Homomeric and Heteromeric α7 Nicotinic Acetylcholine Receptors in Health and Some Central Nervous System Diseases.
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ABSTRACT:
SUBMITTER: Borroni V
PROVIDER: S-EPMC8465519 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Prototoxins are a diverse family of membrane-tethered molecules expressed in the nervous system that modulate nicotinic cholinergic signaling, but their functions and specificity have yet to be completely explored. We tested the selectivity and efficacy of leukocyte antigen, PLAUR (plasminogen activator, urokinase receptor) domain-containing (LYPD)-6B on α3β4-, α3α5β4-, and α7-containing nicotinic acetylcholine receptors (nAChRs). To constrain stoichiometry, fusion proteins encoding concatemers of human α3, β4, and α5 (D and N variants) subunits were expressed in Xenopus laevis oocytes and tested with or without LYPD6B. We used the 2-electrode voltage-clamp method to quantify responses to acetylcholine (ACh): agonist sensitivity (EC50), maximal agonist-induced current (Imax), and time constant (τ) of desensitization. For β4-α3-α3-β4-α3 and β4-α3-β4-α3-α3, LYPD6B decreased EC50 from 631 to 79 μM, reduced Imax by at least 59%, and decreased τ. For β4-α3-α5D-β4-α3 and β4-α3-β4-α-α5D, LYPD6B decreased Imax by 63 and 32%, respectively. Thus, LYPD6B acted only on (α3)3(β4)2 and (α3)2(α5D)(β4)2 and did not affect the properties of (α3)2(β4)3, α7, or (α3)2(α5N)(β4)2 nAChRs. Therefore, LYPD6B acts as a mixed modulator that enhances the sensitivity of (α3)3(β4)2 nAChRs to ACh while reducing ACh-induced whole-cell currents. LYPD6B also negatively modulates α3β4 nAChRs that include the α5D common human variant, but not the N variant associated with nicotine dependence.
| S-EPMC4750422 | biostudies-literature
Project description:Heteromeric nicotinic ACh receptors (nAChRs) were thought to have two orthodox agonist-binding sites at two ?/? subunit interfaces. Highly selective ligands are hard to develop by targeting orthodox agonist sites because of high sequence similarity of this binding pocket among different subunits. Recently, unorthodox ACh-binding sites have been discovered at some ?/? and ?/? subunit interfaces, such as ?4/?4, ?5/?4 and ?3/?4. Targeting unorthodox sites may yield subtype-selective ligands, such as those for (?4?2)2 ?5, (?4?2)2 ?3 and (?6?2)2 ?3 nAChRs. The unorthodox sites have unique pharmacology. Agonist binding at one unorthodox site is not sufficient to activate nAChRs, but it increases activation from the orthodox sites. NS9283, a selective agonist for the unorthodox ?4/?4 site, was initially thought to be a positive allosteric modulator (PAM). NS9283 activates nAChRs with three engineered ?4/?4 sites. PAMs, on the other hand, act at allosteric sites where ACh cannot bind. Known PAM sites include the ACh-homologous non-canonical site (e.g. morantel at ?/?), the C-terminus (e.g. Br-PBTC and 17?-estradiol), a transmembrane domain (e.g. LY2087101) or extracellular and transmembrane domain interfaces (e.g. NS206). Some of these PAMs, such as Br-PBTC and 17?-estradiol, require only one subunit to potentiate activation of nAChRs. In this review, we will discuss differences between activation from orthosteric and allosteric sites, their selective ligands and clinical implications. These studies have advanced understanding of the structure, assembly and pharmacology of heteromeric neuronal nAChRs. LINKED ARTICLES:This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc.
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Project description:Virus entry into animal cells is initiated by attachment to target macromolecules located on host cells. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) trimeric spike glycoprotein targets host angiotensin converting enzyme 2 to gain cellular access. The SARS-CoV-2 glycoprotein contains a neurotoxin-like region that has sequence similarities to the rabies virus and the human immunodeficiency virus glycoproteins, as well as to snake neurotoxins, which interact with nicotinic acetylcholine receptor (nAChR) subtypes via this region. Using a peptide of the neurotoxin-like region of SARS-CoV-2 (SCoV2P), we identified that this area moderately inhibits α3β2, α3β4 and α4β2 subtypes, while potentiating and inhibiting α7 nAChRs. These nAChR subtypes are found in target tissues including the nose, lung, central nervous system and immune cells. Importantly, SCoV2P potentiates and inhibits ACh-induced α7 nAChR responses by an allosteric mechanism, with nicotine enhancing these effects. Live-cell confocal microscopy was used to confirm that SCoV2P interacts with α7 nAChRs in transfected neuronal-like N2a and human embryonic kidney 293 cells. The SARS-CoV-2 ectodomain functionally potentiates and inhibits the α7 subtype with nanomolar potency. Our functional findings identify that the α7 nAChR is a target for the SARS-CoV-2 glycoprotein, providing a new aspect to our understanding of SARS-CoV-2 and host cell interactions, in addition to disease pathogenesis.
| S-EPMC10101490 | biostudies-literature
Project description:A series of 9H-fluoren-9-one substituents were synthesized and evaluated for imaging cerebral α7-nAChRs. Meta-iodine substituted 9-fluorenone 5 with high binding affinity (K i = 9.3 nM) and selectivity was radiolabeled with 125I. Fully in vitro and in vivo studies of [125I]5 have been performed. [125I]5 exhibited well brain uptake with a peak concentration of 7.5 ± 0.9% ID/g in mice brains. Moreover, ex vivo autoradiography studies and micro single-photon emission computed tomography (micro-SPECT/CT) dynamic imaging in mice confirmed its in vivo imaging properties. Besides, molecular docking and MD studies were also performed to interpret the binding mechanisms of the two series of ligands towards α7-nAChRs. To conclude, the meta-iodine substituted 9-fluorenone [125I]5 could be a promising tracer for imaging α7-nAChRs.
| S-EPMC7451070 | biostudies-literature
Project description:The α7 nicotinic acetylcholine receptor (nAChR) is expressed in neuronal and non-neuronal cells and is involved in several physiopathological processes, and is thus an important drug target. We have designed and synthesized novel piperidine derivatives as α7 nAChR antagonists. Thus, we describe here a new series of 1-[2-(4-alkoxy-phenoxy-ethyl)]piperidines and 1-[2-(4-alkyloxy-phenoxy-ethyl)]-1-methylpiperidinium iodides (compounds 11a-11c and 12a-12c), and their actions on α7 nAChRs. The pharmacological activity of these compounds was studied in rat CA1 hippocampal interneurons by using the whole-cell voltage-clamp technique. Inhibition of the choline-induced current was less for 11a-11c than for the methylpiperidinium iodides 12a-12c and depended on the length of the aliphatic chain. Those compounds showing strong effects were studied further using molecular docking and molecular dynamics simulations. The strongest and non-voltage dependent antagonism was shown by 12a, which could establish cation-π interactions with the principal (+)-side and van der Waals interactions with the complementary (-)-side in the α7 nAChRs. Furthermore, compound 11a forms hydrogen bonds with residue Q115 of the complementary (-)-side through water molecules without forming cation-π interactions. Our findings have led to the establishment of a new family of antagonists that interact with the agonist binding cavity of the α7 nAChR, which represent a promising new class of compounds for the treatment of pathologies where these receptors need to be negatively modulated, including neuropsychiatric disorders as well as different types of cancer.
| S-EPMC6048275 | biostudies-literature
Project description:Cholinergic innervation of the hippocampus uses the neurotransmitter acetylcholine (ACh) to coordinate neuronal circuit activity while simultaneously influencing the function of non-neuronal cell types. The α7 nicotinic ACh receptor (nAChR) subtype is highly expressed throughout the hippocampus, has the highest calcium permeability compared with other subtypes of nAChRs, and is of high therapeutic interest due to its association with a variety of neurological disorders and neurodegenerative diseases. In this review, we synthesize research describing α7 nAChR properties, function, and relationship to cognitive dysfunction within the hippocampal circuit and highlight approaches to help improve therapeutic development.
| S-EPMC8914277 | biostudies-literature
Project description:PurposeThe α7 nicotinic acetylcholine receptor (nAChR) is widely expressed in the nervous system, including in the inner retinal neurons in all species studied to date. Although reductions in the expression of α7 nAChRs are thought to contribute to the memory and visual deficits reported in Alzheimer's disease (AD) and schizophrenia , the α7 nAChR knockout (KO) mouse is viable and has only slight visual dysfunction. The absence of a major phenotypic abnormality may be attributable to developmental mechanisms that serve to compensate for α7 nAChR loss. We hypothesized that the upregulation of genes encoding other nAChR subunits or muscarinic acetylcholine receptor (mAChR) subtypes during development partially accounts for the absence of major deficiencies in the α7 nAChR KO mouse. The purpose of this study was to determine whether the deletion of the α7 nAChR subunit in a mouse model resulted in changes in the regulation of other cholinergic receptors or other ion channels in an α7 nAChR KO mouse when compared to a wild-type (WT) mouse.MethodsTo examine gene expression changes, we employed a quantitative real-time polymerase chain reaction (qPCR) using whole retina RNA extracts as well as RNA extracted from selected regions of the retina. These extracts were collected using laser capture microdissection (LCM). The presence of acetylcholine receptor (AChR) subunit and subtype proteins was determined via western blotting. To determine any differences in the number and distribution of choline acetyltransferase (ChAT) amacrine cells, we employed wholemount and vertical immunohistochemistry (IHC) and cell counting. Additionally, in both WT and α7 nAChR KO mouse retinas, the distribution of the nAChR subunit and mAChR subtype proteins were determined via IHC for those KO mice that experienced mRNA changes.ResultsIn the whole retina, there was a statistically significant upregulation of α2, α9, α10, β4, nAChR subunit, and m1 and m4 mAChR subtype transcripts in the α7 nAChR KO mice. However, the retinal layers showed complex patterns of transcript expression. In the ganglion cell layer (GCL), m2 and m4 mAChR subtype transcripts were significantly upregulated, while β3 and β4 nAChR subunit transcripts were significantly downregulated. In the inner portion of the inner nuclear layer (iINL), α2, α9, β4, nAChR subunit, and m3 and m4 mAChR subtype transcripts were significantly downregulated. In the outer portion of the inner nuclear layer (oINL), β2, β4, and m4 AChR subunit transcripts were significantly upregulated. Western blot experiments confirmed the protein expression of α3-α5 and α9-containing nAChR subunits and m1-m2 mAChR subtypes in mouse retinas. IHC results supported many of the mRNA changes observed. Finally, this is the first report of α9 and α10 nAChR subunit expressions in the retina of any species.ConclusionsRather than a simple upregulation of a single AChR subunit or subtype, the absence of the α7 nAChR in the KO mice was associated with complex layer-specific changes in the expression of AChR subunits and subtypes.
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Project description:The α7 nicotinic acetylcholine receptor (α7 nAChR) is involved in various intracellular signaling pathways that mediate addiction, chronic pain, and other diseases, but its intracellular domain structures remain undetermined. The presence of 17 native cysteines in α7 nAChR provides opportunities for extracting structural information through site-directed labeling of chemical probes in strategic locations, but it also creates uncertainties in channel function when those native cysteines must be mutated. Using site-directed mutagenesis and two-electrode voltage clamp electrophysiology measurements, we found that α7 nAChR's function was well tolerated for mutations of all 13 cysteines as long as two pairs of disulfide-bond cysteines remained in the extracellular domain. Furthermore, surface plasmon resonance measurements showed that the cysteine mutations did not affect α7 nAChR binding to the intracellular protein PICK1. The study suggests that a high native cysteine content does not necessarily preclude the use of single cysteine labeling for acquiring structural information on functional proteins.
| S-EPMC7057392 | biostudies-literature
Project description:We investigated assembly and function of nicotinic acetylcholine receptors (nAChRs) composed of α7 and β2 subunits. We measured optical and electrophysiological properties of wild-type and mutant subunits expressed in cell lines and Xenopus laevis oocytes. Laser scanning confocal microscopy indicated that fluorescently tagged α7 and β2 subunits colocalize. Förster resonance energy transfer between fluorescently tagged subunits strongly suggested that α7 and β2 subunits coassemble. Total internal reflection fluorescence microscopy revealed that assemblies localized to filopodia-like processes of SH-EP1 cells. Gain-of-function α7 and β2 subunits confirmed that these subunits coassemble within functional receptors. Moreover, α7β2 nAChRs composed of wild-type subunits or fluorescently tagged subunits had pharmacological properties similar to those of α7 nAChRs, although amplitudes of α7β2 nAChR-mediated, agonist-evoked currents were generally ~2-fold lower than those for α7 nAChRs. It is noteworthy that α7β2 nAChRs displayed sensitivity to low concentrations of the antagonist dihydro-β-erythroidine that was not observed for α7 nAChRs at comparable concentrations. In addition, cysteine mutants revealed that the α7-β2 subunit interface does not bind ligand in a functionally productive manner, partly explaining lower α7β2 nAChR current amplitudes and challenges in identifying the function of native α7β2 nAChRs. On the basis of our findings, we have constructed a model predicting receptor function that is based on stoichiometry and position of β2 subunits within the α7β2 nAChRs.
| S-EPMC3263954 | biostudies-literature
Project description:Nicotinic acetylcholine receptors (nAChRs) present as many different subtypes in the nervous and immune systems, muscles and on the cells of other organs. In the immune system, inflammation is regulated via the vagus nerve through the activation of the non-neuronal α7 nAChR subtype, affecting the production of cytokines. The analgesic properties of α7 nAChR-selective compounds are mostly based on the activation of the cholinergic anti-inflammatory pathway. The molecular mechanism of neuropathic pain relief mediated by the inhibition of α9-containing nAChRs is not fully understood yet, but the role of immune factors in this process is becoming evident. To obtain appropriate drugs, a search of selective agonists, antagonists and modulators of α7- and α9-containing nAChRs is underway. The naturally occurring three-finger snake α-neurotoxins and mammalian Ly6/uPAR proteins, as well as neurotoxic peptides α-conotoxins, are not only sophisticated tools in research on nAChRs but are also considered as potential medicines. In particular, the inhibition of the α9-containing nAChRs by α-conotoxins may be a pathway to alleviate neuropathic pain. nAChRs are involved in the inflammation processes during AIDS and other viral infections; thus they can also be means used in drug design. In this review, we discuss the role of α7- and α9-containing nAChRs in the immune processes and in pain.
| S-EPMC10095066 | biostudies-literature