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https://www.readbyqxmd.com/read/27233449/analysis-of-neuronal-nicotinic-acetylcholine-receptor-%C3%AE-4%C3%AE-2-activation-at-the-single-channel-level
#1
Camila Carignano, Esteban Pablo Barila, Guillermo Spitzmaul
The neuronal nicotinic acetylcholine receptor α4β2 forms pentameric proteins with two alternate stoichiometries. The high-sensitivity receptor is related to (α4)2(β2)3 stoichiometry while the low-sensitivity receptor to (α4)3(β2)2 stoichiometry. Both subtypes share two binding sites at the α4((+))/β2((-)) interface with high affinity for agonists. (α4)3(β2)2 has an additional binding site at the α4((+))/α4((-)) interface with low affinity for agonists. We investigated activation kinetics of both receptor subtypes by patch-clamp recordings of single-channel activity in the presence of several concentrations of acetylcholine (0...
September 2016: Biochimica et Biophysica Acta
https://www.readbyqxmd.com/read/26585318/nicotinic-acetylcholine-receptors-nachrs-at-zebrafish-red-and-white-muscle-show-different-properties-during-development
#2
Kazi T Ahmed, Declan W Ali
Nicotinic acetylcholine receptors (nAChRs) are highly expressed at the vertebrate neuromuscular junction (NMJ) where they are required for muscle activation. Understanding the factors that underlie NMJ development is critical for a full understanding of muscle function. In this study we performed whole cell and outside-out patch clamp recordings, and single-cell RT-qPCR from zebrafish red and white muscle to examine the properties of nAChRs during the first 5 days of development. In red fibers miniature endplate currents (mEPCs) exhibit single exponential time courses at 1...
August 2016: Developmental Neurobiology
https://www.readbyqxmd.com/read/26494801/mechanisms-of-nicotinic-modulation-of-glutamatergic-neuroplasticity-in-humans
#3
Marcelo Di Marcello Valladão Lugon, Giorgi Batsikadze, Shane Fresnoza, Jessica Grundey, Min-Fang Kuo, Walter Paulus, Ester Miyuki Nakamura-Palacios, Michael A Nitsche
The impact of nicotine (NIC) on plasticity is thought to be primarily determined via calcium channel properties of nicotinic receptor subtypes, and glutamatergic plasticity is likewise calcium-dependent. Therefore glutamatergic plasticity is likely modulated by the impact of nicotinic receptor-dependent neuronal calcium influx. We tested this hypothesis for transcranial direct current stimulation (tDCS)-induced long-term potentiation-like plasticity, which is abolished by NIC in nonsmokers. To reduce calcium influx under NIC, we blocked N-methyl-d-aspartate (NMDA) receptors...
October 22, 2015: Cerebral Cortex
https://www.readbyqxmd.com/read/25908400/the-coupling-interface-and-pore-domain-codetermine-the-single-channel-activity-of-the-%C3%AE-7-nicotinic-receptor
#4
Hongxia Yan, Na Pan, Fenqin Xue, Yan Zheng, Chaokun Li, Yongchang Chang, Zhiqing Xu, Hui Yang, Jianliang Zhang
Ligand-gated ion channels play a role in mediating fast synaptic transmission for communication between neurons. However, the structural basis for the functional coupling of the binding and pore domains, resulting in channel opening, remains a topic of intense investigation. Here, a series of α7 nicotinic receptor mutants were constructed for expression in cultured mammalian cells, and their single-channel properties were examined using the patch-clamp technique combined with radio ligand binding and the fluorescence staining technique...
August 2015: Neuropharmacology
https://www.readbyqxmd.com/read/25870122/activation-of-the-macrophage-%C3%AE-7-nicotinic-acetylcholine-receptor-and-control-of-inflammation
#5
REVIEW
Carlos A Báez-Pagán, Manuel Delgado-Vélez, José A Lasalde-Dominicci
Inflammatory responses to stimuli are essential body defenses against foreign threats. However, uncontrolled inflammation may result in serious health problems, which can be life-threatening. The α7 nicotinic acetylcholine receptor, a ligand-gated ion channel expressed in the nervous and immune systems, has an essential role in the control of inflammation. Activation of the macrophage α7 receptor by acetylcholine, nicotine, or other agonists, selectively inhibits production of pro-inflammatory cytokines while leaving anti-inflammatory cytokines undisturbed...
September 2015: Journal of Neuroimmune Pharmacology: the Official Journal of the Society on NeuroImmune Pharmacology
https://www.readbyqxmd.com/read/25548135/mechanism-of-activation-of-the-prokaryotic-channel-elic-by-propylamine-a-single-channel-study
#6
Alessandro Marabelli, Remigijus Lape, Lucia Sivilotti
Prokaryotic channels, such as Erwinia chrysanthemi ligand-gated ion channel (ELIC) and Gloeobacter violaceus ligand-gated ion channel, give key structural information for the pentameric ligand-gated ion channel family, which includes nicotinic acetylcholine receptors. ELIC, a cationic channel from E. chrysanthemi, is particularly suitable for single-channel recording because of its high conductance. Here, we report on the kinetic properties of ELIC channels expressed in human embryonic kidney 293 cells. Single-channel currents elicited by the full agonist propylamine (0...
January 2015: Journal of General Physiology
https://www.readbyqxmd.com/read/24972488/characterization-of-ryanodine-receptor-type-1-single-channel-activity-using-on-nucleus-patch-clamp
#7
Larry E Wagner, Linda A Groom, Robert T Dirksen, David I Yule
In this study, we provide the first description of the biophysical and pharmacological properties of ryanodine receptor type 1 (RyR1) expressed in a native membrane using the on-nucleus configuration of the patch clamp technique. A stable cell line expressing rabbit RyR1 was established (HEK-RyR1) using the FLP-in 293 cell system. In contrast to untransfected cells, RyR1 expression was readily demonstrated by immunoblotting and immunocytochemistry in HEK-RyR1 cells. In addition, the RyR1 agonists 4-CMC and caffeine activated Ca(2+) release that was inhibited by high concentrations of ryanodine...
August 2014: Cell Calcium
https://www.readbyqxmd.com/read/24467848/a-signal-peptide-missense-mutation-associated-with-nicotine-dependence-alters-%C3%AE-2-nicotinic-acetylcholine-receptor-function
#8
Bhagirathi Dash, Ronald J Lukas, Ming D Li
A cytosine to thymidine (C → T) missense mutation in the signal peptide (SP) sequence (rs2472553) of the nicotinic acetylcholine receptor (nAChR) α2 subunit produces a threonine-to-isoleucine substitution (T22I) often associated with nicotine dependence (ND). We assessed effects on function of α2*-nAChR ('*'indicates presence of additional subunits) of this mutation, which could alter SP cleavage, RNA/protein secondary structure, and/or efficiency of transcription, translation, subunit assembly, receptor trafficking or cell surface expression...
April 2014: Neuropharmacology
https://www.readbyqxmd.com/read/23973209/a-presynaptic-enac-channel-drives-homeostatic-plasticity
#9
Meg A Younger, Martin Müller, Amy Tong, Edward C Pym, Graeme W Davis
An electrophysiology-based forward genetic screen has identified two genes, pickpocket11 (ppk11) and pickpocket16 (ppk16), as being necessary for the homeostatic modulation of presynaptic neurotransmitter release at the Drosophila neuromuscular junction (NMJ). Pickpocket genes encode Degenerin/Epithelial Sodium channel subunits (DEG/ENaC). We demonstrate that ppk11 and ppk16 are necessary in presynaptic motoneurons for both the acute induction and long-term maintenance of synaptic homeostasis. We show that ppk11 and ppk16 are cotranscribed as a single mRNA that is upregulated during homeostatic plasticity...
September 18, 2013: Neuron
https://www.readbyqxmd.com/read/23839567/multiple-modes-of-%C3%AE-7-nachr-noncompetitive-antagonism-of-control-agonist-evoked-and-allosterically-enhanced-currents
#10
Can Peng, Matthew R Kimbrell, Chengju Tian, Thomas F Pack, Peter A Crooks, E Kim Fifer, Roger L Papke
Positive allosteric modulators (PAMs) of α7 nicotinic acetylcholine receptors can enhance ion channel currents and downstream effects of α7 stimulation. We investigated the approach of using noncompetitive antagonists to regulate α7 receptor function, potentially distinguishing effects requiring ion channel currents from signaling induced by nonconducting states. Three small readily reversible antagonists, (1S,2R,4R)-N,2,3,3-tetramethylbicyclo[2.2.1]heptan-2-amine (mecamylamine), N-(2.6-dimethylphenylcarbamoylmethyl)triethylammonium bromide (QX-314), and 2-(dimethylamino)ethyl 4-(butylamino)benzoate (tetracaine), as well as three large slowly reversible antagonists, bis-(2,2,6,6-tetramethyl-4-piperidinyl) sebacate (BTMPS), 2,2,6,6-tetramethylpiperidin-4-yl heptanoate (TMPH), and 1,2,4,5-tetra-{5-[1-(3-benzyl)pyridinium]pent-1-yl}benzene tetrabromide (tkP3BzPB), were investigated for their effectiveness and voltage dependence in the inhibition of responses evoked by acetylcholine alone or augmented by the α7-selective PAM N-(5-chloro-2,4-dimethoxyphenyl)-N'-(5-methyl-3-isoxazolyl)-urea (PNU-120596)...
September 2013: Molecular Pharmacology
https://www.readbyqxmd.com/read/23740249/the-minimum-m3-m4-loop-length-of-neurotransmitter-activated-pentameric-receptors-is-critical-for-the-structural-integrity-of-cytoplasmic-portals
#11
Daniel T Baptista-Hon, Tarek Z Deeb, Jeremy J Lambert, John A Peters, Tim G Hales
The 5-HT3A receptor homology model, based on the partial structure of the nicotinic acetylcholine receptor from Torpedo marmorata, reveals an asymmetric ion channel with five portals framed by adjacent helical amphipathic (HA) stretches within the 114-residue loop between the M3 and M4 membrane-spanning domains. The positive charge of Arg-436, located within the HA stretch, is a rate-limiting determinant of single channel conductance (γ). Further analysis reveals that positive charge and volume of residue 436 are determinants of 5-HT3A receptor inward rectification, exposing an additional role for portals...
July 26, 2013: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/23576748/stoichiometry-for-drug-potentiation-of-a-pentameric-ion-channel
#12
Corrie J B daCosta, Steven M Sine
Drug modulation of ion channels is a powerful means to alter physiological responses for therapeutic benefit, yet the structural bases of modulation remain poorly understood. Here we study potentiation of nicotinic α7 acetylcholine receptors, which are emerging drug targets in several neurological disorders. α7 receptors are ligand-gated ion channels composed of five identical subunits, each bearing a site for the potentiating drug PNU-120596 (PNU). How the individual subunits contribute to PNU potentiation is not known...
April 16, 2013: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/23562945/whole-cell-patch-clamp-recording-of-nicotinic-acetylcholine-receptors-in-adult-brugia-malayi-muscle
#13
A P Robertson, S K Buxton, R J Martin
Lymphatic filariasis is a debilitating disease caused by clade III parasites like Brugia malayi and Wuchereria bancrofti. Current recommended treatment regimen for this disease relies on albendazole, ivermectin and diethylcarbamazine, none of which targets the nicotinic acetylcholine receptors in these parasitic nematodes. Our aim therefore has been to develop adult B. malayi for electrophysiological recordings to aid in characterizing the ion channels in this parasite as anthelmintic target sites. In that regard, we recently demonstrated the amenability of adult B...
December 2013: Parasitology International
https://www.readbyqxmd.com/read/23277476/action-of-nicotine-and-analogs-on-acetylcholine-receptors-having-mutations-of-transmitter-binding-site-residue-%C3%AE-g153
#14
Snehal Jadey, Prasad Purohit, Anthony Auerbach
A primary target for nicotine is the acetylcholine receptor channel (AChR). Some of the ability of nicotine to activate differentially AChR subtypes has been traced to a transmitter-binding site amino acid that is glycine in lower affinity and lysine in higher affinity AChRs. We studied the effects of mutations of this residue (αG153) in neuromuscular AChRs activated by nicotine and eight other agonists including nornicotine and anabasine. All of the mutations increased the unliganded gating equilibrium constant...
January 2013: Journal of General Physiology
https://www.readbyqxmd.com/read/23128482/local-application-of-drugs-to-study-nicotinic-acetylcholine-receptor-function-in-mouse-brain-slices
#15
Staci E Engle, Hilary J Broderick, Ryan M Drenan
Tobacco use leads to numerous health problems, including cancer, heart disease, emphysema, and stroke. Addiction to cigarette smoking is a prevalent neuropsychiatric disorder that stems from the biophysical and cellular actions of nicotine on nicotinic acetylcholine receptors (nAChRs) throughout the central nervous system. Understanding the various nAChR subtypes that exist in brain areas relevant to nicotine addiction is a major priority. Experiments that employ electrophysiology techniques such as whole-cell patch clamp or two-electrode voltage clamp recordings are useful for pharmacological characterization of nAChRs of interest...
2012: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/23064317/the-unanticipated-complexity-of-the-selectivity-filter-glutamates-of-nicotinic-receptors
#16
Gisela D Cymes, Claudio Grosman
In ion channels, 'rings' of ionized side chains that decorate the walls of the permeation pathway often lower the energetic barrier to ion conduction. Using single-channel electrophysiological recordings, we studied the poorly understood ring of four glutamates (and one glutamine) that dominates this catalytic effect in the muscle nicotinic acetylcholine receptor ('the intermediate ring of charge'). We show that all four wild-type glutamate side chains are deprotonated in the range of 6.0-9.0 pH, that only two of them contribute to the size of the single-channel current, that these side chains must be able to adopt alternate conformations that either allow or prevent their negative charges from increasing the rate of cation conduction and that the location of these glutamate side chains squarely at one of the ends of the transmembrane pore is critical for their largely unshifted pK(a) values and for the unanticipated impact of their conformational flexibility on cation permeation...
December 2012: Nature Chemical Biology
https://www.readbyqxmd.com/read/23016960/a-nicotinic-acetylcholine-receptor-transmembrane-point-mutation-g275e-associated-with-resistance-to-spinosad-in-frankliniella-occidentalis
#17
Alin M Puinean, Stuart J Lansdell, Toby Collins, Pablo Bielza, Neil S Millar
High levels of resistance to spinosad, a macrocyclic lactone insecticide, have been reported previously in western flower thrips, Frankliniella occidentalis, an economically important insect pest of vegetables, fruit and ornamental crops. We have cloned the nicotinic acetylcholine receptor (nAChR) α6 subunit from F. occidentalis (Foα6) and compared the nucleotide sequence of Foα6 from susceptible and spinosad-resistant insect populations (MLFOM and R1S respectively). A single nucleotide change has been identified in Foα6, resulting in the replacement of a glycine (G) residue in susceptible insects with a glutamic acid (E) in resistant insects...
March 2013: Journal of Neurochemistry
https://www.readbyqxmd.com/read/22874415/activation-of-%C3%AE-7-nicotinic-receptors-by-orthosteric-and-allosteric-agonists-influence-on-single-channel-kinetics-and-conductance
#18
COMPARATIVE STUDY
Magda M Pałczyńska, Marie Jindrichova, Alasdair J Gibb, Neil S Millar
Nicotinic acetylcholine receptors (nAChRs) are oligomeric transmembrane proteins in which five subunits coassemble to form a central ion channel pore. Conventional agonists, such as acetylcholine (ACh), bind to an orthosteric site, located at subunit interfaces in the extracellular domain. More recently, it has been demonstrated that nAChRs can also be activated by ligands binding to an allosteric transmembrane site. In the case of α7 nAChRs, ACh causes rapid activation and almost complete desensitization...
November 2012: Molecular Pharmacology
https://www.readbyqxmd.com/read/22733356/mechanosensitivity-of-nicotinic-receptors
#19
N Clara Pan, Jin Jin Ma, H Benjamin Peng
Nicotinic acetylcholine receptors (nAChRs) are heteropentameric ligand-gated ion channels that mediate excitatory neurotransmission at the neuromuscular junction (NMJ) and other peripheral and central synapses. At the NMJ, acetylcholine receptors (AChRs) are constantly exposed to mechanical stress resulting from muscle contraction. It is therefore of interest to understand if their function is influenced by mechanical stimuli. In this study, patch-clamp recordings showed that AChR channel activity was enhanced upon membrane stretching in both cultured Xenopus muscle cells and C2C12 myotubes...
August 2012: Pflügers Archiv: European Journal of Physiology
https://www.readbyqxmd.com/read/22728938/myasthenic-syndrome-achr%C3%AE-c-loop-mutant-disrupts-initiation-of-channel-gating
#20
Xin-Ming Shen, Joan M Brengman, Steven M Sine, Andrew G Engel
Congenital myasthenic syndromes (CMSs) are neuromuscular disorders that can be caused by defects in ace-tylcholine receptor (AChR) function. Disease-associated point mutants can reveal the unsuspected functional significance of mutated residues. We identified two pathogenic mutations in the extracellular domain of the AChR α subunit (AChRα) in a patient with myasthenic symptoms since birth: a V188M mutation in the C-loop and a heteroallelic G74C mutation in the main immunogenic region. The G74C mutation markedly reduced surface AChR expression in cultured cells, whereas the V188M mutant was expressed robustly but had severely impaired kinetics...
July 2012: Journal of Clinical Investigation
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