 16) for 4 weeks. Spontaneous contractions of detrusor muscle strips and their responses to potassium chloride (KCl) or cumulative dose-contraction curves to CCh were recorded. The effects of muscarinic receptor antagonists (methoctramin and/or 4-diphenylacetoxy-N-methylpiperidine), L-type Ca(+2) channel blocker (nifedipine), and/or rho-kinase inhibitor Y-27632 were investigated. Blood cholesterol level was increased in the HC group with no sign of atherosclerosis. The KCl-induced detrusor smooth muscle contractions were higher in HC, whereas spontaneous and CCh-induced responses were similar in both groups. Preincubation with receptor antagonist for M(3) but not for M(2) attenuated contraction significantly, shifting the dose-response curve to the right. This response was similar in both groups. Among two effector mechanisms of M(3)-mediated detrusor smooth muscle contraction, rho-kinase pathway was not affected by hypercholesterolemia, whereas blockade of L-type Ca(+2) channels potently reduced contractions. The results of this study point out a relation between hypercholesterolemia and contractile mechanism of detrusor smooth muscle likely to change urinary bladder function, via altering L-type Ca(+2) channels. Taken together with escalating incidence of hypercholesterolemia and lower urinary tract symptoms, it is a field which deserves to be investigated further.

Palmitoylation of the β4-Subunit Regulates Surface Expression of Large Conductance Calcium-activated Potassium Channel Splice Variants*

PubMed Central

Chen, Lie; Bi, Danlei; Tian, Lijun; McClafferty, Heather; Steeb, Franziska; Ruth, Peter; Knaus, Hans Guenther; Shipston, Michael J.

2013-01-01

Regulatory β-subunits of large conductance calcium- and voltage-activated potassium (BK) channels play an important role in generating functional diversity and control of cell surface expression of the pore forming α-subunits. However, in contrast to α-subunits, the role of reversible post-translational modification of intracellular residues on β-subunit function is largely unknown. Here we demonstrate that the human β4-subunit is S-acylated (palmitoylated) on a juxtamembrane cysteine residue (Cys-193) in the intracellular C terminus of the regulatory β-subunit. β4-Subunit palmitoylation is important for cell surface expression and endoplasmic reticulum (ER) exit of the β4-subunit alone. Importantly, palmitoylated β4-subunits promote the ER exit and surface expression of the pore-forming α-subunit, whereas β4-subunits that cannot be palmitoylated do not increase ER exit or surface expression of α-subunits. Strikingly, however, this palmitoylation- and β4-dependent enhancement of α-subunit surface expression was only observed in α-subunits that contain a putative trafficking motif (… REVEDEC) at the very C terminus of the α-subunit. Engineering this trafficking motif to other C-terminal α-subunit splice variants results in α-subunits with reduced surface expression that can be rescued by palmitoylated, but not depalmitoylated, β4-subunits. Our data reveal a novel mechanism by which palmitoylated β4-subunit controls surface expression of BK channels through masking of a trafficking motif in the C terminus of the α-subunit. As palmitoylation is dynamic, this mechanism would allow precise control of specific splice variants to the cell surface. Our data provide new insights into how complex interplay between the repertoire of post-transcriptional and post-translational mechanisms controls cell surface expression of BK channels. PMID:23504458

Voltage-Gated Potassium Channels Kv1.3--Potentially New Molecular Target in Cancer Diagnostics and Therapy.

PubMed

Teisseyre, Andrzej; Gąsiorowska, Justyna; Michalak, Krystyna

2015-01-01

Voltage-gated potassium channels, Kv1.3, which were discovered in 1984, are integral membrane proteins which are activated ("open") upon change of the cell membrane potential, enabling a passive flux of potassium ions across the cell membrane. The channels are expressed in many different tissues, both normal and cancer. Since 2005 it has been known that the channels are expressed not only in the plasma membrane, but also in the inner mitochondrial membrane. The activity of Kv1.3 channels plays an important role, among others, in setting the cell resting membrane potential, cell proliferation, apoptosis and volume regulation. For some years, these channels have been considered a potentially new molecular target in both the diagnostics and therapy of some cancer diseases. This review article focuses on: 1) changes of expression of the channels in cancer disorders with special regard to correlations between the channels' expression and stage of the disease, 2) influence of inhibitors of Kv1.3 channels on proliferation and apoptosis of cancer cells, 3) possible future applications of Kv1.3 channels' inhibitors in therapy of some cancer diseases. In the last section, the results of studies performed in our Laboratory of Bioelectricity on the influence of selected biologically active plant-derived compounds from the groups of flavonoids and stilbenes and their natural and synthetic derivatives on the activity of Kv1.3 channels in normal and cancer cells are reviewed. A possible application of some compounds from these groups to support therapy of cancer diseases, such as breast, colon and lymph node cancer, and melanoma or chronic lymphocytic leukemia (B-CLL), is announced.

Leucine-Rich Glioma Inactivated-1 and Voltage-Gated Potassium Channel Autoimmune Encephalitis Associated with Ischemic Stroke: A Case Report

PubMed Central

McGinley, Marisa; Morales-Vidal, Sarkis; Ruland, Sean

2016-01-01

Autoimmune encephalitis is associated with a wide variety of antibodies and clinical presentations. Voltage-gated potassium channel (VGKC) antibodies are a cause of autoimmune non-paraneoplastic encephalitis characterized by memory impairment, psychiatric symptoms, and seizures. We present a case of VGKC encephalitis likely preceding an ischemic stroke. Reports of autoimmune encephalitis associated with ischemic stroke are rare. Several hypotheses linking these two disease processes are proposed. PMID:27242653

The pore-lining region of shaker voltage-gated potassium channels: comparison of beta-barrel and alpha-helix bundle models.

PubMed Central

Kerr, I D; Sansom, M S

1997-01-01

Although there is a large body of site-directed mutagenesis data that identify the pore-lining sequence of the voltage-gated potassium channel, the structure of this region remains unknown. We have interpreted the available biochemical data as a set of topological and orientational restraints and employed these restraints to produce molecular models of the potassium channel pore region, H5. The H5 sequence has been modeled either as a tetramer of membrane-spanning beta-hairpins, thus producing an eight-stranded beta-barrel, or as a tetramer of incompletely membrane-spanning alpha-helical hairpins, thus producing an eight-staved alpha-helix bundle. In total, restraints-directed modeling has produced 40 different configurations of the beta-barrel model, each configuration comprising an ensemble of 20 structures, and 24 different configurations of the alpha-helix bundle model, each comprising an ensemble of 24 structures. Thus, over 1300 model structures for H5 have been generated. Configurations have been ranked on the basis of their predicted pore properties and on the extent of their agreement with the biochemical data. This ranking is employed to identify particular configurations of H5 that may be explored further as models of the pore-lining region of the voltage-gated potassium channel pore. Images FIGURE 7 FIGURE 12 PMID:9251779

Fatty acids and related Kv2 channel blockers: electrophysiology and toxicity on mosquitoes

USDA-ARS?s Scientific Manuscript database

Ligand-gated ion channels form an important superfamily of proteins involved in many biological processes. Among them, the potassium channels constitute a very diverse group involved in neural signaling, neuronal activity and action potential. Among the different types of channel activation, voltage...

Vascular ATP-sensitive potassium channels are over-expressed and partially regulated by nitric oxide in experimental septic shock.

PubMed

Collin, Solène; Sennoun, Nacira; Dron, Anne-Gaëlle; de la Bourdonnaye, Mathilde; Montemont, Chantal; Asfar, Pierre; Lacolley, Patrick; Meziani, Ferhat; Levy, Bruno

2011-05-01

To study the activation and expression of vascular (aorta and small mesenteric arteries) potassium channels during septic shock with or without modulation of the NO pathway. Septic shock was induced in rats by peritonitis. Selective inhibitors of vascular K(ATP) (PNU-37883A) or BK(Ca) [iberiotoxin (IbTX)] channels were used to demonstrate their involvement in vascular hyporeactivity. Vascular response to phenylephrine was measured on aorta and small mesenteric arteries mounted on a wire myograph. Vascular expression of potassium channels was studied by PCR and Western blot, in the presence or absence of 1400W, an inducible NO synthase (iNOS) inhibitor. Aortic activation of the transcriptional factor nuclear factor-kappaB (NF-κB) was assessed by electrophoretic mobility shift assay. Arterial pressure as well as in vivo and ex vivo vascular reactivity were reduced by sepsis and improved by PNU-37883A but not by IbTX. Sepsis was associated with an up-regulation of mRNA and protein expression of vascular K(ATP) channels, while expression of vascular BK(Ca) channels remained unchanged. Selective iNOS inhibition blunted the sepsis-induced increase in aortic NO, decreased NF-κB activation, and down-regulated vascular K(ATP) channel expression. Vascular K(ATP) but not BK(Ca) channels are activated, over-expressed, and partially regulated by NO via NF-κB activation during septic shock. Their selective inhibition restores arterial pressure and vascular reactivity and decreases lactate concentration. The present data suggest that selective vascular K(ATP) channel inhibitors offer potential therapeutic perspectives for septic shock.

A Kv7.2 mutation associated with early onset epileptic encephalopathy with suppression-burst enhances Kv7/M channel activity.

PubMed

Devaux, Jérôme; Abidi, Affef; Roubertie, Agathe; Molinari, Florence; Becq, Hélène; Lacoste, Caroline; Villard, Laurent; Milh, Mathieu; Aniksztejn, Laurent

2016-05-01

Mutations in the KCNQ2 gene encoding the voltage-gated potassium channel subunit Kv7.2 cause early onset epileptic encephalopathy (EOEE). Most mutations have been shown to induce a loss of function or to affect the subcellular distribution of Kv7 channels in neurons. Herein, we investigated functional consequences and subcellular distribution of the p.V175L mutation of Kv7.2 (Kv7.2(V175L) ) found in a patient presenting EOEE. We observed that the mutation produced a 25-40 mV hyperpolarizing shift of the conductance-voltage relationship of both the homomeric Kv7.2(V175L) and heteromeric Kv7.2(V175L) /Kv7.3 channels compared to wild-type channels and a 10 mV hyperpolarizing shift of Kv7.2(V175L) /Kv7.2/Kv7.3 channels in a 1:1:2 ratio mimicking the patient situation. Mutant channels also displayed faster activation kinetics and an increased current density that was prevented by 1 μm linopirdine. The p.V175L mutation did not affect the protein expression of Kv7 channels and its localization at the axon initial segment. We conclude that p.V175L is a gain of function mutation. This confirms previous observations showing that mutations having opposite consequences on M channels can produce EOEE. These findings alert us that drugs aiming to increase Kv7 channel activity might have adverse effects in EOEE in the case of gain-of-function variants. Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.

Complex expression and localization of inactivating Kv channels in cultured hippocampal astrocytes.

PubMed

Bekar, Lane K; Loewen, Matthew E; Cao, Kun; Sun, Xianfeng; Leis, Jerome; Wang, Rui; Forsyth, George W; Walz, Wolfgang

2005-03-01

Voltage-gated potassium channels are well established as critical for setting action potential frequency, membrane potential, and neurotransmitter release in neurons. However, their role in the "nonexcitable" glial cell type is yet to be fully understood. We used whole cell current kinetics, pharmacology, immunocytochemistry, and RT-PCR to characterize A-type current in hippocampal astrocyte cultures to better understand its function. Pharmacological analysis suggests that approximately 70, 10, and <5% of total A current is associated with Kv4, Kv3, and Kv1 channels, respectively. In addition, pharmacology and kinetics provide evidence for a significant contribution of KChIP accessory proteins to astrocytic A-channel composition. Localization of the Shaw Kv3.4 channel to astrocytic processes and the Shal Kv4.3 channel to soma suggest that these channels serve a specific function. Given this complex A-type channel expression pattern, we assessed the role of A currents in membrane voltage oscillations in response to current injections. Although TEA-sensitive delayed-rectifying currents are involved in the extent of repolarization, 4-AP-sensitive A currents serve to increase the rate. As in neurons, this effect may enable astrocytes to respond rapidly to high-frequency synaptic events. Our results indicate that hippocampal astrocytes in vitro express multiple A-type Kv channel alpha-subunits with accessory, possibly Ca(2+)-sensitive, cytoplasmic subunits that appear to be specifically localized to subcellular membrane compartments. Function of these channels remains to be determined in a physiological setting. However, this study suggests that they enable astrocytes to respond rapidly with membrane voltage oscillations to high-frequency incoming signals, possibly synchronizing astrocyte function to neuronal activity.

Modulation of inward rectifier potassium channel by toosendanin, a presynaptic blocker.

PubMed

Wang, Z F; Shi, Y L

2001-07-01

The effect of toosendanin, a presynaptic blocker, on the inward rectifier potassium channel (K(Kir)) of hippocampal CA1 pyramidal neurons of rats was studied by the single-channel patch-clamp technique. The results showed that toosendanin had an inhibitory effect on K(Kir) in an excised inside-out patch of the neuron under a symmetrical 150 mM K(+) condition. By decreasing the slower open time constant and increasing the slower close time constant, toosendanin (1x10(-6)-1x10(-4) g/ml) significantly reduced the open probability of the channel in a concentration-dependent manner. Meanwhile, a dose-dependent reduction in unitary conductance of the channel was also detected after toosendanin application. These data offer an explanation for toosendanin-induced facilitation of neurotransmitter release and antibotulismic effect of the drug.

The stretch-dependent potassium channel TREK-1 and its function in murine myometrium

PubMed Central

Monaghan, Kevin; Baker, Salah A; Dwyer, Laura; Hatton, William C; Sik Park, Kyung; Sanders, Kenton M; Koh, Sang Don

2011-01-01

Smooth muscle of the uterus stays remarkably quiescent during normal pregnancy to allow sufficient time for development of the fetus. At present the mechanisms leading to uterine quiescence during pregnancy and how the suppression of activity is relieved at term are poorly understood. Myometrial excitability is governed by ion channels, and a major hypothesis regarding the regulation of contractility during pregnancy has been that expression of certain channels is regulated by hormonal influences. We have explored the expression and function of stretch-dependent K+ (SDK) channels, which are likely to be due to TREK channels, in murine myometrial tissues and myocytes using PCR, Western blots, patch clamp, intracellular microelectrode and isometric force measurements. TREK-1 is more highly expressed than TREK-2 in myometrium, and there was no detectable expression of TRAAK. Expression of TREK-1 transcripts and protein was regulated during pregnancy and delivery. SDK channels were activated in response to negative pressure applied to patches. SDK channels were insensitive to a broad-spectrum of K+ channel blockers, including tetraethylammonium and 4-aminopyridine, and insensitive to intracellular Ca2+. SDK channels were activated by stretch and arachidonic acid and inhibited by reagents that block TREK-1 channels, l-methionine and/or methioninol. Our data suggest that uterine excitability and contractility during pregnancy is regulated by the expression of SDK/TREK-1 channels. Up-regulation of these channels stabilizes membrane potential and controls contraction during pregnancy and down-regulation of these channels induces the onset of delivery. PMID:21224218

IK channel activation increases tumor growth and induces differential behavioral responses in two breast epithelial cell lines.

PubMed

Thurber, Amy E; Nelson, Michaela; Frost, Crystal L; Levin, Michael; Brackenbury, William J; Kaplan, David L

2017-06-27

Many potassium channel families are over-expressed in cancer, but their mechanistic role in disease progression is poorly understood. Potassium channels modulate membrane potential (Vmem) and thereby influence calcium ion dynamics and other voltage-sensitive signaling mechanisms, potentially acting as transcriptional regulators. This study investigated the differential response to over-expression and activation of a cancer-associated potassium channel, the intermediate conductance calcium-activated potassium channel (IK), on aggressive behaviors in mammary epithelial and breast cancer cell lines. IK was over-expressed in the highly metastatic breast cancer cell line MDA-MB-231 and the spontaneously immortalized breast epithelial cell line MCF-10A, and the effect on cancer-associated behaviors was assessed. IK over-expression increased primary tumor growth and metastasis of MDA-MB-231 in orthotopic xenografts, demonstrating for the first time in any cancer type that increased IK is sufficient to promote cancer aggression. The primary tumors had similar vascularization as determined by CD31 staining and similar histological characteristics. Interestingly, despite the increased in vivo growth and metastasis, neither IK over-expression nor activation with agonist had a significant effect on MDA-MB-231 proliferation, invasion, or migration in vitro. In contrast, IK decreased MCF-10A proliferation and invasion through Matrigel but had no effect on migration in a scratch-wound assay. We conclude that IK activity is sufficient to promote cell aggression in vivo. Our data provide novel evidence supporting IK and downstream signaling networks as potential targets for cancer therapies.

Effects of allocryptopine on outward potassium current and slow delayed rectifier potassium current in rabbit myocardium.

PubMed

Fu, Yi-Cheng; Zhang, Yu; Tian, Liu-Yang; Li, Nan; Chen, Xi; Cai, Zhong-Qi; Zhu, Chao; Li, Yang

2016-05-01

Allocryptopine (ALL) is an effective alkaloid of Corydalis decumbens (Thunb.) Pers. Papaveraceae and has proved to be anti-arrhythmic. The purpose of our study is to investigate the effects of ALL on transmural repolarizing ionic ingredients of outward potassium current (I to) and slow delayed rectifier potassium current (I Ks). The monophasic action potential (MAP) technique was used to record the MAP duration of the epicardium (Epi), myocardium (M) and endocardium (Endo) of the rabbit heart and the whole cell patch clamp was used to record I to and I Ks in cardiomyocytes of Epi, M and Endo layers that were isolated from rabbit ventricles. The effects of ALL on MAP of Epi, M and Endo layers were disequilibrium. ALL could effectively reduce the transmural dispersion of repolarization (TDR) in rabbit transmural ventricular wall. ALL decreased the current densities of I to and I Ks in a voltage and concentration dependent way and narrowed the repolarizing differences among three layers. The analysis of gating kinetics showed ALL accelerated the channel activation of I to in M layers and partly inhibit the channel openings of I to in Epi, M and Endo cells. On the other hand, ALL mainly slowed channel deactivation of I Ks channel in Epi and Endo layers without affecting its activation. Our study gives partially explanation about the mechanisms of transmural inhibition of I to and I Ks channels by ALL in rabbit myocardium. These findings provide novel perspective regarding the anti-arrhythmogenesis application of ALL in clinical settings.

Effect of potassium channel modulators in mouse forced swimming test

PubMed Central

Galeotti, Nicoletta; Ghelardini, Carla; Caldari, Bernardetta; Bartolini, Alessandro

1999-01-01

The effect of intracerebroventricular (i.c.v.) administration of different potassium channel blockers (tetraethylammonium, apamin, charybdotoxin, gliquidone), potassium channel openers (pinacidil, minoxidil, cromakalim) and aODN to mKv1.1 on immobility time was evaluated in the mouse forced swimming test, an animal model of depression. Tetraethylammonium (TEA; 5 μg per mouse i.c.v.), apamin (3 ng per mouse i.c.v.), charybdotoxin (1 μg per mouse i.c.v.) and gliquidone (6 μg per mouse i.c.v.) administered 20 min before the test produced anti-immobility comparable to that induced by the tricyclic antidepressants amitriptyline (15 mg kg−1 s.c.) and imipramine (30 mg kg−1 s.c.). By contrast pinacidil (10–20 μg per mouse i.c.v.), minoxidil (10–20 μg per mouse i.c.v.) and cromakalim (20–30 μg per mouse i.c.v.) increased immobility time when administered in the same experimental conditions. Repeated administration of an antisense oligonucleotide (aODN) to the mKv1.1 gene (1 and 3 nmol per single i.c.v. injection) produced a dose-dependent increase in immobility time of mice 72 h after the last injection. At day 7, the increasing effect produced by aODN disappeared. A degenerate mKv1.1 oligonucleotide (dODN), used as control, did not produce any effect in comparison with saline- and vector-treated mice. At the highest effective dose, potassium channels modulators and the mKv1.1 aODN did not impair motor coordination, as revealed by the rota rod test, nor did they modify spontaneous motility as revealed by the Animex apparatus. These results suggest that modulation of potassium channels plays an important role in the regulation of immobility time in the mouse forced swimming test. PMID:10323599

Potassium Supplementation Prevents Sodium Chloride Cotransporter Stimulation During Angiotensin II Hypertension.

PubMed

Veiras, Luciana C; Han, Jiyang; Ralph, Donna L; McDonough, Alicia A

2016-10-01

Angiotensin II (AngII) hypertension increases distal tubule Na-Cl cotransporter (NCC) abundance and phosphorylation (NCCp), as well as epithelial Na(+) channel abundance and activating cleavage. Acutely raising plasma [K(+)] by infusion or ingestion provokes a rapid decrease in NCCp that drives a compensatory kaliuresis. The first aim tested whether acutely raising plasma [K(+)] with a single 3-hour 2% potassium meal would lower NCCp in Sprague-Dawley rats after 14 days of AngII (400 ng/kg per minute). The potassium-rich meal neither decreased NCCp nor increased K(+) excretion. AngII-infused rats exhibited lower plasma [K(+)] versus controls (3.6±0.2 versus 4.5±0.1 mmol/L; P<0.05), suggesting that AngII-mediated epithelial Na(+) channel activation provokes K(+) depletion. The second aim tested whether doubling dietary potassium intake from 1% (A1K) to 2% (A2K) would prevent K(+) depletion during AngII infusion and, thus, prevent NCC accumulation. A2K-fed rats exhibited normal plasma [K(+)] and 2-fold higher K(+) excretion and plasma [aldosterone] versus A1K. In A1K rats, NCC, NCCpS71, and NCCpT53 abundance increased 1.5- to 3-fold versus controls (P<0.05). The rise in NCC and NCCp abundance was prevented in the A2K rats, yet blood pressure did not significantly decrease. Epithelial Na(+) channel subunit abundance and cleavage increased 1.5- to 3-fold in both A1K and A2K; ROMK (renal outer medulla K(+) channel abundance) abundance was unaffected by AngII or dietary K(+) In summary, the accumulation and phosphorylation of NCC seen during chronic AngII infusion hypertension is likely secondary to potassium deficiency driven by epithelial Na(+) channel stimulation. © 2016 American Heart Association, Inc.

[Response of potassium channels to estrogen and progesterone in the uterine smooth muscle cells of adenomyosis in vitro].

PubMed

Shi, Jinghua; Jin, Li; Leng, Jinhua; Lang, Jinghe

2015-11-01

To investigate the expression of potassium channels and the influence of estrogen and progesterone on the cultured uterine smooth muscle cells (USMC) of adenomyosis in vitro. There were 22 cases of adenomyosis hysterectomy in the adenomyosis group and 12 patients with cervical intraepithelial neoplasia III removal of the uterus in the control group. USMC were separated and cultured in vitro, incubated with different concentrations of estrogen and progesterone. We used reverse transcription-PCR to dectect the expression of large-conductance calcium- and voltage-sensitive potassium channel α subunit (BKCa α) and voltage-gated potassium channel 4.3 (Kv4.3). The mRNA expression of BKCa α and Kv4.3 in the adenomyosis group (4.43±2.05 and 4.52±1.97) were significantly higher than those in the control group (0.83±0.25 and 0.86±0.19, P<0.05). In the control group, Kv4.3 mRNA decreased after treated with 0.1 nmol/L (0.17±0.10) and 1.0 nmol/L (0.13±0.08) estrogen than before (0.55±0.29, P<0.05). In the adenomyosis group, BKCa α mRNA decreased significantly after treated with 10.0 nmol/L estrogen (0.56±0.27 versus 1.01±0.35, P<0.05). 0.1 µmol/L progesterone elevated both BKCa α mRNA (0.44±0.24 versus 0.16±0.09) and Kv4.3 mRNA (1.29±0.51 versus 0.55±0.29) in the control group (all P<0.05); however, there were no significant difference in adenomyosis group of different concentration of progestrone (P>0.05). There is an abnormal expression of potassium channels in the adenomyosis USMC, which is regulated by high concentration of estrogen and might be resistant to progesterone.

Omega-conotoxin- and nifedipine-insensitive voltage-operated calcium channels mediate K(+)-induced release of pro-thyrotropin-releasing hormone-connecting peptides Ps4 and Ps5 from perifused rat hypothalamic slices.

PubMed

Valentijn, K; Tranchand Bunel, D; Vaudry, H

1992-07-01

The rat thyrotropin-releasing hormone (TRH) precursor (prepro-TRH) contains five copies of the TRH progenitor sequence linked together by intervening sequences. Recently, we have shown that the connecting peptides prepro-TRH-(160-169) (Ps4) and prepro-TRH-(178-199) (Ps5) are released from rat hypothalamic neurones in response to elevated potassium concentrations, in a calcium-dependent manner. In the present study, the role of voltage-operated calcium channels in potassium-induced release of Ps4 and Ps5 was investigated, using a perifusion system for rat hypothalamic slices. The release of Ps4 and Ps5 stimulated by potassium (70 mM) was blocked by the inorganic ions Co2+ (2.6 mM) and Ni2+ (5 mM). In contrast, the stimulatory effect of KCl was insensitive to Cd2+ (100 microM). The dihydropyridine antagonist nifedipine (10 microM) had no effect on K(+)-evoked release of Ps4 and Ps5. Furthermore, the response to KCl was not affected by nifedipine (10 microM) in combination with diltiazem (1 microM), a benzothiazepine which increases the affinity of dihydropyridine antagonists for their receptor. The dihydropyridine agonist BAY K 8644, at concentrations as high as 1 mM, did not stimulate the basal secretion of Ps4 and Ps5. In addition, BAY K 8644 had no potentiating effect on K(+)-induced release of Ps4 and Ps5. The marine cone snail toxin omega-conotoxin, a blocker of both L- and N-type calcium channels had no effect on the release of Ps4 and Ps5 stimulated by potassium. Similarly, the omega-conopeptide SNX-111, a selective blocker of N-type calcium channels, did not inhibit the stimulatory effect of potassium. The release of Ps4 and Ps5 evoked by high K+ was insensitive to the non-selective calcium channel blocker verapamil (20 microM). Amiloride (1 microM), a putative blocker of T-type calcium channels, did not affect KCl-induced secretion of the two connecting peptides. Taken together, these results indicate that two connecting peptides derived from the pro-TRH, Ps4 and Ps5, are released by K(+)-induced depolarization through activation of voltage-sensitive calcium channels. The calcium channels appear to have a pharmacological profile different from that of L- and N-type channels. Although, their insensitivity to low Cd2+ concentrations and sensitivity to Ni2+ ions would support the involvement of T-type calcium channels, the lack of effect of amiloride suggests that they belong to a yet undefined class of calcium channels.

The Phe932Ile mutation in KCNT1 channels associated with severe epilepsy, delayed myelination and leukoencephalopathy produces a loss-of-function channel phenotype.

PubMed

Evely, Katherine M; Pryce, Kerri D; Bhattacharjee, Arin

2017-05-20

Sodium-activated potassium (K Na ) channels contribute to firing frequency adaptation and slow after hyperpolarization. The KCNT1 gene (also known as SLACK) encodes a K Na subunit that is expressed throughout the central and peripheral nervous systems. Missense mutations of the SLACK C-terminus have been reported in several patients with rare forms of early onset epilepsy and in some cases severely delayed myelination. To date, such mutations identified in patients with autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), epilepsy of infancy with migrating focal seizures (EIMFS) and Ohtahara syndrome (OS) have been reported to be gain-of-function mutations (Villa and Combi, 2016). An exome sequencing study identified a p.Phe932Ile KCNT1 mutation as the disease-causing change in a child with severe early infantile epileptic encephalopathy and abnormal myelination (Vanderver et al., 2014). We characterized an analogous mutation in the rat Slack channel and unexpectedly found this mutation to produce a loss-of-function phenotype. In an effort to restore current, we tested the known Slack channel opener loxapine. Loxapine exhibited no effect, indicating that this mutation either caused the channel to be insensitive to this established opener or proper translation and trafficking to the membrane was disrupted. Protein analysis confirmed that while total mutant protein did not differ from wild type, membrane expression of the mutant channel was substantially reduced. Although gain-of-function mutations to the Slack channel are linked to epileptic phenotypes, this is the first reported loss-of-function mutation linked to severe epilepsy and delayed myelination. Copyright © 2017 IBRO. Published by Elsevier Ltd. All rights reserved.

A Specific Two-pore Domain Potassium Channel Blocker Defines the Structure of the TASK-1 Open Pore*

PubMed Central

Streit, Anne K.; Netter, Michael F.; Kempf, Franca; Walecki, Magdalena; Rinné, Susanne; Bollepalli, Murali K.; Preisig-Müller, Regina; Renigunta, Vijay; Daut, Jürgen; Baukrowitz, Thomas; Sansom, Mark S. P.; Stansfeld, Phillip J.; Decher, Niels

2011-01-01

Two-pore domain potassium (K2P) channels play a key role in setting the membrane potential of excitable cells. Despite their role as putative targets for drugs and general anesthetics, little is known about the structure and the drug binding site of K2P channels. We describe A1899 as a potent and highly selective blocker of the K2P channel TASK-1. As A1899 acts as an open-channel blocker and binds to residues forming the wall of the central cavity, the drug was used to further our understanding of the channel pore. Using alanine mutagenesis screens, we have identified residues in both pore loops, the M2 and M4 segments, and the halothane response element to form the drug binding site of TASK-1. Our experimental data were used to validate a K2P open-pore homology model of TASK-1, providing structural insights for future rational design of drugs targeting K2P channels. PMID:21362619

Stimulation of Slack K(+) Channels Alters Mass at the Plasma Membrane by Triggering Dissociation of a Phosphatase-Regulatory Complex.

PubMed

Fleming, Matthew R; Brown, Maile R; Kronengold, Jack; Zhang, Yalan; Jenkins, David P; Barcia, Gulia; Nabbout, Rima; Bausch, Anne E; Ruth, Peter; Lukowski, Robert; Navaratnam, Dhasakumar S; Kaczmarek, Leonard K

2016-08-30

Human mutations in the cytoplasmic C-terminal domain of Slack sodium-activated potassium (KNa) channels result in childhood epilepsy with severe intellectual disability. Slack currents can be increased by pharmacological activators or by phosphorylation of a Slack C-terminal residue by protein kinase C. Using an optical biosensor assay, we find that Slack channel stimulation in neurons or transfected cells produces loss of mass near the plasma membrane. Slack mutants associated with intellectual disability fail to trigger any change in mass. The loss of mass results from the dissociation of the protein phosphatase 1 (PP1) targeting protein, Phactr-1, from the channel. Phactr1 dissociation is specific to wild-type Slack channels and is not observed when related potassium channels are stimulated. Our findings suggest that Slack channels are coupled to cytoplasmic signaling pathways and that dysregulation of this coupling may trigger the aberrant intellectual development associated with specific childhood epilepsies. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Stimulation of Slack K+ channels alters mass at the plasma membrane by triggering dissociation of a phosphatase-regulatory complex

PubMed Central

Fleming, Matthew R.; Brown, Maile R.; Kronengold, Jack; Zhang, Yalan; Jenkins, David P.; Barcia, Gulia; Nabbout, Rima; Bausch, Anne E.; Ruth, Peter; Lukowski, Robert; Navaratnam, Dhasakumar S.; Kaczmarek, Leonard K.

2016-01-01

Summary Human mutations in the cytoplasmic C-terminal domain of Slack sodium-activated potassium (KNa) channels result in childhood epilepsy with severe intellectual disability. Slack currents can be increased by pharmacological activators or by phosphorylation of a Slack C-terminal residue by protein kinase C. Using an optical biosensor assay, we find that Slack channel stimulation in neurons or transfected cells produces loss of mass near the plasma membrane. Slack mutants associated with intellectual disability fail to trigger any change in mass. The loss of mass results from the dissociation of the protein phosphatase 1 (PP1) targeting protein, Phactr-1, from the channel. Phactr1 dissociation is specific to wild-type Slack channels and is not observed when related potassium channels are stimulated. Our findings suggest that Slack channels are coupled to cytoplasmic signaling pathways, and that dysregulation of this coupling may trigger the aberrant intellectual development associated with specific childhood epilepsies. PMID:27545877

Gap-junction coupling and ATP-sensitive potassium channels in human β -cell clusters: Effects on emergent dynamics

NASA Astrophysics Data System (ADS)

Loppini, A.; Pedersen, M. G.; Braun, M.; Filippi, S.

2017-09-01

The importance of gap-junction coupling between β cells in pancreatic islets is well established in mouse. Such ultrastructural connections synchronize cellular activity, confine biological heterogeneity, and enhance insulin pulsatility. Dysfunction of coupling has been associated with diabetes and altered β -cell function. However, the role of gap junctions between human β cells is still largely unexplored. By using patch-clamp recordings of β cells from human donors, we previously estimated electrical properties of these channels by mathematical modeling of pairs of human β cells. In this work we revise our estimate by modeling triplet configurations and larger heterogeneous clusters. We find that a coupling conductance in the range 0.005 -0.020 nS/pF can reproduce experiments in almost all the simulated arrangements. We finally explore the consequence of gap-junction coupling of this magnitude between β cells with mutant variants of the ATP-sensitive potassium channels involved in some metabolic disorders and diabetic conditions, translating studies performed on rodents to the human case. Our results are finally discussed from the perspective of therapeutic strategies. In summary, modeling of more realistic clusters with more than two β cells slightly lowers our previous estimate of gap-junction conductance and gives rise to patterns that more closely resemble experimental traces.

Hsp40 Chaperones Promote Degradation of the hERG Potassium Channel*

PubMed Central

Walker, Valerie E.; Wong, Michael J. H.; Atanasiu, Roxana; Hantouche, Christine; Young, Jason C.; Shrier, Alvin

2010-01-01

Loss of function mutations in the hERG (human ether-a-go-go related gene or KCNH2) potassium channel underlie the proarrhythmic cardiac long QT syndrome type 2. Most often this is a consequence of defective trafficking of hERG mutants to the cell surface, with channel retention and degradation at the endoplasmic reticulum. Here, we identify the Hsp40 type 1 chaperones DJA1 (DNAJA1/Hdj2) and DJA2 (DNAJA2) as key modulators of hERG degradation. Overexpression of the DJAs reduces hERG trafficking efficiency, an effect eliminated by the proteasomal inhibitor lactacystin or with DJA mutants lacking their J domains essential for Hsc70/Hsp70 activation. Both DJA1 and DJA2 cause a decrease in the amount of hERG complexed with Hsc70, indicating a preferential degradation of the complex. Similar effects were observed with the E3 ubiquitin ligase CHIP. Both the DJAs and CHIP reduce hERG stability and act differentially on folding intermediates of hERG and the disease-related trafficking mutant G601S. We propose a novel role for the DJA proteins in regulating degradation and suggest that they act at a critical point in secretory pathway quality control. PMID:19940115

A novel muscarinic receptor-independent mechanism of KCNQ2/3 potassium channel blockade by Oxotremorine-M.

PubMed

Zwart, Ruud; Reed, Hannah; Clarke, Sophie; Sher, Emanuele

2016-11-15

Inhibition of KCNQ (Kv7) potassium channels by activation of muscarinic acetylcholine receptors has been well established, and the ion currents through these channels have been long known as M-currents. We found that this cross-talk can be reconstituted in Xenopus oocytes by co-transfection of human recombinant muscarinic M1 receptors and KCNQ2/3 potassium channels. Application of the muscarinic acetylcholine receptor agonist Oxotremorine-methiodide (Oxo-M) between voltage pulses to activate KCNQ2/3 channels caused inhibition of the subsequent KCNQ2/3 responses. This effect of Oxo-M was blocked by the muscarinic acetylcholine receptor antagonist atropine. We also found that KCNQ2/3 currents were inhibited when Oxo-M was applied during an ongoing KCNQ2/3 response, an effect that was not blocked by atropine, suggesting that Oxo-M inhibits KCNQ2/3 channels directly. Indeed, also in oocytes that were transfected with only KCNQ2/3 channels, but not with muscarinic M1 receptors, Oxo-M inhibited the KCNQ2/3 response. These results show that besides the usual muscarinic acetylcholine receptor-mediated inhibition, Oxo-M also inhibits KCNQ2/3 channels by a direct mechanism. We subsequently tested xanomeline, which is a chemically distinct muscarinic acetylcholine receptor agonist, and oxotremorine, which is a close analogue of Oxo-M. Both compounds inhibited KCNQ2/3 currents via activation of M1 muscarinic acetylcholine receptors but, in contrast to Oxo-M, they did not directly inhibit KCNQ2/3 channels. Xanomeline and oxotremorine do not contain a positively charged trimethylammonium moiety that is present in Oxo-M, suggesting that such a charged moiety could be a crucial component mediating this newly described direct inhibition of KCNQ2/3 channels. Copyright © 2016 Elsevier B.V. All rights reserved.

Gene expression of stretch-activated channels and mechanoelectric feedback in the heart.

PubMed

Kelly, D; Mackenzie, L; Hunter, P; Smaill, B; Saint, D A

2006-07-01

1. Mechanoelectric feedback (MEF) in the heart is the process by which mechanical forces on the myocardium can change its electrical properties. Mechanoelectric feedback has been demonstrated in many animal models, ranging from isolated cells, through isolated hearts to whole animals. In humans, MEF has been demonstrated directly in both the atria and the ventricles. It seems likely that MEF provides either the trigger or the substrate for some types of clinically important arrhythmias. 2. Mechanoelectric feedback may arise because of the presence of stretch-sensitive (or mechano-sensitive) ion channels in the cell membrane of the cardiac myocytes. Two types have been demonstrated: (i) a non-specific cation channel (stretch-activated channel (SAC); conductance of approximately 25 pS); and (ii) a potassium channel with a conductance of approximately 100 pS. The gene coding for the SAC has not yet been identified. The gene for the potassium channel is likely to be TREK, a member of the tandem pore potassium channel gene family. We have recorded stretch-sensitive potassium channels in rat isolated myocytes that have the properties of TREK channels expressed in heterologous systems. 3. It has been shown that TREK mRNA is expressed heterogeneously in the rat ventricular wall, with 17-fold more expression in endocardial compared with epicardial cells. This difference is reflected in the TREK currents recorded from endocardial and epicardial cells using whole-cell patch-clamp techniques, although the difference in current density was less pronounced (approximately threefold). Consistent with this, we show here that when the ventricle is stretched by inflation of an intraventricular balloon in a Langendorff perfused rat isolated heart, action potential shortening was more pronounced in the endocardium (30% shortening at 40 mmHg) compared with that in the epicardium (10% shortening at the same pressure). 4. Computer models of the mechanics of the (pig) heart show pronounced spatial variations in strain in the myocardium with large transmural differences (in the left ventricle in particular) and also large differences between the base and apex of the ventricle. 5. The importance of MEF and the non-homogeneous gene expression and strain distribution for arrhythmias is discussed.

Nicorandil, a Nitric Oxide Donor and ATP-Sensitive Potassium Channel Opener, Protects Against Dystrophin-Deficient Cardiomyopathy

PubMed Central

Afzal, Muhammad Z.; Reiter, Melanie; Gastonguay, Courtney; McGivern, Jered V.; Guan, Xuan; Ge, Zhi-Dong; Mack, David L.; Childers, Martin K.; Ebert, Allison D.; Strande, Jennifer L.

2016-01-01

Background Dystrophin-deficient cardiomyopathy is a growing clinical problem without targeted treatments. We investigated whether nicorandil promotes cardioprotection in human dystrophin-deficient induced pluripotent stem cell (iPSC)-derived cardiomyocytes and the muscular dystrophy mdx mouse heart. Methods and Results Dystrophin-deficient iPSC-derived cardiomyocytes had decreased levels of endothelial nitric oxide synthase and neuronal nitric oxide synthase. The dystrophin-deficient cardiomyocytes had increased cell injury and death after 2 hours of stress and recovery. This was associated with increased levels of reactive oxygen species and dissipation of the mitochondrial membrane potential. Nicorandil pretreatment was able to abolish these stress-induced changes through a mechanism that involved the nitric oxide–cyclic guanosine monophosphate pathway and mitochondrial adenosine triphosphate-sensitive potassium channels. The increased reactive oxygen species levels in the dystrophin-deficient cardiomyocytes were associated with diminished expression of select antioxidant genes and increased activity of xanthine oxidase. Furthermore, nicorandil was found to improve the restoration of cardiac function after ischemia and reperfusion in the isolated mdx mouse heart. Conclusion Nicorandil protects against stress-induced cell death in dystrophin-deficient cardiomyocytes and preserves cardiac function in the mdx mouse heart subjected to ischemia and reperfusion injury. This suggests a potential therapeutic role for nicorandil in dystrophin-deficient cardiomyopathy. PMID:26940570

Subclinical Doses of ATP-Sensitive Potassium Channel Modulators Prevent Alterations in Memory and Synaptic Plasticity Induced by Amyloid-β.

PubMed

Salgado-Puga, Karla; Rodríguez-Colorado, Javier; Prado-Alcalá, Roberto A; Peña-Ortega, Fernando

2017-01-01

In addition to coupling cell metabolism and excitability, ATP-sensitive potassium channels (KATP) are involved in neural function and plasticity. Moreover, alterations in KATP activity and expression have been observed in Alzheimer's disease (AD) and during amyloid-β (Aβ)-induced pathology. Thus, we tested whether KATP modulators can influence Aβ-induced deleterious effects on memory, hippocampal network function, and plasticity. We found that treating animals with subclinical doses (those that did not change glycemia) of a KATP blocker (Tolbutamide) or a KATP opener (Diazoxide) differentially restrained Aβ-induced memory deficit, hippocampal network activity inhibition, and long-term synaptic plasticity unbalance (i.e., inhibition of LTP and promotion of LTD). We found that the protective effect of Tolbutamide against Aβ-induced memory deficit was strong and correlated with the reestablishment of synaptic plasticity balance, whereas Diazoxide treatment produced a mild protection against Aβ-induced memory deficit, which was not related to a complete reestablishment of synaptic plasticity balance. Interestingly, treatment with both KATP modulators renders the hippocampus resistant to Aβ-induced inhibition of hippocampal network activity. These findings indicate that KATP are involved in Aβ-induced pathology and they heighten the potential role of KATP modulation as a plausible therapeutic strategy against AD.

Effects of aquaporin 4 and inward rectifier potassium channel 4.1 on medullospinal edema after methylprednisolone treatment to suppress acute spinal cord injury in rats.

PubMed

Li, Ye; Hu, Haifeng; Liu, Jingchen; Zhu, Qingsan; Gu, Rui

2018-02-01

To investigate the effects of aquaporin 4 (AQP4) and inward rectifier potassium channel 4.1 (Kir4.1) on medullospinal edema after treatment with methylprednisolone (MP) to suppress acute spinal cord injury (ASCI) in rats. Sprague Dawley rats were randomly divided into control, sham, ASCI, and MP-treated ASCI groups. After the induction of ASCI, we injected 30 mg/kg MP via the tail vein at various time points. The Tarlov scoring method was applied to evaluate neurological symptoms, and the wet-dry weights method was applied to measure the water content of the spinal cord. The motor function score of the ASCI group was significantly lower than that of the sham group, and the spinal water content was significantly increased. In addition, the levels of AQP4 and Kir4.1 were significantly increased, as was their degree of coexpression. Compared with that in the ASCI group, the motor function score and the water content were significantly increased in the MP group; in addition, the expression and coexpression of AQP4 and Kir4.1 were significantly reduced. Methylprednisolone inhibited medullospinal edema in rats with acute spinal cord injury, possibly by reducing the coexpression of aquaporin 4 and Kir4.1 in medullospinal tissues.

Properties of Single K+ and Cl− Channels in Asclepias tuberosa Protoplasts 1

PubMed Central

Schauf, Charles L.; Wilson, Kathryn J.

1987-01-01

Potassium and chloride channels were characterized in Asclepias tuberosa suspension cell derived protoplasts by patch voltage-clamp. Whole-cell currents and single channels in excised patches had linear instantaneous current-voltage relations, reversing at the Nernst potentials for K+ and Cl−, respectively. Whole cell K+ currents activated exponentially during step depolarizations, while voltage-dependent Cl− channels were activated by hyperpolarizations. Single K+ channel conductance was 40 ± 5 pS with a mean open time of 4.5 milliseconds at 100 millivolts. Potassium channels were blocked by Cs+ and tetraethylammonium, but were insensitive to 4-aminopyridine. Chloride channels had a single-channel conductance of 100 ± 17 picosiemens, mean open time of 8.8 milliseconds, and were blocked by Zn2+ and ethacrynic acid. Whole-cell Cl− currents were inhibited by abscisic acid, and were unaffected by indole-3-acetic acid and 2,4-dichlorophenoxyacetic acid. Since internal and external composition can be controlled, patch-clamped protoplasts are ideal systems for studying the role of ion channels in plant physiology and development. Images Fig. 5 PMID:16665712

Evidence for a protective role of the Gardos channel against hemolysis in murine spherocytosis.

PubMed

De Franceschi, Lucia; Rivera, Alicia; Fleming, Mark D; Honczarenko, Marek; Peters, Luanne L; Gascard, Philippe; Mohandas, Narla; Brugnara, Carlo

2005-08-15

It has been shown that mice with complete deficiency of all 4.1R protein isoforms (4.1-/-) exhibit moderate hemolytic anemia, with abnormal erythrocyte morphology (spherocytosis) and decreased membrane stability. Here, we characterized the Gardos channel function in vitro and in vivo in erythrocytes of 4.1-/- mice. Compared with wild-type, the Gardos channel of 4.1-/- erythrocytes showed an increase in Vmax (9.75 +/- 1.06 vs 6.08 +/- 0.09 mM cell x minute; P < .04) and a decrease in Km (1.01 +/- 0.06 vs 1.47 +/- 1.02 microM; P < .03), indicating an increased sensitivity to activation by intracellular calcium. In vivo function of the Gardos channel was assessed by the oral administration of clotrimazole, a well-characterized Gardos channel blocker. Clotrimazole treatment resulted in worsening of anemia and hemolysis, with decreased red cell survival and increased numbers of circulating hyperchromic spherocytes and microspherocytes. Clotrimazole induced similar changes in 4.2-/- and band 3+/- mice, indicating that these effects of the Gardos channel are shared in different models of murine spherocytosis. Thus, potassium and water loss through the Gardos channel may play an important protective role in compensating for the reduced surface-membrane area of hereditary spherocytosis (HS) erythrocytes and reducing hemolysis in erythrocytes with cytoskeletal impairments.

Effects of protein-protein interactions and ligand binding on the ion permeation in KCNQ1 potassium channel.

PubMed

Jalily Hasani, Horia; Ganesan, Aravindhan; Ahmed, Marawan; Barakat, Khaled H

2018-01-01

The voltage-gated KCNQ1 potassium ion channel interacts with the type I transmembrane protein minK (KCNE1) to generate the slow delayed rectifier (IKs) current in the heart. Mutations in these transmembrane proteins have been linked with several heart-related issues, including long QT syndromes (LQTS), congenital atrial fibrillation, and short QT syndrome. Off-target interactions of several drugs with that of KCNQ1/KCNE1 ion channel complex have been known to cause fatal cardiac irregularities. Thus, KCNQ1/KCNE1 remains an important avenue for drug-design and discovery research. In this work, we present the structural and mechanistic details of potassium ion permeation through an open KCNQ1 structural model using the combined molecular dynamics and steered molecular dynamics simulations. We discuss the processes and key residues involved in the permeation of a potassium ion through the KCNQ1 ion channel, and how the ion permeation is affected by (i) the KCNQ1-KCNE1 interactions and (ii) the binding of chromanol 293B ligand and its derivatives into the complex. The results reveal that interactions between KCNQ1 with KCNE1 causes a pore constriction in the former, which in-turn forms small energetic barriers in the ion-permeation pathway. These findings correlate with the previous experimental reports that interactions of KCNE1 dramatically slows the activation of KCNQ1. Upon ligand-binding onto the complex, the energy-barriers along ion permeation path are more pronounced, as expected, therefore, requiring higher force in our steered-MD simulations. Nevertheless, pulling the ion when a weak blocker is bound to the channel does not necessitate high force in SMD. This indicates that our SMD simulations have been able to discern between strong and week blockers and reveal their influence on potassium ion permeation. The findings presented here will have some implications in understanding the potential off-target interactions of the drugs with the KCNQ1/KCNE1 channel that lead to cardiotoxic effects.

Tetrameric subunit structure of the native brain inwardly rectifying potassium channel Kir 2.2.

PubMed

Raab-Graham, K F; Vandenberg, C A

1998-07-31

Strongly inwardly rectifying potassium channels of the Kir 2 subfamily (IRK1, IRK2, and IRK3) are involved in maintenance and modulation of cell excitability in brain and heart. Electrophysiological studies of channels expressed in heterologous systems have suggested that the pore-conducting pathway contains four subunits. However, inferences from electrophysiological studies have not been tested on native channels and do not address the possibility of nonconducting auxiliary subunits. Here, we investigate the subunit stoichiometry of endogenous inwardly rectifying potassium channel Kir 2.2 (IRK2) from rat brain. Using chemical cross-linking, immunoprecipitiation, and velocity sedimentation, we report physical evidence demonstrating the tetrameric organization of the native channel. Kir 2.2 was sequentially cross-linked to produce bands on SDS-polyacrylamide gel electrophoresis corresponding in size to monomer, dimer, trimer, and three forms of tetramer. Fully cross-linked channel was present as a single band of tetrameric size. Immunoprecipitation of biotinylated membranes revealed a single band corresponding to Kir 2.2, suggesting that the channel is composed of a single type of subunit. Hydrodynamic properties of 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonic acid-solubilized channel were used to calculate the molecular mass of the channel. Velocity sedimentation in H2O or D2O gave a sharp peak with a sedimentation coefficient of 17.3 S. Gel filtration yielded a Stokes radius of 5.92 nm. These data indicate a multisubunit protein with a molecular mass of 193 kDa, calculated to contain 3.98 subunits. Together, these results demonstrate that Kir 2.2 channels are formed by the homotetrameric association of Kir 2.2 subunits and do not contain tightly associated auxiliary subunits. These studies suggest that Kir 2.2 channels differ in structure from related heterooctomeric ATP-sensitive K channels and heterotetrameric G-protein-regulated inward rectifier K channels.

Encephalitis due to antibodies to voltage gated potassium channel (VGKC) with cerebellar involvement in a teenager

PubMed Central

Langille, Megan M.; Desai, Jay

2015-01-01

Encephalitis due to antibodies to voltage gated potassium channel (VGKC) typically presents with limbic encephalitis and medial temporal lobe involvement on neuroimaging. We describe a case of 13 year girl female with encephalitis due to antibodies to VGKC with signal changes in the cerebellar dentate nuclei bilaterally and clinical features that suggested predominant cerebellar involvement. These have never been reported previously in the literature. Our case expands the phenotypic spectrum of this rare condition. PMID:26019428

Encephalitis due to antibodies to voltage gated potassium channel (VGKC) with cerebellar involvement in a teenager.

PubMed

Langille, Megan M; Desai, Jay

2015-01-01

Encephalitis due to antibodies to voltage gated potassium channel (VGKC) typically presents with limbic encephalitis and medial temporal lobe involvement on neuroimaging. We describe a case of 13 year girl female with encephalitis due to antibodies to VGKC with signal changes in the cerebellar dentate nuclei bilaterally and clinical features that suggested predominant cerebellar involvement. These have never been reported previously in the literature. Our case expands the phenotypic spectrum of this rare condition.

Effects of Potassium Channel Blockers on the Negative Inotropic Responses Induced by Cromakalim and Pinacidil in Guinea Pig Atrium

DTIC Science & Technology

1992-01-01

RD-A2•4 875 EFFECTS OF POTASSIUM CHANNEL BLOCKERS ON THE NEGATIVE 1/1 INOTROPIC RESPONSES INDUCED BY CRONAKALIM RND PINACIDIL IN GUINEA PIG ATRIUM(U...INOTROPICTRSPONSES INDUCED BY CROMAKAUM AND PINACIDILIN GUINEA PIG ATRIUM a AUTHOR WAI-MAN LAU 7 FORMING ORG NAMES/ADDRESSES DEFENCE SCIENCE AND a...and Technology Organisaio Aot Val. Negative Inotropic Responses Victoria. Australia Induced by Cromakalim and Pinacidil in Guinea Pig Atrium Key

Slack sodium-activated potassium channel membrane expression requires p38 mitogen-activated protein kinase phosphorylation.

PubMed

Gururaj, Sushmitha; Fleites, John; Bhattacharjee, Arin

2016-04-01

p38 MAPK has long been understood as an inducible kinase under conditions of cellular stress, but there is now increasing evidence to support its role in the regulation of neuronal function. Several phosphorylation targets have been identified, an appreciable number of which are ion channels, implicating the possible involvement of p38 MAPK in neuronal excitability. The KNa channel Slack is an important protein to be studied as it is highly and ubiquitously expressed in DRG neurons and is important in the maintenance of their firing accommodation. We sought to examine if the Slack channel could be a substrate of p38 MAPK activity. First, we found that the Slack C-terminus contains two putative p38 MAPK phosphorylation sites that are highly conserved across species. Second, we show via electrophysiology experiments that KNa currents and further, Slack currents, are subject to tonic modulation by p38 MAPK. Third, biochemical approaches revealed that Slack channel regulation by p38 MAPK occurs through direct phosphorylation at the two putative sites of interaction, and mutating both sites prevented surface expression of Slack channels. Based on these results, we conclude that p38 MAPK is an obligate regulator of Slack channel function via the trafficking of channels into the membrane. The present study identifies Slack KNa channels as p38 MAPK substrates. Copyright © 2015 Elsevier Ltd. All rights reserved.

Slack Sodium-activated Potassium Channel Membrane Expression Requires p38 Mitogen-Activated Protein Kinase Phosphorylation

PubMed Central

Gururaj, Sushmitha; Fleites, John; Bhattacharjee, Arin

2016-01-01

p38 MAPK has long been understood as an inducible kinase under conditions of cellular stress, but there is now increasing evidence to support its role in the regulation of neuronal function. Several phosphorylation targets have been identified, an appreciable number of which are ion channels, implicating the possible involvement of p38 MAPK in neuronal excitability. The KNa channel Slack is an important protein to be studied as it is highly and ubiquitously expressed in DRG neurons and is important in the maintenance of their firing accommodation. We sought to examine if the Slack channel could be a substrate of p38 MAPK activity. First, we found that the Slack C-terminus contains two putative p38 MAPK phosphorylation sites that are highly conserved across species. Second, we show via electrophysiology experiments that KNa currents and further, Slack currents, are subject to tonic modulation by p38 MAPK. Third, biochemical approaches revealed that Slack channel regulation by p38 MAPK occurs through direct phosphorylation at the two putative sites of interaction, and mutating both sites prevented surface expression of Slack channels. Based on these results, we conclude that p38 MAPK is an obligate regulator of Slack channel function via the trafficking of channels into the membrane. The present study identifies Slack KNa channels as p38 MAPK substrates. PMID:26721627

Inward-rectifying potassium (Kir) channels regulate pacemaker activity in spinal nociceptive circuits during early life

PubMed Central

Li, Jie; Blankenship, Meredith L.; Baccei, Mark L.

2013-01-01

Pacemaker neurons in neonatal spinal nociceptive circuits generate intrinsic burst-firing and are distinguished by a lower “leak” membrane conductance compared to adjacent, non-bursting neurons. However, little is known about which subtypes of leak channels regulate the level of pacemaker activity within the developing rat superficial dorsal horn (SDH). Here we demonstrate that a hallmark feature of lamina I pacemaker neurons is a reduced conductance through inward-rectifying potassium (Kir) channels at physiological membrane potentials. Differences in the strength of inward rectification between pacemakers and non-pacemakers indicate the presence of functionally distinct Kir currents in these two populations at room temperature. However, Kir currents in both groups showed high sensitivity to block by extracellular Ba2+ (IC50 ~ 10 µM), which suggests the presence of ‘classical’ Kir (Kir2.x) channels in the neonatal SDH. The reduced Kir conductance within pacemakers is unlikely to be explained by an absence of particular Kir2.x isoforms, as immunohistochemical analysis revealed the expression of Kir2.1, Kir2.2 and Kir2.3 within spontaneously bursting neurons. Importantly, Ba2+ application unmasked rhythmic burst-firing in ~42% of non-bursting lamina I neurons, suggesting that pacemaker activity is a latent property of a sizeable population of SDH cells during early life. In addition, the prevalence of spontaneous burst-firing within lamina I was enhanced in the presence of high internal concentrations of free Mg2+, consistent with its documented ability to block Kir channels from the intracellular side. Collectively, the results indicate that Kir channels are key modulators of pacemaker activity in newborn central pain networks. PMID:23426663

The SK3 channel promotes placental vascularization by enhancing secretion of angiogenic factors.

PubMed

Rada, Cara C; Murray, Grace; England, Sarah K

2014-11-15

Proper placental perfusion is essential for fetal exchange of oxygen, nutrients, and waste with the maternal circulation. Impairment of uteroplacental vascular function can lead to pregnancy complications, including preeclampsia and intrauterine growth restriction (IUGR). Potassium channels have been recognized as regulators of vascular proliferation, angiogenesis, and secretion of vasoactive factors, and their dysfunction may underlie pregnancy-related vascular diseases. Overexpression of one channel in particular, the small-conductance calcium-activated potassium channel 3 (SK3), is known to increase vascularization in mice, and mice overexpressing the SK3 channel (SK3(T/T) mice) have a high rate of fetal demise and IUGR. Here, we show that overexpression of SK3 causes fetal loss through abnormal placental vascularization. We previously reported that, at pregnancy day 14, placentas isolated from SK3(T/T) mice are smaller than those obtained from wild-type mice. In this study, histological analysis reveals that SK3(T/-) placentas at this stage have abnormal placental morphology, and microcomputed tomography shows that these placentas have significantly larger and more blood vessels than those from wild-type mice. To identify the mechanism by which these vascularization defects occur, we measured levels of vascular endothelial growth factor (VEGF), placental growth factor, and the soluble form of VEGF receptor 1 (sFlt-1), which must be tightly regulated to ensure proper placental development. Our data reveal that overexpression of SK3 alters systemic and placental ratios of the angiogenic factor VEGF to antiangiogenic factor sFlt-1 throughout pregnancy. Additionally, we observe increased expression of hypoxia-inducing factor 2α in SK3(T/-) placentas. We conclude that the SK3 channel modulates placental vascular development and fetal health by altering VEGF signaling. Copyright © 2014 the American Physiological Society.

Location of the β4 transmembrane helices in the BK potassium channel

PubMed Central

Wu, Roland S.; Chudasama, Neelesh; Zakharov, Sergey I.; Doshi, Darshan; Motoike, Howard; Liu, Guoxia; Yao, Yongneng; Niu, Xiaowei; Deng, Shi-Xian; Landry, Donald W.; Karlin, Arthur; Marx, Steven O.

2009-01-01

Large-conductance, voltage- and Ca2+-gated potassium (BK) channels control excitability in a number of cell types. BK channels are composed of α subunits, which contain the voltage-sensor domains and the Ca2+- sensor domains, and form the pore, and often one of four types of β subunits, which modulate the channel in a cell-specific manner. β4 is expressed in neurons throughout the brain. Deletion of β4 in mice causes temporal lobe epilepsy. Compared to channels composed of α alone, channels composed of α and β4 activate and deactivate more slowly. We inferred the locations of the two β4 transmembrane (TM) helices, TM1 and TM2, relative to the seven αTM helices, S0-S6, from the extent of disulfide bond formation between cysteines substituted in the extracellular flanks of these TM helices. We found that β4 TM2 is close to α S0 and that β4 TM1 is close to both α S1 and S2. At least at their extracellular ends, TM1 and TM2 are not close to S3 through S6. In six of eight of the most highly crosslinked cysteine pairs, four crosslinks from TM2 to S0 and one each from TM1 to S1 and S2 had small effects on the V50 and on the rates of activation and deactivation. That disulfide crosslinking caused only small functional perturbations is consistent with the proximity of the extracellular ends of TM2 to S0 and of TM1 to S1 and to S2, in both the open and closed states. PMID:19571123

Downregulation of BK Channel Function and Protein Expression in Coronary Arteriolar Smooth Muscle Cells of Type 2 Diabetic Patients.

PubMed

Lu, Tong; Chai, Qiang; Jiao, Guoqing; Wang, Xiao-Li; Sun, Xiaojing; Furuseth, Jonathan D; Stulak, John M; Daly, Richard C; Greason, Kevin L; Cha, Yong-Mei; Lee, Hon-Chi

2018-05-30

Type 2 diabetes (T2D) is strongly associated with cardiovascular morbidity and mortality in patients. Vascular large conductance Ca2+-activated potassium (BK) channels, composed of four pore-forming α subunits (BK-α) and four regulatory β1 subunits (BK-β1), are densely expressed in coronary arterial smooth muscle cells (SMCs) and play an important role in regulating vascular tone and myocardial perfusion. However, the role of BK channels in coronary microvascular dysfunction of human subjects with diabetes is unclear. In this study, we examined BK channel function and protein expression, and BK channel-mediated vasodilation in freshly isolated coronary arterioles from T2D patients. Atrial tissues were obtained from 25 patients with T2D and 16 matched non-diabetic subjects during cardiopulmonary bypass procedure. Microvessel videomicroscopy and immunoblot analysis were performed in freshly dissected coronary arterioles and inside-out single BK channel currents was recorded in enzymatically isolated coronary arteriolar SMCs. We found that BK channel sensitivity to physiological Ca2+ concentration and voltage was downregulated in the coronary arteriolar SMCs of diabetic patients, compared with non-diabetic controls. BK channel kinetics analysis revealed that there was significant shortening of the mean open time and prolongation of the mean closed time in diabetic patients, resulting in a remarkable reduction of the channel open probability. Functional studies showed that BK channel activation by dehydrosoyasaponin-1 was diminished and that BK channel-mediated vasodilation in response to shear stress was impaired in diabetic coronary arterioles. Immunoblot experiments confirmed that the protein expressions of BK-α and BK-β1 subunits were significantly downregulated, but the ratio of BK-α/BK-β1 was unchanged in the coronary arterioles of T2D patients. Our results demonstrated for the first time that BK channel function and BK channel-mediated vasodilation were abnormal in the coronary microvasculature of diabetic patients, due to decreased protein expression and altered intrinsic properties of BK channels.

Molecular mechanism underlying β1 regulation in voltage- and calcium-activated potassium (BK) channels

PubMed Central

Castillo, Karen; Contreras, Gustavo F.; Pupo, Amaury; Torres, Yolima P.; Neely, Alan; González, Carlos; Latorre, Ramon

2015-01-01

Being activated by depolarizing voltages and increases in cytoplasmic Ca2+, voltage- and calcium-activated potassium (BK) channels and their modulatory β-subunits are able to dampen or stop excitatory stimuli in a wide range of cellular types, including both neuronal and nonneuronal tissues. Minimal alterations in BK channel function may contribute to the pathophysiology of several diseases, including hypertension, asthma, cancer, epilepsy, and diabetes. Several gating processes, allosterically coupled to each other, control BK channel activity and are potential targets for regulation by auxiliary β-subunits that are expressed together with the α (BK)-subunit in almost every tissue type where they are found. By measuring gating currents in BK channels coexpressed with chimeras between β1 and β3 or β2 auxiliary subunits, we were able to identify that the cytoplasmic regions of β1 are responsible for the modulation of the voltage sensors. In addition, we narrowed down the structural determinants to the N terminus of β1, which contains two lysine residues (i.e., K3 and K4), which upon substitution virtually abolished the effects of β1 on charge movement. The mechanism by which K3 and K4 stabilize the voltage sensor is not electrostatic but specific, and the α (BK)-residues involved remain to be identified. This is the first report, to our knowledge, where the regulatory effects of the β1-subunit have been clearly assigned to a particular segment, with two pivotal amino acids being responsible for this modulation. PMID:25825713

Endoplasmic reticulum-associated degradation of the renal potassium channel, ROMK, leads to type II Bartter syndrome.

PubMed

O'Donnell, Brighid M; Mackie, Timothy D; Subramanya, Arohan R; Brodsky, Jeffrey L

2017-08-04

Type II Bartter syndrome is caused by mutations in the renal outer medullary potassium (ROMK) channel, but the molecular mechanisms underlying this disease are poorly defined. To rapidly screen for ROMK function, we developed a yeast expression system and discovered that yeast cells lacking endogenous potassium channels could be rescued by WT ROMK but not by ROMK proteins containing any one of four Bartter mutations. We also found that the mutant proteins were significantly less stable than WT ROMK. However, their degradation was slowed in the presence of a proteasome inhibitor or when yeast cells contained mutations in the CDC48 or SSA1 gene, which is required for endoplasmic reticulum (ER)-associated degradation (ERAD). Consistent with these data, sucrose gradient centrifugation and indirect immunofluorescence microscopy indicated that most ROMK protein was ER-localized. To translate these findings to a more relevant cell type, we measured the stabilities of WT ROMK and the ROMK Bartter mutants in HEK293 cells. As in yeast, the Bartter mutant proteins were less stable than the WT protein, and their degradation was slowed in the presence of a proteasome inhibitor. Finally, we discovered that low-temperature incubation increased the steady-state levels of a Bartter mutant, suggesting that the disease-causing mutation traps the protein in a folding-deficient conformation. These findings indicate that the underlying pathology for at least a subset of patients with type II Bartter syndrome is linked to the ERAD pathway and that future therapeutic strategies should focus on correcting deficiencies in ROMK folding. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Crosstalk between KCNK3-Mediated Ion Current and Adrenergic Signaling Regulates Adipose Thermogenesis and Obesity.

PubMed

Chen, Yi; Zeng, Xing; Huang, Xuan; Serag, Sara; Woolf, Clifford J; Spiegelman, Bruce M

2017-11-02

Adrenergic stimulation promotes lipid mobilization and oxidation in brown and beige adipocytes, where the harnessed energy is dissipated as heat in a process known as adaptive thermogenesis. The signaling cascades and energy-dissipating pathways that facilitate thermogenesis have been extensively described, yet little is known about the counterbalancing negative regulatory mechanisms. Here, we identify a two-pore-domain potassium channel, KCNK3, as a built-in rheostat negatively regulating thermogenesis. Kcnk3 is transcriptionally wired into the thermogenic program by PRDM16, a master regulator of thermogenesis. KCNK3 antagonizes norepinephrine-induced membrane depolarization by promoting potassium efflux in brown adipocytes. This limits calcium influx through voltage-dependent calcium channels and dampens adrenergic signaling, thereby attenuating lipolysis and thermogenic respiration. Adipose-specific Kcnk3 knockout mice display increased energy expenditure and are resistant to hypothermia and obesity. These findings uncover a critical K + -Ca 2+ -adrenergic signaling axis that acts to dampen thermogenesis, maintain tissue homeostasis, and reveal an electrophysiological regulatory mechanism of adipocyte function. Copyright © 2017 Elsevier Inc. All rights reserved.

Non-equivalent role of TM2 gating hinges in heteromeric Kir4.1/Kir5.1 potassium channels.

PubMed

Shang, Lijun; Tucker, Stephen J

2008-02-01

Comparison of the crystal structures of the KcsA and MthK potassium channels suggests that the process of opening a K(+) channel involves pivoted bending of the inner pore-lining helices at a highly conserved glycine residue. This bending motion is proposed to splay the transmembrane domains outwards to widen the gate at the "helix-bundle crossing". However, in the inwardly rectifying (Kir) potassium channel family, the role of this "hinge" residue in the second transmembrane domain (TM2) and that of another putative glycine gating hinge at the base of TM2 remain controversial. We investigated the role of these two positions in heteromeric Kir4.1/Kir5.1 channels, which are unique amongst Kir channels in that both subunits lack a conserved glycine at the upper hinge position. Contrary to the effect seen in other channels, increasing the potential flexibility of TM2 by glycine substitutions at the upper hinge position decreases channel opening. Furthermore, the contribution of the Kir4.1 subunit to this process is dominant compared to Kir5.1, demonstrating a non-equivalent contribution of these two subunits to the gating process. A homology model of heteromeric Kir4.1/Kir5.1 shows that these upper "hinge" residues are in close contact with the base of the pore alpha-helix that supports the selectivity filter. Our results also indicate that the highly conserved glycine at the "lower" gating hinge position is required for tight packing of the TM2 helices at the helix-bundle crossing, rather than acting as a hinge residue.

Toxic effects of environmental rare earth elements on delayed outward potassium channels and their mechanisms from a microscopic perspective.

PubMed

Wang, Lihong; He, Jingfang; Xia, Ao; Cheng, Mengzhu; Yang, Qing; Du, Chunlei; Wei, Haiyan; Huang, Xiaohua; Zhou, Qing

2017-08-01

The wide applications cause a large amount of rare earth elements (REEs) to be released into the environment, and ultimately into the human body through food chain. Toxic effects of REEs on humans have been extensively studied, but their toxic effects and binding targets in cells are not understood. Delayed outward potassium channels (K + channels) are good targets for exogenous substances or clinical drugs. To evaluate cellular toxicities of REEs and clarify toxic mechanisms, the toxicities of REEs on the K + channel and their structural basis were investigated. The results showed that delayed outward potassium channels on the plasma membrane are the targets of REEs acting on living organisms, and the changes in the thermodynamic and kinetic characteristics of the K + channel are the reasons of diseases induced by REEs. Two types of REEs, a light REE La 3+ and a heavy REE Tb 3+ , displayed different intensity of toxicities on the K + channel, in which the toxicity of Tb 3+ was stronger than that of La 3+ . More interestingly, in comparison with that of heavy metal Cd 2+ , the cytotoxicities of the light and heavy REEs showed discriminative differences, and the cytotoxicity of Tb 3+ was higher than that of Cd 2+ , while the cytotoxicity of La 3+ was lower than that of Cd 2+ . These different cytotoxicities of La 3+ , Tb 3+ and Cd 2+ on human resulted from the varying binding abilities of the metals to this channel protein. Copyright © 2017 Elsevier Ltd. All rights reserved.

Gating behavior of endoplasmic reticulum potassium channels of rat hepatocytes in diabetes.

PubMed

Ghasemi, Maedeh; Khodaei, Naser; Salari, Sajjad; Eliassi, Afsaneh; Saghiri, Reza

2014-07-01

Defects in endoplasmic reticulum homeostasis are common occurrences in different diseases, such as diabetes, in which the function of endoplasmic reticulum is disrupted. It is now well established that ion channels of endoplasmic reticulum membrane have a critical role in endoplasmic reticulum luminal homeostasis. Our previous studies showed the presence of an ATP-sensitive cationic channel in endoplasmic reticulum. Therefore, in this study, we examined and compared the activities of this channel in control and diabetic rats using single-channel recording techniques. Male Wistar rats were made diabetic for 2 weeks with a single dose injection of streptozotocin (45 mg/kg). Ion channel incorporation of rough endoplasmic reticulum of diabetic hepatocytes into the bilayer lipid membrane allowed the characterization of K+ channel. Ion channel incorporation of rough endoplasmic reticulum vesicles into the bilayer lipid revealed that the channel current-voltage (I-V) relation with a mean slope conductance of 520 ± 19 pS was unaffected in diabetes. Interestingly, the channel Po-voltage relation was significantly lower in diabetic rats at voltages above +30 mV. We concluded that the endoplasmic reticulum cationic channel is involved in diabetes. Also, this finding could be considered as a goal for further therapeutic plans.

Regulation of aldosterone production by ion channels: From basal secretion to primary aldosteronism.

PubMed

Yang, Tingting; He, Min; Hu, Changlong

2018-03-01

Aldosterone is produced by zona glomerulosa (ZG) cells of the adrenal cortex and plays a key role in balancing water and electrolytes levels. Autonomous overproduction of aldosterone leads to primary aldosteronism (PA), which is the most common form of secondary endocrine hypertension. Recently, significant progress has been made towards understanding the genetic basis of PA, where increasing clinical evidence suggests that mutations in ion channels appear to be the major cause of aldosterone-producing adenomas. In this review, we focused on potassium and calcium channels that regulate aldosterone secretion, and their roles in the pathology of PA. Because potassium and calcium channels are differentially expressed in ZG cells in different species of mammals, the limitations of published studies are also discussed. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of genes from pattern formation, tyrosine kinase, and potassium channel families by DNA amplification

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kamb, A.; Weir, M.; Rudy, B.

1989-06-01

The study of gene family members has been aided by the isolation of related genes on the basis of DNA homology. The authors have adapted the polymerase chain reaction to screen animal genomes very rapidly and reliably for likely gene family members. Using conserved amino acid sequences to design degenerate oligonucleotide primers, they have shown that the genome of the nematode Caenorhabditis elegans contains sequences homologous to many Drosophila genes involved in pattern formation, including the segment polarity gene wingless (vertebrate int-1), and homeobox sequences characteristic of the Antennapedia, engrailed, and paired families. In addition, they have used this methodmore » to show that C. elegans contains at least five different sequences homologous to genes in the tyrosine kinase family. Lastly, they have isolated six potassium channel sequences from humans, a result that validates the utility of the method with large genomes and suggests that human potassium channel gene diversity may be extensive.« less

Effects of strychnine on the sodium conductance of the frog node of Ranvier

PubMed Central

1977-01-01

Strychnine blocks sodium conductance in the frog node of Ranvier. This block was studied by reducing and slowing sodium inactivation with scorpion venom. The block is voltage and time dependent. The more positive the axoplasm the greater the block and the faster the approach to equilibrium. Some evidence is presented suggesting that only open channels can be blocked. The block is reduced by raising external sodium or lithium but not impermeant cations. A quaternary derivative of strychnine was synthesized and found to have the same action only when applied intracellularly. We conclude that strychnine blocks sodium channels by a mechanism analogous to that by which it blocks potassium channels. The potassium channel block had previously been found to be identical to that by tetraethylammonium ion derivatives. In addition, strychnine resembles procaine and its derivatives in both its structure and the mechanism of sodium channel block. PMID:302321

The orientation and molecular movement of a k(+) channel voltage-sensing domain.

PubMed

Gandhi, Chris S; Clark, Eliana; Loots, Eli; Pralle, Arnd; Isacoff, Ehud Y

2003-10-30

Voltage-gated channels operate through the action of a voltage-sensing domain (membrane segments S1-S4) that controls the conformation of gates located in the pore domain (membrane segments S5-S6). Recent structural studies on the bacterial K(v)AP potassium channel have led to a new model of voltage sensing in which S4 lies in the lipid at the channel periphery and moves through the membrane as a unit with a portion of S3. Here we describe accessibility probing and disulfide scanning experiments aimed at determining how well the K(v)AP model describes the Drosophila Shaker potassium channel. We find that the S1-S3 helices have one end that is externally exposed, S3 does not undergo a transmembrane motion, and S4 lies in close apposition to the pore domain in the resting and activated state.

Prostaglandin E2 activates channel-mediated calcium entry in human erythrocytes: an indication for a blood clot formation supporting process.

PubMed

Kaestner, Lars; Tabellion, Wiebke; Lipp, Peter; Bernhardt, Ingolf

2004-12-01

Prostaglandin E(2) (PGE(2)) is released from platelets when they are activated. Using fluorescence imaging and the patch-clamp technique, we provide evidence that PGE(2) at physiological concentrations (10(-10) M) activates calcium rises mediated by calcium influx through a non-selective cation-channel in human red blood cells. The extent of calcium increase varied between cells with a total of 45% of the cells responding. It is well known that calcium increases elicited the calcium-activated potassium channel (Gardos channel) in the red cell membrane. Previously, it was shown that the Gardos channel activation results in potassium efflux and shrinkage of the cells. Therefore, we conclude that the PGE(2) responses of red blood cells described here reveal a direct and active participation of erythrocytes in blood clot formation.

Isolation and Characterization of a High Affinity Peptide Inhibitor of ClC-2 Chloride Channels*

PubMed Central

Thompson, Christopher H.; Olivetti, Pedro R.; Fuller, Matthew D.; Freeman, Cody S.; McMaster, Denis; French, Robert J.; Pohl, Jan; Kubanek, Julia; McCarty, Nael A.

2009-01-01

The ClC protein family includes voltage-gated chloride channels and chloride/proton exchangers. In eukaryotes, ClC proteins regulate membrane potential of excitable cells, contribute to epithelial transport, and aid in lysosomal acidification. Although structure/function studies of ClC proteins have been aided greatly by the available crystal structures of a bacterial ClC chloride/proton exchanger, the availability of useful pharmacological tools, such as peptide toxin inhibitors, has lagged far behind that of their cation channel counterparts. Here we report the isolation, from Leiurus quinquestriatus hebraeus venom, of a peptide toxin inhibitor of the ClC-2 chloride channel. This toxin, GaTx2, inhibits ClC-2 channels with a voltage-dependent apparent KD of ∼20 pm, making it the highest affinity inhibitor of any chloride channel. GaTx2 slows ClC-2 activation by increasing the latency to first opening by nearly 8-fold but is unable to inhibit open channels, suggesting that this toxin inhibits channel activation gating. Finally, GaTx2 specifically inhibits ClC-2 channels, showing no inhibitory effect on a battery of other major classes of chloride channels and voltage-gated potassium channels. GaTx2 is the first peptide toxin inhibitor of any ClC protein. The high affinity and specificity displayed by this toxin will make it a very powerful pharmacological tool to probe ClC-2 structure/function. PMID:19574231

Cortisone and hydrocortisone inhibit human Kv1.3 activity in a non-genomic manner.

PubMed

Yu, Jing; Park, Mi-Hyeong; Choi, Se-Young; Jo, Su-Hyun

2015-06-01

Glucocorticoids are hormones released in response to stress that are involved in various physiological processes including immune functions. One immune-modulating mechanism is achieved by the Kv1.3 voltage-dependent potassium channel, which is expressed highly in lymphocytes including effector memory T lymphocytes (TEM). Although glucocorticoids are known to inhibit Kv1.3 function, the detailed inhibitory mechanism is not yet fully understood. Here we studied the rapid non-genomic effects of cortisone and hydrocortisone on the human Kv1.3 channel expressed in Xenopus oocytes. Both cortisone and hydrocortisone reduced the amplitude of the Kv1.3 channel current in a concentration-dependent manner. Both cortisone and hydrocortisone rapidly and irreversibly inhibited Kv1.3 currents, eliminating the possibility of genomic regulation. Inhibition rate was stable relative to the degree of depolarization. Kinetically, cortisone altered the activating gate of Kv1.3 and hydrocortisone interacted with this channel in an open state. These results suggest that cortisone and hydrocortisone inhibit Kv1.3 currents via a non-genomic mechanism, providing a mechanism for the immunosuppressive effects of glucocorticoids.

Dynamic oligomeric conversions of the cytoplasmic RCK domains mediate MthK potassium channel activity.

PubMed

Kuo, Mario Meng-Chiang; Baker, Kent A; Wong, Lee; Choe, Senyon

2007-02-13

The crystal structure of the RCK-containing MthK provides a molecular framework for understanding the ligand gating mechanisms of K+ channels. Here we examined the macroscopic currents of MthK in enlarged Escherichia coli membrane by patch clamp and rapid perfusion techniques and showed that the channel undergoes desensitization in seconds after activation by Ca2+ or Cd2+. Additionally, MthK is inactivated by slightly acidic pH only from the cytoplasmic side. Examinations of isolated RCK domain by size-exclusion chromatography, static light scattering, analytical sedimentation, and stopped-flow spectroscopy show that Ca2+ rapidly converts isolated RCK monomers to multimers at alkaline pH. In contrast, the RCK domain at acidic pH remains firmly dimeric regardless of Ca2+ but restores predominantly to multimer or monomer at basic pH with or without Ca2+, respectively. These functional and biochemical analyses correlate the four functional states of the MthK channel with distinct oligomeric states of its RCK domains and indicate that the RCK domains undergo oligomeric conversions in modulating MthK activities.

Ciguatoxins: Cyclic Polyether Modulators of Voltage-gated Iion Channel Function

PubMed Central

Nicholson, Graham M.; Lewis, Richard J.

2006-01-01

Ciguatoxins are cyclic polyether toxins, derived from marine dinoflagellates, which are responsible for the symptoms of ciguatera poisoning. Ingestion of tropical and subtropical fin fish contaminated by ciguatoxins results in an illness characterised by neurological, cardiovascular and gastrointestinal disorders. The pharmacology of ciguatoxins is characterised by their ability to cause persistent activation of voltage-gated sodium channels, to increase neuronal excitability and neurotransmitter release, to impair synaptic vesicle recycling, and to cause cell swelling. It is these effects, in combination with an action to block voltage-gated potassium channels at high doses, which are believed to underlie the complex of symptoms associated with ciguatera. This review examines the sources, structures and pharmacology of ciguatoxins. In particular, attention is placed on their cellular modes of actions to modulate voltage-gated ion channels and other Na+-dependent mechanisms in numerous cell types and to current approaches for detection and treatment of ciguatera.

Dendritic A-type potassium channel subunit expression in CA1 hippocampal interneurons.

PubMed

Menegola, M; Misonou, H; Vacher, H; Trimmer, J S

2008-06-26

Voltage-gated potassium (Kv) channels are important and diverse determinants of neuronal excitability and exhibit specific expression patterns throughout the brain. Among Kv channels, Kv4 channels are major determinants of somatodendritic A-type current and are essential in controlling the amplitude of backpropagating action potentials (BAPs) into neuronal dendrites. BAPs have been well studied in a variety of neurons, and have been recently described in hippocampal and cortical interneurons, a heterogeneous population of GABAergic inhibitory cells that regulate activity of principal cells and neuronal networks. We used well-characterized mouse monoclonal antibodies against the Kv4.3 and potassium channel interacting protein (KChIP) 1 subunits of A-type Kv channels, and antibodies against different interneuron markers in single- and double-label immunohistochemistry experiments to analyze the expression patterns of Kv4.3 and KChIP1 in hippocampal Ammon's horn (CA1) neurons. Immunohistochemistry was performed on 40 mum rat brain sections using nickel-enhanced diaminobenzidine staining or multiple-label immunofluorescence. Our results show that Kv4.3 and KChIP1 component subunits of A-type channels are co-localized in the soma and dendrites of a large number of GABAergic hippocampal interneurons. These subunits co-localize extensively but not completely with markers defining the four major interneuron subpopulations tested (parvalbumin, calbindin, calretinin, and somatostatin). These results suggest that CA1 hippocampal interneurons can be divided in two groups according to the expression of Kv4.3/KChIP1 channel subunits. Antibodies against Kv4.3 and KChIP1 represent an important new tool for identifying a subpopulation of hippocampal interneurons with a unique dendritic A-type channel complement and ability to control BAPs.

Taurine activates delayed rectifier KV channels via a metabotropic pathway in retinal neurons

PubMed Central

Bulley, Simon; Liu, Yufei; Ripps, Harris; Shen, Wen

2013-01-01

Taurine is one of the most abundant amino acids in the retina, throughout the CNS, and in heart and muscle cells. In keeping with its broad tissue distribution, taurine serves as a modulator of numerous basic processes, such as enzyme activity, cell development, myocardial function and cytoprotection. Despite this multitude of functional roles, the precise mechanism underlying taurine's actions has not yet been identified. In this study we report findings that indicate a novel role for taurine in the regulation of voltage-gated delayed rectifier potassium (KV) channels in retinal neurons by means of a metabotropic receptor pathway. The metabotropic taurine response was insensitive to the Cl− channel blockers, picrotoxin and strychnine, but it was inhibited by a specific serotonin 5-HT2A receptor antagonist, MDL11939. Moreover, we found that taurine enhanced KV channels via intracellular protein kinase C-mediated pathways. When 5-HT2A receptors were expressed in human embryonic kidney cells, taurine and AL34662, a non-specific 5-HT2 receptor activator, produced a similar regulation of KIR channels. In sum, this study provides new evidence that taurine activates a serotonin system, apparently via 5-HT2A receptors and related intracellular pathways. PMID:23045337

Preventive dietary potassium supplementation in young salt-sensitive Dahl rats attenuates development of salt hypertension by decreasing sympathetic vasoconstriction.

PubMed

Zicha, J; Dobešová, Z; Behuliak, M; Kuneš, J; Vaněčková, I

2011-05-01

Increased potassium intake attenuates the development of salt-dependent hypertension, but the detailed mechanisms of blood pressure (BP) reduction are still unclear. The aims of our study were (i) to elucidate these mechanisms, (ii) to compare preventive potassium effects in immature and adult animals and (iii) to evaluate the therapeutic effects of dietary potassium supplementation in rats with established salt hypertension.   Young (4-week-old) and adult (24-week-old) female salt-sensitive Dahl rats were fed a high-salt diet (5% NaCl) or a high-salt diet supplemented with 3% KCl for 5 weeks. The participation of vasoconstrictor (renin-angiotensin and sympathetic nervous systems) and vasodilator systems [prostanoids, Ca(2+) -activated K(+) channels, nitric oxide (NO)] was evaluated using a sequential blockade of these systems. Preventive potassium supplementation attenuated the development of severe salt hypertension in young rats, whereas it had no effects on BP in adult rats with moderate hypertension. Enhanced sympathetic vasoconstriction was responsible for salt hypertension in young rats and its attenuation for potassium-induced BP reduction. Conversely, neither salt hypertension nor its potassium-induced attenuation were associated with significant changes of the vasodilator systems studied. The relative deficiency of vasodilator action of NO and Ca(2+) -activated K(+) channels in salt hypertensive Dahl rats was not improved by potassium supplementation. The attenuation of enhanced sympathetic vasoconstriction is the principal mechanism of antihypertensive action exerted by preventive potassium supplementation in immature Dahl rats. Dietary potassium supplementation has no preventive effects on BP in adult salt-loaded animals or no therapeutic effects on established salt hypertension in young rats. © 2011 The Authors. Acta Physiologica © 2011 Scandinavian Physiological Society.

Dystrophin Is Required for the Normal Function of the Cardio-Protective KATP Channel in Cardiomyocytes

PubMed Central

Graciotti, Laura; Becker, Jodi; Granata, Anna Luisa; Procopio, Antonio Domenico; Tessarollo, Lino; Fulgenzi, Gianluca

2011-01-01

Duchenne and Becker muscular dystrophy patients often develop a cardiomyopathy for which the pathogenesis is still unknown. We have employed the murine animal model of Duchenne muscular dystrophy (mdx), which develops a cardiomyopathy that includes some characteristics of the human disease, to study the molecular basis of this pathology. Here we show that the mdx mouse heart has defects consistent with alteration in compounds that regulate energy homeostasis including a marked decrease in creatine-phosphate (PC). In addition, the mdx heart is more susceptible to anoxia than controls. Since the cardio-protective ATP sensitive potassium channel (KATP) complex and PC have been shown to interact we investigated whether deficits in PC levels correlate with other molecular events including KATP ion channel complex presence, its functionality and interaction with dystrophin. We found that this channel complex is present in the dystrophic cardiac cell membrane but its ability to sense a drop in the intracellular ATP concentration and consequently open is compromised by the absence of dystrophin. We further demonstrate that the creatine kinase muscle isoform (CKm) is displaced from the plasma membrane of the mdx cardiac cells. Considering that CKm is a determinant of KATP channel complex function we hypothesize that dystrophin acts as a scaffolding protein organizing the KATP channel complex and the enzymes necessary for its correct functioning. Therefore, the lack of proper functioning of the cardio-protective KATP system in the mdx cardiomyocytes may be part of the mechanism contributing to development of cardiac disease in dystrophic patients. PMID:22066028

Mutations in Nature Conferred a High Affinity Phosphatidylinositol 4,5-Bisphosphate-binding Site in Vertebrate Inwardly Rectifying Potassium Channels*

PubMed Central

Tang, Qiong-Yao; Larry, Trevor; Hendra, Kalen; Yamamoto, Erica; Bell, Jessica; Cui, Meng; Logothetis, Diomedes E.; Boland, Linda M.

2015-01-01

All vertebrate inwardly rectifying potassium (Kir) channels are activated by phosphatidylinositol 4,5-bisphosphate (PIP2) (Logothetis, D. E., Petrou, V. I., Zhang, M., Mahajan, R., Meng, X. Y., Adney, S. K., Cui, M., and Baki, L. (2015) Annu. Rev. Physiol. 77, 81–104; Fürst, O., Mondou, B., and D'Avanzo, N. (2014) Front. Physiol. 4, 404–404). Structural components of a PIP2-binding site are conserved in vertebrate Kir channels but not in distantly related animals such as sponges and sea anemones. To expand our understanding of the structure-function relationships of PIP2 regulation of Kir channels, we studied AqKir, which was cloned from the marine sponge Amphimedon queenslandica, an animal that represents the phylogenetically oldest metazoans. A requirement for PIP2 in the maintenance of AqKir activity was examined in intact oocytes by activation of a co-expressed voltage-sensing phosphatase, application of wortmannin (at micromolar concentrations), and activation of a co-expressed muscarinic acetylcholine receptor. All three mechanisms to reduce the availability of PIP2 resulted in inhibition of AqKir current. However, time-dependent rundown of AqKir currents in inside-out patches could not be re-activated by direct application to the inside membrane surface of water-soluble dioctanoyl PIP2, and the current was incompletely re-activated by the more hydrophobic arachidonyl stearyl PIP2. When we introduced mutations to AqKir to restore two positive charges within the vertebrate PIP2-binding site, both forms of PIP2 strongly re-activated the mutant sponge channels in inside-out patches. Molecular dynamics simulations validate the additional hydrogen bonding potential of the sponge channel mutants. Thus, nature's mutations conferred a high affinity activation of vertebrate Kir channels by PIP2, and this is a more recent evolutionary development than the structures that explain ion channel selectivity and inward rectification. PMID:25957411

Chloride channels in cancer: Focus on chloride intracellular channel 1 and 4 (CLIC1 AND CLIC4) proteins in tumor development and as novel therapeutic targets.

PubMed

Peretti, Marta; Angelini, Marina; Savalli, Nicoletta; Florio, Tullio; Yuspa, Stuart H; Mazzanti, Michele

2015-10-01

In recent decades, growing scientific evidence supports the role of ion channels in the development of different cancers. Both potassium selective pores and chloride permeabilities are considered the most active channels during tumorigenesis. High rate of proliferation, active migration, and invasiveness into non-neoplastic tissues are specific properties of neoplastic transformation. All these actions require partial or total involvement of chloride channel activity. In this context, this class of membrane proteins could represent valuable therapeutic targets for the treatment of resistant tumors. However, this encouraging premise has not so far produced any valid new channel-targeted antitumoral molecule for cancer treatment. Problematic for drug design targeting ion channels is their vital role in normal cells for essential physiological functions. By targeting these membrane proteins involved in pathological conditions, it is inevitable to cause relevant side effects in healthy organs. In light of this, a new protein family, the chloride intracellular channels (CLICs), could be a promising class of therapeutic targets for its intrinsic individualities: CLIC1 and CLIC4, in particular, not only are overexpressed in specific tumor types or their corresponding stroma but also change localization and function from hydrophilic cytosolic to integral transmembrane proteins as active ionic channels or signal transducers during cell cycle progression in certain cases. These changes in intracellular localization, tissue compartments, and channel function, uniquely associated with malignant transformation, may offer a unique target for cancer therapy, likely able to spare normal cells. This article is part of a special issue itled "Membrane Channels and Transporters in Cancers." Copyright © 2015 Elsevier B.V. All rights reserved.

Ion Channels in Obesity: Pathophysiology and Potential Therapeutic Targets

PubMed Central

Vasconcelos, Luiz H. C.; Souza, Iara L. L.; Pinheiro, Lílian S.; Silva, Bagnólia A.

2016-01-01

Obesity is a multifactorial disease related to metabolic disorders and associated with genetic determinants. Currently, ion channels activity has been linked to many of these disorders, in addition to the central regulation of food intake, energetic balance, hormone release and response, as well as the adipocyte cell proliferation. Therefore, the objective of this work is to review the current knowledge about the influence of ion channels in obesity development. This review used different sources of literature (Google Scholar, PubMed, Scopus, and Web of Science) to assess the role of ion channels in the pathophysiology of obesity. Ion channels present diverse key functions, such as the maintenance of physiological homeostasis and cell proliferation. Cell biology and pharmacological experimental evidences demonstrate that proliferating cells exhibit ion channel expression, conductance, and electrical properties different from the resting cells. Thereby, a large variety of ion channels has been identified in the pathogenesis of obesity such as potassium, sodium, calcium and chloride channels, nicotinic acetylcholine receptor and transient receptor potential channels. The fundamental involvement of these channels on the generation of obesity leads to the progress in the knowledge about the mechanisms responsible for the obesity pathophysiology, consequently emerging as new targets for pharmacological modulation. PMID:27065858

In-Depth Study of the Interaction, Sensitivity, and Gating Modulation by PUFAs on K+ Channels; Interaction and New Targets

PubMed Central

Moreno, Cristina; de la Cruz, Alicia; Valenzuela, Carmen

2016-01-01

Voltage gated potassium channels (KV) are membrane proteins that allow selective flow of K+ ions in a voltage-dependent manner. These channels play an important role in several excitable cells as neurons, cardiomyocytes, and vascular smooth muscle. Over the last 20 years, it has been shown that omega-3 polyunsaturated fatty acids (PUFAs) enhance or decrease the activity of several cardiac KV channels. PUFAs-dependent modulation of potassium ion channels has been reported to be cardioprotective. However, the precise cellular mechanism underlying the cardiovascular benefits remained unclear in part because new PUFAs targets and signaling pathways continue being discovered. In this review, we will focus on recent data available concerning the following aspects of the KV channel modulation by PUFAs: (i) the exact residues involved in PUFAs-KV channels interaction; (ii) the structural PUFAs determinants important for their effects on KV channels; (iii) the mechanism of the gating modulation of KV channels and, finally, (iv) the PUFAs modulation of a few new targets present in smooth muscle cells (SMC), KCa1.1, K2P, and KATP channels, involved in vascular relaxation. PMID:27933000

Discrete change in volatile anesthetic sensitivity in mice with inactivated tandem pore potassium ion channel TRESK.

PubMed

Chae, Yun Jeong; Zhang, Jianan; Au, Paul; Sabbadini, Marta; Xie, Guo-Xi; Yost, C Spencer

2010-12-01

We investigated the role of tandem pore potassium ion channel (K2P) TRESK in neurobehavioral function and volatile anesthetic sensitivity in genetically modified mice. Exon III of the mouse TRESK gene locus was deleted by homologous recombination using a targeting vector. The genotype of bred mice (wild type, knockout, or heterozygote) was determined using polymerase chain reaction. Morphologic and behavioral evaluations of TRESK knockout mice were compared with wild-type littermates. Sensitivity of bred mice to isoflurane, halothane, sevoflurane, and desflurane were studied by determining the minimum alveolar concentration preventing movement to tail clamping in 50% of each genotype. With the exception of decreased number of inactive periods and increased thermal pain sensitivity (20% decrease in latency with hot plate test), TRESK knockout mice had healthy development and behavior. TRESK knockout mice showed a statistically significant 8% increase in isoflurane minimum alveolar concentration compared with wild-type littermates. Sensitivity to other volatile anesthetics was not significantly different. Spontaneous mortality of TRESK knockout mice after initial anesthesia testing was nearly threefold higher than that of wild-type littermates. TRESK alone is not critical for baseline central nervous system function but may contribute to the action of volatile anesthetics. The inhomogeneous change in anesthetic sensitivity corroborates findings in other K2P knockout mice and supports the theory that the mechanism of volatile anesthetic action involves multiple targets. Although it was not shown in this study, a compensatory effect by other K2P channels may also contribute to these observations.

From in silico to in vitro: a trip to reveal flavonoid binding on the Rattus norvegicus Kir6.1 ATP-sensitive inward rectifier potassium channel.

PubMed

Trezza, Alfonso; Cicaloni, Vittoria; Porciatti, Piera; Langella, Andrea; Fusi, Fabio; Saponara, Simona; Spiga, Ottavia

2018-01-01

ATP-sensitive inward rectifier potassium channels (Kir), are a potassium channel family involved in many physiological processes. K ATP dysfunctions are observed in several diseases such as hypoglycaemia, hyperinsulinemia, Prinzmetal angina-like symptoms, cardiovascular diseases. A broader view of the K ATP mechanism is needed in order to operate on their regulation, and in this work we clarify the structure of the Rattus norvegicus ATP-sensitive inward rectifier potassium channel 8 (Kir6.1), which has been obtained through a homology modelling procedure. Due to the medical use of flavonoids, a considerable increase in studies on their influence on human health has recently been observed, therefore our aim is to study, through computational methods, the three-dimensional (3D) conformation together with mechanism of action of Kir6.1 with three flavonoids. Computational analysis by performing molecular dynamics (MD) and docking simulation on rat 3D modelled structure have been completed, in its closed and open conformation state and in complex with Quercetin, 5-Hydroxyflavone and Rutin flavonoids. Our study showed that only Quercetin and 5-Hydroxyflavone were responsible for a significant down-regulation of the Kir6.1 activity, stabilising it in a closed conformation. This hypothesis was supported by in vitro experiments demonstrating that Quercetin and 5-Hydroxyflavone were capable to inhibit K ATP currents of rat tail main artery myocytes recorded by the patch-clamp technique. Combined methodological approaches, such as molecular modelling, docking and MD simulations of Kir6.1 channel, used to elucidate flavonoids intrinsic mechanism of action, are introduced, revealing a new potential druggable protein site.

Dendritic small conductance calcium-activated potassium channels activated by action potentials suppress EPSPs and gate spike-timing dependent synaptic plasticity.

PubMed

Jones, Scott L; To, Minh-Son; Stuart, Greg J

2017-10-23

Small conductance calcium-activated potassium channels (SK channels) are present in spines and can be activated by backpropagating action potentials (APs). This suggests they may play a critical role in spike-timing dependent synaptic plasticity (STDP). Consistent with this idea, EPSPs in both cortical and hippocampal pyramidal neurons were suppressed by preceding APs in an SK-dependent manner. In cortical pyramidal neurons EPSP suppression by preceding APs depended on their precise timing as well as the distance of activated synapses from the soma, was dendritic in origin, and involved SK-dependent suppression of NMDA receptor activation. As a result SK channel activation by backpropagating APs gated STDP induction during low-frequency AP-EPSP pairing, with both LTP and LTD absent under control conditions but present after SK channel block. These findings indicate that activation of SK channels in spines by backpropagating APs plays a key role in regulating both EPSP amplitude and STDP induction.

Voltage-dependent gating of KCNH potassium channels lacking a covalent link between voltage-sensing and pore domains

PubMed Central

Lörinczi, Éva; Gómez-Posada, Juan Camilo; de la Peña, Pilar; Tomczak, Adam P.; Fernández-Trillo, Jorge; Leipscher, Ulrike; Stühmer, Walter; Barros, Francisco; Pardo, Luis A.

2015-01-01

Voltage-gated channels open paths for ion permeation upon changes in membrane potential, but how voltage changes are coupled to gating is not entirely understood. Two modules can be recognized in voltage-gated potassium channels, one responsible for voltage sensing (transmembrane segments S1 to S4), the other for permeation (S5 and S6). It is generally assumed that the conversion of a conformational change in the voltage sensor into channel gating occurs through the intracellular S4–S5 linker that provides physical continuity between the two regions. Using the pathophysiologically relevant KCNH family, we show that truncated proteins interrupted at, or lacking the S4–S5 linker produce voltage-gated channels in a heterologous model that recapitulate both the voltage-sensing and permeation properties of the complete protein. These observations indicate that voltage sensing by the S4 segment is transduced to the channel gate in the absence of physical continuity between the modules. PMID:25818916

Voltage-dependent gating of KCNH potassium channels lacking a covalent link between voltage-sensing and pore domains.

PubMed

Lörinczi, Éva; Gómez-Posada, Juan Camilo; de la Peña, Pilar; Tomczak, Adam P; Fernández-Trillo, Jorge; Leipscher, Ulrike; Stühmer, Walter; Barros, Francisco; Pardo, Luis A

2015-03-30

Voltage-gated channels open paths for ion permeation upon changes in membrane potential, but how voltage changes are coupled to gating is not entirely understood. Two modules can be recognized in voltage-gated potassium channels, one responsible for voltage sensing (transmembrane segments S1 to S4), the other for permeation (S5 and S6). It is generally assumed that the conversion of a conformational change in the voltage sensor into channel gating occurs through the intracellular S4-S5 linker that provides physical continuity between the two regions. Using the pathophysiologically relevant KCNH family, we show that truncated proteins interrupted at, or lacking the S4-S5 linker produce voltage-gated channels in a heterologous model that recapitulate both the voltage-sensing and permeation properties of the complete protein. These observations indicate that voltage sensing by the S4 segment is transduced to the channel gate in the absence of physical continuity between the modules.

Voltage-dependent gating of KCNH potassium channels lacking a covalent link between voltage-sensing and pore domains

NASA Astrophysics Data System (ADS)

Lörinczi, Éva; Gómez-Posada, Juan Camilo; de La Peña, Pilar; Tomczak, Adam P.; Fernández-Trillo, Jorge; Leipscher, Ulrike; Stühmer, Walter; Barros, Francisco; Pardo, Luis A.

2015-03-01

Voltage-gated channels open paths for ion permeation upon changes in membrane potential, but how voltage changes are coupled to gating is not entirely understood. Two modules can be recognized in voltage-gated potassium channels, one responsible for voltage sensing (transmembrane segments S1 to S4), the other for permeation (S5 and S6). It is generally assumed that the conversion of a conformational change in the voltage sensor into channel gating occurs through the intracellular S4-S5 linker that provides physical continuity between the two regions. Using the pathophysiologically relevant KCNH family, we show that truncated proteins interrupted at, or lacking the S4-S5 linker produce voltage-gated channels in a heterologous model that recapitulate both the voltage-sensing and permeation properties of the complete protein. These observations indicate that voltage sensing by the S4 segment is transduced to the channel gate in the absence of physical continuity between the modules.

PKA-induced internalization of slack KNa channels produces dorsal root ganglion neuron hyperexcitability.

PubMed

Nuwer, Megan O; Picchione, Kelly E; Bhattacharjee, Arin

2010-10-20

Inflammatory mediators through the activation of the protein kinase A (PKA) pathway sensitize primary afferent nociceptors to mechanical, thermal, and osmotic stimuli. However, it is unclear which ion conductances are responsible for PKA-induced nociceptor hyperexcitability. We have previously shown the abundant expression of Slack sodium-activated potassium (K(Na)) channels in nociceptive dorsal root ganglion (DRG) neurons. Here we show using cultured DRG neurons, that of the total potassium current, I(K), the K(Na) current is predominantly inhibited by PKA. We demonstrate that PKA modulation of K(Na) channels does not happen at the level of channel gating but arises from the internal trafficking of Slack channels from DRG membranes. Furthermore, we found that knocking down the Slack subunit by RNA interference causes a loss of firing accommodation analogous to that observed during PKA activation. Our data suggest that the change in nociceptive firing occurring during inflammation is the result of PKA-induced Slack channel trafficking.

BAD and KATP channels regulate neuron excitability and epileptiform activity.

PubMed

Martínez-François, Juan Ramón; Fernández-Agüera, María Carmen; Nathwani, Nidhi; Lahmann, Carolina; Burnham, Veronica L; Danial, Nika N; Yellen, Gary

2018-01-25

Brain metabolism can profoundly influence neuronal excitability. Mice with genetic deletion or alteration of Bad ( B CL-2 a gonist of cell d eath) exhibit altered brain-cell fuel metabolism, accompanied by resistance to acutely induced epileptic seizures; this seizure protection is mediated by ATP-sensitive potassium (K ATP ) channels. Here we investigated the effect of BAD manipulation on K ATP channel activity and excitability in acute brain slices. We found that BAD's influence on neuronal K ATP channels was cell-autonomous and directly affected dentate granule neuron (DGN) excitability. To investigate the role of neuronal K ATP channels in the anticonvulsant effects of BAD, we imaged calcium during picrotoxin-induced epileptiform activity in entorhinal-hippocampal slices. BAD knockout reduced epileptiform activity, and this effect was lost upon knockout or pharmacological inhibition of K ATP channels. Targeted BAD knockout in DGNs alone was sufficient for the antiseizure effect in slices, consistent with a 'dentate gate' function that is reinforced by increased K ATP channel activity. © 2018, Martínez-François et al.

Cortisone Dissociates the Shaker Family K Channels from their Beta Subunit

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pan, Y.; Weng, J; Kabaleeswaran, V

2008-01-01

The Shaker family voltage-dependent potassium channels (Kv1) are expressed in a wide variety of cells and are essential for cellular excitability. In humans, loss-of-function mutations of Kv1 channels lead to hyperexcitability and are directly linked to episodic ataxia and atrial fibrillation. All Kv1 channels assemble with {Beta} subunits (Kv{Beta}s), and certain Kv{Beta}s, for example Kv{Beta}1, have an N-terminal segment that closes the channel by the N-type inactivation mechanism. In principle, dissociation of Kv{Beta}1, although never reported, should eliminate inactivation and thus potentiate Kv1 current. We found that cortisone increases rat Kv1 channel activity by binding to Kv{Beta}1. A crystal structuremore » of the K{Beta}v-cortisone complex was solved to 1.82-{angstrom}resolution and revealed novel cortisone binding sites. Further studies demonstrated that cortisone promotes dissociation of Kv{Beta}. The new mode of channel modulation may be explored by native or synthetic ligands to fine-tune cellular excitability.« less

The epilepsy-linked Lgi1 protein assembles into presynaptic Kv1 channels and inhibits inactivation by Kvbeta1.

PubMed

Schulte, Uwe; Thumfart, Jörg-Oliver; Klöcker, Nikolaj; Sailer, Claudia A; Bildl, Wolfgang; Biniossek, Martin; Dehn, Doris; Deller, Thomas; Eble, Silke; Abbass, Karen; Wangler, Tanja; Knaus, Hans-Günther; Fakler, Bernd

2006-03-02

The voltage-gated potassium (Kv) channel subunit Kv1.1 is a major constituent of presynaptic A-type channels that modulate synaptic transmission in CNS neurons. Here, we show that Kv1.1-containing channels are complexed with Lgi1, the functionally unassigned product of the leucine-rich glioma inactivated gene 1 (LGI1), which is causative for an autosomal dominant form of lateral temporal lobe epilepsy (ADLTE). In the hippocampal formation, both Kv1.1 and Lgi1 are coassembled with Kv1.4 and Kvbeta1 in axonal terminals. In A-type channels composed of these subunits, Lgi1 selectively prevents N-type inactivation mediated by the Kvbeta1 subunit. In contrast, defective Lgi1 molecules identified in ADLTE patients fail to exert this effect resulting in channels with rapid inactivation kinetics. The results establish Lgi1 as a novel subunit of Kv1.1-associated protein complexes and suggest that changes in inactivation gating of presynaptic A-type channels may promote epileptic activity.

Regulation of Ion Gradients across Myocardial Ischemic Border Zones: A Biophysical Modelling Analysis

PubMed Central

Niederer, Steven

2013-01-01

The myocardial ischemic border zone is associated with the initiation and sustenance of arrhythmias. The profile of ionic concentrations across the border zone play a significant role in determining cellular electrophysiology and conductivity, yet their spatial-temporal evolution and regulation are not well understood. To investigate the changes in ion concentrations that regulate cellular electrophysiology, a mathematical model of ion movement in the intra and extracellular space in the presence of ionic, potential and material property heterogeneities was developed. The model simulates the spatial and temporal evolution of concentrations of potassium, sodium, chloride, calcium, hydrogen and bicarbonate ions and carbon dioxide across an ischemic border zone. Ischemia was simulated by sodium-potassium pump inhibition, potassium channel activation and respiratory and metabolic acidosis. The model predicted significant disparities in the width of the border zone for each ionic species, with intracellular sodium and extracellular potassium having discordant gradients, facilitating multiple gradients in cellular properties across the border zone. Extracellular potassium was found to have the largest border zone and this was attributed to the voltage dependence of the potassium channels. The model also predicted the efflux of from the ischemic region due to electrogenic drift and diffusion within the intra and extracellular space, respectively, which contributed to depletion in the ischemic region. PMID:23577101

Therapeutic targeting of two-pore-domain potassium (K(2P)) channels in the cardiovascular system.

PubMed

Wiedmann, Felix; Schmidt, Constanze; Lugenbiel, Patrick; Staudacher, Ingo; Rahm, Ann-Kathrin; Seyler, Claudia; Schweizer, Patrick A; Katus, Hugo A; Thomas, Dierk

2016-05-01

The improvement of treatment strategies in cardiovascular medicine is an ongoing process that requires constant optimization. The ability of a therapeutic intervention to prevent cardiovascular pathology largely depends on its capacity to suppress the underlying mechanisms. Attenuation or reversal of disease-specific pathways has emerged as a promising paradigm, providing a mechanistic rationale for patient-tailored therapy. Two-pore-domain K(+) (K(2P)) channels conduct outward K(+) currents that stabilize the resting membrane potential and facilitate action potential repolarization. K(2P) expression in the cardiovascular system and polymodal K2P current regulation suggest functional significance and potential therapeutic roles of the channels. Recent work has focused primarily on K(2P)1.1 [tandem of pore domains in a weak inwardly rectifying K(+) channel (TWIK)-1], K(2P)2.1 [TWIK-related K(+) channel (TREK)-1], and K(2P)3.1 [TWIK-related acid-sensitive K(+) channel (TASK)-1] channels and their role in heart and vessels. K(2P) currents have been implicated in atrial and ventricular arrhythmogenesis and in setting the vascular tone. Furthermore, the association of genetic alterations in K(2P)3.1 channels with atrial fibrillation, cardiac conduction disorders and pulmonary arterial hypertension demonstrates the relevance of the channels in cardiovascular disease. The function, regulation and clinical significance of cardiovascular K(2P) channels are summarized in the present review, and therapeutic options are emphasized. © 2016 Authors; published by Portland Press Limited.

Independent and cooperative motions of the Kv1.2 channel: voltage sensing and gating.

PubMed

Yeheskel, Adva; Haliloglu, Turkan; Ben-Tal, Nir

2010-05-19

Voltage-gated potassium (Kv) channels, such as Kv1.2, are involved in the generation and propagation of action potentials. The Kv channel is a homotetramer, and each monomer is composed of a voltage-sensing domain (VSD) and a pore domain (PD). We analyzed the fluctuations of a model structure of Kv1.2 using elastic network models. The analysis suggested a network of coupled fluctuations of eight rigid structural units and seven hinges that may control the transition between the active and inactive states of the channel. For the most part, the network is composed of amino acids that are known to affect channel activity. The results suggested allosteric interactions and cooperativity between the subunits in the coupling between the motion of the VSD and the selectivity filter of the PD, in accordance with recent empirical data. There are no direct contacts between the VSDs of the four subunits, and the contacts between these and the PDs are loose, suggesting that the VSDs are capable of functioning independently. Indeed, they manifest many inherent fluctuations that are decoupled from the rest of the structure. In general, the analysis suggests that the two domains contribute to the channel function both individually and cooperatively. Copyright 2010 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Expression and Stress-Dependent Induction of Potassium Channel Transcripts in the Common Ice Plant1

PubMed Central

Su, Hua; Golldack, Dortje; Katsuhara, Maki; Zhao, Chengsong; Bohnert, Hans J.

2001-01-01

We have characterized transcripts for three potassium channel homologs in the AKT/KAT subfamily (Shaker type) from the common ice plant (Mesembryanthemum crystallinum), with a focus on their expression during salt stress (up to 500 mm NaCl). Mkt1 and 2, Arabidopsis AKT homologs, and Kmt1, a KAT homolog, are members of small gene families with two to three isoforms each. Mkt1 is root specific; Mkt2 is found in leaves, flowers, and seed capsules; and Kmt1 is expressed in leaves and seed capsules. Mkt1 is present in all cells of the root, and in leaves a highly conserved isoform is detected present in all cells with highest abundance in the vasculature. MKT1 for which antibodies were made is localized to the plasma membrane. Following salt stress, MKT1 (transcripts and protein) is drastically down-regulated, Mkt2 transcripts do not change significantly, and Kmt1 is strongly and transiently (maximum at 6 h) up-regulated in leaves and stems. The detection and stress-dependent behavior of abundant transcripts representing subfamilies of potassium channels provides information about tissue specificity and the complex regulation of genes encoding potassium uptake systems in a halophytic plant. PMID:11161018

Recent advances in distal tubular potassium handling

PubMed Central

Rodan, Aylin R.; Cheng, Chih-Jen

2011-01-01

It is well known that sodium reabsorption and aldosterone play important roles in potassium secretion by the aldosterone-sensitive distal nephron. Sodium- and aldosterone-independent mechanisms also exist. This review focuses on some recent studies that provide novel insights into the sodium- and aldosterone-independent potassium secretion by the aldosterone-sensitive distal nephron. In addition, we discuss a study reporting on the regulation of the mammalian potassium kidney channel ROMK by intracellular and extracellular magnesium, which may be important in the pathogenesis of persistent hypokalemia in patients with concomitant potassium and magnesium deficiency. We also discuss outstanding questions and propose working models for future investigation. PMID:21270092

Calcium-activated potassium channels in basolateral membranes of colon epithelial cells; reconstitution and functional properties.

PubMed

Wiener, H; Turnheim, K

1990-10-26

Using differential sedimentation, isopycnic and Ficoll-400 barrier centrifugation, basolateral membrane vesicles of surface and crypt cells of the rabbit distal colon were enriched 34- and 9-fold, respectively. 86Rb(+)-uptake into these vesicles, driven by an electrical potential difference, was stimulated by submicromolar Ca2+ activities and inhibited by Ba2+. These findings indicate the presence of Ca2(+)-activated K+ channels. The K+ channels in surface and crypt cell membranes differed with respect to inhibition by the bee venom apamin, the scorpion venom charybdotoxin and tetraethylammonium and exhibited a different pH dependence. Fusion of basolateral membrane vesicles with planar phospholipid bilayers revealed the presence of high-conductance Ba2(+)-sensitive K+ channels which were activated by micromolar Ca2+ and inhibited by crude scorpion venom and trifluoperazine. These K+ channels may be involved in the coupling of apical and basolateral membrane conductances during Na+ absorption and Cl- secretion, but they may also play a role in cell volume regulation.

Hypothesized diprotomeric enzyme complex supported by stochastic modelling of palytoxin-induced Na/K pump channels

PubMed Central

Vilallonga, Gabriel D.; de Almeida, Antônio-Carlos G.; Ribeiro, Kelison T.; Campos, Sergio V. A.

2018-01-01

The sodium–potassium pump (Na+/K+ pump) is crucial for cell physiology. Despite great advances in the understanding of this ionic pumping system, its mechanism is not completely understood. We propose the use of a statistical model checker to investigate palytoxin (PTX)-induced Na+/K+ pump channels. We modelled a system of reactions representing transitions between the conformational substates of the channel with parameters, concentrations of the substates and reaction rates extracted from simulations reported in the literature, based on electrophysiological recordings in a whole-cell configuration. The model was implemented using the UPPAAL-SMC platform. Comparing simulations and probabilistic queries from stochastic system semantics with experimental data, it was possible to propose additional reactions to reproduce the single-channel dynamic. The probabilistic analyses and simulations suggest that the PTX-induced Na+/K+ pump channel functions as a diprotomeric complex in which protein–protein interactions increase the affinity of the Na+/K+ pump for PTX. PMID:29657808

Molecular dynamics simulations of a K+ channel blocker: Tc1 toxin from Tityus cambridgei.

PubMed

Grottesi, Alessandro; Sansom, Mark S P

2003-01-30

Toxins that block voltage-gated potassium (Kv) channels provide a possible template for improved homology models of the Kv pore. In assessing the interactions of Kv channels and their toxins it is important to determine the dynamic flexibility of the toxins. Multiple 10 ns duration molecular dynamics simulations combined with essential dynamics analysis have been used to explore the flexibility of four different Kv channel-blocking toxins. Three toxins (Tc1, AgTx and ChTx) share a common fold. They also share a common pattern of conformational dynamics, as revealed by essential dynamics analysis of the simulation results. This suggests that some aspects of dynamic behaviour are conserved across a single protein fold class. In each of these three toxins, the residue exhibiting minimum flexibility corresponds to a conserved lysine residue that is suggested to interact with the filter domain of the channel. Thus, comparative simulations reveal functionally important conservation of molecular dynamics as well as protein fold across a family of related toxins.

The isolated voltage sensing domain of the Shaker potassium channel forms a voltage-gated cation channel.

PubMed

Zhao, Juan; Blunck, Rikard

2016-10-06

Domains in macromolecular complexes are often considered structurally and functionally conserved while energetically coupled to each other. In the modular voltage-gated ion channels the central ion-conducting pore is surrounded by four voltage sensing domains (VSDs). Here, the energetic coupling is mediated by interactions between the S4-S5 linker, covalently linking the domains, and the proximal C-terminus. In order to characterize the intrinsic gating of the voltage sensing domain in the absence of the pore domain, the Shaker Kv channel was truncated after the fourth transmembrane helix S4 (Shaker-iVSD). Shaker-iVSD showed significantly altered gating kinetics and formed a cation-selective ion channel with a strong preference for protons. Ion conduction in Shaker-iVSD developed despite identical primary sequence, indicating an allosteric influence of the pore domain. Shaker-iVSD also displays pronounced 'relaxation'. Closing of the pore correlates with entry into relaxation suggesting that the two processes are energetically related.

The voltage-sensing domain of a phosphatase gates the pore of a potassium channel

PubMed Central

Arrigoni, Cristina; Schroeder, Indra; Romani, Giulia; Van Etten, James L.; Thiel, Gerhard

2013-01-01

The modular architecture of voltage-gated K+ (Kv) channels suggests that they resulted from the fusion of a voltage-sensing domain (VSD) to a pore module. Here, we show that the VSD of Ciona intestinalis phosphatase (Ci-VSP) fused to the viral channel Kcv creates KvSynth1, a functional voltage-gated, outwardly rectifying K+ channel. KvSynth1 displays the summed features of its individual components: pore properties of Kcv (selectivity and filter gating) and voltage dependence of Ci-VSP (V1/2 = +56 mV; z of ∼1), including the depolarization-induced mode shift. The degree of outward rectification of the channel is critically dependent on the length of the linker more than on its amino acid composition. This highlights a mechanistic role of the linker in transmitting the movement of the sensor to the pore and shows that electromechanical coupling can occur without coevolution of the two domains. PMID:23440279

A Cyclic Nucleotide-Gated Channel Mutation Associated with Canine Daylight Blindness Provides Insight into a Role for the S2 Segment Tri-Asp motif in Channel Biogenesis

PubMed Central

Tanaka, Naoto; Delemotte, Lucie; Klein, Michael L.; Komáromy, András M.; Tanaka, Jacqueline C.

2014-01-01

Cone cyclic nucleotide-gated channels are tetramers formed by CNGA3 and CNGB3 subunits; CNGA3 subunits function as homotetrameric channels but CNGB3 exhibits channel function only when co-expressed with CNGA3. An aspartatic acid (Asp) to asparagine (Asn) missense mutation at position 262 in the canine CNGB3 (D262N) subunit results in loss of cone function (daylight blindness), suggesting an important role for this aspartic acid residue in channel biogenesis and/or function. Asp 262 is located in a conserved region of the second transmembrane segment containing three Asp residues designated the Tri-Asp motif. This motif is conserved in all CNG channels. Here we examine mutations in canine CNGA3 homomeric channels using a combination of experimental and computational approaches. Mutations of these conserved Asp residues result in the absence of nucleotide-activated currents in heterologous expression. A fluorescent tag on CNGA3 shows mislocalization of mutant channels. Co-expressing CNGB3 Tri-Asp mutants with wild type CNGA3 results in some functional channels, however, their electrophysiological characterization matches the properties of homomeric CNGA3 channels. This failure to record heteromeric currents suggests that Asp/Asn mutations affect heteromeric subunit assembly. A homology model of S1–S6 of the CNGA3 channel was generated and relaxed in a membrane using molecular dynamics simulations. The model predicts that the Tri-Asp motif is involved in non-specific salt bridge pairings with positive residues of S3/S4. We propose that the D262N mutation in dogs with CNGB3-day blindness results in the loss of these inter-helical interactions altering the electrostatic equilibrium within in the S1–S4 bundle. Because residues analogous to Tri-Asp in the voltage-gated Shaker potassium channel family were implicated in monomer folding, we hypothesize that destabilizing these electrostatic interactions impairs the monomer folding state in D262N mutant CNG channels during biogenesis. PMID:24586388

β-adrenergic Receptor Blocker ICI 118,551 Selectively Increases Intermediate-Conductance Calcium-Activated Potassium Channel (IKCa )-Mediated Relaxations in Rat Main Mesenteric Artery.

PubMed

Ozkan, Melike Hacer; Uma, Serdar

2018-06-01

Endothelial IK C a and/or SK C a channels play an important role in the control of vascular tone by participating in endothelium-dependent relaxation. Whether β-AR antagonists, mainly used in hypertension, affect endothelial K C a channel function is unknown. In this study, we examined the effect of the β2-AR antagonist and inverse agonist ICI 118,551 on the IK C a /SK C a channel activity by assessing functional relaxation responses to several agonists that stimulate these channels. Mesenteric arterial rings isolated from male Sprague Dawley mounted to organ baths. Acetylcholine elicited IK C a - and SK C a -mediated relaxations that were abolished by TRAM-34 and apamin, respectively. ICI 118,551, which did not dilate the arteries per se, increased the IK C a -mediated relaxations, whereas SK C a -mediated relaxations remained unaltered. Same potentiating effect was also detected on the IK C a -mediated relaxations to carbachol and A23187, but not to NS309. Neither acetylcholine-induced nitric oxide-mediated relaxations nor SNP relaxations changed with ICI 118,551. The PKA inhibitor KT-5720, the selective β2-AR agonist salbutamol, the selective β2-AR antagonist butoxamine, the non-selective β-AR antagonist propranolol, and the inverse agonists carvedilol or nadolol failed to affect the IK C a -mediated relaxations. ICI 118,551-induced increase was not reversed by salbutamol or propranolol as well. Besides, low potassium-induced relaxations in endothelium-removed arteries remained the same in the presence of ICI 118,551. These data demonstrate a previously unrecognized action of ICI 118,551, the ability to potentiate endothelial IK C a channel-mediated vasodilation, through a mechanism independent of β2-AR antagonistic or inverse agonistic action. Instead, the enhancement of acetylcholine relaxation seems likely to occur by a mechanism secondary to endothelial calcium increase. © 2017 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Do defects in ion channel glycosylation set the stage for lethal cardiac arrhythmias?

PubMed

Fozzard, Harry A; Kyle, John W

2002-04-30

Many ion channels are modified by the addition of carbohydrate residues. Fozzard and Kyle discuss evidence that sialic acid residues on glycosylated cardiac sodium and potassium channels may be important for preventing early after-depolarizations that can result in cardiac arrhythmias.

Quantum mechanical calculations of charge effects on gating the KcsA channel.

PubMed

Kariev, Alisher M; Znamenskiy, Vasiliy S; Green, Michael E

2007-05-01

A series of ab initio (density functional) calculations were carried out on side chains of a set of amino acids, plus water, from the (intracellular) gating region of the KcsA K(+) channel. Their atomic coordinates, except hydrogen, are known from X-ray structures [D.A. Doyle, J.M. Cabral, R.A. Pfuetzner, A. Kuo, J.M. Gulbis, S.L. Cohen, B.T. Chait, R. MacKinnon, The structure of the potassium channel: molecular basis of K(+) conduction and selectivity, Science 280 (1998) 69-77; R. MacKinnon, S.L. Cohen, A. Kuo, A. Lee, B.T. Chait, Structural conservation in prokaryotic and eukaryotic potassium channels, Science 280 (1998) 106-109; Y. Jiang, A. Lee, J. Chen, M. Cadene, B.T. Chait, R. MacKinnon, The open pore conformation of potassium channels. Nature 417 (2001) 523-526], as are the coordinates of some water oxygen atoms. The 1k4c structure is used for the starting coordinates. Quantum mechanical optimization, in spite of the starting configuration, places the atoms in positions much closer to the 1j95, more tightly closed, configuration. This state shows four water molecules forming a "basket" under the Q119 side chains, blocking the channel. When a hydrated K(+) approaches this "basket", the optimized system shows a strong set of hydrogen bonds with the K(+) at defined positions, preventing further approach of the K(+) to the basket. This optimized structure with hydrated K(+) added shows an ice-like 12 molecule nanocrystal of water. If the water molecules exchange, unless they do it as a group, the channel will remain blocked. The "basket" itself appears to be very stable, although it is possible that the K(+) with its hydrating water molecules may be more mobile, capable of withdrawing from the gate. It is also not surprising that water essentially freezes, or forms a kind of glue, in a nanometer space; this agrees with experimental results on a rather different, but similarly sized (nm dimensions) system [K.B. Jinesh, J.W.M. Frenken, Capillary condensation in atomic scale friction: how water acts like a glue, Phys. Rev. Lett. 96 (2006) 166103/1-4]. It also agrees qualitatively with simulations on channels [A. Anishkin, S. Sukharev, Water dynamics and dewetting transitions in the small mechanosensitive channel MscS, Biophys. J. 86 (2004) 2883-2895; O. Beckstein, M.S.P. Sansom, Liquid-vapor oscillations of water in hydrophobic nanopores, Proc. Natl Acad. Sci. U. S. A. 100 (2003) 7063-7068] and on featureless channel-like systems [J. Lu, M.E. Green, Simulation of water in a pore with charges: application to a gating mechanism for ion channels, Prog. Colloid Polym. Sci. 103 (1997) 121-129], in that it forms a boundary on water that is not obvious from the liquid state. The idea that a structure is stable, even if individual molecules exchange, is well known, for example from the hydration shell of ions. We show that when charges are added in the form of protons to the domains (one proton per domain), the optimized structure is open. No stable water hydrogen bonds hold it together; an opening of 11.0 A appears, measured diagonally between non-neighboring domains as glutamine 119 carbonyl O-O distance. This is comparable to the opening in the MthK potassium channel structure that is generally agreed to be open. The appearance of the opening is in rather good agreement with that found by Perozo and coworkers. In contrast, in the uncharged structure this diagonal distance is 6.5 A, and the water "basket" constricts the uncharged opening still further, with the ice-like structure that couples the K(+) ion to the gating region freezing the entrance to the channel. Comparison with our earlier model for voltage gated channels suggests that a similar mechanism may apply in those channels.

Small-Conductance Ca2+-Activated Potassium Channels Negatively Regulate Aldosterone Secretion in Human Adrenocortical Cells.

PubMed

Yang, Tingting; Zhang, Hai-Liang; Liang, Qingnan; Shi, Yingtang; Mei, Yan-Ai; Barrett, Paula Q; Hu, Changlong

2016-09-01

Aldosterone, which plays a key role in maintaining water and electrolyte balance, is produced by zona glomerulosa cells of the adrenal cortex. Autonomous overproduction of aldosterone from zona glomerulosa cells causes primary hyperaldosteronism. Recent clinical studies have highlighted the pathological role of the KCNJ5 potassium channel in primary hyperaldosteronism. Our objective was to determine whether small-conductance Ca(2+)-activated potassium (SK) channels may also regulate aldosterone secretion in human adrenocortical cells. We found that apamin, the prototypic inhibitor of SK channels, decreased membrane voltage, raised intracellular Ca(2+) and dose dependently increased aldosterone secretion from human adrenocortical H295R cells. By contrast, 1-Ethyl-2-benzimidazolinone, an agonist of SK channels, antagonized apamin's action and decreased aldosterone secretion. Commensurate with an increase in aldosterone production, apamin increased mRNA expression of steroidogenic acute regulatory protein and aldosterone synthase that control the early and late rate-limiting steps in aldosterone biosynthesis, respectively. In addition, apamin increased angiotensin II-stimulated aldosterone secretion, whereas 1-Ethyl-2-benzimidazolinone suppressed both angiotensin II- and high K(+)-stimulated production of aldosterone in H295R cells. These findings were supported by apamin-modulation of basal and angiotensin II-stimulated aldosterone secretion from acutely prepared slices of human adrenals. We conclude that SK channel activity negatively regulates aldosterone secretion in human adrenocortical cells. Genetic association studies are necessary to determine whether mutations in SK channel subtype 2 genes may also drive aldosterone excess in primary hyperaldosteronism. © 2016 American Heart Association, Inc.

An improved synthesis of pyran-3,5-dione: application to the synthesis of ABT-598, a potassium channel opener, via Hantzsch reaction.

PubMed

Li, Wenke; Wayne, Gregory S; Lallaman, John E; Chang, Sou-Jen; Wittenberger, Steven J

2006-02-17

Ketoester 1 is cyclized to give pyran-3,5-dione 2 in 78% yield using a parallel addition of ketoester 1 and base NaO(t)Bu in refluxing THF. Compared to the previously reported procedures, these optimized conditions have significantly increased the yield of this transformation and the quality of pyran 2 and prove to be suitable for large-scale preparation. An application of 2 to the synthesis of ABT-598, a potassium channel opener, is demonstrated.

Targeting solid tumours with potassium channel activators. A return to fundamentals?

PubMed

Trechot, Philippe

2014-01-01

From a pharmacological point of view nicotinamide and minoxidil are potassium channel activators. Nicotinamide is used as a radiosensitizer in ARCON (accelerated radiotherapy combined with carbogen breathing and nicotinamide) therapeutic strategy with promising results but not confirmed so far. Minoxidil has never been considered by radiotherapists. Based from recent pathophysiological considerations we suggest a new perspective for the use of these two "old" molecules in order to target solid tumours. © 2014 Société Française de Pharmacologie et de Thérapeutique.