Nicotine increases GABAergic input on rat dorsal raphe serotonergic neurons through alpha7 nicotinic acetylcholine receptor

2014 ◽  
Vol 112 (12) ◽  
pp. 3154-3163 ◽  
Author(s):  
F. Hernández-Vázquez ◽  
K. Chavarría ◽  
J. Garduño ◽  
S. Hernández-López ◽  
S. P. Mihailescu
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Discharge Rate ◽  
Cell Voltage ◽  
Dorsal Raphe ◽  
Patch Clamp Technique ◽  
Nicotinic Acetylcholine ◽  
Alpha7 Nicotinic Acetylcholine Receptor ◽  
Α7 Nachr ◽  
Dorsal Raphé

The dorsal raphe nucleus (DRN) contains large populations of serotonergic (5-HT) neurons. This nucleus receives GABAergic inhibitory afferents from many brain areas and from DRN interneurons. Both GABAergic and 5-HT DRN neurons express functional nicotinic acetylcholine receptors (nAChRs). Previous studies have demonstrated that nicotine increases 5-HT release and 5-HT DRN neuron discharge rate by stimulating postsynaptic nAChRs and by increasing glutamate and norepinephrine release inside DRN. However, the influence of nicotine on the GABAergic input to 5-HT DRN neurons was poorly investigated. Therefore, the aim of this work was to determine the effect of nicotine on GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) of 5-HT DRN neurons and the subtype of nAChR(s) involved in this response. Experiments were performed in coronal slices obtained from young Wistar rats. GABAergic sIPSCs were recorded from post hoc-identified 5-HT DRN neurons with the whole cell voltage patch-clamp technique. Administration of nicotine (1 μM) increased sIPSC frequency in 72% of identified 5-HT DRN neurons. This effect was not reproduced by the α4β2 nAChR agonist RJR-2403 and was not influenced by TTX (1 μM). It was mimicked by the selective agonist for α7 nAChR, PNU-282987, and exacerbated by the positive allosteric modulator of the same receptor, PNU-120596. The nicotine-induced increase in sIPSC frequency was independent on voltage-gated calcium channels and dependent on Ca2+-induced Ca2+ release (CICR). These results demonstrate that nicotine increases the GABAergic input to most 5-HT DRN neurons, by activating α7 nAChRs and producing CICR in DRN GABAergic terminals.

Serotoninergic dorsal raphe neurons possess functional postsynaptic nicotinic acetylcholine receptors

Synapse ◽  
2008 ◽  
Vol 62 (8) ◽  
pp. 601-615 ◽  
Author(s):  
Luis Galindo-Charles ◽  
Salvador Hernandez-Lopez ◽  
Elvira Galarraga ◽  
Dagoberto Tapia ◽  
José Bargas ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Dorsal Raphe ◽  
Nicotinic Acetylcholine ◽  
Dorsal Raphé ◽  
Raphe Neurons

Expression of functional nicotinic acetylcholine receptors in neuroepithelial bodies of neonatal hamster lung

2003 ◽  
Vol 285 (6) ◽  
pp. L1203-L1212 ◽  
Author(s):  
Xiao Wen Fu ◽  
Colin A. Nurse ◽  
Suzanne M. Farragher ◽  
Ernest Cutz
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Respiratory Control ◽  
Hill Coefficient ◽  
Neuroepithelial Bodies ◽  
Patch Clamp Technique ◽  
Nicotinic Acetylcholine ◽  
Whole Cell Patch Clamp ◽  
Inward Currents ◽  
Double Label

Pulmonary neuroepithelial bodies (NEB) are presumed airway chemoreceptors involved in respiratory control, especially in the neonate. Nicotine is known to affect both lung development and control of breathing. We report expression of functional nicotinic acetylcholine receptors (nAChR) in NEB cells of neonatal hamster lung using a combination of morphological and electrophysiological techniques. Nonisotopic in situ hybridization method was used to localize mRNA for the β2-subunit of nAChR in NEB cells. Double-label immunofluorescence confirmed expression of α4-, α7-, and β2-subunits of nAChR in NEB cells. The electrophysiological characteristics of nAChR in NEB cells were studied using the whole cell patch-clamp technique on fresh lung slices. Application of nicotine (∼0.1-100 μM) evoked inward currents that were concentration dependent (EC50 = 3.8 μM; Hill coefficient = 1.1). ACh (100 μM) and nicotine (50 μM) produced two types of currents. In most NEB cells, nicotine-induced currents had a single desensitizing component that was blocked by mecamylamine (50 μM) and dihydro-β-erythroidine (50 μM). In some NEB cells, nicotine-induced current had two components, with fast- and slow-desensitizing kinetics. The fast component was selectively blocked by methyllcaconitine (MLA, 10 nM), whereas both components were inhibited by mecamylamine. Choline (0.5 mM) also induced an inward current that was abolished by 10 nM MLA. These studies suggest that NEB cells in neonatal hamster lung express functional heteromeric α3β2, α4β2, and α7 nAChR and that cholinergic mechanisms could modulate NEB chemoreceptor function under normal and pathological conditions.

scholarly journals α-Conotoxin as Potential to α7-nAChR Recombinant Expressed in Escherichia coli

Marine Drugs ◽  
2020 ◽  
Vol 18 (8) ◽  
pp. 422
Author(s):  
Yanli Liu ◽  
Yifeng Yin ◽  
Yunyang Song ◽  
Kang Wang ◽  
Fanghui Wu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
High Performance ◽  
Cognitive Disorders ◽  
Coli Strain ◽  
Xenopus Laevis Oocytes ◽  
Obvious Effect ◽  
Nicotinic Acetylcholine ◽  
Α7 Nachr

α7 nicotinic acetylcholine receptors (nAChR) is an important nicotinic acetylcholine receptors subtype and closely associated with cognitive disorders, such as Alzheimer’s and schizophrenia disease. The mutant ArIB (V11L, V16A) of α-conotoxin ArIB with 17-amino acid residues specifically targets α7 nAChR with no obvious effect on other nAChR subtypes. In the study, the synthetic gene encoding mature peptide of ArIB and mutant ArIB (V11L, V16A) carried a fusion protein Trx and 6 × His-tag was separately inserted in pET-32a (+) vector and transformed into Escherichia coli strain BL21(DE3) pLysS for expression. The expressions of Trx-ArIB-His6 and Trx-ArIB (V11L, V16A)-His6 were soluble in Escherichia coli, which were purified by Ni-NTA affinity chromatography column and cleaved by enterokinase to release rArIB and rArIB (V11L, V16A). Then, rArIB and rArIB (V11L, V16A) were purified by high-performance liquid chromatography (HPLC) and identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Bioactivity of rArIB and rArIB (V11L, V16A) was assessed by two-electrode voltage-clamp electrophysiology in Xenopus laevis oocytes expressing human nAChR subtypes. The results indicated that the yield of the fusion proteins was approximately 50 mg/L and rArIB (V11L, V16A) antagonized the α7 nAChR subtype selectively with 8-nM IC50. In summary, this study provides an efficient method to biosynthesize α-conotoxin ArIB and rArIB (V11L, V16A) in Escherichia coli, which could be economical to obtain massively bioactive disulfide-rich polypeptides at fast speed.

Molecular cloning and characterization of a novel human variant of RIC-3, a putative chaperone of nicotinic acetylcholine receptors

2008 ◽  
Vol 28 (6) ◽  
pp. 299-306 ◽  
Author(s):  
Tamara Seredenina ◽  
Teresa Ferraro ◽  
Georg C. Terstappen ◽  
Andrea Caricasole ◽  
Renza Roncarati
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Coiled Coil ◽  
Surface Expression ◽  
Protein Product ◽  
Nicotinic Acetylcholine ◽  
Α7 Nachr ◽  
New Variant ◽  
Novel Variant

Recent reports demonstrate that the RIC-3 (resistant to inhibitors of cholinesterase-3) protein is important for the maturation of nAChRs (nicotinic acetylcholine receptors). In the present study RIC-3e, a novel variant of RIC-3, is described. This variant contains a deletion of exons 4 and 5 of RIC-3, resulting in a protein product lacking a conserved coiled-coil domain. Like RIC-3, the new variant is predominantly, but not exclusively, expressed in the brain. The analysis of expression of variant RIC-3 mRNA and of α7-nAChR mRNA in a set of human tissues shows a similar profile. The RIC-3e protein is functionally active and enables surface expression of mature α7-nAChRs in cell lines not otherwise permissive for the expression of this receptor.

scholarly journals Nicotinic acetylcholine receptor partial antagonist polyamides from tunicates and their predatory sea slugs

2021 ◽  
Author(s):  
Noemi D. Paguigan ◽  
Jortan O. Tun ◽  
Lee S. Leavitt ◽  
Zhenjian Lin ◽  
Kevin Chase ◽  
...  
Keyword(s):  
Nervous System ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Partial Agonist ◽  
Muscarinic Acetylcholine Receptors ◽  
New Drugs ◽  
Calcium Flux ◽  
Nicotinic Acetylcholine ◽  
Chemical Structures ◽  
Α7 Nachr

In our efforts to discover new drugs to treat pain, we identified molleamines A-E (1-5) as major neuroactive components of the sea slug, Pleurobranchus forskalii and their prey, Didemnum molle tunicates. The chemical structures of molleamines were elucidated by spectroscopy and confirmed by the total synthesis of molleamines A (1) and C (3). Synthetic 3 completely blocked acetylcholine-induced calcium flux in peptidergic nociceptors (PNs) in the somatosensory nervous system. Compound 3 affected neither the α7 nAChR nor the muscarinic acetylcholine receptors in calcium flux assays. In addition to nociceptors, 3 partially blocked the acetylcholine-induced calcium flux in the sympathetic nervous system, including neurons from the superior cervical ganglion. Electrophysiology revealed a block of α3β4 (mouse) and α6/α3β4 (rat) nicotinic acetylcholine receptors (nAChRs), with IC50 values of 1.4 and 3.1 μM, respectively. Molleamine C (3) is a partial antagonist, reaching a maximum block of 76-82% of the acetylcholine signal and showing no partial agonist response. Molleamine C (3) may thus provide a lead compound for the development of neuroactive compounds with unique biological properties.

scholarly journals Migration of gastric cancer is suppressed by recombinant Newcastle disease virus (rL-RVG) via regulating α7-nicotinic acetylcholine receptors/ERK- EMT

BMC Cancer ◽  
2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Xuefeng Bu ◽  
Anwei Zhang ◽  
Zhengwei Chen ◽  
Xuanfeng Zhang ◽  
Riting Zhang ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Cell Migration ◽  
Cancer Cells ◽  
Nicotinic Acetylcholine Receptors ◽  
Newcastle Disease ◽  
Acetylcholine Receptors ◽  
Gastric Cancer Cells ◽  
Nicotinic Acetylcholine ◽  
Mesenchymal Transition ◽  
Α7 Nachr

Abstract Background Nicotinic acetylcholine receptors (nAChRs) have been reported to be overexpressed in malignancies in humans and is associated with tumorigenesis and cell migration. In previous studies of gastric cancer, alpha7 nicotinic acetylcholine receptor (α7-nAChR) overexpression leads to epithelial-mesenchymal transition (EMT) and promotes the migration of gastric cancer cells. Recombinant avirulent LaSota strain of Newcastle disease virus (NDV) expressing the rabies virus glycoprotein (rL-RVG) may promote apoptosis of gastric cancer cells and reduces the migration of lung cancer metastasis. However, whether rL-RVG inhibits migration of gastric cancer cells and what the underlying functional mechanism is remains unknown. Methods The gastric cancer cell lines BGC and SGC were randomly divided into 3 groups: rL-RVG, NDV and Phosphate Buffered Solution (PBS) control groups. Furthermore,we adopted ACB and MLA,α7nAChR-siRNA for the overexpression and silencing of α7-nAChR.Corynoxenine was used for inhibiting the MEK-ERK pathway. Western blot, Immunofluoresce,cell proliferation assays,cell migration analyses through wound-healing assays and Transwell assays were used to explore the underlying mechanisms. A mouse xenograft model was used to investigate the effects of rL-RVG,NDV on tumor growth. Results In this study, our findings demonstrate that rL-RVG suppressed the migration of gastric cancer cells and reduced EMT via α7-nAChR in vitro. Furthermore rL-RVG decreased the phosphorylation levels of the MEK/ERK signaling pathway such as down-regulating the expression of P-MEK and P-ERK. Additionally, rL-RVG also reduced the expression level of mesenchymal markers N-cadherin and Vimentin and enhanced the expression of the epithelial marker E-cadherin. Lastly, rL-RVG inhibited nicotinic acetylcholine receptors (nAChRs) to suppress cell migration and epithelial to mesenchymal transition (EMT) in gastric cell. We also found that rL-RVG suppresses the growth of gastric cancer subcutaneous tumor cells in vivo. Conclusion rL-RVG inhibits α7-nAChR-MEK/ERK-EMT to suppress migration of gastric cancer cells.

Stereospecific Interaction of Ketamine with Nicotinic Acetylcholine Receptors in Human Sympathetic Ganglion-like SH-SY5Y Cells

2000 ◽  
Vol 93 (3) ◽  
pp. 818-824 ◽  
Author(s):  
Patrick Friederich ◽  
André Dybek ◽  
Bernd W. Urban
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Maximal Effect ◽  
Patch Clamp Technique ◽  
Nicotinic Acetylcholine ◽  
K Channels ◽  
Channel Inhibition ◽  
Voltage Dependent ◽  
Racemic Ketamine ◽  
Ganglionic Transmission

Background Surprising clinical evidence suggests a block of sympathetic transmission by ketamine. The action of ketamine on nicotinic acetylcholine receptors (nAChRs) in human ganglions is unknown. Because ganglionic transmission depends on nAChRs, such information may help to clarify whether ketamine impairs ganglionic transmission in men. Because racemic ketamine as well as S(+)-ketamine are used clinically, the authors investigated stereospecific effects on human ganglionic nAChRs. Stereospecific psychomimetic effects have been attributed to voltage-dependent Kv channel inhibition; therefore the effects on nAChRs were compared with those on Kv channels present in the same cells. Methods Whole-cell currents through nAChRs and K channels were measured in SH-SY5Y cells with the patch-clamp technique by application of acetylcholine (1 mm, nAChRs) or by a step depolarization from a holding potential of -80 mV to +40 mV (K channels). Electrolyte conditions were identical for both currents. Results Racemic ketamine and the isomers inhibited nAChRs and K channels in a concentration-dependent and reversible manner. Racemic ketamine inhibited nAChRs and K channels, with the anesthetic concentration inducing the half-maximal effect being 1.4 and 300 micrometer, respectively. Only inhibition of the nAChRs was stereoselective. The half-maximal concentrations were 0.8 and 3.6 micrometer for S(+)- and R(-)-ketamine. The K channels were 350 and 70 times less sensitive to the effects of S(+)- and R(-)-ketamine. Conclusion Ketamine at concentrations found during clinical anesthesia exerts stereospecific effects on human ganglionic nAChRs but not on voltage-dependent K channels. Our results support the view that ketamine impairs sympathetic ganglionic transmission. Nonspecific effects on voltage-dependent K channels may underlie psychomimetic side effects.

scholarly journals Effects of C-Terminal Carboxylation on α-Conotoxin LsIA Interactions with Human α7 Nicotinic Acetylcholine Receptor: Molecular Simulation Studies

Marine Drugs ◽  
2019 ◽  
Vol 17 (4) ◽  
pp. 206 ◽  
Author(s):  
Jierong Wen ◽  
Andrew Hung
Keyword(s):  
Conformational Changes ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Salt Bridge ◽  
Md Simulations ◽  
Α7 Nicotinic Acetylcholine Receptor ◽  
Nicotinic Acetylcholine ◽  
Α7 Nachr ◽  
C Terminus ◽  
Toxin Receptor

α-Conotoxins selectively bind to nicotinic acetylcholine receptors (nAChRs), which are therapeutic targets due to their important role in signaling transmission in excitable cells. A previous experimental study has demonstrated that carboxylation of the C-terminal of α-conotoxin LsIA reduces its potency to inhibit human α7 nAChR relative to naturally amidated LsIA. However, little is known about the contribution of conformational changes in the receptor and interactions, induced by C-terminal amidation/carboxylation of conotoxins, to selective binding to nAChRs, since most conotoxins and some disulfide-rich peptides from other conotoxin subfamilies possess a naturally amidated C-terminal. In this study, we employ homology modeling and molecular dynamics (MD) simulations to propose the determinants for differential interactions between amidated and carboxylated LsIAs with α7 nAChR. Our findings indicate an overall increased number of contacts favored by binding of amidated LsIA versus its carboxylated counterpart. Toxin-receptor pairwise interactions, which may play a role in enhancing the potency of the former, include ARG10-TRP77, LEU141 and CYS17-GLN79 via persistent hydrogen bonds and cation-π interactions, which are weakened in the carboxylated form due to a strong intramolecular salt-bridge formed by ARG10 and carboxylated C-terminus. The binding of amidated LsIA also induces enhanced movements in loop C and the juxtamembrane Cys-loop that are closely associated with receptor function. Additionally, the impacts of binding of LsIA on the overall structure and inter-subunit contacts were examined using inter-residue network analysis, suggesting a clockwise tilting of the α7 C and F loops upon binding to carboxylated LsIA, which is absent for amidated LsIA binding. The predicted molecular mechanism of LsIA binding to the α7 receptor may provide new insights into the important role of the C-terminal in the binding potency of conotoxins at neuronal nAChRs for pharmacological purposes.

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