Two-pore channels at the intersection of endolysosomal membrane traffic

PubMed Central

Marchant, Jonathan S.; Patel, Sandip

2016-01-01

Two-pore channels (TPCs) are ancient members of the voltage-gated ion channel superfamily that localize to acidic organelles such as lysosomes. The TPC complex is the proposed target of the Ca2 +-mobilizing messenger NAADP, which releases Ca2 + from these acidic Ca2 + stores. Whereas details of TPC activation and native ion permeation remain unclear, a consensus has emerged around their function in regulating endolysosomal trafficking. This role is supported by recent proteomic data showing that TPCs interact with proteins controlling membrane organization and dynamics, including Rab GTPases and components of the fusion apparatus. Regulation of TPCs by PtdIns(3,5)P2 and/or NAADP (nicotinic acid adenine dinucleotide phosphate) together with their functional and physical association with Rab proteins provides a mechanism for coupling phosphoinositide and trafficking protein cues to local ion fluxes. Therefore, TPCs work at the regulatory cross-roads of (patho)physiological cues to co-ordinate and potentially deregulate traffic flow through the endolysosomal network. This review focuses on the native role of TPCs in trafficking and their emerging contributions to endolysosomal trafficking dysfunction. PMID:26009187

ASIC3 channels in multimodal sensory perception.

PubMed

Li, Wei-Guang; Xu, Tian-Le

2011-01-19

Acid-sensing ion channels (ASICs), which are members of the sodium-selective cation channels belonging to the epithelial sodium channel/degenerin (ENaC/DEG) family, act as membrane-bound receptors for extracellular protons as well as nonproton ligands. At least five ASIC subunits have been identified in mammalian neurons, which form both homotrimeric and heterotrimeric channels. The highly proton sensitive ASIC3 channels are predominantly distributed in peripheral sensory neurons, correlating with their roles in multimodal sensory perception, including nociception, mechanosensation, and chemosensation. Different from other ASIC subunit composing ion channels, ASIC3 channels can mediate a sustained window current in response to mild extracellular acidosis (pH 7.3-6.7), which often occurs accompanied by many sensory stimuli. Furthermore, recent evidence indicates that the sustained component of ASIC3 currents can be enhanced by nonproton ligands including the endogenous metabolite agmatine. In this review, we first summarize the growing body of evidence for the involvement of ASIC3 channels in multimodal sensory perception and then discuss the potential mechanisms underlying ASIC3 activation and mediation of sensory perception, with a special emphasis on its role in nociception. We conclude that ASIC3 activation and modulation by diverse sensory stimuli represent a new avenue for understanding the role of ASIC3 channels in sensory perception. Furthermore, the emerging implications of ASIC3 channels in multiple sensory dysfunctions including nociception allow the development of new pharmacotherapy.

Ion channels in EEG: isolating channel dysfunction in NMDA receptor antibody encephalitis.

PubMed

Symmonds, Mkael; Moran, Catherine H; Leite, M Isabel; Buckley, Camilla; Irani, Sarosh R; Stephan, Klaas Enno; Friston, Karl J; Moran, Rosalyn J

2018-06-01

See Roberts and Breakspear (doi:10.1093/brain/awy136) for a scientific commentary on this article.Neurological and psychiatric practice frequently lack diagnostic probes that can assess mechanisms of neuronal communication non-invasively in humans. In N-methyl-d-aspartate (NMDA) receptor antibody encephalitis, functional molecular assays are particularly important given the presence of NMDA antibodies in healthy populations, the multifarious symptomology and the lack of radiological signs. Recent advances in biophysical modelling techniques suggest that inferring cellular-level properties of neural circuits from macroscopic measures of brain activity is possible. Here, we estimated receptor function from EEG in patients with NMDA receptor antibody encephalitis (n = 29) as well as from encephalopathic and neurological patient controls (n = 36). We show that the autoimmune patients exhibit distinct fronto-parietal network changes from which ion channel estimates can be obtained using a microcircuit model. Specifically, a dynamic causal model of EEG data applied to spontaneous brain responses identifies a selective deficit in signalling at NMDA receptors in patients with NMDA receptor antibody encephalitis but not at other ionotropic receptors. Moreover, though these changes are observed across brain regions, these effects predominate at the NMDA receptors of excitatory neurons rather than at inhibitory interneurons. Given that EEG is a ubiquitously available clinical method, our findings suggest a unique re-purposing of EEG data as an assay of brain network dysfunction at the molecular level.

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.

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

Tau-Dependent Kv4.2 Depletion and Dendritic Hyperexcitability in a Mouse Model of Alzheimer's Disease

PubMed Central

Hall, Alicia M.; Throesch, Benjamin T.; Buckingham, Susan C.; Markwardt, Sean J.; Peng, Yin; Wang, Qin

2015-01-01

Neuronal hyperexcitability occurs early in the pathogenesis of Alzheimer's disease (AD) and contributes to network dysfunction in AD patients. In other disorders with neuronal hyperexcitability, dysfunction in the dendrites often contributes, but dendritic excitability has not been directly examined in AD models. We used dendritic patch-clamp recordings to measure dendritic excitability in the CA1 region of the hippocampus. We found that dendrites, more so than somata, of hippocampal neurons were hyperexcitable in mice overexpressing Aβ. This dendritic hyperexcitability was associated with depletion of Kv4.2, a dendritic potassium channel important for regulating dendritic excitability and synaptic plasticity. The antiepileptic drug, levetiracetam, blocked Kv4.2 depletion. Tau was required, as crossing with tau knock-out mice also prevented both Kv4.2 depletion and dendritic hyperexcitability. Dendritic hyperexcitability induced by Kv4.2 deficiency exacerbated behavioral deficits and increased epileptiform activity in hAPP mice. We conclude that increased dendritic excitability, associated with changes in dendritic ion channels including Kv4.2, may contribute to neuronal dysfunction in early stages AD. PMID:25878292

Lysosomal Physiology

PubMed Central

Xu, Haoxing; Ren, Dejian

2015-01-01

Lysosomes are acidic compartments filled with more than 60 different types of hydrolases. They mediate the degradation of extracellular particles from endocytosis and of intracellular components from autophagy. The digested products are transported out of the lysosome via specific catabolite exporters or via vesicular membrane trafficking. Lysosomes also contain more than 50 membrane proteins and are equipped with the machinery to sense nutrient availability, which determines the distribution, number, size, and activity of lysosomes to control the specificity of cargo flux and timing (the initiation and termination) of degradation. Defects in degradation, export, or trafficking result in lysosomal dysfunction and lysosomal storage diseases (LSDs). Lysosomal channels and transporters mediate ion flux across perimeter membranes to regulate lysosomal ion homeostasis, membrane potential, catabolite export, membrane trafficking, and nutrient sensing. Dysregulation of lysosomal channels underlies the pathogenesis of many LSDs and possibly that of metabolic and common neurodegenerative diseases. PMID:25668017

Genetic autonomic disorders.

PubMed

Axelrod, Felicia B

2013-03-01

Genetic disorders affecting the autonomic nervous system can result in abnormal development of the nervous system or they can be caused by neurotransmitter imbalance, an ion-channel disturbance or by storage of deleterious material. The symptoms indicating autonomic dysfunction, however, will depend upon whether the genetic lesion has disrupted peripheral or central autonomic centers or both. Because the autonomic nervous system is pervasive and affects every organ system in the body, autonomic dysfunction will result in impaired homeostasis and symptoms will vary. The possibility of genetic confirmation by molecular testing for specific diagnosis is increasing but treatments tend to remain only supportive and directed toward particular symptoms. Copyright © 2013 Elsevier Inc. All rights reserved.

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

EFFECTS OF METHYLMERCURY ON SPINAL CORD AFFERENTS AND EFFERENTS—A REVIEW

PubMed Central

Colón-Rodríguez, Alexandra; Hannon, Heidi E.; Atchison, William D.

2017-01-01

Methylmercury (MeHg) is an environmental neurotoxicant of public health concern. It readily accumulates in exposed humans, primarily in neuronal tissue. Exposure to MeHg, either acutely or chronically, causes severe neuronal dysfunction in the central nervous system and spinal neurons; dysfunction of susceptible neuronal populations results in neurodegeneration, at least in part through Ca2+-mediated pathways. Biochemical and morphologic changes in peripheral neurons precede those in central brain regions, despite the fact that MeHg readily crosses the blood-brain barrier. Consequently, it is suggested that unique characteristics of spinal cord afferents and efferents could heighten their susceptibility to MeHg toxicity. Transient receptor potential (TRP) ion channels are a class of Ca2+-permeable cation channels that are highly expressed in spinal afferents, among other sensory and visceral organs. These channels can be activated in numerous ways, including directly via chemical irritants or indirectly via Ca2+ release from intracellular storage organelles. Early studies demonstrated that MeHg interacts with heterologous TRPs, though definitive mechanisms of MeHg toxicity on sensory neurons may involve more complex interaction with, and among, differentially-expressed TRP populations. In spinal efferents, glutamate receptors of the N-methyl-D-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), and possibly kainic acid (KA) classes are thought to play a major role in MeHg-induced neurotoxicity. Specifically, the Ca2+-permeable AMPA receptors, which are abundant in motor neurons, have been identified as being involved in MeHg-induced neurotoxicity. In this review, we will describe the mechanisms that could contribute to MeHg-induced spinal cord afferent and efferent neuronal degeneration, including the possible mediators, such as uniquely expressed Ca2+-permeable ion channels. PMID:28041893

Myokymia and neuromyotonia in veterinary medicine: a comparison with peripheral nerve hyperexcitability syndrome in humans.

PubMed

Vanhaesebrouck, An E; Bhatti, Sofie F M; Franklin, Robin J M; Van Ham, Luc

2013-08-01

Involuntary muscle hyperactivity can result from muscle or peripheral nerve hyperexcitability or central nervous system dysfunction. In humans, diseases causing hyperexcitability of peripheral nerves are grouped together under the term 'peripheral nerve hyperexcitability' (PNH). Hyperexcitability of the peripheral motor nerve can result into five different phenotypic main variants, i.e. fasciculations, myokymia, neuromyotonia, cramps and tetany, each with their own clinical and electromyographic characteristics. This review focuses on the most commonly described expressions of PNH in veterinary medicine, i.e. myokymia and neuromyotonia, in particular in young Jack Russell terriers. Data from 58 veterinary cases with generalized myokymia and neuromyotonia were analyzed, including unpublished treatment and follow-up data on eight Jack Russell terriers from a previous study and seven additional Jack Russell terriers. A dysfunction of the potassium channel or its associated proteins has been found in many human syndromes characterized by PNH, in particular in generalized myokymia and neuromyotonia, and is suspected to occur in veterinary medicine. Potential pathomechanisms of potassium channel dysfunction leading to signs of PNH are broad and include genetic mutations, antibody-mediated attack or ion channel maldistribution due to axonal degeneration or demyelination. A more accurate classification of the different PNH syndromes will facilitate a more rapid diagnosis and guide further research into natural occurring PNH in animals. Copyright © 2013 Elsevier Ltd. All rights reserved.

From Understanding Cellular Function to Novel Drug Discovery: The Role of Planar Patch-Clamp Array Chip Technology

PubMed Central

Py, Christophe; Martina, Marzia; Diaz-Quijada, Gerardo A.; Luk, Collin C.; Martinez, Dolores; Denhoff, Mike W.; Charrier, Anne; Comas, Tanya; Monette, Robert; Krantis, Anthony; Syed, Naweed I.; Mealing, Geoffrey A. R.

2011-01-01

All excitable cell functions rely upon ion channels that are embedded in their plasma membrane. Perturbations of ion channel structure or function result in pathologies ranging from cardiac dysfunction to neurodegenerative disorders. Consequently, to understand the functions of excitable cells and to remedy their pathophysiology, it is important to understand the ion channel functions under various experimental conditions – including exposure to novel drug targets. Glass pipette patch-clamp is the state of the art technique to monitor the intrinsic and synaptic properties of neurons. However, this technique is labor intensive and has low data throughput. Planar patch-clamp chips, integrated into automated systems, offer high throughputs but are limited to isolated cells from suspensions, thus limiting their use in modeling physiological function. These chips are therefore not most suitable for studies involving neuronal communication. Multielectrode arrays (MEAs), in contrast, have the ability to monitor network activity by measuring local field potentials from multiple extracellular sites, but specific ion channel activity is challenging to extract from these multiplexed signals. Here we describe a novel planar patch-clamp chip technology that enables the simultaneous high-resolution electrophysiological interrogation of individual neurons at multiple sites in synaptically connected neuronal networks, thereby combining the advantages of MEA and patch-clamp techniques. Each neuron can be probed through an aperture that connects to a dedicated subterranean microfluidic channel. Neurons growing in networks are aligned to the apertures by physisorbed or chemisorbed chemical cues. In this review, we describe the design and fabrication process of these chips, approaches to chemical patterning for cell placement, and present physiological data from cultured neuronal cells. PMID:22007170

From understanding cellular function to novel drug discovery: the role of planar patch-clamp array chip technology.

PubMed

Py, Christophe; Martina, Marzia; Diaz-Quijada, Gerardo A; Luk, Collin C; Martinez, Dolores; Denhoff, Mike W; Charrier, Anne; Comas, Tanya; Monette, Robert; Krantis, Anthony; Syed, Naweed I; Mealing, Geoffrey A R

2011-01-01

All excitable cell functions rely upon ion channels that are embedded in their plasma membrane. Perturbations of ion channel structure or function result in pathologies ranging from cardiac dysfunction to neurodegenerative disorders. Consequently, to understand the functions of excitable cells and to remedy their pathophysiology, it is important to understand the ion channel functions under various experimental conditions - including exposure to novel drug targets. Glass pipette patch-clamp is the state of the art technique to monitor the intrinsic and synaptic properties of neurons. However, this technique is labor intensive and has low data throughput. Planar patch-clamp chips, integrated into automated systems, offer high throughputs but are limited to isolated cells from suspensions, thus limiting their use in modeling physiological function. These chips are therefore not most suitable for studies involving neuronal communication. Multielectrode arrays (MEAs), in contrast, have the ability to monitor network activity by measuring local field potentials from multiple extracellular sites, but specific ion channel activity is challenging to extract from these multiplexed signals. Here we describe a novel planar patch-clamp chip technology that enables the simultaneous high-resolution electrophysiological interrogation of individual neurons at multiple sites in synaptically connected neuronal networks, thereby combining the advantages of MEA and patch-clamp techniques. Each neuron can be probed through an aperture that connects to a dedicated subterranean microfluidic channel. Neurons growing in networks are aligned to the apertures by physisorbed or chemisorbed chemical cues. In this review, we describe the design and fabrication process of these chips, approaches to chemical patterning for cell placement, and present physiological data from cultured neuronal cells.

Loss of Ca2+-mediated ion transport during colitis correlates with reduced ion transport responses to a Ca2+-activated K+ channel opener

PubMed Central

Hirota, Christina L; McKay, Derek M

2009-01-01

Background and purpose: Epithelial surface hydration is critical for proper gut function. However, colonic tissues from individuals with inflammatory bowel disease or animals with colitis are hyporesponsive to Cl− secretagogues. The Cl− secretory responses to the muscarinic receptor agonist bethanechol are virtually absent in colons of mice with dextran sodium sulphate (DSS)-induced colitis. Our aim was to define the mechanism underlying this cholinergic hyporesponsiveness. Experimental approach: Colitis was induced by 4% DSS water, given orally. Epithelial ion transport was measured in Ussing chambers. Colonic crypts were isolated and processed for mRNA expression via RT-PCR and protein expression via immunoblotting and immunolocalization. Key results: Expression of muscarinic M3 receptors in colonic epithelium was not decreased during colitis. Short-circuit current (ISC) responses to other Ca2+-dependent secretagogues (histamine, thapsigargin, cyclopiazonic acid and calcium ionophore) were either absent or severely attenuated in colonic tissue from DSS-treated mice. mRNA levels of several ion transport molecules (a Ca2+-regulated Cl− channel, the intermediate-conductance Ca2+-activated K+ channel, the cystic fibrosis transmembrane conductance regulator, the Na+/K+-ATPase pump or the Na+/K+/2Cl− co-transporter) were not reduced in colonic crypts from DSS-treated mice. However, protein expression of Na+/K+-ATPase α1 subunits was decreased twofold during colitis. Activation of Ca2+-activated K+ channels increased ISC significantly less in DSS colons compared with control, as did the protein kinase C activator, phorbol 12-myristate 13-acetate. Conclusions and implications: Decreased Na+/K+-ATPase expression probably contributes to overall epithelial hyporesponsiveness during colitis, while dysfunctional K+ channels may account, at least partially, for lack of epithelial secretory responses to Ca2+-mediated secretagogues. PMID:19298254

Astrocyte dysfunction in temporal lobe epilepsy: K+ channels and gap junction coupling.

PubMed

Steinhäuser, Christian; Seifert, Gerald; Bedner, Peter

2012-08-01

Astrocytes are endowed with the machinery to sense and respond to neuronal activity. Recent work has demonstrated that astrocytes play important physiological roles in the CNS, e.g., they synchronize action potential firing, ensure ion homeostasis, transmitter clearance and glucose metabolism, and regulate the vascular tone. Astrocytes are abundantly coupled through gap junctions, which is a prerequisite to redistribute elevated K(+) from sites of excessive neuronal activity to sites of lower extracellular K(+) concentration. Recent studies identified dysfunctional astrocytes as crucial players in epilepsy. Investigation of specimens from patients with pharmacoresistant temporal lobe epilepsy and epilepsy models revealed alterations in expression, localization, and function of astroglial inwardly rectifying K(+) (Kir) channels, particularly Kir4.1, which is suspected to entail impaired K(+) buffering. Gap junctions in astrocytes appear to play a dual role: on the one hand they counteract the generation of hyperactivity by facilitating clearance of elevated extracellular K(+) levels while in contrast, they constitute a pathway for energetic substrate delivery to fuel neuronal (hyper)activity. Recent work suggests that astrocyte dysfunction is causative of the generation or spread of seizure activity. Thus, astrocytes should be considered as promising targets for alternative antiepileptic therapies. Copyright © 2012 Wiley Periodicals, Inc.

New Strategies to Develop Novel Pain Therapies: Addressing Thermoreceptors from Different Points of View

PubMed Central

Fernández-Carvajal, Asia; Fernández-Ballester, Gregorio; Devesa, Isabel; González-Ros, José Manuel; Ferrer-Montiel, Antonio

2011-01-01

One approach to develop successful pain therapies is the modulation of dysfunctional ion channels that contribute to the detection of thermal, mechanical and chemical painful stimuli. These ion channels, known as thermoTRPs, promote the sensitization and activation of primary sensory neurons known as nociceptors. Pharmacological blockade and genetic deletion of thermoTRP have validated these channels as therapeutic targets for pain intervention. Several thermoTRP modulators have progressed towards clinical development, although most failed because of the appearance of unpredicted side effects. Thus, there is yet a need to develop novel channel modulators with improved therapeutic index. Here, we review the current state-of-the art and illustrate new pharmacological paradigms based on TRPV1 that include: (i) the identification of activity-dependent modulators of this thermoTRP channel; (ii) the design of allosteric modulators that interfere with protein-protein interaction involved in the functional coupling of stimulus sensing and gate opening; and (iii) the development of compounds that abrogate the inflammation-mediated increase of receptor expression in the neuronal surface. These new sites of action represent novel strategies to modulate pathologically active TRPV1, while minimizing an effect on the TRPV1 subpopulation involved in physiological and protective roles, thus increasing their potential therapeutic use. PMID:24288041

Novel bandlike signal abnormality suggestive of heterotopia in patient with a KCNQ1 frameshift mutation.

PubMed

Sabharwal, Priyanka; Devinsky, Orrin; M Shepherd, Timothy

2017-12-01

Malformations of cortical development are associated with epilepsy and cognitive dysfunction, and can occur in patients with SCN1A ion channel mutations. We report a novel and subtle bandlike subcortical heterotopia on integrated positron emission tomography-magnetic resonance imaging ( PET-MRI) in a patient with treatment-resistant epilepsy due to a de novo KCNQ1 frameshift mutation. Our case highlights the potential for other channel mutations to cause both epilepsy and cortical malformations. Further scrutiny of high contrast resolution MRI studies is warranted for patients with KCNQ1 and other epilepsy genes to further define their extended phenotype.

The regulation of transient receptor potential canonical 4 (TRPC4) channel by phosphodiesterase 5 inhibitor via the cyclic guanosine 3'5'-monophosphate.

PubMed

Wie, Jinhong; Jeong, SeungJoo; Kwak, Misun; Myeong, Jongyun; Chae, MeeRee; Park, Jong Kwan; Lee, Sung Won; So, Insuk

2017-06-01

The transient receptor potential (TRP) protein superfamily consists of a diverse group of cation channels that bear structural similarities to the fruit fly Drosophila TRP. The TRP superfamily is distinct from other groups of ion channels in displaying a large diversity in ion selectivity, modes of activation, and physiological functions. Classical TRP (transient receptor potential canonical (TRPC)) channels are activated by stimulation of Gq-PLC-coupled receptors and modulated by phosphorylation. The cyclic guanosine monophosphate (cGMP)-PKG pathway is involved in the regulation of TRPC3 and TRPC6 channels. Phosphodiesterase (PDE) 5 inhibitor induced muscle relaxation in corporal smooth muscle cells and was used to treat erectile dysfunction by inhibiting cGMP degradation. Here, we report the functional relationship between TRPC4 and cGMP. In human embryonic kidney (HEK) 293 cells overexpressing TRPC4, cGMP selectively activated TRPC4 channels and increased cytosolic calcium level through TRPC4 channel. We investigated phosphorylation sites in TRPC4 channels and identified S688 as an important phosphorylation site for the cGMP-PKG pathway. Cyclic GMP also activated TRPC4-like current with doubly rectifying current-voltage relationship in prostate smooth muscle cell lines. Taken together, these results show that TRPC4 is phosphorylated by the cGMP-PKG pathway and might be an important target for modulating prostate function by PDE5 inhibitors.

Changes in ion transport in inflammatory disease.

PubMed

Eisenhut, Michael

2006-03-29

Ion transport is essential for maintenance of transmembranous and transcellular electric potential, fluid transport and cellular volume. Disturbance of ion transport has been associated with cellular dysfunction, intra and extracellular edema and abnormalities of epithelial surface liquid volume. There is increasing evidence that conditions characterized by an intense local or systemic inflammatory response are associated with abnormal ion transport. This abnormal ion transport has been involved in the pathogenesis of conditions like hypovolemia due to fluid losses, hyponatremia and hypokalemia in diarrhoeal diseases, electrolyte abnormalities in pyelonephritis of early infancy, septicemia induced pulmonary edema, and in hypersecretion and edema induced by inflammatory reactions of the mucosa of the upper respiratory tract. Components of membranous ion transport systems, which have been shown to undergo a change in function during an inflammatory response include the sodium potassium ATPase, the epithelial sodium channel, the Cystic Fibrosis Transmembrane Conductance Regulator and calcium activated chloride channels and the sodium potassium chloride co-transporter. Inflammatory mediators, which influence ion transport are tumor necrosis factor, gamma interferon, interleukins, transforming growth factor, leukotrienes and bradykinin. They trigger the release of specific messengers like prostaglandins, nitric oxide and histamine which alter ion transport system function through specific receptors, intracellular second messengers and protein kinases. This review summarizes data on in vivo measurements of changes in ion transport in acute inflammatory conditions and in vitro studies, which have explored the underlying mechanisms. Potential interventions directed at a correction of the observed abnormalities are discussed.

Changes in ion transport in inflammatory disease

PubMed Central

Eisenhut, Michael

2006-01-01

Ion transport is essential for maintenance of transmembranous and transcellular electric potential, fluid transport and cellular volume. Disturbance of ion transport has been associated with cellular dysfunction, intra and extracellular edema and abnormalities of epithelial surface liquid volume. There is increasing evidence that conditions characterized by an intense local or systemic inflammatory response are associated with abnormal ion transport. This abnormal ion transport has been involved in the pathogenesis of conditions like hypovolemia due to fluid losses, hyponatremia and hypokalemia in diarrhoeal diseases, electrolyte abnormalites in pyelonephritis of early infancy, septicemia induced pulmonary edema, and in hypersecretion and edema induced by inflammatory reactions of the mucosa of the upper respiratory tract. Components of membranous ion transport systems, which have been shown to undergo a change in function during an inflammatory response include the sodium potassium ATPase, the epithelial sodium channel, the Cystic Fibrosis Transmembrane Conductance Regulator and calcium activated chloride channels and the sodium potassium chloride co-transporter. Inflammatory mediators, which influence ion transport are tumor necrosis factor, gamma interferon, interleukins, transforming growth factor, leukotrienes and bradykinin. They trigger the release of specific messengers like prostaglandins, nitric oxide and histamine which alter ion transport system function through specific receptors, intracellular second messengers and protein kinases. This review summarizes data on in vivo measurements of changes in ion transport in acute inflammatory conditions and in vitro studies, which have explored the underlying mechanisms. Potential interventions directed at a correction of the observed abnormalities are discussed. PMID:16571116

The Voltage-Sensing Domain of K(v)7.2 Channels as a Molecular Target for Epilepsy-Causing Mutations and Anticonvulsants.

PubMed

Miceli, Francesco; Soldovieri, Maria Virginia; Iannotti, Fabio Arturo; Barrese, Vincenzo; Ambrosino, Paolo; Martire, Maria; Cilio, Maria Roberta; Taglialatela, Maurizio

2011-01-01

Understanding the molecular mechanisms underlying voltage-dependent gating in voltage-gated ion channels (VGICs) has been a major effort over the last decades. In recent years, changes in the gating process have emerged as common denominators for several genetically determined channelopathies affecting heart rhythm (arrhythmias), neuronal excitability (epilepsy, pain), or skeletal muscle contraction (periodic paralysis). Moreover, gating changes appear as the main molecular mechanism by which several natural toxins from a variety of species affect ion channel function. In this work, we describe the pathophysiological and pharmacological relevance of the gating process in voltage-gated K(+) channels encoded by the K(v)7 gene family. After reviewing the current knowledge on the molecular mechanisms and on the structural models of voltage-dependent gating in VGICs, we describe the physiological relevance of these channels, with particular emphasis on those formed by K(v)7.2-K(v)7.5 subunits having a well-established role in controlling neuronal excitability in humans. In fact, genetically determined alterations in K(v)7.2 and K(v)7.3 genes are responsible for benign familial neonatal convulsions, a rare seizure disorder affecting newborns, and the pharmacological activation of K(v)7.2/3 channels can exert antiepileptic activity in humans. Both mutation-triggered channel dysfunction and drug-induced channel activation can occur by impeding or facilitating, respectively, channel sensitivity to membrane voltage and can affect overlapping molecular sites within the voltage-sensing domain of these channels. Thus, understanding the molecular steps involved in voltage-sensing in K(v)7 channels will allow to better define the pathogenesis of rare human epilepsy, and to design innovative pharmacological strategies for the treatment of epilepsies and, possibly, other human diseases characterized by neuronal hyperexcitability.

The Voltage-Sensing Domain of Kv7.2 Channels as a Molecular Target for Epilepsy-Causing Mutations and Anticonvulsants

PubMed Central

Miceli, Francesco; Soldovieri, Maria Virginia; Iannotti, Fabio Arturo; Barrese, Vincenzo; Ambrosino, Paolo; Martire, Maria; Cilio, Maria Roberta; Taglialatela, Maurizio

2010-01-01

Understanding the molecular mechanisms underlying voltage-dependent gating in voltage-gated ion channels (VGICs) has been a major effort over the last decades. In recent years, changes in the gating process have emerged as common denominators for several genetically determined channelopathies affecting heart rhythm (arrhythmias), neuronal excitability (epilepsy, pain), or skeletal muscle contraction (periodic paralysis). Moreover, gating changes appear as the main molecular mechanism by which several natural toxins from a variety of species affect ion channel function. In this work, we describe the pathophysiological and pharmacological relevance of the gating process in voltage-gated K+ channels encoded by the Kv7 gene family. After reviewing the current knowledge on the molecular mechanisms and on the structural models of voltage-dependent gating in VGICs, we describe the physiological relevance of these channels, with particular emphasis on those formed by Kv7.2–Kv7.5 subunits having a well-established role in controlling neuronal excitability in humans. In fact, genetically determined alterations in Kv7.2 and Kv7.3 genes are responsible for benign familial neonatal convulsions, a rare seizure disorder affecting newborns, and the pharmacological activation of Kv7.2/3 channels can exert antiepileptic activity in humans. Both mutation-triggered channel dysfunction and drug-induced channel activation can occur by impeding or facilitating, respectively, channel sensitivity to membrane voltage and can affect overlapping molecular sites within the voltage-sensing domain of these channels. Thus, understanding the molecular steps involved in voltage-sensing in Kv7 channels will allow to better define the pathogenesis of rare human epilepsy, and to design innovative pharmacological strategies for the treatment of epilepsies and, possibly, other human diseases characterized by neuronal hyperexcitability. PMID:21687499

Autoantibody-mediated bowel and bladder dysfunction in a patient with chronic, nondiabetic neuropathy.

PubMed

Jackson, Michael W; Gordon, Thomas P; McCombe, Pamela A

2008-04-01

Physiological techniques can be used to detect novel autoantibodies causing alteration of autonomic function after passive transfer to mice. Previously, such antibodies have been detected in patients with type I diabetes mellitus, myasthenia gravis, and Sjogren's syndrome. We now describe a patient with an idiopathic nondiabetic neuropathy with prominent autonomic symptoms, including bladder and bowel dysfunction. Physiological assays of whole colon and bladder were used to determine the presence in the patient serum of functional autoantibodies capable of mediating autonomic dysfunction. Immunoglobulin G (IgG) from this patient was able to disrupt bladder and bowel function on passive transfer to mice. This is a new pattern of autoantibody-mediated abnormality. Although the target antigen is unknown, it is likely to be a cell-surface receptor or ion channel. This case highlights the usefulness of passive transfer studies in detecting functional antibodies in patients with autonomic neuropathy.

Prolactin and teleost ionocytes: new insights into cellular and molecular targets of prolactin in vertebrate epithelia

USGS Publications Warehouse

Breves, Jason P.; McCormick, Stephen D.; Karlstrom, Rolf O.

2014-01-01

The peptide hormone prolactin is a functionally versatile hormone produced by the vertebrate pituitary. Comparative studies over the last six decades have revealed that a conserved function for prolactin across vertebrates is the regulation of ion and water transport in a variety of tissues including those responsible for whole-organism ion homeostasis. In teleost fishes, prolactin was identified as the “freshwater-adapting hormone”, promoting ion-conserving and water-secreting processes by acting on the gill, kidney, gut and urinary bladder. In mammals, prolactin is known to regulate renal, intestinal, mammary and amniotic epithelia, with dysfunction linked to hypogonadism, infertility, and metabolic disorders. Until recently, our understanding of the cellular mechanisms of prolactin action in fishes has been hampered by a paucity of molecular tools to define and study ionocytes, specialized cells that control active ion transport across branchial and epidermal epithelia. Here we review work in teleost models indicating that prolactin regulates ion balance through action on ion transporters, tight-junction proteins, and water channels in ionocytes, and discuss recent advances in our understanding of ionocyte function in the genetically and embryonically accessible zebrafish (Danio rerio). Given the high degree of evolutionary conservation in endocrine and osmoregulatory systems, these studies in teleost models are contributing novel mechanistic insight into how prolactin participates in the development, function, and dysfunction of osmoregulatory systems across the vertebrate lineage.

Transient Receptor Potential Vanilloid 1 Expression Mediates Capsaicin-Induced Cell Death.

PubMed

Ramírez-Barrantes, Ricardo; Córdova, Claudio; Gatica, Sebastian; Rodriguez, Belén; Lozano, Carlo; Marchant, Ivanny; Echeverria, Cesar; Simon, Felipe; Olivero, Pablo

2018-01-01

The transient receptor potential (TRP) ion channel family consists of a broad variety of non-selective cation channels that integrate environmental physicochemical signals for dynamic homeostatic control. Involved in a variety of cellular physiological processes, TRP channels are fundamental to the control of the cell life cycle. TRP channels from the vanilloid (TRPV) family have been directly implicated in cell death. TRPV1 is activated by pain-inducing stimuli, including inflammatory endovanilloids and pungent exovanilloids, such as capsaicin (CAP). TRPV1 activation by high doses of CAP (>10 μM) leads to necrosis, but also exhibits apoptotic characteristics. However, CAP dose-response studies are lacking in order to determine whether CAP-induced cell death occurs preferentially via necrosis or apoptosis. In addition, it is not known whether cytosolic Ca 2+ and mitochondrial dysfunction participates in CAP-induced TRPV1-mediated cell death. By using TRPV1-transfected HeLa cells, we investigated the underlying mechanisms involved in CAP-induced TRPV1-mediated cell death, the dependence of CAP dose, and the participation of mitochondrial dysfunction and cytosolic Ca 2+ increase. Together, our results contribute to elucidate the pathophysiological steps that follow after TRPV1 stimulation with CAP. Low concentrations of CAP (1 μM) induce cell death by a mechanism involving a TRPV1-mediated rapid and transient intracellular Ca 2+ increase that stimulates plasma membrane depolarization, thereby compromising plasma membrane integrity and ultimately leading to cell death. Meanwhile, higher doses of CAP induce cell death via a TRPV1-independent mechanism, involving a slow and persistent intracellular Ca 2+ increase that induces mitochondrial dysfunction, plasma membrane depolarization, plasma membrane loss of integrity, and ultimately, cell death.

Dual role of K ATP channel C-terminal motif in membrane targeting and metabolic regulation.

PubMed

Kline, Crystal F; Kurata, Harley T; Hund, Thomas J; Cunha, Shane R; Koval, Olha M; Wright, Patrick J; Christensen, Matthew; Anderson, Mark E; Nichols, Colin G; Mohler, Peter J

2009-09-29

The coordinated sorting of ion channels to specific plasma membrane domains is necessary for excitable cell physiology. K(ATP) channels, assembled from pore-forming (Kir6.x) and regulatory sulfonylurea receptor subunits, are critical electrical transducers of the metabolic state of excitable tissues, including skeletal and smooth muscle, heart, brain, kidney, and pancreas. Here we show that the C-terminal domain of Kir6.2 contains a motif conferring membrane targeting in primary excitable cells. Kir6.2 lacking this motif displays aberrant channel targeting due to loss of association with the membrane adapter ankyrin-B (AnkB). Moreover, we demonstrate that this Kir6.2 C-terminal AnkB-binding motif (ABM) serves a dual role in K(ATP) channel trafficking and membrane metabolic regulation and dysfunction in these pathways results in human excitable cell disease. Thus, the K(ATP) channel ABM serves as a previously unrecognized bifunctional touch-point for grading K(ATP) channel gating and membrane targeting and may play a fundamental role in controlling excitable cell metabolic regulation.

Big-conductance Ca2+-activated K+ channels in physiological and pathophysiological urinary bladder smooth muscle cells

PubMed Central

Parajuli, Shankar P.; Zheng, Yun-Min; Levin, Robert; Wang, Yong-Xiao

2016-01-01

ABSTRACT Contraction and relaxation of urinary bladder smooth muscle cells (UBSMCs) represent the important physiological functions of the bladder. Contractile responses in UBSMCs are regulated by a number of ion channels including big-conductance Ca2+- activated K+ (BK) channels. Great progress has been made in studies of BK channels in UBSMCs. The intent of this review is to summarize recent exciting findings with respect to the functional interactions of BK channels with muscarinic receptors, ryanodine receptors (RyRs) and inositol triphosphate receptors (IP3Rs) as well as their functional importance under normal and pathophysiological conditions. BK channels are highly expressed in UBSMCs. Activation of muscarinic M3 receptors inhibits the BK channel activity, facilitates opening of voltage-dependent Ca2+ (CaV) channels, and thereby enhances excitability and contractility of UBSMCs. Signaling molecules and regulatory mechanisms involving RyRs and IP3Rs have a significant effect on functions of BK channels and thereby regulate cellular responses in UBSMCs under normal and pathophysiological conditions including overactive bladders. Moreover, BK channels may represent a novel target for the treatment of bladder dysfunctions. PMID:27101440

β1-Adrenergic blocker bisoprolol reverses down-regulated ion channels in sinoatrial node of heart failure rats.

PubMed

Du, Yuan; Zhang, Junbo; Xi, Yutao; Wu, Geru; Han, Ke; Huang, Xin; Ma, Aiqun; Wang, Tingzhong

2016-06-01

Bisoprolol, an antagonist of β1-adrenergic receptors, is effective in reducing the morbidity and mortality in patients with heart failure (HF). It has been found that HF is accompanied with dysfunction of the sinoatrial node (SAN). However, whether bisoprolol reverses the decreased SAN function in HF and how the relevant ion channels in SAN change were relatively less studied. SAN function and messenger RNA (mRNA) expression of sodium channels and hyperpolarization-activated cyclic nucleotide-gated (HCN) channel subunits were assessed in sham-operated rats, abdominal arterio-venous shunt (volume overload)-induced HF rats, and bisoprolol- treated HF rats. SAN cells of rats were isolated by laser capture microdissection. Quantitative real-time PCR analysis was used to quantify mRNA expression of sodium channels and HCN channel subunits in SAN. Intrinsic heart rate declined and sinus node recovery time prolonged in HF rats, indicating the suppressed SAN function, which could be improved by bisoprolol treatment. Nav1.1, Nav1.6, and HCN4 mRNA expressions were reduced in SAN in HF rats compared with that in control rats. Treatment with bisoprolol could reverse both the SAN function and the Nav1.1, Nav1.6, and HCN4 mRNA expression partially. These data indicated that bisoprolol is effective in HF treatment partially due to improved SAN function by reversing the down-regulation of sodium channels (Nav1.1 and Nav1.6) and HCN channel (HCN4) subunits in SAN in failing hearts.

Redox and Nitric Oxide-Mediated Regulation of Sensory Neuron Ion Channel Function

PubMed Central

2015-01-01

Abstract Significance: Reactive oxygen and nitrogen species (ROS and RNS, respectively) can intimately control neuronal excitability and synaptic strength by regulating the function of many ion channels. In peripheral sensory neurons, such regulation contributes towards the control of somatosensory processing; therefore, understanding the mechanisms of such regulation is necessary for the development of new therapeutic strategies and for the treatment of sensory dysfunctions, such as chronic pain. Recent Advances: Tremendous progress in deciphering nitric oxide (NO) and ROS signaling in the nervous system has been made in recent decades. This includes the recognition of these molecules as important second messengers and the elucidation of their metabolic pathways and cellular targets. Mounting evidence suggests that these targets include many ion channels which can be directly or indirectly modulated by ROS and NO. However, the mechanisms specific to sensory neurons are still poorly understood. This review will therefore summarize recent findings that highlight the complex nature of the signaling pathways involved in redox/NO regulation of sensory neuron ion channels and excitability; references to redox mechanisms described in other neuron types will be made where necessary. Critical Issues: The complexity and interplay within the redox, NO, and other gasotransmitter modulation of protein function are still largely unresolved. Issues of specificity and intracellular localization of these signaling cascades will also be addressed. Future Directions: Since our understanding of ROS and RNS signaling in sensory neurons is limited, there is a multitude of future directions; one of the most important issues for further study is the establishment of the exact roles that these signaling pathways play in pain processing and the translation of this understanding into new therapeutics. Antioxid. Redox Signal. 22, 486–504. PMID:24735331

Cytokine–Ion Channel Interactions in Pulmonary Inflammation

PubMed Central

Hamacher, Jürg; Hadizamani, Yalda; Borgmann, Michèle; Mohaupt, Markus; Männel, Daniela Narcissa; Moehrlen, Ueli; Lucas, Rudolf; Stammberger, Uz

2018-01-01

The lungs conceptually represent a sponge that is interposed in series in the bodies’ systemic circulation to take up oxygen and eliminate carbon dioxide. As such, it matches the huge surface areas of the alveolar epithelium to the pulmonary blood capillaries. The lung’s constant exposure to the exterior necessitates a competent immune system, as evidenced by the association of clinical immunodeficiencies with pulmonary infections. From the in utero to the postnatal and adult situation, there is an inherent vital need to manage alveolar fluid reabsorption, be it postnatally, or in case of hydrostatic or permeability edema. Whereas a wealth of literature exists on the physiological basis of fluid and solute reabsorption by ion channels and water pores, only sparse knowledge is available so far on pathological situations, such as in microbial infection, acute lung injury or acute respiratory distress syndrome, and in the pulmonary reimplantation response in transplanted lungs. The aim of this review is to discuss alveolar liquid clearance in a selection of lung injury models, thereby especially focusing on cytokines and mediators that modulate ion channels. Inflammation is characterized by complex and probably time-dependent co-signaling, interactions between the involved cell types, as well as by cell demise and barrier dysfunction, which may not uniquely determine a clinical picture. This review, therefore, aims to give integrative thoughts and wants to foster the unraveling of unmet needs in future research. PMID:29354115

Genetics and epithelial cell dysfunction in cystic fibrosis

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

Riordan, J.R.; Buchwald, M.

1987-01-01

This book examines the advances being made in the study of the physiology, cell biology, and molecular genetics of cystic fibrosis. Emphasis is placed on various areas of research that involve epithelial cells (e.g., the CF-specific phenotypes exhibited by epithelial cells, abnormalities in epithelium ion transport, chloride channel regulation in CF epithelial.) Coverage is presented on the current status of CF, including data on the incidence of the disease, its mode of inheritance, chromosomal localization, genetic heterogeneity, and screening and management.

Peripheral neuropathy in the twitcher mouse: accumulation of extracellular matrix in the endoneurium and aberrant expression of ion channels.

PubMed

Kagitani-Shimono, Kuriko; Mohri, Ikuko; Yagi, Takashi; Taniike, Masako; Suzuki, Kinuko

2008-05-01

Globoid cell leukodystrophy (GLD; Krabbe's disease), caused by a genetic galactosylceramidase deficiency, affects both the central and peripheral nervous systems (CNS and PNS). Allogenic hematopoietic stem-cell transplantation (HSCT) has been beneficial for clinical improvement of this disease. However, recent reports by Siddiqi et al. suggested that none of their transplanted patients achieved complete normalization of their peripheral nerve function, despite the well-documented remyelination of the CNS and PNS in the treated patients. We hypothesized that the PNS dysfunction in GLD is due to altered Schwann cell-axon interactions, resulting in structural abnormalities of the node of Ranvier and aberrant expression of ion channels caused by demyelination and that the persistence of this altered interaction is responsible for the dysfunction of the PNS after HSCT. Since there has not been any investigation of the Schwann cell-axonal relationship in twitcher mice, an authentic model of GLD, we first investigated structural abnormalities, focusing on the node of Ranvier in untreated twitcher mice, and compared the results with those obtained after receiving bone marrow transplantation (BMT). As expected, we found numerous supernumerary Schwann cells that formed structurally abnormal nodes of Ranvier. Similar findings, though at somewhat variable extent, were detected in mice treated with BMT. Activated supernumerary Schwann cells expressed GFAP immunoreactivity and generated Alcian blue-positive extracellular matrix (ECM) in the endoneurial space. The processes of these supernumerary Schwann cells often covered and obliterated the nodal regions. Furthermore, the distribution of Na(+) channel immunoreactivity was diffuse without the concentration at the nodes of Ranvier as seen in wild-type mice. Neither K(+) channels nor Neurexin IV/ Caspr/ Paranoidin (NCP-1) were detected in the twi/twi sciatic nerve. The results of our study suggest the importance of normalization of the Schwann cell-axon relationship for the functional recovery of peripheral nerves, when one considers therapeutic strategies for PNS pathology in GLD.

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

David Triggle: Research collaborations and scientific exchanges with the China Pharmaceutical University, Nanjing, China.

PubMed

Dai, De-Zai

2015-11-15

Over the period 1995-2012, David Triggle was a frequent visitor to the China Pharmaceutical University in Nanjing, China making many important contributions that enhanced the activities of the Research Division of Pharmacology at the University. In addition to providing collegial advice and facilitating interactions with the international pharmacological community, Professor Triggle's international reputation as a thought leader in the field of ion channel research and drug discovery provided important insights into the potential pathophysiological and therapeutic effects of targeting ion channels. This included the L-type calcium channel and the outward delayed rectified potassium currents of rapid (IKr) and slow (IKs) components in the myocardium. The Nanjing research team had been particularly interested in ion channel dysfunction in the context of cardiac arrhythmias, remodeling and drug discovery. With Professor Triggle's assistance, the relationship between an increase in ICa.L and other biological events including an enhancement of IKr and IKr currents, NADPH oxidase and endothelin receptor activation, down regulation of calcium modulating protein FKBP12.6, sarco/endoplasmic reticulum Ca(2+)ATPse (SERCA2A) and calsequens 2 (CASQ2), calcium leak at the diastole and endoplasmic reticulum stress, were evaluated and are discussed. Additionally, the organization of several international symposia was greatly enhanced by input from Professor Triggle as were the published research manuscripts in international pharmacology journals. During his association with the China Pharmaceutical University, Professor Triggle aided in enhancing the scientific standing of the Pharmacology department and was a highly effective ambassador for international research cooperation. Copyright © 2015. Published by Elsevier Inc.

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.

[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.

Rescuing cardiac automaticity in L-type Cav1.3 channelopathies and beyond.

PubMed

Mesirca, Pietro; Bidaud, Isabelle; Mangoni, Matteo E

2016-10-15

Pacemaker activity of the sino-atrial node generates the heart rate. Disease of the sinus node and impairment of atrioventricular conduction induce an excessively low ventricular rate (bradycardia), which cannot meet the needs of the organism. Bradycardia accounts for about half of the total workload of clinical cardiologists. The 'sick sinus' syndrome (SSS) is characterized by sinus bradycardia and periods of intermittent atrial fibrillation. Several genetic or acquired risk factors or pathologies can lead to SSS. Implantation of an electronic pacemaker constitutes the only available therapy for SSS. The incidence of SSS is forecast to double over the next 50 years, with ageing of the general population thus urging the development of complementary or alternative therapeutic strategies. In recent years an increasing number of mutations affecting ion channels involved in sino-atrial automaticity have been reported to underlie inheritable SSS. L-type Ca v 1.3 channels play a major role in the generation and regulation of sino-atrial pacemaker activity and atrioventricular conduction. Mutation in the CACNA1D gene encoding Ca v 1.3 channels induces loss-of-function in channel activity and underlies the sino-atrial node dysfunction and deafness syndrome (SANDD). Mice lacking Ca v 1.3 channels (Ca v 1.3 -/- ) fairly recapitulate SSS and constitute a precious model to test new therapeutic approaches to handle this disease. Work in our laboratory shows that targeting G protein-gated K + (I KACh ) channels effectively rescues SSS of Ca v 1.3 -/- mice. This new concept of 'compensatory' ion channel targeting shines new light on the principles underlying the pacemaker mechanism and may open the way to new therapies for SSS. © 2016 The Authors. The Journal of Physiology © 2016 The Physiological Society.

Structural basis for the selective permeability of channels made of communicating junction proteins.

PubMed

Ek-Vitorin, Jose F; Burt, Janis M

2013-01-01

The open state(s) of gap junction channels is evident from their permeation by small ions in response to an applied intercellular (transjunctional/transchannel) voltage gradient. That an open channel allows variable amounts of current to transit from cell-to-cell in the face of a constant intercellular voltage difference indicates channel open/closing can be complete or partial. The physiological significance of such open state options is, arguably, the main concern of junctional regulation. Because gap junctions are permeable to many substances, it is sensible to inquire whether and how each open state influences the intercellular diffusion of molecules as valuable as, but less readily detected than current-carrying ions. Presumably, structural changes perceived as shifts in channel conductivity would significantly alter the transjunctional diffusion of molecules whose limiting diameter approximates the pore's limiting diameter. Moreover, changes in junctional permeability to some molecules might occur without evident changes in conductivity, either at macroscopic or single channel level. Open gap junction channels allow the exchange of cytoplasmic permeants between contacting cells by simple diffusion. The identity of such permeants, and the functional circumstances and consequences of their junctional exchange presently constitute the most urgent (and demanding) themes of the field. Here, we consider the necessity for regulating this exchange, the possible mechanism(s) and structural elements likely involved in such regulation, and how regulatory phenomena could be perceived as changes in chemical vs. electrical coupling; an overall reflection on our collective knowledge of junctional communication is then applied to suggest new avenues of research. This article is part of a Special Issue entitled: The Communicating junctions, roles and dysfunctions. Copyright © 2012 Elsevier B.V. All rights reserved.

Utility of the Electrocardiogram in Drug Overdose and Poisoning: Theoretical Considerations and Clinical Implications

PubMed Central

Yates, Christopher; Manini, Alex F

2012-01-01

The ECG is a rapidly available clinical tool that can help clinicians manage poisoned patients. Specific myocardial effects of cardiotoxic drugs have well-described electrocardiographic manifestations. In the practice of clinical toxicology, classic ECG changes may hint at blockade of ion channels, alterations of adrenergic tone, or dysfunctional metabolic activity of the myocardium. This review will offer a structured approach to ECG interpretation in poisoned patients with a focus on clinical implications and ECG-based management recommendations in the initial evaluation of patients with acute cardiotoxicity. PMID:22708912

Ion Channels in the Heart

PubMed Central

Bartos, Daniel C.; Grandi, Eleonora; Ripplinger, Crystal M.

2015-01-01

Optimal cardiac function depends on proper timing of excitation and contraction in various regions of the heart, as well as on appropriate heart rate. This is accomplished via specialized electrical properties of various components of the system, including the sinoatrial node, atria, atrioventricular node, His-Purkinje system, and ventricles. Here we review the major regionally-determined electrical properties of these cardiac regions and present the available data regarding the molecular and ionic bases of regional cardiac function and dysfunction. Understanding these differences is of fundamental importance for the investigation of arrhythmia mechanisms and pharmacotherapy. PMID:26140724

The Gárdos channel: a review of the Ca2+-activated K+ channel in human erythrocytes.

PubMed

Maher, Anthony D; Kuchel, Philip W

2003-08-01

Ca(2+)-dependent K(+) efflux from human erythrocytes was first described in the 1950s. Subsequent studies revealed that a K(+)-specific membrane protein (the Gárdos channel) was responsible for this phenomenon (the Gárdos effect). In recent years several types of Ca-activated K(+) channel have been identified and studied in a wide range of cells, with the erythrocyte Gárdos channel serving as both a model for a broader physiological perspective, and an intriguing component of erythrocyte function. The existence of this channel has raised a number of questions. For example, what is its role in the establishment and maintenance of ionic distribution across the red cell membrane? What role might it play in erythrocyte development? To what extent is it active in circulating erythrocytes? What are the cell-physiological implications of its dysfunction?This review summarises current knowledge of this membrane protein with respect to its function and structure, its physiological roles (some putative) and its contribution to various disease states, and it provides an introduction to adaptable NMR methods, which is our own area of technical expertise, for such ion transport analysis.

Cryo-EM structure of the cytoplasmic domain of murine transient receptor potential cation channel subfamily C member 6 (TRPC6).

PubMed

Azumaya, Caleigh M; Sierra-Valdez, Francisco; Cordero-Morales, Julio F; Nakagawa, Terunaga

2018-05-11

The kidney maintains the internal milieu by regulating the retention and excretion of proteins, ions, and small molecules. The glomerular podocyte forms the slit diaphragm of the ultrafiltration filter, whose damage leads to progressive kidney failure and focal segmental glomerulosclerosis (FSGS). The canonical transient receptor potential 6 (TRPC6) ion channel is expressed in the podocyte and mutations in its cytoplasmic domain cause FSGS in humans. In vitro evaluation of disease-causing mutations in TRPC6 has revealed that these genetic alterations result in abnormal ion channel gating. However, the mechanism whereby the cytoplasmic domain modulates TRPC6 function is largely unknown. Here we report a cryoEM structure of the cytoplasmic domain of murine TRPC6 at 3.8Å resolution. The cytoplasmic fold of TRPC6 is characterized by an inverted dome-like chamber pierced by four radial horizontal helices that converge into a vertical coiled-coil at the central axis. Unlike in other TRP channels, TRPC6 displays a unique domain swap that occurs at the junction of the horizontal helices and coiled-coil. Multiple FSGS mutations converge at the buried interface between the vertical coiled-coil and the ankyrin repeats, which form the dome, suggesting these regions are critical for allosteric gating modulation. This functionally critical interface is a potential target for drug design. Importantly, dysfunction in other family members leads to learning deficits (TRPC1/4/5) and ataxia (TRPC3). Our data provide a structural framework for the mechanistic investigation of the TRPC family. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Kindling and Oxidative Stress as Contributors to Myalgic Encephalomyelitis/Chronic Fatigue Syndrome

PubMed Central

Jason, L. A.; Porter, N.; Herrington, J.; Sorenson, M.; Kubow, S.

2010-01-01

Myalgic Encephalomyelitis/chronic fatigue syndrome (ME/CFS) is one of the more complex illnesses involving multiple systems within the body. Onset of ME/CFS frequently occurs quickly, and many patients report a prior exposure to a viral infection. This debilitating illness can affect the immune, neuroendocrine, autonomic, and neurologic systems. Abnormal biological findings among some patients have included aberrant ion transport and ion channel activity, cortisol deficiency, sympathetic nervous system hyperactivity, EEG spike waves, left ventricular dysfunction in the heart, low natural killer cell cytotoxicity, and a shift from Th1 to Th2 cytokines. We propose that the kindling and oxidative stress theories provide a heuristic template for better understanding the at times conflicting findings regarding the etiology and pathophysiology of this illness. PMID:21253446

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.

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.

βIV Spectrinopathies Cause Profound Intellectual Disability, Congenital Hypotonia, and Motor Axonal Neuropathy.

PubMed

Wang, Chih-Chuan; Ortiz-González, Xilma R; Yum, Sabrina W; Gill, Sara M; White, Amy; Kelter, Erin; Seaver, Laurie H; Lee, Sansan; Wiley, Graham; Gaffney, Patrick M; Wierenga, Klaas J; Rasband, Matthew N

2018-06-07

βIV spectrin links ankyrinG (AnkG) and clustered ion channels at axon initial segments (AISs) and nodes of Ranvier to the axonal cytoskeleton. Here, we report bi-allelic pathogenic SPTBN4 variants (three homozygous and two compound heterozygous) that cause a severe neurological syndrome that includes congenital hypotonia, intellectual disability, and motor axonal and auditory neuropathy. We introduced these variants into βIV spectrin, expressed these in neurons, and found that 5/7 were loss-of-function variants disrupting AIS localization or abolishing phosphoinositide binding. Nerve biopsies from an individual with a loss-of-function variant had reduced nodal Na + channels and no nodal KCNQ2 K + channels. Modeling the disease in mice revealed that although ankyrinR (AnkR) and βI spectrin can cluster Na + channels and partially compensate for the loss of AnkG and βIV spectrin at nodes of Ranvier, AnkR and βI spectrin cannot cluster KCNQ2- and KCNQ3-subunit-containing K + channels. Our findings define a class of spectrinopathies and reveal the molecular pathologies causing nervous-system dysfunction. Copyright © 2018 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

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.

Enhancement of proton transfer in ion channels by membrane phosphate headgroups.

PubMed

Wyatt, Debra L; de Godoy, Carlos Marcelo G; Cukierman, Samuel

2009-05-14

The transfer of protons (H+) in gramicidin (gA) channels is markedly distinct in monoglyceride and phospholipid membranes. In this study, the molecular groups that account for those differences were investigated using a new methodology. The rates of H+ transfer were measured in single gA channels reconstituted in membranes made of plain ceramides or sphingomyelins and compared to those in monoglyceride and phospholipid bilayers. Single-channel conductances to protons (gH) were significantly larger in sphingomyelin than in ceramide membranes. A novel and unsuspected finding was that H+ transfer was heavily attenuated or completely blocked in ceramide (but not in sphingomyelin) membranes in low-ionic-strength solutions. It is reasoned that H-bond dynamics at low ionic strengths between membrane ceramides and gA makes channels dysfunctional. The rate of H+ transfer in gA channels in ceramide membranes is significantly higher than that in monoglyceride bilayers. This suggests that solvation of the hydrophobic surface of gA channels by two acyl chains in ceramides stabilizes the gA channels and the water wire inside the pore, leading to an enhancement of H+ transfer in relation to that occurring in monoglyceride membranes. gH values in gA channels are similar in ceramide and monoglyceride bilayers and in sphingomyelin and phospholipid membranes. It is concluded that phospho headgroups in membranes have significant effects on the rate of H+ transfer at the membrane gA channel/solution interfaces, enhancing the entry and exit rates of protons in channels.

Calcineurin Dysregulation Underlies Spinal Cord Injury-Induced K+ Channel Dysfunction in DRG Neurons

PubMed Central

Zemel, Benjamin M.; Brown, Eric V.; Urban, Mark W.; Tymanskyj, Stephen R.; Lepore, Angelo C.

2017-01-01

Dysfunction of the fast-inactivating Kv3.4 potassium current in dorsal root ganglion (DRG) neurons contributes to the hyperexcitability associated with persistent pain induced by spinal cord injury (SCI). However, the underlying mechanism is not known. In light of our previous work demonstrating modulation of the Kv3.4 channel by phosphorylation, we investigated the role of the phosphatase calcineurin (CaN) using electrophysiological, molecular, and imaging approaches in adult female Sprague Dawley rats. Pharmacological inhibition of CaN in small-diameter DRG neurons slowed repolarization of the somatic action potential (AP) and attenuated the Kv3.4 current. Attenuated Kv3.4 currents also exhibited slowed inactivation. We observed similar effects on the recombinant Kv3.4 channel heterologously expressed in Chinese hamster ovary cells, supporting our findings in DRG neurons. Elucidating the molecular basis of these effects, mutation of four previously characterized serines within the Kv3.4 N-terminal inactivation domain eliminated the effects of CaN inhibition on the Kv3.4 current. SCI similarly induced concurrent Kv3.4 current attenuation and slowing of inactivation. Although there was little change in CaN expression and localization after injury, SCI induced upregulation of the native regulator of CaN 1 (RCAN1) in the DRG at the transcript and protein levels. Consistent with CaN inhibition resulting from RCAN1 upregulation, overexpression of RCAN1 in naive DRG neurons recapitulated the effects of pharmacological CaN inhibition on the Kv3.4 current and the AP. Overall, these results demonstrate a novel regulatory pathway that links CaN, RCAN1, and Kv3.4 in DRG neurons. Dysregulation of this pathway might underlie a peripheral mechanism of pain sensitization induced by SCI. SIGNIFICANCE STATEMENT Pain sensitization associated with spinal cord injury (SCI) involves poorly understood maladaptive modulation of neuronal excitability. Although central mechanisms have received significant attention, recent studies have identified peripheral nerve hyperexcitability as a driver of persistent pain signaling after SCI. However, the ion channels and signaling molecules responsible for this change in primary sensory neuron excitability are still not well defined. To address this problem, this study used complementary electrophysiological and molecular methods to determine how Kv3.4, a voltage-gated K+ channel robustly expressed in dorsal root ganglion neurons, becomes dysfunctional upon calcineurin (CaN) inhibition. The results strongly suggest that CaN inhibition underlies SCI-induced dysfunction of Kv3.4 and the associated excitability changes through upregulation of the native regulator of CaN 1 (RCAN1). PMID:28751455

Cell Membrane Transport Mechanisms: Ion Channels and Electrical Properties of Cell Membranes.

PubMed

Kulbacka, Julita; Choromańska, Anna; Rossowska, Joanna; Weżgowiec, Joanna; Saczko, Jolanta; Rols, Marie-Pierre

2017-01-01

Cellular life strongly depends on the membrane ability to precisely control exchange of solutes between the internal and external (environmental) compartments. This barrier regulates which types of solutes can enter and leave the cell. Transmembrane transport involves complex mechanisms responsible for passive and active carriage of ions and small- and medium-size molecules. Transport mechanisms existing in the biological membranes highly determine proper cellular functions and contribute to drug transport. The present chapter deals with features and electrical properties of the cell membrane and addresses the questions how the cell membrane accomplishes transport functions and how transmembrane transport can be affected. Since dysfunctions of plasma membrane transporters very often are the cause of human diseases, we also report how specific transport mechanisms can be modulated or inhibited in order to enhance the therapeutic effect.

Do androgens play a beneficial role in the regulation of vascular tone? Nongenomic vascular effects of testosterone metabolites

PubMed Central

Perusquía, Mercedes

2010-01-01

The marked sexual dimorphism that exists in human cardiovascular diseases has led to the dogmatic concept that testosterone (Tes) has deleterious effects and exacerbates the development of cardiovascular disease in males. While some animal studies suggest that Tes does exert deleterious effects by enhancing vascular tone through acute or chronic mechanisms, accumulating evidence suggests that Tes and other androgens exert beneficial effects by inducing rapid vasorelaxation of vascular smooth muscle through nongenomic mechanisms. While this effect frequently has been observed in large arteries at micromolar concentrations, more recent studies have reported vasorelaxation of smaller resistance arteries at nanomolar (physiological) concentrations. The key mechanism underlying Tes-induced vasorelaxation appears to be the modulation of vascular smooth muscle ion channel function, particularly the inactivation of L-type voltage-operated Ca2+ channels and/or the activation of voltage-operated and Ca2+-activated K+ channels. Studies employing Tes analogs and metabolites reveal that androgen-induced vasodilation is a structurally specific nongenomic effect that is fundamentally different than the genomic effects on reproductive targets. For example, 5α-dihydrotestosterone exhibits potent genomic-androgenic effects but only moderate vasorelaxing activity, whereas its isomer 5β-dihydrotestosterone is devoid of androgenic effects but is a highly efficacious vasodilator. These findings suggest that the dihydro-metabolites of Tes or other androgen analogs devoid of androgenic or estrogenic effects could have useful therapeutic roles in hypertension, erectile dysfunction, prostatic ischemia, or other vascular dysfunctions. PMID:20228257

An ion channel library for drug discovery and safety screening on automated platforms.

PubMed

Wible, Barbara A; Kuryshev, Yuri A; Smith, Stephen S; Liu, Zhiqi; Brown, Arthur M

2008-12-01

Ion channels represent the third largest class of targets in drug discovery after G-protein coupled receptors and kinases. In spite of this ranking, ion channels continue to be under exploited as drug targets compared with the other two groups for several reasons. First, with 400 ion channel genes and an even greater number of functional channels due to mixing and matching of individual subunits, a systematic collection of ion channel-expressing cell lines for drug discovery and safety screening has not been available. Second, the lack of high-throughput functional assays for ion channels has limited their use as drug targets. Now that automated electrophysiology has come of age and provided the technology to assay ion channels at medium to high throughput, we have addressed the need for a library of ion channel cell lines by constructing the Ion Channel Panel (ChanTest Corp., Cleveland, OH). From 400 ion channel genes, a collection of 82 of the most relevant human ion channels for drug discovery, safety, and human disease has been assembled.Each channel has been stably overexpressed in human embryonic kidney 293 or Chinese hamster ovary cells. Cell lines have been selected and validated on automated electrophysiology systems to facilitate cost-effective screening for safe and selective compounds at earlier stages in the drug development process. The screening and validation processes as well as the relative advantages of different screening platforms are discussed.

Tuning the ion selectivity of tetrameric cation channels by changing the number of ion binding sites

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

Derebe, Mehabaw G.; Sauer, David B.; Zeng, Weizhong

2015-11-30

Selective ion conduction across ion channel pores is central to cellular physiology. To understand the underlying principles of ion selectivity in tetrameric cation channels, we engineered a set of cation channel pores based on the nonselective NaK channel and determined their structures to high resolution. These structures showcase an ensemble of selectivity filters with a various number of contiguous ion binding sites ranging from 2 to 4, with each individual site maintaining a geometry and ligand environment virtually identical to that of equivalent sites in K{sup +} channel selectivity filters. Combined with single channel electrophysiology, we show that only themore » channel with four ion binding sites is K{sup +} selective, whereas those with two or three are nonselective and permeate Na{sup +} and K{sup +} equally well. These observations strongly suggest that the number of contiguous ion binding sites in a single file is the key determinant of the channel's selectivity properties and the presence of four sites in K{sup +} channels is essential for highly selective and efficient permeation of K{sup +} ions.« less

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.

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

Pyruvate Dehydrogenase Kinase-mediated Glycolytic Metabolic Shift in the Dorsal Root Ganglion Drives Painful Diabetic Neuropathy*

PubMed Central

Rahman, Md Habibur; Jha, Mithilesh Kumar; Kim, Jong-Heon; Nam, Youngpyo; Lee, Maan Gee; Go, Younghoon; Harris, Robert A.; Park, Dong Ho; Kook, Hyun; Lee, In-Kyu; Suk, Kyoungho

2016-01-01

The dorsal root ganglion (DRG) is a highly vulnerable site in diabetic neuropathy. Under diabetic conditions, the DRG is subjected to tissue ischemia or lower ambient oxygen tension that leads to aberrant metabolic functions. Metabolic dysfunctions have been documented to play a crucial role in the pathogenesis of diverse pain hypersensitivities. However, the contribution of diabetes-induced metabolic dysfunctions in the DRG to the pathogenesis of painful diabetic neuropathy remains ill-explored. In this study, we report that pyruvate dehydrogenase kinases (PDK2 and PDK4), key regulatory enzymes in glucose metabolism, mediate glycolytic metabolic shift in the DRG leading to painful diabetic neuropathy. Streptozotocin-induced diabetes substantially enhanced the expression and activity of the PDKs in the DRG, and the genetic ablation of Pdk2 and Pdk4 attenuated the hyperglycemia-induced pain hypersensitivity. Mechanistically, Pdk2/4 deficiency inhibited the diabetes-induced lactate surge, expression of pain-related ion channels, activation of satellite glial cells, and infiltration of macrophages in the DRG, in addition to reducing central sensitization and neuroinflammation hallmarks in the spinal cord, which probably accounts for the attenuated pain hypersensitivity. Pdk2/4-deficient mice were partly resistant to the diabetes-induced loss of peripheral nerve structure and function. Furthermore, in the experiments using DRG neuron cultures, lactic acid treatment enhanced the expression of the ion channels and compromised cell viability. Finally, the pharmacological inhibition of DRG PDKs or lactic acid production substantially attenuated diabetes-induced pain hypersensitivity. Taken together, PDK2/4 induction and the subsequent lactate surge induce the metabolic shift in the diabetic DRG, thereby contributing to the pathogenesis of painful diabetic neuropathy. PMID:26769971

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.

Fe(2+) substrate transport through ferritin protein cage ion channels influences enzyme activity and biomineralization.

PubMed

Behera, Rabindra K; Torres, Rodrigo; Tosha, Takehiko; Bradley, Justin M; Goulding, Celia W; Theil, Elizabeth C

2015-09-01

Ferritins, complex protein nanocages, form internal iron-oxy minerals (Fe2O3·H2O), by moving cytoplasmic Fe(2+) through intracage ion channels to cage-embedded enzyme (2Fe(2+)/O2 oxidoreductase) sites where ferritin biomineralization is initiated. The products of ferritin enzyme activity are diferric oxy complexes that are mineral precursors. Conserved, carboxylate amino acid side chains of D127 from each of three cage subunits project into ferritin ion channels near the interior ion channel exits and, thus, could direct Fe(2+) movement to the internal enzyme sites. Ferritin D127E was designed and analyzed to probe properties of ion channel size and carboxylate crowding near the internal ion channel opening. Glu side chains are chemically equivalent to, but longer by one -CH2 than Asp, side chains. Ferritin D127E assembled into normal protein cages, but diferric peroxo formation (enzyme activity) was not observed, when measured at 650 nm (DFP λ max). The caged biomineral formation, measured at 350 nm in the middle of the broad, nonspecific Fe(3+)-O absorption band, was slower. Structural differences (protein X-ray crystallography), between ion channels in wild type and ferritin D127E, which correlate with the inhibition of ferritin D127E enzyme activity include: (1) narrower interior ion channel openings/pores; (2) increased numbers of ion channel protein-metal binding sites, and (3) a change in ion channel electrostatics due to carboxylate crowding. The contributions of ion channel size and structure to ferritin activity reflect metal ion transport in ion channels are precisely regulated both in ferritin protein nanocages and membranes of living cells.

Fe2+ Substrate Transport through Ferritin Protein Cage Ion Channels Influences Enzyme Activity and Biomineralization

PubMed Central

Behera, Rabindra K.; Torres, Rodrigo; Tosha, Takehiko; Bradley, Justin M.; Goulding, Celia W.; Theil, Elizabeth C.

2015-01-01

Ferritins, complex protein nanocages, form internal iron-oxy minerals (Fe2O3.H2O), by moving cytoplasmic Fe2+ through intracage ion channels to cage-embedded enzyme (2Fe2+/O2 oxidoreductase) sites where ferritin biomineralization is initiated. The products of ferritin enzyme activity are diferric oxy complexes that are mineral precursors. Conserved, carboxylate amino acid side chains of D127 from each of three cage subunits project into ferritin ion channels near the interior ion channel exits and, thus, could direct Fe2+ movement to the internal enzyme sites. Ferritin D127E was designed and analyzed to probe properties of ion channel size and carboxylate crowding near the internal ion channel opening. Glu side chains are chemically equivalent to, but longer by one – CH2 than Asp, side chains. Ferritin D127E assembled into normal protein cages, but diferric peroxo formation (enzyme activity) was not observed, when measured at 650nm (DFP λmax). The caged biomineral formation, measured at 350 nm in the middle of the broad, nonspecific Fe3+-O absorption band, was slower. Structural differences (protein X-ray crystallography), between ion channels in wild type and ferritin D127E, which correlate with the inhibition of ferritin D127E enzyme activity include: 1. narrower interior ion channel openings/pores, 2. increased numbers of ion channel protein-metal binding sites, and 3. a change in ion channel electrostatics due to carboxylate crowding. The contributions of ion channel size and structure to ferritin activity reflect metal ion transport in ion channels are precisely regulated both in ferritin protein nanocages and membranes of living cells. PMID:26202907

Dynamic ion-ion and water-ion interactions in ion channels.

PubMed Central

Wu, J V

1992-01-01

The dynamic interactions among ions and water molecules in ion channels are treated based on an assumption that ions at binding sites can be knocked off by both transient entering ions and local water molecules. The theory, when applied to a single-site model K+ channel, provides solutions for super- and subsaturations, flux-ratio exponent (n') greater than 1, osmotic streaming current, activity-dependent reversal potentials, and anomalous mole-fraction behavior. The analysis predicts that: (a) the saturation may but, in general, does not follow the Michaelis-Menten relation; (b) streaming current results from imbalanced water-ion knock-off interactions; (c) n' greater than 1 even for single-site channels, but it is unlikely to exceed 1.4 unless the pore is occupied by one or more ion(s); (d) in the calculation involving two permeant ion species with similar radii, the heavier ions show higher affinity; the ion-ion knock-off dissociation from the site is more effective when two interacting ions are identical. Therefore, the "multi-ion behaviors" found in most ion channels are the consequences of dynamic ion-ion and water-ion interactions. The presence of these interactions does not require two or more binding sites in channels. PMID:1376158

The Control of Male Fertility by Spermatozoan Ion Channels

PubMed Central

Lishko, Polina V.; Kirichok, Yuriy; Ren, Dejian; Navarro, Betsy; Chung, Jean-Ju

2014-01-01

Ion channels control the sperm ability to fertilize the egg by regulating sperm maturation in the female reproductive tract and by triggering key sperm physiological responses required for successful fertilization such as hyperactivated motility, chemotaxis, and the acrosome reaction. CatSper, a pH-regulated, calcium-selective ion channel, and KSper (Slo3) are core regulators of sperm tail calcium entry and sperm hyperactivated motility. Many other channels had been proposed as regulating sperm activity without direct measurements. With the development of the sperm patch-clamp technique, CatSper and KSper have been confirmed as the primary spermatozoan ion channels. In addition, the voltage-gated proton channel Hv1 has been identified in human sperm tail, and the P2X2 ion channel has been identified in the midpiece of mouse sperm. Mutations and deletions in sperm-specific ion channels affect male fertility in both mice and humans without affecting other physiological functions. The uniqueness of sperm ion channels makes them ideal pharmaceutical targets for contraception. In this review we discuss how ion channels regulate sperm physiology. PMID:22017176

Phosphorylation-Dependent Regulation of Ryanodine Receptors

PubMed Central

Marx, Steven O.; Reiken, Steven; Hisamatsu, Yuji; Gaburjakova, Marta; Gaburjakova, Jana; Yang, Yi-Ming; Rosemblit, Nora; Marks, Andrew R.

2001-01-01

Ryanodine receptors (RyRs), intracellular calcium release channels required for cardiac and skeletal muscle contraction, are macromolecular complexes that include kinases and phosphatases. Phosphorylation/dephosphorylation plays a key role in regulating the function of many ion channels, including RyRs. However, the mechanism by which kinases and phosphatases are targeted to ion channels is not well understood. We have identified a novel mechanism involved in the formation of ion channel macromolecular complexes: kinase and phosphatase targeting proteins binding to ion channels via leucine/isoleucine zipper (LZ) motifs. Activation of kinases and phosphatases bound to RyR2 via LZs regulates phosphorylation of the channel, and disruption of kinase binding via LZ motifs prevents phosphorylation of RyR2. Elucidation of this new role for LZs in ion channel macromolecular complexes now permits: (a) rapid mapping of kinase and phosphatase targeting protein binding sites on ion channels; (b) predicting which kinases and phosphatases are likely to regulate a given ion channel; (c) rapid identification of novel kinase and phosphatase targeting proteins; and (d) tools for dissecting the role of kinases and phosphatases as modulators of ion channel function. PMID:11352932

Ion channel blockers for the treatment of neuropathic pain.

PubMed

Colombo, Elena; Francisconi, Simona; Faravelli, Laura; Izzo, Emanuela; Pevarello, Paolo

2010-05-01

Neuropathic pain, a severe chronic pain condition characterized by a complex pathophysiology, is a largely unmet medical need. Ion channels, which underlie cell excitability, are heavily implicated in the biological mechanisms that generate and sustain neuropathic pain. This review highlights the biological evidence supporting the involvement of voltage-, proton- and ligand-gated ion channels in the neuropathic pain setting. Ion channel modulators at different research or development stages are reviewed and referenced. Ion channel modulation is one of the main avenues to achieve novel, improved neuropathic pain treatments. Voltage-gated sodium and calcium channel and glutamate receptor modulators are likely to produce new, improved agents in the future. Rationally targeting subtypes of known ion channels, tackling recently discovered ion channel targets or combining drugs with different mechanism of action will be primary sources of new drugs in the longer term.

Micro- and nanofabrication methods for ion channel reconstitution in bilayer lipid membranes

NASA Astrophysics Data System (ADS)

Tadaki, Daisuke; Yamaura, Daichi; Arata, Kohei; Ohori, Takeshi; Ma, Teng; Yamamoto, Hideaki; Niwano, Michio; Hirano-Iwata, Ayumi

2018-03-01

The self-assembled bilayer lipid membrane (BLM) forms the basic structure of the cell membrane and serves as a major barrier against ion movement. Ion channel proteins function as gated pores that permit ion permeation across the BLM. The reconstitution of ion channel proteins in artificially formed BLMs represents a well-defined system for investigating channel functions and screening drug effects on ion channels. In this review, we will discuss our recent microfabrication approaches to the formation of stable BLMs containing ion channel proteins as a potential platform for next-generation drug screening systems. BLMs formed in a microaperture having a tapered edge exhibited highly stable properties, such as a lifetime of ∼65 h and tolerance to solution changes even after the incorporation of the human ether-a-go-go-related gene (hERG) channel. We also explore a new method of efficiently incorporating human ion channels into BLMs by centrifugation. Our approaches to the formation of stable BLMs and efficient channel incorporation markedly improve the experimental efficiency of BLM reconstitution systems, leading to the realization of a BLM-based high-throughput platform for functional assays of various ion channels.

Thermodynamic limitations on the temperature sensitivity of cell-membrane ion channels: Trouble with enthalpy uncertainty

NASA Astrophysics Data System (ADS)

Zheltikov, A. M.

2018-06-01

Energy exchange between a thermodynamic ensemble of heat- and cold-activated cell-membrane ion channels and the surrounding heat reservoir is shown to impose fundamental limitations on the performance of such channels as temperature-controlled gates for thermal cell activation. Analysis of unavoidable thermodynamic internal-energy fluctuations caused by energy exchange between the ion channels and the heat bath suggests that the resulting enthalpy uncertainty is too high for a robust ion-current gating by a single ion channel, implying that large ensembles of ion channels are needed for thermal cell activation. We argue, based on this thermodynamic analysis, that, had thermosensitive cell-membrane ion channels operated individually, rather than as large ensembles, robust thermal cell activation would have been impossible because of thermodynamic fluctuations.

Targeting ion channels for the treatment of gastrointestinal motility disorders

PubMed Central

Beyder, Arthur

2012-01-01

Gastrointestinal (GI) functional and motility disorders are highly prevalent and responsible for long-term morbidity and sometimes mortality in the affected patients. It is estimated that one in three persons has a GI functional or motility disorder. However, diagnosis and treatment of these widespread conditions remains challenging. This partly stems from the multisystem pathophysiology, including processing abnormalities in the central and peripheral (enteric) nervous systems and motor dysfunction in the GI wall. Interstitial cells of Cajal (ICCs) are central to the generation and propagation of the cyclical electrical activity and smooth muscle cells (SMCs) are responsible for electromechanical coupling. In these and other excitable cells voltage-sensitive ion channels (VSICs) are the main molecular units that generate and regulate electrical activity. Thus, VSICs are potential targets for intervention in GI motility disorders. Research in this area has flourished with advances in the experimental methods in molecular and structural biology and electrophysiology. However, our understanding of the molecular mechanisms responsible for the complex and variable electrical behavior of ICCs and SMCs remains incomplete. In this review, we focus on the slow waves and action potentials in ICCs and SMCs. We describe the constituent VSICs, which include voltage-gated sodium (NaV), calcium (CaV), potassium (KV, KCa), chloride (Cl–) and nonselective ion channels (transient receptor potentials [TRPs]). VSICs have significant structural homology and common functional mechanisms. We outline the approaches and limitations and provide examples of targeting VSICs at the pores, voltage sensors and alternatively spliced sites. Rational drug design can come from an integrated view of the structure and mechanisms of gating and activation by voltage or mechanical stress. PMID:22282704

Matrix metalloproteinase-9 and -2 enhance the ligand sensitivity of photoreceptor cyclic nucleotide-gated channels.

PubMed

Meighan, Peter C; Meighan, Starla E; Rich, Elizabeth D; Brown, R Lane; Varnum, Michael D

2012-01-01

Photoreceptor cyclic nucleotide-gated (CNG) channels are the principal ion channels responsible for transduction of the light-induced change in cGMP concentration into an electrical signal. The ligand sensitivity of photoreceptor CNG channels is subject to regulation by intracellular signaling effectors, including calcium-calmodulin, tyrosine kinases and phosphoinositides. Little is known, however, about regulation of channel activity by modification to extracellular regions of CNG channel subunits. Extracellular proteases MMP9 and -2 are present in the interphotoreceptor matrix adjacent to photoreceptor outer segments. Given that MMPs have been implicated in retinal dysfunction and degeneration, we hypothesized that MMP activity may alter the functional properties of photoreceptor CNG channels. For heterologously expressed rod and cone CNG channels, extracellular exposure to MMPs dramatically increased the apparent affinity for cGMP and the efficacy of cAMP. These changes to ligand sensitivity were not prevented by destabilization of the actin cytoskeleton or by disruption of integrin mediated cell adhesion, but could be attenuated by inhibition of MMP catalytic activity. MMP-mediated gating changes exhibited saturable kinetic properties consistent with enzymatic processing of the CNG channels. In addition, exposure to MMPs decreased the abundance of full-length expressed CNGA3 subunits, with a concomitant increase in putative degradation products. Similar gating effects and apparent proteolysis were observed also for native rod photoreceptor CNG channels. Furthermore, constitutive apparent proteolysis of retinal CNGA1 and retinal MMP9 levels were both elevated in aged mice compared with young mice. Together, these results provide evidence that MMP-mediated proteolysis can regulate the ligand sensitivity of CNG channels.

Matrix metalloproteinase-9 and -2 enhance the ligand sensitivity of photoreceptor cyclic nucleotide-gated channels

PubMed Central

Meighan, Peter C.; Meighan, Starla E.; Rich, Elizabeth D.; Brown, R. Lane; Varnum, Michael D.

2012-01-01

Photoreceptor cyclic nucleotide-gated (CNG) channels are the principal ion channels responsible for transduction of the light-induced change in cGMP concentration into an electrical signal. The ligand sensitivity of photoreceptor CNG channels is subject to regulation by intracellular signaling effectors, including calcium-calmodulin, tyrosine kinases and phosphoinositides. Little is known, however, about regulation of channel activity by modification to extracellular regions of CNG channel subunits. Extracellular proteases MMP9 and -2 are present in the interphotoreceptor matrix adjacent to photoreceptor outer segments. Given that MMPs have been implicated in retinal dysfunction and degeneration, we hypothesized that MMP activity may alter the functional properties of photoreceptor CNG channels. For heterologously expressed rod and cone CNG channels, extracellular exposure to MMPs dramatically increased the apparent affinity for cGMP and the efficacy of cAMP. These changes to ligand sensitivity were not prevented by destabilization of the actin cytoskeleton or by disruption of integrin mediated cell adhesion, but could be attenuated by inhibition of MMP catalytic activity. MMP-mediated gating changes exhibited saturable kinetic properties consistent with enzymatic processing of the CNG channels. In addition, exposure to MMPs decreased the abundance of full-length expressed CNGA3 subunits, with a concomitant increase in putative degradation products. Similar gating effects and apparent proteolysis were observed also for native rod photoreceptor CNG channels. Furthermore, constitutive apparent proteolysis of retinal CNGA1 and retinal MMP9 levels were both elevated in aged mice compared with young mice. Together, these results provide evidence that MMP-mediated proteolysis can regulate the ligand sensitivity of CNG channels. PMID:22699690

Non-Equilibrium Dynamics Contribute to Ion Selectivity in the KcsA Channel

PubMed Central

Haas, Stephan; Farley, Robert A.

2014-01-01

The ability of biological ion channels to conduct selected ions across cell membranes is critical for the survival of both animal and bacterial cells. Numerous investigations of ion selectivity have been conducted over more than 50 years, yet the mechanisms whereby the channels select certain ions and reject others are not well understood. Here we report a new application of Jarzynski’s Equality to investigate the mechanism of ion selectivity using non-equilibrium molecular dynamics simulations of Na+ and K+ ions moving through the KcsA channel. The simulations show that the selectivity filter of KcsA adapts and responds to the presence of the ions with structural rearrangements that are different for Na+ and K+. These structural rearrangements facilitate entry of K+ ions into the selectivity filter and permeation through the channel, and rejection of Na+ ions. A mechanistic model of ion selectivity by this channel based on the results of the simulations relates the structural rearrangement of the selectivity filter to the differential dehydration of ions and multiple-ion occupancy and describes a mechanism to efficiently select and conduct K+. Estimates of the K+/Na+ selectivity ratio and steady state ion conductance for KcsA from the simulations are in good quantitative agreement with experimental measurements. This model also accurately describes experimental observations of channel block by cytoplasmic Na+ ions, the “punch through” relief of channel block by cytoplasmic positive voltages, and is consistent with the knock-on mechanism of ion permeation. PMID:24465882

Ion concentrations and velocity profiles in nanochannel electroosmotic flows

NASA Astrophysics Data System (ADS)

Qiao, R.; Aluru, N. R.

2003-03-01

Ion distributions and velocity profiles for electroosmotic flow in nanochannels of different widths are studied in this paper using molecular dynamics and continuum theory. For the various channel widths studied in this paper, the ion distribution near the channel wall is strongly influenced by the finite size of the ions and the discreteness of the solvent molecules. The classical Poisson-Boltzmann equation fails to predict the ion distribution near the channel wall as it does not account for the molecular aspects of the ion-wall and ion-solvent interactions. A modified Poisson-Boltzmann equation based on electrochemical potential correction is introduced to account for ion-wall and ion-solvent interactions. The electrochemical potential correction term is extracted from the ion distribution in a smaller channel using molecular dynamics. Using the electrochemical potential correction term extracted from molecular dynamics (MD) simulation of electroosmotic flow in a 2.22 nm channel, the modified Poisson-Boltzmann equation predicts the ion distribution in larger channel widths (e.g., 3.49 and 10.00 nm) with good accuracy. Detailed studies on the velocity profile in electro-osmotic flow indicate that the continuum flow theory can be used to predict bulk fluid flow in channels as small as 2.22 nm provided that the viscosity variation near the channel wall is taken into account. We propose a technique to embed the velocity near the channel wall obtained from MD simulation of electroosmotic flow in a narrow channel (e.g., 2.22 nm wide channel) into simulation of electroosmotic flow in larger channels. Simulation results indicate that such an approach can predict the velocity profile in larger channels (e.g., 3.49 and 10.00 nm) very well. Finally, simulation of electroosmotic flow in a 0.95 nm channel indicates that viscosity cannot be described by a local, linear constitutive relationship that the continuum flow theory is built upon and thus the continuum flow theory is not applicable for electroosmotic flow in such small channels.

Down regulated expression of the beta1 subunit of the big-conductance Ca2+ sensitive K+ channel in sphincter of Oddi cells from rabbits fed with a high cholesterol diet.

PubMed

Du, Pang; Cui, Guang-Bin; Wang, Ya-Rong; Zhang, Xiao-Yong; Ma, Ke-Jun; Wei, Jing-Guo

2006-12-01

Hypercholesterolemia, which is closely related to gallbladder bile stasis, can cause sphincter of Oddi dysfunction (SOD) by increasing the tension of sphincter of Oddi (SO). Intracellular calcium ion concentration ([Ca(2+)](i)) could influence the tension of SO. The beta1 subunit of the big-conductance Ca(2+) sensitive K(+) channel (BK(Ca)) can enhance the sensitivity of the BK(Ca) channel to [Ca(2+)](i). Absence and decline of the BKCa channel subunit beta1 could lead to many diseases. However, the relationship between hypercholesterolemia and the expression of beta1 subunit is not well understood. In this study, we successfully expressed and purified the rabbit BK(Ca) beta1 subunit protein and prepared its polyclonal antibody. The specificity of the prepared antibody was determined by Western blotting. A SOD rabbit model induced by a high cholesterol diet was established and the expression of the beta1 subunit of SO was determined by immunohistochemical staining and western blotting. Compared with the controls, our results demonstrated that hypercholesterolemia could decrease the expression of the beta1 subunit in the SO cells from rabbits. This indicates that lower expression of BKCa channel beta1 subunit might induce SOD.

Receptors, channels, and signalling in the urothelial sensory system in the bladder

PubMed Central

Merrill, Liana; Gonzalez, Eric J.; Girard, Beatrice M.; Vizzard, Margaret A.

2017-01-01

The storage and periodic elimination of urine, termed micturition, requires a complex neural control system to coordinate the activities of the urinary bladder, urethra, and urethral sphincters. At the level of the lumbosacral spinal cord, lower urinary tract reflex mechanisms are modulated by supraspinal controls with mechanosensory input from the urothelium, resulting in regulation of bladder contractile activity. The specific identity of the mechanical sensor is not yet known, but considerable interest exists in the contribution of transient receptor potential (TRP) channels to the mechanosensory functions of the urothelium. The sensory, transduction, and signalling properties of the urothelium can influence adjacent urinary bladder tissues including the suburothelial nerve plexus, interstitial cells of Cajal, and detrusor smooth muscle cells. Diverse stimuli, including those that activate TRP channels expressed by the urothelium, can influence urothelial release of chemical mediators (such as ATP). Changes to the urothelium are associated with a number of bladder pathologies that underlie urinary bladder dysfunction. Urothelial receptor and/or ion channel expression and the release of signalling molecules (such as ATP and nitric oxide) can be altered with bladder disease, neural injury, target organ inflammation, or psychogenic stress. Urothelial receptors and channels represent novel targets for potential therapies that are intended to modulate micturition function or bladder sensation. PMID:26926246

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

Ion Channel Gene Expression in Lung Adenocarcinoma: Potential Role in Prognosis and Diagnosis

PubMed Central

Ko, Jae-Hong; Gu, Wanjun; Lim, Inja; Bang, Hyoweon; Ko, Eun A.; Zhou, Tong

2014-01-01

Ion channels are known to regulate cancer processes at all stages. The roles of ion channels in cancer pathology are extremely diverse. We systematically analyzed the expression patterns of ion channel genes in lung adenocarcinoma. First, we compared the expression of ion channel genes between normal and tumor tissues in patients with lung adenocarcinoma. Thirty-seven ion channel genes were identified as being differentially expressed between the two groups. Next, we investigated the prognostic power of ion channel genes in lung adenocarcinoma. We assigned a risk score to each lung adenocarcinoma patient based on the expression of the differentially expressed ion channel genes. We demonstrated that the risk score effectively predicted overall survival and recurrence-free survival in lung adenocarcinoma. We also found that the risk scores for ever-smokers were higher than those for never-smokers. Multivariate analysis indicated that the risk score was a significant prognostic factor for survival, which is independent of patient age, gender, stage, smoking history, Myc level, and EGFR/KRAS/ALK gene mutation status. Finally, we investigated the difference in ion channel gene expression between the two major subtypes of non-small cell lung cancer: adenocarcinoma and squamous-cell carcinoma. Thirty ion channel genes were identified as being differentially expressed between the two groups. We suggest that ion channel gene expression can be used to improve the subtype classification in non-small cell lung cancer at the molecular level. The findings in this study have been validated in several independent lung cancer cohorts. PMID:24466154

Mechanisms defining the electrotonic potential abnormalities in simulated amyotrophic lateral sclerosis.

PubMed

Stephanova, D I; Krustev, S M; Negrev, N

2012-06-01

Electrotonic potentials allow the accommodative processes to polarizing stimuli to be assessed. Electrotonic potential transients in response to applied polarizing stimuli are caused by the kinetics of underlying axonal conductances. Here, we study these transients using our multi-layered model of the human motor nerve, in three simulated cases of the motor neuron disease amyotrophic lateral sclerosis (ALS): ALS1, ALS2 and ALS3 are three consecutively greater degrees of uniform axonal dysfunctions along the human motor nerve fibre. The results show that the responses in the ALS1 case are quite similar to the normal case. In contrast, in the ALS2 and ALS3 cases, long-lasting (100 ms) subthreshold depolarizing stimuli activate the classical "transient" Na(+) channels in the nodal and in the internodal axolemma beneath the myelin sheath; this leads to action potential generation during the early parts of the electrotonic responses in all compartments along the fibre length. The results also show that the electrotonic potentials in response to long-lasting (100 ms) subthreshold hyperpolarizing stimuli in the ALS1 and ALS2 cases are quiet similar to those of the normal case. However, the current kinetics in the ALS3 case differs from the normal case after the termination of the long-lasting hyperpolarizing stimuli. In the most abnormal ALS3 case, the activation of the Na(+) channels in the nodal and in the internodal axolemma leads to repetitive action potential generation in the late parts (100-200 ms) of the hyperpolarizing electrotonic responses. The results show that the repetitive firing, due to the progressively increased nodal and internodal ion channel dysfunction, are consistent with the loss of functional potassium channels involving both the fast and the slow potassium channel types. The results confirm that the electrotonic potentials in the three simulated ALS cases are specific indicators for the motor neuron disease ALS. The mechanisms underlying the simulated ALS are also discussed.

Cardiac ion channels

PubMed Central

Priest, Birgit T; McDermott, Jeff S

2015-01-01

Ion channels are critical for all aspects of cardiac function, including rhythmicity and contractility. Consequently, ion channels are key targets for therapeutics aimed at cardiac pathophysiologies such as atrial fibrillation or angina. At the same time, off-target interactions of drugs with cardiac ion channels can be the cause of unwanted side effects. This manuscript aims to review the physiology and pharmacology of key cardiac ion channels. The intent is to highlight recent developments for therapeutic development, as well as elucidate potential mechanisms for drug-induced cardiac side effects, rather than present an in-depth review of each channel subtype. PMID:26556552

CFTR dysfunction in cystic fibrosis and chronic obstructive pulmonary disease.

PubMed

Fernandez Fernandez, Elena; de Santi, Chiara; De Rose, Virginia; Greene, Catherine M

2018-05-11

Obstructive lung diseases such as cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) are causes of high morbidity and mortality worldwide. CF is a multiorgan genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene and is characterized by progressive chronic obstructive lung disease. Most cases of COPD are a result of noxious particles, mainly cigarette smoke but also other environmental pollutants. Areas covered: Although the pathogenesis and pathophysiology of CF and COPD differ, they do share key phenotypic features and because of these similarities there is great interest in exploring common mechanisms and/or factors affected by CFTR mutations and environmental insults involved in COPD. Various molecular, cellular and clinical studies have confirmed that CFTR protein dysfunction is common in both the CF and COPD airways. This review provides an update of our understanding of the role of dysfunctional CFTR in both respiratory diseases. Expert Commentary: Drugs developed for people with CF to improve mutant CFTR function and enhance CFTR ion channel activity might also be beneficial in patients with COPD. A move toward personalized therapy using, for example, microRNA modulators in conjunction with CFTR potentiators or correctors, could enhance treatment of both diseases.

Mapping the function of neuronal ion channels in model and experiment

PubMed Central

Podlaski, William F; Seeholzer, Alexander; Groschner, Lukas N; Miesenböck, Gero; Ranjan, Rajnish; Vogels, Tim P

2017-01-01

Ion channel models are the building blocks of computational neuron models. Their biological fidelity is therefore crucial for the interpretation of simulations. However, the number of published models, and the lack of standardization, make the comparison of ion channel models with one another and with experimental data difficult. Here, we present a framework for the automated large-scale classification of ion channel models. Using annotated metadata and responses to a set of voltage-clamp protocols, we assigned 2378 models of voltage- and calcium-gated ion channels coded in NEURON to 211 clusters. The IonChannelGenealogy (ICGenealogy) web interface provides an interactive resource for the categorization of new and existing models and experimental recordings. It enables quantitative comparisons of simulated and/or measured ion channel kinetics, and facilitates field-wide standardization of experimentally-constrained modeling. DOI: http://dx.doi.org/10.7554/eLife.22152.001 PMID:28267430

Calcineurin Dysregulation Underlies Spinal Cord Injury-Induced K+ Channel Dysfunction in DRG Neurons.

PubMed

Zemel, Benjamin M; Muqeem, Tanziyah; Brown, Eric V; Goulão, Miguel; Urban, Mark W; Tymanskyj, Stephen R; Lepore, Angelo C; Covarrubias, Manuel

2017-08-23

Dysfunction of the fast-inactivating Kv3.4 potassium current in dorsal root ganglion (DRG) neurons contributes to the hyperexcitability associated with persistent pain induced by spinal cord injury (SCI). However, the underlying mechanism is not known. In light of our previous work demonstrating modulation of the Kv3.4 channel by phosphorylation, we investigated the role of the phosphatase calcineurin (CaN) using electrophysiological, molecular, and imaging approaches in adult female Sprague Dawley rats. Pharmacological inhibition of CaN in small-diameter DRG neurons slowed repolarization of the somatic action potential (AP) and attenuated the Kv3.4 current. Attenuated Kv3.4 currents also exhibited slowed inactivation. We observed similar effects on the recombinant Kv3.4 channel heterologously expressed in Chinese hamster ovary cells, supporting our findings in DRG neurons. Elucidating the molecular basis of these effects, mutation of four previously characterized serines within the Kv3.4 N-terminal inactivation domain eliminated the effects of CaN inhibition on the Kv3.4 current. SCI similarly induced concurrent Kv3.4 current attenuation and slowing of inactivation. Although there was little change in CaN expression and localization after injury, SCI induced upregulation of the native regulator of CaN 1 (RCAN1) in the DRG at the transcript and protein levels. Consistent with CaN inhibition resulting from RCAN1 upregulation, overexpression of RCAN1 in naive DRG neurons recapitulated the effects of pharmacological CaN inhibition on the Kv3.4 current and the AP. Overall, these results demonstrate a novel regulatory pathway that links CaN, RCAN1, and Kv3.4 in DRG neurons. Dysregulation of this pathway might underlie a peripheral mechanism of pain sensitization induced by SCI. SIGNIFICANCE STATEMENT Pain sensitization associated with spinal cord injury (SCI) involves poorly understood maladaptive modulation of neuronal excitability. Although central mechanisms have received significant attention, recent studies have identified peripheral nerve hyperexcitability as a driver of persistent pain signaling after SCI. However, the ion channels and signaling molecules responsible for this change in primary sensory neuron excitability are still not well defined. To address this problem, this study used complementary electrophysiological and molecular methods to determine how Kv3.4, a voltage-gated K + channel robustly expressed in dorsal root ganglion neurons, becomes dysfunctional upon calcineurin (CaN) inhibition. The results strongly suggest that CaN inhibition underlies SCI-induced dysfunction of Kv3.4 and the associated excitability changes through upregulation of the native regulator of CaN 1 (RCAN1). Copyright © 2017 the authors 0270-6474/17/378257-17$15.00/0.

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

Ion channel gene expression predicts survival in glioma patients

PubMed Central

Wang, Rong; Gurguis, Christopher I.; Gu, Wanjun; Ko, Eun A; Lim, Inja; Bang, Hyoweon; Zhou, Tong; Ko, Jae-Hong

2015-01-01

Ion channels are important regulators in cell proliferation, migration, and apoptosis. The malfunction and/or aberrant expression of ion channels may disrupt these important biological processes and influence cancer progression. In this study, we investigate the expression pattern of ion channel genes in glioma. We designate 18 ion channel genes that are differentially expressed in high-grade glioma as a prognostic molecular signature. This ion channel gene expression based signature predicts glioma outcome in three independent validation cohorts. Interestingly, 16 of these 18 genes were down-regulated in high-grade glioma. This signature is independent of traditional clinical, molecular, and histological factors. Resampling tests indicate that the prognostic power of the signature outperforms random gene sets selected from human genome in all the validation cohorts. More importantly, this signature performs better than the random gene signatures selected from glioma-associated genes in two out of three validation datasets. This study implicates ion channels in brain cancer, thus expanding on knowledge of their roles in other cancers. Individualized profiling of ion channel gene expression serves as a superior and independent prognostic tool for glioma patients. PMID:26235283

Detection of single ion channel activity with carbon nanotubes

NASA Astrophysics Data System (ADS)

Zhou, Weiwei; Wang, Yung Yu; Lim, Tae-Sun; Pham, Ted; Jain, Dheeraj; Burke, Peter J.

2015-03-01

Many processes in life are based on ion currents and membrane voltages controlled by a sophisticated and diverse family of membrane proteins (ion channels), which are comparable in size to the most advanced nanoelectronic components currently under development. Here we demonstrate an electrical assay of individual ion channel activity by measuring the dynamic opening and closing of the ion channel nanopores using single-walled carbon nanotubes (SWNTs). Two canonical dynamic ion channels (gramicidin A (gA) and alamethicin) and one static biological nanopore (α-hemolysin (α-HL)) were successfully incorporated into supported lipid bilayers (SLBs, an artificial cell membrane), which in turn were interfaced to the carbon nanotubes through a variety of polymer-cushion surface functionalization schemes. The ion channel current directly charges the quantum capacitance of a single nanotube in a network of purified semiconducting nanotubes. This work forms the foundation for a scalable, massively parallel architecture of 1d nanoelectronic devices interrogating electrophysiology at the single ion channel level.

Conductivity noise in transmembrane ion channels due to ion concentration fluctuations via diffusion.

PubMed

Mak, D O; Webb, W W

1997-03-01

A Green's function approach is developed from first principles to evaluate the power spectral density of conductance fluctuations caused by ion concentration fluctuations via diffusion in an electrolyte system. This is applied to simple geometric models of transmembrane ion channels to obtain an estimate of the magnitude of ion concentration fluctuation noise in the channel current. Pure polypeptide alamethicin forms stable ion channels with multiple conductance states in artificial phospholipid bilayers isolated onto tips of micropipettes with gigaohm seals. In the single-channel current recorded by voltage-clamp techniques, excess noise was found after the background instrumental noise and the intrinsic Johnson and shot noises were removed. The noise que to ion concentration fluctuations via diffusion was isolated by the dependence of the excess current noise on buffer ion concentration. The magnitude of the concentration fluctuation noise derived from experimental data lies within limits estimated using our simple geometric channel models. Variation of the noise magnitude for alamethicin channels in various conductance states agrees with theoretical prediction.

Characterization of Membrane Patch-Ion Channel Probes for Scanning Ion Conductance Microscopy.

PubMed

Shi, Wenqing; Zeng, Yuhan; Zhu, Cheng; Xiao, Yucheng; Cummins, Theodore R; Hou, Jianghui; Baker, Lane A

2018-05-01

Integration of dual-barrel membrane patch-ion channel probes (MP-ICPs) to scanning ion conductance microscopy (SICM) holds promise of providing a revolutionized approach of spatially resolved chemical sensing. A series of experiments are performed to further the understanding of the system and to answer some fundamental questions, in preparation for future developments of this approach. First, MP-ICPs are constructed that contain different types of ion channels including transient receptor potential vanilloid 1 and large conductance Ca2 + -activated K + channels to establish the generalizability of the methods. Next, the capability of the MP-ICP platforms in single ion channel activity measurements is proved. In addition, the interplay between the SICM barrel and the ICP barrel is studied. For ion channels gated by uncharged ligands, channel activity at the ICP barrel is unaffected by the SICM barrel potential; whereas for ion channels that are gated by charged ligands, enhanced channel activity can be obtained by biasing the SICM barrel at potentials with opposite polarity to the charge of the ligand molecules. Finally, a proof-of-principle experiment is performed and site-specific molecular/ionic flux sensing is demonstrated at single-ion-channel level, which show that the MP-ICP platform can be used to quantify local molecular/ionic concentrations. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Ion channel expression patterns in glioblastoma stem cells with functional and therapeutic implications for malignancy.

PubMed

Pollak, Julia; Rai, Karan G; Funk, Cory C; Arora, Sonali; Lee, Eunjee; Zhu, Jun; Price, Nathan D; Paddison, Patrick J; Ramirez, Jan-Marino; Rostomily, Robert C

2017-01-01

Ion channels and transporters have increasingly recognized roles in cancer progression through the regulation of cell proliferation, migration, and death. Glioblastoma stem-like cells (GSCs) are a source of tumor formation and recurrence in glioblastoma multiforme, a highly aggressive brain cancer, suggesting that ion channel expression may be perturbed in this population. However, little is known about the expression and functional relevance of ion channels that may contribute to GSC malignancy. Using RNA sequencing, we assessed the enrichment of ion channels in GSC isolates and non-tumor neural cell types. We identified a unique set of GSC-enriched ion channels using differential expression analysis that is also associated with distinct gene mutation signatures. In support of potential clinical relevance, expression of selected GSC-enriched ion channels evaluated in human glioblastoma databases of The Cancer Genome Atlas and Ivy Glioblastoma Atlas Project correlated with patient survival times. Finally, genetic knockdown as well as pharmacological inhibition of individual or classes of GSC-enriched ion channels constrained growth of GSCs compared to normal neural stem cells. This first-in-kind global examination characterizes ion channels enriched in GSCs and explores their potential clinical relevance to glioblastoma molecular subtypes, gene mutations, survival outcomes, regional tumor expression, and experimental responses to loss-of-function. Together, the data support the potential biological and therapeutic impact of ion channels on GSC malignancy and provide strong rationale for further examination of their mechanistic and therapeutic importance.

Ion channel expression patterns in glioblastoma stem cells with functional and therapeutic implications for malignancy

PubMed Central

Pollak, Julia; Rai, Karan G.; Funk, Cory C.; Arora, Sonali; Lee, Eunjee; Zhu, Jun; Price, Nathan D.; Paddison, Patrick J.; Ramirez, Jan-Marino; Rostomily, Robert C.

2017-01-01

Ion channels and transporters have increasingly recognized roles in cancer progression through the regulation of cell proliferation, migration, and death. Glioblastoma stem-like cells (GSCs) are a source of tumor formation and recurrence in glioblastoma multiforme, a highly aggressive brain cancer, suggesting that ion channel expression may be perturbed in this population. However, little is known about the expression and functional relevance of ion channels that may contribute to GSC malignancy. Using RNA sequencing, we assessed the enrichment of ion channels in GSC isolates and non-tumor neural cell types. We identified a unique set of GSC-enriched ion channels using differential expression analysis that is also associated with distinct gene mutation signatures. In support of potential clinical relevance, expression of selected GSC-enriched ion channels evaluated in human glioblastoma databases of The Cancer Genome Atlas and Ivy Glioblastoma Atlas Project correlated with patient survival times. Finally, genetic knockdown as well as pharmacological inhibition of individual or classes of GSC-enriched ion channels constrained growth of GSCs compared to normal neural stem cells. This first-in-kind global examination characterizes ion channels enriched in GSCs and explores their potential clinical relevance to glioblastoma molecular subtypes, gene mutations, survival outcomes, regional tumor expression, and experimental responses to loss-of-function. Together, the data support the potential biological and therapeutic impact of ion channels on GSC malignancy and provide strong rationale for further examination of their mechanistic and therapeutic importance. PMID:28264064

Organic toxins as tools to understand ion channel mechanisms and structure.

PubMed

Morales-Lázaro, Sara Luz; Hernández-García, Enrique; Serrano-Flores, Barbara; Rosenbaum, Tamara

2015-01-01

Ion channels constitute a varied class of membrane proteins with pivotal roles in cellular physiology and that are fundamental for neuronal signaling, hormone secretion and muscle contractility. Hence, it is not unanticipated that toxins from diverse organisms have evolved to modulate the activity of ion channels. For instance, animals such as cone snails, scorpions, spiders and snakes use toxins to immobilize and capture their prey by affecting ion channel function. This is a beautiful example of an evolutionary process that has led to the development of an injection apparatus from predators and to the existence of toxins with high affinity and specificity for a given target. Toxins have been used in the field of ion channel biophysics for several decades to gain insight into the gating mechanisms and the structure of ion channels. Through the use of these peptides, much has been learned about the ion conduction pathways, voltage-sensing mechanisms, pore sizes, kinetics, inactivation processes, etc. This review examines an assortment of toxins that have been used to study different ion channels and describes some key findings about the structure-function relationships in these proteins through the details of the toxin-ion channel interactions.

Mechanically Activated Ion Channels

PubMed Central

Ranade, Sanjeev S.; Syeda, Ruhma; Patapoutian, Ardem

2015-01-01

Mechanotransduction, the conversion of physical forces into biochemical signals, is an essential component of numerous physiological processes including not only conscious senses of touch and hearing, but also unconscious senses such as blood pressure regulation. Mechanically activated (MA) ion channels have been proposed as sensors of physical force, but the identity of these channels and an understanding of how mechanical force is transduced has remained elusive. A number of recent studies on previously known ion channels along with the identification of novel MA ion channels have greatly transformed our understanding of touch and hearing in both vertebrates and invertebrates. Here, we present an updated review of eukaryotic ion channel families that have been implicated in mechanotransduction processes and evaluate the qualifications of the candidate genes according to specified criteria. We then discuss the proposed gating models for MA ion channels and highlight recent structural studies of mechanosensitive potassium channels. PMID:26402601

Systematic analysis of the contributions of stochastic voltage gated channels to neuronal noise

PubMed Central

O'Donnell, Cian; van Rossum, Mark C. W.

2014-01-01

Electrical signaling in neurons is mediated by the opening and closing of large numbers of individual ion channels. The ion channels' state transitions are stochastic and introduce fluctuations in the macroscopic current through ion channel populations. This creates an unavoidable source of intrinsic electrical noise for the neuron, leading to fluctuations in the membrane potential and spontaneous spikes. While this effect is well known, the impact of channel noise on single neuron dynamics remains poorly understood. Most results are based on numerical simulations. There is no agreement, even in theoretical studies, on which ion channel type is the dominant noise source, nor how inclusion of additional ion channel types affects voltage noise. Here we describe a framework to calculate voltage noise directly from an arbitrary set of ion channel models, and discuss how this can be use to estimate spontaneous spike rates. PMID:25360105

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

Nootropic α7 nicotinic receptor allosteric modulator derived from GABAA receptor modulators

PubMed Central

Ng, Herman J.; Whittemore, Edward R.; Tran, Minhtam B.; Hogenkamp, Derk J.; Broide, Ron S.; Johnstone, Timothy B.; Zheng, Lijun; Stevens, Karen E.; Gee, Kelvin W.

2007-01-01

Activation of brain α7 nicotinic acetylcholine receptors (α7 nAChRs) has broad therapeutic potential in CNS diseases related to cognitive dysfunction, including Alzheimer's disease and schizophrenia. In contrast to direct agonist activation, positive allosteric modulation of α7 nAChRs would deliver the clinically validated benefits of allosterism to these indications. We have generated a selective α7 nAChR-positive allosteric modulator (PAM) from a library of GABAA receptor PAMs. Compound 6 (N-(4-chlorophenyl)-α-[[(4-chloro-phenyl)amino]methylene]-3-methyl-5-isoxazoleacet-amide) evokes robust positive modulation of agonist-induced currents at α7 nAChRs, while preserving the rapid native characteristics of desensitization, and has little to no efficacy at other ligand-gated ion channels. In rodent models, it corrects sensory-gating deficits and improves working memory, effects consistent with cognitive enhancement. Compound 6 represents a chemotype for allosteric activation of α7 nAChRs, with therapeutic potential in CNS diseases with cognitive dysfunction. PMID:17470817

The electronic stopping powers and angular energy-loss dependence of helium and lithium ions in the silicon crystal

NASA Astrophysics Data System (ADS)

Mikšová, R.; Macková, A.; Malinský, P.

2017-09-01

We have measured the electronic stopping powers of helium and lithium ions in the channelling direction of the Si〈1 0 0〉 crystal. The energy range used (2.0-8.0 MeV) was changed by 200 and 400-keV steps. The ratio α between the channelling and random stopping powers was determined as a function of the angle for 2, 3 and 4 MeV 4He+ ions and for 3 and 6 MeV 7Li+,2+ ions. The measurements were carried out using the Rutherford backscattering spectrometry in the channelling mode (RBS-C) in a silicon-on-insulator material. The experimental channelling stopping-power values measured in the channelling direction were then discussed in the frame of the random energy stopping predictions calculated using SRIM-2013 code and the theoretical unitary convolution approximation (UCA) model. The experimental channelling stopping-power values decrease with increasing ion energy. The stopping-power difference between channelled and randomly moving ions increases with the enhanced initial ion energy. The ratio between the channelling and random ion stopping powers α as a function of the ion beam incoming angle for 2, 3 and 4 MeV He+ ions and for 3 and 6 MeV Li+,2+ ions was observed in the range 0.5-1.

High throughput screening technologies for ion channels

PubMed Central

Yu, Hai-bo; Li, Min; Wang, Wei-ping; Wang, Xiao-liang

2016-01-01

Ion channels are involved in a variety of fundamental physiological processes, and their malfunction causes numerous human diseases. Therefore, ion channels represent a class of attractive drug targets and a class of important off-targets for in vitro pharmacological profiling. In the past decades, the rapid progress in developing functional assays and instrumentation has enabled high throughput screening (HTS) campaigns on an expanding list of channel types. Chronologically, HTS methods for ion channels include the ligand binding assay, flux-based assay, fluorescence-based assay, and automated electrophysiological assay. In this review we summarize the current HTS technologies for different ion channel classes and their applications. PMID:26657056

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 ) .

Ion channels versus ion pumps: the principal difference, in principle.

PubMed

Gadsby, David C

2009-05-01

The incessant traffic of ions across cell membranes is controlled by two kinds of border guards: ion channels and ion pumps. Open channels let selected ions diffuse rapidly down electrical and concentration gradients, whereas ion pumps labour tirelessly to maintain the gradients by consuming energy to slowly move ions thermodynamically uphill. Because of the diametrically opposed tasks and the divergent speeds of channels and pumps, they have traditionally been viewed as completely different entities, as alike as chalk and cheese. But new structural and mechanistic information about both of these classes of molecular machines challenges this comfortable separation and forces its re-evaluation.

Cirrhotic cardiomyopathy

PubMed Central

Ruiz-del-Árbol, Luis; Serradilla, Regina

2015-01-01

During the course of cirrhosis, there is a progressive deterioration of cardiac function manifested by the disappearance of the hyperdynamic circulation due to a failure in heart function with decreased cardiac output. This is due to a deterioration in inotropic and chronotropic function which takes place in parallel with a diastolic dysfunction and cardiac hypertrophy in the absence of other known cardiac disease. Other findings of this specific cardiomyopathy include impaired contractile responsiveness to stress stimuli and electrophysiological abnormalities with prolonged QT interval. The pathogenic mechanisms of cirrhotic cardiomyopathy include impairment of the b-adrenergic receptor signalling, abnormal cardiomyocyte membrane lipid composition and biophysical properties, ion channel defects and overactivity of humoral cardiodepressant factors. Cirrhotic cardiomyopathy may be difficult to determine due to the lack of a specific diagnosis test. However, an echocardiogram allows the detection of the diastolic dysfunction and the E/e′ ratio may be used in the follow-up progression of the illness. Cirrhotic cardiomyopathy plays an important role in the pathogenesis of the impairment of effective arterial blood volume and correlates with the degree of liver failure. A clinical consequence of cardiac dysfunction is an inadequate cardiac response in the setting of vascular stress that may result in renal hypoperfusion leading to renal failure. The prognosis is difficult to establish but the severity of diastolic dysfunction may be a marker of mortality risk. Treatment is non-specific and liver transplantation may normalize the cardiac function. PMID:26556983

Structure, function and translational relevance of aquaporin dual water and ion channels.

PubMed

Yool, Andrea J; Campbell, Ewan M

2012-01-01

Aquaporins have been assumed to be selective for water alone, and aquaglyceroporins are accepted as carrying water and small uncharged solutes including glycerol. This review presents an expanded view of aquaporins as channels with more complex mechanisms of regulation and diverse repertoires of substrate permeabilities than were originally appreciated in the early establishment of the field. The role of aquaporins as dual water and gated ion channels is likely to have physiological and potentially translational relevance, and can be evaluated with newly developed molecular and pharmacological tools. Ion channel activity has been shown for Aquaporins -0, -1, and -6, Drosphila Big Brain, and plant Nodulin-26. Although the concept of ion channel function in aquaporins remains controversial, research advances are beginning to define not only the ion channel function but also the detailed molecular mechanisms that govern and mediate the multifunctional capabilities. With regard to physiological relevance, the adaptive benefit of expression of ion channel activity in aquaporins, implied by amino acid sequence conservation of the ion channel gating domains, suggests they provide more than water or glycerol and solute transport. Dual ion and water channels are of interest for understanding the modulation of transmembrane fluid gradients, volume regulation, and possible signal transduction in tissues expressing classes of aquaporins that have the dual function capability. Other aquaporin classes might be found in future work to have ion channel activities, pending identification of the possible signaling pathways that could govern activation. Copyright © 2012 Elsevier Ltd. All rights reserved.

Mitochondrial Ion Channels/Transporters as Sensors and Regulators of Cellular Redox Signaling

PubMed Central

Ryu, Shin-Young; Jhun, Bong Sook; Hurst, Stephen

2014-01-01

Abstract Significance: Mitochondrial ion channels/transporters and the electron transport chain (ETC) serve as key sensors and regulators for cellular redox signaling, the production of reactive oxygen species (ROS) and nitrogen species (RNS) in mitochondria, and balancing cell survival and death. Although the functional and pharmacological characteristics of mitochondrial ion transport mechanisms have been extensively studied for several decades, the majority of the molecular identities that are responsible for these channels/transporters have remained a mystery until very recently. Recent Advances: Recent breakthrough studies uncovered the molecular identities of the diverse array of major mitochondrial ion channels/transporters, including the mitochondrial Ca2+ uniporter pore, mitochondrial permeability transition pore, and mitochondrial ATP-sensitive K+ channel. This new information enables us to form detailed molecular and functional characterizations of mitochondrial ion channels/transporters and their roles in mitochondrial redox signaling. Critical Issues: Redox-mediated post-translational modifications of mitochondrial ion channels/transporters and ETC serve as key mechanisms for the spatiotemporal control of mitochondrial ROS/RNS generation. Future Directions: Identification of detailed molecular mechanisms for redox-mediated regulation of mitochondrial ion channels will enable us to find novel therapeutic targets for many diseases that are associated with cellular redox signaling and mitochondrial ion channels/transporters. Antioxid. Redox Signal. 21, 987–1006. PMID:24180309

Marine Toxins Targeting Ion Channels

PubMed Central

Arias, Hugo R.

2006-01-01

This introductory minireview points out the importance of ion channels for cell communication. The basic concepts on the structure and function of ion channels triggered by membrane voltage changes, the so-called voltage-gated ion channels (VGICs), as well as those activated by neurotransmitters, the so-called ligand-gated ion channel (LGICs), are introduced. Among the most important VGIC superfamiles, we can name the voltage-gated Na+ (NaV), Ca2+ (CaV), and K+ (KV) channels. Among the most important LGIC super families, we can include the Cys-loop or nicotinicoid, the glutamate-activated (GluR), and the ATP-activated (P2XnR) receptor superfamilies. Ion channels are transmembrane proteins that allow the passage of different ions in a specific or unspecific manner. For instance, the activation of NaV, CaV, or KV channels opens a pore that is specific for Na+, Ca2+, or K+, respectively. On the other hand, the activation of certain LGICs such as nicotinic acetylcholine receptors, GluRs, and P2XnRs allows the passage of cations (e.g., Na+, K+, and/or Ca2+), whereas the activation of other LGICs such as type A γ-butyric acid and glycine receptors allows the passage of anions (e.g., Cl− and/or HCO3−). In this regard, the activation of NaV and CaV as well as ligand-gated cation channels produce membrane depolarization, which finally leads to stimulatory effects in the cell, whereas the activation of KV as well as ligand-gated anion channels induce membrane hyperpolarization that finally leads to inhibitory effects in the cell. The importance of these ion channel superfamilies is emphasized by considering their physiological functions throughout the body as well as their pathophysiological implicance in several neuronal diseases. In this regard, natural molecules, and especially marine toxins, can be potentially used as modulators (e.g., inhibitors or prolongers) of ion channel functions to treat or to alleviate a specific ion channel-linked disease (e.g., channelopaties).

Acid stress mediated adaptive divergence in ion channel function during embryogenesis in Rana arvalis

PubMed Central

Shu, Longfei; Laurila, Anssi; Räsänen, Katja

2015-01-01

Ion channels and pumps are responsible for ion flux in cells, and are key mechanisms mediating cellular function. Many environmental stressors, such as salinity and acidification, are known to severely disrupt ionic balance of organisms thereby challenging fitness of natural populations. Although ion channels can have several vital functions during early life-stages (e.g. embryogenesis), it is currently not known i) how developing embryos maintain proper intracellular conditions when exposed to environmental stress and ii) to what extent environmental stress can drive intra-specific divergence in ion channels. Here we studied the moor frog, Rana arvalis, from three divergent populations to investigate the role of different ion channels and pumps for embryonic survival under acid stress (pH 4 vs 7.5) and whether populations adapted to contrasting acidities differ in the relative role of different ion channel/pumps. We found that ion channels that mediate Ca2+ influx are essential for embryonic survival under acidic pH, and, intriguingly, that populations differ in calcium channel function. Our results suggest that adaptive divergence in embryonic acid stress tolerance of amphibians may in part be mediated by Ca2+ balance. We suggest that ion flux may mediate adaptive divergence of natural populations at early life-stages in the face of environmental stress. PMID:26381453

QPatch: the missing link between HTS and ion channel drug discovery.

PubMed

Mathes, Chris; Friis, Søren; Finley, Michael; Liu, Yi

2009-01-01

The conventional patch clamp has long been considered the best approach for studying ion channel function and pharmacology. However, its low throughput has been a major hurdle to overcome for ion channel drug discovery. The recent emergence of higher throughput, automated patch clamp technology begins to break this bottleneck by providing medicinal chemists with high-quality, information-rich data in a more timely fashion. As such, these technologies have the potential to bridge a critical missing link between high-throughput primary screening and meaningful ion channel drug discovery programs. One of these technologies, the QPatch automated patch clamp system developed by Sophion Bioscience, records whole-cell ion channel currents from 16 or 48 individual cells in a parallel fashion. Here, we review the general applicability of the QPatch to studying a wide variety of ion channel types (voltage-/ligand-gated cationic/anionic channels) in various expression systems. The success rate of gigaseals, formation of the whole-cell configuration and usable cells ranged from 40-80%, depending on a number of factors including the cell line used, ion channel expressed, assay development or optimization time and expression level in these studies. We present detailed analyses of the QPatch features and results in case studies in which secondary screening assays were successfully developed for a voltage-gated calcium channel and a ligand-gated TRP channel. The increase in throughput compared to conventional patch clamp with the same cells was approximately 10-fold. We conclude that the QPatch, combining high data quality and speed with user friendliness and suitability for a wide array of ion channels, resides on the cutting edge of automated patch clamp technology and plays a pivotal role in expediting ion channel drug discovery.

TRPP2 ion channels: Critical regulators of organ morphogenesis in health and disease.

PubMed

Busch, Tilman; Köttgen, Michael; Hofherr, Alexis

2017-09-01

Ion channels control the membrane potential and mediate transport of ions across membranes. Archetypical physiological functions of ion channels include processes such as regulation of neuronal excitability, muscle contraction, or transepithelial ion transport. In that regard, transient receptor potential ion channel polycystin 2 (TRPP2) is remarkable, because it controls complex morphogenetic processes such as the establishment of properly shaped epithelial tubules and left-right-asymmetry of organs. The fascinating question of how an ion channel regulates morphogenesis has since captivated the attention of scientists in different disciplines. Four loosely connected key insights on different levels of biological complexity ranging from protein to whole organism have framed our understanding of TRPP2 physiology: 1) TRPP2 is a non-selective cation channel; 2) TRPP2 is part of a receptor-ion channel complex; 3) TRPP2 localizes to primary cilia; and 4) TRPP2 is required for organ morphogenesis. In this review, we will discuss the current knowledge in these key areas and highlight some of the challenges ahead. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Differential subcellular distribution of ion channels and the diversity of neuronal function.

PubMed

Nusser, Zoltan

2012-06-01

Following the astonishing molecular diversity of voltage-gated ion channels that was revealed in the past few decades, the ion channel repertoire expressed by neurons has been implicated as the major factor governing their functional heterogeneity. Although the molecular structure of ion channels is a key determinant of their biophysical properties, their subcellular distribution and densities on the surface of nerve cells are just as important for fulfilling functional requirements. Recent results obtained with high resolution quantitative localization techniques revealed complex, subcellular compartment-specific distribution patterns of distinct ion channels. Here I suggest that within a given neuron type every ion channel has a unique cell surface distribution pattern, with the functional consequence that this dramatically increases the computational power of nerve cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

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

The T1R2/T1R3 sweet receptor and TRPM5 ion channel taste targets with therapeutic potential.

PubMed

Sprous, Dennis; Palmer, Kyle R

2010-01-01

Taste signaling is a critical determinant of ingestive behaviors and thereby linked to obesity and related metabolic dysfunctions. Recent evidence of taste signaling pathways in the gut suggests the link to be more direct, raising the possibility that taste receptor systems could be regarded as therapeutic targets. T1R2/T1R3, the G protein coupled receptor that mediates sweet taste, and the TRPM5 ion channel have been the focus of discovery programs seeking novel compounds that could be useful in modifying taste. We review in this chapter the hypothesis of gastrointestinal taste signaling and discuss the potential for T1R2/T1R3 and TRPM5 as targets of therapeutic intervention in obesity and diabetes. Critical to the development of a drug discovery program is the creation of libraries that enhance the likelihood of identifying novel compounds that modulate the target of interest. We advocate a computer-based chemoinformatic approach for assembling natural and synthetic compound libraries as well as for supporting optimization of structure activity relationships. Strategies for discovering modulators of T1R2/T1R3 and TRPM5 using methods of chemoinformatics are presented herein. Copyright 2010 Elsevier Inc. All rights reserved.

Ion Trapping with Fast-Response Ion-Selective Microelectrodes Enhances Detection of Extracellular Ion Channel Gradients

PubMed Central

Messerli, Mark A.; Collis, Leon P.; Smith, Peter J.S.

2009-01-01

Previously, functional mapping of channels has been achieved by measuring the passage of net charge and of specific ions with electrophysiological and intracellular fluorescence imaging techniques. However, functional mapping of ion channels using extracellular ion-selective microelectrodes has distinct advantages over the former methods. We have developed this method through measurement of extracellular K+ gradients caused by efflux through Ca2+-activated K+ channels expressed in Chinese hamster ovary cells. We report that electrodes constructed with short columns of a mechanically stable K+-selective liquid membrane respond quickly and measure changes in local [K+] consistent with a diffusion model. When used in close proximity to the plasma membrane (<4 μm), the ISMs pose a barrier to simple diffusion, creating an ion trap. The ion trap amplifies the local change in [K+] without dramatically changing the rise or fall time of the [K+] profile. Measurement of extracellular K+ gradients from activated rSlo channels shows that rapid events, 10–55 ms, can be characterized. This method provides a noninvasive means for functional mapping of channel location and density as well as for characterizing the properties of ion channels in the plasma membrane. PMID:19217875

Calculating tracer currents through narrow ion channels: Beyond the independent particle model.

PubMed

Coalson, Rob D; Jasnow, David

2018-06-01

Discrete state models of single-file ion permeation through a narrow ion channel pore are employed to analyze the ratio of forward to backward tracer current. Conditions under which the well-known Ussing formula for this ratio hold are explored in systems where ions do not move independently through the channel. Building detailed balance into the rate constants for the model in such a way that under equilibrium conditions (equal rate of forward vs. backward permeation events) the Nernst Equation is satisfied, it is found that in a model where only one ion can occupy the channel at a time, the Ussing formula is always obeyed for any number of binding sites, reservoir concentrations of the ions and electric potential difference across the membrane which the ion channel spans, independent of the internal details of the permeation pathway. However, numerical analysis demonstrates that when multiple ions can occupy the channel at once, the nonequilibrium forward/backward tracer flux ratio deviates from the prediction of the Ussing model. Assuming an appropriate effective potential experienced by ions in the channel, we provide explicit formulae for the rate constants in these models. © 2018 IOP Publishing Ltd.

ModFossa: A library for modeling ion channels using Python.

PubMed

Ferneyhough, Gareth B; Thibealut, Corey M; Dascalu, Sergiu M; Harris, Frederick C

2016-06-01

The creation and simulation of ion channel models using continuous-time Markov processes is a powerful and well-used tool in the field of electrophysiology and ion channel research. While several software packages exist for the purpose of ion channel modeling, most are GUI based, and none are available as a Python library. In an attempt to provide an easy-to-use, yet powerful Markov model-based ion channel simulator, we have developed ModFossa, a Python library supporting easy model creation and stimulus definition, complete with a fast numerical solver, and attractive vector graphics plotting.

Ion-binding properties of the ClC chloride selectivity filter

PubMed Central

Lobet, Séverine; Dutzler, Raimund

2006-01-01

The ClC channels are members of a large protein family of chloride (Cl−) channels and secondary active Cl− transporters. Despite their diverse functions, the transmembrane architecture within the family is conserved. Here we present a crystallographic study on the ion-binding properties of the ClC selectivity filter in the close homolog from Escherichia coli (EcClC). The ClC selectivity filter contains three ion-binding sites that bridge the extra- and intracellular solutions. The sites bind Cl− ions with mM affinity. Despite their close proximity within the filter, the three sites can be occupied simultaneously. The ion-binding properties are found conserved from the bacterial transporter EcClC to the human Cl− channel ClC-1, suggesting a close functional link between ion permeation in the channels and active transport in the transporters. In resemblance to K+ channels, ions permeate the ClC channel in a single file, with mutual repulsion between the ions fostering rapid conduction. PMID:16341087

Simulation of the effect of a magnetically insulated anode on a low-power cylindrical Hall thruster

NASA Astrophysics Data System (ADS)

Yongjie, DING; Hong, LI; Boyang, JIA; Peng, LI; Liqiu, WEI; Yu, XU; Wuji, PENG; Hezhi, SUN; Yong, CAO; Daren, YU

2018-03-01

The intersection point of the characteristic magnetic field line (CMFL) crossing the anode boundary with the discharge channel wall, and its influence on thruster performance and the energy and flux of ions bombarding the channel wall, have been studied numerically. The simulation results demonstrate that with the increase in distance from the crossover point of the CMFL with the channel wall to the bottom of the thruster channel, the ionization rate in the discharge channel gradually increases; meanwhile, the ion energy and ion current density bombarding the channel wall decreases. When the point of the CMFL with the channel wall is at the channel outlet, the thrust, specific impulse, and efficiency are at a maximum, while the ion energy and ion current density bombarding the channel wall are at a minimum. Therefore, to improve the performance and lifetime of the thruster, it is important to control the point of intersection of the CMFL with the channel wall.

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.

Brownian dynamics study of ion transport in the vestibule of membrane channels.

PubMed

Li, S C; Hoyles, M; Kuyucak, S; Chung, S H

1998-01-01

Brownian dynamics simulations have been carried out to study the transport of ions in a vestibular geometry, which offers a more realistic shape for membrane channels than cylindrical tubes. Specifically, we consider a torus-shaped channel, for which the analytical solution of Poisson's equation is possible. The system is composed of the toroidal channel, with length and radius of the constricted region of 80 A and 4 A, respectively, and two reservoirs containing 50 sodium ions and 50 chloride ions. The positions of each of these ions executing Brownian motion under the influence of a stochastic force and a systematic electric force are determined at discrete time steps of 50 fs for up to 2.5 ns. All of the systematic forces acting on an ion due to the other ions, an external electric field, fixed charges in the channel protein, and the image charges induced at the water-protein boundary are explicitly included in the calculations. We find that the repulsive dielectric force arising from the induced surface charges plays a dominant role in channel dynamics. It expels an ion from the vestibule when it is deliberately put in it. Even in the presence of an applied electric potential of 100 mV, an ion cannot overcome this repulsive force and permeate the channel. Only when dipoles of a favorable orientation are placed along the sides of the transmembrane segment can an ion traverse the channel under the influence of a membrane potential. When the strength of the dipoles is further increased, an ion becomes detained in a potential well, and the driving force provided by the applied field is not sufficient to drive the ion out of the well. The trajectory of an ion navigating across the channel mostly remains close to the central axis of the pore lumen. Finally, we discuss the implications of these findings for the transport of ions across the membrane.

Task Dependent Prefrontal Dysfunction in Persons with Asperger's Disorder Investigated with Multi-Channel Near-Infrared Spectroscopy

ERIC Educational Resources Information Center

Iwanami, Akira; Okajima, Yuka; Ota, Haruhisa; Tani, Masayuki; Yamada, Takashi; Hashimoro, Ryuichiro; Kanai, Chieko; Watanabe, Hiromi; Yamasue, Hidenori; Kawakubo, Yuki; Kato, Nobumasa

2011-01-01

Dysfunction of the prefrontal cortex has been previously reported in individuals with Asperger's disorder. In the present study, we used multi-channel near-infrared spectroscopy (NIRS) to detect changes in the oxygenated hemoglobin concentration ([oxy-Hb]) during two verbal fluency tasks. The subjects were 20 individuals with Asperger's disorder…

Saturation of conductance in single ion channels: the blocking effect of the near reaction field.

PubMed

Nadler, Boaz; Schuss, Zeev; Hollerbach, Uwe; Eisenberg, R S

2004-11-01

The ionic current flowing through a protein channel in the membrane of a biological cell depends on the concentration of the permeant ion, as well as on many other variables. As the concentration increases, the rate of arrival of bath ions to the channel's entrance increases, and typically so does the net current. This concentration dependence is part of traditional diffusion and rate models that predict Michaelis-Menten current-concentration relations for a single ion channel. Such models, however, neglect other effects of bath concentrations on the net current. The net current depends not only on the entrance rate of ions into the channel, but also on forces acting on ions inside the channel. These forces, in turn, depend not only on the applied potential and charge distribution of the channel, but also on the long-range Coulombic interactions with the surrounding bath ions. In this paper, we study the effects of bath concentrations on the average force on an ion in a single ion channel. We show that the force of the reaction field on a discrete ion inside a channel embedded in an uncharged lipid membrane contains a blocking (shielding) term that is proportional to the square root of the ionic bath concentration. We then show that different blocking strengths yield different behavior of the current-concentration and conductance-concentration curves. Our theory shows that at low concentrations, when the blocking force is weak, conductance grows linearly with concentration, as in traditional models, e.g., Michaelis-Menten formulations. As the concentration increases to a range of moderate shielding, conductance grows as the square root of concentration, whereas at high concentrations, with high shielding, conductance may actually decrease with increasing concentrations: the conductance-concentration curve can invert. Therefore, electrostatic interactions between bath ions and the single ion inside the channel can explain the different regimes of conductance-concentration relations observed in experiments.

Comparison of Nernst-Planck and reaction rate models for multiply occupied channels.

PubMed Central

Levitt, D G

1982-01-01

The Nernst-Planck continuum equation for a channel that can be occupied by at most two ions is solved for two different physical cases. The first case is for the assumption that the water and ion cannot get around each other anywhere in the channel, so that if there are two ions in the channel the distance between them is fixed by the number of water molecules between them. The second case is for the assumption that there are regions at he ends of the channel where the ions and water can get around each other. For these two cases, the validity of the simple two-site reaction-rate approximation when there is a continuously varying central energy barrier was evaluated by comparing it with the exact Nernst-Planck solution. For the first continuum case, the kinetics for the continuum and reaction-rate models are nearly identical. For the second case, the agreement depends on the strength of the ion-ion interaction energy. For a low interaction energy (large channel diameter) a high ion concentrations, there is a large difference in the flux as a function of voltage for the two models-with the continuum flux becoming more than four times larger at 250 mV. Simple analytical expressions are derived for the two-ion continuum channel for the case where the ends are in equilibrium with the bulk solution and for the case where ion mobility becomes zero when there are two ions in the channel. The implications of these results for biological channels are discussed. PMID:6280783

Investigating ion channel conformational changes using voltage clamp fluorometry.

PubMed

Talwar, Sahil; Lynch, Joseph W

2015-11-01

Ion channels are membrane proteins whose functions are governed by conformational changes. The widespread distribution of ion channels, coupled with their involvement in most physiological and pathological processes and their importance as therapeutic targets, renders the elucidation of these conformational mechanisms highly compelling from a drug discovery perspective. Thanks to recent advances in structural biology techniques, we now have high-resolution static molecular structures for members of the major ion channel families. However, major questions remain to be resolved about the conformational states that ion channels adopt during activation, drug modulation and desensitization. Patch-clamp electrophysiology has long been used to define ion channel conformational states based on functional criteria. It achieves this by monitoring conformational changes at the channel gate and cannot detect conformational changes occurring in regions distant from the gate. Voltage clamp fluorometry involves labelling cysteines introduced into domains of interest with environmentally sensitive fluorophores and inferring structural rearrangements from voltage or ligand-induced fluorescence changes. Ion channel currents are monitored simultaneously to verify the conformational status. By defining real time conformational changes in domains distant from the gate, this technique provides unexpected new insights into ion channel structure and function. This review aims to summarise the methodology and highlight recent innovative applications of this powerful technique. This article is part of the Special Issue entitled 'Fluorescent Tools in Neuropharmacology'. Copyright © 2015 Elsevier Ltd. All rights reserved.

Simulation of charge transport in ion channels and nanopores with anisotropic permittivity

PubMed Central

Mashl, R. Jay; Lee, Kyu Il; Jakobsson, Eric; Ravaioli, Umberto

2010-01-01

Ion channels are part of nature's solution for regulating biological environments. Every ion channel consists of a chain of amino acids carrying a strong and sharply varying permanent charge, folded in such a way that it creates a nanoscopic aqueous pore spanning the otherwise mostly impermeable membranes of biological cells. These naturally occurring proteins are particularly interesting to device engineers seeking to understand how such nanoscale systems realize device-like functions. Availability of high-resolution structural information from X-ray crystallography, as well as large-scale computational resources, makes it possible to conduct realistic ion channel simulations. In general, a hierarchy of simulation methodologies is needed to study different aspects of a biological system like ion channels. Biology Monte Carlo (BioMOCA), a three-dimensional coarse-grained particle ion channel simulator, offers a powerful and general approach to study ion channel permeation. BioMOCA is based on the Boltzmann Transport Monte Carlo (BTMC) and Particle-Particle-Particle-Mesh (P3M) methodologies developed at the University of Illinois at Urbana-Champaign. In this paper we briefly discuss the various approaches to simulating ion flow in channel systems that are currently being pursued by the biophysics and engineering communities, and present the effect of having anisotropic dielectric constants on ion flow through a number of nanopores with different effective diameters. PMID:20445807

Novel screening techniques for ion channel targeting drugs

PubMed Central

Obergrussberger, Alison; Stölzle-Feix, Sonja; Becker, Nadine; Brüggemann, Andrea; Fertig, Niels; Möller, Clemens

2015-01-01

Ion channels are integral membrane proteins that regulate the flux of ions across the cell membrane. They are involved in nearly all physiological processes, and malfunction of ion channels has been linked to many diseases. Until recently, high-throughput screening of ion channels was limited to indirect, e.g. fluorescence-based, readout technologies. In the past years, direct label-free biophysical readout technologies by means of electrophysiology have been developed. Planar patch-clamp electrophysiology provides a direct functional label-free readout of ion channel function in medium to high throughput. Further electrophysiology features, including temperature control and higher-throughput instruments, are continually being developed. Electrophysiological screening in a 384-well format has recently become possible. Advances in chip and microfluidic design, as well as in cell preparation and handling, have allowed challenging cell types to be studied by automated patch clamp. Assays measuring action potentials in stem cell-derived cardiomyocytes, relevant for cardiac safety screening, and neuronal cells, as well as a large number of different ion channels, including fast ligand-gated ion channels, have successfully been established by automated patch clamp. Impedance and multi-electrode array measurements are particularly suitable for studying cardiomyocytes and neuronal cells within their physiological network, and to address more complex physiological questions. This article discusses recent advances in electrophysiological technologies available for screening ion channel function and regulation. PMID:26556400

Novel screening techniques for ion channel targeting drugs.

PubMed

Obergrussberger, Alison; Stölzle-Feix, Sonja; Becker, Nadine; Brüggemann, Andrea; Fertig, Niels; Möller, Clemens

2015-01-01

Ion channels are integral membrane proteins that regulate the flux of ions across the cell membrane. They are involved in nearly all physiological processes, and malfunction of ion channels has been linked to many diseases. Until recently, high-throughput screening of ion channels was limited to indirect, e.g. fluorescence-based, readout technologies. In the past years, direct label-free biophysical readout technologies by means of electrophysiology have been developed. Planar patch-clamp electrophysiology provides a direct functional label-free readout of ion channel function in medium to high throughput. Further electrophysiology features, including temperature control and higher-throughput instruments, are continually being developed. Electrophysiological screening in a 384-well format has recently become possible. Advances in chip and microfluidic design, as well as in cell preparation and handling, have allowed challenging cell types to be studied by automated patch clamp. Assays measuring action potentials in stem cell-derived cardiomyocytes, relevant for cardiac safety screening, and neuronal cells, as well as a large number of different ion channels, including fast ligand-gated ion channels, have successfully been established by automated patch clamp. Impedance and multi-electrode array measurements are particularly suitable for studying cardiomyocytes and neuronal cells within their physiological network, and to address more complex physiological questions. This article discusses recent advances in electrophysiological technologies available for screening ion channel function and regulation.

Highly Sensitive and Patchable Pressure Sensors Mimicking Ion-Channel-Engaged Sensory Organs.

PubMed

Chun, Kyoung-Yong; Son, Young Jun; Han, Chang-Soo

2016-04-26

Biological ion channels have led to much inspiration because of their unique and exquisite operational functions in living cells. Specifically, their extreme and dynamic sensing abilities can be realized by the combination of receptors and nanopores coupled together to construct an ion channel system. In the current study, we demonstrated that artificial ion channel pressure sensors inspired by nature for detecting pressure are highly sensitive and patchable. Our ion channel pressure sensors basically consisted of receptors and nanopore membranes, enabling dynamic current responses to external forces for multiple applications. The ion channel pressure sensors had a sensitivity of ∼5.6 kPa(-1) and a response time of ∼12 ms at a frequency of 1 Hz. The power consumption was recorded as less than a few μW. Moreover, a reliability test showed stability over 10 000 loading-unloading cycles. Additionally, linear regression was performed in terms of temperature, which showed no significant variations, and there were no significant current variations with humidity. The patchable ion channel pressure sensors were then used to detect blood pressure/pulse in humans, and different signals were clearly observed for each person. Additionally, modified ion channel pressure sensors detected complex motions including pressing and folding in a high-pressure range (10-20 kPa).

Orientation independence of single-vacancy and single-ion permeability ratios.

PubMed Central

McGill, P; Schumaker, M F

1995-01-01

Single-vacancy models have been proposed as open channel permeation mechanisms for K+ channels. Single-ion models have been used to describe permeation through Na+ channels. This paper demonstrates that these models have a distinctive symmetry property. Their permeability ratios, measured under biionic conditions, are independent of channel orientation when the reversal potential is zero. This symmetry is a property of general m-site single-vacancy channels, m-site shaking-stack channels, as well as m-site single-ion channels. An experimental finding that the permeability ratios of a channel did not have this symmetry would provide evidence that a single-vacancy or single-ion model is an incorrect or incomplete description of permeation. Images FIGURE 1 PMID:7669913

Pore size matters for potassium channel conductance

PubMed Central

Moldenhauer, Hans; Pincuntureo, Matías

2016-01-01

Ion channels are membrane proteins that mediate efficient ion transport across the hydrophobic core of cell membranes, an unlikely process in their absence. K+ channels discriminate K+ over cations with similar radii with extraordinary selectivity and display a wide diversity of ion transport rates, covering differences of two orders of magnitude in unitary conductance. The pore domains of large- and small-conductance K+ channels share a general architectural design comprising a conserved narrow selectivity filter, which forms intimate interactions with permeant ions, flanked by two wider vestibules toward the internal and external openings. In large-conductance K+ channels, the inner vestibule is wide, whereas in small-conductance channels it is narrow. Here we raise the idea that the physical dimensions of the hydrophobic internal vestibule limit ion transport in K+ channels, accounting for their diversity in unitary conductance. PMID:27619418

Ion-imprinted nanoparticles for the concurrent estimation of Pb(II) and Cu(II) ions over a two channel surface plasmon resonance-based fiber optic platform

NASA Astrophysics Data System (ADS)

Shrivastav, Anand Mohan; Gupta, Banshi D.

2018-01-01

We report the design, fabrication, and characterization of an optical fiber sensor based on the surface plasmon resonance (SPR) technique for the simultaneous determination of lead (Pb) and copper (Cu) metal ions in aqueous samples. Two cascade channels over a single optical fiber are fabricated by removing cladding from two well-separated regions of the fiber. SPR working as a transducing mechanism for the sensor is realized by coating thin films of copper and silver over unclad cores of channel I and channel II, respectively. Ion-imprinted nanoparticles for both ions are separately synthesized and coated over the metal-coated unclad cores of the fiber as the recognition layers for sensor fabrication. A first channel having layer of Pb(II) ion-imprinted nanoparticles detects Pb(II) ions and a second channel having layer of Cu(II) ion-imprinted nanoparticles are used for the detection of Cu(II) ions. Both channels are characterized using the wavelength interrogation method. The sensor operates in the range between 0 to 1000 μg/L and 0 to 1000 mg/L for Pb(II) and Cu(II) ions, respectively. These ranges cover water resources and the human body for these ions. The sensitivities of channel I and channel II are found to be 8.19×104 nm/(μg/L) and 4.07×105 nm/(mg/L) near the lowest concentration of Pb(II) and Cu(II) ions, respectively. The sensor can detect concentrations of Pb(II) and Cu(II) ions as low as 4.06 × 10-12 g/L and 8.18 × 10-10 g/L, respectively, which are the least among the reported values in the literature. Further, the probe is simple, cost effective, highly selective, and applicable for online monitoring and remote sensing.

Selectivity and permeation of alkali metal ions in K+-channels.

PubMed

Furini, Simone; Domene, Carmen

2011-06-24

Ion conduction in K(+)-channels is usually described in terms of concerted movements of K(+) progressing in a single file through a narrow pore. Permeation is driven by an incoming ion knocking on those ions already inside the protein. A fine-tuned balance between high-affinity binding and electrostatic repulsive forces between permeant ions is needed to achieve efficient conduction. While K(+)-channels are known to be highly selective for K(+) over Na(+), some K(+) channels conduct Na(+) in the absence of K(+). Other ions are known to permeate K(+)-channels with a more moderate preference and unusual conduction features. We describe an extensive computational study on ion conduction in K(+)-channels rendering free energy profiles for the translocation of three different alkali ions and some of their mixtures. The free energy maps for Rb(+) translocation show at atomic level why experimental Rb(+) conductance is slightly lower than that of K(+). In contrast to K(+) or Rb(+), external Na(+) block K(+) currents, and the sites where Na(+) transport is hindered are characterized. Translocation of K(+)/Na(+) mixtures is energetically unfavorable owing to the absence of equally spaced ion-binding sites for Na(+), excluding Na(+) from a channel already loaded with K(+). Copyright © 2011 Elsevier Ltd. All rights reserved.

In silico design of novel hERG-neutral sildenafil-like PDE5 inhibitors.

PubMed

Kayık, Gülru; Tüzün, Nurcan Ş; Durdagi, Serdar

2017-10-01

Cyclic nucleotide phosphodiesterase enzymes (PDEs) have functions in regulating the levels of intracellular second messengers, 3', 5'-cyclic adenosine monophosphate (cAMP) and 3', 5'-cyclic guanosine monophosphate (cGMP), via hydrolysis and decomposing mechanisms in cells. They take essential roles in modulating various cellular activities such as memory and smooth muscle functions. PDE type 5 (PDE5) inhibitors enhance the vasodilatory effects of cGMP in the corpus cavernosum and they are used to treat erectile dysfunction. Patch clamp experiments showed that the IC 50 values of the human ether-à-go-go-related gene (hERG1) potassium (K) ion channel blocking affinity of PDE5 inhibitors sildenafil, vardenafil, and tadalafil as 33, 12, and 100 μM, respectively. hERG1 channel is responsible for the regulation of the action potential of human ventricular myocyte by contributing the rapid component of delayed rectifier K + current (I Kr ) component of the cardiac action potential. In this work, interaction patterns and binding affinity predictions of selected PDE5 inhibitors against the hERG1 channel are studied. It is attempted to develop PDE5 inhibitor analogs with lower binding affinity to hERG1 ion channel while keeping their pharmacological activity against their principal target PDE5 using in silico methods. Based on detailed analyses of docking poses and predicted interaction energies, novel analogs of PDE5 inhibitors with lower predicted binding affinity to hERG1 channels without loosing their principal target activity were proposed. Moreover, molecular dynamics (MD) simulations and post-processing MD analyses (i.e. Molecular Mechanics/Generalized Born Surface Area calculations) were performed. Detailed analysis of molecular simulations helped us to better understand the PDE5 inhibitor-target binding interactions in the atomic level. Results of this study can be useful for designing of novel and safe PDE5 inhibitors with enhanced activity and other tailored properties.

A New Poisson-Nernst-Planck Model with Ion-Water Interactions for Charge Transport in Ion Channels.

PubMed

Chen, Duan

2016-08-01

In this work, we propose a new Poisson-Nernst-Planck (PNP) model with ion-water interactions for biological charge transport in ion channels. Due to narrow geometries of these membrane proteins, ion-water interaction is critical for both dielectric property of water molecules in channel pore and transport dynamics of mobile ions. We model the ion-water interaction energy based on realistic experimental observations in an efficient mean-field approach. Variation of a total energy functional of the biological system yields a new PNP-type continuum model. Numerical simulations show that the proposed model with ion-water interaction energy has the new features that quantitatively describe dielectric properties of water molecules in narrow pores and are possible to model the selectivity of some ion channels.

An evolutionarily conserved gene family encodes proton-selective ion channels.

PubMed

Tu, Yu-Hsiang; Cooper, Alexander J; Teng, Bochuan; Chang, Rui B; Artiga, Daniel J; Turner, Heather N; Mulhall, Eric M; Ye, Wenlei; Smith, Andrew D; Liman, Emily R

2018-03-02

Ion channels form the basis for cellular electrical signaling. Despite the scores of genetically identified ion channels selective for other monatomic ions, only one type of proton-selective ion channel has been found in eukaryotic cells. By comparative transcriptome analysis of mouse taste receptor cells, we identified Otopetrin1 (OTOP1), a protein required for development of gravity-sensing otoconia in the vestibular system, as forming a proton-selective ion channel. We found that murine OTOP1 is enriched in acid-detecting taste receptor cells and is required for their zinc-sensitive proton conductance. Two related murine genes, Otop2 and Otop3 , and a Drosophila ortholog also encode proton channels. Evolutionary conservation of the gene family and its widespread tissue distribution suggest a broad role for proton channels in physiology and pathophysiology. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Therapeutic Approaches to Genetic Ion Channelopathies and Perspectives in Drug Discovery

PubMed Central

Imbrici, Paola; Liantonio, Antonella; Camerino, Giulia M.; De Bellis, Michela; Camerino, Claudia; Mele, Antonietta; Giustino, Arcangela; Pierno, Sabata; De Luca, Annamaria; Tricarico, Domenico; Desaphy, Jean-Francois; Conte, Diana

2016-01-01

In the human genome more than 400 genes encode ion channels, which are transmembrane proteins mediating ion fluxes across membranes. Being expressed in all cell types, they are involved in almost all physiological processes, including sense perception, neurotransmission, muscle contraction, secretion, immune response, cell proliferation, and differentiation. Due to the widespread tissue distribution of ion channels and their physiological functions, mutations in genes encoding ion channel subunits, or their interacting proteins, are responsible for inherited ion channelopathies. These diseases can range from common to very rare disorders and their severity can be mild, disabling, or life-threatening. In spite of this, ion channels are the primary target of only about 5% of the marketed drugs suggesting their potential in drug discovery. The current review summarizes the therapeutic management of the principal ion channelopathies of central and peripheral nervous system, heart, kidney, bone, skeletal muscle and pancreas, resulting from mutations in calcium, sodium, potassium, and chloride ion channels. For most channelopathies the therapy is mainly empirical and symptomatic, often limited by lack of efficacy and tolerability for a significant number of patients. Other channelopathies can exploit ion channel targeted drugs, such as marketed sodium channel blockers. Developing new and more specific therapeutic approaches is therefore required. To this aim, a major advancement in the pharmacotherapy of channelopathies has been the discovery that ion channel mutations lead to change in biophysics that can in turn specifically modify the sensitivity to drugs: this opens the way to a pharmacogenetics strategy, allowing the development of a personalized therapy with increased efficacy and reduced side effects. In addition, the identification of disease modifiers in ion channelopathies appears an alternative strategy to discover novel druggable targets. PMID:27242528

Therapeutic Approaches to Genetic Ion Channelopathies and Perspectives in Drug Discovery.

PubMed

Imbrici, Paola; Liantonio, Antonella; Camerino, Giulia M; De Bellis, Michela; Camerino, Claudia; Mele, Antonietta; Giustino, Arcangela; Pierno, Sabata; De Luca, Annamaria; Tricarico, Domenico; Desaphy, Jean-Francois; Conte, Diana

2016-01-01

In the human genome more than 400 genes encode ion channels, which are transmembrane proteins mediating ion fluxes across membranes. Being expressed in all cell types, they are involved in almost all physiological processes, including sense perception, neurotransmission, muscle contraction, secretion, immune response, cell proliferation, and differentiation. Due to the widespread tissue distribution of ion channels and their physiological functions, mutations in genes encoding ion channel subunits, or their interacting proteins, are responsible for inherited ion channelopathies. These diseases can range from common to very rare disorders and their severity can be mild, disabling, or life-threatening. In spite of this, ion channels are the primary target of only about 5% of the marketed drugs suggesting their potential in drug discovery. The current review summarizes the therapeutic management of the principal ion channelopathies of central and peripheral nervous system, heart, kidney, bone, skeletal muscle and pancreas, resulting from mutations in calcium, sodium, potassium, and chloride ion channels. For most channelopathies the therapy is mainly empirical and symptomatic, often limited by lack of efficacy and tolerability for a significant number of patients. Other channelopathies can exploit ion channel targeted drugs, such as marketed sodium channel blockers. Developing new and more specific therapeutic approaches is therefore required. To this aim, a major advancement in the pharmacotherapy of channelopathies has been the discovery that ion channel mutations lead to change in biophysics that can in turn specifically modify the sensitivity to drugs: this opens the way to a pharmacogenetics strategy, allowing the development of a personalized therapy with increased efficacy and reduced side effects. In addition, the identification of disease modifiers in ion channelopathies appears an alternative strategy to discover novel druggable targets.

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

Progesterone Inhibition of Neuronal Calcium Signaling Underlies Aspects of Progesterone-Mediated Neuroprotection

PubMed Central

Luoma, Jessie I; Stern, Christopher M; Mermelstein, Paul G.

2011-01-01

Progesterone is being utilized as a therapeutic means to ameliorate neuron loss and cognitive dysfunction following traumatic brain injury Although there have been numerous attempts to determine the means by which progesterone exerts neuroprotective effects, studies describing the underlying molecular mechanisms are lacking What has become clear, however, is the notion that progesterone can thwart several physiological processes that are detrimental to neuron function and survival, including inflammation, edema, demyelination and excitotoxicity One clue regarding the means by which progesterone has restorative value comes from the notion that these aforementioned biological processes all share the common theme of eliciting pronounced increases in intracellular calcium. Thus, we propose the hypothesis that progesterone regulation of calcium signaling underlies its ability to mitigate these cellular insults, ultimately leading to neuroprotection. Further, we describe recent findings that indicate neuroprotection is achieved via progesterone block of voltage-gated calcium channels, although additional outcomes may arise from blockade of various other ion channels and neurotransmitter receptors. PMID:22101209

Moving Fe2+ from ferritin ion channels to catalytic OH centers depends on conserved protein cage carboxylates.

PubMed

Behera, Rabindra K; Theil, Elizabeth C

2014-06-03

Ferritin biominerals are protein-caged metabolic iron concentrates used for iron-protein cofactors and oxidant protection (Fe(2+) and O2 sequestration). Fe(2+) passage through ion channels in the protein cages, like membrane ion channels, required for ferritin biomineral synthesis, is followed by Fe(2+) substrate movement to ferritin enzyme (Fox) sites. Fe(2+) and O2 substrates are coupled via a diferric peroxo (DFP) intermediate, λmax 650 nm, which decays to [Fe(3+)-O-Fe(3+)] precursors of caged ferritin biominerals. Structural studies show multiple conformations for conserved, carboxylate residues E136 and E57, which are between ferritin ion channel exits and enzymatic sites, suggesting functional connections. Here we show that E136 and E57 are required for ferritin enzyme activity and thus are functional links between ferritin ion channels and enzymatic sites. DFP formation (Kcat and kcat/Km), DFP decay, and protein-caged hydrated ferric oxide accumulation decreased in ferritin E57A and E136A; saturation required higher Fe(2+) concentrations. Divalent cations (both ion channel and intracage binding) selectively inhibit ferritin enzyme activity (block Fe(2+) access), Mn(2+) < Co(2+) < Cu(2+) < Zn(2+), reflecting metal ion-protein binding stabilities. Fe(2+)-Cys126 binding in ferritin ion channels, observed as Cu(2+)-S-Cys126 charge-transfer bands in ferritin E130D UV-vis spectra and resistance to Cu(2+) inhibition in ferritin C126S, was unpredicted. Identifying E57 and E136 links in Fe(2+) movement from ferritin ion channels to ferritin enzyme sites completes a bucket brigade that moves external Fe(2+) into ferritin enzymatic sites. The results clarify Fe(2+) transport within ferritin and model molecular links between membrane ion channels and cytoplasmic destinations.

Global versus local mechanisms of temperature sensing in ion channels.

PubMed

Arrigoni, Cristina; Minor, Daniel L

2018-05-01

Ion channels turn diverse types of inputs, ranging from neurotransmitters to physical forces, into electrical signals. Channel responses to ligands generally rely on binding to discrete sensor domains that are coupled to the portion of the channel responsible for ion permeation. By contrast, sensing physical cues such as voltage, pressure, and temperature arises from more varied mechanisms. Voltage is commonly sensed by a local, domain-based strategy, whereas the predominant paradigm for pressure sensing employs a global response in channel structure to membrane tension changes. Temperature sensing has been the most challenging response to understand and whether discrete sensor domains exist for pressure and temperature has been the subject of much investigation and debate. Recent exciting advances have uncovered discrete sensor modules for pressure and temperature in force-sensitive and thermal-sensitive ion channels, respectively. In particular, characterization of bacterial voltage-gated sodium channel (BacNa V ) thermal responses has identified a coiled-coil thermosensor that controls channel function through a temperature-dependent unfolding event. This coiled-coil thermosensor blueprint recurs in other temperature sensitive ion channels and thermosensitive proteins. Together with the identification of ion channel pressure sensing domains, these examples demonstrate that "local" domain-based solutions for sensing force and temperature exist and highlight the diversity of both global and local strategies that channels use to sense physical inputs. The modular nature of these newly discovered physical signal sensors provides opportunities to engineer novel pressure-sensitive and thermosensitive proteins and raises new questions about how such modular sensors may have evolved and empowered ion channel pores with new sensibilities.

Ion-binding properties of a K+ channel selectivity filter in different conformations.

PubMed

Liu, Shian; Focke, Paul J; Matulef, Kimberly; Bian, Xuelin; Moënne-Loccoz, Pierre; Valiyaveetil, Francis I; Lockless, Steve W

2015-12-08

K(+) channels are membrane proteins that selectively conduct K(+) ions across lipid bilayers. Many voltage-gated K(+) (KV) channels contain two gates, one at the bundle crossing on the intracellular side of the membrane and another in the selectivity filter. The gate at the bundle crossing is responsible for channel opening in response to a voltage stimulus, whereas the gate at the selectivity filter is responsible for C-type inactivation. Together, these regions determine when the channel conducts ions. The K(+) channel from Streptomyces lividians (KcsA) undergoes an inactivation process that is functionally similar to KV channels, which has led to its use as a practical system to study inactivation. Crystal structures of KcsA channels with an open intracellular gate revealed a selectivity filter in a constricted conformation similar to the structure observed in closed KcsA containing only Na(+) or low [K(+)]. However, recent work using a semisynthetic channel that is unable to adopt a constricted filter but inactivates like WT channels challenges this idea. In this study, we measured the equilibrium ion-binding properties of channels with conductive, inactivated, and constricted filters using isothermal titration calorimetry (ITC). EPR spectroscopy was used to determine the state of the intracellular gate of the channel, which we found can depend on the presence or absence of a lipid bilayer. Overall, we discovered that K(+) ion binding to channels with an inactivated or conductive selectivity filter is different from K(+) ion binding to channels with a constricted filter, suggesting that the structures of these channels are different.

Large fraction of crystal directions leads to ion channeling

NASA Astrophysics Data System (ADS)

Nordlund, K.; Djurabekova, F.; Hobler, G.

2016-12-01

It is well established that when energetic ions are moving in crystals, they may penetrate much deeper if they happen to be directed in some specific crystal directions. This `channeling' effect is utilized for instance in certain ion beam analysis methods and has been described by analytical theories and atomistic computer simulations. However, there have been very few systematic studies of channeling in directions other than the principal low-index ones. We present here a molecular dynamics-based approach to calculate ion channeling systematically over all crystal directions, providing ion `channeling maps' that easily show in which directions channeling is expected. The results show that channeling effects can be quite significant even at energies below 1 keV, and that in many cases, significant planar channeling occurs also in a wide range of crystal directions between the low-index principal ones. In all of the cases studied, a large fraction (˜20 -60 % ) of all crystal directions show channeling. A practical implication of this is that modern experiments on randomly oriented nanostructures will have a large probability of channeling. It also means that when ion irradiations are carried out on polycrystalline samples, channeling effects on the results cannot a priori be assumed to be negligible. The maps allow for easy selection of good `nonchanneling' directions in experiments or alternatively finding wide channels for beneficial uses of channeling. We implement channeling theory to also give the fraction of channeling directions in a manner directly comparable to the simulations. The comparison shows good qualitative agreement. In particular, channeling theory is very good at predicting which channels are active at a given energy. This is true down to sub-keV energies, provided the penetration depth is not too small.

Molecular mechanism of ATP binding and ion channel activation in P2X receptors

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

Hattori, Motoyuki; Gouaux, Eric

P2X receptors are trimeric ATP-activated ion channels permeable to Na{sup +}, K{sup +} and Ca{sup 2+}. The seven P2X receptor subtypes are implicated in physiological processes that include modulation of synaptic transmission, contraction of smooth muscle, secretion of chemical transmitters and regulation of immune responses. Despite the importance of P2X receptors in cellular physiology, the three-dimensional composition of the ATP-binding site, the structural mechanism of ATP-dependent ion channel gating and the architecture of the open ion channel pore are unknown. Here we report the crystal structure of the zebrafish P2X4 receptor in complex with ATP and a new structure ofmore » the apo receptor. The agonist-bound structure reveals a previously unseen ATP-binding motif and an open ion channel pore. ATP binding induces cleft closure of the nucleotide-binding pocket, flexing of the lower body {beta}-sheet and a radial expansion of the extracellular vestibule. The structural widening of the extracellular vestibule is directly coupled to the opening of the ion channel pore by way of an iris-like expansion of the transmembrane helices. The structural delineation of the ATP-binding site and the ion channel pore, together with the conformational changes associated with ion channel gating, will stimulate development of new pharmacological agents.« less

Calcium signalling in salivary gland physiology and dysfunction

PubMed Central

2015-01-01

Abstract Studies over the past four decades have established that Ca2+ is a critical factor in control of salivary gland function and have led to identification of the critical components of this process. The major ion transport mechanisms and ion channels that are involved in fluid secretion have also been established. The key event in activation of fluid secretion is an increase in [Ca2+]i triggered by inositol 1,4,5‐trisphosphate (IP3)‐induced release of Ca2+ from ER via the IP3 receptor (IP3R). IP3Rs determine the site of initiation and the pattern of the [Ca2+]i signal in the cell. However, Ca2+ entry into the cell is required to sustain the elevation of [Ca2+]i and fluid secretion and is mediated by the store‐operated Ca2+ entry (SOCE) mechanism. Orai1, TRPC1, TRPC3 and STIM1 have been identified as critical components of SOCE in these cells. Cells finely tune the generation and amplification of [Ca2+]i signals for regulation of cell function. An important emerging area is the concept that unregulated [Ca2+]i signals in cells can directly cause cell damage, dysfunction and disease. Alternatively, aberrant [Ca2+]i signals can also amplify and increase the rates of cell damage. Such defects in Ca2+ signalling have been described in salivary glands in conjunction with radiation‐induced loss of salivary gland function as well as in the salivary defects associated with the autoimmune exocrinopathy Sjögren's syndrome. Such defects have been associated with altered function or expression of key Ca2+ signalling components, such as STIM proteins and TRP channels. These studies offer new avenues for examining the mechanisms underlying the disease and development of novel clinical targets and therapeutic strategies. PMID:26592972

Calcium signalling in salivary gland physiology and dysfunction.

PubMed

Ambudkar, Indu S

2016-06-01

Studies over the past four decades have established that Ca(2+) is a critical factor in control of salivary gland function and have led to identification of the critical components of this process. The major ion transport mechanisms and ion channels that are involved in fluid secretion have also been established. The key event in activation of fluid secretion is an increase in [Ca(2+) ]i triggered by inositol 1,4,5-trisphosphate (IP3 )-induced release of Ca(2+) from ER via the IP3 receptor (IP3 R). IP3 Rs determine the site of initiation and the pattern of the [Ca(2+) ]i signal in the cell. However, Ca(2+) entry into the cell is required to sustain the elevation of [Ca(2+) ]i and fluid secretion and is mediated by the store-operated Ca(2+) entry (SOCE) mechanism. Orai1, TRPC1, TRPC3 and STIM1 have been identified as critical components of SOCE in these cells. Cells finely tune the generation and amplification of [Ca(2+) ]i signals for regulation of cell function. An important emerging area is the concept that unregulated [Ca(2+) ]i signals in cells can directly cause cell damage, dysfunction and disease. Alternatively, aberrant [Ca(2+) ]i signals can also amplify and increase the rates of cell damage. Such defects in Ca(2+) signalling have been described in salivary glands in conjunction with radiation-induced loss of salivary gland function as well as in the salivary defects associated with the autoimmune exocrinopathy Sjögren's syndrome. Such defects have been associated with altered function or expression of key Ca(2+) signalling components, such as STIM proteins and TRP channels. These studies offer new avenues for examining the mechanisms underlying the disease and development of novel clinical targets and therapeutic strategies. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

Ion Channels in Brain Metastasis

PubMed Central

Klumpp, Lukas; Sezgin, Efe C.; Eckert, Franziska; Huber, Stephan M.

2016-01-01

Breast cancer, lung cancer and melanoma exhibit a high metastatic tropism to the brain. Development of brain metastases severely worsens the prognosis of cancer patients and constrains curative treatment options. Metastasizing to the brain by cancer cells can be dissected in consecutive processes including epithelial–mesenchymal transition, evasion from the primary tumor, intravasation and circulation in the blood, extravasation across the blood–brain barrier, formation of metastatic niches, and colonization in the brain. Ion channels have been demonstrated to be aberrantly expressed in tumor cells where they regulate neoplastic transformation, malignant progression or therapy resistance. Moreover, many ion channel modulators are FDA-approved drugs and in clinical use proposing ion channels as druggable targets for future anti-cancer therapy. The present review article aims to summarize the current knowledge on the function of ion channels in the different processes of brain metastasis. The data suggest that certain channel types involving voltage-gated sodium channels, ATP-release channels, ionotropic neurotransmitter receptors and gap junction-generating connexins interfere with distinct processes of brain metastazation. PMID:27618016

From mucolipidosis type IV to Ebola: TRPML and two-pore channels at the crossroads of endo-lysosomal trafficking and disease.

PubMed

Grimm, Christian; Butz, Elisabeth; Chen, Cheng-Chang; Wahl-Schott, Christian; Biel, Martin

2017-11-01

What do lysosomal storage disorders such as mucolipidosis type IV have in common with Ebola, cancer cell migration, or LDL-cholesterol trafficking? LDL-cholesterol, certain bacterial toxins and viruses, growth factors, receptors, integrins, macromolecules destined for degradation or secretion are all sorted and transported via the endolysosomal system (ES). There are several pathways known in the ES, e.g. the degradation, the recycling, or the retrograde trafficking pathway. The ES comprises early and late endosomes, lysosomes and recycling endosomes as well as autophagosomes and lysosome related organelles. Contact sites between the ES and the endoplasmic reticulum or the Golgi apparatus may also be considered part of it. Dysfunction of this complex intracellular machinery can cause or contribute to the development of a number of diseases ranging from neurodegenerative, infectious, or metabolic diseases to retinal and pigmentation disorders as well as cancer and autophagy-related diseases. Endolysosomal ion channels such as mucolipins (TRPMLs) and two-pore channels (TPCs) play an important role in intracellular cation/calcium signaling and homeostasis and appear to critically contribute to the proper function of the endolysosomal trafficking network. Copyright © 2017 Elsevier Ltd. All rights reserved.

New light on ion channel imaging by total internal reflection fluorescence (TIRF) microscopy.

PubMed

Yamamura, Hisao; Suzuki, Yoshiaki; Imaizumi, Yuji

2015-05-01

Ion channels play pivotal roles in a wide variety of cellular functions; therefore, their physiological characteristics, pharmacological responses, and molecular structures have been extensively investigated. However, the mobility of an ion channel itself in the cell membrane has not been examined in as much detail. A total internal reflection fluorescence (TIRF) microscope allows fluorophores to be imaged in a restricted region within an evanescent field of less than 200 nm from the interface of the coverslip and plasma membrane in living cells. Thus the TIRF microscope is useful for selectively visualizing the plasmalemmal surface and subplasmalemmal zone. In this review, we focused on a single-molecule analysis of the dynamic movement of ion channels in the plasma membrane using TIRF microscopy. We also described two single-molecule imaging techniques under TIRF microscopy: fluorescence resonance energy transfer (FRET) for the identification of molecules that interact with ion channels, and subunit counting for the determination of subunit stoichiometry in a functional channel. TIRF imaging can also be used to analyze spatiotemporal Ca(2+) events in the subplasmalemma. Single-molecule analyses of ion channels and localized Ca(2+) signals based on TIRF imaging provide beneficial pharmacological and physiological information concerning the functions of ion channels. Copyright © 2015 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

Streaming potentials in gramicidin channels measured with ion-selective microelectrodes.

PubMed Central

Tripathi, S; Hladky, S B

1998-01-01

Streaming potentials have been measured for gramicidin channels with a new method employing ion-selective microelectrodes. It is shown that ideally ion-selective electrodes placed at the membrane surface record the true streaming potential. Using this method for ion concentrations below 100 mM, approximately seven water molecules are transported whenever a sodium, potassium, or cesium ion, passes through the channel. This new method confirms earlier measurements (Rosenberg, P.A., and A. Finkelstein. 1978. Interaction of ions and water in gramicidin A channels. J. Gen. Physiol. 72:327-340) in which the streaming potentials were calculated as the difference between electrical potentials measured in the presence of gramicidin and in the presence of the ion carriers valinomycin and nonactin. PMID:9635745

Ion Selectivity Mechanism in a Bacterial Pentameric Ligand-Gated Ion Channel

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

Fritsch, Sebastian; Ivanov, Ivaylo; Wang, Hailong

2010-01-01

The proton-gated ion channel from Gloeobacter violaceus (GLIC) is a prokaryotic homolog of the eukaryotic nicotinic acetylcholine receptor that responds to the binding of neurotransmitter acetylcholine and mediates fast signal transmission. Recent emergence of a high-resolution crystal structure of GLIC captured in a potentially open state allowed detailed, atomic-level insight into ion conduction and selectivity mechanisms in these channels. Herein, we have examined the barriers to ion conduction and origins of ion selectivity in the GLIC channel by the construction of potential-of-mean-force profiles for sodium and chloride ions inside the transmembrane region. Our calculations reveal that the GLIC channel ismore » open for a sodium ion to transport, but presents a 11 kcal/mol free energy barrier for a chloride ion. Our collective findings identify three distinct contributions to the observed preference for the permeant ions. First, there is a substantial contribution due to a ring of negatively charged glutamate residues (E-2 ) at the narrow intracellular end of the channel. The negative electrostatics of this region and the ability of the glutamate side chains to directly bind cations would strongly favor the passage of sodium ions while hindering translocation of chloride ions. Second, our results imply a significant hydrophobic contribution to selectivity linked to differences in the desolvation penalty for the sodium versus chloride ions in the central hydrophobic region of the pore. This hydrophobic contribution is evidenced by the large free energy barriers experienced by Cl in the middle of the pore for both GLIC and the E-2 A mutant. Finally, there is a distinct contribution arising from the overall negative electrostatics of the channel.« less

Small Ion Channel Linking Molecular Simulations and Electrophysiology

NASA Technical Reports Server (NTRS)

Pohorille, Andrzej

2017-01-01

Ion channels are pore-forming protein assemblies that mediate the transport of small ions across cell membranes. Otherwise, membrane bilayers would be almost impermeable to ions incapable to traverse the low dielectric constant, hydrophobic membrane core. Ion channels are ubiquitous to all life forms. In humans and other higher organisms they play the central role in conducting nerve impulses, cardiac functions, muscle contraction and apoptosis. On the other extreme of biological complexity, viral ion channels (viroporins) influence many stages of the virus infection cycle either through regulating virus replication, such as entry, assembly and release or modulating the electrochemical balance in the subcellular compartments of host cells. Ion channels were crucial components of protocells. Their emergence facilitated adaptation of nascent life to different environmental conditions. The earliest ion channels must have been much simpler than most of their modern ancestors. Viral channels are among only a few naturally occurring models to study the structure, function and evolution of primordial channels. Experimental studies of these properties are difficult and often unreliable. In principle, computational methods, and molecular dynamics (MD) simulations in particular, can aid in providing information about both the structure and the function of ion channels. However, MD suffers from its own problems, such as inability to access sufficiently long time scales or limited accuracy of force fields. It is, therefore, essential to determine the reliability of MD simulations. We propose to do so on the basis of two criteria. One is channel stability on time scales that extend for several microseconds or longer. The other is the ability to reproduce the measured ionic conductance as a function of applied voltage. If both the stability and the calculated ionic conductance are satisfactory it will greatly increase our confidence that the structure and the function of a channel are described sufficiently accurately. To our knowledge, long time scale stability (approx.10 micro-sec) and the correct electrophysiology have been shown so far for only one channel - the synthetic LS3 hexamer). In this presentation, this approach will be discussed in application to two viral channels - Vpu, encoded by the HIV-1 genome and p7 of hepatitis C.

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

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

Never at rest: insights into the conformational dynamics of ion channels from cryo-electron microscopy.

PubMed

Lau, Carus; Hunter, Mark J; Stewart, Alastair; Perozo, Eduardo; Vandenberg, Jamie I

2018-04-01

The tightly regulated opening and closure of ion channels underlies the electrical signals that are vital for a wide range of physiological processes. Two decades ago the first atomic level view of ion channel structures led to a detailed understanding of ion selectivity and conduction. In recent years, spectacular developments in the field of cryo-electron microscopy have resulted in cryo-EM superseding crystallography as the technique of choice for determining near-atomic resolution structures of ion channels. Here, we will review the recent developments in cryo-EM and its specific application to the study of ion channel gating. We will highlight the advantages and disadvantages of the current technology and where the field is likely to head in the next few years. © 2018 The Authors. The Journal of Physiology © 2018 The Physiological Society.

Channels Active in the Excitability of Nerves and Skeletal Muscles across the Neuromuscular Junction: Basic Function and Pathophysiology

ERIC Educational Resources Information Center

Goodman, Barbara E.

2008-01-01

Ion channels are essential for the basic physiological function of excitable cells such as nerve, skeletal, cardiac, and smooth muscle cells. Mutations in genes that encode ion channels have been identified to cause various diseases and disorders known as channelopathies. An understanding of how individual ion channels are involved in the…

The Origin and Early Evolution of Membrane Proteins

NASA Technical Reports Server (NTRS)

Pohorille, Andrew; Schweighofter, Karl; Wilson, Michael A.

2006-01-01

The origin and early evolution of membrane proteins, and in particular ion channels, are considered from the point of view that the transmembrane segments of membrane proteins are structurally quite simple and do not require specific sequences to fold. We argue that the transport of solute species, especially ions, required an early evolution of efficient transport mechanisms, and that the emergence of simple ion channels was protobiologically plausible. We also argue that, despite their simple structure, such channels could possess properties that, at the first sight, appear to require markedly larger complexity. These properties can be subtly modulated by local modifications to the sequence rather than global changes in molecular architecture. In order to address the evolution and development of ion channels, we focus on identifying those protein domains that are commonly associated with ion channel proteins and are conserved throughout the three main domains of life (Eukarya, Prokarya, and Archaea). We discuss the potassium-sodium-calcium superfamily of voltage-gated ion channels, mechanosensitive channels, porins, and ABC-transporters and argue that these families of membrane channels have sufficiently universal architectures that they can readily adapt to the diverse functional demands arising during evolution.

Dominant-negative suppression of big brain ion channel activity by mutation of a conserved glutamate in the first transmembrane domain.

PubMed

Yool, Andrea J

2007-01-01

The neurogenic protein Drosophila big brain (BIB), which is involved in the process of neuroblast determination, and the water channel aquaporin-1 (AQP1) are among a subset of the major intrinsic protein (MIP) channels that have been found to show gated monovalent cation channel activity. A glutamate residue in the first transmembrane (M1) domain is conserved throughout the MIP family. Mutation of this residue to asparagine in BIB (E71N) knocks out ion channel activity, and when coexpressed with BIB wild-type as shown here generates a dominant-negative effect on ion channel function, measured in the Xenopus oocyte expression system using two-electrode voltage clamp. cRNAs for wild-type and mutant BIB or AQP1 channels were injected individually or as mixtures. The magnitude of the BIB ionic conductance response was greatly reduced by coexpression of the mutant E71N subunit, suggesting a dominant-negative mechanism of action. The analogous mutation in AQP1 (E17N) did not impair ion channel activation by cGMP, but did knock out water channel function, although not via a dominant-negative effect. This contrast in sensitivity between BIB and AQP1 to mutation of the M1 glutamate suggests the possibility of interesting structural differences in the molecular basis of the ion permeation between these two classes of channels. The dominant-negative construct of BIB could be a tool for testing a role for BIB ion channels during nervous system development in Drosophila. The neurogenic protein Drosophila big brain (BIB), which is involved in the process of neuroblast determination, and the water channel aquaporin-1 (AQP1) are among a subset of the major intrinsic protein (MIP) channels that have been found to show gated monovalent cation channel activity. A glutamate residue in the first transmembrane (M1) domain is conserved throughout the MIP family. Mutation of this residue to asparagine in BIB (E71N) knocks out ion channel activity, and when coexpressed with BIB wild-type as shown here generates a dominant-negative effect on ion channel function, measured in the Xenopus oocyte expression system using two-electrode voltage clamp. cRNAs for wild-type and mutant BIB or AQP1 channels were injected individually or as mixtures. The magnitude of the BIB ionic conductance response was greatly reduced by coexpression of the mutant E71N subunit, suggesting a dominant-negative mechanism of action. The analogous mutation in AQP1 (E17N) did not impair ion channel activation by cGMP, but did knock out water channel function, although not via a dominant-negative effect. This contrast in sensitivity between BIB and AQP1 to mutation of the M1 glutamate suggests the possibility of interesting structural differences in the molecular basis of the ion permeation between these two classes of channels. The dominant-negative construct of BIB could be a tool for testing a role for BIB ion channels during nervous system development in Drosophila.

Dielectric Self-Energy in Poisson-Boltzmann and Poisson-Nernst-Planck Models of Ion Channels

PubMed Central

Corry, Ben; Kuyucak, Serdar; Chung, Shin-Ho

2003-01-01

We demonstrated previously that the two continuum theories widely used in modeling biological ion channels give unreliable results when the radius of the conduit is less than two Debye lengths. The reason for this failure is the neglect of surface charges on the protein wall induced by permeating ions. Here we attempt to improve the accuracy of the Poisson-Boltzmann and Poisson-Nernst-Planck theories, when applied to channel-like environments, by including a specific dielectric self-energy term to overcome spurious shielding effects inherent in these theories. By comparing results with Brownian dynamics simulations, we show that the inclusion of an additional term in the equations yields significant qualitative improvements. The modified theories perform well in very wide and very narrow channels, but are less successful at intermediate sizes. The situation is worse in multi-ion channels because of the inability of the continuum theories to handle the ion-to-ion interactions correctly. Thus, further work is required if these continuum theories are to be reliably salvaged for quantitative studies of biological ion channels in all situations. PMID:12770869

Crystal structure of the ATP-gated P2X[subscript 4] ion channel in the closed state

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

Kawate, Toshimitsu; Michel, Jennifer Carlisle; Birdsong, William T.

2009-08-13

P2X receptors are cation-selective ion channels gated by extracellular ATP, and are implicated in diverse physiological processes, from synaptic transmission to inflammation to the sensing of taste and pain. Because P2X receptors are not related to other ion channel proteins of known structure, there is at present no molecular foundation for mechanisms of ligand-gating, allosteric modulation and ion permeation. Here we present crystal structures of the zebrafish P2X{sub 4} receptor in its closed, resting state. The chalice-shaped, trimeric receptor is knit together by subunit-subunit contacts implicated in ion channel gating and receptor assembly. Extracellular domains, rich in {beta}-strands, have largemore » acidic patches that may attract cations, through fenestrations, to vestibules near the ion channel. In the transmembrane pore, the 'gate' is defined by an {approx}8 {angstrom} slab of protein. We define the location of three non-canonical, intersubunit ATP-binding sites, and suggest that ATP binding promotes subunit rearrangement and ion channel opening.« less

Unraveling the mechanism of selective ion transport in hydrophobic subnanometer channels.

PubMed

Li, Hui; Francisco, Joseph S; Zeng, Xiao Cheng

2015-09-01

Recently reported synthetic organic nanopore (SONP) can mimic a key feature of natural ion channels, i.e., selective ion transport. However, the physical mechanism underlying the K(+)/Na(+) selectivity for the SONPs is dramatically different from that of natural ion channels. To achieve a better understanding of the selective ion transport in hydrophobic subnanometer channels in general and SONPs in particular, we perform a series of ab initio molecular dynamics simulations to investigate the diffusivity of aqua Na(+) and K(+) ions in two prototype hydrophobic nanochannels: (i) an SONP with radius of 3.2 Å, and (ii) single-walled carbon nanotubes (CNTs) with radii of 3-5 Å (these radii are comparable to those of the biological potassium K(+) channels). We find that the hydration shell of aqua Na(+) ion is smaller than that of aqua K(+) ion but notably more structured and less yielding. The aqua ions do not lower the diffusivity of water molecules in CNTs, but in SONP the diffusivity of aqua ions (Na(+) in particular) is strongly suppressed due to the rugged inner surface. Moreover, the aqua Na(+) ion requires higher formation energy than aqua K(+) ion in the hydrophobic nanochannels. As such, we find that the ion (K(+) vs. Na(+)) selectivity of the (8, 8) CNT is ∼20× higher than that of SONP. Hence, the (8, 8) CNT is likely the most efficient artificial K(+) channel due in part to its special interior environment in which Na(+) can be fully solvated, whereas K(+) cannot. This work provides deeper insights into the physical chemistry behind selective ion transport in nanochannels.

Promotion of Water Channels for Enhanced Ion Transport in 14 nm Diameter Carbon Nanotubes.

PubMed

Sheng, Jiadong; Zhu, Qi; Zeng, Xian; Yang, Zhaohui; Zhang, Xiaohua

2017-03-29

Ion transport plays an important role in solar-to-electricity conversion, drug delivery, and a variety of biological processes. Carbon nanotube (CNT) is a promising material as an ion transporter in the applications of the mimicking of natural ion channels, desalination, and energy harvesting. Here, we demonstrate a unique, enhanced ion transport through a vertically aligned multiwall CNT membrane after the application of an electric potential across CNT membranes. Interestingly, electrowetting arising from the application of an electric potential is critical for the enhancement of overall ion transport rate through CNT membranes. The wettability of a liquid with high surface tension on the interior channel walls of CNTs increases during an electric potential treatment and promotes the formation of water channels in CNTs. The formation of water channels in CNTs induces an increase in overall ion diffusion through CNT membranes. This phenomenon is also related to a decrease in the charge transfer resistance of CNTs (R ct ) after an electric potential is applied. Correspondingly, the enhanced ion flow rate gives rise to an enhancement in the capacitive performance of CNT based membranes. Our observations might have profound impact on the development of CNT based energy storage devices as well as artificial ion channels.

A Comparison of the Performance and Application Differences Between Manual and Automated Patch-Clamp Techniques

PubMed Central

Yajuan, Xiao; Xin, Liang; Zhiyuan, Li

2012-01-01

The patch clamp technique is commonly used in electrophysiological experiments and offers direct insight into ion channel properties through the characterization of ion channel activity. This technique can be used to elucidate the interaction between a drug and a specific ion channel at different conformational states to understand the ion channel modulators’ mechanisms. The patch clamp technique is regarded as a gold standard for ion channel research; however, it suffers from low throughput and high personnel costs. In the last decade, the development of several automated electrophysiology platforms has greatly increased the screen throughput of whole cell electrophysiological recordings. New advancements in the automated patch clamp systems have aimed to provide high data quality, high content, and high throughput. However, due to the limitations noted above, automated patch clamp systems are not capable of replacing manual patch clamp systems in ion channel research. While automated patch clamp systems are useful for screening large amounts of compounds in cell lines that stably express high levels of ion channels, the manual patch clamp technique is still necessary for studying ion channel properties in some research areas and for specific cell types, including primary cells that have mixed cell types and differentiated cells that derive from induced pluripotent stem cells (iPSCs) or embryonic stem cells (ESCs). Therefore, further improvements in flexibility with regard to cell types and data quality will broaden the applications of the automated patch clamp systems in both academia and industry. PMID:23346269

Ion channel remodeling in vascular smooth muscle during hypertension: Implications for novel therapeutic approaches

PubMed Central

Joseph, Biny K.; Thakali, Keshari M.; Moore, Christopher L.; Rhee, Sung W.

2013-01-01

Ion channels are multimeric, transmembrane proteins that selectively mediate ion flux across the plasma membrane in a variety of cells including vascular smooth muscle cells (VSMCs). The dynamic interplay of Ca2+ and K+ channels on the plasma membrane of VSMCs plays a pivotal role in modulating the vascular tone of small arteries and arterioles. The abnormally-elevated arterial tone observed in hypertension thus points to an aberrant expression and function of Ca2+ and K+ channels in the VSMCs. In this short review, we focus on the three well-studied ion channels in VSMCs, namely the L-type Ca2+ (CaV1.2) channels, the voltage-gated K+ (KV) channels, and the large-conductance Ca2+-activated K+ (BK) channels. First, we provide a brief overview on the physiological role of vascular CaV1.2, KV and BK channels in regulating arterial tone. Second, we discuss the current understanding of the expression changes and regulation of CaV1.2, KV and BK channels in the vasculature during hypertension. Third, based on available proof-of-concept studies, we describe the potential therapeutic approaches targeting these vascular ion channels in order to restore blood pressure to normotensive levels. PMID:23376354

Trafficking Mechanisms Underlying Neuronal Voltage-gated Ion Channel Localization at the Axon Initial Segment

PubMed Central

Vacher, Helene; Trimmer, James S.

2012-01-01

Summary Voltage-gated ion channels are diverse and fundamental determinants of neuronal intrinsic excitability. Voltage-gated K+ (Kv) and Na+ (Nav) channels play complex yet fundamentally important roles in determining intrinsic excitability. The Kv and Nav channels located at the axon initial segment (AIS) play a unique and especially important role in generating neuronal output in the form of anterograde axonal and backpropagating action potentials, Aberrant intrinsic excitability in individual neurons within networks contributes to synchronous neuronal activity leading to seizures. Mutations in ion channel genes gives rise to a variety of seizure-related “Channelopathies”, and many of the ion channel subunits associated with epilepsy mutations are localized at the AIS, making this a hotspot for epileptogenesis. Here we review the cellular mechanisms that underlie the trafficking of Kv and Nav channels found at the AIS, and how Kv and Nav channel mutations associated with epilepsy can alter these processes. PMID:23216576

A Change in the Ion Selectivity of Ligand-Gated Ion Channels Provides a Mechanism to Switch Behavior.

PubMed

Pirri, Jennifer K; Rayes, Diego; Alkema, Mark J

2015-01-01

Behavioral output of neural networks depends on a delicate balance between excitatory and inhibitory synaptic connections. However, it is not known whether network formation and stability is constrained by the sign of synaptic connections between neurons within the network. Here we show that switching the sign of a synapse within a neural circuit can reverse the behavioral output. The inhibitory tyramine-gated chloride channel, LGC-55, induces head relaxation and inhibits forward locomotion during the Caenorhabditis elegans escape response. We switched the ion selectivity of an inhibitory LGC-55 anion channel to an excitatory LGC-55 cation channel. The engineered cation channel is properly trafficked in the native neural circuit and results in behavioral responses that are opposite to those produced by activation of the LGC-55 anion channel. Our findings indicate that switches in ion selectivity of ligand-gated ion channels (LGICs) do not affect network connectivity or stability and may provide an evolutionary and a synthetic mechanism to change behavior.

Relevance of quantum mechanics on some aspects of ion channel function

PubMed Central

Roy, Sisir

2010-01-01

Mathematical modeling of ionic diffusion along K ion channels indicates that such diffusion is oscillatory, at the weak non-Markovian limit. This finding leads us to derive a Schrödinger–Langevin equation for this kind of system within the framework of stochastic quantization. The Planck’s constant is shown to be relevant to the Lagrangian action at the level of a single ion channel. This sheds new light on the issue of applicability of quantum formalism to ion channel dynamics and to the physical constraints of the selectivity filter. PMID:19520314

HTS techniques for patch clamp-based ion channel screening - advances and economy.

PubMed

Farre, Cecilia; Fertig, Niels

2012-06-01

Ten years ago, the first publication appeared showing patch clamp recordings performed on a planar glass chip instead of using a conventional patch clamp pipette. "Going planar" proved to revolutionize ion channel drug screening as we know it, by allowing high quality measurements of ion channels and their effectors at a higher throughput and at the same time de-skilling the highly laborious technique. Over the years, platforms evolved in response to user requirements regarding experimental features, data handling plus storage, and suitable target diversity. This article gives a snapshot image of patch clamp-based ion channel screening with focus on platforms developed to meet requirements of high-throughput screening environments. The commercially available platforms are described, along with their benefits and drawbacks in ion channel drug screening. Automated patch clamp (APC) platforms allow faster investigation of a larger number of ion channel active compounds or cell clones than previously possible. Since patch clamp is the only method allowing direct, real-time measurements of ion channel activity, APC holds the promise of picking up high quality leads, where they otherwise would have been overseen using indirect methods. In addition, drug candidate safety profiling can be performed earlier in the drug discovery process, avoiding late-phase compound withdrawal due to safety liability issues, which is highly costly and inefficient.

Relevance of Viroporin Ion Channel Activity on Viral Replication and Pathogenesis

PubMed Central

Nieto-Torres, Jose L.; Verdiá-Báguena, Carmina; Castaño-Rodriguez, Carlos; Aguilella, Vicente M.; Enjuanes, Luis

2015-01-01

Modification of host-cell ionic content is a significant issue for viruses, as several viral proteins displaying ion channel activity, named viroporins, have been identified. Viroporins interact with different cellular membranes and self-assemble forming ion conductive pores. In general, these channels display mild ion selectivity, and, eventually, membrane lipids play key structural and functional roles in the pore. Viroporins stimulate virus production through different mechanisms, and ion channel conductivity has been proved particularly relevant in several cases. Key stages of the viral cycle such as virus uncoating, transport and maturation are ion-influenced processes in many viral species. Besides boosting virus propagation, viroporins have also been associated with pathogenesis. Linking pathogenesis either to the ion conductivity or to other functions of viroporins has been elusive for a long time. This article summarizes novel pathways leading to disease stimulated by viroporin ion conduction, such as inflammasome driven immunopathology. PMID:26151305

Theory and simulation of ion conduction in the pentameric GLIC channel.

PubMed

Zhu, Fangqiang; Hummer, Gerhard

2012-10-09

GLIC is a bacterial member of the large family of pentameric ligand-gated ion channels. To study ion conduction through GLIC and other membrane channels, we combine the one-dimensional potential of mean force for ion passage with a Smoluchowski diffusion model, making it possible to calculate single-channel conductance in the regime of low ion concentrations from all-atom molecular dynamics (MD) simulations. We then perform MD simulations to examine sodium ion conduction through the GLIC transmembrane pore in two systems with different bulk ion concentrations. The ion potentials of mean force, calculated from umbrella sampling simulations with Hamiltonian replica exchange, reveal a major barrier at the hydrophobic constriction of the pore. The relevance of this barrier for ion transport is confirmed by a committor function that rises sharply in the barrier region. From the free evolution of Na(+) ions starting at the barrier top, we estimate the effective diffusion coefficient in the barrier region, and subsequently calculate the conductance of the pore. The resulting diffusivity compares well with the position-dependent ion diffusion coefficient obtained from restrained simulations. The ion conductance obtained from the diffusion model agrees with the value determined via a reactive-flux rate calculation. Our results show that the conformation in the GLIC crystal structure, with an estimated conductance of ~1 picosiemens at 140 mM ion concentration, is consistent with a physiologically open state of the channel.

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.

Contractile dysfunctions in ATP-dependent K+ channel-deficient mouse muscle during fatigue involve excessive depolarization and Ca2+ influx through L-type Ca2+ channels.

PubMed

Cifelli, Carlo; Boudreault, Louise; Gong, Bing; Bercier, Jean-Philippe; Renaud, Jean-Marc

2008-10-01

Muscles deficient in ATP-dependent potassium (KATP) channels develop contractile dysfunctions during fatigue that may explain their apparently faster rate of fatigue compared with wild-type muscles. The objectives of this study were to determine: (1) whether the contractile dysfunctions, namely unstimulated force and depressed force recovery, result from excessive membrane depolarization and Ca2+ influx through L-type Ca2+ channels; and (2) whether reducing the magnitude of these two contractile dysfunctions reduces the rate of fatigue in KATP channel-deficient muscles. To reduce Ca2+ influx, we lowered the extracellular Ca2+ concentration ([Ca2+]o) from 2.4 to 0.6 mM or added 1 microM verapamil, an L-type Ca2+ channel blocker. Flexor digitorum brevis (FDB) muscles deficient in KATP channels were obtained by exposing wild-type muscles to 10 microM glibenclamide or by using FDB from Kir6.2-/- mice. Fatigue was elicited with one contraction per second for 3 min at 37 degrees C. In wild-type FDB, lowered [Ca2+]o or verapamil did not affect the decrease in peak tetanic force and unstimulated force during fatigue and force recovery following fatigue. In KATP channel-deficient FDB, lowered [Ca2+]o or verapamil slowed down the decrease in peak tetanic force recovery, reduced unstimulated force and improved force recovery. In Kir6.2-/- FDB, the rate of fatigue became slower than in wild-type FDB in the presence of verapamil. The cell membrane depolarized from -83 to -57 mV in normal wild-type FDB. The depolarizations in some glibenclamide-exposed fibres were similar to those of normal FDB, while in other fibres the cell membrane depolarized to -31 mV in 80 s, which was also the time when these fibres supercontracted. It is concluded that: (1) KATP channels are crucial in preventing excessive membrane depolarization and Ca2+ influx through L-type Ca2+ channels; and (2) they contribute to the decrease in force during fatigue.

Unraveling the mechanism of selective ion transport in hydrophobic subnanometer channels

PubMed Central

Li, Hui; Francisco, Joseph S.; Zeng, Xiao Cheng

2015-01-01

Recently reported synthetic organic nanopore (SONP) can mimic a key feature of natural ion channels, i.e., selective ion transport. However, the physical mechanism underlying the K+/Na+ selectivity for the SONPs is dramatically different from that of natural ion channels. To achieve a better understanding of the selective ion transport in hydrophobic subnanometer channels in general and SONPs in particular, we perform a series of ab initio molecular dynamics simulations to investigate the diffusivity of aqua Na+ and K+ ions in two prototype hydrophobic nanochannels: (i) an SONP with radius of 3.2 Å, and (ii) single-walled carbon nanotubes (CNTs) with radii of 3–5 Å (these radii are comparable to those of the biological potassium K+ channels). We find that the hydration shell of aqua Na+ ion is smaller than that of aqua K+ ion but notably more structured and less yielding. The aqua ions do not lower the diffusivity of water molecules in CNTs, but in SONP the diffusivity of aqua ions (Na+ in particular) is strongly suppressed due to the rugged inner surface. Moreover, the aqua Na+ ion requires higher formation energy than aqua K+ ion in the hydrophobic nanochannels. As such, we find that the ion (K+ vs. Na+) selectivity of the (8, 8) CNT is ∼20× higher than that of SONP. Hence, the (8, 8) CNT is likely the most efficient artificial K+ channel due in part to its special interior environment in which Na+ can be fully solvated, whereas K+ cannot. This work provides deeper insights into the physical chemistry behind selective ion transport in nanochannels. PMID:26283377

Rediscovering sperm ion channels with the patch-clamp technique

PubMed Central

Kirichok, Yuriy; Lishko, Polina V.

2011-01-01

Upon ejaculation, mammalian spermatozoa have to undergo a sequence of physiological transformations within the female reproductive tract that will allow them to reach and fertilize the egg. These include initiation of motility, hyperactivation of motility and perhaps chemotaxis toward the egg, and culminate in the acrosome reaction that permits sperm to penetrate the protective vestments of the egg. These physiological responses are triggered through the activation of sperm ion channels that cause elevations of sperm intracellular pH and Ca2+ in response to certain cues within the female reproductive tract. Despite their key role in sperm physiology and their absolute requirement for the process of fertilization, sperm ion channels remain poorly understood due to the extreme difficulty in application of the patch-clamp technique to spermatozoa. This review covers the topic of sperm ion channels in the following order: first, we discuss how the intracellular Ca2+ and pH signaling mediated by sperm ion channels controls sperm behavior during the process of fertilization. Then, we briefly cover the history of the methodology to study sperm ion channels, which culminated in the recent development of a reproducible whole-cell patch-clamp technique for mouse and human cells. We further discuss the main approaches used to patch-clamp mature mouse and human spermatozoa. Finally, we focus on the newly discovered sperm ion channels CatSper, KSper (Slo3) and HSper (Hv1), identified by the sperm patch-clamp technique. We conclude that the patch-clamp technique has markedly improved and shifted our understanding of the sperm ion channels, in addition to revealing significant species-specific differences in these channels. This method is critical for identification of the molecular mechanisms that control sperm behavior within the female reproductive tract and make fertilization possible. PMID:21642646

Amphotericin B channels in phospholipid membrane-coated nanoporous silicon surfaces: implications for photovoltaic driving of ions across membranes.

PubMed

Yilma, Solomon; Liu, Nangou; Samoylov, Alexander; Lo, Ting; Brinker, C Jeffrey; Vodyanoy, Vitaly

2007-03-15

The antimycotic agent amphotericin B (AmB) functions by forming complexes with sterols to form ion channels that cause membrane leakage. When AmB and cholesterol mixed at 2:1 ratio were incorporated into phospholipid bilayer membranes formed on the tip of patch pipettes, ion channel current fluctuations with characteristic open and closed states were observed. These channels were also functional in phospholipid membranes formed on nanoporous silicon surfaces. Electrophysiological studies of AmB-cholesterol mixtures that were incorporated into phospholipid membranes formed on the surface of nanoporous (6.5 nm pore diameter) silicon plates revealed large conductance ion channels ( approximately 300 pS) with distinct open and closed states. Currents through the AmB-cholesterol channels on nanoporous silicon surfaces can be driven by voltage applied via conventional electrical circuits or by photovoltaic electrical potential entirely generated when the nanoporous silicon surface is illuminated with a narrow laser beam. Electrical recordings made during laser illumination of AmB-cholesterol containing membrane-coated nanoporous silicon surfaces revealed very large conductance ion channels with distinct open and closed states. Our findings indicate that nanoporous silicon surfaces can serve as mediums for ion-channel-based biosensors. The photovoltaic properties of nanoporous silicon surfaces show great promise for making such biosensors addressable via optical technologies.

Skin Barrier and Calcium.

PubMed

Lee, Sang Eun; Lee, Seung Hun

2018-06-01

Epidermal barrier formation and the maintenance of barrier homeostasis are essential to protect us from the external environments and organisms. Moreover, impaired keratinocytes differentiation and dysfunctional skin barrier can be the primary causes or aggravating factors for many inflammatory skin diseases including atopic dermatitis and psoriasis. Therefore, understanding the regulation mechanisms of keratinocytes differentiation and skin barrier homeostasis is important to understand many skin diseases and establish an effective treatment strategy. Calcium ions (Ca 2+ ) and their concentration gradient in the epidermis are essential in regulating many skin functions, including keratinocyte differentiation, skin barrier formation, and permeability barrier homeostasis. Recent studies have suggested that the intracellular Ca 2+ stores such as the endoplasmic reticulum (ER) are the major components that form the epidermal calcium gradient and the ER calcium homeostasis is crucial for regulating keratinocytes differentiation, intercellular junction formation, antimicrobial barrier, and permeability barrier homeostasis. Thus, both Ca 2+ release from intracellular stores, such as the ER and Ca 2+ influx mechanisms are important in skin barrier. In addition, growing evidences identified the functional existence and the role of many types of calcium channels which mediate calcium flux in keratinocytes. In this review, the origin of epidermal calcium gradient and their role in the formation and regulation of skin barrier are focused. We also focus on the role of ER calcium homeostasis in skin barrier. Furthermore, the distribution and role of epidermal calcium channels, including transient receptor potential channels, store-operated calcium entry channel Orai1, and voltage-gated calcium channels in skin barrier are discussed.

Free-Energy Calculations. A Mathematical Perspective

NASA Technical Reports Server (NTRS)

Pohorille, Andrzej

2015-01-01

Ion channels are pore-forming assemblies of transmembrane proteins that mediate and regulate ion transport through cell walls. They are ubiquitous to all life forms. In humans and other higher organisms they play the central role in conducting nerve impulses. They are also essential to cardiac processes, muscle contraction and epithelial transport. Ion channels from lower organisms can act as toxins or antimicrobial agents, and in a number of cases are involved in infectious diseases. Because of their important and diverse biological functions they are frequent targets of drug action. Also, simple natural or synthetic channels find numerous applications in biotechnology. For these reasons, studies of ion channels are at the forefront of biophysics, structural biology and cellular biology. In the last decade, the increased availability of X-ray structures has greatly advanced our understanding of ion channels. However, their mechanism of action remains elusive. This is because, in order to assist controlled ion transport, ion channels are dynamic by nature, but X-ray crystallography captures the channel in a single, sometimes non-native state. To explain how ion channels work, X-ray structures have to be supplemented with dynamic information. In principle, molecular dynamics (MD) simulations can aid in providing this information, as this is precisely what MD has been designed to do. However, MD simulations suffer from their own problems, such as inability to access sufficiently long time scales or limited accuracy of force fields. To assess the reliability of MD simulations it is only natural to turn to the main function of channels - conducting ions - and compare calculated ionic conductance with electrophysiological data, mainly single channel recordings, obtained under similar conditions. If this comparison is satisfactory it would greatly increase our confidence that both the structures and our computational methodologies are sufficiently accurate. Channel conductance, defined as the ratio of ionic current through the channel to applied voltage, can be calculated in MD simulations by way of applying an external electric field to the system and counting the number of ions that traverse the channel per unit time. If the current is small, a voltage significantly higher than the experimental one needs to be applied to collect sufficient statistics of ion crossing events. Then, the calculated conductance has to be extrapolated to the experimental voltage using procedures of unknown accuracy. Instead, we propose an alternative approach that applies if ion transport through channels can be described with sufficient accuracy by the one-dimensional diffusion equation in the potential given by the free energy profile and applied voltage. Then, it is possible to test the assumptions of the equation, recover the full voltage/current dependence, determine the reliability of the calculated conductance and reconstruct the underlying (equilibrium) free energy profile, all from MD simulations at a single voltage. We will present the underlying theory, model calculations that test this theory and simulations on ion conductance through a channel that has been extensively studied experimentally. To our knowledge this is the first case in which the complete, experimentally measured dependence of the current on applied voltage has been reconstructed from MD simulations.

Molecular Dynamics Simulation of the Antiamoebin Ion Channel: Linking Structure and Conductance

NASA Technical Reports Server (NTRS)

Wilson, Michael A.; Wei, Chenyu; Bjelkmar, Paer; Wallace, B. A.; Pohorille, Andrew

2011-01-01

Molecular dynamics simulations were carried out in order to ascertain which of the potential multimeric forms of the transmembrane peptaibol channel, antiamoebin, is consistant with its measured conductance. Estimates of the conductance obtained through counting ions that cross the channel and by solving the Nernst-Planck equation yield consistent results, indicating that the motion of ions inside the channel can be satisfactorily described as diffusive.The calculated conductance of octameric channels is markedly higher than the conductance measured in single channel recordings, whereas the tetramer appears to be non-conducting. The conductance of the hexamer was estimated to be 115+/-34 pS and 74+/-20 pS, at 150 mV and 75 mV, respectively, in satisfactory agreement with the value of 90 pS measured at 75 mV. On this basis we propose that the antiamoebin channel consists of six monomers. Its pore is large enough to accommodate K(+) and Cl(-) with their first solvation shells intact. The free energy barrier encountered by K(+) is only 2.2 kcal/mol whereas Cl(-) encounters a substantially higher barrier of nearly 5 kcal/mol. This difference makes the channel selective for cations. Ion crossing events are shown to be uncorrelated and follow Poisson statistics. keywords: ion channels, peptaibols, channel conductance, molecular dynamics

Toxic β-Amyloid (Aβ) Alzheimer's Ion Channels: From Structure to Function and Design

NASA Astrophysics Data System (ADS)

Nussinov, Ruth

2012-02-01

Full-length amyloid beta peptides (Aβ1-40/42) form neuritic amyloid plaques in Alzheimer's disease (AD) patients and are implicated in AD pathology. Recent biophysical and cell biological studies suggest a direct mechanism of amyloid beta toxicity -- ion channel mediated loss of calcium homeostasis. Truncated amyloid beta fragments (Aβ11-42 and Aβ17-42), commonly termed as non-amyloidogenic are also found in amyloid plaques of Alzheimer's disease (AD) and in the preamyloid lesions of Down's syndrome (DS), a model system for early onset AD study. Very little is known about the structure and activity of these smaller peptides although they could be key AD and DS pathological agents. Using complementary techniques of explicit solvent molecular dynamics (MD) simulations, atomic force microscopy (AFM), channel conductance measurements, cell calcium uptake assays, neurite degeneration and cell death assays, we have shown that non-amyloidogenic Aβ9-42 and Aβ17-42 peptides form ion channels with loosely attached subunits and elicit single channel conductances. The subunits appear mobile suggesting insertion of small oligomers, followed by dynamic channel assembly and dissociation. These channels allow calcium uptake in APP-deficient cells and cause neurite degeneration in human cortical neurons. Channel conductance, calcium uptake and neurite degeneration are selectively inhibited by zinc, a blocker of amyloid ion channel activity. Thus truncated Aβ fragments could account for undefined roles played by full length Aβs and provide a novel mechanism of AD and DS pathology. The emerging picture from our large-scale simulations is that toxic ion channels formed by β-sheets are highly polymorphic, and spontaneously break into loosely interacting dynamic units (though still maintaining ion channel structures as imaged with AFM), that associate and dissociate leading to toxic ion flux. This sharply contrasts intact conventional gated ion channels that consist of tightly interacting α-helices that robustly prevent ion leakage, rather than hydrogen-bonded β-strands. Moreover, in comparison with β-rich antimicrobial peptide (AMP) such as a protegrin-1 (PG-1), both Aβ and PG-1 are cytotoxic, and capable of forming fibrils and dynamic channels which consist of subunits with similar dimensions. These combined properties support a functional relationship between amyloidogenic peptides and β-sheet-rich cytolytic AMPs, suggesting that PG-1 is amyloidogenic and amyloids may have an antimicrobial function.

Divalent ions are potential permeating blockers of the non-selective NaK ion channel: combined QM and MD based investigations.

PubMed

Sadhu, Biswajit; Sundararajan, Mahesh; Bandyopadhyay, Tusar

2017-10-18

The bacterial NaK ion channel is distinctly different from other known ion channels due to its inherent non-selective feature. One of the unexplored and rather interesting features is its ability to permeate divalent metal ions (such as Ca 2+ and Ba 2+ ) and not monovalent alkali metal ions. Several intriguing questions about the energetics and structural aspects still remain unanswered. For instance, what causes Ca 2+ to permeate as well as block the selectivity filter (SF) of the NaK ion channel and act as a "permeating blocker"? How and at what energetic cost does another chemical congener, Sr 2+ , as well as Ba 2+ , a potent blocker of the K + ion channel, permeate through the SF of the NaK ion channel? Finally, how do their translocation energetics differ from those of monovalent ions such as K + ? Here, in an attempt to address these outstanding issues, we elucidate the structure, binding and selectivity of divalent ions (Ca 2+ , Sr 2+ and Ba 2+ ) as they permeate through the SF of the NaK ion channel using all-atom molecular dynamics simulations and density functional theory based calculations. We unveil mechanistic insight into this translocation event using well-tempered metadynamics simulations in a polarizable environment using the mean-field model of water and incorporating electronic continuum corrections for ions via charge rescaling. The results show that, akin to K + coordination, Sr 2+ and Ba 2+ bind at the SF in a very similar fashion and remain octa-coordinated at all sites. Interestingly, differing from its local hydration structure, Ca 2+ interacts with eight carbonyls to remain at the middle of the S3 site. Furthermore, the binding of divalent metals at SF binding sites is more favorable than the binding of K + . However, their permeation through the extracellular entrance faces a considerably higher energetic barrier compared to that for K + , which eventually manifests their inherent blocking feature.

Sperm-specific ion channels: targets holding the most potential for male contraceptives in development.

PubMed

Zheng, Li-Ping; Wang, Hua-Feng; Li, Bao-Ming; Zeng, Xu-Hui

2013-10-01

There is a global need for an ideal method of male contraception. However, the development of male contraceptives has not been well successful. Research on sperm-specific ion channels, especially the recent advance obtained from electrophysiological studies, has emphasized the conception that those channels are targets with the most potential to develop non-hormonal male contraceptives. While summarizing the general options for male contraception, this review focuses on the properties and functions of sperm ion channels together with the attempts of utilizing these channels to develop male contraceptives. We believe that a deeper insight into the signaling and molecular mechanisms by which ion channels regulate sperm functions will pave the way for developing novel male-based contraceptives. Copyright © 2013 Elsevier Inc. All rights reserved.

Population patch clamp electrophysiology: a breakthrough technology for ion channel screening.

PubMed

Dale, Tim J; Townsend, Claire; Hollands, Emma C; Trezise, Derek J

2007-10-01

Population patch clamp (PPC) is a novel high throughput planar array electrophysiology technique that allows ionic currents to be recorded from populations of cells under voltage clamp. For the drug discovery pharmacologist, PPC promises greater speed and precision than existing methods for screening compounds at voltage-gated ion channel targets. Moreover, certain constitutively active or slow-ligand gated channels that have hitherto proved challenging to screen with planar array electrophysiology (e.g. SK/IK channels) are now more accessible. In this article we review early findings using PPC and provide a perspective on its likely impact on ion channel drug discovery. To support this, we include some new data on ion channel assay duplexing and on modulator assays, approaches that have thus far not been described.

A membrane-embedded pathway delivers general anesthetics to two interacting binding sites in the Gloeobacter violaceus ion channel.

PubMed

Arcario, Mark J; Mayne, Christopher G; Tajkhorshid, Emad

2017-06-09

General anesthetics exert their effects on the central nervous system by acting on ion channels, most notably pentameric ligand-gated ion channels. Although numerous studies have focused on pentameric ligand-gated ion channels, the details of anesthetic binding and channel modulation are still debated. A better understanding of the anesthetic mechanism of action is necessary for the development of safer and more efficacious drugs. Herein, we present a computational study identifying two anesthetic binding sites in the transmembrane domain of the Gloeobacter violaceus ligand-gated ion channel (GLIC) channel, characterize the putative binding pathway, and observe structural changes associated with channel function. Molecular simulations of desflurane reveal a binding pathway to GLIC via a membrane-embedded tunnel using an intrasubunit protein lumen as the conduit, an observation that explains the Meyer-Overton hypothesis, or why the lipophilicity of an anesthetic and its potency are generally proportional. Moreover, employing high concentrations of ligand led to the identification of a second transmembrane site (TM2) that inhibits dissociation of anesthetic from the TM1 site and is consistent with the high concentrations of anesthetics required to achieve clinical effects. Finally, asymmetric binding patterns of anesthetic to the channel were found to promote an iris-like conformational change that constricts and dehydrates the ion pore, creating a 13.5 kcal/mol barrier to ion translocation. Together with previous studies, the simulations presented herein demonstrate a novel anesthetic binding site in GLIC that is accessed through a membrane-embedded tunnel and interacts with a previously known site, resulting in conformational changes that produce a non-conductive state of the channel. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Transcriptomic analysis of the ion channelome of human platelets and megakaryocytic cell lines.

PubMed

Wright, Joy R; Amisten, Stefan; Goodall, Alison H; Mahaut-Smith, Martyn P

2016-08-01

Ion channels have crucial roles in all cell types and represent important therapeutic targets. Approximately 20 ion channels have been reported in human platelets; however, no systematic study has been undertaken to define the platelet channelome. These membrane proteins need only be expressed at low copy number to influence function and may not be detected using proteomic or transcriptomic microarray approaches. In our recent work, quantitative real-time PCR (qPCR) provided key evidence that Kv1.3 is responsible for the voltage-dependent K+ conductance of platelets and megakaryocytes. The present study has expanded this approach to assess relative expression of 402 ion channels and channel regulatory genes in human platelets and three megakaryoblastic/erythroleukaemic cell lines. mRNA levels in platelets are low compared to other blood cells, therefore an improved method of isolating platelets was developed. This used a cocktail of inhibitors to prevent formation of leukocyte-platelet aggregates, and a combination of positive and negative immunomagnetic cell separation, followed by rapid extraction of mRNA. Expression of 34 channel-related transcripts was quantified in platelets, including 24 with unknown roles in platelet function, but that were detected at levels comparable to ion channels with established roles in haemostasis or thrombosis. Trace expression of a further 50 ion channel genes was also detected. More extensive channelomes were detected in MEG-01, CHRF-288-11 and HEL cells (195, 185 and 197 transcripts, respectively), but lacked several channels observed in the platelet. These "channelome" datasets provide an important resource for further studies of ion channel function in the platelet and megakaryocyte.

Band gap engineering by swift heavy ions irradiation induced amorphous nano-channels in LiNbO3

DOE PAGES

Sachan, Ritesh; Pakarinen, Olli H.; Liu, Peng; ...

2015-04-01

The irradiation of lithium niobate with swift heavy ions results in the creation of amorphous nano-sized channels along the incident ion path. These nano-channels are on the order of a hundred microns in length and could be useful for photonic applications. However, there are two major challenges in these nano-channels characterization; (i) it is difficult to investigate the structural characteristics of these nano-channels due to their very long length, and (ii) the analytical electron microscopic analysis of individual ion track is complicated due to electron beam sensitive nature of lithium niobate. Here, we report the first high resolution microscopic characterizationmore » of these amorphous nano-channels, widely known as ion-tracks, by direct imaging them at different depths in the material, and subsequently correlating the key characteristics with Se of ions. Energetic Kr ions ( 84Kr 22 with 1.98 GeV energy) are used to irradiate single crystal lithium niobate with a fluence of 2x10 10 ions/cm 2, which results in the formation of individual ion tracks with a penetration depth of ~180 μm. Along the ion path, electron energy loss of the ions, which is responsible for creating the ion tracks, increases with depth under these conditions in LiNbO 3, resulting in increases in track diameter of a factor of ~2 with depth. This diameter increase with electronic stopping power is consistent with predictions of the inelastic thermal spike model. We also show a new method to measure the band gap in individual ion track by using electron energy-loss spectroscopy.« less

Malaria parasite mutants with altered erythrocyte permeability: a new drug resistance mechanism and important molecular tool

PubMed Central

Hill, David A; Desai, Sanjay A

2010-01-01

Erythrocytes infected with plasmodia, including those that cause human malaria, have increased permeability to a diverse collection of organic and inorganic solutes. While these increases have been known for decades, their mechanistic basis was unclear until electrophysiological studies revealed flux through one or more ion channels on the infected erythrocyte membrane. Current debates have centered on the number of distinct ion channels, which channels mediate the transport of each solute and whether the channels represent parasite-encoded proteins or human channels activated after infection. This article reviews the identification of the plasmodial surface anion channel and other proposed channels with an emphasis on two distinct channel mutants generated through in vitro selection. These mutants implicate parasite genetic elements in the parasite-induced permeability, reveal an important new antimalarial drug resistance mechanism and provide tools for molecular studies. We also critically examine the technical issues relevant to the detection of ion channels by electrophysiological methods; these technical considerations have general applicability for interpreting studies of various ion channels proposed for the infected erythrocyte membrane. PMID:20020831

Expression and distribution of voltage-gated ion channels in ferret sinoatrial node.

PubMed

Brahmajothi, Mulugu V; Morales, Michael J; Campbell, Donald L; Steenbergen, Charles; Strauss, Harold C

2010-10-01

Spontaneous diastolic depolarization in the sinoatrial (SA) node enables it to serve as pacemaker of the heart. The variable cell morphology within the SA node predicts that ion channel expression would be heterogeneous and different from that in the atrium. To evaluate ion channel heterogeneity within the SA node, we used fluorescent in situ hybridization to examine ion channel expression in the ferret SA node region and atrial appendage. SA nodal cells were distinguished from surrounding cardiac myocytes by expression of the slow (SA node) and cardiac (surrounding tissue) forms of troponin I. Nerve cells in the sections were identified by detection of GAP-43 and cytoskeletal middle neurofilament. Transcript expression was characterized for the 4 hyperpolarization-activated cation channels, 6 voltage-gated Na(+) channels, 3 voltage-gated Ca(2+) channels, 24 voltage-gated K(+) channel α-subunits, and 3 ancillary subunits. To ensure that transcript expression was representative of protein expression, immunofluorescence was used to verify localization patterns of voltage-dependent K(+) channels. Colocalizations were performed to observe any preferential patterns. Some overlapping and nonoverlapping binding patterns were observed. Measurement of different cation channel transcripts showed heterogeneous expression with many different patterns of expression, attesting to the complexity of electrical activity in the SA node. This study provides insight into the possible role ion channel heterogeneity plays in SA node pacemaker activity.

Electric Field-Controlled Ion Transport In TiO2 Nanochannel.

PubMed

Li, Dan; Jing, Wenheng; Li, Shuaiqiang; Shen, Hao; Xing, Weihong

2015-06-03

On the basis of biological ion channels, we constructed TiO2 membranes with rigid channels of 2.3 nm to mimic biomembranes with flexible channels; an external electric field was employed to regulate ion transport in the confined channels at a high ionic strength in the absence of electrical double layer overlap. Results show that transport rates for both Na+ and Mg2+ were decreased irrespective of the direction of the electric field. Furthermore, a voltage-gated selective ion channel was formed, the Mg2+ channel closed at -2 V, and a reversed relative electric field gradient was at the same order of the concentration gradient, whereas the Na+ with smaller Stokes radius and lower valence was less sensitive to the electric field and thus preferentially occupied and passed the channel. Thus, when an external electric field is applied, membranes with larger nanochannels have promising applications in selective separation of mixture salts at a high concentration.

Moving Fe2+ from ferritin ion channels to catalytic OH centers depends on conserved protein cage carboxylates

PubMed Central

Behera, Rabindra K.; Theil, Elizabeth C.

2014-01-01

Ferritin biominerals are protein-caged metabolic iron concentrates used for iron–protein cofactors and oxidant protection (Fe2+ and O2 sequestration). Fe2+ passage through ion channels in the protein cages, like membrane ion channels, required for ferritin biomineral synthesis, is followed by Fe2+ substrate movement to ferritin enzyme (Fox) sites. Fe2+ and O2 substrates are coupled via a diferric peroxo (DFP) intermediate, λmax 650 nm, which decays to [Fe3+–O–Fe3+] precursors of caged ferritin biominerals. Structural studies show multiple conformations for conserved, carboxylate residues E136 and E57, which are between ferritin ion channel exits and enzymatic sites, suggesting functional connections. Here we show that E136 and E57 are required for ferritin enzyme activity and thus are functional links between ferritin ion channels and enzymatic sites. DFP formation (Kcat and kcat/Km), DFP decay, and protein-caged hydrated ferric oxide accumulation decreased in ferritin E57A and E136A; saturation required higher Fe2+ concentrations. Divalent cations (both ion channel and intracage binding) selectively inhibit ferritin enzyme activity (block Fe2+ access), Mn2+ << Co2+ < Cu2+ < Zn2+, reflecting metal ion–protein binding stabilities. Fe2+–Cys126 binding in ferritin ion channels, observed as Cu2+–S–Cys126 charge-transfer bands in ferritin E130D UV-vis spectra and resistance to Cu2+ inhibition in ferritin C126S, was unpredicted. Identifying E57 and E136 links in Fe2+ movement from ferritin ion channels to ferritin enzyme sites completes a bucket brigade that moves external Fe2+ into ferritin enzymatic sites. The results clarify Fe2+ transport within ferritin and model molecular links between membrane ion channels and cytoplasmic destinations. PMID:24843174

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

Different structural requirements for functional ion pore transplantation suggest different gating mechanisms of NMDA and kainate receptors.

PubMed

Villmann, Carmen; Hoffmann, Jutta; Werner, Markus; Kott, Sabine; Strutz-Seebohm, Nathalie; Nilsson, Tanja; Hollmann, Michael

2008-10-01

Although considerable progress has been made in characterizing the physiological function of the high-affinity kainate (KA) receptor subunits KA1 and KA2, no homomeric ion channel function has been shown. An ion channel transplantation approach was employed in this study to directly test if homomerically expressed KA1 and KA2 pore domains are capable of conducting currents. Transplantation of the ion pore of KA1 or KA2 into GluR6 generated perfectly functional ion channels that allowed characterization of those electrophysiological and pharmacological properties that are determined exclusively by the ion pore of KA1 or KA2. This demonstrates for the first time that KA1 and KA2 ion pore domains are intrinsically capable of conducting ions even in homomeric pore assemblies. NMDA receptors, similar to KA1- or KA2-containing receptors, function only as heteromeric complexes. They are composed of NR1 and NR2 subunits, which both are non-functional when expressed homomerically. In contrast to NR1, the homomeric NR2B ion pore failed to translate ligand binding into pore opening when transplanted into GluR6. Similarly, heteromeric coexpression of the ion channel domains of both NR1 and NR2 inserted into GluR6 failed to produce functional channels. Therefore, we conclude that the mechanism underlying the ion channel opening in the obligatorily heterotetrameric NMDA receptors differs significantly from that in the facultatively heterotetrameric alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate and KA receptors.

Functional K(ATP) channels in the rat retinal microvasculature: topographical distribution, redox regulation, spermine modulation and diabetic alteration.

PubMed

Ishizaki, Eisuke; Fukumoto, Masanori; Puro, Donald G

2009-05-15

The essential task of the circulatory system is to match blood flow to local metabolic demand. However, much remains to be learned about this process. To better understand how local perfusion is regulated, we focused on the functional organization of the retinal microvasculature, which is particularly well adapted for the local control of perfusion. Here, we assessed the distribution and regulation of functional K(ATP) channels whose activation mediates the hyperpolarization induced by adenosine. Using microvascular complexes freshly isolated from the rat retina, we found a topographical heterogeneity in the distribution of functional K(ATP) channels; capillaries generate most of the K(ATP) current. The initiation of K(ATP)-induced responses in the capillaries supports the concept that the regulation of retinal perfusion is highly decentralized. Additional study revealed that microvascular K(ATP) channels are redox sensitive, with oxidants increasing their activity. Furthermore, the oxidant-mediated activation of these channels is driven by the polyamine spermine, whose catabolism produces oxidants. In addition, our observation that spermine-dependent oxidation occurs predominately in the capillaries accounts for why they generate most of the K(ATP) current detected in retinal microvascular complexes. Here, we also analysed retinal microvessels of streptozotocin-injected rats. We found that soon after the onset of diabetes, an increase in spermine-dependent oxidation at proximal microvascular sites boosts their K(ATP) current and thereby virtually eliminates the topographical heterogeneity of functional K(ATP) channels. We conclude that spermine-dependent oxidation is a previously unrecognized mechanism by which this polyamine modulates ion channels; in addition to a physiological role, spermine-dependent oxidation may also contribute to microvascular dysfunction in the diabetic retina.

Ion channel electrophysiology via integrated planar patch-clamp chip with on-demand drug exchange.

PubMed

Chen, Chang-Yu; Tu, Ting-Yuan; Jong, De-Shien; Wo, Andrew M

2011-06-01

Planar patch clamp has revolutionized characterization of ion channel behavior in drug discovery primarily via advancement in high throughput. Lab use of planar technology, however, addresses different requirements and suffers from inflexibility to enable wide range of interrogation via a single cell. This work presents integration of planar patch clamp with microfluidics, achieving multiple solution exchanges for tailor-specific measurement and allowing rapid replacement of the cell-contacting aperture. Studies via endogenously expressed ion channels in HEK 293T cells were commenced to characterize the device. Results reveal the microfluidic concentration generator produces distinct solution/drug combination/concentrations on-demand. Volume-regulated chloride channel and voltage-gated potassium channels in HEK 293T cells immersed in generated solutions under various osmolarities or drug concentrations show unique channel signature under specific condition. Excitation and blockage of ion channels in a single cell was demonstrated via serial solution exchange. Robustness of the reversible bonding and ease of glass substrate replacement were proven via repeated usage of the integrated device. The present approach reveals the capability and flexibility of integrated microfluidic planar patch-clamp system for ion channel assays. Copyright © 2011 Wiley Periodicals, Inc.

Activation of the mechanosensitive ion channel MscL by mechanical stimulation of supported Droplet-Hydrogel bilayers

PubMed Central

Rosholm, Kadla R.; Baker, Matthew A. B.; Ridone, Pietro; Nakayama, Yoshitaka; Rohde, Paul R.; Cuello, Luis G.; Lee, Lawrence K.; Martinac, Boris

2017-01-01

The droplet on hydrogel bilayer (DHB) is a novel platform for investigating the function of ion channels. Advantages of this setup include tight control of all bilayer components, which is compelling for the investigation of mechanosensitive (MS) ion channels, since they are highly sensitive to their lipid environment. However, the activation of MS ion channels in planar supported lipid bilayers, such as the DHB, has not yet been established. Here we present the activation of the large conductance MS channel of E. coli, (MscL), in DHBs. By selectively stretching the droplet monolayer with nanolitre injections of buffer, we induced quantifiable DHB tension, which could be related to channel activity. The MscL activity response revealed that the droplet monolayer tension equilibrated over time, likely by insertion of lipid from solution. Our study thus establishes a method to controllably activate MS channels in DHBs and thereby advances studies of MS channels in this novel platform. PMID:28345591

One-channel Cell-attached Patch-clamp Recording

PubMed Central

Maki, Bruce A.; Cummings, Kirstie A.; Paganelli, Meaghan A.; Murthy, Swetha E.; Popescu, Gabriela K.

2014-01-01

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate depends on properties intrinsic to each channel protein as well as the mechanism by which it is generated and controlled and represents an important area of current research. Information about the operational dynamics of ion channel proteins can be obtained by observing long stretches of current produced by a single molecule. Described here is a protocol for obtaining one-channel cell-attached patch-clamp current recordings for a ligand gated ion channel, the NMDA receptor, expressed heterologously in HEK293 cells or natively in cortical neurons. Also provided are instructions on how to adapt the method to other ion channels of interest by presenting the example of the mechano-sensitive channel PIEZO1. This method can provide data regarding the channel’s conductance properties and the temporal sequence of open-closed conformations that make up the channel’s activation mechanism, thus helping to understand their functions in health and disease. PMID:24961614

Piezo proteins are pore-forming subunits of mechanically activated channels.

PubMed

Coste, Bertrand; Xiao, Bailong; Santos, Jose S; Syeda, Ruhma; Grandl, Jörg; Spencer, Kathryn S; Kim, Sung Eun; Schmidt, Manuela; Mathur, Jayanti; Dubin, Adrienne E; Montal, Mauricio; Patapoutian, Ardem

2012-02-19

Mechanotransduction has an important role in physiology. Biological processes including sensing touch and sound waves require as-yet-unidentified cation channels that detect pressure. Mouse Piezo1 (MmPiezo1) and MmPiezo2 (also called Fam38a and Fam38b, respectively) induce mechanically activated cationic currents in cells; however, it is unknown whether Piezo proteins are pore-forming ion channels or modulate ion channels. Here we show that Drosophila melanogaster Piezo (DmPiezo, also called CG8486) also induces mechanically activated currents in cells, but through channels with remarkably distinct pore properties including sensitivity to the pore blocker ruthenium red and single channel conductances. MmPiezo1 assembles as a ∼1.2-million-dalton homo-oligomer, with no evidence of other proteins in this complex. Purified MmPiezo1 reconstituted into asymmetric lipid bilayers and liposomes forms ruthenium-red-sensitive ion channels. These data demonstrate that Piezo proteins are an evolutionarily conserved ion channel family involved in mechanotransduction.

Existence of efficient divalent metal ion-catalyzed and inefficient divalent metal ion-independent channels in reactions catalyzed by a hammerhead ribozyme

PubMed Central

Zhou, Jing-Min; Zhou, De-Min; Takagi, Yasuomi; Kasai, Yasuhiro; Inoue, Atsushi; Baba, Tadashi; Taira, Kazunari

2002-01-01

The hammerhead ribozyme is generally accepted as a well characterized metalloenzyme. However, the precise nature of the interactions of the RNA with metal ions remains to be fully defined. Examination of metal ion-catalyzed hammerhead reactions at limited concentrations of metal ions is useful for evaluation of the role of metal ions, as demonstrated in this study. At concentrations of Mn2+ ions from 0.3 to 3 mM, addition of the ribozyme to the reaction mixture under single-turnover conditions enhances the reaction with the product reaching a fixed maximum level. Further addition of the ribozyme inhibits the reaction, demonstrating that a certain number of divalent metal ions is required for proper folding and also for catalysis. At extremely high concentrations, monovalent ions, such as Na+ ions, can also serve as cofactors in hammerhead ribozyme-catalyzed reactions. However, the catalytic efficiency of monovalent ions is extremely low and, thus, high concentrations are required. Furthermore, addition of monovalent ions to divalent metal ion-catalyzed hammerhead reactions inhibits the divalent metal ion-catalyzed reactions, suggesting that the more desirable divalent metal ion–ribozyme complexes are converted to less desirable monovalent metal ion–ribozyme complexes via removal of divalent metal ions, which serve as a structural support in the ribozyme complex. Even though two channels appear to exist, namely an efficient divalent metal ion-catalyzed channel and an inefficient monovalent metal ion-catalyzed channel, it is clear that, under physiological conditions, hammerhead ribozymes are metalloenzymes that act via the significantly more efficient divalent metal ion-dependent channel. Moreover, the observed kinetic data are consistent with Lilley’s and DeRose’s two-phase folding model that was based on ground state structure analyses. PMID:12034824

Using iPSC Models to Probe Regulation of Cardiac Ion Channel Function.

PubMed

Bruyneel, Arne A N; McKeithan, Wesley L; Feyen, Dries A M; Mercola, Mark

2018-05-25

Cardiovascular disease is the leading contributor to mortality and morbidity. Many deaths of heart failure patients can be attributed to sudden cardiac death due primarily to ventricular arrhythmia. Currently, most anti-arrhythmics modulate ion channel conductivity or β-adrenergic signaling, but these drugs have limited efficacy for some indications, and can potentially be proarrhythmic. Recent studies have shown that mutations in proteins other than cardiac ion channels may confer susceptibility to congenital as well as acquired arrhythmias. Additionally, ion channels themselves are subject to regulation at the levels of channel expression, trafficking and post-translational modification; thus, research into the regulation of ion channels may elucidate disease mechanisms and potential therapeutic targets for future drug development. This review summarizes the current knowledge of the molecular mechanisms of arrhythmia susceptibility and discusses technological advances such as induced pluripotent stem cell-derived cardiomyocytes, gene editing, functional genomics, and physiological screening platforms that provide a new paradigm for discovery of new therapeutic targets to treat congenital and acquired diseases of the heart rhythm.

A Secondary Structural Transition in the C-helix Promotes Gating of Cyclic Nucleotide-regulated Ion Channels*

PubMed Central

Puljung, Michael C.; Zagotta, William N.

2013-01-01

Cyclic nucleotide-regulated ion channels bind second messengers like cAMP to a C-terminal domain, consisting of a β-roll, followed by two α-helices (B- and C-helices). We monitored the cAMP-dependent changes in the structure of the C-helix of a C-terminal fragment of HCN2 channels using transition metal ion FRET between fluorophores on the C-helix and metal ions bound between histidine pairs on the same helix. cAMP induced a change in the dimensions of the C-helix and an increase in the metal binding affinity of the histidine pair. cAMP also caused an increase in the distance between a fluorophore on the C-helix and metal ions bound to the B-helix. Stabilizing the C-helix of intact CNGA1 channels by metal binding to a pair of histidines promoted channel opening. These data suggest that ordering of the C-helix is part of the gating conformational change in cyclic nucleotide-regulated channels. PMID:23525108

Ion Permeation and Mechanotransduction Mechanisms of Mechanosensitive Piezo Channels.

PubMed

Zhao, Qiancheng; Wu, Kun; Geng, Jie; Chi, Shaopeng; Wang, Yanfeng; Zhi, Peng; Zhang, Mingmin; Xiao, Bailong

2016-03-16

Piezo proteins have been proposed as the long-sought-after mechanosensitive cation channels in mammals that play critical roles in various mechanotransduction processes. However, the molecular bases that underlie their ion permeation and mechanotransduction have remained functionally undefined. Here we report our finding of the miniature pore-forming module of Piezo1 that resembles the pore architecture of other trimeric channels and encodes the essential pore properties. We further identified specific residues within the pore module that determine unitary conductance, pore blockage and ion selectivity for divalent and monovalent cations and anions. The non-pore-containing region of Piezo1 confers mechanosensitivity to mechano-insensitive trimeric acid-sensing ion channels, demonstrating that Piezo1 channels possess intrinsic mechanotransduction modules separate from their pore modules. In conclusion, this is the first report on the bona fide pore module and mechanotransduction components of Piezo channels, which define their ion-conducting properties and gating by mechanical stimuli, respectively. Copyright © 2016 Elsevier Inc. All rights reserved.

Functional Adaptation of the N-Methyl-d-aspartate Receptor to Inhibition by Ethanol Is Modulated by Striatal-Enriched Protein Tyrosine Phosphatase and p38 Mitogen-Activated Protein Kinase

PubMed Central

Coultrap, Steven J.; Browning, Michael D.; Proctor, William R.

2011-01-01

The hippocampal N-methyl-d-aspartate receptor (NMDAR) activity plays important roles in cognition and is a major substrate for ethanol-induced memory dysfunction. This receptor is a glutamate-gated ion channel, which is composed of NR1 and NR2 subunits in various brain areas. Although homomeric NR1 subunits form an active ion channel that conducts Na+ and Ca2+ currents, the incorporation of NR2 subunits allows this channel to be modulated by the Src family of kinases, phosphatases, and by simple molecules such as ethanol. We have found that short-term ethanol application inhibits the NMDAR activity via striatal enriched protein tyrosine phosphatase (STEP)-regulated mechanisms. The genetic deletion of the active form of STEP, STEP61, leads to marked attenuation of ethanol inhibition of NMDAR currents. In addition, STEP61 negatively regulates Fyn and p38 mitogen-activated protein kinase (MAPK), and these proteins are members of the NMDAR super molecular complex. Here we demonstrate, using whole-cell electrophysiological recording, Western blot analysis, and pharmacological manipulations, that neurons exposed to a 3-h, 45 mM ethanol treatment develop an adaptive attenuation of short-term ethanol inhibition of NMDAR currents in brain slices. Our results suggest that this adaptation of NMDAR responses is associated with a partial inactivation of STEP61, an activation of p38 MAPK, and a requirement for NR2B activity. Together, these data indicate that altered STEP61 and p38 MAPK signaling contribute to the modulation of ethanol inhibition of NMDARs in brain neurons. PMID:21680777

Harnessing the Flow of Excitation: TRP, Voltage-Gated Na(+), and Voltage-Gated Ca(2+) Channels in Contemporary Medicine.

PubMed

Frolov, Roman V; Weckström, Matti

2016-01-01

Cellular signaling in both excitable and nonexcitable cells involves several classes of ion channels. Some of them are of minor importance, with very specialized roles in physiology, but here we concentrate on three major channel classes: TRP (transient receptor potential channels), voltage-gated sodium channels (Nav), and voltage-gated calcium channels (Cav). Here, we first propose a conceptual framework binding together all three classes of ion channels, a "flow-of-excitation model" that takes into account the inputs mediated by TRP and other similar channels, the outputs invariably provided by Cav channels, and the regenerative transmission of signals in the neural networks, for which Nav channels are responsible. We use this framework to examine the function, structure, and pharmacology of these channel classes both at cellular and also at whole-body physiological level. Building on that basis we go through the pathologies arising from the direct or indirect malfunction of the channels, utilizing ion channel defects, the channelopathies. The pharmacological interventions affecting these channels are numerous. Part of those are well-established treatments, like treatment of hypertension or some forms of epilepsy, but many other are deeply problematic due to poor drug specificity, ion channel diversity, and widespread expression of the channels in tissues other than those actually targeted. © 2016 Elsevier Inc. All rights reserved.

Experiments with crystal deflectors for high energy ion beams: Electromagnetic dissociation probability for well channeled ions

NASA Astrophysics Data System (ADS)

Scandale, W.; Taratin, A. M.; Kovalenko, A. D.

2013-01-01

The paper presents the current status with the use of the crystal defectors for high energy ion beams. The channeling properties of multicharged ions are discussed. The results of the experiments on the deflection and extraction (collimation) of high energy ion beams with bent crystals performed in the accelerator centers are shortly considered. The analysis of the recent collimation experiment with a Pb nuclei of 270GeV/c per charge at the CERN Super Proton Synchrotron showed that the channeling efficiency was as large as about 90%. For Pb ions of the LHC energies a new mechanism, which can reduce the channeling efficiency, appears. The electromagnetic dissociation (ED) becomes possible for well channeled particles. However, the estimations performed in the paper show that the ED probability is small and should not visibly reduce the collimation efficiency. On the other hand, the aligned crystal gives the possibility to study the ED processes of heavy nuclei in the conditions when nuclear interactions are fully suppressed.

Ion channel signaling influences cellular proliferation and phagocyte activity during axolotl tail regeneration.

PubMed

Franklin, Brandon M; Voss, S Randal; Osborn, Jeffrey L

2017-08-01

Little is known about the potential for ion channels to regulate cellular behaviors during tissue regeneration. Here, we utilized an amphibian tail regeneration assay coupled with a chemical genetic screen to identify ion channel antagonists that altered critical cellular processes during regeneration. Inhibition of multiple ion channels either partially (anoctamin1/Tmem16a, anoctamin2/Tmem16b, K V 2.1, K V 2.2, L-type Ca V channels and H/K ATPases) or completely (GlyR, GABA A R, K V 1.5 and SERCA pumps) inhibited tail regeneration. Partial inhibition of tail regeneration by blocking the calcium activated chloride channels, anoctamin1&2, was associated with a reduction of cellular proliferation in tail muscle and mesenchymal regions. Inhibition of anoctamin 1/2 also altered the post-amputation transcriptional response of p44/42 MAPK signaling pathway genes, including decreased expression of erk1/erk2. We also found that complete inhibition via voltage gated K + channel blockade was associated with diminished phagocyte recruitment to the amputation site. The identification of H + pumps as required for axolotl tail regeneration supports findings in Xenopus and Planaria models, and more generally, the conservation of ion channels as regulators of tissue regeneration. This study provides a preliminary framework for an in-depth investigation of the mechanistic role of ion channels and their potential involvement in regulating cellular proliferation and other processes essential to wound healing, appendage regeneration, and tissue repair. Copyright © 2017 Elsevier B.V. All rights reserved.

Three homologous subunits form a high affinity peptide-gated ion channel in Hydra.

PubMed

Dürrnagel, Stefan; Kuhn, Anne; Tsiairis, Charisios D; Williamson, Michael; Kalbacher, Hubert; Grimmelikhuijzen, Cornelis J P; Holstein, Thomas W; Gründer, Stefan

2010-04-16

Recently, three ion channel subunits of the degenerin (DEG)/epithelial Na(+) channel (ENaC) gene family have been cloned from the freshwater polyp Hydra magnipapillata, the Hydra Na(+) channels (HyNaCs) 2-4. Two of them, HyNaC2 and HyNaC3, co-assemble to form an ion channel that is gated by the neuropeptides Hydra-RFamides I and II. The HyNaC2/3 channel is so far the only cloned ionotropic receptor from cnidarians and, together with the related ionotropic receptor FMRFamide-activated Na(+) channel (FaNaC) from snails, the only known peptide-gated ionotropic receptor. The HyNaC2/3 channel has pore properties, like a low Na(+) selectivity and a low amiloride affinity, that are different from other channels of the DEG/ENaC gene family, suggesting that a component of the native Hydra channel might still be lacking. Here, we report the cloning of a new ion channel subunit from Hydra, HyNaC5. The new subunit is closely related to HyNaC2 and -3 and co-localizes with HyNaC2 and -3 to the base of the tentacles. Coexpression in Xenopus oocytes of HyNaC5 with HyNaC2 and -3 largely increases current amplitude after peptide stimulation and affinity of the channel to Hydra-RFamides I and II. Moreover, the HyNaC2/3/5 channel has altered pore properties and amiloride affinity, more similarly to other DEG/ENaC channels. Collectively, our results suggest that the three homologous subunits HyNaC2, -3, and -5 form a peptide-gated ion channel in Hydra that could contribute to fast synaptic transmission.

Trails of Kilovolt Ions Created by Subsurface Channeling

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

Redinger, Alex; Standop, Sebastian; Michely, Thomas

2010-02-19

Using scanning tunneling microscopy, we observe the damage trails produced by keV noble-gas ions incident at glancing angles onto Pt(111). Surface vacancies and adatoms aligned along the ion trajectory constitute the ion trails. Atomistic simulations reveal that these straight trails are produced by nuclear (elastic) collisions with surface layer atoms during subsurface channeling of the projectiles. In a small energy window around 5 keV, Xe{sup +} ions create vacancy grooves that mark the ion trajectory with atomic precision. The asymmetry of the adatom production on the two sides of the projectile path is traced back to the asymmetry of themore » ion's subsurface channel.« less

Multiple-channel detection of cellular activities by ion-sensitive transistors

NASA Astrophysics Data System (ADS)

Machida, Satoru; Shimada, Hideto; Motoyama, Yumi

2018-04-01

An ion-sensitive field-effect transistor to record cellular activities was demonstrated. This field-effect transistor (bio transistor) includes cultured cells on the gate insulator instead of gate electrode. The bio transistor converts a change in potential underneath the cells into variation of the drain current when ion channels open. The bio transistor has high detection sensitivity to even minute variations in potential utilizing a subthreshold swing region. To open ion channels, a reagent solution (acetylcholine) was added to a human-originating cell cultured on the bio transistor. The drain current was successfully decreased with the addition of acetylcholine. Moreover, we attempted to detect the opening of ion channels using a multiple-channel measurement circuit containing several bio transistors. As a consequence, the drain current distinctly decreased only after the addition of acetylcholine. We confirmed that this measurement system including bio transistors enables to observation of cellular activities sensitively and simultaneously.

Fundamental properties of local anesthetics: half-maximal blocking concentrations for tonic block of Na+ and K+ channels in peripheral nerve.

PubMed

Bräu, M E; Vogel, W; Hempelmann, G

1998-10-01

Local anesthetics suppress excitability by interfering with ion channel function. Ensheathment of peripheral nerve fibers, however, impedes diffusion of drugs to the ion channels and may influence the evaluation of local anesthetic potencies. Investigating ion channels in excised membrane patches avoids these diffusion barriers. We investigated the effect of local anesthetics with voltage-dependent Na+ and K+ channels in enzymatically dissociated sciatic nerve fibers of Xenopus laevis using the patch clamp method. The outside-out configuration was chosen to apply drugs to the external face of the membrane. Local anesthetics reversibly blocked the transient Na+ inward current, as well as the steady-state K+ outward current. Half-maximal tonic inhibiting concentrations (IC50), as obtained from concentration-effect curves for Na+ current block were: tetracaine 0.7 microM, etidocaine 18 microM, bupivacaine 27 microM, procaine 60 microM, mepivacaine 149 microM, and lidocaine 204 microM. The values for voltage-dependent K+ current block were: bupivacaine 92 microM, etidocaine 176 microM, tetracaine 946 microM, lidocaine 1118 microM, mepivacaine 2305 microM, and procaine 6302 microM. Correlation of potencies with octanol:buffer partition coefficients (logP0) revealed that ester-bound local anesthetics were more potent in blocking Na+ channels than amide drugs. Within these groups, lipophilicity governed local anesthetic potency. We conclude that local anesthetic action on peripheral nerve ion channels is mediated via lipophilic drug-channel interactions. Half-maximal blocking concentrations of commonly used local anesthetics for Na+ and K+ channel block were determined on small membrane patches of peripheral nerve fibers. Because drugs can directly diffuse to the ion channel in this model, these data result from direct interactions of the drugs with ion channels.

Sub-cellular distribution and translocation of TRP channels.

PubMed

Toro, Carlos A; Arias, Luis A; Brauchi, Sebastian

2011-01-01

Cellular electrical activity is the result of a highly complex processes that involve the activation of ion channel proteins. Ion channels make pores on cell membranes that rapidly transit between conductive and non-conductive states, allowing different ions to flow down their electrochemical gradients across cell membranes. In the case of neuronal cells, ion channel activity orchestrates action potentials traveling through axons, enabling electrical communication between cells in distant parts of the body. Somatic sensation -our ability to feel touch, temperature and noxious stimuli- require ion channels able to sense and respond to our peripheral environment. Sensory integration involves the summing of various environmental cues and their conversion into electrical signals. Members of the Transient Receptor Potential (TRP) family of ion channels have emerged as important mediators of both cellular sensing and sensory integration. The regulation of the spatial and temporal distribution of membrane receptors is recognized as an important mechanism for controlling the magnitude of the cellular response and the time scale on which cellular signaling occurs. Several studies have shown that this mechanism is also used by TRP channels to modulate cellular response and ultimately fulfill their physiological function as sensors. However, the inner-working of this mode of control for TRP channels remains poorly understood. The question of whether TRPs intrinsically regulate their own vesicular trafficking or weather the dynamic regulation of TRP channel residence on the cell surface is caused by extrinsic changes in the rates of vesicle insertion or retrieval remain open. This review will examine the evidence that sub-cellular redistribution of TRP channels plays an important role in regulating their activity and explore the mechanisms that control the trafficking of vesicles containing TRP channels.

Cellular defibrillation: interaction of micro-scale electric fields with voltage-gated ion channels.

PubMed

Kargol, Armin; Malkinski, Leszek; Eskandari, Rahmatollah; Carter, Maya; Livingston, Daniel

2015-09-01

We study the effect of micro-scale electric fields on voltage-gated ion channels in mammalian cell membranes. Such micro- and nano-scale electric fields mimic the effects of multiferroic nanoparticles that were recently proposed [1] as a novel way of controlling the function of voltage-sensing biomolecules such as ion channels. This article describes experimental procedures and initial results that reveal the effect of the electric field, in close proximity of cells, on the ion transport through voltage-gated ion channels. We present two configurations of the whole-cell patch-clamping apparatus that were used to detect the effect of external stimulation on ionic currents and discuss preliminary results that indicate modulation of the ionic currents consistent with the applied stimulus.

Distinct regions that control ion selectivity and calcium-dependent activation in the bestrophin ion channel.

PubMed

Vaisey, George; Miller, Alexandria N; Long, Stephen B

2016-11-22

Cytoplasmic calcium (Ca 2+ ) activates the bestrophin anion channel, allowing chloride ions to flow down their electrochemical gradient. Mutations in bestrophin 1 (BEST1) cause macular degenerative disorders. Previously, we determined an X-ray structure of chicken BEST1 that revealed the architecture of the channel. Here, we present electrophysiological studies of purified wild-type and mutant BEST1 channels and an X-ray structure of a Ca 2+ -independent mutant. From these experiments, we identify regions of BEST1 responsible for Ca 2+ activation and ion selectivity. A "Ca 2+ clasp" within the channel's intracellular region acts as a sensor of cytoplasmic Ca 2+ . Alanine substitutions within a hydrophobic "neck" of the pore, which widen it, cause the channel to be constitutively active, irrespective of Ca 2+ . We conclude that the primary function of the neck is as a "gate" that controls chloride permeation in a Ca 2+ -dependent manner. In contrast to what others have proposed, we find that the neck is not a major contributor to the channel's ion selectivity. We find that mutation of a cytosolic "aperture" of the pore does not perturb the Ca 2+ dependence of the channel or its preference for anions over cations, but its mutation dramatically alters relative permeabilities among anions. The data suggest that the aperture functions as a size-selective filter that permits the passage of small entities such as partially dehydrated chloride ions while excluding larger molecules such as amino acids. Thus, unlike ion channels that have a single "selectivity filter," in bestrophin, distinct regions of the pore govern anion-vs.-cation selectivity and the relative permeabilities among anions.

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

Ngo, Van; Wang, Yibo; Haas, Stephan

Crystal structures of several bacterial Na v channels have been recently published and molecular dynamics simulations of ion permeation through these channels are consistent with many electrophysiological properties of eukaryotic channels. Bacterial Na v channels have been characterized as functionally asymmetric, and the mechanism of this asymmetry has not been clearly understood. To address this question, we combined non-equilibrium simulation data with two-dimensional equilibrium unperturbed landscapes generated by umbrella sampling and Weighted Histogram Analysis Methods for multiple ions traversing the selectivity filter of bacterial Na vAb channel. This approach provided new insight into the mechanism of selective ion permeation inmore » bacterial Nav channels. The non-equilibrium simulations indicate that two or three extracellular K + ions can block the entrance to the selectivity filter of Na vAb in the presence of applied forces in the inward direction, but not in the outward direction. The block state occurs in an unstable local minimum of the equilibrium unperturbed free-energy landscape of two K+ ions that can be ‘locked’ in place bymodest applied forces. In contrast to K +, three Na + ions move favorably through the selectivity filter together as a unit in a loose “knock-on” mechanism of permeation in both inward and outward directions, and there is no similar local minimum in the two-dimensional free-energy landscape of two Na + ions for a block state. The useful work predicted by the non-equilibrium simulations that is required to break the K + block is equivalent to large applied potentials experimentally measured for two bacterial Na v channels to induce inward currents of K + ions. Here, these results illustrate how inclusion of non-equilibrium factors in the simulations can provide detailed information about mechanisms of ion selectivity that is missing from mechanisms derived from either crystal structures or equilibrium unperturbed free-energy landscapes.« less

Combining molecular dynamics and an electrodiffusion model to calculate ion channel conductance

NASA Astrophysics Data System (ADS)

Wilson, Michael A.; Nguyen, Thuy Hien; Pohorille, Andrew

2014-12-01

Establishing the relation between the structures and functions of protein ion channels, which are protein assemblies that facilitate transmembrane ion transport through water-filled pores, is at the forefront of biological and medical sciences. A reliable way to determine whether our understanding of this relation is satisfactory is to reproduce the measured ionic conductance over a broad range of applied voltages. This can be done in molecular dynamics simulations by way of applying an external electric field to the system and counting the number of ions that traverse the channel per unit time. Since this approach is computationally very expensive we develop a markedly more efficient alternative in which molecular dynamics is combined with an electrodiffusion equation. This alternative approach applies if steady-state ion transport through channels can be described with sufficient accuracy by the one-dimensional diffusion equation in the potential given by the free energy profile and applied voltage. The theory refers only to line densities of ions in the channel and, therefore, avoids ambiguities related to determining the surface area of the channel near its endpoints or other procedures connecting the line and bulk ion densities. We apply the theory to a simple, model system based on the trichotoxin channel. We test the assumptions of the electrodiffusion equation, and determine the precision and consistency of the calculated conductance. We demonstrate that it is possible to calculate current/voltage dependence and accurately reconstruct the underlying (equilibrium) free energy profile, all from molecular dynamics simulations at a single voltage. The approach developed here applies to other channels that satisfy the conditions of the electrodiffusion equation.

Human Digital Meissner Corpuscles Display Immunoreactivity for the Multifunctional Ion Channels Trpc6 and Trpv4.

PubMed

Alonso-González, Paula; Cabo, Roberto; San José, Isabel; Gago, Angel; Suazo, Iván C; García-Suárez, Olivia; Cobo, Juan; Vega, José A

2017-06-01

Ion channels are at the basis of the sensory processes including mechanosensing. Some members of the transient receptor potential (TRP) ion channel superfamily have been proposed as mechanosensors, but their putative role in mechanotransduction is controversial. Among them there are TRP canonical 6 (TRPC6) and TRP vanilloid 4 (TRPV4) ion channels, which are known to cooperate in mechanical hyperalgesia. Here, we investigated the occurrence, distribution, and possible colocalization of TRPC6 and TRPV4 in human digital Meissner sensory corpuscles using immunohistochemistry and double immunofluorescence (associate with markers for specific corpuscular constituents). TRPC6 immunoreactivity was restricted to the axon of Meissner corpuscles, whereas TRPV4 was detected in the axon but also in the lamellar cells. Moreover, axonal colocalization of TRPV4 and TRPC6 was found in the digital Meissner corpuscles. Present results demonstrate for the first time the occurrence and colocalization of two ion channels candidates to mechanosensors in human cutaneous mechanoreceptors. The functional significance of these ion channels in that place remains to be clarified, but should be related to different properties of mechanosensitivity. Anat Rec, 300:1022-1031, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Two barriers for sodium in vascular endothelium?

PubMed Central

Oberleithner, Hans

2012-01-01

Vascular endothelium plays a key role in blood pressure regulation. Recently, it has been shown that a 5% increase of plasma sodium concentration (sodium excess) stiffens endothelial cells by about 25%, leading to cellular dysfunction. Surface measurements demonstrated that the endothelial glycocalyx (eGC), an anionic biopolymer, deteriorates when sodium is elevated. In view of these results, a two-barrier model for sodium exiting the circulation across the endothelium is suggested. The first sodium barrier is the eGC which selectively buffers sodium ions with its negatively charged prote-oglycans.The second sodium barrier is the endothelial plasma membrane which contains sodium channels. Sodium excess, in the presence of aldosterone, leads to eGC break-down and, in parallel, to an up-regulation of plasma membrane sodium channels. The following hypothesis is postulated: Sodium excess increases vascular sodium permeability. Under such con-ditions (e.g. high-sodium diet), day-by-day ingested sodium, instead of being readily buffered by the eGC and then rapidly excreted by the kidneys, is distributed in the whole body before being finally excreted. Gradually, the sodium overload damages the organism. PMID:22471931

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.

In silico assessment of the conduction mechanism of the Ryanodine Receptor 1 reveals previously unknown exit pathways.

PubMed

Heinz, Leonard P; Kopec, Wojciech; de Groot, Bert L; Fink, Rainer H A

2018-05-02

The ryanodine receptor 1 is a large calcium ion channel found in mammalian skeletal muscle. The ion channel gained a lot of attention recently, after multiple independent authors published near-atomic cryo electron microscopy data. Taking advantage of the unprecedented quality of structural data, we performed molecular dynamics simulations on the entire ion channel as well as on a reduced model. We calculated potentials of mean force for Ba 2+ , Ca 2+ , Mg 2+ , K + , Na + and Cl - ions using umbrella sampling to identify the key residues involved in ion permeation. We found two main binding sites for the cations, whereas the channel is strongly repulsive for chloride ions. Furthermore, the data is consistent with the model that the receptor achieves its ion selectivity by over-affinity for divalent cations in a calcium-block-like fashion. We reproduced the experimental conductance for potassium ions in permeation simulations with applied voltage. The analysis of the permeation paths shows that ions exit the pore via multiple pathways, which we suggest to be related to the experimental observation of different subconducting states.

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.

Intravesical TRPV4 blockade reduces repeated variate stress-induced bladder dysfunction by increasing bladder capacity and decreasing voiding frequency in male rats

PubMed Central

Merrill, Liana

2014-01-01

Individuals with functional lower urinary tract disorders including interstitial cystitis (IC)/bladder pain syndrome (BPS) and overactive bladder (OAB) often report symptom (e.g., urinary frequency) worsening due to stress. One member of the transient receptor potential ion channel vanilloid family, TRPV4, has recently been implicated in urinary bladder dysfunction disorders including OAB and IC/BPS. These studies address the role of TRPV4 in stress-induced bladder dysfunction using an animal model of stress in male rats. To induce stress, rats were exposed to 7 days of repeated variate stress (RVS). Quantitative PCR data demonstrated significant (P ≤ 0.01) increases in TRPV4 transcript levels in urothelium but not detrusor smooth muscle. Western blot analyses of split urinary bladders (i.e., urothelium and detrusor) showed significant (P ≤ 0.01) increases in TRPV4 protein expression levels in urothelial tissues but not detrusor smooth muscle. We previously showed that RVS produces bladder dysfunction characterized by decreased bladder capacity and increased voiding frequency. The functional role of TRPV4 in RVS-induced bladder dysfunction was evaluated using continuous, open outlet intravesical infusion of saline in conjunction with administration of a TRPV4 agonist, GSK1016790A (3 μM), a TRPV4 antagonist, HC067047 (1 μM), or vehicle (0.1% DMSO in saline) in control and RVS-treated rats. Bladder capacity, void volume, and intercontraction interval significantly decreased following intravesical instillation of GSK1016790A in control rats and significantly (P ≤ 0.01) increased following administration of HC067047 in RVS-treated rats. These results demonstrate increased TRPV4 expression in the urothelium following RVS and that TRPV4 blockade ameliorates RVS-induced bladder dysfunction consistent with the role of TRPV4 as a promising target for bladder function disorders. PMID:24965792

Potential Roles of Amiloride-Sensitive Sodium Channels in Cancer Development.

PubMed

Xu, Siguang; Liu, Cui; Ma, Yana; Ji, Hong-Long; Li, Xiumin

2016-01-01

The ENaC/degenerin ion channel superfamily includes the amiloride-sensitive epithelial sodium channel (ENaC) and acid sensitive ionic channel (ASIC). ENaC is a multimeric ion channel formed by heteromultimeric membrane glycoproteins, which participate in a multitude of biological processes by mediating the transport of sodium (Na(+)) across epithelial tissues such as the kidney, lungs, bladder, and gut. Aberrant ENaC functions contribute to several human disease states including pseudohypoaldosteronism, Liddle syndrome, cystic fibrosis, and salt-sensitive hypertension. Increasing evidence suggests that ion channels not only regulate ion homeostasis and electric signaling in excitable cells but also play important roles in cancer cell behaviors such as proliferation, apoptosis, invasion, and migration. Indeed, ENaCs/ASICs had been reported to be associated with cancer characteristics. Given their cell surface localization and pharmacology, pharmacological strategies to target ENaC/ASIC family members may be promising cancer therapeutics.

Thin membrane sensor with biochemical switch

NASA Technical Reports Server (NTRS)

Worley, III, Jennings F. (Inventor); Case, George D. (Inventor)

1994-01-01

A modular biosensor system for chemical or biological agent detection utilizes electrochemical measurement of an ion current across a gate membrane triggered by the reaction of the target agent with a recognition protein conjugated to a channel blocker. The sensor system includes a bioresponse simulator or biochemical switch module which contains the recognition protein-channel blocker conjugate, and in which the detection reactions occur, and a transducer module which contains a gate membrane and a measuring electrode, and in which the presence of agent is sensed electrically. In the poised state, ion channels in the gate membrane are blocked by the recognition protein-channel blocker conjugate. Detection reactions remove the recognition protein-channel blocker conjugate from the ion channels, thus eliciting an ion current surge in the gate membrane which subsequently triggers an output alarm. Sufficiently large currents are generated that simple direct current electronics are adequate for the measurements. The biosensor has applications for environmental, medical, and industrial use.

K + block is the mechanism of functional asymmetry in bacterial Na v channels

DOE PAGES

Ngo, Van; Wang, Yibo; Haas, Stephan; ...

2016-01-04

Crystal structures of several bacterial Na v channels have been recently published and molecular dynamics simulations of ion permeation through these channels are consistent with many electrophysiological properties of eukaryotic channels. Bacterial Na v channels have been characterized as functionally asymmetric, and the mechanism of this asymmetry has not been clearly understood. To address this question, we combined non-equilibrium simulation data with two-dimensional equilibrium unperturbed landscapes generated by umbrella sampling and Weighted Histogram Analysis Methods for multiple ions traversing the selectivity filter of bacterial Na vAb channel. This approach provided new insight into the mechanism of selective ion permeation inmore » bacterial Nav channels. The non-equilibrium simulations indicate that two or three extracellular K + ions can block the entrance to the selectivity filter of Na vAb in the presence of applied forces in the inward direction, but not in the outward direction. The block state occurs in an unstable local minimum of the equilibrium unperturbed free-energy landscape of two K+ ions that can be ‘locked’ in place bymodest applied forces. In contrast to K +, three Na + ions move favorably through the selectivity filter together as a unit in a loose “knock-on” mechanism of permeation in both inward and outward directions, and there is no similar local minimum in the two-dimensional free-energy landscape of two Na + ions for a block state. The useful work predicted by the non-equilibrium simulations that is required to break the K + block is equivalent to large applied potentials experimentally measured for two bacterial Na v channels to induce inward currents of K + ions. Here, these results illustrate how inclusion of non-equilibrium factors in the simulations can provide detailed information about mechanisms of ion selectivity that is missing from mechanisms derived from either crystal structures or equilibrium unperturbed free-energy landscapes.« less

K+ Block Is the Mechanism of Functional Asymmetry in Bacterial Nav Channels

PubMed Central

Ngo, Van; Wang, Yibo; Haas, Stephan; Noskov, Sergei Y.; Farley, Robert A.

2016-01-01

Crystal structures of several bacterial Nav channels have been recently published and molecular dynamics simulations of ion permeation through these channels are consistent with many electrophysiological properties of eukaryotic channels. Bacterial Nav channels have been characterized as functionally asymmetric, and the mechanism of this asymmetry has not been clearly understood. To address this question, we combined non-equilibrium simulation data with two-dimensional equilibrium unperturbed landscapes generated by umbrella sampling and Weighted Histogram Analysis Methods for multiple ions traversing the selectivity filter of bacterial NavAb channel. This approach provided new insight into the mechanism of selective ion permeation in bacterial Nav channels. The non-equilibrium simulations indicate that two or three extracellular K+ ions can block the entrance to the selectivity filter of NavAb in the presence of applied forces in the inward direction, but not in the outward direction. The block state occurs in an unstable local minimum of the equilibrium unperturbed free-energy landscape of two K+ ions that can be ‘locked’ in place by modest applied forces. In contrast to K+, three Na+ ions move favorably through the selectivity filter together as a unit in a loose “knock-on” mechanism of permeation in both inward and outward directions, and there is no similar local minimum in the two-dimensional free-energy landscape of two Na+ ions for a block state. The useful work predicted by the non-equilibrium simulations that is required to break the K+ block is equivalent to large applied potentials experimentally measured for two bacterial Nav channels to induce inward currents of K+ ions. These results illustrate how inclusion of non-equilibrium factors in the simulations can provide detailed information about mechanisms of ion selectivity that is missing from mechanisms derived from either crystal structures or equilibrium unperturbed free-energy landscapes. PMID:26727271

Molecular dynamics study of ion transport through an open model of voltage-gated sodium channel.

PubMed

Li, Yang; Sun, Ruining; Liu, Huihui; Gong, Haipeng

2017-05-01

Voltage-gated sodium (Na V ) channels are critical in the signal transduction of excitable cells. In this work, we modeled the open conformation for the pore domain of a prokaryotic Na V channel (Na V Rh), and used molecular dynamics simulations to track the translocation of dozens of Na + ions through the channel in the presence of a physiological transmembrane ion concentration gradient and a transmembrane electrical field that was closer to the physiological one than previous studies. Channel conductance was then estimated from simulations on the wide-type and DEKA mutant of Na V Rh. Interestingly, the conductivity predicted from the DEKA mutant agrees well with experimental measurement on eukaryotic Na V 1.4 channel. Moreover, the wide-type and DEKA mutant of Na V Rh exhibited markedly distinct ion permeation patterns, which thus implies the mechanistic difference between prokaryotic and eukaryotic Na V channels. Copyright © 2017 Elsevier B.V. All rights reserved.

Permeation Mechanisms in the TMEM16B Calcium-Activated Chloride Channels

PubMed Central

2017-01-01

TMEM16A and TMEM16B encode for Ca2+-activated Cl− channels (CaCC) and are expressed in many cell types and play a relevant role in many physiological processes. Here, I performed a site-directed mutagenesis study to understand the molecular mechanisms of ion permeation of TMEM16B. I mutated two positive charged residues R573 and K540, respectively located at the entrance and inside the putative channel pore and I measured the properties of wild-type and mutant TMEM16B channels expressed in HEK-293 cells using whole-cell and excised inside-out patch clamp experiments. I found evidence that R573 and K540 control the ion permeability of TMEM16B depending both on which side of the membrane the ion substitution occurs and on the level of channel activation. Moreover, these residues contribute to control blockage or activation by permeant anions. Finally, R573 mutation abolishes the anomalous mole fraction effect observed in the presence of a permeable anion and it alters the apparent Ca2+-sensitivity of the channel. These findings indicate that residues facing the putative channel pore are responsible both for controlling the ion selectivity and the gating of the channel, providing an initial understanding of molecular mechanism of ion permeation in TMEM16B. PMID:28046119

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.

On the estimation of cooperativity in ion channel kinetics: activation free energy and kinetic mechanism of Shaker K+ channel.

PubMed

Banerjee, Kinshuk; Das, Biswajit; Gangopadhyay, Gautam

2013-04-28

In this paper, we have explored generic criteria of cooperative behavior in ion channel kinetics treating it on the same footing with multistate receptor-ligand binding in a compact theoretical framework. We have shown that the characterization of cooperativity of ion channels in terms of the Hill coefficient violates the standard Hill criteria defined for allosteric cooperativity of ligand binding. To resolve the issue, an alternative measure of cooperativity is proposed here in terms of the cooperativity index that sets a unified criteria for both the systems. More importantly, for ion channel this index can be very useful to describe the cooperative kinetics as it can be readily determined from the experimentally measured ionic current combined with theoretical modelling. We have analyzed the correlation between the voltage value and slope of the voltage-activation curve at the half-activation point and consequently determined the standard free energy of activation of the ion channel using two well-established mechanisms of cooperativity, namely, Koshland-Nemethy-Filmer (KNF) and Monod-Wyman-Changeux (MWC) models. Comparison of the theoretical results for both the models with appropriate experimental data of mutational perturbation of Shaker K(+) channel supports the experimental fact that the KNF model is more suitable to describe the cooperative behavior of this class of ion channels, whereas the performance of the MWC model is unsatisfactory. We have also estimated the mechanistic performance through standard free energy of channel activation for both the models and proposed a possible functional disadvantage in the MWC scheme.

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.

Ion Transport in Confined Geometries below the Nanoscale: Access Resistance Dominates Protein Channel Conductance in Diluted Solutions.

PubMed

Alcaraz, Antonio; López, M Lidón; Queralt-Martín, María; Aguilella, Vicente M

2017-10-24

Synthetic nanopores and mesoscopic protein channels have common traits like the importance of electrostatic interactions between the permeating ions and the nanochannel. Ion transport at the nanoscale occurs under confinement conditions so that the usual assumptions made in microfluidics are challenged, among others, by interfacial effects such as access resistance (AR). Here, we show that a sound interpretation of electrophysiological measurements in terms of channel ion selective properties requires the consideration of interfacial effects, up to the point that they dominate protein channel conductance in diluted solutions. We measure AR in a large ion channel, the bacterial porin OmpF, by means of single-channel conductance measurements in electrolyte solutions containing varying concentrations of high molecular weight PEG, sterically excluded from the pore. Comparison of experiments performed in charged and neutral planar membranes shows that lipid surface charges modify the ion distribution and determine the value of AR, indicating that lipid molecules are more than passive scaffolds even in the case of large transmembrane proteins. We also found that AR may reach up to 80% of the total channel conductance in diluted solutions, where electrophysiological recordings register essentially the AR of the system and depend marginally on the pore characteristics. These findings may have implications for several low aspect ratio biological channels that perform their physiological function in a low ionic strength and macromolecule crowded environment, just the two conditions enhancing the AR contribution.

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

Ion channels in artificial bolaamphiphile membranes deposited on sensor chips: optical detection in an ion-channel-based biosensor

NASA Astrophysics Data System (ADS)

Schalkhammer, Thomas G. M.; Weiss-Wichert, Christof; Smetazko, Michaela M.; Valina-Saba, Miriam

1997-06-01

Signal amplification using labels should be replaced by a technique monitoring the biochemical binding event directly. The use of a ligand coupled to an artificial gated membrane ion channel is a new promising strategy. Binding of protein- or DNA/RNA-analytes at ligand modified peptide channels results in an on/off-response of the channel current due to channel closure or distortion. The sensor consists of stable transmembrane channels with a ligand bound covalently at the peptide channel entrance, a sensor chip with a photostructurized hydrophobic polymer frame, a hydrophilic ion conducting membrane support, a lipid membrane incorporating the engineered ion channels, and a current amplifier or a sensitive fluorescence monitor. Detection of channel opening or closure can ether be obtained by directly monitoring membrane conductivity or a transient change of pH or ion concentration within the membrane compartment. This change can be induced by electrochemical or optical means and its decay is directly correlated to the permeability of the membrane. The ion concentration in the sub membrane compartment was monitored by incorporation of fluorescent indicator dyes. To obtain the stable sensor membrane the lipid layer had to be attached on a support and the floating of the second lipid membrane on top of the first one had to be prevented. Both problems do not occur using our new circular C44-C76 bolaamphiphilic lipids consisting of a long hydrophobic core region and two hydrophilic heads. Use of maleic ester-head groups enabled us to easily modify the lipids with amines, thioles, alcohols, phosphates, boronic acid as well as fluorescent dyes. The properties of these membranes were studied using LB and fluorescence techniques. Based on this detection principle miniaturized sensor chips with significantly enhanced sensitivity and large multi analyte arrays are under construction.

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

Koenig, Xaver; Kovar, Michael; Rubi, Lena

The plant alkaloid ibogaine has promising anti-addictive properties. Albeit not licenced as a therapeutic drug, and despite hints that ibogaine may perturb the heart rhythm, this alkaloid is used to treat drug addicts. We have recently reported that ibogaine inhibits human ERG (hERG) potassium channels at concentrations similar to the drugs affinity for several of its known brain targets. Thereby the drug may disturb the heart's electrophysiology. Here, to assess the drug's cardiac ion channel profile in more detail, we studied the effects of ibogaine and its congener 18-Methoxycoronaridine (18-MC) on various cardiac voltage-gated ion channels. We confirmed that heterologouslymore » expressed hERG currents are reduced by ibogaine in low micromolar concentrations. Moreover, at higher concentrations, the drug also reduced human Na{sub v}1.5 sodium and Ca{sub v}1.2 calcium currents. Ion currents were as well reduced by 18-MC, yet with diminished potency. Unexpectedly, although blocking hERG channels, ibogaine did not prolong the action potential (AP) in guinea pig cardiomyocytes at low micromolar concentrations. Higher concentrations (≥ 10 μM) even shortened the AP. These findings can be explained by the drug's calcium channel inhibition, which counteracts the AP-prolonging effect generated by hERG blockade. Implementation of ibogaine's inhibitory effects on human ion channels in a computer model of a ventricular cardiomyocyte, on the other hand, suggested that ibogaine does prolong the AP in the human heart. We conclude that therapeutic concentrations of ibogaine have the propensity to prolong the QT interval of the electrocardiogram in humans. In some cases this may lead to cardiac arrhythmias. - Highlights: • We study effects of anti-addiction drug ibogaine on ionic currents in cardiomyocytes. • We assess the cardiac ion channel profile of ibogaine. • Ibogaine inhibits hERG potassium, sodium and calcium channels. • Ibogaine’s effects on ion channels are a potential source of cardiac arrhythmias. • 18-Methoxycoronaridine has a lower affinity for cardiac ion channels than ibogaine.« less

Ion Selectivity Mechanism in a Bacterial Pentameric Ligand-Gated Ion Channel

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

Fritsch, Sebastian M; Ivanov, Ivaylo N; Wang, Hailong

2011-01-01

The proton-gated ion channel from Gloeobacter violaceus (GLIC) is a prokaryotic homolog of the eukaryotic nicotinic acetylcholine receptor (nAChR) that responds to the binding of neurotransmitter acetylcholine and mediates fast signal transmission. Recent emergence of a high resolution crystal structure of GLIC captured in a potentially open state allowed detailed, atomic-level insight into ion conduction and selectivity mechanisms in these channels. Herein, we have examined the barriers to ion conduction and origins of ion selectivity in the GLIC channel by the construction of potential of mean force (PMF) profiles for sodium and chloride ions inside the transmembrane region. Our calculationsmore » reveal that the GLIC channel is open for a sodium ion to transport, but presents a ~10 kcal/mol free energy barrier for a chloride ion, which arises primarily from the unfavorable interactions with a ring of negatively charged glutamate residues (E-2 ) at the intracellular end and a ring of hydrophobic residues (I9 ) in the middle of the transmembrane domain. Our collective findings further suggest that the charge selection mechanism can, to a large extent, be attributed to the narrow intracellular end and a ring of glutamate residues in this position their strong negative electrostatics and ability to bind cations. By contrast, E19 at the extracellular entrance only plays a minor role in ion selectivity of GLIC. In addition to electrostatics, both ion hydration and protein dynamics are found to be crucial for ion conduction as well, which explains why a chloride ion experiences a much greater barrier than a sodium ion in the hydrophobic region of the pore.« less

A study of quantum mechanical probabilities in the classical Hodgkin-Huxley model.

PubMed

Moradi, N; Scholkmann, F; Salari, V

2015-03-01

The Hodgkin-Huxley (HH) model is a powerful model to explain different aspects of spike generation in excitable cells. However, the HH model was proposed in 1952 when the real structure of the ion channel was unknown. It is now common knowledge that in many ion-channel proteins the flow of ions through the pore is governed by a gate, comprising a so-called "selectivity filter" inside the ion channel, which can be controlled by electrical interactions. The selectivity filter (SF) is believed to be responsible for the selection and fast conduction of particular ions across the membrane of an excitable cell. Other (generally larger) parts of the molecule such as the pore-domain gate control the access of ions to the channel protein. In fact, two types of gates are considered here for ion channels: the "external gate", which is the voltage sensitive gate, and the "internal gate" which is the selectivity filter gate (SFG). Some quantum effects are expected in the SFG due to its small dimensions, which may play an important role in the operation of an ion channel. Here, we examine parameters in a generalized model of HH to see whether any parameter affects the spike generation. Our results indicate that the previously suggested semi-quantum-classical equation proposed by Bernroider and Summhammer (BS) agrees strongly with the HH equation under different conditions and may even provide a better explanation in some cases. We conclude that the BS model can refine the classical HH model substantially.

[RyR-bound FKBP12.6 and the modulation].

PubMed

Yano, M; Matsuzaki, M

2001-06-01

In the pathogenesis of cardiac dysfunction in heart failure, a decrease in the activity of the sarcoplasmic reticulum (SR) Ca(2+) -ATPase is believed to be a major determinant. Recently, a novel mechanism of cardiac dysfunction in heart failure has been reported on the basis of the following findings:1) PKA hyperphosphorylation of RyR causes a dissociation of FKBP12.6 from RyR, resulting in the abnormal single-channel properties (increased Ca(2+) sensitivity for activation and elevated channel activity associated with destabilization of RyR (Marx et al, Cell 101:365, 2000), 2) a prominent abnormal Ca(2+) leak occurs through RyR, following a partial loss of RyR-bound FKBP12.6 and the resultant conformational change in RyR (Yano M et al, Circulation 102:2131, 2000). This abnormal Ca(2+) leak might possibly cause Ca(2+) overload and consequent diastolic dysfunction, as well as systolic dysfunction.

Structure of conducting channel of lightning

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

Alanakyan, Yu. R.

2013-08-15

The spatial distribution of the plasma density in a lightning channel is studied theoretically. It is shown that the electric-field double layer is formed at the channel boundary. In this case, the electron temperature changes abruptly and ions are accelerated by the electric field of the double layer. The ion momentum flux density is close to the surrounding gas pressure. Cleaning of the channel from heavy particles occurs in particle-exchange processes between the plasma channel and the surrounding air. Hydrogen ions are accumulated inside the expanding channel from the surrounding air, which is enriched by hydrogen-contained molecules. In this case,more » the plasma channel is unstable and splits to a chain of equidistant bunches of plasma. The hydrogen-enrich bunches burn diffusely after recombination exhibiting the bead lightning behavior.« less

SIMULATION OF ION CONDUCTION IN α-HEMOLYSIN NANOPORES WITH COVALENTLY ATTACHED β-CYCLODEXTRIN BASED ON BOLTZMANN TRANSPORT MONTE CARLO MODEL

PubMed Central

Toghraee, Reza; Lee, Kyu-Il; Papke, David; Chiu, See-Wing; Jakobsson, Eric; Ravaioli, Umberto

2009-01-01

Ion channels, as natures’ solution to regulating biological environments, are particularly interesting to device engineers seeking to understand how natural molecular systems realize device-like functions, such as stochastic sensing of organic analytes. What’s more, attaching molecular adaptors in desired orientations inside genetically engineered ion channels, enhances the system functionality as a biosensor. In general, a hierarchy of simulation methodologies is needed to study different aspects of a biological system like ion channels. Biology Monte Carlo (BioMOCA), a three-dimensional coarse-grained particle ion channel simulator, offers a powerful and general approach to study ion channel permeation. BioMOCA is based on the Boltzmann Transport Monte Carlo (BTMC) and Particle-Particle-Particle-Mesh (P3M) methodologies developed at the University of Illinois at Urbana-Champaign. In this paper, we have employed BioMOCA to study two engineered mutations of α-HL, namely (M113F)6(M113C-D8RL2)1-β-CD and (M113N)6(T117C-D8RL3)1-β-CD. The channel conductance calculated by BioMOCA is slightly higher than experimental values. Permanent charge distributions and the geometrical shape of the channels gives rise to selectivity towards anions and also an asymmetry in I-V curves, promoting a rectification largely for cations. PMID:20938493

Radio-frequency response of single pores and artificial ion channels

NASA Astrophysics Data System (ADS)

Kim, H. S.; Ramachandran, S.; Stava, E.; van der Weide, D. W.; Blick, R. H.

2011-09-01

Intercellular communication relies on ion channels and pores in cell membranes. These protein-formed channels enable the exchange of ions and small molecules to electrically and/or chemically interact with the cells. Traditionally, recordings on single-ion channels and pores are performed in the dc regime, due to the extremely high impedance of these molecular junctions. This paper is intended as an introduction to radio-frequency (RF) recordings of single-molecule junctions in bilipid membranes. First, we demonstrate how early approaches to using microwave circuitry as readout devices for ion channel formation were realized. The second step will then focus on how to engineer microwave coupling into the high-impedance channel by making use of bio-compatible micro-coaxial lines. We then demonstrate integration of an ultra-broadband microwave circuit for the direct sampling of single α-hemolysin pores in a suspended bilipid membrane. Simultaneous direct current recordings reveal that we can monitor and correlate the RF transmission signal. This enables us to relate the open-close states of the direct current to the RF signal. Altogether, our experiments lay the ground for an RF-readout technique to perform real-time in vitro recordings of pores. The technique thus holds great promise for research and drug screening applications. The possible enhancement of sampling rates of single channels and pores by the large recording bandwidth will allow us to track the passage of single ions.

A laser microsurgical method of cell wall removal allows detection of large-conductance ion channels in the guard cell plasma membrane

NASA Technical Reports Server (NTRS)

Miedema, H.; Henriksen, G. H.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

1999-01-01

Application of patch clamp techniques to higher-plant cells has been subject to the limitation that the requisite contact of the patch electrode with the cell membrane necessitates prior enzymatic removal of the plant cell wall. Because the wall is an integral component of plant cells, and because cell-wall-degrading enzymes can disrupt membrane properties, such enzymatic treatments may alter ion channel behavior. We compared ion channel activity in enzymatically isolated protoplasts of Vicia faba guard cells with that found in membranes exposed by a laser microsurgical technique in which only a tiny portion of the cell wall is removed while the rest of the cell remains intact within its tissue environment. "Laser-assisted" patch clamping reveals a new category of high-conductance (130 to 361 pS) ion channels not previously reported in patch clamp studies on plant plasma membranes. These data indicate that ion channels are present in plant membranes that are not detected by conventional patch clamp techniques involving the production of individual plant protoplasts isolated from their tissue environment by enzymatic digestion of the cell wall. Given the large conductances of the channels revealed by laser-assisted patch clamping, we hypothesize that these channels play a significant role in the regulation of ion content and electrical signalling in guard cells.

Disease-associated changes in the expression of ion channels, ion receptors, ion exchangers and Ca{sup 2+}-handling proteins in heart hypertrophy

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

Zwadlo, Carolin; Borlak, Juergen

2005-09-15

The molecular pathology of cardiac hypertrophy is multifactorial with transcript regulation of ion channels, ion exchangers and Ca{sup 2+}-handling proteins being speculative. We therefore investigated disease-associated changes in gene expression of various ion channels and their receptors as well as ion exchangers, cytoskeletal proteins and Ca{sup 2+}-handling proteins in normotensive and spontaneously hypertensive (SHR) rats. We also compared experimental findings with results from hypertrophic human hearts, previously published (Borlak, J., and Thum, T., 2003. Hallmarks of ion channel gene expression in end-stage heart failure. FASEB J. 17, 1592-1608). We observed significant (P < 0.05) induction in transcript level of ATP-drivenmore » ion exchangers (Atp1A1, NCX-1, SERCA2a), ion channels (L-type Ca{sup 2+}-channel, K{sub ir}3.4, Na{sub v}1.5) and RyR-2 in hypertrophic hearts, while gene expression was repressed in diseased human hearts. Further, the genes coding for calreticulin and calmodulin, PMCA 1 and 4 as well as {alpha}-skeletal actin were significantly (P < 0.05) changed in hypertrophic human heart, but were unchanged in hypertrophic left ventricles of the rat heart. Notably, transcript level of {alpha}- and {beta}-MHC, calsequestrin, K{sub ir}6.1 (in the right ventricle only), phospholamban as well as troponin T were repressed in both diseased human and rat hearts. Our study enabled an identification of disease-associated candidate genes. Their regulation is likely to be the result of an imbalance between pressure load/stretch force and vascular tonus and the observed changes may provide a rational for the rhythm disturbances observed in patients with cardiac hypertrophy.« less

Regulatory cascade of neuronal loss and glucose metabolism.

PubMed

Hassan, Mubashir; Sehgal, Sheikh A; Rashid, Sajid

2014-01-01

During recent years, numerous lines of research including proteomics and molecular biology have highlighted multiple targets and signaling pathways involved in metabolic abnormalities and neurodegeneration. However, correlation studies of individual neurodegenerative disorders (ND) including Alzheimer, Parkinson, Huntington and Amyotrophic lateral sclerosis in association with Diabetes type 2 Mellitus (D2M) are demanding tasks. Here, we report a comprehensive mechanistic overview of major contributors involved in process-based co-regulation of D2M and NDs. D2M is linked with Alzheimer's disease through deregulation of calcium ions thereby leading to metabolic fluctuations of glucose and insulin. Parkinson-associated proteins disturb insulin level through ATP-sensitive potassium ion channels and extracellular signal-regulated kinases to enhance glucose level. Similarly, proteins which perturb carbohydrate metabolism for disturbing glucose homeostasis link Huntington, Amyotrophic lateral sclerosis and D2M. Other misleading processes which interconnect D2M and NDs include oxidative stress, mitochondrial dysfunctions and microRNAs (miRNA29a/b and miRNA-9). Overall, the collective listing of pathway-specific targets would help in establishing novel connections between NDs and D2M to explore better therapeutic interventions.

Hyperglycemia and Diabetes Downregulate the Functional Expression of TRPV4 Channels in Retinal Microvascular Endothelium

PubMed Central

Monaghan, Kevin; McNaughten, Jennifer; McGahon, Mary K.; Kelly, Catriona; Kyle, Daniel; Yong, Phaik Har

2015-01-01

Retinal endothelial cell dysfunction is believed to play a key role in the etiology and pathogenesis of diabetic retinopathy. Numerous studies have shown that TRPV4 channels are critically involved in maintaining normal endothelial cell function. In the current paper, we demonstrate that TRPV4 is functionally expressed in the endothelium of the retinal microcirculation and that both channel expression and activity is downregulated by hyperglycaemia. Quantitative PCR and immunostaining demonstrated molecular expression of TRPV4 in cultured bovine retinal microvascular endothelial cells (RMECs). Functional TRPV4 activity was assessed in cultured RMECs from endothelial Ca2+-responses recorded using fura-2 microfluorimetry and electrophysiological recordings of membrane currents. The TRPV4 agonist 4α-phorbol 12,13-didecanoate (4-αPDD) increased [Ca2+]i in RMECs and this response was largely abolished using siRNA targeted against TRPV4. These Ca2+-signals were completely inhibited by removal of extracellular Ca2+, confirming their dependence on influx of extracellular Ca2+. The 4-αPDD Ca2+-response recorded in the presence of cyclopiazonic acid (CPA), which depletes the intracellular stores preventing any signal amplification through store release, was used as a measure of Ca2+-influx across the cell membrane. This response was blocked by HC067047, a TRPV4 antagonist. Under voltage clamp conditions, the TRPV4 agonist GSK1016790A stimulated a membrane current, which was again inhibited by HC067047. Following incubation with 25mM D-glucose TRPV4 expression was reduced in comparison with RMECs cultured under control conditions, as were 4αPDD-induced Ca2+-responses in the presence of CPA and ion currents evoked by GSK1016790A. Molecular expression of TRPV4 in the retinal vascular endothelium of 3 months’ streptozotocin-induced diabetic rats was also reduced in comparison with that in age-matched controls. We conclude that hyperglycaemia and diabetes reduce the molecular and functional expression of TRPV4 channels in retinal microvascular endothelial cells. These changes may contribute to diabetes induced endothelial dysfunction and retinopathy. PMID:26047504

Role of Protein Flexibility in Ion Permeation: A Case Study in Gramicidin A

PubMed Central

Baştuğ, Turgut; Gray-Weale, Angus; Patra, Swarna M.; Kuyucak, Serdar

2006-01-01

Proteins have a flexible structure, and their atoms exhibit considerable fluctuations under normal operating conditions. However, apart from some enzyme reactions involving ligand binding, our understanding of the role of flexibility in protein function remains mostly incomplete. Here we investigate this question in the realm of membrane proteins that form ion channels. Specifically, we consider ion permeation in the gramicidin A channel, and study how the energetics of ion conduction changes as the channel structure is progressively changed from completely flexible to a fixed one. For each channel structure, the potential of mean force for a permeating potassium ion is determined from molecular dynamics (MD) simulations. Using the same molecular dynamics data for completely flexible gramicidin A, we also calculate the average densities and fluctuations of the peptide atoms and investigate the correlations between these fluctuations and the motion of a permeating ion. Our results show conclusively that peptide flexibility plays an important role in ion permeation in the gramicidin A channel, thus providing another reason—besides the well-known problem with the description of single file pore water—why this channel cannot be modeled using continuum electrostatics with a fixed structure. The new method developed here for studying the role of protein flexibility on its function clarifies the contributions of the fluctuations to energy and entropy, and places limits on the level of detail required in a coarse-grained model. PMID:16415054

Understanding the conductive channel evolution in Na:WO3-x-based planar devices

NASA Astrophysics Data System (ADS)

Shang, Dashan; Li, Peining; Wang, Tao; Carria, Egidio; Sun, Jirong; Shen, Baogen; Taubner, Thomas; Valov, Ilia; Waser, Rainer; Wuttig, Matthias

2015-03-01

An ion migration process in a solid electrolyte is important for ion-based functional devices, such as fuel cells, batteries, electrochromics, gas sensors, and resistive switching systems. In this study, a planar sandwich structure is prepared by depositing tungsten oxide (WO3-x) films on a soda-lime glass substrate, from which Na+ diffuses into the WO3-x films during the deposition. The entire process of Na+ migration driven by an alternating electric field is visualized in the Na-doped WO3-x films in the form of conductive channel by in situ optical imaging combined with infrared spectroscopy and near-field imaging techniques. A reversible change of geometry between a parabolic and a bar channel is observed with the resistance change of the devices. The peculiar channel evolution is interpreted by a thermal-stress-induced mechanical deformation of the films and an asymmetric Na+ mobility between the parabolic and the bar channels. These results exemplify a typical ion migration process driven by an alternating electric field in a solid electrolyte with a low ion mobility and are expected to be beneficial to improve the controllability of the ion migration in ion-based functional devices, such as resistive switching devices.An ion migration process in a solid electrolyte is important for ion-based functional devices, such as fuel cells, batteries, electrochromics, gas sensors, and resistive switching systems. In this study, a planar sandwich structure is prepared by depositing tungsten oxide (WO3-x) films on a soda-lime glass substrate, from which Na+ diffuses into the WO3-x films during the deposition. The entire process of Na+ migration driven by an alternating electric field is visualized in the Na-doped WO3-x films in the form of conductive channel by in situ optical imaging combined with infrared spectroscopy and near-field imaging techniques. A reversible change of geometry between a parabolic and a bar channel is observed with the resistance change of the devices. The peculiar channel evolution is interpreted by a thermal-stress-induced mechanical deformation of the films and an asymmetric Na+ mobility between the parabolic and the bar channels. These results exemplify a typical ion migration process driven by an alternating electric field in a solid electrolyte with a low ion mobility and are expected to be beneficial to improve the controllability of the ion migration in ion-based functional devices, such as resistive switching devices. Electronic supplementary information (ESI) available. See DOI: 10.1039/c4nr07545e

Nimodipine, an L-type calcium channel blocker attenuates mitochondrial dysfunctions to protect against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinsonism in mice.

PubMed

Singh, Alpana; Verma, Poonam; Balaji, Gillela; Samantaray, Supriti; Mohanakumar, Kochupurackal P

2016-10-01

Parkinson's disease (PD), the most common progressive neurodegenerative movement disorder, results from loss of dopaminergic neurons of substantia nigra pars compacta. These neurons exhibit Cav1.3 channel-dependent pacemaking activity. Epidemiological studies suggest reduced risk for PD in population under long-term antihypertensive therapy with L-type calcium channel antagonists. These prompted us to investigate nimodipine, an L-type calcium channel blocker for neuroprotective effect in cellular and animal models of PD. Nimodipine (0.1-10 μM) significantly attenuated 1-methyl-4-phenyl pyridinium ion-induced loss in mitochondrial morphology, mitochondrial membrane potential and increases in intracellular calcium levels in SH-SY5Y neuroblastoma cell line as measured respectively employing Mitotracker green staining, TMRM, and Fura-2 fluorescence, but only a feeble neuroprotective effect was observed in MTT assay. Nimodipine dose-dependently reduced 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced parkinsonian syndromes (akinesia and catalepsy) and loss in swimming ability in Balb/c mice. It attenuated MPTP-induced loss of dopaminergic tyrosine hydroxylase positive neurons in substantia nigra, improved mitochondrial oxygen consumption and inhibited reactive oxygen species production in the striatal mitochondria measured using dichlorodihydrofluorescein fluorescence, but failed to block striatal dopamine depletion. These results point to an involvement of L-type calcium channels in MPTP-induced dopaminergic neuronal death in experimental parkinsonism and more importantly provide evidences for nimodipine to improve mitochondrial integrity and function. Copyright © 2016 Elsevier Ltd. All rights reserved.

[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.

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

Computational model based approach to analysis ventricular arrhythmias: Effects of dysfunction calcium channels

NASA Astrophysics Data System (ADS)

Gulothungan, G.; Malathi, R.

2018-04-01

Disturbed sodium (Na+) and calcium (Ca2+) handling is known to be a major predisposing factor for life-threatening cardiac arrhythmias. Cardiac contractility in ventricular tissue is prominent by Ca2+ channels like voltage dependent Ca2+ channels, sodium-calcium exchanger (Na+-Ca2+x) and sacroplasmicrecticulum (SR) Ca2+ pump and leakage channels. Experimental and clinical possibilities for studying cardiac arrhythmias in human ventricular myocardium are very limited. Therefore, the use of alternative methods such as computer simulations is of great importance. Our aim of this article is to study the impact on action potential (AP) generation and propagation in single ventricular myocyte and ventricular tissue under different dysfunction Ca2+ channels condition. In enhanced activity of Na+-Ca2+x, single myocyte produces AP duration (APD90) and APD50 is significantly smaller (266 ms and 235 ms). Its Na+-Ca2+x current at depolarization is increases 60% from its normal level and repolarization current goes more negative (nonfailing= -0.28 pA/pF and failing= -0.47 pA/pF). Similarly, same enhanced activity of Na+-Ca2+x in 10 mm region of ventricular sheet, raises the plateau potential abruptly, which ultimately affects the diastolic repolarization. Compare with normal ventricular sheet region of 10 mm, 10% of ventricular sheet resting state is reduces and ventricular sheet at time 250 ms is goes to resting state very early. In hypertrophy condition, single myocyte produces APD90 and APD50 is worthy of attention smaller (232 mS and 198 ms). Its sodium-potassium (Na+-K+) pump current is 75% reduces from its control conditions (0.13 pA/pF). Hypertrophy condition, 50% of ventricular sheet is reduces to minimum plateau potential state, that starts the repolarization process very early and reduces the APD. In a single failing SR Ca2+ channels myocyte, recovery of Ca2+ concentration level in SR reduces upto 15% from its control myocytes. At time 290 ms, 70% of ventricular sheet is in dysfunction resting potential state in the range -83 mV and ventricular sheet at time 295 ms is goes to 65% dysfunction resting state. Therefore we concluded that shorter APD, instability resting potential and affected calcium induced calcium release (CICR) due to dysfunction Ca2+ channels is potentially have a substantial effect on cardiac contractility and relaxation. Computational study on ventricular tissue AP and its underlying ionic channel currents could help to elucidate possible arrhythmogenic mechanism on a cellular level.

Charging the quantum capacitance of graphene with a single biological ion channel.

PubMed

Wang, Yung Yu; Pham, Ted D; Zand, Katayoun; Li, Jinfeng; Burke, Peter J

2014-05-27

The interaction of cell and organelle membranes (lipid bilayers) with nanoelectronics can enable new technologies to sense and measure electrophysiology in qualitatively new ways. To date, a variety of sensing devices have been demonstrated to measure membrane currents through macroscopic numbers of ion channels. However, nanoelectronic based sensing of single ion channel currents has been a challenge. Here, we report graphene-based field-effect transistors combined with supported lipid bilayers as a platform for measuring, for the first time, individual ion channel activity. We show that the supported lipid bilayers uniformly coat the single layer graphene surface, acting as a biomimetic barrier that insulates (both electrically and chemically) the graphene from the electrolyte environment. Upon introduction of pore-forming membrane proteins such as alamethicin and gramicidin A, current pulses are observed through the lipid bilayers from the graphene to the electrolyte, which charge the quantum capacitance of the graphene. This approach combines nanotechnology with electrophysiology to demonstrate qualitatively new ways of measuring ion channel currents.

Charging the Quantum Capacitance of Graphene with a Single Biological Ion Channel

PubMed Central

2015-01-01

The interaction of cell and organelle membranes (lipid bilayers) with nanoelectronics can enable new technologies to sense and measure electrophysiology in qualitatively new ways. To date, a variety of sensing devices have been demonstrated to measure membrane currents through macroscopic numbers of ion channels. However, nanoelectronic based sensing of single ion channel currents has been a challenge. Here, we report graphene-based field-effect transistors combined with supported lipid bilayers as a platform for measuring, for the first time, individual ion channel activity. We show that the supported lipid bilayers uniformly coat the single layer graphene surface, acting as a biomimetic barrier that insulates (both electrically and chemically) the graphene from the electrolyte environment. Upon introduction of pore-forming membrane proteins such as alamethicin and gramicidin A, current pulses are observed through the lipid bilayers from the graphene to the electrolyte, which charge the quantum capacitance of the graphene. This approach combines nanotechnology with electrophysiology to demonstrate qualitatively new ways of measuring ion channel currents. PMID:24754625

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

Superstatistics analysis of the ion current distribution function: Met3PbCl influence study.

PubMed

Miśkiewicz, Janusz; Trela, Zenon; Przestalski, Stanisław; Karcz, Waldemar

2010-09-01

A novel analysis of ion current time series is proposed. It is shown that higher (second, third and fourth) statistical moments of the ion current probability distribution function (PDF) can yield new information about ion channel properties. The method is illustrated on a two-state model where the PDF of the compound states are given by normal distributions. The proposed method was applied to the analysis of the SV cation channels of vacuolar membrane of Beta vulgaris and the influence of trimethyllead chloride (Met(3)PbCl) on the ion current probability distribution. Ion currents were measured by patch-clamp technique. It was shown that Met(3)PbCl influences the variance of the open-state ion current but does not alter the PDF of the closed-state ion current. Incorporation of higher statistical moments into the standard investigation of ion channel properties is proposed.

Dielectric boundary force and its crucial role in gramicidin

NASA Astrophysics Data System (ADS)

Nadler, Boaz; Hollerbach, Uwe; Eisenberg, R. S.

2003-08-01

In an electrostatic problem with nonuniform geometry, a charge Q in one region induces surface charges [called dielectric boundary charges (DBC)] at boundaries between different dielectrics. These induced surface charges, in return, exert a force [called dielectric boundary force (DBF)] on the charge Q that induced them. The DBF is often overlooked. It is not present in standard continuum theories of (point) ions in or near membranes and proteins, such as Gouy-Chapman, Debye-Huckel, Poisson-Boltzmann or Poisson-Nernst- Planck. The DBF is important when a charge Q is near dielectric interfaces, for example, when ions permeate through protein channels embedded in biological membranes. In this paper, we define the DBF and calculate it explicitly for a planar dielectric wall and for a tunnel geometry resembling the ionic channel gramicidin. In general, we formulate the DBF in a form useful for continuum theories, namely, as a solution of a partial differential equation with boundary conditions. The DBF plays a crucial role in the permeation of ions through the gramicidin channel. A positive ion in the channel produces a DBF of opposite sign to that of the fixed charge force (FCF) produced by the permanent charge of the gramicidin polypeptide, and so the net force on the positive ion is reduced. A negative ion creates a DBF of the same sign as the FCF and so the net (repulsive) force on the negative ion is increased. Thus, a positive ion can permeate the channel, while a negative ion is excluded from it. In gramicidin, it is this balance between the FCF and DBF that allows only singly charged positive ions to move into and through the channel. The DBF is not directly responsible, however, for selectivity between the alkali metal ions (e.g., Li+, Na+, K+): we prove that the DBF on a mobile spherical ion is independent of the ion’s radius.

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.

Self-Assembling Organic Nanopores as Synthetic Transmembrane Channels with Tunable Functions

NASA Astrophysics Data System (ADS)

Wei, Xiaoxi

A long-standing goal in the area of supramolecular self-assembly involves the development of synthetic ion/water channels capable of mimicking the mass-transport characteristics of biological channels and pores. Few examples of artificial transmembrane channels with large lumen, high conductivity and selectivity are known. A review of pronounced biological transmembrane protein channels and some representative synthetic models have been provided in Chapter 1, followed by our discovery and initial investigation of shape-persistent oligoamide and phenylene ethynylene macrocycles as synthetic ion/water channels. In Chapter 2, the systematic structural modification of oligoamide macrocycles 1, the so-called first-generation of these shape-persistent macrocycles, has led to third-generation macrocycles 3. The third generation was found to exhibit unprecedented, strong intermolecular association in both the solid state and solution via multiple techniques including X-ray diffraction (XRD), SEM, and 1H NMR. Fluorescence spectroscopy paired with dynamic light scattering (DLS) revealed that macrocycles 3 can assemble into a singly dispersed nanotubular structure in solution. The resultant self-assembling pores consisting of 3 were examined by HPTS-LUVs assays and BLM studies (Chapter 3) and found to form cation-selective (PK+/PCl- = 69:1) transmembrane ion channels with large conductance (200 ˜ 2000 pS for alkali cations) and high stability with open times reaching to 103 seconds. Tuning the aggregation state of macrocycles by choosing an appropriate polar solvent mixture (i.e., 3:1, THF:DMF, v/v) and concentration led to the formation of ion channels with well-defined square top behavior. A parallel study using DLS to examine the size of aggregates was used in conjunction with channel activity assays (LUVs/BLM) to reveal the effects of the aggregation state on channel activity. Empirical evidence now clearly indicates that a preassembled state, perhaps that of a nanotubular assembly, rather than the individual molecules of 3, is required to partition into the lipid bilayer in order for these macrocycles to act as channels. Further structural modification has led to fourth-generation macrocycles 4 having readily-tunable cavities (Chapter 4). Macrocycles 4 , with a hybrid backbone composed half of the oligoamide and half of the phenylene ethynylene moieties, exhibits similar self-assembling behavior by forming nanotubular stacks. The results of a preliminary study based on LUVs-assays and BLM single channel recording experiments are summarized and clearly indicate that ion channels formed by this fourth-generation exhibit high stability and differing ion selectivity largely consistent with the corresponding structural modification of the interior cavity. Especially, the increased anion conductance observed for 4d indicates that our strategy of tuning supramolecular function based on synthetic modification of the backbone and pore is effective. In Chapter 5, our four-residue tetraurea macrocycles 5 have shown significant potency to selectively interact with the G-quadruplex, leading to a strong stabilization effect for G-quadruplex without binding to duplex DNA as observed by UV-melt assays. The ready synthetic availability of these macrocycles makes them amenable to future chemical modification, which allows systematic improvement of binding affinity and specificity. Moreover, it has been discovered that these macrocycles can partition into lipid bilayers and form very stable transmembrane ion channels with a pore size of ˜5 A. Preliminary data shows that this smaller ion channel may lead to exceptional ion conducting selectivity, which is rarely seen in the field of synthetic ion pores. These molecules may serve as a unique platform for the rational development of potent and versatile therapeutic agents. The exceptional ion conducting properties of these channels place aromatic oligoamide macrocycles 3 and 4 at a unique position with both high conductance and long channel-opening duration. These results demonstrate that oligoamide macrocycles provide us a reliable platform based on which further development of highly conducting and selective synthetic mass-transporting channels, with functions that are comparable to or even rival those of natural channels and pores, may be developed. Further improvement of these synthetic channels could lead to numerous applications, such as those for complementing ion channel deficiency in clinical medicine, designing biosensors, and the development of new materials, as well as their use in separation and purifications.

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

Hepatitis C virus p7 mediates membrane-to-membrane adhesion.

PubMed

Lee, Gi Young; Lee, Sora; Lee, Hye-Ra; Yoo, Young Do

2016-09-01

Viroporin p7 of the hepatitis C virus (HCV) acts as an ion channel for pH equilibration to stabilize HCV particles; most studies of p7 have focused on this role. However, pH equilibration by p7 via its ion channel activity does not fully explain the importance of p7 in HCV particle production. Indeed, several researchers have suggested p7 to have an unidentified ion channel-independent function. Here, we show that p7 has a novel role as a lipid raft adhesion factor, which is independent of its ion channel activity. We found that p7 targets not only the liquid-disordered (Ld) phase, but also the negatively-charged liquid-ordered (Lo) phase that can be represented as a lipid raft. p7 clusters at the phase boundary of the neutral Ld phase and the negatively-charged Lo phase. Interestingly, p7 targeting the Lo phase facilitates membrane-to-membrane adhesion, and this activity is not inhibited by p7 ion channel inhibitors. Our results demonstrated that HCV p7 has dual roles as a viroporin and as a lipid raft adhesion factor. This ion channel-independent function of p7 might be an attractive target for development of anti-HCV compounds. Copyright © 2016 Elsevier B.V. All rights reserved.

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.

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.

Turning a Poor Ion Channel into a Good Pump

NASA Astrophysics Data System (ADS)

Astumian, Dean

2003-05-01

We consider a membrane protein that can exist in two configurations, either one of which acts as a poor ion channel, allowing ions to slowly leak across the membrane from high to low elctrochemical potential. We show that random external fluctuations can provide the energy to turn this poor channel into a good pump that drives ion transport from low to high electrochemical potential. We discuss this result in terms of a gambling analogy, and point to possible implications for fields as far ranging as population biology, economics, and actuarial science.

Computational study of a calcium release-activated calcium channel

NASA Astrophysics Data System (ADS)

Talukdar, Keka; Shantappa, Anil

2016-05-01

The naturally occurring proteins that form hole in membrane are commonly known as ion channels. They play multiple roles in many important biological processes. Deletion or alteration of these channels often leads to serious problems in the physiological processes as it controls the flow of ions through it. The proper maintenance of the flow of ions, in turn, is required for normal health. Here we have investigated the behavior of a calcium release-activated calcium ion channel with pdb entry 4HKR in Drosophila Melanogaster. The equilibrium energy as well as molecular dynamics simulation is performed first. The protein is subjected to molecular dynamics simulation to find their energy minimized value. Simulation of the protein in the environment of water and ions has given us important results too. The solvation energy is also found using Charmm potential.

Subacute motor neuron hyperexcitability with mercury poisoning: a case series and literature review.

PubMed

Zhou, Zhibin; Zhang, Xingwen; Cui, Fang; Liu, Ruozhuo; Dong, Zhao; Wang, Xiaolin; Yu, Shengyuan

2014-01-01

Motor neuron hyperexcitability (MNH) indicates a disorder characterized by an ectopic motor nerve discharge on electromyogram (EMG). Here, we present a series of three cases of subacute MNH with mercury poisoning. The first case showed hyperhidrosis, insomnia, generalied myokymia, cramps, tremor, weight loss, and myokymic and neuromyotonic discharges, followed by encephalopathy with confusion, hallucinations, and memory decrease. The second case was similar to the former but without encephalopathic features. The third case showed widespread fasciculation, fatigue, insomnia, weight loss, and autonomic dysfunction, including constipation, micturition difficulty, and impotence, with multiple fibrillation, unstable fasciculation, widened motor neuron potential, and an incremental response at high-rate stimulation in repetitive nerve stimulation. Based on the symptoms, the three cases were diagnosed as Morvan's syndrome, Isaacs' syndrome, and Lambert-Eaton myasthenic syndrome with ALS-like syndrome, respectively. Mercury poisoning in the three cases was confirmed by analysis of blood and urine samples. All cases recovered several months after chelation therapy and were in good condition at follow-up. Very few cases of MNH linked with mercury exposure have been reported in the literature. The mechanism of mercury-induced MNH may be associated with ion channel dysfunction. © 2014 S. Karger AG, Basel.

Gsk3 Signalling and Redox Status in Bipolar Disorder: Evidence from Lithium Efficacy

PubMed Central

2016-01-01

Objective. To discuss the link between glycogen synthase kinase-3 (GSK3) and the main biological alterations demonstrated in bipolar disorder (BD), with special attention to the redox status and the evidence supporting the efficacy of lithium (a GSK3 inhibitor) in the treatment of BD. Methods. A literature research on the discussed topics, using Pubmed and Google Scholar, has been conducted. Moreover, a manual selection of interesting references from the identified articles has been performed. Results. The main biological alterations of BD, pertaining to inflammation, oxidative stress, membrane ion channels, and circadian system, seem to be intertwined. The dysfunction of the GSK3 signalling pathway is involved in all the aforementioned “biological causes” of BD. In a complex scenario, it can be seen as the common denominator linking them all. Lithium inhibition of GSK3 could, at least in part, explain its positive effect on these biological dysfunctions and its superiority in terms of clinical efficacy. Conclusions. Deepening the knowledge on the molecular bases of BD is fundamental to identifying the biochemical pathways that must be targeted in order to provide patients with increasingly effective therapeutic tools against an invalidating disorder such as BD. PMID:27630757

Gsk3 Signalling and Redox Status in Bipolar Disorder: Evidence from Lithium Efficacy.

PubMed

Luca, Antonina; Calandra, Carmela; Luca, Maria

2016-01-01

Objective. To discuss the link between glycogen synthase kinase-3 (GSK3) and the main biological alterations demonstrated in bipolar disorder (BD), with special attention to the redox status and the evidence supporting the efficacy of lithium (a GSK3 inhibitor) in the treatment of BD. Methods. A literature research on the discussed topics, using Pubmed and Google Scholar, has been conducted. Moreover, a manual selection of interesting references from the identified articles has been performed. Results. The main biological alterations of BD, pertaining to inflammation, oxidative stress, membrane ion channels, and circadian system, seem to be intertwined. The dysfunction of the GSK3 signalling pathway is involved in all the aforementioned "biological causes" of BD. In a complex scenario, it can be seen as the common denominator linking them all. Lithium inhibition of GSK3 could, at least in part, explain its positive effect on these biological dysfunctions and its superiority in terms of clinical efficacy. Conclusions. Deepening the knowledge on the molecular bases of BD is fundamental to identifying the biochemical pathways that must be targeted in order to provide patients with increasingly effective therapeutic tools against an invalidating disorder such as BD.

Optogenetic approaches to evaluate striatal function in animal models of Parkinson disease.

PubMed

Parker, Krystal L; Kim, Youngcho; Alberico, Stephanie L; Emmons, Eric B; Narayanan, Nandakumar S

2016-03-01

Optogenetics refers to the ability to control cells that have been genetically modified to express light-sensitive ion channels. The introduction of optogenetic approaches has facilitated the dissection of neural circuits. Optogenetics allows for the precise stimulation and inhibition of specific sets of neurons and their projections with fine temporal specificity. These techniques are ideally suited to investigating neural circuitry underlying motor and cognitive dysfunction in animal models of human disease. Here, we focus on how optogenetics has been used over the last decade to probe striatal circuits that are involved in Parkinson disease, a neurodegenerative condition involving motor and cognitive abnormalities resulting from degeneration of midbrain dopaminergic neurons. The precise mechanisms underlying the striatal contribution to both cognitive and motor dysfunction in Parkinson disease are unknown. Although optogenetic approaches are somewhat removed from clinical use, insight from these studies can help identify novel therapeutic targets and may inspire new treatments for Parkinson disease. Elucidating how neuronal and behavioral functions are influenced and potentially rescued by optogenetic manipulation in animal models could prove to be translatable to humans. These insights can be used to guide future brain-stimulation approaches for motor and cognitive abnormalities in Parkinson disease and other neuropsychiatric diseases.

Ion channel-mediated uptake of cationic vital dyes into live cells: a potential source of error when assessing cell viability.

PubMed

Bukhari, Maurish; Burm, Hayley; Samways, Damien S K

2016-10-01

Ionic "vital dyes" are commonly used to assess cell viability based on the idea that their permeation is contingent on a loss of membrane integrity. However, the possibility that dye entry is conducted into live cells by endogenous membrane transporters must be recognized and controlled for. Several cation-selective plasma membrane-localized ion channels, including the adenosine 5'-triphosphate (ATP)-gated P2X receptors, have been reported to conduct entry of the DNA-binding fluorescence dye, YO-PRO-1, into live cells. Extracellular ATP often becomes elevated as a result of release from dying cells, and so it is possible that activation of P2X channels on neighboring live cells could lead to exaggerated estimation of cytotoxicity. Here, we screened a number of fluorescent vital dyes for ion channel-mediated uptake in HEK293 cells expressing recombinant P2X2, P2X7, or TRPV1 channels. Our data shows that activation of all three channels caused substantial uptake and nuclear accumulation of YO-PRO-1, 4',6-diamidino-2-phenylindole (DAPI), and Hoechst 33258 into transfected cells and did so well within the time period usually used for incubation of cells with vital dyes. In contrast, channel activation in the presence of propidium iodide and SYTOX Green caused no measurable uptake and accumulation during a 20-min exposure, suggesting that these dyes are not likely to exhibit measurable uptake through these particular ion channels during a conventional cell viability assay. Caution is encouraged when choosing and employing cationic dyes for the purpose of cell viability assessment, particularly when there is a likelihood of cells expressing ion channels permeable to large ions.

Allosteric modulation balances thermodynamic stability and restores function of ΔF508 CFTR

PubMed Central

Aleksandrov, Andrei A.; Kota, Pradeep; Cui, Liying; Jensen, Tim; Alekseev, Alexey E.; Reyes, Santiago; He, Lihua; Gentzsch, Martina; Aleksandrov, Luba A.; Dokholyan, Nikolay V.; Riordan, John R.

2013-01-01

Most cystic fibrosis is caused by a deletion of a single residue (F508) in CFTR that disrupts the folding and biosynthetic maturation of the ion channel protein. Progress towards understanding the underlying mechanisms and overcoming the defect remain incomplete. Here we show that the thermal instability of human ΔF508 CFTR channel activity evident in both cell-attached membrane patches and planar phospholipid bilayers is not observed in corresponding mutant CFTRs of several non-mammalian species. These more stable orthologs are distinguished from their mammalian counterparts by the substitution of proline residues at several key dynamic locations in the first nucleotide domain (NBD1), including the structurally diverse region (SDR), the gamma phosphate switch loop and the Regulatory Insertion (RI). Molecular Dynamic analyses revealed that addition of the prolines could reduce flexibility at these locations and increase the temperatures of unfolding transitions of ΔF508 NBD1 to that of the wild-type. Introduction of these prolines experimentally into full-length human ΔF508 CFTR together with the already recognized I539T suppressor mutation, also in the SDR, restored channel function and thermodynamic stability as well as its trafficking to and lifetime at the cell surface. Thus, while cellular manipulations that circumvent its culling by quality control systems leave ΔF508 CFTR dysfunctional at physiological temperature, restoration of the delicate balance between the dynamic protein’s inherent stability and channel activity returns a near-normal state. PMID:22406676

Projection structure of a ClC-type chloride channel at 6.5Å resolution

NASA Astrophysics Data System (ADS)

Mindell, Joseph A.; Maduke, Merritt; Miller, Christopher; Grigorieff, Nikolaus

2001-01-01

Virtually all cells in all eukaryotic organisms express ion channels of the ClC type, the only known molecular family of chloride-ion-selective channels. The diversity of ClC channels highlights the multitude and range of functions served by gated chloride-ion conduction in biological membranes, such as controlling electrical excitability in skeletal muscle, maintaining systemic blood pressure, acidifying endosomal compartments, and regulating electrical responses of GABA (γ-aminobutyric acid)-containing interneurons in the central nervous system. Previously, we expressed and purified a prokaryotic ClC channel homologue. Here we report the formation of two-dimensional crystals of this ClC channel protein reconstituted into phospholipid bilayer membranes. Cryo-electron microscopic analysis of these crystals yields a projection structure at 6.5Å resolution, which shows off-axis water-filled pores within the dimeric channel complex.

The Transcription Factors Islet and Lim3 Combinatorially Regulate Ion Channel Gene Expression

PubMed Central

Wolfram, Verena; Southall, Tony D.; Günay, Cengiz; Prinz, Astrid A.; Brand, Andrea H.

2014-01-01

Expression of appropriate ion channels is essential to allow developing neurons to form functional networks. Our previous studies have identified LIM-homeodomain (HD) transcription factors (TFs), expressed by developing neurons, that are specifically able to regulate ion channel gene expression. In this study, we use the technique of DNA adenine methyltransferase identification (DamID) to identify putative gene targets of four such TFs that are differentially expressed in Drosophila motoneurons. Analysis of targets for Islet (Isl), Lim3, Hb9, and Even-skipped (Eve) identifies both ion channel genes and genes predicted to regulate aspects of dendritic and axonal morphology. Significantly, some ion channel genes are bound by more than one TF, consistent with the possibility of combinatorial regulation. One such gene is Shaker (Sh), which encodes a voltage-dependent fast K+ channel (Kv1.1). DamID reveals that Sh is bound by both Isl and Lim3. We used body wall muscle as a test tissue because in conditions of low Ca2+, the fast K+ current is carried solely by Sh channels (unlike neurons in which a second fast K+ current, Shal, also contributes). Ectopic expression of isl, but not Lim3, is sufficient to reduce both Sh transcript and Sh current level. By contrast, coexpression of both TFs is additive, resulting in a significantly greater reduction in both Sh transcript and current compared with isl expression alone. These observations provide evidence for combinatorial activity of Isl and Lim3 in regulating ion channel gene expression. PMID:24523544

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

High-throughput electrophysiological assays for voltage gated ion channels using SyncroPatch 768PE.

PubMed

Li, Tianbo; Lu, Gang; Chiang, Eugene Y; Chernov-Rogan, Tania; Grogan, Jane L; Chen, Jun

2017-01-01

Ion channels regulate a variety of physiological processes and represent an important class of drug target. Among the many methods of studying ion channel function, patch clamp electrophysiology is considered the gold standard by providing the ultimate precision and flexibility. However, its utility in ion channel drug discovery is impeded by low throughput. Additionally, characterization of endogenous ion channels in primary cells remains technical challenging. In recent years, many automated patch clamp (APC) platforms have been developed to overcome these challenges, albeit with varying throughput, data quality and success rate. In this study, we utilized SyncroPatch 768PE, one of the latest generation APC platforms which conducts parallel recording from two-384 modules with giga-seal data quality, to push these 2 boundaries. By optimizing various cell patching parameters and a two-step voltage protocol, we developed a high throughput APC assay for the voltage-gated sodium channel Nav1.7. By testing a group of Nav1.7 reference compounds' IC50, this assay was proved to be highly consistent with manual patch clamp (R > 0.9). In a pilot screening of 10,000 compounds, the success rate, defined by > 500 MΩ seal resistance and >500 pA peak current, was 79%. The assay was robust with daily throughput ~ 6,000 data points and Z' factor 0.72. Using the same platform, we also successfully recorded endogenous voltage-gated potassium channel Kv1.3 in primary T cells. Together, our data suggest that SyncroPatch 768PE provides a powerful platform for ion channel research and drug discovery.

Revealing dynamically-organized receptor ion channel clusters in live cells by a correlated electric recording and super-resolution single-molecule imaging approach.

PubMed

Yadav, Rajeev; Lu, H Peter

2018-03-28

The N-methyl-d-aspartate (NMDA) receptor ion-channel is activated by the binding of ligands, along with the application of action potential, important for synaptic transmission and memory functions. Despite substantial knowledge of the structure and function, the gating mechanism of the NMDA receptor ion channel for electric on-off signals is still a topic of debate. We investigate the NMDA receptor partition distribution and the associated channel's open-close electric signal trajectories using a combined approach of correlating single-molecule fluorescence photo-bleaching, single-molecule super-resolution imaging, and single-channel electric patch-clamp recording. Identifying the compositions of NMDA receptors, their spatial organization and distributions over live cell membranes, we observe that NMDA receptors are organized inhomogeneously: nearly half of the receptor proteins are individually dispersed; whereas others exist in heterogeneous clusters of around 50 nm in size as well as co-localized within the diffraction limited imaging area. We demonstrate that inhomogeneous interactions and partitions of the NMDA receptors can be a cause of the heterogeneous gating mechanism of NMDA receptors in living cells. Furthermore, comparing the imaging results with the ion-channel electric current recording, we propose that the clustered NMDA receptors may be responsible for the variation in the current amplitude observed in the on-off two-state ion-channel electric signal trajectories. Our findings shed new light on the fundamental structure-function mechanism of NMDA receptors and present a conceptual advancement of the ion-channel mechanism in living cells.

Obtaining spheroplasts of armored dinoflagellates and first single-channel recordings of their ion channels using patch-clamping.

PubMed

Pozdnyakov, Ilya; Matantseva, Olga; Negulyaev, Yuri; Skarlato, Sergei

2014-09-05

Ion channels are tightly involved in various aspects of cell physiology, including cell signaling, proliferation, motility, endo- and exo-cytosis. They may be involved in toxin production and release by marine dinoflagellates, as well as harmful algal bloom proliferation. So far, the patch-clamp technique, which is the most powerful method to study the activity of ion channels, has not been applied to dinoflagellate cells, due to their complex cellulose-containing cell coverings. In this paper, we describe a new approach to overcome this problem, based on the preparation of spheroplasts from armored bloom-forming dinoflagellate Prorocentrum minimum. We treated the cells of P. minimum with a cellulose synthesis inhibitor, 2,6-dichlorobenzonitrile (DCB), and found out that it could also induce ecdysis and arrest cell shape maintenance in these microalgae. Treatment with 100-250 µM DCB led to an acceptable 10% yield of P. minimum spheroplasts and was independent of the incubation time in the range of 1-5 days. We show that such spheroplasts are suitable for patch-clamping in the cell-attached mode and can form 1-10 GOhm patch contact with a glass micropipette, allowing recording of ion channel activity. The first single-channel recordings of dinoflagellate ion channels are presented.

Obtaining Spheroplasts of Armored Dinoflagellates and First Single-Channel Recordings of Their Ion Channels Using Patch-Clamping

PubMed Central

Pozdnyakov, Ilya; Matantseva, Olga; Negulyaev, Yuri; Skarlato, Sergei

2014-01-01

Ion channels are tightly involved in various aspects of cell physiology, including cell signaling, proliferation, motility, endo- and exo-cytosis. They may be involved in toxin production and release by marine dinoflagellates, as well as harmful algal bloom proliferation. So far, the patch-clamp technique, which is the most powerful method to study the activity of ion channels, has not been applied to dinoflagellate cells, due to their complex cellulose-containing cell coverings. In this paper, we describe a new approach to overcome this problem, based on the preparation of spheroplasts from armored bloom-forming dinoflagellate Prorocentrum minimum. We treated the cells of P. minimum with a cellulose synthesis inhibitor, 2,6-dichlorobenzonitrile (DCB), and found out that it could also induce ecdysis and arrest cell shape maintenance in these microalgae. Treatment with 100–250 µM DCB led to an acceptable 10% yield of P. minimum spheroplasts and was independent of the incubation time in the range of 1–5 days. We show that such spheroplasts are suitable for patch-clamping in the cell-attached mode and can form 1–10 GOhm patch contact with a glass micropipette, allowing recording of ion channel activity. The first single-channel recordings of dinoflagellate ion channels are presented. PMID:25199048

Mechanism underlying impaired cardiac pacemaking rhythm during ischemia: A simulation study

NASA Astrophysics Data System (ADS)

Bai, Xiangyun; Wang, Kuanquan; Yuan, Yongfeng; Li, Qince; Dobrzynski, Halina; Boyett, Mark R.; Hancox, Jules C.; Zhang, Henggui

2017-09-01

Ischemia in the heart impairs function of the cardiac pacemaker, the sinoatrial node (SAN). However, the ionic mechanisms underlying the ischemia-induced dysfunction of the SAN remain elusive. In order to investigate the ionic mechanisms by which ischemia causes SAN dysfunction, action potential models of rabbit SAN and atrial cells were modified to incorporate extant experimental data of ischemia-induced changes to membrane ion channels and intracellular ion homeostasis. The cell models were incorporated into an anatomically detailed 2D model of the intact SAN-atrium. Using the multi-scale models, the functional impact of ischemia-induced electrical alterations on cardiac pacemaking action potentials (APs) and their conduction was investigated. The effects of vagal tone activity on the regulation of cardiac pacemaker activity in control and ischemic conditions were also investigated. The simulation results showed that at the cellular level ischemia slowed the SAN pacemaking rate, which was mainly attributable to the altered Na+-Ca2+ exchange current and the ATP-sensitive potassium current. In the 2D SAN-atrium tissue model, ischemia slowed down both the pacemaking rate and the conduction velocity of APs into the surrounding atrial tissue. Simulated vagal nerve activity, including the actions of acetylcholine in the model, amplified the effects of ischemia, leading to possible SAN arrest and/or conduction exit block, which are major features of the sick sinus syndrome. In conclusion, this study provides novel insights into understanding the mechanisms by which ischemia alters SAN function, identifying specific conductances as contributors to bradycardia and conduction block.

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.

Overcharging below the nanoscale: Multivalent cations reverse the ion selectivity of a biological channel

NASA Astrophysics Data System (ADS)

García-Giménez, Elena; Alcaraz, Antonio; Aguilella, Vicente M.

2010-02-01

We report charge inversion within a nanoscopic biological protein ion channel in salts of multivalent ions. The presence of positive divalent and trivalent counterions reverses the cationic selectivity of the OmpF channel, a general diffusion porin located in the outer membrane of E. coli. We discuss the conditions under which charge inversion can be inferred from the change in sign of the measured quantity, the channel zero current potential. By comparing experimental results in protein channels whose charge has been modified after site-directed mutagenesis, the predictions of current theories of charge inversion are critically examined. It is emphasized that charge inversion does not necessarily increase with the bare surface charge density of the interface and that even this concept of surface charge density may become meaningless in some biological ion channels. Thus, any theory based on electrostatic correlations or chemical binding should explicitly take into account the particular structure of the charged interface.

Optical Waveguide Lightmode Spectroscopic Techniques for Investigating Membrane-Bound Ion Channel Activities

PubMed Central

Székács, Inna; Kaszás, Nóra; Gróf, Pál; Erdélyi, Katalin; Szendrő, István; Mihalik, Balázs; Pataki, Ágnes; Antoni, Ferenc A.; Madarász, Emilia

2013-01-01

Optical waveguide lightmode spectroscopic (OWLS) techniques were probed for monitoring ion permeation through channels incorporated into artificial lipid environment. A novel sensor set-up was developed by depositing liposomes or cell-derived membrane fragments onto hydrophilic polytetrafluoroethylene (PTFE) membrane. The fibrous material of PTFE membrane could entrap lipoid vesicles and the water-filled pores provided environment for the hydrophilic domains of lipid-embedded proteins. The sensor surface was kept clean from the lipid holder PTFE membrane by a water- and ion-permeable polyethylene terephthalate (PET) mesh. The sensor set-up was tested with egg yolk lecithin liposomes containing gramicidin ion channels and with cell-derived membrane fragments enriched in GABA-gated anion channels. The method allowed monitoring the move of Na+ and organic cations through gramicidin channels and detecting the Cl–-channel functions of the (α5β2γ2) GABAA receptor in the presence or absence of GABA and the competitive GABA-blocker bicuculline. PMID:24339925

Ion Transport through Membrane-Spanning Nanopores Studied by Molecular Dynamics Simulations and Continuum Electrostatics Calculations

PubMed Central

Peter, Christine; Hummer, Gerhard

2005-01-01

Narrow hydrophobic regions are a common feature of biological channels, with possible roles in ion-channel gating. We study the principles that govern ion transport through narrow hydrophobic membrane pores by molecular dynamics simulation of model membranes formed of hexagonally packed carbon nanotubes. We focus on the factors that determine the energetics of ion translocation through such nonpolar nanopores and compare the resulting free-energy barriers for pores with different diameters corresponding to the gating regions in closed and open forms of potassium channels. Our model system also allows us to compare the results from molecular dynamics simulations directly to continuum electrostatics calculations. Both simulations and continuum calculations show that subnanometer wide pores pose a huge free-energy barrier for ions, but a small increase in the pore diameter to ∼1 nm nearly eliminates that barrier. We also find that in those wider channels the ion mobility is comparable to that in the bulk phase. By calculating local electrostatic potentials, we show that the long range Coulomb interactions of ions are strongly screened in the wide water-filled channels. Whereas continuum calculations capture the overall energetics reasonably well, the local water structure, which is not accounted for in this model, leads to interesting effects such as the preference of hydrated ions to move along the pore wall rather than through the center of the pore. PMID:16006629

[Application of Brownian dynamics to the description of transmembrane ion flow as exemplified by the chloride channel of glycine receptor].

PubMed

Boronovskiĭ, S E; Nartsissov, Ia R

2009-01-01

Using the Brownian dynamics of the movement of hydrated ion in a viscous water solution, a mathematical model has been built, which describes the transport of charged particles through a single protein pore in a lipid membrane. The dependences of transmembrane ion currents on ion concentrations in solution have been obtained. It was shown that, if the geometry of a membrane pore is identical to that of the inner part of the glycine receptor channel and there is no ion selectivity, then the values of both chloride and sodium currents are not greater than 0.5 pA at the physiological concentrations of these ions. If local charge heterogeneity caused by charged amino acid residues of transmembrane protein segments is included into the model calculations, the chloride current increases to about 3.7 pA, which exceeds more than seven times the value for sodium ions under the conditions of the complex channel geometry in the range of physiological concentrations of ions in the solution. The model takes changes in the density of charge distribution both inside the channel and near the protein surface into account. The alteration of pore geometry can be also considered as a parameter at the researcher's option. Thus, the model appears as an effective tool for the description of transmembrane currents for other types of membrane channels.

A common pathway for charge transport through voltage-sensing domains.

PubMed

Chanda, Baron; Bezanilla, Francisco

2008-02-07

Voltage-gated ion channels derive their voltage sensitivity from the movement of specific charged residues in response to a change in transmembrane potential. Several studies on mechanisms of voltage sensing in ion channels support the idea that these gating charges move through a well-defined permeation pathway. This gating pathway in a voltage-gated ion channel can also be mutated to transport free cations, including protons. The recent discovery of proton channels with sequence homology to the voltage-sensing domains suggests that evolution has perhaps exploited the same gating pathway to generate a bona fide voltage-dependent proton transporter. Here we will discuss implications of these findings on the mechanisms underlying charge (and ion) transport by voltage-sensing domains.

Investigation of Ion Channel Activities of Gramicidin A in the Presence of Ionic Liquids Using Model Cell Membranes

PubMed Central

Ryu, Hyunil; Lee, Hwankyu; Iwata, Seigo; Choi, Sangbaek; Ki Kim, Moon; Kim, Young-Rok; Maruta, Shinsaku; Min Kim, Sun; Jeon, Tae-Joon

2015-01-01

Ionic liquids (ILs) are considered to be green solvents because of their non-volatility. Although ILs are relatively safe in the atmospheric environment, they may be toxic in other environments. Our previous research showed that the cytotoxicity of ILs to biological organisms is attributable to interference with cell membranes by IL insertion. However, the effects of ILs on ion channels, which play important roles in cell homeostasis, have not been comprehensively studied to date. In this work, we studied the interactions between ILs and lipid bilayer membranes with gramicidin A ion channels. We used two methods, namely electrical and fluorescence measurements of ions that permeate the membrane. The lifetimes of channels were increased by all the ILs tested in this work via stabilizing the compressed structure of the lipid bilayer and the rate of ion flux through gA channels was decreased by changing the membrane surface charge. The former effect, which increased the rate of ion flux, was dominant at high salt concentrations, whereas the latter, which decreased the rate of ion flux, was dominant at low salt concentrations. The effects of ILs increased with increasing concentration and alkyl chain length. The experimental results were further studied using molecular dynamics simulations. PMID:26189604

Nanobodies that block gating of the P2X7 ion channel ameliorate inflammation.

PubMed

Danquah, Welbeck; Meyer-Schwesinger, Catherine; Rissiek, Björn; Pinto, Carolina; Serracant-Prat, Arnau; Amadi, Miriam; Iacenda, Domenica; Knop, Jan-Hendrik; Hammel, Anna; Bergmann, Philine; Schwarz, Nicole; Assunção, Joana; Rotthier, Wendy; Haag, Friedrich; Tolosa, Eva; Bannas, Peter; Boué-Grabot, Eric; Magnus, Tim; Laeremans, Toon; Stortelers, Catelijne; Koch-Nolte, Friedrich

2016-11-23

Ion channels are desirable therapeutic targets, yet ion channel-directed drugs with high selectivity and few side effects are still needed. Unlike small-molecule inhibitors, antibodies are highly selective for target antigens but mostly fail to antagonize ion channel functions. Nanobodies-small, single-domain antibody fragments-may overcome these problems. P2X7 is a ligand-gated ion channel that, upon sensing adenosine 5'-triphosphate released by damaged cells, initiates a proinflammatory signaling cascade, including release of cytokines, such as interleukin-1β (IL-1β). To further explore its function, we generated and characterized nanobodies against mouse P2X7 that effectively blocked (13A7) or potentiated (14D5) gating of the channel. Systemic injection of nanobody 13A7 in mice blocked P2X7 on T cells and macrophages in vivo and ameliorated experimental glomerulonephritis and allergic contact dermatitis. We also generated nanobody Dano1, which specifically inhibited human P2X7. In endotoxin-treated human blood, Dano1 was 1000 times more potent in preventing IL-1β release than small-molecule P2X7 antagonists currently in clinical development. Our results show that nanobody technology can generate potent, specific therapeutics against ion channels, confirm P2X7 as a therapeutic target for inflammatory disorders, and characterize a potent new drug candidate that targets P2X7. Copyright © 2016, American Association for the Advancement of Science.

The Split Personality of Glutamate Transporters: A Chloride Channel and a Transporter.

PubMed

Cater, Rosemary J; Ryan, Renae M; Vandenberg, Robert J

2016-03-01

Transporters and ion channels are conventionally categorised into distinct classes of membrane proteins. However, some membrane proteins have a split personality and can function as both transporters and ion channels. The excitatory amino acid transporters (EAATs) in particular, function as both glutamate transporters and chloride (Cl(-)) channels. The EAATs couple the transport of glutamate to the co-transport of three Na(+) ions and one H(+) ion into the cell, and the counter-transport of one K(+) ion out of the cell. The EAAT Cl(-) channel is activated by the binding of glutamate and Na(+), but is thermodynamically uncoupled from glutamate transport and involves molecular determinants distinct from those responsible for glutamate transport. Several crystal structures of an EAAT archaeal homologue, GltPh, at different stages of the transport cycle, alongside numerous functional studies and molecular dynamics simulations, have provided extensive insights into the mechanism of substrate transport via these transporters. However, the molecular determinants involved in Cl(-) permeation, and the mechanism by which this channel is activated are not entirely understood. Here we will discuss what is currently known about the molecular determinants involved in EAAT-mediated Cl(-) permeation and the mechanisms that underlie their split personality.

Current-voltage characteristics influenced by the nanochannel diameter and surface charge density in a fluidic field-effect-transistor.

PubMed

Singh, Kunwar Pal; Guo, Chunlei

2017-06-21

The nanochannel diameter and surface charge density have a significant impact on current-voltage characteristics in a nanofluidic transistor. We have simulated the effect of the channel diameter and surface charge density on current-voltage characteristics of a fluidic nanochannel with positive surface charge on its walls and a gate electrode on its surface. Anion depletion/enrichment leads to a decrease/increase in ion current with gate potential. The ion current tends to increase linearly with gate potential for narrow channels at high surface charge densities and narrow channels are more effective to control the ion current at high surface charge densities. The current-voltage characteristics are highly nonlinear for wide channels at low surface charge densities and they show different regions of current change with gate potential. The ion current decreases with gate potential after attaining a peak value for wide channels at low values of surface charge densities. At low surface charge densities, the ion current can be controlled by a narrow range of gate potentials for wide channels. The current change with source drain voltage shows ohmic, limiting and overlimiting regions.

A parallel finite element simulator for ion transport through three-dimensional ion channel systems.

PubMed

Tu, Bin; Chen, Minxin; Xie, Yan; Zhang, Linbo; Eisenberg, Bob; Lu, Benzhuo

2013-09-15

A parallel finite element simulator, ichannel, is developed for ion transport through three-dimensional ion channel systems that consist of protein and membrane. The coordinates of heavy atoms of the protein are taken from the Protein Data Bank and the membrane is represented as a slab. The simulator contains two components: a parallel adaptive finite element solver for a set of Poisson-Nernst-Planck (PNP) equations that describe the electrodiffusion process of ion transport, and a mesh generation tool chain for ion channel systems, which is an essential component for the finite element computations. The finite element method has advantages in modeling irregular geometries and complex boundary conditions. We have built a tool chain to get the surface and volume mesh for ion channel systems, which consists of a set of mesh generation tools. The adaptive finite element solver in our simulator is implemented using the parallel adaptive finite element package Parallel Hierarchical Grid (PHG) developed by one of the authors, which provides the capability of doing large scale parallel computations with high parallel efficiency and the flexibility of choosing high order elements to achieve high order accuracy. The simulator is applied to a real transmembrane protein, the gramicidin A (gA) channel protein, to calculate the electrostatic potential, ion concentrations and I - V curve, with which both primitive and transformed PNP equations are studied and their numerical performances are compared. To further validate the method, we also apply the simulator to two other ion channel systems, the voltage dependent anion channel (VDAC) and α-Hemolysin (α-HL). The simulation results agree well with Brownian dynamics (BD) simulation results and experimental results. Moreover, because ionic finite size effects can be included in PNP model now, we also perform simulations using a size-modified PNP (SMPNP) model on VDAC and α-HL. It is shown that the size effects in SMPNP can effectively lead to reduced current in the channel, and the results are closer to BD simulation results. Copyright © 2013 Wiley Periodicals, Inc.

Role of TRP ion channels in cancer and tumorigenesis.

PubMed

Shapovalov, George; Ritaine, Abigael; Skryma, Roman; Prevarskaya, Natalia

2016-05-01

Transient receptor potential (TRP) channels are recently identified proteins that form a versatile family of ion channels, the majority of which are calcium permeable and exhibit complex regulatory patterns with sensitivity to multiple environmental factors. While this sensitivity has captured early attention, leading to recognition of TRP channels as environmental and chemical sensors, many later studies concentrated on the regulation of intracellular calcium by TRP channels. Due to mutations, dysregulation of ion channel gating or expression levels, normal spatiotemporal patterns of local Ca(2+) distribution become distorted. This causes deregulation of downstream effectors sensitive to changes in Ca(2+) homeostasis that, in turn, promotes pathophysiological cancer hallmarks, such as enhanced survival, proliferation and invasion. These observations give rise to the appreciation of the important contributions that TRP channels make to many cellular processes controlling cell fate and positioning these channels as important players in cancer regulation. This review discusses the accumulated scientific knowledge focused on TRP channel involvement in regulation of cell fate in various transformed tissues.

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

Structure of a CLC chloride ion channel by cryo-electron microscopy

PubMed Central

Park, Eunyong; Campbell, Ernest B.; MacKinnon, Roderick

2017-01-01

CLC proteins transport chloride (Cl−) ions across cellular membranes to regulate muscle excitability, electrolyte movement across epithelia, and acidification of intracellular organelles. Some CLC proteins are channels that conduct Cl− ions passively, whereas others are secondary active transporters that exchange two Cl− ions for one H+. The structural basis underlying these distinctive transport mechanisms is puzzling because CLC channels and transporters are expected to share the same architecture based on sequence homology. To solve this puzzle we determined the structure of a mammalian CLC channel (CLC-K) using cryo-electron microscopy. A conserved loop in the Cl− transport pathway shows a structure markedly different from that of CLC transporters. Consequently, the cytosolic constriction for Cl− passage is widened in CLC-K such that the kinetic barrier previously postulated for Cl−/H+ transporter function would be reduced. Thus, reduction of a kinetic barrier in CLC channels enables fast flow of Cl− down its electrochemical gradient. PMID:28002411

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

The ClC-0 chloride channel is a ‘broken’ Cl−/H+ antiporter

PubMed Central

Lísal, Jiří; Maduke, Merritt

2008-01-01

Ion channels have historically been viewed as distinct from secondary active transporters. However, the recent discovery that the CLC ‘chloride channel’ family is made up of both channels and active transporters has led to the hypothesis that the ion-transport mechanisms of these two types of membrane proteins may be similar. Here we use single-channel analysis to demonstrate that ClC-0 channel gating (opening and closing) involves the transmembrane movement of protons. This result indicates that ClC-0 is a ‘broken’ Cl−/H+ antiporter in which one of the conformational states has become leaky for chloride ions. This finding clarifies the evolutionary relationship between the channels and transporters and conveys that similar mechanisms and analogous protein movements are used by both. PMID:18641661

Understanding the conductive channel evolution in Na:WO(3-x)-based planar devices.

PubMed

Shang, Dashan; Li, Peining; Wang, Tao; Carria, Egidio; Sun, Jirong; Shen, Baogen; Taubner, Thomas; Valov, Ilia; Waser, Rainer; Wuttig, Matthias

2015-04-14

An ion migration process in a solid electrolyte is important for ion-based functional devices, such as fuel cells, batteries, electrochromics, gas sensors, and resistive switching systems. In this study, a planar sandwich structure is prepared by depositing tungsten oxide (WO(3-x)) films on a soda-lime glass substrate, from which Na(+) diffuses into the WO(3-x) films during the deposition. The entire process of Na(+) migration driven by an alternating electric field is visualized in the Na-doped WO(3-x) films in the form of conductive channel by in situ optical imaging combined with infrared spectroscopy and near-field imaging techniques. A reversible change of geometry between a parabolic and a bar channel is observed with the resistance change of the devices. The peculiar channel evolution is interpreted by a thermal-stress-induced mechanical deformation of the films and an asymmetric Na(+) mobility between the parabolic and the bar channels. These results exemplify a typical ion migration process driven by an alternating electric field in a solid electrolyte with a low ion mobility and are expected to be beneficial to improve the controllability of the ion migration in ion-based functional devices, such as resistive switching devices.

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.

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.

Mass-dependent channel electron multiplier operation. [for ion detection

NASA Technical Reports Server (NTRS)

Fields, S. A.; Burch, J. L.; Oran, W. A.

1977-01-01

The absolute counting efficiency and pulse height distributions of a continuous-channel electron multiplier used in the detection of hydrogen, argon and xenon ions are assessed. The assessment technique, which involves the post-acceleration of 8-eV ion beams to energies from 100 to 4000 eV, provides information on counting efficiency versus post-acceleration voltage characteristics over a wide range of ion mass. The charge pulse height distributions for H2 (+), A (+) and Xe (+) were measured by operating the experimental apparatus in a marginally gain-saturated mode. It was found that gain saturation occurs at lower channel multiplier operating voltages for light ions such as H2 (+) than for the heavier ions A (+) and Xe (+), suggesting that the technique may be used to discriminate between these two classes of ions in electrostatic analyzers.

Hysteresis in voltage-gated channels.

PubMed

Villalba-Galea, Carlos A

2017-03-04

Ion channels constitute a superfamily of membrane proteins found in all living creatures. Their activity allows fast translocation of ions across the plasma membrane down the ion's transmembrane electrochemical gradient, resulting in a difference in electrical potential across the plasma membrane, known as the membrane potential. A group within this superfamily, namely voltage-gated channels, displays activity that is sensitive to the membrane potential. The activity of voltage-gated channels is controlled by the membrane potential, while the membrane potential is changed by these channels' activity. This interplay produces variations in the membrane potential that have evolved into electrical signals in many organisms. These signals are essential for numerous biological processes, including neuronal activity, insulin release, muscle contraction, fertilization and many others. In recent years, the activity of the voltage-gated channels has been observed not to follow a simple relationship with the membrane potential. Instead, it has been shown that the activity of voltage-gated channel displays hysteresis. In fact, a growing number of evidence have demonstrated that the voltage dependence of channel activity is dynamically modulated by activity itself. In spite of the great impact that this property can have on electrical signaling, hysteresis in voltage-gated channels is often overlooked. Addressing this issue, this review provides examples of voltage-gated ion channels displaying hysteretic behavior. Further, this review will discuss how Dynamic Voltage Dependence in voltage-gated channels can have a physiological role in electrical signaling. Furthermore, this review will elaborate on the current thoughts on the mechanism underlying hysteresis in voltage-gated channels.

Artificial transmembrane ion channels from self-assembling peptide nanotubes

NASA Astrophysics Data System (ADS)

Ghadiri, M. Reza; Granja, Juan R.; Buehler, Lukas K.

1994-05-01

NATURALLY occurring membrane channels and pores are formed from a large family of diverse proteins, peptides and organic secon-dary metabolites whose vital biological functions include control of ion flow, signal transduction, molecular transport and produc-tion of cellular toxins. But despite the availability of a large amount of biochemical information about these molecules1, the design and synthesis of artificial systems that can mimic the bio-logical function of natural compounds remains a formidable task2-12. Here we present a simple strategy for the design of artifi-cial membrane ion channels based on a self-assembled cylindrical β-sheet peptide architecture13. Our systems-essentially stacks of peptide rings-display good channel-mediated ion-transport activ-ity with rates exceeding 107 ions s-1, rivalling the performance of many naturally occurring counterparts. Such molecular assemblies should find use in the design of novel cytotoxic agents, membrane transport vehicles and drug-delivery systems.

Atomistic Detailed Mechanism and Weak Cation-Conducting Activity of HIV-1 Vpu Revealed by Free Energy Calculations

PubMed Central

Padhi, Siladitya; Burri, Raghunadha Reddy; Jameel, Shahid; Priyakumar, U. Deva

2014-01-01

The viral protein U (Vpu) encoded by HIV-1 has been shown to assist in the detachment of virion particles from infected cells. Vpu forms cation-specific ion channels in host cells, and has been proposed as a potential drug target. An understanding of the mechanism of ion transport through Vpu is desirable, but remains limited because of the unavailability of an experimental structure of the channel. Using a structure of the pentameric form of Vpu – modeled and validated based on available experimental data – umbrella sampling molecular dynamics simulations (cumulative simulation time of more than 0.4 µs) were employed to elucidate the energetics and the molecular mechanism of ion transport in Vpu. Free energy profiles corresponding to the permeation of Na+ and K+ were found to be similar to each other indicating lack of ion selection, consistent with previous experimental studies. The Ser23 residue is shown to enhance ion transport via two mechanisms: creating a weak binding site, and increasing the effective hydrophilic length of the channel, both of which have previously been hypothesized in experiments. A two-dimensional free energy landscape has been computed to model multiple ion permeation, based on which a mechanism for ion conduction is proposed. It is shown that only one ion can pass through the channel at a time. This, along with a stretch of hydrophobic residues in the transmembrane domain of Vpu, explains the slow kinetics of ion conduction. The results are consistent with previous conductance studies that showed Vpu to be a weakly conducting ion channel. PMID:25392993

Electric Dipolar Kondo Effect Emerging from a Vibrating Magnetic Ion

NASA Astrophysics Data System (ADS)

Hotta, Takashi; Ueda, Kazuo

2012-06-01

When a magnetic ion vibrates in a metal, it inevitably introduces a new channel of hybridization with conduction electrons, and in general, the vibrating ion induces an electric dipole moment. In such a situation, we find that magnetic and nonmagnetic Kondo effects alternatively occur due to the screening of the spin moment and electric dipole moment of the vibrating ion. In particular, the electric dipolar two-channel Kondo effect is found to occur for a weak Coulomb interaction. We also show that a magnetically robust heavy-electron state appears near the fixed point of the electric dipolar two-channel Kondo effect. We believe that the vibrating magnetic ion opens a new door in Kondo physics.

Fragile X mental retardation protein controls synaptic vesicle exocytosis by modulating N-type calcium channel density

NASA Astrophysics Data System (ADS)

Ferron, Laurent; Nieto-Rostro, Manuela; Cassidy, John S.; Dolphin, Annette C.

2014-04-01

Fragile X syndrome (FXS), the most common heritable form of mental retardation, is characterized by synaptic dysfunction. Synaptic transmission depends critically on presynaptic calcium entry via voltage-gated calcium (CaV) channels. Here we show that the functional expression of neuronal N-type CaV channels (CaV2.2) is regulated by fragile X mental retardation protein (FMRP). We find that FMRP knockdown in dorsal root ganglion neurons increases CaV channel density in somata and in presynaptic terminals. We then show that FMRP controls CaV2.2 surface expression by targeting the channels to the proteasome for degradation. The interaction between FMRP and CaV2.2 occurs between the carboxy-terminal domain of FMRP and domains of CaV2.2 known to interact with the neurotransmitter release machinery. Finally, we show that FMRP controls synaptic exocytosis via CaV2.2 channels. Our data indicate that FMRP is a potent regulator of presynaptic activity, and its loss is likely to contribute to synaptic dysfunction in FXS.

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

Moczydlowski, Edward G.

Ion channel proteins regulate complex patterns of cellular electrical activity and ionic signaling. Certain K+ channels play an important role in immunological biodefense mechanisms of adaptive and innate immunity. Most ion channel proteins are oligomeric complexes with the conductive pore located at the central subunit interface. The long-term activity of many K+ channel proteins is dependent on the concentration of extracellular K+; however, the mechanism is unclear. Thus, this project focused on mechanisms underlying structural stability of tetrameric K+ channels. Using KcsA of Streptomyces lividans as a model K+ channel of known structure, the molecular basis of tetramer stability wasmore » investigated by: 1. Bioinformatic analysis of the tetramer interface. 2. Effect of two local anesthetics (lidocaine, tetracaine) on tetramer stability. 3. Molecular simulation of drug docking to the ion conduction pore. The results provide new insights regarding the structural stability of K+ channels and its possible role in cell physiology.« less

On the simple random-walk models of ion-channel gate dynamics reflecting long-term memory.

PubMed

Wawrzkiewicz, Agata; Pawelek, Krzysztof; Borys, Przemyslaw; Dworakowska, Beata; Grzywna, Zbigniew J

2012-06-01

Several approaches to ion-channel gating modelling have been proposed. Although many models describe the dwell-time distributions correctly, they are incapable of predicting and explaining the long-term correlations between the lengths of adjacent openings and closings of a channel. In this paper we propose two simple random-walk models of the gating dynamics of voltage and Ca(2+)-activated potassium channels which qualitatively reproduce the dwell-time distributions, and describe the experimentally observed long-term memory quite well. Biological interpretation of both models is presented. In particular, the origin of the correlations is associated with fluctuations of channel mass density. The long-term memory effect, as measured by Hurst R/S analysis of experimental single-channel patch-clamp recordings, is close to the behaviour predicted by our models. The flexibility of the models enables their use as templates for other types of ion channel.

Fusion of Night Vision and Thermal Images

DTIC Science & Technology

2006-12-01

with the walls of the MCP channels. Thus, a thin metal oxide coating commonly known as an ion barrier film is added to the input side of the MCP to...with film ion barrier to filmless gated tubes. An important improvement for Gen 4 products is a greater target identification range and higher target...Metal Seals with S-25 Cathode Mircro-channel plate Ceramic/Metal Seals with GaAS Cathode Mircro-channel plate with ion barrier film Ceramic

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.

Energetics of discrete selectivity bands and mutation-induced transitions in the calcium-sodium ion channels family.

PubMed

Kaufman, I; Luchinsky, D G; Tindjong, R; McClintock, P V E; Eisenberg, R S

2013-11-01

We use Brownian dynamics (BD) simulations to study the ionic conduction and valence selectivity of a generic electrostatic model of a biological ion channel as functions of the fixed charge Q(f) at its selectivity filter. We are thus able to reconcile the discrete calcium conduction bands recently revealed in our BD simulations, M0 (Q(f)=1e), M1 (3e), M2 (5e), with a set of sodium conduction bands L0 (0.5e), L1 (1.5e), thereby obtaining a completed pattern of conduction and selectivity bands vs Q(f) for the sodium-calcium channels family. An increase of Q(f) leads to an increase of calcium selectivity: L0 (sodium-selective, nonblocking channel) → M0 (nonselective channel) → L1 (sodium-selective channel with divalent block) → M1 (calcium-selective channel exhibiting the anomalous mole fraction effect). We create a consistent identification scheme where the L0 band is putatively identified with the eukaryotic sodium channel The scheme created is able to account for the experimentally observed mutation-induced transformations between nonselective channels, sodium-selective channels, and calcium-selective channels, which we interpret as transitions between different rows of the identification table. By considering the potential energy changes during permeation, we show explicitly that the multi-ion conduction bands of calcium and sodium channels arise as the result of resonant barrierless conduction. The pattern of periodic conduction bands is explained on the basis of sequential neutralization taking account of self-energy, as Q(f)(z,i)=ze(1/2+i), where i is the order of the band and z is the valence of the ion. Our results confirm the crucial influence of electrostatic interactions on conduction and on the Ca(2+)/Na(+) valence selectivity of calcium and sodium ion channels. The model and results could be also applicable to biomimetic nanopores with charged walls.

Energetics of discrete selectivity bands and mutation-induced transitions in the calcium-sodium ion channels family

NASA Astrophysics Data System (ADS)

Kaufman, I.; Luchinsky, D. G.; Tindjong, R.; McClintock, P. V. E.; Eisenberg, R. S.

2013-11-01

We use Brownian dynamics (BD) simulations to study the ionic conduction and valence selectivity of a generic electrostatic model of a biological ion channel as functions of the fixed charge Qf at its selectivity filter. We are thus able to reconcile the discrete calcium conduction bands recently revealed in our BD simulations, M0 (Qf=1e), M1 (3e), M2 (5e), with a set of sodium conduction bands L0 (0.5e), L1 (1.5e), thereby obtaining a completed pattern of conduction and selectivity bands vs Qf for the sodium-calcium channels family. An increase of Qf leads to an increase of calcium selectivity: L0 (sodium-selective, nonblocking channel) → M0 (nonselective channel) → L1 (sodium-selective channel with divalent block) → M1 (calcium-selective channel exhibiting the anomalous mole fraction effect). We create a consistent identification scheme where the L0 band is putatively identified with the eukaryotic sodium channel The scheme created is able to account for the experimentally observed mutation-induced transformations between nonselective channels, sodium-selective channels, and calcium-selective channels, which we interpret as transitions between different rows of the identification table. By considering the potential energy changes during permeation, we show explicitly that the multi-ion conduction bands of calcium and sodium channels arise as the result of resonant barrierless conduction. The pattern of periodic conduction bands is explained on the basis of sequential neutralization taking account of self-energy, as Qf(z,i)=ze(1/2+i), where i is the order of the band and z is the valence of the ion. Our results confirm the crucial influence of electrostatic interactions on conduction and on the Ca2+/Na+ valence selectivity of calcium and sodium ion channels. The model and results could be also applicable to biomimetic nanopores with charged walls.

X-ray Synchrotron Radiation in a Plasma Wiggler

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

Wang, Shuoquin; /UCLA /SLAC, SSRL

2005-09-27

A relativistic electron beam can radiate due to its betatron motion inside an ion channel. The ion channel is induced by the electron bunch as it propagates through an underdense plasma. In the theory section of this thesis the formation of the ion channel, the trajectories of beam electrons inside the ion channel, the radiation power and the radiation spectrum of the spontaneous emission are studied. The comparison between different plasma wiggler schemes is made. The difficulties in realizing stimulated emission as the beam traverses the ion channel are investigated, with particular emphasis on the bunching mechanism, which is importantmore » for the ion channel free electron laser. This thesis reports an experiment conducted at the Stanford Linear Accelerator Center (SLAC) to measure the betatron X-ray radiations for the first time. They first describe the construction and characterization of the lithium plasma source. In the experiment, the transverse oscillations of the SLAC 28.5 GeV electron beam traversing through a 1.4 meter long lithium plasma source are clearly seen. These oscillations lead to a quadratic density dependence of the spontaneously emitted betatron X-ray radiation. The divergence angle of the X-ray radiation is measured. The absolute photon yield and the spectral brightness at 14.2 KeV photon energy are estimated and seen to be in reasonable agreement with theory.« less

Acid-sensing ion channels and transient-receptor potential ion channels in zebrafish taste buds.

PubMed

Levanti, M; Randazzo, B; Viña, E; Montalbano, G; Garcia-Suarez, O; Germanà, A; Vega, J A; Abbate, F

2016-09-01

Sensory information from the environment is required for life and survival, and it is detected by specialized cells which together make up the sensory system. The fish sensory system includes specialized organs that are able to detect mechanical and chemical stimuli. In particular, taste buds are small organs located on the tongue in terrestrial vertebrates that function in the perception of taste. In fish, taste buds occur on the lips, the flanks, and the caudal (tail) fins of some species and on the barbels of others. In fish taste receptor cells, different classes of ion channels have been detected which, like in mammals, presumably participate in the detection and/or transduction of chemical gustatory signals. However, since some of these ion channels are involved in the detection of additional sensory modalities, it can be hypothesized that taste cells sense stimuli other than those specific for taste. This mini-review summarizes current knowledge on the presence of transient-receptor potential (TRP) and acid-sensing (ASIC) ion channels in the taste buds of teleosts, especially adult zebrafish. Up to now ASIC4, TRPC2, TRPA1, TRPV1 and TRPV4 ion channels have been found in the sensory cells, while ASIC2 was detected in the nerves supplying the taste buds. Copyright © 2016 Elsevier GmbH. All rights reserved.

Non-ionotropic signaling by the NMDA receptor: controversy and opportunity.

PubMed

Gray, John A; Zito, Karen; Hell, Johannes W

2016-01-01

Provocative emerging evidence suggests that the N-methyl-d-aspartate (NMDA) receptor can signal in the absence of ion flux through the receptor. This non-ionotropic signaling is thought to be due to agonist-induced conformational changes in the receptor, independently of channel opening. Non-ionotropic NMDA receptor signaling has been proposed to be sufficient to induce synaptic long-term depression (LTD), directly challenging the decades-old model that prolonged low-level calcium influx is required to induce LTD. Here, we briefly review these recent findings, focusing primarily on the potential role of non-ionotropic signaling in NMDA receptor-mediated LTD. Further reports concerning additional roles of non-ionotropic NMDA receptor signaling are also discussed. If validated, this new view of NMDA receptor-mediated signaling will usher in an exciting new era of exploring synapse function and dysfunction.

Non-ionotropic signaling by the NMDA receptor: controversy and opportunity

PubMed Central

Gray, John A.; Zito, Karen; Hell, Johannes W.

2016-01-01

Provocative emerging evidence suggests that the N-methyl-d-aspartate (NMDA) receptor can signal in the absence of ion flux through the receptor. This non-ionotropic signaling is thought to be due to agonist-induced conformational changes in the receptor, independently of channel opening. Non-ionotropic NMDA receptor signaling has been proposed to be sufficient to induce synaptic long-term depression (LTD), directly challenging the decades-old model that prolonged low-level calcium influx is required to induce LTD. Here, we briefly review these recent findings, focusing primarily on the potential role of non-ionotropic signaling in NMDA receptor-mediated LTD. Further reports concerning additional roles of non-ionotropic NMDA receptor signaling are also discussed. If validated, this new view of NMDA receptor-mediated signaling will usher in an exciting new era of exploring synapse function and dysfunction. PMID:27303637

Sickle cell anemia mice develop a unique cardiomyopathy with restrictive physiology

PubMed Central

Bakeer, Nihal; James, Jeanne; Roy, Swarnava; Wansapura, Janaka; Shanmukhappa, Shiva Kumar; Lorenz, John N.; Osinska, Hanna; Backer, Kurt; Huby, Anne-Cecile; Shrestha, Archana; Niss, Omar; Fleck, Robert; Quinn, Charles T.; Taylor, Michael D.; Purevjav, Enkhsaikhan; Aronow, Bruce J.; Towbin, Jeffrey A.; Malik, Punam

2016-01-01

Cardiopulmonary complications are the leading cause of mortality in sickle cell anemia (SCA). Elevated tricuspid regurgitant jet velocity, pulmonary hypertension, diastolic, and autonomic dysfunction have all been described, but a unifying pathophysiology and mechanism explaining the poor prognosis and propensity to sudden death has been elusive. Herein, SCA mice underwent a longitudinal comprehensive cardiac analysis, combining state-of-the-art cardiac imaging with electrocardiography, histopathology, and molecular analysis to determine the basis of cardiac dysfunction. We show that in SCA mice, anemia-induced hyperdynamic physiology was gradually superimposed with restrictive physiology, characterized by progressive left atrial enlargement and diastolic dysfunction with preserved systolic function. This phenomenon was absent in WT mice with experimentally induced chronic anemia of similar degree and duration. Restrictive physiology was associated with microscopic cardiomyocyte loss and secondary fibrosis detectable as increased extracellular volume by cardiac-MRI. Ultrastructural mitochondrial changes were consistent with severe chronic hypoxia/ischemia and sarcomere diastolic-length was shortened. Transcriptome analysis revealed up-regulation of genes involving angiogenesis, extracellular-matrix, circadian-rhythm, oxidative stress, and hypoxia, whereas ion-channel transport and cardiac conduction were down-regulated. Indeed, progressive corrected QT prolongation, arrhythmias, and ischemic changes were noted in SCA mice before sudden death. Sudden cardiac death is common in humans with restrictive cardiomyopathies and long QT syndromes. Our findings may thus provide a unifying cardiac pathophysiology that explains the reported cardiac abnormalities and sudden death seen in humans with SCA. PMID:27503873

Potential therapeutic targets for ATP-gated P2X receptor ion channels.

PubMed

Li, Zhiyuan; Liang, Dong; Chen, Ling

2008-04-01

P2X receptors make up a novel family of ligand-gated ion channels that are activated by binding of extracellular ATP. These receptors can form a number of homomeric and heteromeric ion channels, which are widely distributed throughout the human body. They are thought to play an important role in many cellular processes, including synaptic transmission and thrombocyte aggregation. These ion channels are also involved in the pathology of several disease states, including chronic inflammation and neuropathic pain, and thus are the potential targets for drug development. The recent discovery of potent and highly selective antagonists for P2X(7) receptors, through the use of high-throughput screening, has helped to further understand the P2X receptor pharmacology and provided new evidence that P2X(7) receptors play a specific role in chronic pain states. In this review, we discuss how the P2X family of ion channels has distinguished itself as a potential new drug target. We are optimistic that safe and effective candidate drugs will be suitable for progression into clinical development.

A Component-Based FPGA Design Framework for Neuronal Ion Channel Dynamics Simulations

PubMed Central

Mak, Terrence S. T.; Rachmuth, Guy; Lam, Kai-Pui; Poon, Chi-Sang

2008-01-01

Neuron-machine interfaces such as dynamic clamp and brain-implantable neuroprosthetic devices require real-time simulations of neuronal ion channel dynamics. Field Programmable Gate Array (FPGA) has emerged as a high-speed digital platform ideal for such application-specific computations. We propose an efficient and flexible component-based FPGA design framework for neuronal ion channel dynamics simulations, which overcomes certain limitations of the recently proposed memory-based approach. A parallel processing strategy is used to minimize computational delay, and a hardware-efficient factoring approach for calculating exponential and division functions in neuronal ion channel models is used to conserve resource consumption. Performances of the various FPGA design approaches are compared theoretically and experimentally in corresponding implementations of the AMPA and NMDA synaptic ion channel models. Our results suggest that the component-based design framework provides a more memory economic solution as well as more efficient logic utilization for large word lengths, whereas the memory-based approach may be suitable for time-critical applications where a higher throughput rate is desired. PMID:17190033

TMEM16 proteins: unknown structure and confusing functions.

PubMed

Picollo, Alessandra; Malvezzi, Mattia; Accardi, Alessio

2015-01-16

The TMEM16 family of membrane proteins, also known as anoctamins, plays key roles in a variety of physiological functions that range from ion transport to phospholipid scrambling and to regulating other ion channels. The first two family members to be functionally characterized, TMEM16A (ANO1) and TMEM16B (ANO2), form Ca(2+)-activated Cl(-) channels and are important for transepithelial ion transport, olfaction, phototransduction, smooth muscle contraction, nociception, cell proliferation and control of neuronal excitability. The roles of other family members, such as TMEM16C (ANO3), TMEM16D (ANO4), TMEM16F (ANO6), TMEM16G (ANO7) and TMEM16J (ANO9), remain poorly understood and controversial. These homologues were reported to be phospholipid scramblases, ion channels, to have both functions or to be regulatory subunits of other channels. Mutations in TMEM16F cause Scott syndrome, a bleeding disorder caused by impaired Ca(2+)-dependent externalization of phosphatidylserine in activated platelets, suggesting that this homologue might be a scramblase. However, overexpression of TMEM16F has also been associated with a remarkable number of different ion channel types, raising the possibility that this protein might be involved in both ion and lipid transports. The recent identification of an ancestral TMEM16 homologue with intrinsic channel and scramblase activities supports this hypothesis. Thus, the TMEM16 family might have diverged in two or three different subclasses, channels, scramblases and dual-function channel/scramblases. The structural bases and functional implication of such a functional diversity within a single protein family remain to be elucidated and the links between TMEM16 functions and human physiology and pathologies need to be investigated. Copyright © 2014 Elsevier Ltd. All rights reserved.

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.

Anion-Cation Permeability Correlates with Hydrated Counterion Size in Glycine Receptor Channels

PubMed Central

Sugiharto, Silas; Lewis, Trevor M.; Moorhouse, Andrew J.; Schofield, Peter R.; Barry, Peter H.

2008-01-01

The functional role of ligand-gated ion channels depends critically on whether they are predominantly permeable to cations or anions. However, these, and other ion channels, are not perfectly selective, allowing some counterions to also permeate. To address the mechanisms by which such counterion permeation occurs, we measured the anion-cation permeabilities of different alkali cations, Li+ Na+, and Cs+, relative to either Cl− or \\documentclass[10pt]{article} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{pmc} \\usepackage[Euler]{upgreek} \\pagestyle{empty} \\oddsidemargin -1.0in \\begin{document} \\begin{equation*}{\\mathrm{NO}}_{3}^{-}\\end{equation*}\\end{document} anions in both a wild-type glycine receptor channel (GlyR) and a mutant GlyR with a wider pore diameter. We hypothesized and showed that counterion permeation in anionic channels correlated inversely with an equivalent or effective hydrated size of the cation relative to the channel pore radius, with larger counterion permeabilities being observed in the wider pore channel. We also showed that the anion component of conductance was independent of the nature of the cation. We suggest that anions and counterion cations can permeate through the pore as neutral ion pairs, to allow the cations to overcome the large energy barriers resulting from the positively charged selectivity filter in small GlyR channels, with the permeability of such ion pairs being dependent on the effective hydrated diameter of the ion pair relative to the pore diameter. PMID:18708455

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

Transport proteins of the plant plasma membrane

NASA Technical Reports Server (NTRS)

Assmann, S. M.; Haubrick, L. L.; Evans, M. L. (Principal Investigator)

1996-01-01

Recently developed molecular and genetic approaches have enabled the identification and functional characterization of novel genes encoding ion channels, ion carriers, and water channels of the plant plasma membrane.

Properties of an inward rectifying K channel in the membrane of guinea-pig atrial cardioballs.

PubMed

Bechem, M; Glitsch, H G; Pott, L

1983-11-01

Single channel outward current fluctuations are recorded in excised (outside-out) membrane patches of isolated atrial cells in culture (cardioballs) from hearts of adult guinea-pigs. The ionic channel displays a high selectivity to K ions. Accordingly the reversal potential of the single channel current is close to the K equilibrium potential. The open channel conductance is unaffected by the membrane potential but depends on the K concentration of the outside solution (19.7pS at 2 mM Ko to 30.7pS at 20 mM Ko). The open state probability (Po) of the channel shows a marked voltage dependence. Po amounts to c.0.9 at -40 mV and decreases to c.0.1 at +40 mV. Under the assumption of no channel interaction a macroscopic steady state current voltage relationship is reconstructed from the single channel data. The relationship displays inward-going rectification. The rectification is due to the voltage dependence of Po. The I-V curve displays a negative slope at membrane potentials positive to -15 mV. In bathing solutions containing Ba ions (0.2 mM) Po is reduced by rapid closures which interrupt the open state events. The unit channel conductance is unaffected by Ba ions. The channel block exerted by Ba ions is augmented with increasing membrane hyperpolarization. The results suggest that the channel studied may represent a background K conductance.

The Earliest Ion Channels in Protocellular Membranes

NASA Technical Reports Server (NTRS)

Mijajlovic, Milan; Pohorille, Andrew; Wilson, Michael; Wei, Chenyu

2010-01-01

Cellular membranes with their hydrophobic interior are virtually impermeable to ions. Bulk of ion transport through them is enabled through ion channels. Ion channels of contemporary cells are complex protein molecules which span the membrane creating a cylindrical pore filled with water. Protocells, which are widely regarded as precursors to modern cells, had similarly impermeable membranes, but the set of proteins in their disposal was much simpler and more limited. We have been, therefore, exploring an idea that the first ion channels in protocellular membranes were formed by much smaller peptide molecules that could spontaneously selfassemble into short-lived cylindrical bundles in a membrane. Earlier studies have shown that a group of peptides known as peptaibols is capable of forming ion channels in lipid bilayers when they are exposed to an electric field. Peptaibols are small, non-genetically encoded peptides produced by some fungi as a part of their system of defense against bacteria. They are usually only 14-20 residues long, which is just enough to span the membrane. Their sequence is characterized by the presence of non-standard amino acids which, interestingly, are also expected to have existed on the early earth. In particular, the presence of 2-aminoisobutyric acid (AIB) gives peptaibols strong helix forming propensities. Association of the helices inside membranes leads to the formation of cylindrical bundles, typically containing 4 to 10 monomers. Although peptaibols are excellent candidates for models of the earliest ion channels their structures, which are stabilized only by van der Waals forces and occasional hydrogen bonds between neighboring helices, are not very stable. Although it might properly reflect protobiological reality, it is also a major obstacle in studying channel behavior. For this reason we focused on two members of the peptaibol family, trichotoxin and antiamoebin, which are characterized by a single conductance level. This indicates that their structures are unique and stable. In addition, it is also believed that the trichotoxin channel displays some selectivity between potassium and chloride ions. This makes trichotoxin and antiamoebin ideal models of the earliest ion channels that could provide insight into the origins of ion conductance and selectivity. In the absence of crystal structure of the trichotoxin and antiamoebin channels, we propose their molecular models based on experimentally determined number of monomers forming the bundles. We use molecular dynamics simulations to validate the models in terms of their conductance and selectivity. On the basis of our simulations we show that the emergence of channels built of small, alpha-helical peptides was protobiologically plausible and did not require highly specific amino acid sequences, which is a convenient evolutionary trait. Despite their simple structure, such channels could possess properties that, at the first sight, appear to require markedly larger complexity. To this end, we will discuss how the amino acid sequence and structure of primitive channels give rise to the phenomena of ionic conductance and selectivity across the earliest cell walls, which were essential functions for the emergence and early evolution of protocells. Furthermore, we will argue that even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during evolution.

The earliest ion channels in protocellular membranes

NASA Astrophysics Data System (ADS)

Mijajlovic, Milan; Pohorille, Andrew; Wilson, Michael; Wei, Chenyu

Cellular membranes with their hydrophobic interior are virtually impermeable to ions. Bulk of ion transport through them is enabled through ion channels. Ion channels of contemporary cells are complex protein molecules which span the membrane creating a cylindrical pore filled with water. Protocells, which are widely regarded as precursors to modern cells, had similarly impermeable membranes, but the set of proteins in their disposal was much simpler and more limited. We have been, therefore, exploring an idea that the first ion channels in protocellular membranes were formed by much smaller peptide molecules that could spontaneously self-assemble into short-lived cylindrical bundles in a membrane. Earlier studies have shown that a group of peptides known as peptaibols is capable of forming ion channels in lipid bilayers when they are exposed to an electric field. Peptaibols are small, non-genetically encoded peptides produced by some fungi as a part of their system of defense against bacteria. They are usually only 14-20 residues long, which is just enough to span the membrane. Their sequence is characterized by the presence of non-standard amino acids which, interestingly, are also expected to have existed on the early earth. In particular, the presence of 2-aminoisobutyric acid (AIB) gives peptaibols strong helix forming propensities. Association of the helices inside membranes leads to the formation of cylindrical bundles, typically containing 4 to 10 monomers. Although peptaibols are excellent candidates for models of the earliest ion channels their struc-tures, which are stabilized only by van der Waals forces and occasional hydrogen bonds between neighboring helices, are not very stable. Although it might properly reflect protobiological real-ity, it is also a major obstacle in studying channel behavior. For this reason we focused on two members of the peptaibol family, trichotoxin and antiamoebin, which are characterized by a single conductance level. This indicates that their structures are unique and stable. In addition, it is also believed that the trichotoxin channel displays some selectivity between potassium and chloride ions. This makes trichotoxin and antiamoebin ideal models of the earliest ion channels that could provide insight into the origins of ion conductance and selectivity. In the absence of crystal structure of the trichotoxin and antiamoebin channels, we propose their molecular models based on experimentally determined number of monomers forming the bundles. We use molecular dynamics simulations to validate the models in terms of their conductance and selectivity. On the basis of our simulations we show that the emergence of channels built of small, α-helical peptides was protobiologically plausible and did not require highly specific amino acid sequences, which is a convenient evolutionary trait. Despite their simple structure, such channels could possess properties that, at the first sight, appear to require markedly larger complexity. To this end, we will discuss how the amino acid sequence and structure of primitive channels give rise to the phenomena of ionic conductance and selectivity across the earliest cell walls, which were essential functions for the emergence and early evolution of protocells. Furthermore, we will argue that even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during evolution.

Hydrogen peroxide scavenger, catalase, alleviates ion transport dysfunction in murine colitis.

PubMed

Barrett, Kim E; McCole, Declan F

2016-11-01

Reactive oxygen species (ROS) such as hydrogen peroxide (H 2 O 2 ) contribute to epithelial damage and ion transport dysfunction (key events in inflammatory diarrhoea) in inflammatory bowel disease (IBD). The aim of this study was to identify if H 2 O 2 mediates suppression of colonic ion transport function in the murine dextran sulfate sodium (DSS) colitis model by using the H 2 O 2 degrading enzyme, catalase. Colitis was induced by administering DSS (4%) in drinking water for 5 days followed by 3 days on normal H 2 O. Mice were administered either pegylated catalase or saline at day -1, 0 and +1 of DSS treatment. Ion transport responses to the Ca 2+ -dependent agonist, carbachol (CCh), or the cAMP-dependent agonist, forskolin, were measured across distal colonic mucosa mounted in Ussing chambers. Parameters of DSS-induced inflammation (loss in body weight, decreased colon length, altered stool consistency), were only partially alleviated by catalase while histology was only minimally improved. However, catalase significantly reversed the DSS-induced reduction in baseline ion transport as well as colonic I sc responses to CCh. However, ion transport responses to forskolin were not significantly restored. Catalase also reduced activation of ERK MAP kinase in the setting of colitis, and increased expression of the Na + -K + -2Cl - cotransporter, NKCC1, consistent with restoration of ion transport function. Ex vivo treatment of inflamed colonic mucosae with catalase also partially restored ion transport function. Therefore, catalase partially prevents, and rescues, the loss of ion transport properties in DSS colitis even in the setting of unresolved tissue inflammation. These findings indicate a prominent role for ROS in ion transport dysfunction in colitis and may suggest novel strategies for the treatment of inflammatory diarrhoea. © 2016 John Wiley & Sons Australia, Ltd.

The Hydrogen Peroxide Scavenger, Catalase, Alleviates Ion Transport Dysfunction in Murine Colitis

PubMed Central

Barrett, Kim E.; McCole, Declan F.

2016-01-01

Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) contribute to epithelial damage and ion transport dysfunction (key events in inflammatory diarrhea) in inflammatory bowel disease (IBD). The aim of this study was to identify if H2O2 mediates suppression of colonic ion transport function in the murine dextran sulfate sodium (DSS) colitis model by using the H2O2 degrading enzyme, catalase. Colitis was induced by administering DSS (4%) in drinking water for 5 days followed by 3 days on normal H2O. Mice were administered either pegylated-catalase or saline at day −1, 0 and +1 of DSS treatment. Ion transport responses to the Ca2+-dependent agonist, carbachol (CCh), or the cAMP-dependent agonist, forskolin, were measured across distal colonic mucosa mounted in Ussing chambers. Parameters of DSS-induced inflammation (loss in body weight, decreased colon length, altered stool consistency), were only partially alleviated by catalase while histology was only minimally improved. However, catalase significantly reversed the DSS-induced reduction in baseline ion transport as well as colonic Isc responses to CCh. However, ion transport responses to forskolin were not significantly restored. Catalase also reduced activation of ERK MAP kinase in the setting of colitis, and increased expression of the Na+-K+-2Cl− cotransporter, NKCC1, consistent with restoration of ion transport function. Ex vivo treatment of inflamed colonic mucosae with catalase also partially restored ion transport function. Therefore, catalase partially prevents, and rescues, the loss of ion transport properties in DSS colitis even in the setting of unresolved tissue inflammation. These findings indicate a prominent role for ROS in ion transport dysfunction in colitis and may suggest novel strategies for the treatment of inflammatory diarrhea. PMID:27543846

Molecular dynamics and brownian dynamics investigation of ion permeation and anesthetic halothane effects on a proton-gated ion channel.

PubMed

Cheng, Mary Hongying; Coalson, Rob D; Tang, Pei

2010-11-24

Bacterial Gloeobacter violaceus pentameric ligand-gated ion channel (GLIC) is activated to cation permeation upon lowering the solution pH. Its function can be modulated by anesthetic halothane. In the present work, we integrate molecular dynamics (MD) and Brownian dynamics (BD) simulations to elucidate the ion conduction, charge selectivity, and halothane modulation mechanisms in GLIC, based on recently resolved X-ray crystal structures of the open-channel GLIC. MD calculations of the potential of mean force (PMF) for a Na(+) revealed two energy barriers in the extracellular domain (R109 and K38) and at the hydrophobic gate of transmembrane domain (I233), respectively. An energy well for Na(+) was near the intracellular entrance: the depth of this energy well was modulated strongly by the protonation state of E222. The energy barrier for Cl(-) was found to be 3-4 times higher than that for Na(+). Ion permeation characteristics were determined through BD simulations using a hybrid MD/continuum electrostatics approach to evaluate the energy profiles governing the ion movement. The resultant channel conductance and a near-zero permeability ratio (P(Cl)/P(Na)) were comparable to experimental data. On the basis of these calculations, we suggest that a ring of five E222 residues may act as an electrostatic gate. In addition, the hydrophobic gate region may play a role in charge selectivity due to a higher dehydration energy barrier for Cl(-) ions. The effect of halothane on the Na(+) PMF was also evaluated. Halothane was found to perturb salt bridges in GLIC that may be crucial for channel gating and open-channel stability, but had no significant impact on the single ion PMF profiles.

Computational Methods of Studying the Binding of Toxins From Venomous Animals to Biological Ion Channels: Theory and Applications

PubMed Central

Chen, Rong; Chung, Shin-Ho

2013-01-01

The discovery of new drugs that selectively block or modulate ion channels has great potential to provide new treatments for a host of conditions. One promising avenue revolves around modifying or mimicking certain naturally occurring ion channel modulator toxins. This strategy appears to offer the prospect of designing drugs that are both potent and specific. The use of computational modeling is crucial to this endeavor, as it has the potential to provide lower cost alternatives for exploring the effects of new compounds on ion channels. In addition, computational modeling can provide structural information and theoretical understanding that is not easily derivable from experimental results. In this review, we look at the theory and computational methods that are applicable to the study of ion channel modulators. The first section provides an introduction to various theoretical concepts, including force-fields and the statistical mechanics of binding. We then look at various computational techniques available to the researcher, including molecular dynamics, Brownian dynamics, and molecular docking systems. The latter section of the review explores applications of these techniques, concentrating on pore blocker and gating modifier toxins of potassium and sodium channels. After first discussing the structural features of these channels, and their modes of block, we provide an in-depth review of past computational work that has been carried out. Finally, we discuss prospects for future developments in the field. PMID:23589832

PhcrTx2, a New Crab-Paralyzing Peptide Toxin from the Sea Anemone Phymanthus crucifer

PubMed Central

Garateix, Anoland; Salceda, Emilio; Zaharenko, André Junqueira; Pons, Tirso; Santos, Yúlica; Arreguín, Roberto; Ständker, Ludger; Forssmann, Wolf-Georg; Tytgat, Jan; Vega, Rosario

2018-01-01

Sea anemones produce proteinaceous toxins for predation and defense, including peptide toxins that act on a large variety of ion channels of pharmacological and biomedical interest. Phymanthus crucifer is commonly found in the Caribbean Sea; however, the chemical structure and biological activity of its toxins remain unknown, with the exception of PhcrTx1, an acid-sensing ion channel (ASIC) inhibitor. Therefore, in the present work, we focused on the isolation and characterization of new P. crucifer toxins by chromatographic fractionation, followed by a toxicity screening on crabs, an evaluation of ion channels, and sequence analysis. Five groups of toxic chromatographic fractions were found, and a new paralyzing toxin was purified and named PhcrTx2. The toxin inhibited glutamate-gated currents in snail neurons (maximum inhibition of 35%, IC50 4.7 µM), and displayed little or no influence on voltage-sensitive sodium/potassium channels in snail and rat dorsal root ganglion (DRG) neurons, nor on a variety of cloned voltage-gated ion channels. The toxin sequence was fully elucidated by Edman degradation. PhcrTx2 is a new β-defensin-fold peptide that shares a sequence similarity to type 3 potassium channels toxins. However, its low activity on the evaluated ion channels suggests that its molecular target remains unknown. PhcrTx2 is the first known paralyzing toxin in the family Phymanthidae. PMID:29414882

Pore dilatation increases the bicarbonate permeability of CFTR, ANO1 and glycine receptor anion channels

PubMed Central

Jun, Ikhyun; Cheng, Mary Hongying; Sim, Eunji; Jung, Jinsei; Suh, Bong Lim; Kim, Yonjung; Son, Hankil; Park, Kyungsoo; Kim, Chul Hoon; Yoon, Joo‐Heon; Whitcomb, David C.; Bahar, Ivet

2016-01-01

Key points Cellular stimuli can modulate the ion selectivity of some anion channels, such as CFTR, ANO1 and the glycine receptor (GlyR), by changing pore size.Ion selectivity of CFTR, ANO1 and GlyR is critically affected by the electric permittivity and diameter of the channel pore.Pore size change affects the energy barriers of ion dehydration as well as that of size‐exclusion of anion permeation.Pore dilatation increases the bicarbonate permeability (P HC O3/ Cl ) of CFTR, ANO1 and GlyR.Dynamic change in P HC O3/ Cl may mediate many physiological and pathological processes. Abstract Chloride (Cl−) and bicarbonate (HCO3 −) are two major anions and their permeation through anion channels plays essential roles in our body. However, the mechanism of ion selection by the anion channels is largely unknown. Here, we provide evidence that pore dilatation increases the bicarbonate permeability (P HC O3/ Cl ) of anion channels by reducing energy barriers of size‐exclusion and ion dehydration of HCO3 − permeation. Molecular, physiological and computational analyses of major anion channels, such as cystic fibrosis transmembrane conductance regulator (CFTR), anoctamin‐1(ANO1/TMEM16A) and the glycine receptor (GlyR), revealed that the ion selectivity of anion channels is basically determined by the electric permittivity and diameter of the pore. Importantly, cellular stimuli dynamically modulate the anion selectivity of CFTR and ANO1 by changing the pore size. In addition, pore dilatation by a mutation in the pore‐lining region alters the anion selectivity of GlyR. Changes in pore size affected not only the energy barriers of size exclusion but that of ion dehydration by altering the electric permittivity of water‐filled cavity in the pore. The dynamic increase in P HC O3/ Cl by pore dilatation may have many physiological and pathophysiological implications ranging from epithelial HCO3 − secretion to neuronal excitation. PMID:26663196

Self-organised criticality and 1/f noise in single-channel current of voltage-dependent anion channel

NASA Astrophysics Data System (ADS)

Banerjee, J.; Verma, M. K.; Manna, S.; Ghosh, S.

2006-02-01

Noise profile of Voltage Dependent Anion Channel (VDAC) is investigated in open channel state. Single-channel currents through VDAC from mitochondria of rat brain reconstituted into a planar lipid bilayer are recorded under different voltage clamped conditions across the membrane. Power spectrum analysis of current indicates power law noise of 1/f nature. Moreover, this 1/f nature of the open channel noise is seen throughout the range of applied membrane potential from -30 to +30 mV. It is being proposed that 1/f noise in open ion channel arises out of obstruction in the passage of ions across the membrane. The process is recognised as a phenomenon of self-organized criticality (SOC) like sandpile avalanche and other physical systems. Based on SOC it has been theoretically established that the system of ion channel follows power law noise as observed in our experiments. We also show that the first-time return probability of current fluctuations obeys a power law distribution.

Demystifying Mechanosensitive Piezo Ion Channels.

PubMed

Xu, X Z Shawn

2016-06-01

Mechanosensitive channels mediate touch, hearing, proprioception, and blood pressure regulation. Piezo proteins, including Piezo1 and Piezo2, represent a new class of mechanosensitive channels that have been reported to play key roles in most, if not all, of these modalities. The structural architecture and molecular mechanisms by which Piezos act as mechanosensitive channels, however, remain mysterious. Two new studies have now provided critical insights into the atomic structure and molecular basis of the ion permeation and mechano-gating properties of the Piezo1 channel.

Ionic blockade of the rat connexin40 gap junction channel by large tetraalkylammonium ions.

PubMed

Musa, H; Gough, J D; Lees, W J; Veenstra, R D

2001-12-01

The rat connexin40 gap junction channel is permeable to monovalent cations including tetramethylammonium and tetraethylammonium ions. Larger tetraalkyammonium (TAA(+)) ions beginning with tetrabutylammonium (TBA(+)) reduced KCl junctional currents disproportionately. Ionic blockade by tetrapentylammonium (TPeA(+)) and tetrahexylammonium (THxA(+)) ions were concentration- and voltage-dependent and occurred only when TAA(+) ions were on the same side as net K(+) efflux across the junction, indicative of block of the ionic permeation pathway. The voltage-dependent dissociation constants (K(m)(V(j))) were lower for THxA(+) than TPeA(+), consistent with steric effects within the pore. The K(m)-V(j) relationships for TPeA(+) and THxA(+) were fit with different reaction rate models for a symmetrical (homotypic) connexin gap junction channel and were described by either a one- or two-site model that assumed each ion traversed the entire V(j) field. Bilateral addition of TPeA(+) ions confirmed a common site of interaction within the pore that possessed identical K(m)(V(j)) values for cis-trans concentrations of TPeA(+) ions as indicated by the modeled I-V relations and rapid channel block that precluded unitary current measurements. The TAA(+) block of K(+) currents and bilateral TPeA(+) interactions did not alter V(j)-gating of Cx40 gap junctions. N-octyl-tributylammonium and -triethylammonium also blocked rCx40 channels with higher affinity and faster kinetics than TBA(+) or TPeA(+), indicative of a hydrophobic site within the pore near the site of block.

Stimulation of TRPC5 cationic channels by low micromolar concentrations of lead ions (Pb{sup 2+})

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

Sukumar, Piruthivi; Beech, David J., E-mail: d.j.beech@leeds.ac.uk

2010-02-26

Lead toxicity is long-recognised but continues to be a major public health problem. Its effects are wide-ranging and include induction of hyper-anxiety states. In general it is thought to act by interfering with Ca{sup 2+} signalling but specific targets are not clearly identified. Transient receptor potential canonical 5 (TRPC5) is a Ca{sup 2+}-permeable ion channel that is linked positively to innate fear responses and unusual amongst ion channels in being stimulated by trivalent lanthanides, which include gadolinium. Here we show investigation of the effect of lead, which is a divalent ion (Pb{sup 2+}). Intracellular Ca{sup 2+} and whole-cell patch-clamp recordingsmore » were performed on HEK 293 cells conditionally over-expressing TRPC5 or other TRP channels. Extracellular application of Pb{sup 2+} stimulated TRPC5 at concentrations greater than 1 {mu}M. Control cells without TRPC5 showed little or no response to Pb{sup 2+} and expression of other TRP channels (TRPM2 or TRPM3) revealed partial inhibition by 10 {mu}M Pb{sup 2+}. The stimulatory effect on TRPC5 depended on an extracellular residue (E543) near the ion pore: similar to gadolinium action, E543Q TRPC5 was resistant to Pb{sup 2+} but showed normal stimulation by the receptor agonist sphingosine-1-phosphate. The study shows that Pb{sup 2+} is a relatively potent stimulator of the TRPC5 channel, generating the hypothesis that a function of the channel is to sense metal ion poisoning.« less

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.

Brownian Dynamics Simulations of Ion Transport through the VDAC

PubMed Central

Lee, Kyu Il; Rui, Huan; Pastor, Richard W.; Im, Wonpil

2011-01-01

It is important to gain a physical understanding of ion transport through the voltage-dependent anion channel (VDAC) because this channel provides primary permeation pathways for metabolites and electrolytes between the cytosol and mitochondria. We performed grand canonical Monte Carlo/Brownian dynamics (GCMC/BD) simulations to explore the ion transport properties of human VDAC isoform 1 (hVDAC1; PDB:2K4T) embedded in an implicit membrane. When the MD-derived, space-dependent diffusion constant was used in the GCMC/BD simulations, the current-voltage characteristics and ion number profiles inside the pore showed excellent agreement with those calculated from all-atom molecular-dynamics (MD) simulations, thereby validating the GCMC/BD approach. Of the 20 NMR models of hVDAC1 currently available, the third one (NMR03) best reproduces both experimental single-channel conductance and ion selectivity (i.e., the reversal potential). In addition, detailed analyses of the ion trajectories, one-dimensional multi-ion potential of mean force, and protein charge distribution reveal that electrostatic interactions play an important role in the channel structure and ion transport relationship. Finally, the GCMC/BD simulations of various mutants based on NMR03 show good agreement with experimental ion selectivity. The difference in ion selectivity between the wild-type and the mutants is the result of altered potential of mean force profiles that are dominated by the electrostatic interactions. PMID:21281575

Gasotransmitter Regulation of Ion Channels: A Key Step in O2 Sensing By the Carotid Body

PubMed Central

Prabhakar, Nanduri R.

2014-01-01

Carotid bodies detect hypoxia in arterial blood, translating this stimulus into physiological responses via the CNS. It is long established that ion channels are critical to this process. More recent evidence indicates that gasotransmitters exert powerful influences on O2 sensing by the carotid body. Here, we review current understanding of hypoxia-dependent production of gasotransmitters, how they regulate ion channels in the carotid body, and how this impacts carotid body function. PMID:24382871

Validity of the Electrodiffusion Model for Calculating Conductance of Simple Ion Channels.

PubMed

Pohorille, Andrew; Wilson, Michael A; Wei, Chenyu

2017-04-20

We examine the validity and utility of the electrodiffusion (ED) equation, i.e., the generalized Nernst-Planck equation, to characterize, in combination with molecular dynamics, the electrophysiological behavior of simple ion channels. As models, we consider three systems-two naturally occurring channels formed by α-helical bundles of peptaibols, trichotoxin, and alamethicin, and a synthetic, hexameric channel, formed by a peptide that contains only leucine and serine. All these channels mediate transport of potassium and chloride ions. Starting with equilibrium properties, such as the potential of mean force experienced by an ion traversing the channel and diffusivity, obtained from molecular dynamics simulations, the ED equation can be used to determine the full current-voltage dependence with modest or no additional effort. The potential of mean force can be obtained not only from equilibrium simulations, but also, with comparable accuracy, from nonequilibrium simulations at a single voltage. The main assumptions underlying the ED equation appear to hold well for the channels and voltages studied here. To expand the utility of the ED equation, we examine what are the necessary and sufficient conditions for Ohmic and nonrectifying behavior and relate deviations from this behavior to the shape of the ionic potential of mean force.

The permeability of endplate channels to monovalent and divalent metal cations

PubMed Central

1980-01-01

The relative permeability of endplate channels to monovalent and divalent metal ions was determined from reversal potentials. Thallium is the most permeant ion with a permeability ratio relative to Na+ of 2.5. The selectivity among alkali metals is weak with a sequence, Cs+ greater than Rb+ greater than K+ greater than Na+ greater than Li+, and permeability ratios of 1.4, 1.3, 1.1, 1.0, and 0.9. The selectivity among divalent ions is also weak, with a sequence for alkaline earths of Mg++ greater than Ca++ greater than Ba++ greater than Sr++. The transition metal ions Mn++, Co++, Ni++, Zn++, and Cd++ are also permeant. Permeability ratios for divalent ions decreased as the concentration of divalent ion was increased in a manner consistent with the negative surface potential theory of Lewis (1979 J. Physiol. (Lond.). 286: 417--445). With 20 mM XCl2 and 85.5 mM glucosamine.HCl in the external solution, the apparent permeability ratios for the alkaline earth cations (X++) are in the range 0.18--0.25. Alkali metal ions see the endplate channel as a water-filled, neutral pore without high-field-strength sites inside. Their permeability sequence is the same as their aqueous mobility sequence. Divalent ions, however, have a permeability sequence almost opposite from their mobility sequence and must experience some interaction with groups in the channel. In addition, the concentrations of monovalent and divalent ions are increased near the channel mouth by a weak negative surface potential. PMID:6247423

Structural dynamics of the cell nucleus

PubMed Central

Wiegert, Simon; Bading, Hilmar

2011-01-01

Neuronal morphology plays an essential role in signal processing in the brain. Individual neurons can undergo use-dependent changes in their shape and connectivity, which affects how intracellular processes are regulated and how signals are transferred from one cell to another in a neuronal network. Calcium is one of the most important intracellular second messengers regulating cellular morphologies and functions. In neurons, intracellular calcium levels are controlled by ion channels in the plasma membrane such as NMDA receptors (NMDARs), voltage-gated calcium channels (VGCCs) and certain α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) as well as by calcium exchange pathways between the cytosol and internal calcium stores including the endoplasmic reticulum and mitochondria. Synaptic activity and the subsequent opening of ligand and/or voltage-gated calcium channels can initiate cytosolic calcium transients which propagate towards the cell soma and enter the nucleus via its nuclear pore complexes (NPCs) embedded in the nuclear envelope. We recently described the discovery that in hippocampal neurons the morphology of the nucleus affects the calcium dynamics within the nucleus. Here we propose that nuclear infoldings determine whether a nucleus functions as an integrator or detector of oscillating calcium signals. We outline possible ties between nuclear mophology and transcriptional activity and discuss the importance of extending the approach to whole cell calcium signal modeling in order to understand synapse-to-nucleus communication in healthy and dysfunctional neurons. PMID:21738832

Cloning and characterization of CLCN5, the human kidney chloride channel gene implicated in Dent disease (an X-linked hereditary nephrolithiasis).

PubMed

Fisher, S E; van Bakel, I; Lloyd, S E; Pearce, S H; Thakker, R V; Craig, I W

1995-10-10

Dent disease, an X-linked familial renal tubular disorder, is a form of Fanconi syndrome associated with proteinuria, hypercalciuria, nephrocalcinosis, kidney stones, and eventual renal failure. We have previously used positional cloning to identify the 3' part of a novel kidney-specific gene (initially termed hClC-K2, but now referred to as CLCN5), which is deleted in patients from one pedigree segregating Dent disease. Mutations that disrupt this gene have been identified in other patients with this disorder. Here we describe the isolation and characterization of the complete open reading frame of the human CLCN5 gene, which is predicted to encode a protein of 746 amino acids, with significant homology to all known members of the ClC family of voltage-gated chloride channels. CLCN5 belongs to a distinct branch of this family, which also includes the recently identified genes CLCN3 and CLCN4. We have shown that the coding region of CLCN5 is organized into 12 exons, spanning 25-30 kb of genomic DNA, and have determined the sequence of each exon-intron boundary. The elucidation of the coding sequence and exon-intron organization of CLCN5 will both expedite the evaluation of structure/function relationships of these ion channels and facilitate the screening of other patients with renal tubular dysfunction for mutations at this locus.

Discrete-state representation of ion permeation coupled to fast gating in a model of ClC chloride channels: comparison to multi-ion continuous space Brownian dynamics simulations.

PubMed

Coalson, Rob D; Cheng, Mary Hongying

2010-01-28

A discrete-state model of chloride ion motion in a ClC chloride channel is constructed, following a previously developed multi-ion continuous space model of the same system (Cheng, M. H.; Mamonov, A. B.; Dukes, J. W.; Coalson, R. D. J. Phys. Chem. B 2007, 111, 5956) that included a simplistic representation of the fast gate in this channel. The reducibility of the many-body continuous space to the eight discrete-state model considered in the present work is examined in detail by performing three-dimensional Brownian dynamics simulations of each allowed state-to-state transition in order to extract the appropriate rate constant for this process, and then inserting the pairwise rate constants thereby obtained into an appropriate set of kinetic master equations. Experimental properties of interest, including the rate of Cl(-) ion permeation through the open channel and the average rate of closing of the fast gate as a function of bulk Cl(-) ion concentrations in the intracellular and extracellular electrolyte reservoirs are computed. Good agreement is found between the results obtained via the eight discrete-state model versus the multi-ion continuous space model, thereby encouraging continued development of the discrete-state model to include more complex behaviors observed experimentally in these channels.

Computational, Experimental and Engineering Foundations of Ionic Channels as Miniaturized Sensors, Devices and Systems

DTIC Science & Technology

2003-10-01

made in an ensemble of channels of unknown orientation and number, preventing quantitative analysis . • Currents have been compared among continuum PNP...microfluidic) analysis of ion channels to obtain fundamental insights into the selectivity, conductivity, and sensitivity of ion channels [19], [6...1.1 Develop fast and efficient simulators for steady-state analysis of continuum model for extraction of I-V curves. 1.2 Create

Molecular Targets for Antiepileptic Drug Development

PubMed Central

Meldrum, Brian S.; Rogawski, Michael A.

2007-01-01

Summary This review considers how recent advances in the physiology of ion channels and other potential molecular targets, in conjunction with new information on the genetics of idiopathic epilepsies, can be applied to the search for improved antiepileptic drugs (AEDs). Marketed AEDs predominantly target voltage-gated cation channels (the α subunits of voltage-gated Na+ channels and also T-type voltage-gated Ca2+ channels) or influence GABA-mediated inhibition. Recently, α2–δ voltage-gated Ca2+ channel subunits and the SV2A synaptic vesicle protein have been recognized as likely targets. Genetic studies of familial idiopathic epilepsies have identified numerous genes associated with diverse epilepsy syndromes, including genes encoding Na+ channels and GABAA receptors, which are known AED targets. A strategy based on genes associated with epilepsy in animal models and humans suggests other potential AED targets, including various voltage-gated Ca2+ channel subunits and auxiliary proteins, A- or M-type voltage-gated K+ channels, and ionotropic glutamate receptors. Recent progress in ion channel research brought about by molecular cloning of the channel subunit proteins and studies in epilepsy models suggest additional targets, including G-protein-coupled receptors, such as GABAB and metabotropic glutamate receptors; hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channel subunits, responsible for hyperpolarization-activated current Ih; connexins, which make up gap junctions; and neurotransmitter transporters, particularly plasma membrane and vesicular transporters for GABA and glutamate. New information from the structural characterization of ion channels, along with better understanding of ion channel function, may allow for more selective targeting. For example, Na+ channels underlying persistent Na+ currents or GABAA receptor isoforms responsible for tonic (extrasynaptic) currents represent attractive targets. The growing understanding of the pathophysiology of epilepsy and the structural and functional characterization of the molecular targets provide many opportunities to create improved epilepsy therapies. PMID:17199015

Creating Lithium-Ion Electrolytes with Biomimetic Ionic Channels in Metal-Organic Frameworks.

PubMed

Shen, Li; Wu, Hao Bin; Liu, Fang; Brosmer, Jonathan L; Shen, Gurong; Wang, Xiaofeng; Zink, Jeffrey I; Xiao, Qiangfeng; Cai, Mei; Wang, Ge; Lu, Yunfeng; Dunn, Bruce

2018-06-01

Solid-state electrolytes are the key to the development of lithium-based batteries with dramatically improved energy density and safety. Inspired by ionic channels in biological systems, a novel class of pseudo solid-state electrolytes with biomimetic ionic channels is reported herein. This is achieved by complexing the anions of an electrolyte to the open metal sites of metal-organic frameworks (MOFs), which transforms the MOF scaffolds into ionic-channel analogs with lithium-ion conduction and low activation energy. This work suggests the emergence of a new class of pseudo solid-state lithium-ion conducting electrolytes. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Nano-Patch-Clamp Array: Microfabricated Glass Chips for High-Throughput Electrophysiology

NASA Astrophysics Data System (ADS)

Fertig, Niels

2003-03-01

Electrophysiology (i.e. patch clamping) remains the gold standard for pharmacological testing of putative ion channel active drugs (ICADs), but suffers from low throughput. A new ion channel screening technology based on microfabricated glass chip devices will be presented. The glass chips contain very fine apertures, which are used for whole-cell voltage clamp recordings as well as single channel recordings from mammalian cell lines. Chips containing multiple patch clamp wells will be used in a first bench-top device, which will allow perfusion and electrical readout of each well. This scalable technology will allow for automated, rapid and parallel screening on ion channel drug targets.

Ion channels in key marine invertebrates; their diversity and potential for applications in biotechnology.

PubMed

Brown, Euan R; Piscopo, Stefania

2011-01-01

Of the intra-membrane proteins, the class that comprises voltage and ligand-gated ion channels represents the major substrate whereby signals pass between and within cells in all organisms. It has been presumed that vertebrate and particularly mammalian ion channels represent the apex of evolutionary complexity and diversity and much effort has been focused on understanding their function. However, the recent availability of cheap high throughput genome sequencing has massively broadened and deepened the quality of information across phylogeny and is radically changing this view. Here we review current knowledge on such channels in key marine invertebrates where physiological evidence is backed up by molecular sequences and expression/functional studies. As marine invertebrates represent a much greater range of phyla than terrestrial vertebrates and invertebrates together, we argue that these animals represent a highly divergent, though relatively underused source of channel novelty. As ion channels are exquisitely selective sensors for voltage and ligands, their potential and actual applications in biotechnology are manifold. Copyright © 2011 Elsevier Inc. All rights reserved.

Laser-Driven Ion Acceleration from Plasma Micro-Channel Targets

PubMed Central

Zou, D. B.; Pukhov, A.; Yi, L. Q.; Zhou, H. B.; Yu, T. P.; Yin, Y.; Shao, F. Q.

2017-01-01

Efficient energy boost of the laser-accelerated ions is critical for their applications in biomedical and hadron research. Achiev-able energies continue to rise, with currently highest energies, allowing access to medical therapy energy windows. Here, a new regime of simultaneous acceleration of ~100 MeV protons and multi-100 MeV carbon-ions from plasma micro-channel targets is proposed by using a ~1020 W/cm2 modest intensity laser pulse. It is found that two trains of overdense electron bunches are dragged out from the micro-channel and effectively accelerated by the longitudinal electric-field excited in the plasma channel. With the optimized channel size, these “superponderomotive” energetic electrons can be focused on the front surface of the attached plastic substrate. The much intense sheath electric-field is formed on the rear side, leading to up to ~10-fold ionic energy increase compared to the simple planar geometry. The analytical prediction of the optimal channel size and ion maximum energies is derived, which shows good agreement with the particle-in-cell simulations. PMID:28218247

Autoimmune Channelopathies of the Nervous System

PubMed Central

Kleopa, Kleopas A

2011-01-01

Ion channels are complex transmembrane proteins that orchestrate the electrical signals necessary for normal function of excitable tissues, including the central nervous system, peripheral nerve, and both skeletal and cardiac muscle. Progress in molecular biology has allowed cloning and expression of genes that encode channel proteins, while comparable advances in biophysics, including patch-clamp electrophysiology and related techniques, have made the functional assessment of expressed proteins at the level of single channel molecules possible. The role of ion channel defects in the pathogenesis of numerous disorders has become increasingly apparent over the last two decades. Neurological channelopathies are frequently genetically determined but may also be acquired through autoimmune mechanisms. All of these autoimmune conditions can arise as paraneoplastic syndromes or independent from malignancies. The pathogenicity of autoantibodies to ion channels has been demonstrated in most of these conditions, and patients may respond well to immunotherapies that reduce the levels of the pathogenic autoantibodies. Autoimmune channelopathies may have a good prognosis, especially if diagnosed and treated early, and if they are non-paraneoplastic. This review focuses on clinical, pathophysiologic and therapeutic aspects of autoimmune ion channel disorders of the nervous system. PMID:22379460

Laser-Driven Ion Acceleration from Plasma Micro-Channel Targets

NASA Astrophysics Data System (ADS)

Zou, D. B.; Pukhov, A.; Yi, L. Q.; Zhou, H. B.; Yu, T. P.; Yin, Y.; Shao, F. Q.

2017-02-01

Efficient energy boost of the laser-accelerated ions is critical for their applications in biomedical and hadron research. Achiev-able energies continue to rise, with currently highest energies, allowing access to medical therapy energy windows. Here, a new regime of simultaneous acceleration of ~100 MeV protons and multi-100 MeV carbon-ions from plasma micro-channel targets is proposed by using a ~1020 W/cm2 modest intensity laser pulse. It is found that two trains of overdense electron bunches are dragged out from the micro-channel and effectively accelerated by the longitudinal electric-field excited in the plasma channel. With the optimized channel size, these “superponderomotive” energetic electrons can be focused on the front surface of the attached plastic substrate. The much intense sheath electric-field is formed on the rear side, leading to up to ~10-fold ionic energy increase compared to the simple planar geometry. The analytical prediction of the optimal channel size and ion maximum energies is derived, which shows good agreement with the particle-in-cell simulations.

Kv7.1 ion channels require a lipid to couple voltage sensing to pore opening.

PubMed

Zaydman, Mark A; Silva, Jonathan R; Delaloye, Kelli; Li, Yang; Liang, Hongwu; Larsson, H Peter; Shi, Jingyi; Cui, Jianmin

2013-08-06

Voltage-gated ion channels generate dynamic ionic currents that are vital to the physiological functions of many tissues. These proteins contain separate voltage-sensing domains, which detect changes in transmembrane voltage, and pore domains, which conduct ions. Coupling of voltage sensing and pore opening is critical to the channel function and has been modeled as a protein-protein interaction between the two domains. Here, we show that coupling in Kv7.1 channels requires the lipid phosphatidylinositol 4,5-bisphosphate (PIP2). We found that voltage-sensing domain activation failed to open the pore in the absence of PIP2. This result is due to loss of coupling because PIP2 was also required for pore opening to affect voltage-sensing domain activation. We identified a critical site for PIP2-dependent coupling at the interface between the voltage-sensing domain and the pore domain. This site is actually a conserved lipid-binding site among different K(+) channels, suggesting that lipids play an important role in coupling in many ion channels.

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.

Design and development of a fast ion mass spectrometer

NASA Technical Reports Server (NTRS)

Burch, J. L.

1983-01-01

Two Fast Ion Mass Spectrometers (FIMS A and FIMS B) were developed. The design, development, construction, calibration, integration, and flight of these instruments, along with early results from the data analysis efforts are summarized. A medium energy ion mass spectrometer that covers mass velocity space with significantly higher time resolution, improved mass resolution, (particularly for heavier ions), and wider energy range than existing instruments had achieved was completed. The initial design consisted of a dual channel cylindrical electrostatic analyzer followed by a dual channel cylindrical velocity filter. The gain versus count rate characteristics of the high current channel electron multipliers (CEM's), which were chosen for ion detection, revealed a systematic behavior that can be used as a criterion for selection of CEM's for long counting lifetimes.

A chimera encoding the fusion of an acetylcholine-binding protein to an ion channel is stabilized in a state close to the desensitized form of ligand-gated ion channels.

PubMed

Grutter, Thomas; Prado de Carvalho, Lia; Virginie, Dufresne; Taly, Antoine; Fischer, Markus; Changeux, Jean-Pierre

2005-03-01

To understand the mechanism of allosteric coupling between the ligand-binding domain and the ion channel of the Cys-loop ligand-gated ion channels (LGICs), we fused the soluble acetylcholine-binding protein (AChBP), which lacks an ion channel, to either the cationic serotonin type-3A ion channel (5HT(3A)) or the anionic glycine ion channel. Both linear chimeras expressed in HEK-293 cells display high affinity for the nicotinic agonist epibatidine (K(D) = 0.2-0.5 nM), but are not targeted to the cell surface. Only after substituting a ring of three loops located at the putative membrane side of the AChBP three-dimensional structure by the homologous residues of 5HT(3A), the resulting chimera AChBP(ring)/5HT(3A) (i) still displayed on intact cells an apparent high affinity for epibatidine, yet with a fourfold decrease (K(D) = 2.1 nM), (ii) displayed a high proportion of low affinity sites (11 +/- 7 microM) for the resting state stabilizing competitive antagonist alpha-bungarotoxin and (iii) was successfully targeted to the cell surface, as seen by immunofluorescence labelling. The AChBP(ring)/5HT(3A) chimera forms a pentameric structure, as revealed by sucrose gradient sedimentation. However, no whole-cell patch-clamp currents were detectable. Interestingly, binding assays with membrane fragments prepared from cells expressing AChBP(ring)/5HT(3A) showed a decrease in the apparent affinity for the agonists nicotine and epibatidine (5-fold), concomitant with an increase in the proportion of high-affinity sites (48 +/- 1 nM) for alpha-bungarotoxin. These results indicate that fusion of AChBP to an ion channel forms a pentameric receptor exposed to the cell surface and able to convert between discrete allosteric states, but stabilized in a high affinity state for epibatidine that likely corresponds to a desensitized form of LGICs. These artificial chimeras might offer a useful system to investigate signal transduction in LGICs.

Hysteresis in voltage-gated channels

PubMed Central

2017-01-01

ABSTRACT Ion channels constitute a superfamily of membrane proteins found in all living creatures. Their activity allows fast translocation of ions across the plasma membrane down the ion's transmembrane electrochemical gradient, resulting in a difference in electrical potential across the plasma membrane, known as the membrane potential. A group within this superfamily, namely voltage-gated channels, displays activity that is sensitive to the membrane potential. The activity of voltage-gated channels is controlled by the membrane potential, while the membrane potential is changed by these channels' activity. This interplay produces variations in the membrane potential that have evolved into electrical signals in many organisms. These signals are essential for numerous biological processes, including neuronal activity, insulin release, muscle contraction, fertilization and many others. In recent years, the activity of the voltage-gated channels has been observed not to follow a simple relationship with the membrane potential. Instead, it has been shown that the activity of voltage-gated channel displays hysteresis. In fact, a growing number of evidence have demonstrated that the voltage dependence of channel activity is dynamically modulated by activity itself. In spite of the great impact that this property can have on electrical signaling, hysteresis in voltage-gated channels is often overlooked. Addressing this issue, this review provides examples of voltage-gated ion channels displaying hysteretic behavior. Further, this review will discuss how Dynamic Voltage Dependence in voltage-gated channels can have a physiological role in electrical signaling. Furthermore, this review will elaborate on the current thoughts on the mechanism underlying hysteresis in voltage-gated channels. PMID:27689426

Finite element simulation of the gating mechanism of mechanosensitive ion channels

NASA Astrophysics Data System (ADS)

Bavi, Navid; Qin, Qinghua; Martinac, Boris

2013-08-01

In order to eliminate limitations of existing experimental or computational methods (such as patch-clamp technique or molecular dynamic analysis) a finite element (FE) model for multi length-scale and time-scale investigation on the gating mechanism of mechanosensitive (MS) ion channels has been established. Gating force value (from typical patch clamping values) needed to activate Prokaryotic MS ion channels was applied as tensional force to the FE model of the lipid bilayer. Making use of the FE results, we have discussed the effects of the geometrical and the material properties of the Escherichia coli MscL mechanosensitive ion channel opening in relation to the membrane's Young's modulus (which will vary depending on the cell type or cholesterol density in an artificial membrane surrounding the MscL ion channel). The FE model has shown that when the cell membrane stiffens the required channel activation force increases considerably. This is in agreement with experimental results taken from the literature. In addition, the present study quantifies the relationship between the membrane stress distribution around a `hole' for modeling purposes and the stress concentration in the place transmembrane proteins attached to the hole by applying an appropriate mesh refinement as well as well defining contact condition in these areas.

A Low-Noise Transimpedance Amplifier for BLM-Based Ion Channel Recording.

PubMed

Crescentini, Marco; Bennati, Marco; Saha, Shimul Chandra; Ivica, Josip; de Planque, Maurits; Morgan, Hywel; Tartagni, Marco

2016-05-19

High-throughput screening (HTS) using ion channel recording is a powerful drug discovery technique in pharmacology. Ion channel recording with planar bilayer lipid membranes (BLM) is scalable and has very high sensitivity. A HTS system based on BLM ion channel recording faces three main challenges: (i) design of scalable microfluidic devices; (ii) design of compact ultra-low-noise transimpedance amplifiers able to detect currents in the pA range with bandwidth >10 kHz; (iii) design of compact, robust and scalable systems that integrate these two elements. This paper presents a low-noise transimpedance amplifier with integrated A/D conversion realized in CMOS 0.35 μm technology. The CMOS amplifier acquires currents in the range ±200 pA and ±20 nA, with 100 kHz bandwidth while dissipating 41 mW. An integrated digital offset compensation loop balances any voltage offsets from Ag/AgCl electrodes. The measured open-input input-referred noise current is as low as 4 fA/√Hz at ±200 pA range. The current amplifier is embedded in an integrated platform, together with a microfluidic device, for current recording from ion channels. Gramicidin-A, α-haemolysin and KcsA potassium channels have been used to prove both the platform and the current-to-digital converter.

A Low-Noise Transimpedance Amplifier for BLM-Based Ion Channel Recording

PubMed Central

Crescentini, Marco; Bennati, Marco; Saha, Shimul Chandra; Ivica, Josip; de Planque, Maurits; Morgan, Hywel; Tartagni, Marco

2016-01-01

High-throughput screening (HTS) using ion channel recording is a powerful drug discovery technique in pharmacology. Ion channel recording with planar bilayer lipid membranes (BLM) is scalable and has very high sensitivity. A HTS system based on BLM ion channel recording faces three main challenges: (i) design of scalable microfluidic devices; (ii) design of compact ultra-low-noise transimpedance amplifiers able to detect currents in the pA range with bandwidth >10 kHz; (iii) design of compact, robust and scalable systems that integrate these two elements. This paper presents a low-noise transimpedance amplifier with integrated A/D conversion realized in CMOS 0.35 μm technology. The CMOS amplifier acquires currents in the range ±200 pA and ±20 nA, with 100 kHz bandwidth while dissipating 41 mW. An integrated digital offset compensation loop balances any voltage offsets from Ag/AgCl electrodes. The measured open-input input-referred noise current is as low as 4 fA/√Hz at ±200 pA range. The current amplifier is embedded in an integrated platform, together with a microfluidic device, for current recording from ion channels. Gramicidin-A, α-haemolysin and KcsA potassium channels have been used to prove both the platform and the current-to-digital converter. PMID:27213382

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.

Endotoxin induces fibrosis in vascular endothelial cells through a mechanism dependent on transient receptor protein melastatin 7 activity.

PubMed

Echeverría, Cesar; Montorfano, Ignacio; Hermosilla, Tamara; Armisén, Ricardo; Velásquez, Luis A; Cabello-Verrugio, Claudio; Varela, Diego; Simon, Felipe

2014-01-01

The pathogenesis of systemic inflammatory diseases, including endotoxemia-derived sepsis syndrome, is characterized by endothelial dysfunction. It has been demonstrated that the endotoxin lipopolysaccharide (LPS) induces the conversion of endothelial cells (ECs) into activated fibroblasts through endothelial-to-mesenchymal transition mechanism. Fibrogenesis is highly dependent on intracellular Ca2+ concentration increases through the participation of calcium channels. However, the specific molecular identity of the calcium channel that mediates the Ca2+ influx during endotoxin-induced endothelial fibrosis is still unknown. Transient receptor potential melastatin 7 (TRPM7) is a calcium channel that is expressed in many cell types, including ECs. TRPM7 is involved in a number of crucial processes such as the conversion of fibroblasts into activated fibroblasts, or myofibroblasts, being responsible for the development of several characteristics of them. However, the role of the TRPM7 ion channel in endotoxin-induced endothelial fibrosis is unknown. Thus, our aim was to study whether the TRPM7 calcium channel participates in endotoxin-induced endothelial fibrosis. Using primary cultures of ECs, we demonstrated that TRPM7 is a crucial protein involved in endotoxin-induced endothelial fibrosis. Suppression of TRPM7 expression protected ECs from the fibrogenic process stimulated by endotoxin. Downregulation of TRPM7 prevented the endotoxin-induced endothelial markers decrease and fibrotic genes increase in ECs. In addition, TRPM7 downregulation abolished the endotoxin-induced increase in ECM proteins in ECs. Furthermore, we showed that intracellular Ca2+ levels were greatly increased upon LPS challenge in a mechanism dependent on TRPM7 expression. These results demonstrate that TRPM7 is a key protein involved in the mechanism underlying endotoxin-induced endothelial fibrosis.

Placing Ion Channels into a Signaling Network of T Cells: From Maturing Thymocytes to Healthy T Lymphocytes or Leukemic T Lymphoblasts

PubMed Central

Delgado-Enciso, Iván; Best-Aguilera, Carlos; Rojas-Sotelo, Rocío Monserrat; Pottosin, Igor

2015-01-01

T leukemogenesis is a multistep process, where the genetic errors during T cell maturation cause the healthy progenitor to convert into the leukemic precursor that lost its ability to differentiate but possesses high potential for proliferation, self-renewal, and migration. A new misdirecting “leukemogenic” signaling network appears, composed by three types of participants which are encoded by (1) genes implicated in determined stages of T cell development but deregulated by translocations or mutations, (2) genes which normally do not participate in T cell development but are upregulated, and (3) nondifferentially expressed genes which become highly interconnected with genes expressed differentially. It appears that each of three groups may contain genes coding ion channels. In T cells, ion channels are implicated in regulation of cell cycle progression, differentiation, activation, migration, and cell death. In the present review we are going to reveal a relationship between different genetic defects, which drive the T cell neoplasias, with calcium signaling and ion channels. We suggest that changes in regulation of various ion channels in different types of the T leukemias may provide the intracellular ion microenvironment favorable to maintain self-renewal capacity, arrest differentiation, induce proliferation, and enhance motility. PMID:25866806

Single-Walled Carbon Nanotubes: Mimics of Biological Ion Channels.

PubMed

Amiri, Hasti; Shepard, Kenneth L; Nuckolls, Colin; Hernández Sánchez, Raúl

2017-02-08

Here we report on the ion conductance through individual, small diameter single-walled carbon nanotubes. We find that they are mimics of ion channels found in natural systems. We explore the factors governing the ion selectivity and permeation through single-walled carbon nanotubes by considering an electrostatic mechanism built around a simplified version of the Gouy-Chapman theory. We find that the single-walled carbon nanotubes preferentially transported cations and that the cation permeability is size-dependent. The ionic conductance increases as the absolute hydration enthalpy decreases for monovalent cations with similar solid-state radii, hydrated radii, and bulk mobility. Charge screening experiments using either the addition of cationic or anionic polymers, divalent metal cations, or changes in pH reveal the enormous impact of the negatively charged carboxylates at the entrance of the single-walled carbon nanotubes. These observations were modeled in the low-to-medium concentration range (0.1-2.0 M) by an electrostatic mechanism that mimics the behavior observed in many biological ion channel-forming proteins. Moreover, multi-ion conduction in the high concentration range (>2.0 M) further reinforces the similarity between single-walled carbon nanotubes and protein ion channels.

Channeling STIM analysis of radiation damage in single crystal diamond membrane

NASA Astrophysics Data System (ADS)

Sudić, I.; Cosic, D.; Ditalia Tchernij, S.; Olivero, P.; Pomorski, M.; Skukan, N.; Jakšić, M.

2017-08-01

The use of focused ion beam transmission channeling patterns to monitor the damage creation process in thin diamond single crystal membrane is described. A 0.8 MeV proton beam from the Ruđer Bošković Institute nuclear microprobe was used to perform Channeling Scanning Transmission Ion Microscopy (CSTIM) measurements. CSTIM was used instead of RBS channeling because of (several orders of magnitude) lower damage done to the sample during the measurements. Damage was introduced in selected areas by 15 MeV carbon beam in range of fluences 3·1015-2·1017 ions/cm2. Contrary to Ion Beam Induced Charge (IBIC), CSTIM is shown to be sensitive to the large fluences of ion beam radiation. Complementary studies of both IBIC and CSTIM are presented to show that very high fluence range can be covered by these two microprobe techniques, providing much wider information about the diamond radiation hardness. In addition micro Raman measurements were performed and the height of the GR 1 peak was correlated to the ion beam fluence.

IBiSA_Tools: A Computational Toolkit for Ion-Binding State Analysis in Molecular Dynamics Trajectories of Ion Channels.

PubMed

Kasahara, Kota; Kinoshita, Kengo

2016-01-01

Ion conduction mechanisms of ion channels are a long-standing conundrum. Although the molecular dynamics (MD) method has been extensively used to simulate ion conduction dynamics at the atomic level, analysis and interpretation of MD results are not straightforward due to complexity of the dynamics. In our previous reports, we proposed an analytical method called ion-binding state analysis to scrutinize and summarize ion conduction mechanisms by taking advantage of a variety of analytical protocols, e.g., the complex network analysis, sequence alignment, and hierarchical clustering. This approach effectively revealed the ion conduction mechanisms and their dependence on the conditions, i.e., ion concentration and membrane voltage. Here, we present an easy-to-use computational toolkit for ion-binding state analysis, called IBiSA_tools. This toolkit consists of a C++ program and a series of Python and R scripts. From the trajectory file of MD simulations and a structure file, users can generate several images and statistics of ion conduction processes. A complex network named ion-binding state graph is generated in a standard graph format (graph modeling language; GML), which can be visualized by standard network analyzers such as Cytoscape. As a tutorial, a trajectory of a 50 ns MD simulation of the Kv1.2 channel is also distributed with the toolkit. Users can trace the entire process of ion-binding state analysis step by step. The novel method for analysis of ion conduction mechanisms of ion channels can be easily used by means of IBiSA_tools. This software is distributed under an open source license at the following URL: http://www.ritsumei.ac.jp/~ktkshr/ibisa_tools/.

Neuroimmune interactions in itch: Do chronic itch, chronic pain, and chronic cough share similar mechanisms?

PubMed

Ji, Ru-Rong

2015-12-01

Itch and pain are closely related but also clearly distinct sensations. Pain is known to suppress itch, while analgesics such as morphine can provoke itch. However, in pathological and chronic conditions, pain and itch also have similarities. Dysfunction of the nervous system, as manifested by neural plastic changes in primary sensory neurons of the peripheral nervous system (peripheral sensitization) and spinal cord and brain stem neurons in the central nervous system (central sensitization) will result in chronic pain and itch. Importantly, these diseases also result from immune dysfunction, since inflammatory mediators can directly activate or sensitize nociceptive and pruriceptive neurons in the peripheral and central nervous system, leading to pain and itch hypersensitivity. In this mini-review, I discuss the roles of Toll-like receptors (TLRs), transient receptor potential ankyrin 1 (TRPA1) ion channel, and Nav1.7 sodium channel in regulating itch and inflammation, with special emphasis of neuronal TLR signaling and the interaction of TLR7 and TRPA1. Chronic pain and chronic itch are debilitating diseases and dramatically impact the life quality of patients. Targeting TLRs for the control of inflammation, neuroinflammation (inflammation restricted in the nervous system), and hyperexcitability of nociceptors and pruriceptors will lead to new therapeutics for the relief of chronic pain and chronic itch. Finally, given the shared mechanisms among chronic cough, chronic pain, and chronic itch and the demonstrated efficacy of the neuropathic pain drug gabapentin in treating chronic cough, novel therapeutics targeting TRPA1, Nav1.7, and TLRs may also help to alleviate refractory cough via modulating neuron-immune interaction. Copyright © 2015 Elsevier Ltd. All rights reserved.

New Theoretical Model of Nerve Conduction in Unmyelinated Nerves

PubMed Central

Akaishi, Tetsuya

2017-01-01

Nerve conduction in unmyelinated fibers has long been described based on the equivalent circuit model and cable theory. However, without the change in ionic concentration gradient across the membrane, there would be no generation or propagation of the action potential. Based on this concept, we employ a new conductive model focusing on the distribution of voltage-gated sodium ion channels and Coulomb force between electrolytes. Based on this new model, the propagation of the nerve conduction was suggested to take place far before the generation of action potential at each channel. We theoretically showed that propagation of action potential, which is enabled by the increasing Coulomb force produced by inflowing sodium ions, from one sodium ion channel to the next sodium channel would be inversely proportionate to the density of sodium channels on the axon membrane. Because the longitudinal number of sodium ion channel would be proportionate to the square root of channel density, the conduction velocity of unmyelinated nerves is theoretically shown to be proportionate to the square root of channel density. Also, from a viewpoint of equilibrium state of channel importation and degeneration, channel density was suggested to be proportionate to axonal diameter. Based on these simple basis, conduction velocity in unmyelinated nerves was theoretically shown to be proportionate to the square root of axonal diameter. This new model would also enable us to acquire more accurate and understandable vision on the phenomena in unmyelinated nerves in addition to the conventional electric circuit model and cable theory. PMID:29081751

Fractional Poisson-Nernst-Planck Model for Ion Channels I: Basic Formulations and Algorithms.

PubMed

Chen, Duan

2017-11-01

In this work, we propose a fractional Poisson-Nernst-Planck model to describe ion permeation in gated ion channels. Due to the intrinsic conformational changes, crowdedness in narrow channel pores, binding and trapping introduced by functioning units of channel proteins, ionic transport in the channel exhibits a power-law-like anomalous diffusion dynamics. We start from continuous-time random walk model for a single ion and use a long-tailed density distribution function for the particle jump waiting time, to derive the fractional Fokker-Planck equation. Then, it is generalized to the macroscopic fractional Poisson-Nernst-Planck model for ionic concentrations. Necessary computational algorithms are designed to implement numerical simulations for the proposed model, and the dynamics of gating current is investigated. Numerical simulations show that the fractional PNP model provides a more qualitatively reasonable match to the profile of gating currents from experimental observations. Meanwhile, the proposed model motivates new challenges in terms of mathematical modeling and computations.

Absence of ion-binding affinity in the putatively inactivated low-[K+] structure of the KcsA potassium channel.

PubMed

Boiteux, Céline; Bernèche, Simon

2011-01-12

Potassium channels are membrane proteins that selectively conduct K(+) across cellular membranes. The narrowest part of their pore, the selectivity filter, is responsible for distinguishing K(+) from Na(+), and can also act as a gate through a mechanism known as C-type inactivation. It has been proposed that a conformation of the KcsA channel obtained by crystallization in presence of low concentration of K(+) (PDB 1K4D) could correspond to the C-type inactivated state. Here, we show using molecular mechanics simulations that such conformation has little ion-binding affinity and that ions do not contribute to its stability. The simulations suggest that, in this conformation, the selectivity filter is mostly occupied by water molecules. Whether such ion-free state of the KcsA channel is physiologically accessible and representative of the inactivated state of eukaryotic channels remains unclear. Copyright © 2011 Elsevier Ltd. All rights reserved.

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.

Molecular Basis of Cardiac Delayed Rectifier Potassium Channel Function and Pharmacology.

PubMed

Wu, Wei; Sanguinetti, Michael C

2016-06-01

Human cardiomyocytes express 3 distinct types of delayed rectifier potassium channels. Human ether-a-go-go-related gene (hERG) channels conduct the rapidly activating current IKr; KCNQ1/KCNE1 channels conduct the slowly activating current IKs; and Kv1.5 channels conduct an ultrarapid activating current IKur. Here the authors provide a general overview of the mechanistic and structural basis of ion selectivity, gating, and pharmacology of the 3 types of cardiac delayed rectifier potassium ion channels. Most blockers bind to S6 residues that line the central cavity of the channel, whereas activators interact with the channel at 4 symmetric binding sites outside the cavity. Copyright © 2016 Elsevier Inc. All rights reserved.

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.

Functional ion channels in human pulmonary artery smooth muscle cells: Voltage-dependent cation channels

PubMed Central

Firth, Amy L.; Remillard, Carmelle V.; Platoshyn, Oleksandr; Fantozzi, Ivana; Ko, Eun A.; Yuan, Jason X.-J.

2011-01-01

The activity of voltage-gated ion channels is critical for the maintenance of cellular membrane potential and generation of action potentials. In turn, membrane potential regulates cellular ion homeostasis, triggering the opening and closing of ion channels in the plasma membrane and, thus, enabling ion transport across the membrane. Such transmembrane ion fluxes are important for excitation–contraction coupling in pulmonary artery smooth muscle cells (PASMC). Families of voltage-dependent cation channels known to be present in PASMC include voltage-gated K+ (Kv) channels, voltage-dependent Ca2+-activated K+ (Kca) channels, L- and T- type voltage-dependent Ca2+ channels, voltage-gated Na+ channels and voltage-gated proton channels. When cells are dialyzed with Ca2+-free K+- solutions, depolarization elicits four components of 4-aminopyridine (4-AP)-sensitive Kvcurrents based on the kinetics of current activation and inactivation. In cell-attached membrane patches, depolarization elicits a wide range of single-channel K+ currents, with conductances ranging between 6 and 290 pS. Macroscopic 4-AP-sensitive Kv currents and iberiotoxin-sensitive Kca currents are also observed. Transcripts of (a) two Na+ channel α-subunit genes (SCN5A and SCN6A), (b) six Ca2+ channel α–subunit genes (α1A, α1B, α1X, α1D, α1Eand α1G) and many regulatory subunits (α2δ1, β1-4, and γ6), (c) 22 Kv channel α–subunit genes (Kv1.1 - Kv1.7, Kv1.10, Kv2.1, Kv3.1, Kv3.3, Kv3.4, Kv4.1, Kv4.2, Kv5.1, Kv 6.1-Kv6.3, Kv9.1, Kv9.3, Kv10.1 and Kv11.1) and three Kv channel β-subunit genes (Kvβ1-3) and (d) four Kca channel α–subunit genes (Sloα1 and SK2-SK4) and four Kca channel β-subunit genes (Kcaβ1-4) have been detected in PASMC. Tetrodotoxin-sensitive and rapidly inactivating Na+ currents have been recorded with properties similar to those in cardiac myocytes. In the presence of 20 mM external Ca2+, membrane depolarization from a holding potential of -100 mV elicits a rapidly inactivating T-type Ca2+ current, while depolarization from a holding potential of -70 mV elicits a slowly inactivating dihydropyridine-sensitive L-type Ca2+ current. This review will focus on describing the electrophysiological properties and molecular identities of these voltage-dependent cation channels in PASMC and their contribution to the regulation of pulmonary vascular function and its potential role in the pathogenesis of pulmonary vascular disease. PMID:21927714

Dynamic hybrid materials for constitutional self-instructed membranes.

PubMed

Cazacu, Adinela; Legrand, Yves-Marie; Pasc, Andreea; Nasr, Gihane; Van der Lee, Arie; Mahon, Eugene; Barboiu, Mihail

2009-05-19

Constitutional self-instructed membranes were developed and used for mimicking the adaptive structural functionality of natural ion-channel systems. These membranes are based on dynamic hybrid materials in which the functional self-organized macrocycles are reversibly connected with the inorganic silica through hydrophobic noncovalent interactions. Supramolecular columnar ion-channel architectures can be generated by reversible confinement within scaffolding hydrophobic silica mesopores. They can be structurally determined by using X-ray diffraction and morphologically tuned by alkali-salts templating. From the conceptual point of view, these membranes express a synergistic adaptive behavior: the simultaneous binding of the fittest cation and its anion would be a case of "homotropic allosteric interactions," because in time it increases the transport efficiency of the pore-contained superstructures by a selective evolving process toward the fittest ion channel. The hybrid membranes presented here represent dynamic constitutional systems evolving over time to form the fittest ion channels from a library of molecular and supramolecular components, or selecting the fittest ion pairs from a mixture of salts demonstrating flexible adaptation.

High Mobility SiGe/Si Transistor Structures on Sapphire Substrates Using Ion Implantation

NASA Technical Reports Server (NTRS)

Alterovitz, S. A.; Mueller, C. H.; Croke, E. T.

2003-01-01

High mobility n-type SiGe/Si transistor structures have been fabricated on sapphire substrates by ion implanting phosphorus ions into strained 100 Angstrom thick silicon channels for the first time. The strained Si channels were sandwiched between Si(sub 0.7)Ge(sub 0.3) layers, which, in turn, were deposited on Si(sub 0.7)Ge(sub 0.3) virtual substrates and graded SiGe buffer layers. After the molecular beam epitaxy (MBE) film growth process was completed, ion thick silicon channels implantation and post-annealing were used to introduce donors. The phosphorous ions were preferentially located in the Si channel at a peak concentration of approximately 1x10(exp 18)/cu cm. Room temperature electron mobilities exceeding 750 sq cm/V-sec at carrier densities of 1x10(exp 12)/sq cm were measured. Electron concentration appears to be the key factor that determines mobility, with the highest mobility observed for electron densities in the 1 - 2x10(exp 12)/sq cm range.

Coronary arterial BK channel dysfunction exacerbates ischemia/reperfusion-induced myocardial injury in diabetic mice.

PubMed

Lu, Tong; Jiang, Bin; Wang, Xiao-Li; Lee, Hon-Chi

2016-09-01

The large conductance Ca(2+)-activated K(+) (BK) channels, abundantly expressed in coronary artery smooth muscle cells (SMCs), play a pivotal role in regulating coronary circulation. A large body of evidence indicates that coronary arterial BK channel function is diminished in both type 1 and type 2 diabetes. However, the consequence of coronary BK channel dysfunction in diabetes is not clear. We hypothesized that impaired coronary BK channel function exacerbates myocardial ischemia/reperfusion (I/R) injury in streptozotocin-induced diabetic mice. Combining patch-clamp techniques and cellular biological approaches, we found that diabetes facilitated the colocalization of angiotensin II (Ang II) type 1 receptors and BK channel α-subunits (BK-α), but not BK channel β1-subunits (BK-β1), in the caveolae of coronary SMCs. This caveolar compartmentation in vascular SMCs not only enhanced Ang II-mediated inhibition of BK-α but also produced a physical disassociation between BK-α and BK-β1, leading to increased infarct size in diabetic hearts. Most importantly, genetic ablation of caveolae integrity or pharmacological activation of coronary BK channels protected the cardiac function of diabetic mice from experimental I/R injury in both in vivo and ex vivo preparations. Our results demonstrate a vascular ionic mechanism underlying the poor outcome of myocardial injury in diabetes. Hence, activation of coronary BK channels may serve as a therapeutic target for cardiovascular complications of diabetes.

Automated Electrophysiology Makes the Pace for Cardiac Ion Channel Safety Screening

PubMed Central

Möller, Clemens; Witchel, Harry

2011-01-01

The field of automated patch-clamp electrophysiology has emerged from the tension between the pharmaceutical industry’s need for high-throughput compound screening versus its need to be conservative due to regulatory requirements. On the one hand, hERG channel screening was increasingly requested for new chemical entities, as the correlation between blockade of the ion channel coded by hERG and torsades de pointes cardiac arrhythmia gained increasing attention. On the other hand, manual patch-clamping, typically quoted as the “gold-standard” for understanding ion channel function and modulation, was far too slow (and, consequently, too expensive) for keeping pace with the numbers of compounds submitted for hERG channel investigations from pharmaceutical R&D departments. In consequence it became more common for some pharmaceutical companies to outsource safety pharmacological investigations, with a focus on hERG channel interactions. This outsourcing has allowed those pharmaceutical companies to build up operational flexibility and greater independence from internal resources, and allowed them to obtain access to the latest technological developments that emerged in automated patch-clamp electrophysiology – much of which arose in specialized biotech companies. Assays for nearly all major cardiac ion channels are now available by automated patch-clamping using heterologous expression systems, and recently, automated action potential recordings from stem-cell derived cardiomyocytes have been demonstrated. Today, most of the large pharmaceutical companies have acquired automated electrophysiology robots and have established various automated cardiac ion channel safety screening assays on these, in addition to outsourcing parts of their needs for safety screening. PMID:22131974

Toward Personalized Medicine: Using Cardiomyocytes Differentiated From Urine-Derived Pluripotent Stem Cells to Recapitulate Electrophysiological Characteristics of Type 2 Long QT Syndrome.

PubMed

Jouni, Mariam; Si-Tayeb, Karim; Es-Salah-Lamoureux, Zeineb; Latypova, Xenia; Champon, Benoite; Caillaud, Amandine; Rungoat, Anais; Charpentier, Flavien; Loussouarn, Gildas; Baró, Isabelle; Zibara, Kazem; Lemarchand, Patricia; Gaborit, Nathalie

2015-09-01

Human genetically inherited cardiac diseases have been studied mainly in heterologous systems or animal models, independent of patients' genetic backgrounds. Because sources of human cardiomyocytes (CMs) are extremely limited, the use of urine samples to generate induced pluripotent stem cell-derived CMs would be a noninvasive method to identify cardiac dysfunctions that lead to pathologies within patients' specific genetic backgrounds. The objective was to validate the use of CMs differentiated from urine-derived human induced pluripotent stem (UhiPS) cells as a new cellular model for studying patients' specific arrhythmia mechanisms. Cells obtained from urine samples of a patient with long QT syndrome who harbored the HERG A561P gene mutation and his asymptomatic noncarrier mother were reprogrammed using the episomal-based method. UhiPS cells were then differentiated into CMs using the matrix sandwich method.UhiPS-CMs showed proper expression of atrial and ventricular myofilament proteins and ion channels. They were electrically functional, with nodal-, atrial- and ventricular-like action potentials recorded using high-throughput optical and patch-clamp techniques. Comparison of HERG expression from the patient's UhiPS-CMs to the mother's UhiPS-CMs showed that the mutation led to a trafficking defect that resulted in reduced delayed rectifier K(+) current (IKr). This phenotype gave rise to action potential prolongation and arrhythmias. UhiPS cells from patients carrying ion channel mutations can be used as novel tools to differentiate functional CMs that recapitulate cardiac arrhythmia phenotypes. © 2015 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Ion channel-transporter interactions

PubMed Central

Neverisky, Daniel L.; Abbott, Geoffrey W.

2016-01-01

All living cells require membrane proteins that act as conduits for the regulated transport of ions, solutes and other small molecules across the cell membrane. Ion channels provide a pore that permits often rapid, highly selective, and tightly regulated movement of ions down their electrochemical gradient. In contrast, active transporters can move moieties up their electrochemical gradient. The secondary active transporters (such as SLC superfamily solute transporters) achieve this by coupling uphill movement of the substrate to downhill movement of another ion, such as sodium. The primary active transporters (including H+/K+-ATPases and Na+/K+-ATPases) utilize ATP hydrolysis as an energy source to power uphill transport. It is well known that proteins in each of these classes work in concert with members of the other classes to ensure, for example, ion homeostasis, ion secretion, and restoration of ion balance following action potentials. More recently, evidence is emerging of direct physical interaction between true ion channels, and some primary or secondary active transporters. Here, we review the first known members of this new class of macromolecular complexes that we term “chansporters”, explore their biological roles, and discuss the pathophysiological consequences of their disruption. We compare functional and/or physical interactions between the ubiquitous KCNQ1 potassium channel and various active transporters, and examine other newly discovered chansporter complexes that suggest we may be seeing the tip of the iceberg in a newly emerging signaling modality. PMID:27098917

All-d-Enantiomer of β-Amyloid Peptide Forms Ion Channels in Lipid Bilayers

PubMed Central

2012-01-01

Alzheimer’s disease (AD) is the most common type of senile dementia in aging populations. Amyloid β (Aβ)-mediated dysregulation of ionic homeostasis is the prevailing underlying mechanism leading to synaptic degeneration and neuronal death. Aβ-dependent ionic dysregulation most likely occurs either directly via unregulated ionic transport through the membrane or indirectly via Aβ binding to cell membrane receptors and subsequent opening of existing ion channels or transporters. Receptor binding is expected to involve a high degree of stereospecificity. Here, we investigated whether an Aβ peptide enantiomer, whose entire sequence consists of d-amino acids, can form ion-conducting channels; these channels can directly mediate Aβ effects even in the absence of receptor–peptide interactions. Using complementary approaches of planar lipid bilayer (PLB) electrophysiological recordings and molecular dynamics (MD) simulations, we show that the d-Aβ isomer exhibits ion conductance behavior in the bilayer indistinguishable from that described earlier for the l-Aβ isomer. The d isomer forms channel-like pores with heterogeneous ionic conductance similar to the l-Aβ isomer channels, and the d-isomer channel conductance is blocked by Zn2+, a known blocker of l-Aβ isomer channels. MD simulations further verify formation of β-barrel-like Aβ channels with d- and l-isomers, illustrating that both d- and l-Aβ barrels can conduct cations. The calculated values of the single-channel conductance are approximately in the range of the experimental values. These findings are in agreement with amyloids forming Ca2+ leaking, unregulated channels in AD, and suggest that Aβ toxicity is mediated through a receptor-independent, nonstereoselective mechanism. PMID:22423218

Regulation of Ion Channels by Pyridine Nucleotides

PubMed Central

Kilfoil, Peter J.; Tipparaju, Srinivas M.; Barski, Oleg A.; Bhatnagar, Aruni

2014-01-01

Recent research suggests that in addition to their role as soluble electron carriers, pyridine nucleotides [NAD(P)(H)] also regulate ion transport mechanisms. This mode of regulation seems to have been conserved through evolution. Several bacterial ion–transporting proteins or their auxiliary subunits possess nucleotide-binding domains. In eukaryotes, the Kv1 and Kv4 channels interact with pyridine nucleotide–binding β-subunits that belong to the aldo-keto reductase superfamily. Binding of NADP+ to Kvβ removes N-type inactivation of Kv currents, whereas NADPH stabilizes channel inactivation. Pyridine nucleotides also regulate Slo channels by interacting with their cytosolic regulator of potassium conductance domains that show high sequence homology to the bacterial TrkA family of K+ transporters. These nucleotides also have been shown to modify the activity of the plasma membrane KATP channels, the cystic fibrosis transmembrane conductance regulator, the transient receptor potential M2 channel, and the intracellular ryanodine receptor calcium release channels. In addition, pyridine nucleotides also modulate the voltage-gated sodium channel by supporting the activity of its ancillary subunit—the glycerol-3-phosphate dehydrogenase-like protein. Moreover, the NADP+ metabolite, NAADP+, regulates intracellular calcium homeostasis via the 2-pore channel, ryanodine receptor, or transient receptor potential M2 channels. Regulation of ion channels by pyridine nucleotides may be required for integrating cell ion transport to energetics and for sensing oxygen levels or metabolite availability. This mechanism also may be an important component of hypoxic pulmonary vasoconstriction, memory, and circadian rhythms, and disruption of this regulatory axis may be linked to dysregulation of calcium homeostasis and cardiac arrhythmias. PMID:23410881

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

A Cytosolic Amphiphilic α-Helix Controls the Activity of the Bile Acid-sensitive Ion Channel (BASIC)*

PubMed Central

Schmidt, Axel; Löhrer, Daniel; Alsop, Richard J.; Lenzig, Pia; Oslender-Bujotzek, Adrienne; Wirtz, Monika; Rheinstädter, Maikel C.; Gründer, Stefan; Wiemuth, Dominik

2016-01-01

The bile acid-sensitive ion channel (BASIC) is a member of the degenerin/epithelial Na+ channel (Deg/ENaC) family of ion channels. It is mainly found in bile duct epithelial cells, the intestinal tract, and the cerebellum and is activated by alterations of its membrane environment. Bile acids, one class of putative physiological activators, exert their effect by changing membrane properties, leading to an opening of the channel. The physiological function of BASIC, however, is unknown. Deg/ENaC channels are characterized by a trimeric subunit composition. Each subunit is composed of two transmembrane segments, which are linked by a large extracellular domain. The termini of the channels protrude into the cytosol. Many Deg/ENaC channels contain regulatory domains and sequence motifs within their cytosolic domains. In this study, we show that BASIC contains an amphiphilic α-helical structure within its N-terminal domain. This α-helix binds to the cytosolic face of the plasma membrane and stabilizes a closed state. Truncation of this domain renders the channel hyperactive. Collectively, we identify a cytoplasmic domain, unique to BASIC, that controls channel activity via membrane interaction. PMID:27679529

A Cytosolic Amphiphilic α-Helix Controls the Activity of the Bile Acid-sensitive Ion Channel (BASIC).

PubMed

Schmidt, Axel; Löhrer, Daniel; Alsop, Richard J; Lenzig, Pia; Oslender-Bujotzek, Adrienne; Wirtz, Monika; Rheinstädter, Maikel C; Gründer, Stefan; Wiemuth, Dominik

2016-11-18

The bile acid-sensitive ion channel (BASIC) is a member of the degenerin/epithelial Na + channel (Deg/ENaC) family of ion channels. It is mainly found in bile duct epithelial cells, the intestinal tract, and the cerebellum and is activated by alterations of its membrane environment. Bile acids, one class of putative physiological activators, exert their effect by changing membrane properties, leading to an opening of the channel. The physiological function of BASIC, however, is unknown. Deg/ENaC channels are characterized by a trimeric subunit composition. Each subunit is composed of two transmembrane segments, which are linked by a large extracellular domain. The termini of the channels protrude into the cytosol. Many Deg/ENaC channels contain regulatory domains and sequence motifs within their cytosolic domains. In this study, we show that BASIC contains an amphiphilic α-helical structure within its N-terminal domain. This α-helix binds to the cytosolic face of the plasma membrane and stabilizes a closed state. Truncation of this domain renders the channel hyperactive. Collectively, we identify a cytoplasmic domain, unique to BASIC, that controls channel activity via membrane interaction. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Channel function reconstitution and re-animation: a single-channel strategy in the postcrystal age

PubMed Central

Oiki, Shigetoshi

2015-01-01

The most essential properties of ion channels for their physiologically relevant functions are ion-selective permeation and gating. Among the channel species, the potassium channel is primordial and the most ubiquitous in the biological world, and knowledge of this channel underlies the understanding of features of other ion channels. The strategy applied to studying channels changed dramatically after the crystal structure of the potassium channel was resolved. Given the abundant structural information available, we exploited the bacterial KcsA potassium channel as a simple model channel. In the postcrystal age, there are two effective frameworks with which to decipher the functional codes present in the channel structure, namely reconstitution and re-animation. Complex channel proteins are decomposed into essential functional components, and well-examined parts are rebuilt for integrating channel function in the membrane (reconstitution). Permeation and gating are dynamic operations, and one imagines the active channel by breathing life into the ‘frozen’ crystal (re-animation). Capturing the motion of channels at the single-molecule level is necessary to characterize the behaviour of functioning channels. Advanced techniques, including diffracted X-ray tracking, lipid bilayer methods and high-speed atomic force microscopy, have been used. Here, I present dynamic pictures of the KcsA potassium channel from the submolecular conformational changes to the supramolecular collective behaviour of channels in the membrane. These results form an integrated picture of the active channel and offer insights into the processes underlying the physiological function of the channel in the cell membrane. PMID:25833254

Channel function reconstitution and re-animation: a single-channel strategy in the postcrystal age.

PubMed

Oiki, Shigetoshi

2015-06-15

The most essential properties of ion channels for their physiologically relevant functions are ion-selective permeation and gating. Among the channel species, the potassium channel is primordial and the most ubiquitous in the biological world, and knowledge of this channel underlies the understanding of features of other ion channels. The strategy applied to studying channels changed dramatically after the crystal structure of the potassium channel was resolved. Given the abundant structural information available, we exploited the bacterial KcsA potassium channel as a simple model channel. In the postcrystal age, there are two effective frameworks with which to decipher the functional codes present in the channel structure, namely reconstitution and re-animation. Complex channel proteins are decomposed into essential functional components, and well-examined parts are rebuilt for integrating channel function in the membrane (reconstitution). Permeation and gating are dynamic operations, and one imagines the active channel by breathing life into the 'frozen' crystal (re-animation). Capturing the motion of channels at the single-molecule level is necessary to characterize the behaviour of functioning channels. Advanced techniques, including diffracted X-ray tracking, lipid bilayer methods and high-speed atomic force microscopy, have been used. Here, I present dynamic pictures of the KcsA potassium channel from the submolecular conformational changes to the supramolecular collective behaviour of channels in the membrane. These results form an integrated picture of the active channel and offer insights into the processes underlying the physiological function of the channel in the cell membrane. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

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.

Multi-channel retarding field analyzer for EAST

NASA Astrophysics Data System (ADS)

M, HENKEL; D, HÖSCHEN; Y, LIANG; Y, LI; S, C. LIU; D, NICOLAI; N, SANDRI; G, SATHEESWARAN; N, YAN; H, X. ZHANG; the EAST, team2

2018-05-01

A multi-channel retarding field analyzer (MC-RFA) including two RFA modules and two Langmuir probes to measure the ion and electron temperature profiles within the scrape-off layer was developed for investigations of the interplay between magnetic topology and plasma transport at the plasma boundary. The MC-RFA probe for the stellarator W7-X and first measurements at the tokamak EAST was designed. The probe head allows simultaneous multi-channel ion temperature as well as for electron temperature measurements. The usability for radial correlation measurements of the measured ion currents is also given.

Near-membrane electric field calcium ion dehydration.

PubMed

Barger, James P; Dillon, Patrick F

2016-12-01

The dehydration of ion-water complexes prior to ion channel transit has focused on channel protein-mediated dissociation of water. Ion dehydration by the membrane electric field has not previously been considered. Near membrane electric fields have previously been shown to cause the disassociation of non-covalently bound small molecule-small molecule, small molecule-protein, and protein-protein complexes. It is well known that cosmotropic, structure making ions such as calcium and sodium significantly bind multiple water ions in solution. It is also known that these ions are often not hydrated as they pass through membrane ion channels. Using capillary electrophoresis, the range of electric fields needed to strip water molecules from calcium ions has been measured. Ion migration velocity is a linear function of the electric field. At low electric fields, the migration rate of calcium ion was shown to be linearly related to the applied electric field. Using a form of the Stoke's equation applicable to ion migration, the hydrated calcium radius was found to be 0.334nm, corresponding to a water hydration shell of 5.09 water molecules. At higher electric fields, the slope of the calcium migration velocity as a function of the electric field increased, which was modeled as a decrease in the radius of the migrating ion as the water was removed. Using a tanh function to model the transition of the ion from a hydrated to a stripped state, the transition had a midpoint at 446V/cm, and was 88% complete at 587V/cm with a correlation coefficient of 0.9996. The migration velocity of the stripped calcium ion was found to be a function of both the decrease in radius and an increase in the effective, electronic viscosity of the dipole medium through which the dehydrated ion moved. The size of the electric field needed to dehydrate calcium occurs 6-7nm from the cell membrane. Calcium ions within this distance from the membrane will be devoid of water molecules when they reach the calcium selective channel pore entrances, all known to be approximately 1-2nm from the membrane. No matter what the calcium pore structure, calcium ions reaching the channel entrance will be devoid of a water shell. Copyright © 2016 Elsevier Ltd. All rights reserved.

Channeling technique to make nanoscale ion beams

NASA Astrophysics Data System (ADS)

Biryukov, V. M.; Bellucci, S.; Guidi, V.

2005-04-01

Particle channeling in a bent crystal lattice has led to an efficient instrument for beam steering at accelerators [Biryukov et al., Crystal Channeling and its Application at High Energy Accelerators, Springer, Berlin, 1997], demonstrated from MeV to TeV energies. In particular, crystal focusing of high-energy protons to micron size has been demonstrated at IHEP with the results well in match with Lindhard (critical angle) prediction. Channeling in crystal microstructures has been proposed as a unique source of a microbeam of high-energy particles [Bellucci et al., Phys. Rev. ST Accel. Beams 6 (2003) 033502]. Channeling in nanostructures (single-wall and multi-wall nanotubes) offers the opportunities to produce ion beams on nanoscale. Particles channeled in a nanotube (with typical diameter of about 1 nm) are trapped in two dimensions and can be steered (deflected, focused) with the efficiency similar to that of crystal channeling or better. This technique has been a subject of computer simulations, with experimental efforts under way in several high-energy labs, including IHEP. We present the theoretical outlook for making channeling-based nanoscale ion beams and report the experience with crystal-focused microscale proton beams.

Lipid microdomains and the regulation of ion channel function

PubMed Central

Dart, Caroline

2010-01-01

Many types of ion channel localize to cholesterol and sphingolipid-enriched regions of the plasma membrane known as lipid microdomains or ‘rafts’. The precise physiological role of these unique lipid microenvironments remains elusive due largely to difficulties associated with studying these potentially extremely small and dynamic domains. Nevertheless, increasing evidence suggests that membrane rafts regulate channel function in a number of different ways. Raft-enriched lipids such as cholesterol and sphingolipids exert effects on channel activity either through direct protein–lipid interactions or by influencing the physical properties of the bilayer. Rafts also appear to selectively recruit interacting signalling molecules to generate subcellular compartments that may be important for efficient and selective signal transduction. Direct interaction with raft-associated scaffold proteins such as caveolin can also influence channel function by altering gating kinetics or by affecting trafficking and surface expression. Selective association of ion channels with specific lipid microenvironments within the membrane is thus likely to be an important and fundamental regulatory aspect of channel physiology. This brief review highlights some of the existing evidence for raft modulation of channel function. PMID:20519314

Ca2+-dependent phospholipid scrambling by a reconstituted TMEM16 ion channel.

PubMed

Malvezzi, Mattia; Chalat, Madhavan; Janjusevic, Radmila; Picollo, Alessandra; Terashima, Hiroyuki; Menon, Anant K; Accardi, Alessio

2013-01-01

Phospholipid (PL) scramblases disrupt the lipid asymmetry of the plasma membrane, externalizing phosphatidylserine to trigger blood coagulation and mark apoptotic cells. Recently, members of the TMEM16 family of Ca(2+)-gated channels have been shown to be involved in Ca(2+)-dependent scrambling. It is however controversial whether they are scramblases or channels regulating scrambling. Here we show that purified afTMEM16, from Aspergillus fumigatus, is a dual-function protein: it is a Ca(2+)-gated channel, with characteristics of other TMEM16 homologues, and a Ca(2+)-dependent scramblase, with the expected properties of mammalian PL scramblases. Remarkably, we find that a single Ca(2+) site regulates separate transmembrane pathways for ions and lipids. Two other purified TMEM16-channel homologues do not mediate scrambling, suggesting that the family diverged into channels and channel/scramblases. We propose that the spatial separation of the ion and lipid pathways underlies the evolutionary divergence of the TMEM16 family, and that other homologues, such as TMEM16F, might also be dual-function channel/scramblases.

Interaction of a dinoflagellate neurotoxin with voltage-activated ion channels in a marine diatom.

PubMed

Kitchen, Sheila A; Bourdelais, Andrea J; Taylor, Alison R

2018-01-01

The potent neurotoxins produced by the harmful algal bloom species Karenia brevis are activators of sodium voltage-gated channels (VGC) in animals, resulting in altered channel kinetics and membrane hyperexcitability. Recent biophysical and genomic evidence supports widespread presence of homologous sodium (Na + ) and calcium (Ca 2+ ) permeable VGCs in unicellular algae, including marine phytoplankton. We therefore hypothesized that VGCs of these phytoplankton may be an allelopathic target for waterborne neurotoxins produced by K. brevis blooms that could lead to ion channel dysfunction and disruption of signaling in a similar manner to animal Na + VGCs. We examined the interaction of brevetoxin-3 (PbTx-3), a K. brevis neurotoxin, with the Na + /Ca 2+ VGC of the non-toxic diatom Odontella sinensi s using electrophysiology. Single electrode current- and voltage- clamp recordings from O. sinensis in the presence of PbTx-3 were used to examine the toxin's effect on voltage gated Na + /Ca 2+ currents. In silico analysis was used to identify the putative PbTx binding site in the diatoms. We identified Na + /Ca 2+ VCG homologs from the transcriptomes and genomes of 12 diatoms, including three transcripts from O. sinensis and aligned them with site-5 of Na + VGCs, previously identified as the PbTx binding site in animals. Up to 1 µM PbTx had no effect on diatom resting membrane potential or membrane excitability. The kinetics of fast inward Na + /Ca 2+ currents that underlie diatom action potentials were also unaffected. However, the peak inward current was inhibited by 33%, delayed outward current was inhibited by 25%, and reversal potential of the currents shifted positive, indicating a change in permeability of the underlying channels. Sequence analysis showed a lack of conservation of the PbTx binding site in diatom VGC homologs, many of which share molecular features more similar to single-domain bacterial Na + /Ca 2+ VGCs than the 4-domain eukaryote channels. Although membrane excitability and the kinetics of action potential currents were unaffected, the permeation of the channels underlying the diatom action potential was significantly altered in the presence of PbTx-3. However, at environmentally relevant concentrations the effects of PbTx- on diatom voltage activated currents and interference of cell signaling through this pathway may be limited. The relative insensitivity of phytoplankton VGCs may be due to divergence of site-5 (the putative PbTx binding site), and in some cases, such as O. sinensis , resistance to toxin effects may be because of evolutionary loss of the 4-domain eukaryote channel, while retaining a single domain bacterial-like VGC that can substitute in the generation of fast action potentials.

The Transport of Salt and Water across Isolated Rat Ileum

PubMed Central

Clarkson, T. W.

1967-01-01

The flows of sodium, potassium, and chloride under electrical and chemical gradients and of salt and water in the presence of osmotic pressure gradients are described by phenomenological equations based on the thermodynamics of irreversible processes. The aim was to give the simplest possible description, that is to postulate the least number of active transport processes and the least number of separate pathways across the intestine. On this basis, the results were consistent with the following picture of the intestine: Two channels exist across this tissue, one allowing only passive transport of ions and the other only active. In the passive channel, the predominant resistance to ion flow is friction with the water in the channel. The electroosmotic flow indicates that the passive channel is lined with negative fixed charged groups having a surface charge density of 3000 esu cm-2. The values of the ion-water frictional coefficients, and the relationship between ionic concentrations and flows indicate that the passive channel is extracellular. The active channel behaves as two membranes in series, the first membrane being semipermeable but allowing active transport of sodium, and the second membrane being similar to the passive channel. Friction with the ions in the second "membrane" is the predominant resistance to water flow. PMID:11526854

Testing the Applicability of Nernst-Planck Theory in Ion Channels: Comparisons with Brownian Dynamics Simulations

PubMed Central

Song, Chen; Corry, Ben

2011-01-01

The macroscopic Nernst-Planck (NP) theory has often been used for predicting ion channel currents in recent years, but the validity of this theory at the microscopic scale has not been tested. In this study we systematically tested the ability of the NP theory to accurately predict channel currents by combining and comparing the results with those of Brownian dynamics (BD) simulations. To thoroughly test the theory in a range of situations, calculations were made in a series of simplified cylindrical channels with radii ranging from 3 to 15 Å, in a more complex ‘catenary’ channel, and in a realistic model of the mechanosensitive channel MscS. The extensive tests indicate that the NP equation is applicable in narrow ion channels provided that accurate concentrations and potentials can be input as the currents obtained from the combination of BD and NP match well with those obtained directly from BD simulations, although some discrepancies are seen when the ion concentrations are not radially uniform. This finding opens a door to utilising the results of microscopic simulations in continuum theory, something that is likely to be useful in the investigation of a range of biophysical and nano-scale applications and should stimulate further studies in this direction. PMID:21731672

Testing the applicability of Nernst-Planck theory in ion channels: comparisons with Brownian dynamics simulations.

PubMed

Song, Chen; Corry, Ben

2011-01-01

The macroscopic Nernst-Planck (NP) theory has often been used for predicting ion channel currents in recent years, but the validity of this theory at the microscopic scale has not been tested. In this study we systematically tested the ability of the NP theory to accurately predict channel currents by combining and comparing the results with those of Brownian dynamics (BD) simulations. To thoroughly test the theory in a range of situations, calculations were made in a series of simplified cylindrical channels with radii ranging from 3 to 15 Å, in a more complex 'catenary' channel, and in a realistic model of the mechanosensitive channel MscS. The extensive tests indicate that the NP equation is applicable in narrow ion channels provided that accurate concentrations and potentials can be input as the currents obtained from the combination of BD and NP match well with those obtained directly from BD simulations, although some discrepancies are seen when the ion concentrations are not radially uniform. This finding opens a door to utilising the results of microscopic simulations in continuum theory, something that is likely to be useful in the investigation of a range of biophysical and nano-scale applications and should stimulate further studies in this direction.

Identification and characterization of a bacterial hydrosulphide ion channel

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

Czyzewski, Bryan K.; Wang, Da-Neng

2012-10-26

The hydrosulphide ion (HS{sup -}) and its undissociated form, hydrogen sulphide (H{sub 2}S), which are believed to have been critical to the origin of life on Earth, remain important in physiology and cellular signalling. As a major metabolite in anaerobic bacterial growth, hydrogen sulphide is a product of both assimilatory and dissimilatory sulphate reduction. These pathways can reduce various oxidized sulphur compounds including sulphate, sulphite and thiosulphate. The dissimilatory sulphate reduction pathway uses this molecule as the terminal electron acceptor for anaerobic respiration, in which process it produces excess amounts of H{sub 2}S. The reduction of sulphite is a keymore » intermediate step in all sulphate reduction pathways. In Clostridium and Salmonella, an inducible sulphite reductase is directly linked to the regeneration of NAD{sup +}, which has been suggested to have a role in energy production and growth, as well as in the detoxification of sulphite. Above a certain concentration threshold, both H{sub 2}S and HS{sup -} inhibit cell growth by binding the metal centres of enzymes and cytochrome oxidase, necessitating a release mechanism for the export of this toxic metabolite from the cell. Here we report the identification of a hydrosulphide ion channel in the pathogen Clostridium difficile through a combination of genetic, biochemical and functional approaches. The HS{sup -} channel is a member of the formate/nitrite transport family, in which about 50 hydrosulphide ion channels form a third subfamily alongside those for formate (FocA) and for nitrite (NirC). The hydrosulphide ion channel is permeable to formate and nitrite as well as to HS{sup -} ions. Such polyspecificity can be explained by the conserved ion selectivity filter observed in the channel's crystal structure. The channel has a low open probability and is tightly regulated, to avoid decoupling of the membrane proton gradient.« less

Probing pore constriction in a ligand-gated ion channel by trapping a metal ion in the pore upon agonist dissociation.

PubMed

Pittel, Ilya; Witt-Kehati, Dvora; Degani-Katzav, Nurit; Paas, Yoav

2010-08-20

Eukaryotic pentameric ligand-gated ion channels (pLGICs) are receptors activated by neurotransmitters to rapidly transport ions across cell membranes, down their electrochemical gradients. Recent crystal structures of two prokaryotic pLGICs were interpreted to imply that the extracellular side of the transmembrane pore constricts to close the channel (Hilf, R. J., and Dutzler, R. (2009) Nature 457, 115-118; Bocquet, N., Nury, H., Baaden, M., Le Poupon, C., Changeux, J. P., Delarue, M., and Corringer, P. J. (2009) Nature 457, 111-114). Here, we utilized a eukaryotic acetylcholine (ACh)-serotonin chimeric pLGIC that was engineered with histidines to coordinate a metal ion within the channel pore, at its cytoplasmic side. In a previous study, the access of Zn(2+) ions to the engineered histidines had been explored when the channel was either at rest (closed) or active (open) (Paas, Y., Gibor, G., Grailhe, R., Savatier-Duclert, N., Dufresne, V., Sunesen, M., de Carvalho, L. P., Changeux, J. P., and Attali, B. (2005) Proc. Natl. Acad. Sci. U.S.A. 102, 15877-15882). In this study, the interactions of Zn(2+) with the pore were probed upon agonist (ACh) dissociation that triggers the transition of the receptor from the active conformation to the resting conformation (i.e. during deactivation). Application of Zn(2+) onto ACh-bound open receptors obstructed their pore and prevented ionic flow. Removing ACh from its extracellular binding sites to trigger deactivation while Zn(2+) is still bound led to tight trapping of Zn(2+) within the pore. Together with single-channel recordings, made to explore single pore-blocking events, we show that dissociation of ACh causes the gate to shut on a Zn(2+) ion that effectively acts as a "foot in the door." We infer that, upon deactivation, the cytoplasmic side of the pore of the ACh-serotonin receptor chimera constricts to close the channel.

A uniquely adaptable pore is consistent with NALCN being an ion sensor

PubMed Central

Senatore, Adriano; Spafford, J. David

2013-01-01

NALCN is an intriguing, orphan ion channel among the 4x6TM family of related voltage-gated cation channels, sharing a common architecture of four homologous domains consisting of six transmembrane helices, separated by three cytoplasmic linkers and delimited by N and C-terminal ends. NALCN is one of the shortest 4x6TM family members, lacking much of the variation that provides the diverse palate of gating features, and tissue specific adaptations of sodium and calcium channels. NALCN’s most distinctive feature is that that it possesses a highly adaptable pore with a calcium-like EEEE selectivity filter in radially symmetrical animals and a more sodium-like EEKE or EKEE selectivity filter in bilaterally symmetrical animals including vertebrates. Two lineages of animals evolved alternative calcium-like EEEE and sodium-like EEKE / EKEE pores, spliced to regulate NALCN functions in differing cellular environments, such as muscle (heart and skeletal) and secretory tissue (brain and glands), respectively. A highly adaptable pore in an otherwise conserved ion channel in the 4x6TM channel family is not consistent with a role for NALCN in directly gating a significant ion conductance that can be either sodium ions or calcium ions. NALCN was proposed to be an expressible Gd3+-sensitive, NMDG+-impermeant, non-selective and ohmic leak conductance in HEK-293T cells, but we were unable to distinguish these reported currents from leaky patch currents (ILP) in control HEK-293T cells. We suggest that NALCN functions as a sensor for the much larger UNC80/UNC79 complex, in a manner consistent with the coupling mechanism known for other weakly or non-conducting 4x6TM channel sensor proteins such as Nax or Cav1.1. We propose that NALCN serves as a variable sensor that responds to calcium or sodium ion flux, depending on whether the total cellular current density is generated more from calcium-selective or sodium-selective channels. PMID:23442378

A uniquely adaptable pore is consistent with NALCN being an ion sensor.

PubMed

Senatore, Adriano; Spafford, J David

2013-01-01

NALCN is an intriguing, orphan ion channel among the 4x6TM family of related voltage-gated cation channels, sharing a common architecture of four homologous domains consisting of six transmembrane helices, separated by three cytoplasmic linkers and delimited by N and C-terminal ends. NALCN is one of the shortest 4x6TM family members, lacking much of the variation that provides the diverse palate of gating features, and tissue specific adaptations of sodium and calcium channels. NALCN's most distinctive feature is that that it possesses a highly adaptable pore with a calcium-like EEEE selectivity filter in radially symmetrical animals and a more sodium-like EEKE or EKEE selectivity filter in bilaterally symmetrical animals including vertebrates. Two lineages of animals evolved alternative calcium-like EEEE and sodium-like EEKE / EKEE pores, spliced to regulate NALCN functions in differing cellular environments, such as muscle (heart and skeletal) and secretory tissue (brain and glands), respectively. A highly adaptable pore in an otherwise conserved ion channel in the 4x6TM channel family is not consistent with a role for NALCN in directly gating a significant ion conductance that can be either sodium ions or calcium ions. NALCN was proposed to be an expressible Gd ( 3+) -sensitive, NMDG (+) -impermeant, non-selective and ohmic leak conductance in HEK-293T cells, but we were unable to distinguish these reported currents from leaky patch currents (ILP) in control HEK-293T cells. We suggest that NALCN functions as a sensor for the much larger UNC80/UNC79 complex, in a manner consistent with the coupling mechanism known for other weakly or non-conducting 4x6TM channel sensor proteins such as Nax or Cav 1.1. We propose that NALCN serves as a variable sensor that responds to calcium or sodium ion flux, depending on whether the total cellular current density is generated more from calcium-selective or sodium-selective channels.

Chemoselective tarantula toxins report voltage activation of wild-type ion channels in live cells

PubMed Central

Tilley, Drew C.; Eum, Kenneth S.; Fletcher-Taylor, Sebastian; Austin, Daniel C.; Dupré, Christophe; Patrón, Lilian A.; Garcia, Rita L.; Lam, Kit; Yarov-Yarovoy, Vladimir; Cohen, Bruce E.; Sack, Jon T.

2014-01-01

Electrically excitable cells, such as neurons, exhibit tremendous diversity in their firing patterns, a consequence of the complex collection of ion channels present in any specific cell. Although numerous methods are capable of measuring cellular electrical signals, understanding which types of ion channels give rise to these signals remains a significant challenge. Here, we describe exogenous probes which use a novel mechanism to report activity of voltage-gated channels. We have synthesized chemoselective derivatives of the tarantula toxin guangxitoxin-1E (GxTX), an inhibitory cystine knot peptide that binds selectively to Kv2-type voltage gated potassium channels. We find that voltage activation of Kv2.1 channels triggers GxTX dissociation, and thus GxTX binding dynamically marks Kv2 activation. We identify GxTX residues that can be replaced by thiol- or alkyne-bearing amino acids, without disrupting toxin folding or activity, and chemoselectively ligate fluorophores or affinity probes to these sites. We find that GxTX–fluorophore conjugates colocalize with Kv2.1 clusters in live cells and are released from channels activated by voltage stimuli. Kv2.1 activation can be detected with concentrations of probe that have a trivial impact on cellular currents. Chemoselective GxTX mutants conjugated to dendrimeric beads likewise bind live cells expressing Kv2.1, and the beads are released by channel activation. These optical sensors of conformational change are prototype probes that can indicate when ion channels contribute to electrical signaling. PMID:25331865

Density-dependent changes of the pore properties of the P2X2 receptor channel

PubMed Central

Fujiwara, Yuichiro; Kubo, Yoshihiro

2004-01-01

Ligand-gated ion channels underlie and play important roles in synaptic transmission, and it is generally accepted that the ion channel pores have a rigid structure that enables strict regulation of ion permeation. One exception is the P2X ATP-gated channel. After application of ATP, the ion selectivity of the P2X2 channel time-dependently changes, i.e. permeability to large cations gradually increases, and there is significant cell-to-cell variation in the intensity of inward rectification. Here we show P2X2 channel properties are correlated with the expression level: increasing P2X2 expression level in oocytes increases permeability to large cations, decreases inward rectification and increases ligand sensitivity. We also observed that the inward rectification changed in a dose-dependent manner, i.e. when low concentration of ATP was applied to an oocyte with a high expression level, the intensity of inward rectification of the evoked current was weak. Taken together, these results show that the pore properties of P2X2 channel are not static but change dynamically depending on the open channel density. Furthermore, we identified by mutagenesis study that Ile328 located at the outer mouth of the pore is critical for the density-dependent changes of P2X2. Our findings suggest synaptic transmission can be modulated by the local density-dependent changes of channel properties caused, for example, by the presence of clustering molecules. PMID:15107474

Monitoring ion-channel function in real time through quantum decoherence

PubMed Central

Hall, Liam T.; Hill, Charles D.; Cole, Jared H.; Städler, Brigitte; Caruso, Frank; Mulvaney, Paul; Wrachtrup, Jörg; Hollenberg, Lloyd C. L.

2010-01-01

In drug discovery, there is a clear and urgent need for detection of cell-membrane ion-channel operation with wide-field capability. Existing techniques are generally invasive or require specialized nanostructures. We show that quantum nanotechnology could provide a solution. The nitrogen-vacancy (NV) center in nanodiamond is of great interest as a single-atom quantum probe for nanoscale processes. However, until now nothing was known about the quantum behavior of a NV probe in a complex biological environment. We explore the quantum dynamics of a NV probe in proximity to the ion channel, lipid bilayer, and surrounding aqueous environment. Our theoretical results indicate that real-time detection of ion-channel operation at millisecond resolution is possible by directly monitoring the quantum decoherence of the NV probe. With the potential to scan and scale up to an array-based system, this conclusion may have wide-ranging implications for nanoscale biology and drug discovery. PMID:20937908

Seeking structural specificity: direct modulation of pentameric ligand-gated ion channels by alcohols and general anesthetics.

PubMed

Howard, Rebecca J; Trudell, James R; Harris, R Adron

2014-01-01

Alcohols and other anesthetic agents dramatically alter neurologic function in a wide range of organisms, yet their molecular sites of action remain poorly characterized. Pentameric ligand-gated ion channels, long implicated in important direct effects of alcohol and anesthetic binding, have recently been illuminated in renewed detail thanks to the determination of atomic-resolution structures of several family members from lower organisms. These structures provide valuable models for understanding and developing anesthetic agents and for allosteric modulation in general. This review surveys progress in this field from function to structure and back again, outlining early evidence for relevant modulation of pentameric ligand-gated ion channels and the development of early structural models for ion channel function and modulation. We highlight insights and challenges provided by recent crystal structures and resulting simulations, as well as opportunities for translation of these newly detailed models back to behavior and therapy.

Electronic control of H+ current in a bioprotonic device with Gramicidin A and Alamethicin

NASA Astrophysics Data System (ADS)

Hemmatian, Zahra; Keene, Scott; Josberger, Erik; Miyake, Takeo; Arboleda, Carina; Soto-Rodríguez, Jessica; Baneyx, François; Rolandi, Marco

2016-10-01

In biological systems, intercellular communication is mediated by membrane proteins and ion channels that regulate traffic of ions and small molecules across cell membranes. A bioelectronic device with ion channels that control ionic flow across a supported lipid bilayer (SLB) should therefore be ideal for interfacing with biological systems. Here, we demonstrate a biotic-abiotic bioprotonic device with Pd contacts that regulates proton (H+) flow across an SLB incorporating the ion channels Gramicidin A (gA) and Alamethicin (ALM). We model the device characteristics using the Goldman-Hodgkin-Katz (GHK) solution to the Nernst-Planck equation for transport across the membrane. We derive the permeability for an SLB integrating gA and ALM and demonstrate pH control as a function of applied voltage and membrane permeability. This work opens the door to integrating more complex H+ channels at the Pd contact interface to produce responsive biotic-abiotic devices with increased functionality.

Monitoring ion-channel function in real time through quantum decoherence.

PubMed

Hall, Liam T; Hill, Charles D; Cole, Jared H; Städler, Brigitte; Caruso, Frank; Mulvaney, Paul; Wrachtrup, Jörg; Hollenberg, Lloyd C L

2010-11-02

In drug discovery, there is a clear and urgent need for detection of cell-membrane ion-channel operation with wide-field capability. Existing techniques are generally invasive or require specialized nanostructures. We show that quantum nanotechnology could provide a solution. The nitrogen-vacancy (NV) center in nanodiamond is of great interest as a single-atom quantum probe for nanoscale processes. However, until now nothing was known about the quantum behavior of a NV probe in a complex biological environment. We explore the quantum dynamics of a NV probe in proximity to the ion channel, lipid bilayer, and surrounding aqueous environment. Our theoretical results indicate that real-time detection of ion-channel operation at millisecond resolution is possible by directly monitoring the quantum decoherence of the NV probe. With the potential to scan and scale up to an array-based system, this conclusion may have wide-ranging implications for nanoscale biology and drug discovery.

Seeking Structural Specificity: Direct Modulation of Pentameric Ligand-Gated Ion Channels by Alcohols and General Anesthetics

PubMed Central

Trudell, James R.; Harris, R. Adron

2014-01-01

Alcohols and other anesthetic agents dramatically alter neurologic function in a wide range of organisms, yet their molecular sites of action remain poorly characterized. Pentameric ligand-gated ion channels, long implicated in important direct effects of alcohol and anesthetic binding, have recently been illuminated in renewed detail thanks to the determination of atomic-resolution structures of several family members from lower organisms. These structures provide valuable models for understanding and developing anesthetic agents and for allosteric modulation in general. This review surveys progress in this field from function to structure and back again, outlining early evidence for relevant modulation of pentameric ligand-gated ion channels and the development of early structural models for ion channel function and modulation. We highlight insights and challenges provided by recent crystal structures and resulting simulations, as well as opportunities for translation of these newly detailed models back to behavior and therapy. PMID:24515646

Beyond voltage-gated ion channels: Voltage-operated membrane proteins and cellular processes.

PubMed

Zhang, Jianping; Chen, Xingjuan; Xue, Yucong; Gamper, Nikita; Zhang, Xuan

2018-04-18

Voltage-gated ion channels were believed to be the only voltage-sensitive proteins in excitable (and some non-excitable) cells for a long time. Emerging evidence indicates that the voltage-operated model is shared by some other transmembrane proteins expressed in both excitable and non-excitable cells. In this review, we summarize current knowledge about voltage-operated proteins, which are not classic voltage-gated ion channels as well as the voltage-dependent processes in cells for which single voltage-sensitive proteins have yet to be identified. Particularly, we will focus on the following. (1) Voltage-sensitive phosphoinositide phosphatases (VSP) with four transmembrane segments homologous to the voltage sensor domain (VSD) of voltage-gated ion channels; VSPs are the first family of proteins, other than the voltage-gated ion channels, for which there is sufficient evidence for the existence of the VSD domain; (2) Voltage-gated proton channels comprising of a single voltage-sensing domain and lacking an identified pore domain; (3) G protein coupled receptors (GPCRs) that mediate the depolarization-evoked potentiation of Ca 2+ mobilization; (4) Plasma membrane (PM) depolarization-induced but Ca 2+ -independent exocytosis in neurons. (5) Voltage-dependent metabolism of phosphatidylinositol 4,5-bisphosphate (PtdIns[4,5]P 2 , PIP 2 ) in the PM. These recent discoveries expand our understanding of voltage-operated processes within cellular membranes. © 2018 Wiley Periodicals, Inc.

Strontium and barium in aqueous solution and a potassium channel binding site

NASA Astrophysics Data System (ADS)

Chaudhari, Mangesh I.; Rempe, Susan B.

2018-06-01

Ion hydration structure and free energy establish criteria for understanding selective ion binding in potassium (K+) ion channels and may be significant to understanding blocking mechanisms as well. Recently, we investigated the hydration properties of Ba2+, the most potent blocker of K+ channels among the simple metal ions. Here, we use a similar method of combining ab initio molecular dynamics simulations, statistical mechanical theory, and electronic structure calculations to probe the fundamental hydration properties of Sr2+, which does not block bacterial K+ channels. The radial distribution of water around Sr2+ suggests a stable 8-fold geometry in the local hydration environment, similar to Ba2+. While the predicted hydration free energy of -331.8 kcal/mol is comparable with the experimental result of -334 kcal/mol, the value is significantly more favorable than the -305 kcal/mol hydration free energy of Ba2+. When placed in the innermost K+ channel blocking site, the solvation free energies and lowest energy structures of both Sr2+ and Ba2+ are nearly unchanged compared with their respective hydration properties. This result suggests that the block is not attributable to ion trapping due to +2 charge, and differences in blocking behavior arise due to free energies associated with the exchange of water ligands for channel ligands instead of free energies of transfer from water to the binding site.

Novel pharmacological targets for the rhythm control management of atrial fibrillation.

PubMed

Burashnikov, Alexander; Antzelevitch, Charles

2011-12-01

Atrial fibrillation (AF) is a growing clinical problem associated with increased morbidity and mortality. Development of safe and effective pharmacological treatments for AF is one of the greatest unmet medical needs facing our society. In spite of significant progress in non-pharmacological AF treatments (largely due to the use of catheter ablation techniques), anti-arrhythmic agents (AADs) remain first line therapy for rhythm control management of AF for most AF patients. When considering efficacy, safety and tolerability, currently available AADs for rhythm control of AF are less than optimal. Ion channel inhibition remains the principal strategy for termination of AF and prevention of its recurrence. Practical clinical experience indicates that multi-ion channel blockers are generally more optimal for rhythm control of AF compared to ion channel-selective blockers. Recent studies suggest that atrial-selective sodium channel block can lead to safe and effective suppression of AF and that concurrent inhibition of potassium ion channels may potentiate this effect. An important limitation of the ion channel block approach for AF treatment is that non-electrical factors (largely structural remodeling) may importantly determine the generation of AF, so that "upstream therapy", aimed at preventing or reversing structural remodeling, may be required for effective rhythm control management. This review focuses on novel pharmacological targets for the rhythm control management of AF. Copyright © 2011 Elsevier Inc. All rights reserved.

Equivalence of two approaches for modeling ion permeation through a transmembrane channel with an internal binding site

NASA Astrophysics Data System (ADS)

Zhou, Huan-Xiang

2011-04-01

Ion permeation through transmembrane channels has traditionally been modeled using two different approaches. In one approach, the translocation of the permeant ion through the channel pore is modeled as continuous diffusion and the rate of ion transport is obtained from solving the steady-state diffusion equation. In the other approach, the translocation of the permeant ion through the pore is modeled as hopping along a discrete set of internal binding sites and the rate of ion transport is obtained from solving a set of steady-state rate equations. In a recent work [Zhou, J. Phys. Chem. Lett. 1, 1973 (2010)], the rate constants for binding to an internal site were further calculated by modeling binding as diffusion-influenced reactions. That work provided the foundation for bridging the two approaches. Here we show that, by representing a binding site as an energy well, the two approaches indeed give the same result for the rate of ion transport.

Gating Behavior of Endoplasmic Reticulum Potassium Channels of Rat Hepatocytes in Diabetes

PubMed Central

Ghasemi, Maedeh; Khodaei, Naser; Salari, Sajjad; Eliassi, Afsaneh; Saghiri, Reza

2014-01-01

Background: 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. Method: 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. Results: 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. Conclusion: 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. PMID:24842143

Energetic and dynamic analysis of transport of Na + and K + through a cyclic peptide nanotube in water and in lipid bilayers

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

Song, Yeonho; Lee, Ji Hye; Hwang, Hoon

Potential of mean force (PMF) profiles and position-dependent diffusion coefficients of Na + and K + are calculated to elucidate the translocation of ions through a cyclic peptide nanotube, composed of 8 × cyclo[-(D-Leu-Trp) 4-] rings, in water and in hydrated DMPC bilayers. The PMF profiles and PMF decomposition analysis for the monovalent cations show that favorable interactions of the cations with the CPN as well as the lipid bilayer and dehydration free energy penalties are two major competing factors which determine the free energy surface for ion transport through CPNs both in water and lipid bilayers, and that themore » selectivity of CPNs to cations mainly arises from favorable interaction energies of cations with CPNs and lipid bilayers that are more dominant than the dehydration penalties. Calculations of the position-dependent diffusion coefficients and dynamic friction kernels of the cations indicate that the dehydration process along with the molecular rearrangements occurring outside the channel and the coupling of the ion motions with the chain-structured water movements inside the channel lead to decrease of the diffusion coefficients far away from the channel entrance and also reduced coefficients inside the channel. Here the PMF and diffusivity profiles for Na + and K + reveal that the energetics of ion transport through the CPN are governed by global interactions of ions with all the components in the system while the diffusivity of ions through the channel is mostly determined by local interactions of ions with the confined water molecules inside the channel. Comparison of Na + and K + ion distributions based on overdamped Brownian dynamics simulations based on the PMF and diffusivity profiles with the corresponding results from molecular dynamics shows good agreement, indicating accuracy of the Bayesian inference method for determining diffusion coefficients in this application. In addition this work shows that position-dependent diffusion coefficients of ions are required to explain the dynamics and conductance of ions through the CPN properly.« less

Energetic and dynamic analysis of transport of Na + and K + through a cyclic peptide nanotube in water and in lipid bilayers

DOE PAGES

Song, Yeonho; Lee, Ji Hye; Hwang, Hoon; ...

2016-11-04

Potential of mean force (PMF) profiles and position-dependent diffusion coefficients of Na + and K + are calculated to elucidate the translocation of ions through a cyclic peptide nanotube, composed of 8 × cyclo[-(D-Leu-Trp) 4-] rings, in water and in hydrated DMPC bilayers. The PMF profiles and PMF decomposition analysis for the monovalent cations show that favorable interactions of the cations with the CPN as well as the lipid bilayer and dehydration free energy penalties are two major competing factors which determine the free energy surface for ion transport through CPNs both in water and lipid bilayers, and that themore » selectivity of CPNs to cations mainly arises from favorable interaction energies of cations with CPNs and lipid bilayers that are more dominant than the dehydration penalties. Calculations of the position-dependent diffusion coefficients and dynamic friction kernels of the cations indicate that the dehydration process along with the molecular rearrangements occurring outside the channel and the coupling of the ion motions with the chain-structured water movements inside the channel lead to decrease of the diffusion coefficients far away from the channel entrance and also reduced coefficients inside the channel. Here the PMF and diffusivity profiles for Na + and K + reveal that the energetics of ion transport through the CPN are governed by global interactions of ions with all the components in the system while the diffusivity of ions through the channel is mostly determined by local interactions of ions with the confined water molecules inside the channel. Comparison of Na + and K + ion distributions based on overdamped Brownian dynamics simulations based on the PMF and diffusivity profiles with the corresponding results from molecular dynamics shows good agreement, indicating accuracy of the Bayesian inference method for determining diffusion coefficients in this application. In addition this work shows that position-dependent diffusion coefficients of ions are required to explain the dynamics and conductance of ions through the CPN properly.« less

Inactivation of Endothelial Small/Intermediate Conductance of Calcium-Activated Potassium Channels Contributes to Coronary Arteriolar Dysfunction in Diabetic Patients.

PubMed

Liu, Yuhong; Xie, An; Singh, Arun K; Ehsan, Afshin; Choudhary, Gaurav; Dudley, Samuel; Sellke, Frank W; Feng, Jun

2015-08-24

Diabetes is associated with coronary arteriolar endothelial dysfunction. We investigated the role of the small/intermediate (SK(Ca)/IK(Ca)) conductance of calcium-activated potassium channels in diabetes-related endothelial dysfunction. Coronary arterioles (80 to 150 μm in diameter) were dissected from discarded right atrial tissues of diabetic (glycosylated hemoglobin = 9.6±0.25) and nondiabetic patients (glycosylated hemoglobin 5.4±0.12) during coronary artery bypass graft surgery (n=8/group). In-vitro relaxation response of precontracted arterioles was examined in the presence of the selective SK(Ca)/IK(Ca) activator NS309 and other vasodilatory agents. The channel density and membrane potential of diabetic and nondiabetic endothelial cells was measured by using the whole cell patch-clamp technique. The protein expression and distribution of the SK(Ca)/IK(Ca) in the human myocardium and coronary arterioles was examined by Western blotting and immunohistochemistry. Our results indicate that diabetes significantly reduced the coronary arteriolar response to the SK(Ca)/IK(Ca) activator NS309 compared to the respective responses of nondiabetic vessels (P<0.05 versus nondiabetes). The relaxation response of diabetic arterioles to NS309 was prevented by denudation of endothelium (P=0.001 versus endothelium-intact). Diabetes significantly decreased endothelial SK(Ca)/IK(Ca) currents and hyperpolarization induced by the SK(Ca)/IK(Ca) activator NS309 as compared with that of nondiabetics. There were no significant differences in the expression and distribution of SK(Ca)/IK(Ca) proteins in the coronary microvessels. Diabetes is associated with inactivation of endothelial SK(Ca)/IK(Ca) channels, which may contribute to endothelial dysfunction in diabetic patients. © 2015 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Electrogenic transport and K+ ion channel expression by the human endolymphatic sac epithelium

PubMed Central

Kim, Sung Huhn; Kim, Bo Gyung; Kim, Jin Young; Roh, Kyung Jin; Suh, Michelle J.; Jung, JinSei; Moon, In Seok; Moon, Sung K.; Choi, Jae Young

2015-01-01

The endolymphatic sac (ES) is a cystic organ that is a part of the inner ear and is connected to the cochlea and vestibule. The ES is thought to be involved in inner ear ion homeostasis and fluid volume regulation for the maintenance of hearing and balance function. Many ion channels, transporters, and exchangers have been identified in the ES luminal epithelium, mainly in animal studies, but there has been no functional study investigating ion transport using human ES tissue. We designed the first functional experiments on electrogenic transport in human ES and investigated the contribution of K+ channels in the electrogenic transport, which has been rarely identified, even in animal studies, using electrophysiological/pharmacological and molecular biological methods. As a result, we identified functional and molecular evidence for the essential participation of K+ channels in the electrogenic transport of human ES epithelium. The identified K+ channels involved in the electrogenic transport were KCNN2, KCNJ14, KCNK2, and KCNK6, and the K+ transports via those channels are thought to play an important role in the maintenance of the unique ionic milieu of the inner ear fluid. PMID:26655723

Molecular basis of the interaction between gating modifier spider toxins and the voltage sensor of voltage-gated ion channels

NASA Astrophysics Data System (ADS)

Lau, Carus H. Y.; King, Glenn F.; Mobli, Mehdi

2016-09-01

Voltage-sensor domains (VSDs) are modular transmembrane domains of voltage-gated ion channels that respond to changes in membrane potential by undergoing conformational changes that are coupled to gating of the ion-conducting pore. Most spider-venom peptides function as gating modifiers by binding to the VSDs of voltage-gated channels and trapping them in a closed or open state. To understand the molecular basis underlying this mode of action, we used nuclear magnetic resonance to delineate the atomic details of the interaction between the VSD of the voltage-gated potassium channel KvAP and the spider-venom peptide VSTx1. Our data reveal that the toxin interacts with residues in an aqueous cleft formed between the extracellular S1-S2 and S3-S4 loops of the VSD whilst maintaining lipid interactions in the gaps formed between the S1-S4 and S2-S3 helices. The resulting network of interactions increases the energetic barrier to the conformational changes required for channel gating, and we propose that this is the mechanism by which gating modifier toxins inhibit voltage-gated ion channels.

Grafting voltage and pharmacological sensitivity in potassium channels.

PubMed

Lan, Xi; Fan, Chunyan; Ji, Wei; Tian, Fuyun; Xu, Tao; Gao, Zhaobing

2016-08-01

A classical voltage-gated ion channel consists of four voltage-sensing domains (VSDs). However, the roles of each VSD in the channels remain elusive. We developed a GVTDT (Graft VSD To Dimeric TASK3 channels that lack endogenous VSDs) strategy to produce voltage-gated channels with a reduced number of VSDs. TASK3 channels exhibit a high host tolerance to VSDs of various voltage-gated ion channels without interfering with the intrinsic properties of the TASK3 selectivity filter. The constructed channels, exemplified by the channels grafted with one or two VSDs from Kv7.1 channels, exhibit classical voltage sensitivity, including voltage-dependent opening and closing. Furthermore, the grafted Kv7.1 VSD transfers the potentiation activity of benzbromarone, an activator that acts on the VSDs of the donor channels, to the constructed channels. Our study indicates that one VSD is sufficient to voltage-dependently gate the pore and provides new insight into the roles of VSDs.

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

Modeling the human Nav1.5 sodium channel: structural and mechanistic insights of ion permeation and drug blockade

PubMed Central

Ahmed, Marawan; Jalily Hasani, Horia; Ganesan, Aravindhan; Houghton, Michael; Barakat, Khaled

2017-01-01

Abnormalities in the human Nav1.5 (hNav1.5) voltage-gated sodium ion channel (VGSC) are associated with a wide range of cardiac problems and diseases in humans. Current structural models of hNav1.5 are still far from complete and, consequently, their ability to study atomistic interactions of this channel is very limited. Here, we report a comprehensive atomistic model of the hNav1.5 ion channel, constructed using homology modeling technique and refined through long molecular dynamics simulations (680 ns) in the lipid membrane bilayer. Our model was comprehensively validated by using reported mutagenesis data, comparisons with previous models, and binding to a panel of known hNav1.5 blockers. The relatively long classical MD simulation was sufficient to observe a natural sodium permeation event across the channel’s selectivity filters to reach the channel’s central cavity, together with the identification of a unique role of the lysine residue. Electrostatic potential calculations revealed the existence of two potential binding sites for the sodium ion at the outer selectivity filters. To obtain further mechanistic insight into the permeation event from the central cavity to the intracellular region of the channel, we further employed “state-of-the-art” steered molecular dynamics (SMD) simulations. Our SMD simulations revealed two different pathways through which a sodium ion can be expelled from the channel. Further, the SMD simulations identified the key residues that are likely to control these processes. Finally, we discuss the potential binding modes of a panel of known hNav1.5 blockers to our structural model of hNav1.5. We believe that the data presented here will enhance our understanding of the structure–property relationships of the hNav1.5 ion channel and the underlying molecular mechanisms in sodium ion permeation and drug interactions. The results presented here could be useful for designing safer drugs that do not block the hNav1.5 channel. PMID:28831242

Ferritin ion channel disorder inhibits Fe(II)/O2 reactivity at distant sites.

PubMed

Tosha, Takehiko; Behera, Rabindra K; Theil, Elizabeth C

2012-11-05

Ferritins, a complex, mineralized, protein nanocage family essential for life, provide iron concentrates and oxidant protection. Protein-based ion channels and Fe(II)/O(2) catalysis initiate conversion of thousands of Fe atoms to caged, ferritin Fe(2)O(3)·H(2)O minerals. The ion channels consist of six helical segments, contributed by 3 of 12 or 24 polypeptide subunits, around the 3-fold cage axes. The channel structure guides entering Fe(II) ions toward multiple, catalytic, diiron sites buried inside ferritin protein helices, ~20 Å away from channel internal exits. The catalytic product, Fe(III)-O(H)-Fe(III), is a mineral precursor; mineral nucleation begins inside the protein cage with mineral growth in the central protein cavity (5-8 nm diameter). Amino acid substitutions that changed ionic or hydrophobic channel interactions R72D, D122R, and L134P increased ion channel structural disorder (protein crystallographic analyses) and increased Fe(II) exit [chelated Fe(II) after ferric mineral reduction/dissolution]. Since substitutions of some channel carboxylate residues diminished ferritin catalysis with no effect on Fe(II) exit, such as E130A and D127A, we investigated catalysis in ferritins with altered Fe(II) exit, R72D, D122R and L134P. The results indicate that simply changing the ionic properties of the channels, as in the R72D variant, need not change the forward catalytic rate. However, both D122R and L134P, which had dramatic effects on ferritin catalysis, also caused larger effects on channel structure and order, contrasting with R72D. All three amino acid substitutions, however, decreased the stability of the catalytic intermediate, diferric peroxo, even though overall ferritin cage structure is very stable, resisting 80 °C and 6 M urea. The localized structural changes in ferritin subdomains that affect ferritin function over long distances illustrate new properties of the protein cage in natural ferritin function and for applied ferritin uses.

Nonhuman Primate Optogenetics: Recent Advances and Future Directions

PubMed Central

Acker, Leah

2017-01-01

Optogenetics is the use of genetically coded, light-gated ion channels or pumps (opsins) for millisecond resolution control of neural activity. By targeting opsin expression to specific cell types and neuronal pathways, optogenetics can expand our understanding of the neural basis of normal and pathological behavior. To maximize the potential of optogenetics to study human cognition and behavior, optogenetics should be applied to the study of nonhuman primates (NHPs). The homology between NHPs and humans makes these animals the best experimental model for understanding human brain function and dysfunction. Moreover, for genetic tools to have translational promise, their use must be demonstrated effectively in large, wild-type animals such as Rhesus macaques. Here, we review recent advances in primate optogenetics. We highlight the technical hurdles that have been cleared, challenges that remain, and summarize how optogenetic experiments are expanding our understanding of primate brain function. PMID:29118219

Improved understanding of the pathophysiology of atrial fibrillation through the lens of discrete pathological pathways

PubMed Central

Balouch, Muhammad A.; Kolek, Matthew J.; Darbar, Dawood

2014-01-01

Atrial fibrillation (AF) is a common disorder with a complex and incompletely understood pathophysiology. Genetic approaches to understanding the pathophysiology of AF have led to the identification of several biological pathways important in the pathogenesis of the arrhythmia. These include pathways important for cardiac development, generation and propagation of atrial electrical impulses, and atrial remodeling and fibrosis. While common and rare genetic variants in these pathways are associated with increased susceptibility to AF, they differ substantially among patients with lone versus typical AF. Furthermore, how these pathways converge to a final common clinical phenotype of AF is unclear and might also vary among different patient populations. Here, we review the contemporary knowledge of AF pathogenesis and discuss how derangement in cardiac development, ion channel dysfunction, and promotion of atrial fibrosis may contribute to this common and important clinical disorder. PMID:25054116

Roles of Ion Channels in the Environmental Responses of Plants

NASA Astrophysics Data System (ADS)

Furuichi, Takuya; Kawano, Tomonori; Tatsumi, Hitoshi; Sokabe, Masahiro

When plant cells are exposed to environmental stresses or perceive internal signal molecules involved in growth and development, ion channels are transiently activated to convert these stimuli into intracellular signals. Among the ions taken up by plant cells, Ca2+ plays an essential role as an intracellular second messenger in plants; the cytoplasmic free Ca2+ concentration ([Ca2+]c) is therefore strictly regulated. Signal transduction pathways mediated by changes in [Ca2+]c - termed Ca2+ signaling - are initiated by the activation of Ca2+-permeable channels in many cases. To date, a large body of electrophysiological and recent molecular biological studies have revealed that plants possess Ca2+ channels belonging to distinct types with different gating mechanisms, and a variety of genes for Ca2+-permeable channels have been isolated and functionally characterized. Topics in this chapter focus on long-distance signal translocation in plants and the characteristics of a variety of plant Ca2+-permeable channels including voltage-dependent Ca2+-permeable channels, cyclic nucleotide-gated cation channels, ionotropic glutamate receptors and mechanosensitive channels. We discuss their roles in environmental responses and in the regulation of growth and development.

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.

Ion channel pharmacology under flow: automation via well-plate microfluidics.

PubMed

Spencer, C Ian; Li, Nianzhen; Chen, Qin; Johnson, Juliette; Nevill, Tanner; Kammonen, Juha; Ionescu-Zanetti, Cristian

2012-08-01

Automated patch clamping addresses the need for high-throughput screening of chemical entities that alter ion channel function. As a result, there is considerable utility in the pharmaceutical screening arena for novel platforms that can produce relevant data both rapidly and consistently. Here we present results that were obtained with an innovative microfluidic automated patch clamp system utilizing a well-plate that eliminates the necessity of internal robotic liquid handling. Continuous recording from cell ensembles, rapid solution switching, and a bench-top footprint enable a number of assay formats previously inaccessible to automated systems. An electro-pneumatic interface was employed to drive the laminar flow of solutions in a microfluidic network that delivered cells in suspension to ensemble recording sites. Whole-cell voltage clamp was applied to linear arrays of 20 cells in parallel utilizing a 64-channel voltage clamp amplifier. A number of unique assays requiring sequential compound applications separated by a second or less, such as rapid determination of the agonist EC(50) for a ligand-gated ion channel or the kinetics of desensitization recovery, are enabled by the system. In addition, the system was validated via electrophysiological characterizations of both voltage-gated and ligand-gated ion channel targets: hK(V)2.1 and human Ether-à-go-go-related gene potassium channels, hNa(V)1.7 and 1.8 sodium channels, and (α1) hGABA(A) and (α1) human nicotinic acetylcholine receptor receptors. Our results show that the voltage dependence, kinetics, and interactions of these channels with pharmacological agents were matched to reference data. The results from these IonFlux™ experiments demonstrate that the system provides high-throughput automated electrophysiology with enhanced reliability and consistency, in a user-friendly format.

Hypoxia Sensing in Plants: On a Quest for Ion Channels as Putative Oxygen Sensors.

PubMed

Wang, Feifei; Chen, Zhong-Hua; Shabala, Sergey

2017-07-01

Over 17 million km2 of land is affected by soil flooding every year, resulting in substantial yield losses and jeopardizing food security across the globe. A key step in resolving this problem and creating stress-tolerant cultivars is an understanding of the mechanisms by which plants sense low-oxygen stress. In this work, we review the current knowledge about the oxygen-sensing and signaling pathway in mammalian and plant systems and postulate the potential role of ion channels as putative oxygen sensors in plant roots. We first discuss the definition and requirements for the oxygen sensor and the difference between sensing and signaling. We then summarize the literature and identify several known candidates for oxygen sensing in the mammalian literature. This includes transient receptor potential (TRP) channels; K+-permeable channels (Kv, BK and TASK); Ca2+ channels (RyR and TPC); and various chemo- and reactive oxygen species (ROS)-dependent oxygen sensors. Identified key oxygen-sensing domains (PAS, GCS, GAF and PHD) in mammalian systems are used to predict the potential plant counterparts in Arabidopsis. Finally, the sequences of known mammalian ion channels with reported roles in oxygen sensing were employed to BLAST the Arabidopsis genome for the candidate genes. Several plasma membrane and tonoplast ion channels (such as TPC, AKT and KCO) and oxygen domain-containing proteins with predicted oxygen-sensing ability were identified and discussed. We propose a testable model for potential roles of ion channels in plant hypoxia sensing. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

The role of cystic fibrosis transmembrane conductance regulator phenylalanine 508 side chain in ion channel gating

PubMed Central

Cui, Liying; Aleksandrov, Luba; Hou, Yue-Xian; Gentzsch, Martina; Chen, Jey-Hsin; Riordan, John R; Aleksandrov, Andrei A

2006-01-01

Cystic fibrosis transmembrane conductance regulator (CFTR) is an ion channel employing the ABC transporter structural motif. Deletion of a single residue (Phe508) in the first nucleotide-binding domain (NBD1), which occurs in most patients with cystic fibrosis, impairs both maturation and function of the protein. However, substitution of the Phe508 with small uncharged amino acids, including cysteine, is permissive for maturation. To explore the possible role of the phenylalanine aromatic side chain in channel gating we introduced a cysteine at this position in cysless CFTR, enabling its selective chemical modification by sulfhydryl reagents. Both cysless and wild-type CFTR ion channels have identical mean open times when activated by different nucleotide ligands. Moreover, both channels could be locked in an open state by introducing an ATPase inhibiting mutation (E1371S). However, the introduction of a single cysteine (F508C) prevented the cysless E1371S channel from maintaining the permanently open state, allowing closing to occur. Chemical modification of cysless E1371S/F508C by sulfhydryl reagents was used to probe the role of the side chain in ion channel function. Specifically, benzyl-methanethiosulphonate modification of this variant restored the gating behaviour to that of cysless E1371S containing the wild-type phenylalanine at position 508. This provides the first direct evidence that a specific interaction of the Phe508 aromatic side chain plays a role in determining the residency time in the closed state. Thus, despite the fact that this aromatic side chain is not essential for CFTR folding, it is important in the ion channel function. PMID:16484308

Innovative polymer nanocomposite electrolytes: nanoscale manipulation of ion channels by functionalized graphenes.

PubMed

Choi, Bong Gill; Hong, Jinkee; Park, Young Chul; Jung, Doo Hwan; Hong, Won Hi; Hammond, Paula T; Park, Hoseok

2011-06-28

The chemistry and structure of ion channels within the polymer electrolytes are of prime importance for studying the transport properties of electrolytes as well as for developing high-performance electrochemical devices. Despite intensive efforts on the synthesis of polymer electrolytes, few studies have demonstrated enhanced target ion conduction while suppressing unfavorable ion or mass transport because the undesirable transport occurs through an identical pathway. Herein, we report an innovative, chemical strategy for the synthesis of polymer electrolytes whose ion-conducting channels are physically and chemically modulated by the ionic (not electronic) conductive, functionalized graphenes and for a fundamental understanding of ion and mass transport occurring in nanoscale ionic clusters. The functionalized graphenes controlled the state of water by means of nanoscale manipulation of the physical geometry and chemical functionality of ionic channels. Furthermore, the confinement of bound water within the reorganized nanochannels of composite membranes was confirmed by the enhanced proton conductivity at high temperature and the low activation energy for ionic conduction through a Grotthus-type mechanism. The selectively facilitated transport behavior of composite membranes such as high proton conductivity and low methanol crossover was attributed to the confined bound water, resulting in high-performance fuel cells.

Investigation of the role of non-selective calcium channel blocker (flunarizine) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice.

PubMed

Gulati, Puja; Muthuraman, Arunachalam; Kaur, Parneet

2015-04-01

The present study was designed to investigate the role of flunarizine (a non-selective calcium channel blocker) on cerebral ischemic-reperfusion associated cognitive dysfunction in aged mice. Bilateral carotid artery occlusion of 12min followed by reperfusion for 24h was given to induce cerebral injury in male Swiss mice. The assessment of learning & memory was performed by Morris water maze test; motor in-coordination was evaluated by rota rod, lateral push and inclined beam walking tests; cerebral infarct size was quantified by triphenyltetrazolium chloride staining. In addition, reduced glutathione (GSH), total calcium and acetylcholinesterase (AChE) activity were also estimated in aged brain tissue. Donepezil treated group served as a positive control in this study. Ischemia reperfusion (I/R) injury produced significant increase in cerebral infarct size. A significant loss of memory along with impairment of motor performance was also noted. Further, I/R injury also produced significant increase in levels of total calcium, AChE activity and decrease in GSH levels. Pretreatment of flunarizine significantly attenuated I/R induced infarct size, behavioral and biochemical changes. Hence, it may be concluded that, a non-selective calcium channel blocker can be useful in I/R associated cognitive dysfunction due to its anti-oxidant, anti-infarct and modulatory actions of neurotransmitters & calcium channels. Copyright © 2015 Elsevier Inc. All rights reserved.

Structural plasticity and dynamic selectivity of acid-sensing ion channel-spider toxin complexes

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

Baconguis, Isabelle; Gouaux, Eric

2012-07-29

Acid-sensing ion channels (ASICs) are voltage-independent, amiloride-sensitive channels involved in diverse physiological processes ranging from nociception to taste. Despite the importance of ASICs in physiology, we know little about the mechanism of channel activation. Here we show that psalmotoxin activates non-selective and Na +-selective currents in chicken ASIC1a at pH7.25 and 5.5, respectively. Crystal structures of ASIC1a–psalmotoxin complexes map the toxin binding site to the extracellular domain and show how toxin binding triggers an expansion of the extracellular vestibule and stabilization of the open channel pore. At pH7.25 the pore is approximately 10Å in diameter, whereas at pH5.5 the poremore » is largely hydrophobic and elliptical in cross-section with dimensions of approximately 5 by 7Å, consistent with a barrier mechanism for ion selectivity. These studies define mechanisms for activation of ASICs, illuminate the basis for dynamic ion selectivity and provide the blueprints for new therapeutic agents.« less

The isolated voltage sensing domain of the Shaker potassium channel forms a voltage-gated cation channel

PubMed Central

Zhao, Juan; Blunck, Rikard

2016-01-01

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. DOI: http://dx.doi.org/10.7554/eLife.18130.001 PMID:27710769

Endothelial dysfunction and amyloid-β-induced neurovascular alterations

PubMed Central

Koizumi, Kenzo; Wang, Gang; Park, Laibaik

2015-01-01

Alzheimer's disease (AD) and cerebrovascular diseases share common vascular risk factors that have disastrous effects on cerebrovascular regulation. Endothelial cells, lining inner walls of cerebral blood vessels, form a dynamic interface between the blood and the brain and are critical for the maintenance of neurovascular homeostasis. Accordingly, injury in endothelial cells is regarded as one of the earliest symptoms of impaired vasoregulatory mechanisms. Extracellular buildup of amyloid-β (Aβ) is a central pathogenic factor in AD. Aβ exerts potent detrimental effects on cerebral blood vessels and impairs endothelial structure and function. Recent evidence implicates vascular oxidative stress and activation of the nonselective cationic channel transient receptor potential melastatin (TRPM)-2 on endothelial cells in the mechanisms of Aβ-induced neurovascular dysfunction. Thus, Aβ triggers opening of TRPM2 channels in endothelial cells leading to intracellular Ca2+ overload and vasomotor dysfunction. The cerebrovascular dysfunction may contribute to AD pathogenesis by reducing the cerebral blood supply, leading to increased susceptibility to vascular insufficiency, and by promoting Aβ accumulation. The recent realization that vascular factors contribute to AD pathobiology suggests new targets for the prevention and treatment of this devastating disease. PMID:26328781

The desensitization gate of inhibitory Cys-loop receptors

NASA Astrophysics Data System (ADS)

Gielen, Marc; Thomas, Philip; Smart, Trevor G.

2015-04-01

Cys-loop neurotransmitter-gated ion channels are vital for communication throughout the nervous system. Following activation, these receptors enter into a desensitized state in which the ion channel shuts even though the neurotransmitter molecules remain bound. To date, the molecular determinants underlying this most fundamental property of Cys-loop receptors have remained elusive. Here we present a generic mechanism for the desensitization of Cys-loop GABAA (GABAARs) and glycine receptors (GlyRs), which both mediate fast inhibitory synaptic transmission. Desensitization is regulated by interactions between the second and third transmembrane segments, which affect the ion channel lumen near its intracellular end. The GABAAR and GlyR pore blocker picrotoxin prevented desensitization, consistent with its deep channel-binding site overlapping a physical desensitization gate.

Nanostructuring of conduction channels in (In,Ga)As-InP heterostructures: Overcoming carrier generation caused by Ar ion milling

NASA Astrophysics Data System (ADS)

Hortelano, V.; Weidlich, H.; Semtsiv, M. P.; Masselink, W. T.; Ramsteiner, M.; Jahn, U.; Biermann, K.; Takagaki, Y.

2018-04-01

Nanometer-sized channels are fabricated in (In,Ga)As-InP heterostructures using Ar ion milling. The ion milling causes spontaneous creation of nanowires, and moreover, electrical conduction of the surface as carriers is generated by sputtering-induced defects. We demonstrate a method to restore electrical isolation in the etched area that is compatible with the presence of the nanochannels. We remove the heavily damaged surface layer using a diluted HCl solution and subsequently recover the crystalline order in the moderately damaged part by annealing. We optimize the HCl concentration to make the removal stop on its own before reaching the conduction channel part. The lateral depletion in the channels is shown to be almost absent.

A limited 4 Å radial displacement of the S4-S5 linker is sufficient for internal gate closing in Kv channels.

PubMed

Faure, Élise; Starek, Greg; McGuire, Hugo; Bernèche, Simon; Blunck, Rikard

2012-11-16

Voltage-gated ion channels are responsible for the generation of action potentials in our nervous system. Conformational rearrangements in their voltage sensor domains in response to changes of the membrane potential control pore opening and thus ion conduction. Crystal structures of the open channel in combination with a wealth of biophysical data and molecular dynamics simulations led to a consensus on the voltage sensor movement. However, the coupling between voltage sensor movement and pore opening, the electromechanical coupling, occurs at the cytosolic face of the channel, from where no structural information is available yet. In particular, the question how far the cytosolic pore gate has to close to prevent ion conduction remains controversial. In cells, spectroscopic methods are hindered because labeling of internal sites remains difficult, whereas liposomes or detergent solutions containing purified ion channels lack voltage control. Here, to overcome these problems, we controlled the state of the channel by varying the lipid environment. This way, we directly measured the position of the S4-S5 linker in both the open and the closed state of a prokaryotic Kv channel (KvAP) in a lipid environment using Lanthanide-based resonance energy transfer. We were able to reconstruct the movement of the covalent link between the voltage sensor and the pore domain and used this information as restraints for molecular dynamics simulations of the closed state structure. We found that a small decrease of the pore radius of about 3-4 Å is sufficient to prevent ion permeation through the pore.

Effect of cholesterol depletion on the pore dilation of TRPV1.

PubMed

Jansson, Erik T; Trkulja, Carolina L; Ahemaiti, Aikeremu; Millingen, Maria; Jeffries, Gavin Dm; Jardemark, Kent; Orwar, Owe

2013-01-02

The TRPV1 ion channel is expressed in nociceptors, where pharmacological modulation of its function may offer a means of alleviating pain and neurogenic inflammation processes in the human body. The aim of this study was to investigate the effects of cholesterol depletion of the cell on ion-permeability of the TRPV1 ion channel. The ion-permeability properties of TRPV1 were assessed using whole-cell patch-clamp and YO-PRO uptake rate studies on a Chinese hamster ovary (CHO) cell line expressing this ion channel. Prolonged capsaicin-induced activation of TRPV1 with N-methyl-D-glucamine (NMDG) as the sole extracellular cation, generated a biphasic current which included an initial outward current followed by an inward current. Similarly, prolonged proton-activation (pH 5.5) of TRPV1 under hypocalcemic conditions also generated a biphasic current including a fast initial current peak followed by a larger second one. Patch-clamp recordings of reversal potentials of TRPV1 revealed an increase of the ion-permeability for NMDG during prolonged activation of this ion channel under hypocalcemic conditions. Our findings show that cholesterol depletion inhibited both the second current, and the increase in ion-permeability of the TRPV1 channel, resulting from sustained agonist-activation with capsaicin and protons (pH 5.5). These results were confirmed with YO-PRO uptake rate studies using laser scanning confocal microscopy, where cholesterol depletion was found to decrease TRPV1 mediated uptake rates of YO-PRO. Hence, these results propose a novel mechanism by which cellular cholesterol depletion modulates the function of TRPV1, which may constitute a novel approach for treatment of neurogenic pain.

Fabrication of Amorphous Indium Gallium Zinc Oxide Thin Film Transistor by using Focused Ion Beam

NASA Astrophysics Data System (ADS)

Zhu, Wencong

Compared with other transparent semiconductors, amorphous indium gallium zinc oxide (a-IGZO) has both good uniformity and high electron mobility, which make it as a good candidate for displays or large-scale transparent circuit. The goal of this research is to fabricate alpha-IGZO thin film transistor (TFT) with channel milled by focused ion beam (FIB). TFTs with different channel geometries can be achieved by applying different milling strategies, which facilitate modifying complex circuit. Technology Computer-Aided Design (TCAD) was also introduced to understand the effect of trapped charges on the device performance. The investigation of the trapped charge at IGZO/SiO2 interface was performed on the IGZO TFT on p-Silicon substrate with thermally grown SiO2 as dielectric. The subgap density-of-state model was used for the simulation, which includes conduction band-tail trap states and donor-like state in the subgap. The result shows that the de-trapping and donor-state ionization determine the interface trapped charge density at various gate biases. Simulation of IGZO TFT with FIB defined channel on the same substrate was also applied. The drain and source were connected intentionally during metal deposition and separated by FIB milling. Based on the simulation, the Ga ions in SiO2 introduced by the ion beam was drifted by gate bias and affects the saturation drain current. Both side channel and direct channel transparent IGZO TFTs were fabricated on the glass substrate with coated ITO. Higher ion energy (30 keV) was used to etch through the substrate between drain and source and form side channels at the corner of milled trench. Lower ion energy (16 keV) was applied to stop the milling inside IGZO thin film and direct channel between drain and source was created. Annealing after FIB milling removed the residual Ga ions and the devices show switch feature. Direct channel shows higher saturation drain current (~10-6 A) compared with side channel (~10-7 A) because of its shorter channel length and wider width, however, it also exhibit higher gate leakage current (>10-7 A) than side channel (<10-7 A) due to larger Ga ion implantation and diffusion region in SiO2 after annealing. Hysteresis window increase and positive VON shift were also observed due to the interface trap density increase and carrier density suppression both by Ga ions. Laser interference lithography was applied to define the IGZO active region, which gives more flexibility on TFT channel dimension and circuit modification. He-Cd laser with 325 nm wavelength was used to define 2D array of IGZO islands with period of 2.5 im. Logic gate array was designed and fabricated by combining this 2D array of IGZO islands and FIB direct channel milling. After annealing, device shows on-off feature, but high temperature (400 °C) release more free carrier and results in negative shift of VON. The row selection voltage was also introduced in the design of logic gate array to act as switch of input signals to each row separately. However, due to the long input signal sweeping time, the leakage current cannot be overlooked. The idea can be verified by AC or short pulse input signal.

The Origins of Transmembrane Ion Channels

NASA Technical Reports Server (NTRS)

Pohorille, Andrew; Wilson, Michael A.

2012-01-01

Even though membrane proteins that mediate transport of ions and small molecules across cell walls are among the largest and least understood biopolymers in contemporary cells, it is still possible to shed light on their origins and early evolution. The central observation is that transmembrane portions of most ion channels are simply bundles of -helices. By combining results of experimental and computer simulation studies on synthetic models and natural channels, mostly of non-genomic origin, we show that the emergence of -helical channels was protobiologically plausible, and did not require highly specific amino acid sequences. Despite their simple structure, such channels could possess properties that, at the first sight, appear to require markedly larger complexity. Specifically, we explain how the antiamoebin channels, which are made of identical helices, 16 amino acids in length, achieve efficiency comparable to that of highly evolved channels. We further show that antiamoebin channels are extremely flexible, compared to modern, genetically coded channels. On the basis of our results, we propose that channels evolved further towards high structural complexity because they needed to acquire stable rigid structures and mechanisms for precise regulation rather than improve efficiency. In general, even though architectures of membrane proteins are not nearly as diverse as those of water-soluble proteins, they are sufficiently flexible to adapt readily to the functional demands arising during evolution.

Mitochondrial Ion Channels in Cancer Transformation

PubMed Central

Madamba, Stephen M.; Damri, Kevin N.; Dejean, Laurent M.; Peixoto, Pablo M.

2015-01-01

Cancer transformation involves reprograming of mitochondrial function to avert cell death mechanisms, monopolize energy metabolism, accelerate mitotic proliferation, and promote metastasis. Mitochondrial ion channels have emerged as promising therapeutic targets because of their connection to metabolic and apoptotic functions. This mini review discusses how mitochondrial channels may be associated with cancer transformation and expands on the possible involvement of mitochondrial protein import complexes in pathophysiological process. PMID:26090338

Chasing the open-state structure of pentameric ligand-gated ion channels

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

Gonzalez-Gutierrez, Giovanni; Wang, Yuhang; Cymes, Gisela D.

Remarkable advances have been made toward the structural characterization of ion channels in the last two decades. However, the unambiguous assignment of well-defined functional states to the obtained structural models has proved challenging. In the case of the superfamily of nicotinic-receptor channels (also referred to as pentameric ligand-gated ion channels [pLGICs]), for example, two different types of model of the open-channel conformation have been proposed on the basis of structures solved to resolutions better than 4.0 Å. At the level of the transmembrane pore, the open-state models of the proton-gated pLGIC fromGloeobacter violaceus(GLIC) and the invertebrate glutamate-gated Cl –channel (GluCl)more » are very similar to each other, but that of the glycine receptor (GlyR) is considerably wider. Indeed, the mean distances between the axis of ion permeation and the Cα atoms at the narrowest constriction of the pore (position -2') differ by ~2 Å in these two classes of model, a large difference when it comes to understanding the physicochemical bases of ion conduction and charge selectivity. Here, we take advantage of the extreme open-channel stabilizing effect of mutations at pore-facing position 9'. We find that the I9'A mutation slows down entry into desensitization of GLIC to the extent that macroscopic currents decay only slightly by the end of pH 4.5 solution applications to the extracellular side for several minutes. We crystallize (at pH 4.5) two variants of GLIC carrying this mutation and solve their structures to resolutions of 3.12 Å and 3.36 Å. Furthermore, we perform all-atom molecular dynamics simulations of ion permeation and picrotoxinin block, using the different open-channel structural models. On the basis of these results, we favor the notion that the open-channel structure of pLGICs from animals is much closer to that of the narrow models (of GLIC and GluCl) than it is to that of the GlyR.« less

Photocontrol of Voltage-Gated Ion Channel Activity by Azobenzene Trimethylammonium Bromide in Neonatal Rat Cardiomyocytes

PubMed Central

Frolova, Sheyda R.; Gaiko, Olga; Tsvelaya, Valeriya A.; Pimenov, Oleg Y.; Agladze, Konstantin I.

2016-01-01

The ability of azobenzene trimethylammonium bromide (azoTAB) to sensitize cardiac tissue excitability to light was recently reported. The dark, thermally relaxed trans- isomer of azoTAB suppressed spontaneous activity and excitation propagation speed, whereas the cis- isomer had no detectable effect on the electrical properties of cardiomyocyte monolayers. As the membrane potential of cardiac cells is mainly controlled by activity of voltage-gated ion channels, this study examined whether the sensitization effect of azoTAB was exerted primarily via the modulation of voltage-gated ion channel activity. The effects of trans- and cis- isomers of azoTAB on voltage-dependent sodium (INav), calcium (ICav), and potassium (IKv) currents in isolated neonatal rat cardiomyocytes were investigated using the whole-cell patch-clamp technique. The experiments showed that azoTAB modulated ion currents, causing suppression of sodium (Na+) and calcium (Ca2+) currents and potentiation of net potassium (K+) currents. This finding confirms that azoTAB-effect on cardiac tissue excitability do indeed result from modulation of voltage-gated ion channels responsible for action potential. PMID:27015602

The twisted ion-permeation pathway of a resting voltage-sensing domain.

PubMed

Tombola, Francesco; Pathak, Medha M; Gorostiza, Pau; Isacoff, Ehud Y

2007-02-01

Proteins containing voltage-sensing domains (VSDs) translate changes in membrane potential into changes in ion permeability or enzymatic activity. In channels, voltage change triggers a switch in conformation of the VSD, which drives gating in a separate pore domain, or, in channels lacking a pore domain, directly gates an ion pathway within the VSD. Neither mechanism is well understood. In the Shaker potassium channel, mutation of the first arginine residue of the S4 helix to a smaller uncharged residue makes the VSD permeable to ions ('omega current') in the resting conformation ('S4 down'). Here we perform a structure-guided perturbation analysis of the omega conductance to map its VSD permeation pathway. We find that there are four omega pores per channel, which is consistent with one conduction path per VSD. Permeating ions from the extracellular medium enter the VSD at its peripheral junction with the pore domain, and then plunge into the core of the VSD in a curved conduction pathway. Our results provide a model of the resting conformation of the VSD.

Dynamic hybrid materials for constitutional self-instructed membranes

PubMed Central

Cazacu, Adinela; Legrand, Yves-Marie; Pasc, Andreea; Nasr, Gihane; Van der Lee, Arie; Mahon, Eugene; Barboiu, Mihail

2009-01-01

Constitutional self-instructed membranes were developed and used for mimicking the adaptive structural functionality of natural ion-channel systems. These membranes are based on dynamic hybrid materials in which the functional self-organized macrocycles are reversibly connected with the inorganic silica through hydrophobic noncovalent interactions. Supramolecular columnar ion-channel architectures can be generated by reversible confinement within scaffolding hydrophobic silica mesopores. They can be structurally determined by using X-ray diffraction and morphologically tuned by alkali-salts templating. From the conceptual point of view, these membranes express a synergistic adaptive behavior: the simultaneous binding of the fittest cation and its anion would be a case of “homotropic allosteric interactions,” because in time it increases the transport efficiency of the pore-contained superstructures by a selective evolving process toward the fittest ion channel. The hybrid membranes presented here represent dynamic constitutional systems evolving over time to form the fittest ion channels from a library of molecular and supramolecular components, or selecting the fittest ion pairs from a mixture of salts demonstrating flexible adaptation. PMID:19416909

Voltage Gated Ion Channel Function: Gating, Conduction, and the Role of Water and Protons

PubMed Central

Kariev, Alisher M.; Green, Michael E.

2012-01-01

Ion channels, which are found in every biological cell, regulate the concentration of electrolytes, and are responsible for multiple biological functions, including in particular the propagation of nerve impulses. The channels with the latter function are gated (opened) by a voltage signal, which allows Na+ into the cell and K+ out. These channels have several positively charged amino acids on a transmembrane domain of their voltage sensor, and it is generally considered, based primarily on two lines of experimental evidence, that these charges move with respect to the membrane to open the channel. At least three forms of motion, with greatly differing extents and mechanisms of motion, have been proposed. There is a “gating current”, a capacitative current preceding the channel opening, that corresponds to several charges (for one class of channel typically 12–13) crossing the membrane field, which may not require protein physically crossing a large fraction of the membrane. The coupling to the opening of the channel would in these models depend on the motion. The conduction itself is usually assumed to require the “gate” of the channel to be pulled apart to allow ions to enter as a section of the protein partially crosses the membrane, and a selectivity filter at the opposite end of the channel determines the ion which is allowed to pass through. We will here primarily consider K+ channels, although Na+ channels are similar. We propose that the mechanism of gating differs from that which is generally accepted, in that the positively charged residues need not move (there may be some motion, but not as gating current). Instead, protons may constitute the gating current, causing the gate to open; opening consists of only increasing the diameter at the gate from approximately 6 Å to approximately 12 Å. We propose in addition that the gate oscillates rather than simply opens, and the ion experiences a barrier to its motion across the channel that is tuned by the water present within the channel. Our own quantum calculations as well as numerous experiments of others are interpreted in terms of this hypothesis. It is also shown that the evidence that supports the motion of the sensor as the gating current can also be consistent with the hypothesis we present. PMID:22408417

Improved Ion-Channel Biosensors

NASA Technical Reports Server (NTRS)

Nadeau, Jay; White, Victor; Dougherty, Dennis; Maurer, Joshua

2004-01-01

An effort is underway to develop improved biosensors of a type based on ion channels in biomimetic membranes. These sensors are microfabricated from silicon and other materials compatible with silicon. As described, these sensors offer a number of advantages over prior sensors of this type.

Ion channel drug discovery and research: the automated Nano-Patch-Clamp technology.

PubMed

Brueggemann, A; George, M; Klau, M; Beckler, M; Steindl, J; Behrends, J C; Fertig, N

2004-01-01

Unlike the genomics revolution, which was largely enabled by a single technological advance (high throughput sequencing), rapid advancement in proteomics will require a broader effort to increase the throughput of a number of key tools for functional analysis of different types of proteins. In the case of ion channels -a class of (membrane) proteins of great physiological importance and potential as drug targets- the lack of adequate assay technologies is felt particularly strongly. The available, indirect, high throughput screening methods for ion channels clearly generate insufficient information. The best technology to study ion channel function and screen for compound interaction is the patch clamp technique, but patch clamping suffers from low throughput, which is not acceptable for drug screening. A first step towards a solution is presented here. The nano patch clamp technology, which is based on a planar, microstructured glass chip, enables automatic whole cell patch clamp measurements. The Port-a-Patch is an automated electrophysiology workstation, which uses planar patch clamp chips. This approach enables high quality and high content ion channel and compound evaluation on a one-cell-at-a-time basis. The presented automation of the patch process and its scalability to an array format are the prerequisites for any higher throughput electrophysiology instruments.

Chansporter complexes in cell signaling.

PubMed

Abbott, Geoffrey W

2017-09-01

Ion channels facilitate diffusion of ions across cell membranes for such diverse purposes as neuronal signaling, muscular contraction, and fluid homeostasis. Solute transporters often utilize ionic gradients to move aqueous solutes up their concentration gradient, also fulfilling a wide variety of tasks. Recently, an increasing number of ion channel-transporter ('chansporter') complexes have been discovered. Chansporter complex formation may overcome what could otherwise be considerable spatial barriers to rapid signal integration and feedback between channels and transporters, the ions and other substrates they transport, and environmental factors to which they must respond. Here, current knowledge in this field is summarized, covering both heterologous expression structure/function findings and potential mechanisms by which chansporter complexes fulfill contrasting roles in cell signaling in vivo. © 2017 Federation of European Biochemical Societies.

Thinking in cycles: MWC is a good model for acetylcholine receptor-channels

PubMed Central

Auerbach, Anthony

2012-01-01

Abstract Neuromuscular acetylcholine receptors have long been a model system for understanding the mechanisms of operation of ligand-gated ion channels and fast chemical synapses. These five subunit membrane proteins have two allosteric (transmitter) binding sites and a distant ion channel domain. Occupation of the binding sites by agonist molecules transiently increases the probability that the channel is ion-permeable. Recent experiments show that the Monod, Wyman and Changeux formalism for allosteric proteins, originally developed for haemoglobin, is an excellent model for acetylcholine receptors. By using mutations and single-channel electrophysiology, the gating equilibrium constants for receptors with zero, one or two bound agonist molecules, and the agonist association and dissociation rate constants from both the closed- and open-channel conformations, have been estimated experimentally. The change in affinity for each transmitter molecule between closed and open conformations provides ∼–5.1 kcal mol−1 towards the global gating isomerization of the protein. PMID:21807612

Multi-Dielectric Brownian Dynamics and Design-Space-Exploration Studies of Permeation in Ion Channels.

PubMed

Siksik, May; Krishnamurthy, Vikram

2017-09-01

This paper proposes a multi-dielectric Brownian dynamics simulation framework for design-space-exploration (DSE) studies of ion-channel permeation. The goal of such DSE studies is to estimate the channel modeling-parameters that minimize the mean-squared error between the simulated and expected "permeation characteristics." To address this computational challenge, we use a methodology based on statistical inference that utilizes the knowledge of channel structure to prune the design space. We demonstrate the proposed framework and DSE methodology using a case study based on the KcsA ion channel, in which the design space is successfully reduced from a 6-D space to a 2-D space. Our results show that the channel dielectric map computed using the framework matches with that computed directly using molecular dynamics with an error of 7%. Finally, the scalability and resolution of the model used are explored, and it is shown that the memory requirements needed for DSE remain constant as the number of parameters (degree of heterogeneity) increases.

Nanometer-scale water- and proton-diffusion heterogeneities across water channels in polymer electrolyte membranes.

PubMed

Song, Jinsuk; Han, Oc Hee; Han, Songi

2015-03-16

Nafion, the most widely used polymer for electrolyte membranes (PEMs) in fuel cells, consists of a fluorocarbon backbone and acidic groups that, upon hydration, swell to form percolated channels through which water and ions diffuse. Although the effects of the channel structures and the acidic groups on water/ion transport have been studied before, the surface chemistry or the spatially heterogeneous diffusivity across water channels has never been shown to directly influence water/ion transport. By the use of molecular spin probes that are selectively partitioned into heterogeneous regions of the PEM and Overhauser dynamic nuclear polarization relaxometry, this study reveals that both water and proton diffusivity are significantly faster near the fluorocarbon and the acidic groups lining the water channels than within the water channels. The concept that surface chemistry at the (sub)nanometer scale dictates water and proton diffusivity invokes a new design principle for PEMs. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Towards a Unified Understanding of Lithium Action in Basic Biology and its Significance for Applied Biology.

PubMed

Jakobsson, Eric; Argüello-Miranda, Orlando; Chiu, See-Wing; Fazal, Zeeshan; Kruczek, James; Nunez-Corrales, Santiago; Pandit, Sagar; Pritchet, Laura

2017-12-01

Lithium has literally been everywhere forever, since it is one of the three elements created in the Big Bang. Lithium concentration in rocks, soil, and fresh water is highly variable from place to place, and has varied widely in specific regions over evolutionary and geologic time. The biological effects of lithium are many and varied. Based on experiments in which animals are deprived of lithium, lithium is an essential nutrient. At the other extreme, at lithium ingestion sufficient to raise blood concentration significantly over 1 mM/, lithium is acutely toxic. There is no consensus regarding optimum levels of lithium intake for populations or individuals-with the single exception that lithium is a generally accepted first-line therapy for bipolar disorder, and specific dosage guidelines for sufferers of that condition are generally agreed on. Epidemiological evidence correlating various markers of social dysfunction and disease vs. lithium level in drinking water suggest benefits of moderately elevated lithium compared to average levels of lithium intake. In contrast to other biologically significant ions, lithium is unusual in not having its concentration in fluids of multicellular animals closely regulated. For hydrogen ions, sodium ions, potassium ions, calcium ions, chloride ions, and magnesium ions, blood and extracellular fluid concentrations are closely and necessarily regulated by systems of highly selective channels, and primary and secondary active transporters. Lithium, while having strong biological activity, is tolerated over body fluid concentrations ranging over many orders of magnitude. The lack of biological regulation of lithium appears due to lack of lithium-specific binding sites and selectivity filters. Rather lithium exerts its myriad physiological and biochemical effects by competing for macromolecular sites that are relatively specific for other cations, most especially for sodium and magnesium. This review will consider what is known about the nature of this competition and suggest using and extending this knowledge towards the goal of a unified understanding of lithium in biology and the application of that understanding in medicine and nutrition.

Beneficial effects of intracoronary nicorandil on microvascular dysfunction after primary percutaneous coronary intervention: demonstration of its superiority to nitroglycerin in a cross-over study.

PubMed

Ito, Noritoshi; Nanto, Shinsuke; Doi, Yasuji; Kurozumi, Yuma; Natsukawa, Tomoaki; Shibata, Hiroyuki; Morita, Masaya; Kawata, Atsushi; Tsuruoka, Ayumu; Sawano, Hirotaka; Okada, Ken-ichiro; Sakata, Yasuhiko; Kai, Tatsuro; Hayashi, Toru

2013-08-01

In patients undergoing primary percutaneous coronary intervention (PCI) for the treatment of ST-segment elevation myocardial infarction (STEMI), coronary microvascular dysfunction is associated with poor prognosis. Coronary microvascular resistance is predominantly regulated by ATP-sensitive potassium (KATP) channels. The aim of this study was to clarify whether nicorandil, a hybrid KATP channel opener and nitric oxide donor, may be a good candidate for improving microvascular dysfunction even when administered after primary PCI. We compared the beneficial effects of nicorandil and nitroglycerin on microvascular function in 60 consecutive patients with STEMI. After primary PCI, all patients received single intracoronary administrations of nitroglycerin (250 μg) and nicorandil (2 mg) in a randomized order; 30 received nicorandil first, while the other 30 received nitroglycerin first. Microvascular dysfunction was evaluated with the index of microcirculatory resistance (IMR), defined as the distal coronary pressure multiplied by the hyperemic mean transit time. As a first administration, nicorandil decreased IMR significantly more than did nitroglycerin (median [interquartile ranges]: 10.8[5.2-20.7] U vs. 2.1[1.0-6.0] U, p=0.0002).As a second administration, nicorandil further decreased IMR, while nitroglycerin did not (median [interquartile ranges]: 6.0[1.3-12.7] U vs. -1.4[-2.6 to 1.3] U, p<0.0001). The IMR after the second administration was significantly associated with myocardial blush grade, angiographic TIMI frame count after the procedure, and peak creatine kinase level. Intracoronary nicorandil reduced microvascular dysfunction after primary PCI more effectively than did nitroglycerin in patients with STEMI, probably via its KATP channel-opening effect.

A Non-canonical Voltage-Sensing Mechanism Controls Gating in K2P K(+) Channels.

PubMed

Schewe, Marcus; Nematian-Ardestani, Ehsan; Sun, Han; Musinszki, Marianne; Cordeiro, Sönke; Bucci, Giovanna; de Groot, Bert L; Tucker, Stephen J; Rapedius, Markus; Baukrowitz, Thomas

2016-02-25

Two-pore domain (K2P) K(+) channels are major regulators of excitability that endow cells with an outwardly rectifying background "leak" conductance. In some K2P channels, strong voltage-dependent activation has been observed, but the mechanism remains unresolved because they lack a canonical voltage-sensing domain. Here, we show voltage-dependent gating is common to most K2P channels and that this voltage sensitivity originates from the movement of three to four ions into the high electric field of an inactive selectivity filter. Overall, this ion-flux gating mechanism generates a one-way "check valve" within the filter because outward movement of K(+) induces filter opening, whereas inward movement promotes inactivation. Furthermore, many physiological stimuli switch off this flux gating mode to convert K2P channels into a leak conductance. These findings provide insight into the functional plasticity of a K(+)-selective filter and also refine our understanding of K2P channels and the mechanisms by which ion channels can sense voltage. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Two-pore channels (TPCs): Novel voltage-gated ion channels with pleiotropic functions

PubMed Central

Feijóo-Bandín, Sandra; García-Vence, María; García-Rúa, Vanessa; Roselló-Lletí, Esther; Portolés, Manuel; Rivera, Miguel; González-Juanatey, José Ramón; Lago, Francisca

2017-01-01

ABSTRACT Two-pore channels (TPC1-3) comprise a subfamily of the eukaryotic voltage-gated ion channels (VGICs) superfamily that are mainly expressed in acidic stores in plants and animals. TPCS are widespread across the animal kingdom, with primates, mice and rats lacking TPC3, and mainly act as Ca+ and Na+ channels, although it was also suggested that they could be permeable to other ions. Nowadays, TPCs have been related to the development of different diseases, including Parkinson´s disease, obesity or myocardial ischemia. Due to this, their study has raised the interest of the scientific community to try to understand their mechanism of action in order to be able to develop an efficient drug that could regulate TPCs activity. In this review, we will provide an updated view regarding TPCs structure, function and activation, as well as their role in different pathophysiological processes. PMID:27440385

Design of high-energy high-current linac with focusing by superconducting solenoids

NASA Astrophysics Data System (ADS)

Batskikh, Guennady I.; Belugin, Vladimir M.; Bondarev, Boris I.; Fedotov, Arkady P.; Durkin, Alexander P.; Ivanov, Yury D.; Mikhailov, Vladimir N.; Murin, Boris P.; Mustafin, Kharis Kh.; Shumakov, Igor V.; Uksusov, Nikolay I.

1995-09-01

The advancement of MRTI design for 1.5 GeV and 250 mA ion CW linac is presented in the report. In new linac version all the way from input to output the ions are focused by magnetic fields of superconducting solenoids. The ion limit current is far beyond the needed value. The linac focusing channel offers major advantages over the more conventional ones. The acceptance is 1.7 times as large for such focusing channel as for quadrupole one. Concurrently, a random perturbation sensitivity for such channel is one order of magnitude smaller than in quadrupole channel. These focusing channel features allow to decrease beam matched radius and increase a linac radiation purity without aperture growth. ``Regotron'' is used as high power generator in linac main part. But D&W cavities need not be divided into sections connected by RF-bridges which denuded them of high coupling factor.

Aluminium and hydrogen ions inhibit a mechanosensory calcium-selective cation channel

NASA Technical Reports Server (NTRS)

Ding, J. P.; Pickard, B. G.

1993-01-01

The tension-dependent activity of mechanosensory calcium-selective cation channels in excised plasmalemmal patches from onion bulb scale epidermis is modulated by pH in the physiologically meaningful range between 4.5 and 7.2. It is rapidly lowered by lowering pH and rapidly raised by raising pH. Channel activity is effectively inhibited by low levels of aluminium ions and activity can be partially restored by washing for a few minutes. We suggest that under normal conditions the sensitivity of the mechanosensory channels to pH of the wall free space plays important roles in regulation of plant activities such as growth. We further suggest that, when levels of acid and aluminium ions in the soil solution are high, they might inhibit similar sensory channels in cells of the root tip, thus contributing critically to the acid soil syndrome.

Coarse Architecture of the Transient Receptor Potential Vanilloid 1 (TRPV1) Ion Channel Determined by Fluorescence Resonance Energy Transfer*

PubMed Central

De-la-Rosa, Víctor; Rangel-Yescas, Gisela E.; Ladrón-de-Guevara, Ernesto; Rosenbaum, Tamara; Islas, León D.

2013-01-01

The transient receptor potential vanilloid 1 ion channel is responsible for the perception of high temperatures and low extracellular pH, and it is also involved in the response to some pungent compounds. Importantly, it is also associated with the perception of pain and noxious stimuli. Here, we attempt to discern the molecular organization and location of the N and C termini of the transient receptor potential vanilloid 1 ion channel by measuring FRET between genetically attached enhanced yellow and cyan fluorescent protein to the N or C terminus of the channel protein, expressed in transfected HEK 293 cells or Xenopus laevis oocytes. The static measurements of the domain organization were mapped into an available cryo-electron microscopy density of the channel with good agreement. These measurements also provide novel insights into the organization of terminal domains and their proximity to the plasma membrane. PMID:23965996

Coarse architecture of the transient receptor potential vanilloid 1 (TRPV1) ion channel determined by fluorescence resonance energy transfer.

PubMed

De-la-Rosa, Víctor; Rangel-Yescas, Gisela E; Ladrón-de-Guevara, Ernesto; Rosenbaum, Tamara; Islas, León D

2013-10-11

The transient receptor potential vanilloid 1 ion channel is responsible for the perception of high temperatures and low extracellular pH, and it is also involved in the response to some pungent compounds. Importantly, it is also associated with the perception of pain and noxious stimuli. Here, we attempt to discern the molecular organization and location of the N and C termini of the transient receptor potential vanilloid 1 ion channel by measuring FRET between genetically attached enhanced yellow and cyan fluorescent protein to the N or C terminus of the channel protein, expressed in transfected HEK 293 cells or Xenopus laevis oocytes. The static measurements of the domain organization were mapped into an available cryo-electron microscopy density of the channel with good agreement. These measurements also provide novel insights into the organization of terminal domains and their proximity to the plasma membrane.

High throughput ion-channel pharmacology: planar-array-based voltage clamp.

PubMed

Kiss, Laszlo; Bennett, Paul B; Uebele, Victor N; Koblan, Kenneth S; Kane, Stefanie A; Neagle, Brad; Schroeder, Kirk

2003-02-01

Technological advances often drive major breakthroughs in biology. Examples include PCR, automated DNA sequencing, confocal/single photon microscopy, AFM, and voltage/patch-clamp methods. The patch-clamp method, first described nearly 30 years ago, was a major technical achievement that permitted voltage-clamp analysis (membrane potential control) of ion channels in most cells and revealed a role for channels in unimagined areas. Because of the high information content, voltage clamp is the best way to study ion-channel function; however, throughput is too low for drug screening. Here we describe a novel breakthrough planar-array-based HT patch-clamp technology developed by Essen Instruments capable of voltage-clamping thousands of cells per day. This technology provides greater than two orders of magnitude increase in throughput compared with the traditional voltage-clamp techniques. We have applied this method to study the hERG K(+) channel and to determine the pharmacological profile of QT prolonging drugs.

Structural and functional characterization of a calcium-activated cation channel from Tsukamurella paurometabola

NASA Astrophysics Data System (ADS)

Dhakshnamoorthy, Balasundaresan; Rohaim, Ahmed; Rui, Huan; Blachowicz, Lydia; Roux, Benoît

2016-09-01

The selectivity filter is an essential functional element of K+ channels that is highly conserved both in terms of its primary sequence and its three-dimensional structure. Here, we investigate the properties of an ion channel from the Gram-positive bacterium Tsukamurella paurometabola with a selectivity filter formed by an uncommon proline-rich sequence. Electrophysiological recordings show that it is a non-selective cation channel and that its activity depends on Ca2+ concentration. In the crystal structure, the selectivity filter adopts a novel conformation with Ca2+ ions bound within the filter near the pore helix where they are coordinated by backbone oxygen atoms, a recurrent motif found in multiple proteins. The binding of Ca2+ ion in the selectivity filter controls the widening of the pore as shown in crystal structures and in molecular dynamics simulations. The structural, functional and computational data provide a characterization of this calcium-gated cationic channel.

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

Sensory Transduction in Caenorhabditis elegans

NASA Astrophysics Data System (ADS)

Brown, Austin L.; Ramot, Daniel; Goodman, Miriam B.

The roundworm Caenorhabditis elegans has a well-defined and comparatively simple repertoire of sensory-guided behaviors, all of which rely on its ability to detect chemical, mechanical or thermal stimuli. In this chapter, we review what is known about the ion channels that mediate sensation in this remarkable model organism. Genetic screens for mutants defective in sensory-guided behaviors have identified genes encoding channel proteins, which are likely transducers of chemical, thermal, and mechanical stimuli. Such classical genetic approaches are now being coupled with molecular genetics and in vivo cellular physiology to elucidate how these channels are activated in specific sensory neurons. The ion channel superfamilies implicated in sensory transduction in C. elegans - CNG, TRP, and DEG/ENaC - are conserved across phyla and also appear to contribute to sensory transduction in other organisms, including vertebrates. What we learn about the role of these ion channels in C. elegans sensation is likely to illuminate analogous processes in other animals, including humans.

Effect of multiply charged ions on the performance and beam characteristics in annular and cylindrical type Hall thruster plasmas

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

Kim, Holak; Lim, Youbong; Choe, Wonho, E-mail: wchoe@kaist.ac.kr

2014-10-06

Plasma plume and thruster performance characteristics associated with multiply charged ions in a cylindrical type Hall thruster (CHT) and an annular type Hall thruster are compared under identical conditions such as channel diameter, channel depth, propellant mass flow rate. A high propellant utilization in a CHT is caused by a high ionization rate, which brings about large multiply charged ions. Ion currents and utilizations are much different due to the presence of multiply charged ions. A high multiply charged ion fraction and a high ionization rate in the CHT result in a higher specific impulse, thrust, and discharge current.

A Voltage Dependent Non-Inactivating Na+ Channel Activated during Apoptosis in Xenopus Oocytes

PubMed Central

Englund, Ulrika H.; Gertow, Jens; Kågedal, Katarina; Elinder, Fredrik

2014-01-01

Ion channels in the plasma membrane are important for the apoptotic process. Different types of voltage-gated ion channels are up-regulated early in the apoptotic process and block of these channels prevents or delays apoptosis. In the present investigation we examined whether ion channels are up-regulated in oocytes from the frog Xenopus laevis during apoptosis. The two-electrode voltage-clamp technique was used to record endogenous ion currents in the oocytes. During staurosporine-induced apoptosis a voltage-dependent Na+ current increased three-fold. This current was activated at voltages more positive than 0 mV (midpoint of the open-probability curve was +55 mV) and showed almost no sign of inactivation during a 1-s pulse. The current was resistant to the Na+-channel blockers tetrodotoxin (1 µM) and amiloride (10 µM), while the Ca2+-channel blocker verapamil (50 µM) in the bath solution completely blocked the current. The intracellular Na+ concentration increased in staurosporine-treated oocytes, but could be prevented by replacing extracellular Na+ whith either K+ or Choline+. Prevention of this influx of Na+ also prevented the STS-induced up-regulation of the caspase-3 activity, suggesting that the intracellular Na+ increase is required to induce apoptosis. Taken together, we have found that a voltage dependent Na+ channel is up-regulated during apoptosis and that influx of Na+ is a crucial step in the apoptotic process in Xenopus oocytes. PMID:24586320

The probability of quantal secretion near a single calcium channel of an active zone.

PubMed Central

Bennett, M R; Farnell, L; Gibson, W G

2000-01-01

A Monte Carlo analysis has been made of calcium dynamics and quantal secretion at microdomains in which the calcium reaches very high concentrations over distances of <50 nm from a channel and for which calcium dynamics are dominated by diffusion. The kinetics of calcium ions in microdomains due to either the spontaneous or evoked opening of a calcium channel, both of which are stochastic events, are described in the presence of endogenous fixed and mobile buffers. Fluctuations in the number of calcium ions within 50 nm of a channel are considerable, with the standard deviation about half the mean. Within 10 nm of a channel these numbers of ions can give rise to calcium concentrations of the order of 100 microM. The temporal changes in free calcium and calcium bound to different affinity indicators in the volume of an entire varicosity or bouton following the opening of a single channel are also determined. A Monte Carlo analysis is also presented of how the dynamics of calcium ions at active zones, after the arrival of an action potential and the stochastic opening of a calcium channel, determine the probability of exocytosis from docked vesicles near the channel. The synaptic vesicles in active zones are found docked in a complex with their calcium-sensor associated proteins and a voltage-sensitive calcium channel, forming a secretory unit. The probability of quantal secretion from an isolated secretory unit has been determined for different distances of an open calcium channel from the calcium sensor within an individual unit: a threefold decrease in the probability of secretion of a quantum occurs with a doubling of the distance from 25 to 50 nm. The Monte Carlo analysis also shows that the probability of secretion of a quantum is most sensitive to the size of the single-channel current compared with its sensitivity to either the binding rates of the sites on the calcium-sensor protein or to the number of these sites that must bind a calcium ion to trigger exocytosis of a vesicle. PMID:10777721

SIGMAR1 Regulates Membrane Electrical Activity in Response to Extracellular Matrix Stimulation to Drive Cancer Cell Invasiveness.

PubMed

Crottès, David; Rapetti-Mauss, Raphael; Alcaraz-Perez, Francisca; Tichet, Mélanie; Gariano, Giuseppina; Martial, Sonia; Guizouarn, Hélène; Pellissier, Bernard; Loubat, Agnès; Popa, Alexandra; Paquet, Agnès; Presta, Marco; Tartare-Deckert, Sophie; Cayuela, Maria Luisa; Martin, Patrick; Borgese, Franck; Soriani, Olivier

2016-02-01

The sigma 1 receptor (Sig1R) is a stress-activated chaperone that regulates ion channels and is associated with pathologic conditions, such as stroke, neurodegenerative diseases, and addiction. Aberrant expression levels of ion channels and Sig1R have been detected in tumors and cancer cells, such as myeloid leukemia and colorectal cancer, but the link between ion channel regulation and Sig1R overexpression during malignancy has not been established. In this study, we found that Sig1R dynamically controls the membrane expression of the human voltage-dependent K(+) channel human ether-à-go-go-related gene (hERG) in myeloid leukemia and colorectal cancer cell lines. Sig1R promoted the formation of hERG/β1-integrin signaling complexes upon extracellular matrix stimulation, triggering the activation of the PI3K/AKT pathway. Consequently, the presence of Sig1R in cancer cells increased motility and VEGF secretion. In vivo, Sig1R expression enhanced the aggressiveness of tumor cells by potentiating invasion and angiogenesis, leading to poor survival. Collectively, our findings highlight a novel function for Sig1R in mediating cross-talk between cancer cells and their microenvironment, thus driving oncogenesis by shaping cellular electrical activity in response to extracellular signals. Given the involvement of ion channels in promoting several hallmarks of cancer, our study also offers a potential strategy to therapeutically target ion channel function through Sig1R inhibition. ©2015 American Association for Cancer Research.

Non-synaptic ion channels in insects--basic properties of currents and their modulation in neurons and skeletal muscles.

PubMed

Wicher, D; Walther, C; Wicher, C

2001-08-01

Insects are favoured objects for studying information processing in restricted neuronal networks, e.g. motor pattern generation or sensory perception. The analysis of the underlying processes requires knowledge of the electrical properties of the cells involved. These properties are determined by the expression pattern of ionic channels and by the regulation of their function, e.g. by neuromodulators. We here review the presently available knowledge on insect non-synaptic ion channels and ionic currents in neurons and skeletal muscles. The first part of this article covers genetic and structural informations, the localization of channels, their electrophysiological and pharmacological properties, and known effects of second messengers and modulators such as neuropeptides or biogenic amines. In a second part we describe in detail modulation of ionic currents in three particularly well investigated preparations, i.e. Drosophila photoreceptor, cockroach DUM (dorsal unpaired median) neuron and locust jumping muscle. Ion channel structures are almost exclusively known for the fruitfly Drosophila, and most of the information on their function has also been obtained in this animal, mainly based on mutational analysis and investigation of heterologously expressed channels. Now the entire genome of Drosophila has been sequenced, it seems almost completely known which types of channel genes--and how many of them--exist in this animal. There is much knowledge of the various types of channels formed by 6-transmembrane--spanning segments (6TM channels) including those where four 6TM domains are joined within one large protein (e.g. classical Na+ channel). In comparison, two TM channels and 4TM (or tandem) channels so far have hardly been explored. There are, however, various well characterized ionic conductances, e.g. for Ca2+, Cl- or K+, in other insect preparations for which the channels are not yet known. In some of the larger insects, i.e. bee, cockroach, locust and moth, rather detailed information has been established on the role of ionic currents in certain physiological or behavioural contexts. On the whole, however, knowledge of non-synaptic ion channels in such insects is still fragmentary. Modulation of ion currents usually involves activation of more or less elaborate signal transduction cascades. The three detailed examples for modulation presented in the second part indicate, amongst other things, that one type of modulator usually leads to concerted changes of several ion currents and that the effects of different modulators in one type of cell may overlap. Modulators participate in the adaptive changes of the various cells responsible for different physiological or behavioural states. Further study of their effects on the single cell level should help to understand how small sets of cells cooperate in order to produce the appropriate output.

Ion transport in pigmentation.

PubMed

Bellono, Nicholas W; Oancea, Elena V

2014-12-01

Skin melanocytes and ocular pigment cells contain specialized organelles called melanosomes, which are responsible for the synthesis of melanin, the major pigment in mammals. Defects in the complex mechanisms involved in melanin synthesis and regulation result in vision and pigmentation deficits, impaired development of the visual system, and increased susceptibility to skin and eye cancers. Ion transport across cellular membranes is critical for many biological processes, including pigmentation, but the molecular mechanisms by which it regulates melanin synthesis, storage, and transfer are not understood. In this review we first discuss ion channels and transporters that function at the plasma membrane of melanocytes; in the second part we consider ion transport across the membrane of intracellular organelles, with emphasis on melanosomes. We discuss recently characterized lysosomal and endosomal ion channels and transporters associated with pigmentation phenotypes. We then review the evidence for melanosomal channels and transporters critical for pigmentation, discussing potential molecular mechanisms mediating their function. The studies investigating ion transport in pigmentation physiology open new avenues for future research and could reveal novel molecular mechanisms underlying melanogenesis.

Ion transport in pigmentation

PubMed Central

Bellono, Nicholas W.; Oancea, Elena V.

2014-01-01

Skin melanocytes and ocular pigment cells contain specialized organelles called melanosomes, which are responsible for the synthesis of melanin, the major pigment in mammals. Defects in the complex mechanisms involved in melanin synthesis and regulation result in vision and pigmentation deficits, impaired development of the visual system,, and increased susceptibility to skin and eye cancers. Ion transport across cellular membranes is critical for many biological processes, including pigmentation, but the molecular mechanisms by which it regulates melanin synthesis, storage, and transfer are not understood. In this review we first discuss ion channels and transporters that function at the plasma membrane of melanocytes; in the second part we consider ion transport across the membrane of intracellular organelles, with emphasis on melanosomes. We discuss recently characterized lysosomal and endosomal ion channels and transporters associated with pigmentation phenotypes. We then review the evidence for melanosomal channels and transporters critical for pigmentation, discussing potential molecular mechanisms mediating their function. The studies investigating ion transport in pigmentation physiology open new avenues for future research and could reveal novel molecular mechanisms underlying melanogenesis. PMID:25034214

Far-field analysis of coupled bulk and boundary layer diffusion toward an ion channel entrance.

PubMed Central

Schumaker, M F; Kentler, C J

1998-01-01

We present a far-field analysis of ion diffusion toward a channel embedded in a membrane with a fixed charge density. The Smoluchowski equation, which represents the 3D problem, is approximated by a system of coupled three- and two-dimensional diffusions. The 2D diffusion models the quasi-two-dimensional diffusion of ions in a boundary layer in which the electrical potential interaction with the membrane surface charge is important. The 3D diffusion models ion transport in the bulk region outside the boundary layer. Analytical expressions for concentration and flux are developed that are accurate far from the channel entrance. These provide boundary conditions for a numerical solution of the problem. Our results are used to calculate far-field ion flows corresponding to experiments of Bell and Miller (Biophys. J. 45:279, 1984). PMID:9591651

Atomic Force Microscopy and MD Simulations Reveal Pore-Like Structures of All-D-Enantiomer of Alzheimer’s β-Amyloid Peptide: Relevance to the Ion Channel Mechanism of AD Pathology

PubMed Central

Connelly, Laura; Arce, Fernando Teran; Jang, Hyunbum; Capone, Ricardo; Kotler, Samuel A.; Ramachandran, Srinivasan; Kagan, Bruce L.; Nussinov, Ruth; Lal, Ratnesh

2012-01-01

Alzheimer’s disease (AD) is a protein misfolding disease characterized by a build-up of β-amyloid (Aβ) peptide as senile plaques, uncontrolled neurodegeneration, and memory loss. AD pathology is linked to the destabilization of cellular ionic homeostasis and involves Aβ peptide-plasma membrane interactions. In principle, there are two possible ways through which disturbance of the ionic homeostasis can take place: directly, where the Aβ peptide either inserts into the membrane and creates ion-conductive pores or destabilizes the membrane organization; or, indirectly, where the Aβ peptide interacts with existing cell membrane receptors. To distinguish between these two possible types of Aβ-membrane interactions, we took advantage of the biochemical tenet that ligand-receptor interactions are stereospecific; L-amino acid peptides, but not their D-counterparts, bind to cell membrane receptors. However, with respect to the ion channel-mediated mechanism, like L-amino acids, D-amino acid peptides will also form ion channel-like structures. Using atomic force microscopy (AFM) we imaged the structures of both D- and L-enantiomers of the full length Aβ1-42 when reconstituted in lipid bilayers. AFM imaging shows that both L- and D-Aβ isomers form similar channel-like structures. Molecular dynamics (MD) simulations support the AFM imaged 3D structures. Earlier we have shown that D-Aβ1-42 channels conduct ions similarly to their L-counter parts. Taken together, our results support the direct mechanism of Aβ ion channel-mediated destabilization of ionic homeostasis rather than the indirect mechanism through Aβ interaction with membrane receptors. PMID:22217000

Characterization of Rat Meibomian Gland Ion and Fluid Transport

PubMed Central

Yu, Dongfang; Davis, Richard M.; Aita, Megumi; Burns, Kimberlie A.; Clapp, Phillip W.; Gilmore, Rodney C.; Chua, Michael; O'Neal, Wanda K.; Schlegel, Richard; Randell, Scott H.; C. Boucher, Richard

2016-01-01

Purpose We establish novel primary rat meibomian gland (MG) cell culture systems and explore the ion transport activities of the rat MG. Methods Freshly excised rat MG tissues were characterized as follows: (1) mRNA expression of selected epithelial ion channels/transporters were measured by RT-PCR, (2) localization of epithelial sodium channel (ENaC) mRNAs was performed by in situ hybridization, and (3) protein expression and localization of βENaC, the Na+/K+/Cl− cotransporter (NKCC), and the Na+/K+ ATPase were evaluated by immunofluorescence. Primary isolated rat MG cells were cocultured with 3T3 feeder cells and a Rho-associated kinase (ROCK) inhibitor (Y-27632) for expansion. Passaged rat MG cells were cultured as planar sheets under air-liquid interface (ALI) conditions for gene expression and electrophysiologic studies. Passaged rat MG cells also were cultured in matrigel matrices to form spheroids, which were examined ultrastructurally by transmission electron microscopy (TEM) and functionally using swelling assays. Results Expression of multiple ion channel/transporter genes was detected in rat MG tissues. β-ENaC mRNA and protein were localized more to MG peripheral acinar cells than central acinar cells or ductular epithelial cells. Electrophysiologic studies of rat MG cell planar cultures demonstrated functional sodium, chloride, and potassium channels, and cotransporters activities. Transmission electron microscopic analyses of rat MG spheroids revealed highly differentiated MG cells with abundant lysosomal lamellar bodies. Rat MG spheroids culture-based measurements demonstrated active volume regulation by ion channels. Conclusions This study demonstrates the presence and function of ion channels and volume transport by rat MG. Two novel primary MG cell culture models that may be useful for MG research were established. PMID:27127933

Investigation of polarization effects in the gramicidin A channel from ab initio molecular dynamics simulations.

PubMed

Timko, Jeff; Kuyucak, Serdar

2012-11-28

Polarization is an important component of molecular interactions and is expected to play a particularly significant role in inhomogeneous environments such as pores and interfaces. Here we investigate the effects of polarization in the gramicidin A ion channel by performing quantum mechanics/molecular mechanics molecular dynamics (MD) simulations and comparing the results with those obtained from classical MD simulations with non-polarizable force fields. We consider the dipole moments of backbone carbonyl groups and channel water molecules as well as a number of structural quantities of interest. The ab initio results show that the dipole moments of the carbonyl groups and water molecules are highly sensitive to the hydrogen bonds (H-bonds) they participate in. In the absence of a K(+) ion, water molecules in the channel are quite mobile, making the H-bond network highly dynamic. A central K(+) ion acts as an anchor for the channel waters, stabilizing the H-bond network and thereby increasing their average dipole moments. In contrast, the K(+) ion has little effect on the dipole moments of the neighboring carbonyl groups. The weakness of the ion-peptide interactions helps to explain the near diffusion-rate conductance of K(+) ions through the channel. We also address the sampling issue in relatively short ab initio MD simulations. Results obtained from a continuous 20 ps ab initio MD simulation are compared with those generated by sampling ten windows from a much longer classical MD simulation and running each window for 2 ps with ab initio MD. Both methods yield similar results for a number of quantities of interest, indicating that fluctuations are fast enough to justify the short ab initio MD simulations.

Structural dynamics of the cell nucleus: basis for morphology modulation of nuclear calcium signaling and gene transcription.

PubMed

Queisser, Gillian; Wiegert, Simon; Bading, Hilmar

2011-01-01

Neuronal morphology plays an essential role in signal processing in the brain. Individual neurons can undergo use-dependent changes in their shape and connectivity, which affects how intracellular processes are regulated and how signals are transferred from one cell to another in a neuronal network. Calcium is one of the most important intracellular second messengers regulating cellular morphologies and functions. In neurons, intracellular calcium levels are controlled by ion channels in the plasma membrane such as NMDA receptors (NMDARs), voltage-gated calcium channels (VGCCs) and certain α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) as well as by calcium exchange pathways between the cytosol and internal calcium stores including the endoplasmic reticulum and mitochondria. Synaptic activity and the subsequent opening of ligand and/or voltage-gated calcium channels can initiate cytosolic calcium transients which propagate towards the cell soma and enter the nucleus via its nuclear pore complexes (NPCs) embedded in the nuclear envelope. We recently described the discovery that in hippocampal neurons the morphology of the nucleus affects the calcium dynamics within the nucleus. Here we propose that nuclear infoldings determine whether a nucleus functions as an integrator or detector of oscillating calcium signals. We outline possible ties between nuclear mophology and transcriptional activity and discuss the importance of extending the approach to whole cell calcium signal modeling in order to understand synapse-to-nucleus communication in healthy and dysfunctional neurons.

Conductance of Ion Channels - Theory vs. Experiment

NASA Technical Reports Server (NTRS)

Pohorille, Andrew; Wilson, Michael; Mijajlovic, Milan

2013-01-01

Transmembrane ion channels mediate a number of essential physiological processes in a cell ranging from regulating osmotic pressure to transmission of neural signals. Kinetics and selectivity of ion transport is of critical importance to a cell and, not surprisingly, it is a subject of numerous experimental and theoretical studies. In this presentation we will analyze in detail computer simulations of two simple channels from fungi - antiamoebin and trichotoxin. Each of these channels is made of an alpha-helical bundle of small, nongenomically synthesized peptides containing a number of rare amino acids and exhibits strong antimicrobial activity. We will focus on calculating ionic conductance defined as the ratio of ionic current through the channel to applied voltage. From molecular dynamics simulations, conductance can be calculated in at least two ways, each involving different approximations. Specifically, the current, given as the number of charges transferred through the channel per unit of time, can be obtained from the number of events in which ions cross the channel during the simulation. This method works well for large currents (high conductance values and/or applied voltages). If the number of crossing events is small, reliable estimates of current are difficult to achieve. Alternatively, conductance can be estimated assuming that ion transport can be well approximated as diffusion in the external potential given by the free energy profile. Then, the current can be calculated by solving the one-dimensional diffusion equation in this external potential and applied voltage (the generalized Nernst-Planck equation). To do so three ingredients are needed: the free energy profile, the position-dependent diffusion coefficient and the diffusive flux of ions into the channel. All these quantities can be obtained from molecular dynamics simulations. An important advantage of this method is that it can be used equally well to estimating large and small currents. In addition, once the free energy profile becomes available the full current-voltage dependence can be readily obtained. For both channels we carried out calculations using both approaches. We also tested the main assumptions underlying the diffusive model, such as uncorrelated nature of individual crossing events and Fickian diffusion. The accuracy and consistency of different methods will be discussed. Finally we will discuss how comparisons between calculated and measured ionic conductance and selectivity of transport can be used for determining structural models of the channels.

Use of planar array electrophysiology for the development of robust ion channel cell lines.

PubMed

Clare, Jeffrey J; Chen, Mao Xiang; Downie, David L; Trezise, Derek J; Powell, Andrew J

2009-01-01

The tractability of ion channels as drug targets has been significantly improved by the advent of planar array electrophysiology platforms which have dramatically increased the capacity for electrophysiological profiling of lead series compounds. However, the data quality and through-put obtained with these platforms is critically dependent on the robustness of the expression reagent being used. The generation of high quality, recombinant cell lines is therefore a key step in the early phase of ion channel drug discovery and this can present significant challenges due to the diversity and organisational complexity of many channel types. This article focuses on several complex and difficult to express ion channels and illustrates how improved stable cell lines can be obtained by integration of planar array electrophysiology systems into the cell line generation process per se. By embedding this approach at multiple stages (e.g., during development of the expression strategy, during screening and validation of clonal lines, and during characterisation of the final cell line), the cycle time and success rate in obtaining robust expression of complex multi-subunit channels can be significantly improved. We also review how recent advances in this technology (e.g., population patch clamp) have further widened the versatility and applicability of this approach.

Binding site in eag voltage sensor accommodates a variety of ions and is accessible in closed channel.

PubMed

Silverman, William R; Bannister, John P A; Papazian, Diane M

2004-11-01

In ether-a-go-go K+ channels, voltage-dependent activation is modulated by ion binding to a site located in an extracellular-facing crevice between transmembrane segments S2 and S3 in the voltage sensor. We find that acidic residues D278 in S2 and D327 in S3 are able to coordinate a variety of divalent cations, including Mg2+, Mn2+, and Ni2+, which have qualitatively similar functional effects, but different half-maximal effective concentrations. Our data indicate that ions binding to individual voltage sensors in the tetrameric channel act without cooperativity to modulate activation gating. We have taken advantage of the unique phenotype of Ni2+ in the D274A channel, which contains a mutation of a nonbinding site residue, to demonstrate that ions can access the binding site from the extracellular solution when the voltage sensor is in the resting conformation. Our results are difficult to reconcile with the x-ray structure of the KvAP K+ channel, in which the binding site residues are widely separated, and with the hydrophobic paddle model for voltage-dependent activation, in which the voltage sensor domain, including the S3-S4 loop, is near the cytoplasmic side of the membrane in the closed channel.

Glutamate Signaling and Mitochondrial Dysfunction in Models of Parkinson’s Disease

DTIC Science & Technology

2014-03-01

stages of PD, an elevation in synaptically released glutamate leads to persistent activation of NMDARs that synergizes with Cav1 calcium channels to...neurons is attributable to activity -dependent calcium entry through Cav1 channels, resulting in mitochondrial oxidant stress. Although this mechanism...glutamate leads to persistent activation of NMDARs that synergizes with Cav1 calcium channels to significantly increase mitochondrial oxidant stress and

Calcium Homeostatasis and Mitochondrial Dysfunction in Dopaminergic Neurons of the Substantia Nigra

DTIC Science & Technology

2010-03-01

discovery that calcium entry through L-type channels during normal pacemaking elevates the sensitivity of SNc dopaminergic neurons to toxins; • the...discovery that L-type calcium channels participate in but are not necessary for pacemaking; • the discovery that serum concentration of the...FDA approved doses; • the discovery that calcium entry through L-type channels during pacemaking elevates mitochondrial oxidant stress and leads

Laser Induced Fluorescence Measurements in a Hall Thruster Plume as a Function of Background Pressure

NASA Technical Reports Server (NTRS)

Spektor, R.; Tighe, W. G.; Kamhawi, H.

2016-01-01

A set of Laser Induced Fluorescence (LIF) measurements in the near-field region of the NASA- 173M Hall thruster plume is presented at four background pressure conditions varying from 9.4 x 10(exp -6) torr to 3.3 x 10(exp -5) torr. The xenon ion velocity distribution function was measured simultaneously along the axial and radial directions. An ultimate exhaust velocity of 19.6+/-0.25 km/s achieved at a distance of 20 mm was measured, and that value was not sensitive to pressure. On the other hand, the ion axial velocity at the thruster exit was strongly influenced by pressure, indicating that the accelerating electric field moved inward with increased pressure. The shift in electric field corresponded to an increase in measured thrust. Pressure had a minor effect on the radial component of ion velocity, mainly affecting ions exiting close to the channel inner wall. At that radial location the radial component of ion velocity was approximately 1000 m/s greater at the lowest pressure than at the highest pressure. A reduction of the inner magnet coil current by 0.6 A resulted in a lower axial ion velocity at the channel exit while the radial component of ion velocity at the channel inner wall location increased by 1300 m/s, and at the channel outer wall location the radial ion velocity remained unaffected. The ultimate exhaust velocity was not significantly affected by the inner magnet current.

Atomistic Modeling of Ion Conduction through the Voltage-Sensing Domain of the Shaker K+ Ion Channel.

PubMed

Wood, Mona L; Freites, J Alfredo; Tombola, Francesco; Tobias, Douglas J

2017-04-20

Voltage-sensing domains (VSDs) sense changes in the membrane electrostatic potential and, through conformational changes, regulate a specific function. The VSDs of wild-type voltage-dependent K + , Na + , and Ca 2+ channels do not conduct ions, but they can become ion-permeable through pathological mutations in the VSD. Relatively little is known about the underlying mechanisms of conduction through VSDs. The most detailed studies have been performed on Shaker K + channel variants in which ion conduction through the VSD is manifested in electrophysiology experiments as a voltage-dependent inward current, the so-called omega current, which appears when the VSDs are in their resting state conformation. Only monovalent cations appear to permeate the Shaker VSD via a pathway that is believed to be, at least in part, the same as that followed by the S4 basic side chains during voltage-dependent activation. We performed μs-time scale atomistic molecular dynamics simulations of a cation-conducting variant of the Shaker VSD under applied electric fields in an experimentally validated resting-state conformation, embedded in a lipid bilayer surrounded by solutions containing guanidinium chloride or potassium chloride. Our simulations provide insights into the Shaker VSD permeation pathway, the protein-ion interactions that control permeation kinetics, and the mechanism of voltage-dependent activation of voltage-gated ion channels.

Easily Accessible Polycyclic Amines that Inhibit the Wild-Type and Amantadine-Resistant Mutants of the M2 Channel of Influenza A Virus

PubMed Central

2015-01-01

Amantadine inhibits the M2 proton channel of influenza A virus, yet most of the currently circulating strains of the virus carry mutations in the M2 protein that render the virus amantadine-resistant. While most of the research on novel amantadine analogues has revolved around the synthesis of novel adamantane derivatives, we have recently found that other polycyclic scaffolds effectively block the M2 proton channel, including amantadine-resistant mutant channels. In this work, we have synthesized and characterized a series of pyrrolidine derivatives designed as analogues of amantadine. Inhibition of the wild-type M2 channel and the A/M2-S31N, A/M2-V27A, and A/M2-L26F mutant forms of the channel were measured in Xenopus oocytes using two-electrode voltage clamp assays. Most of the novel compounds inhibited the wild-type ion channel in the low micromolar range. Of note, two of the compounds inhibited the amantadine-resistant A/M2-V27A and A/M2-L26F mutant ion channels with submicromolar and low micromolar IC50, respectively. None of the compounds was found to inhibit the S31N mutant ion channel. PMID:24941437

Isolated pores dissected from human two-pore channel 2 are functional

PubMed Central

Penny, Christopher J.; Rahman, Taufiq; Sula, Altin; Miles, Andrew J.; Wallace, B. A.; Patel, Sandip

2016-01-01

Multi-domain voltage-gated ion channels appear to have evolved through sequential rounds of intragenic duplication from a primordial one-domain precursor. Whereas modularity within one-domain symmetrical channels is established, little is known about the roles of individual regions within more complex asymmetrical channels where the domains have undergone substantial divergence. Here we isolated and characterised both of the divergent pore regions from human TPC2, a two-domain channel that holds a key intermediate position in the evolution of voltage-gated ion channels. In HeLa cells, each pore localised to the ER and caused Ca2+ depletion, whereas an ER-targeted pore mutated at a residue that inactivates full-length TPC2 did not. Additionally, one of the pores expressed at high levels in E. coli. When purified, it formed a stable, folded tetramer. Liposomes reconstituted with the pore supported Ca2+ and Na+ uptake that was inhibited by known blockers of full-length channels. Computational modelling of the pore corroborated cationic permeability and drug interaction. Therefore, despite divergence, both pores are constitutively active in the absence of their partners and retain several properties of the wild-type pore. Such symmetrical ‘pore-only’ proteins derived from divergent channel domains may therefore provide tractable tools for probing the functional architecture of complex ion channels. PMID:27941820

On the mechanism of TBA block of the TRPV1 channel.

PubMed

Oseguera, Andrés Jara; Islas, León D; García-Villegas, Refugio; Rosenbaum, Tamara

2007-06-01

The transient receptor potential vanilloid 1 (TRPV1) channel is a nonselective cation channel activated by capsaicin and responsible for thermosensation. To date, little is known about the gating characteristics of these channels. Here we used tetrabutylammonium (TBA) to determine whether this molecule behaves as an ion conduction blocker in TRPV1 channels and to gain insight into the nature of the activation gate of this protein. TBA belongs to a family of classic potassium channel blockers that have been widely used as tools for determining the localization of the activation gate and the properties of the pore of several ion channels. We found TBA to be a voltage-dependent pore blocker and that the properties of block are consistent with an open-state blocker, with the TBA molecule binding to multiple open states, each with different blocker affinities. Kinetics of channel closure and burst-length analysis in the presence of blocker are consistent with a state-dependent blocking mechanism, with TBA interfering with closing of an activation gate. This activation gate may be located cytoplasmically with respect to the binding site of TBA ions, similar to what has been observed in potassium channels. We propose an allosteric model for TRPV1 activation and block by TBA, which explains our experimental data.

Isolated pores dissected from human two-pore channel 2 are functional.

PubMed

Penny, Christopher J; Rahman, Taufiq; Sula, Altin; Miles, Andrew J; Wallace, B A; Patel, Sandip

2016-12-12

Multi-domain voltage-gated ion channels appear to have evolved through sequential rounds of intragenic duplication from a primordial one-domain precursor. Whereas modularity within one-domain symmetrical channels is established, little is known about the roles of individual regions within more complex asymmetrical channels where the domains have undergone substantial divergence. Here we isolated and characterised both of the divergent pore regions from human TPC2, a two-domain channel that holds a key intermediate position in the evolution of voltage-gated ion channels. In HeLa cells, each pore localised to the ER and caused Ca 2+ depletion, whereas an ER-targeted pore mutated at a residue that inactivates full-length TPC2 did not. Additionally, one of the pores expressed at high levels in E. coli. When purified, it formed a stable, folded tetramer. Liposomes reconstituted with the pore supported Ca 2+ and Na + uptake that was inhibited by known blockers of full-length channels. Computational modelling of the pore corroborated cationic permeability and drug interaction. Therefore, despite divergence, both pores are constitutively active in the absence of their partners and retain several properties of the wild-type pore. Such symmetrical 'pore-only' proteins derived from divergent channel domains may therefore provide tractable tools for probing the functional architecture of complex ion channels.

TPC Proteins Are Phosphoinositide-activated Sodium-selective Ion Channels in Endosomes and Lysosomes

PubMed Central

Wang, Xiang; Zhang, Xiaoli; Dong, Xian-ping; Samie, Mohammad; Li, Xinran; Cheng, Xiping; Goschka, Andrew; Shen, Dongbiao; Zhou, Yandong; Harlow, Janice; Zhu, Michael X.; Clapham, David E.; Ren, Dejian; Xu, Haoxing

2012-01-01

Summary Mammalian Two-Pore Channels (TPC1, 2; TPCN1, TPCN2) encode ion channels in intracellular endosomes and lysosomes and were proposed to mediate endolysosomal calcium release triggered by the second messenger, nicotinic acid adenine dinucleotide phosphate (NAADP). By directly recording TPCs in endolysosomes from wild-type and TPC double knockout mice, here we show that, in contrast to previous conclusions, TPCs are in fact sodium-selective channels activated by PI(3,5)P2, and are not activated by NAADP. Moreover, the primary endolysosomal ion is Na+, not K+, as had been previously assumed. These findings suggest that the organellar membrane potential may undergo large regulatory changes, and may explain the specificity of PI(3,5)P2 in regulating the fusogenic potential of intracellular organelles. PMID:23063126

Cation Selectivity in Biological Cation Channels Using Experimental Structural Information and Statistical Mechanical Simulation.

PubMed

Finnerty, Justin John; Peyser, Alexander; Carloni, Paolo

2015-01-01

Cation selective channels constitute the gate for ion currents through the cell membrane. Here we present an improved statistical mechanical model based on atomistic structural information, cation hydration state and without tuned parameters that reproduces the selectivity of biological Na+ and Ca2+ ion channels. The importance of the inclusion of step-wise cation hydration in these results confirms the essential role partial dehydration plays in the bacterial Na+ channels. The model, proven reliable against experimental data, could be straightforwardly used for designing Na+ and Ca2+ selective nanopores.