scholarly journals Peimine, an Anti-Inflammatory Compound from Chinese Herbal Extracts, Modulates Muscle-Type Nicotinic Receptors

2021 ◽  
Vol 22 (20) ◽  
pp. 11287
Author(s):  
Armando Alberola-Die ◽  
José Antonio Encinar ◽  
Raúl Cobo ◽  
Gregorio Fernández-Ballester ◽  
José Manuel González-Ros ◽  
...  
Keyword(s):  
Molecular Dynamics ◽  
Ion Channels ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Transmembrane Domain ◽  
Muscle Type ◽  
Functional Studies ◽  
Voltage Dependent ◽  
Neuronal Tissues ◽  
Anti Inflammatory

Fritillaria bulbs are used in Traditional Chinese Medicine to treat several illnesses. Peimine (Pm), an anti-inflammatory compound from Fritillaria, is known to inhibit some voltage-dependent ion channels and muscarinic receptors, but its interaction with ligand-gated ion channels remains unexplored. We have studied if Pm affects nicotinic acetylcholine receptors (nAChRs), since they play broad functional roles, both in the nervous system and non-neuronal tissues. Muscle-type nAChRs were incorporated to Xenopus oocytes and the action of Pm on the membrane currents elicited by ACh (IAChs) was assessed. Functional studies were combined with virtual docking and molecular dynamics assays. Co-application of ACh and Pm reversibly blocked IACh, with an IC50 in the low micromolar range. Pm inhibited nAChR by: (i) open-channel blockade, evidenced by the voltage-dependent inhibition of IAch, (ii) enhancement of nAChR desensitization, revealed by both an accelerated IACh decay and a decelerated IACh deactivation, and (iii) resting-nAChR blockade, deduced from the IACh inhibition elicited by Pm when applied before ACh superfusion. In good concordance, virtual docking and molecular dynamics assays demonstrated that Pm binds to different sites at the nAChR, mostly at the transmembrane domain. Thus, Pm from Fritillaria bulbs, considered therapeutic herbs, targets nAChRs with high affinity, which might account for its anti-inflammatory actions.

scholarly journals Variants in CHRNB2 and CHRNA4 Identified in Patients with Insular Epilepsy

2020 ◽  
Vol 47 (6) ◽  
pp. 800-809 ◽  
Author(s):  
Maxime Cadieux-Dion ◽  
Simone Meneghini ◽  
Chiara Villa ◽  
Dènahin Hinnoutondji Toffa ◽  
Ronny Wickstrom ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Functional Characterization ◽  
Reversal Potential ◽  
Missense Variant ◽  
Functional Expression ◽  
Functional Studies ◽  
Sorting Intolerant From Tolerant ◽  
First Case ◽  
And Function

Abstract:Purpose:Our purpose was to determine the role of CHRNA4 and CHRNB2 in insular epilepsy.Method:We identified two patients with drug-resistant predominantly sleep-related hypermotor seizures, one harboring a heterozygous missense variant (c.77C>T; p. Thr26Met) in the CHRNB2 gene and the other a heterozygous missense variant (c.1079G>A; p. Arg360Gln) in the CHRNA4 gene. The patients underwent electrophysiological and neuroimaging studies, and we performed functional characterization of the p. Thr26Met (c.77C>T) in the CHRNB2 gene.Results:We localized the epileptic foci to the left insula in the first case (now seizure-free following epilepsy surgery) and to both insulae in the second case. Based on tools predicting the possible impact of amino acid substitutions on the structure and function of proteins (sorting intolerant from tolerant and PolyPhen-2), variants identified in this report could be deleterious. Functional expression in human cell lines of α4β2 (wild-type), α4β2-Thr26Met (homozygote), and α4β2/β2-Thr26Met (heterozygote) nicotinic acetylcholine receptors revealed that the mutant subunit led to significantly higher whole-cell nicotinic currents. This feature was observed in both homo- and heterozygous conditions and was not accompanied by major alterations of the current reversal potential or the shape of the concentration-response relation.Conclusions:This study suggests that variants in CHRNB2 and CHRNA4, initially linked to autosomal dominant nocturnal frontal lobe epilepsy, are also found in patients with predominantly sleep-related insular epilepsy. Although the reported variants should be considered of unknown clinical significance for the moment, identification of additional similar cases and further functional studies could eventually strengthen this association.

scholarly journals Mechanism of Tacrine Block at Adult Human Muscle Nicotinic Acetylcholine Receptors

2002 ◽  
Vol 120 (3) ◽  
pp. 369-393 ◽  
Author(s):  
Richard J. Prince ◽  
Richard A. Pennington ◽  
Steven M. Sine
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Binding Sites ◽  
Acetylcholine Receptors ◽  
Open Channel ◽  
Single Channel ◽  
Dependent Manner ◽  
Open Probability ◽  
Sequential Model ◽  
Hek 293 ◽  
Voltage Dependent

We used single-channel kinetic analysis to study the inhibitory effects of tacrine on human adult nicotinic receptors (nAChRs) transiently expressed in HEK 293 cells. Single channel recording from cell-attached patches revealed concentration- and voltage-dependent decreases in mean channel open probability produced by tacrine (IC50 4.6 μM at −70 mV, 1.6 μM at −150 mV). Two main effects of tacrine were apparent in the open- and closed-time distributions. First, the mean channel open time decreased with increasing tacrine concentration in a voltage-dependent manner, strongly suggesting that tacrine acts as an open-channel blocker. Second, tacrine produced a new class of closings whose duration increased with increasing tacrine concentration. Concentration dependence of closed-times is not predicted by sequential models of channel block, suggesting that tacrine blocks the nAChR by an unusual mechanism. To probe tacrine's mechanism of action we fitted a series of kinetic models to our data using maximum likelihood techniques. Models incorporating two tacrine binding sites in the open receptor channel gave dramatically improved fits to our data compared with the classic sequential model, which contains one site. Improved fits relative to the sequential model were also obtained with schemes incorporating a binding site in the closed channel, but only if it is assumed that the channel cannot gate with tacrine bound. Overall, the best description of our data was obtained with a model that combined two binding sites in the open channel with a single site in the closed state of the receptor.

scholarly journals Role of the Cys Loop and Transmembrane Domain in the Allosteric Modulation of α4β2 Nicotinic Acetylcholine Receptors

2016 ◽  
Vol 292 (2) ◽  
pp. 551-562 ◽  
Author(s):  
Constanza Alcaino ◽  
Maria Musgaard ◽  
Teresa Minguez ◽  
Simone Mazzaferro ◽  
Manuel Faundez ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Transmembrane Domain ◽  
Allosteric Modulation ◽  
Nicotinic Acetylcholine

scholarly journals Identification of Crucial Residues in α-Conotoxin EI Inhibiting Muscle Nicotinic Acetylcholine Receptor

Toxins ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 603 ◽  
Author(s):  
Jiong Ning ◽  
Jie Ren ◽  
Yang Xiong ◽  
Yong Wu ◽  
Manqi Zhangsun ◽  
...  
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Fold Increase ◽  
Cd Spectroscopy ◽  
Xenopus Laevis Oocytes ◽  
Muscle Type ◽  
Nicotinic Acetylcholine ◽  
Activity Loss ◽  
Spectroscopy Studies ◽  
Key Residues

α-Conotoxins (α-CTxs) are small disulfide-rich peptides from venom of Conus species that target nicotinic acetylcholine receptors (nAChRs). The muscle-type nAChRs have been recognized as a potential target for several diseases, such as myogenic disorders, muscle dystrophies, and myasthenia gravis. EI, an α4/7-CTx, mainly blocks α1β1δε nAChRs and has an extra N-terminal extension of three amino acids. In this study, the alanine scanning (Ala-scan) mutagenesis was applied in order to identify key residues of EI for binding with mouse α1β1δε nAChR. The Ala-substituted analogues were tested for their abilities of modulating muscle and neuronal nAChRs in Xenopus laevis oocytes using two-electrode voltage clamp (TEVC) recordings. Electrophysiological results indicated that the vital residues for functional activity of EI were His-7, Pro-8, Met-12, and Pro-15. These changes exhibited a significant decrease in potency of EI against mouse α1β1δε nAChR. Interestingly, replacing the critical serine (Ser) at position 13 with an alanine (Ala) residue resulted in a 2-fold increase in potency at the α1β1δε nAChR, and showed loss of activity on α3β2 and α3β4 nAChRs. Selectivity and potency of [S13A] EI was improved compared with wild-type EI (WT EI). In addition, the structure–activity relationship (SAR) of EI revealed that the “Arg1–Asn2–Hyp3” residues at the N-terminus conferred potency at the muscle-type nAChRs, and the deletion analogue △1–3 EI caused a total loss of activity at the α1β1δε nAChR. Circular dichroism (CD) spectroscopy studies demonstrated that activity loss of truncated analogue △1–3 EI for α1β1δε nAChR is attributed to disturbance of the secondary structure. In this report, an Ala-scan mutagenesis strategy is presented to identify crucial residues that are significantly affecting potency of E1 for mouse α1β1δε nAChR. It may also be important in remodeling of some novel ligands for inhibiting muscle-type nAChRs.

scholarly journals Activation of Nicotinic Acetylcholine Receptors Augments Calcium Channel-mediated Exocytosis in Rat Pheochromocytoma (PC12) Cells

1998 ◽  
Vol 111 (2) ◽  
pp. 257-269 ◽  
Author(s):  
Amy B. Harkins ◽  
Aaron P. Fox
Keyword(s):  
Pc12 Cells ◽  
Nicotinic Acetylcholine Receptors ◽  
Acetylcholine Receptors ◽  
Single Cells ◽  
Cellular Functions ◽  
Nicotinic Acetylcholine ◽  
Cellular Processes ◽  
Voltage Dependent ◽  
Vesicle Secretion ◽  
Ca2 Channels

The functional effect of activating Ca2+-permeable neuronal nicotinic acetylcholine receptors (nAChRs) on vesicle secretion was studied in PC12 cells. Single cells were patch-clamped in the whole-cell configuration and stimulated with either brief pulses of nicotine to activate the Ca2+-permeable nAChRs or with voltage steps to activate voltage-dependent Ca2+ channels. Membrane capacitance was used as a measure of vesicle secretion. Activation of nAChRs by nicotine application to cells voltage clamped at −80 mV evoked secretion. This secretion was completely abolished by nicotinic antagonists. When the cells were voltage clamped at +20 mV in the presence of Cd2+ to block voltage-activated Ca2+ channels, nicotine elicited a small amount of secretion. Most interestingly, when the nAChRs were activated coincidentally with voltage-dependent Ca2+ channels, secretion was augmented approximately twofold over the secretion elicited with voltage-dependent Ca2+ channels alone. Our data suggest that Ca2+ influx via nAChRs affects Ca2+-dependent cellular functions, including vesicle secretion. In addition to the secretion evoked by nAChR activation at hyperpolarized potentials, we demonstrate that even at depolarized potentials, nAChRs provide an important Ca2+ entry pathway underlying Ca2+-dependent cellular processes such as exocytosis.

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