Α4β2 Nicotinic Receptors Research Articles

The MA Helix Is Important for Receptor Assembly and Function in the α4β2 nACh Receptor.

Pentameric ligand-gated ion channels (pLGICs) are expressed throughout the central and peripheral nervous systems of vertebrates and modulate many aspects of human health and disease. Recent structural and computational data indicate that cation-selective pLGICs contain a long helical extension (MA) of one of the transmembrane helices. The MA helix has been shown to affect both the membrane expression of, and ion conductance levels through, these pLGICs. Here we probe the functional effects of 68 mutations in the MA region of the α4β2 nicotinic acetylcholine receptor (nAChR), using a voltage-sensitive membrane dye and radioligand binding to measure receptor function and expression/assembly. We found seven alanine mutations in a stretch of the MA helix that prevent correct receptor folding and/or assembly, as evidenced by the lack of both function and ligand binding. A further two alanine mutations resulted in receptors that were capable of binding ligand but showed no functional response, and we propose that, in these mutants, ligand binding is insufficient to trigger channel opening. The data clarify the effect of the MA helix, and as the effects of some of our mutations in the α4β2 nAChR differ from the effects of equivalent mutations in other cation-selective pLGICs, we suggest that residues in the MA helix may play subtly different roles in different receptors.

A simplified protocol for the automated production of 2-[18 F]fluoro-3-[2-((S)-3-pyrrolinyl)methoxy]pyridine ([18 F]nifene) on an IBA Synthera® module.

The α4β2 nicotinic acetylcholine receptor (nAChR) ligand 2-[18 F]fluoro-3-[2-((S)-3-pyrrolinyl)methoxy]pyridine ([18 F]nifene) has been synthesized in 10% decay-corrected radiochemical yield using the IBA Synthera® platform (IBA Cyclotron Solutions, Louvain-la-Neuve, Belgium) with an integrated fluidic processor (IFP). Boc-nitronifene served as the precursor, and 20% trifluoroacetic acid (TFA) was used to deprotect the Boc-group after radiolabeling. By omitting the solvent extraction step after radiolabeling, the process was simplified to a single step with no manual intervention. [18 F]Nifene was obtained in decay-corrected radiochemical yields of 10 ± 2% (n = 20) and radiochemical purity >99%. Typical specific radioactivities of 2700-4865 mCi/μmole (100-180 GBq/μmol) were measured at the end of synthesis; total synthesis times were about 1h 40 min.

Differential tyrosine and serine/threonine phosphorylation/dephosphorylation pathways regulate the expression of α7 versus α3β4 nicotinic receptor subtypes in mouse hippocampal neurons

We have recently reported that α7 and α3β4 nicotinic acetylcholine receptor (nAChR) subtypes are expressed in human chromaffin cells in the plasma membrane where they colocalize and physically interact. The present study was designed to evaluate whether those receptor subtypes also colocalize at the central nervous system to mutually interact, and whether their expression and colocalization are regulated by phosphorylation/dephosphorylation processes, as they are in human chromaffin cells. We have here found that in isolated and maintained in culture mouse hippocampal neurons, nAChR expression and colocalization of α7, but not α3β4, nAChR subtypes decreased by tyrosine (Tyr)- and serine/threonine (Ser/Thr)-phosphatase inhibition. However, Tyr-kinase inhibition or protein-phosphatase 2A (PP2A) activation increased α3β4 nAChR expression, diminishing receptor subtypes colocalization. Furthermore, colocalization is not recovered if the inhibitors of Tyr-phosphatase and kinases, or the inhibitor of Ser/Thr-phosphatases and the activator of PP2A are applied together. Therefore, regulation of α7 and α3β4 nAChR subtypes expression by Tyr- and Ser/Thr kinases and phosphatases exhibit differential mechanisms in mouse hippocampal neurons. Colocalization of nAChR subtypes, however, is altered by any maneuver that affects these kinases or phosphatases, which might have consequences in the functional activity of nAChR subtypes.

Nicotine-mediated effects in neuronal and mouse models of synucleinopathy.

Alpha-synuclein (α-Syn) aggregation, transmission, and contribution to neurotoxicity represent central mechanisms underlying Parkinson's disease. The plant alkaloid "nicotine" was reported to attenuate α-Syn aggregation in different models, but its precise mode of action remains unclear. In this study, we investigated the effect of 2-week chronic nicotine treatment on α-Syn aggregation, neuroinflammation, neurodegeneration, and motor deficits in D-line α-Syn transgenic mice. We also established a novel humanized neuronal model of α-Syn aggregation and toxicity based on treatment of dopaminergic neurons derived from human induced pluripotent stem cells (iPSC) with α-Syn preformed fibrils (PFF) and applied this model to investigate the effects of nicotine and other compounds and their modes of action. Overall, our results showed that nicotine attenuated α-Syn-provoked neuropathology in both models. Moreover, when investigating the role of nicotinic acetylcholine receptor (nAChR) signaling in nicotine's neuroprotective effects in iPSC-derived dopaminergic neurons, we observed that while α4-specific antagonists reduced the nicotine-induced calcium response, α4 agonists (e.g., AZD1446 and anatabine) mediated similar neuroprotective responses against α-Syn PFF-provoked neurodegeneration. Our results show that nicotine attenuates α-Syn-provoked neuropathology in vivo and in a humanized neuronal model of synucleinopathy and that activation of α4β2 nicotinic receptors might mediate these neuroprotective effects.

Varenicline enhances recognition memory via α7 nicotinic acetylcholine receptors in the medial prefrontal cortex in male mice

Previous studies postulated that chronic administration of varenicline, a partial and full agonist at α4β2 and α7 nicotinic acetylcholine receptors (nAChRs), respectively, enhances recognition memory. However, whether its acute administration is effective, on which brain region(s) it acts, and in what signaling it is involved, remain unknown. To address these issues, we conducted a novel object recognition test using male C57BL/6J mice, focusing on the medial prefrontal cortex (mPFC), a brain region associated with nicotine-induced enhancement of recognition memory. Systemic administration of varenicline before the training dose-dependently enhanced recognition memory. Intra-mPFC varenicline infusion also enhanced recognition memory, and this enhancement was blocked by intra-mPFC co-infusion of a selective α7, but not α4β2, nAChR antagonist. Consistent with this, intra-mPFC infusion of a selective α7 nAChR agonist augmented object recognition memory. Furthermore, intra-mPFC co-infusion of U-73122, a phospholipase C (PLC) inhibitor, or 2-aminoethoxydiphenylborane (2-APB), an inositol trisphosphate (IP3) receptor inhibitor, suppressed the varenicline-induced memory enhancement, suggesting that α7 nAChRs may also act as Gq-coupled metabotropic receptors. Additionally, whole-cell recordings from mPFC layer V pyramidal neurons in vitro revealed that varenicline significantly increased the summation of evoked excitatory postsynaptic potentials, and this effect was suppressed by U-73122 or 2-APB. These findings suggest that varenicline might acutely enhance recognition memory via mPFC α7 nAChR stimulation, followed by mPFC neuronal excitation, which is mediated by the activation of PLC and IP3 receptor signaling. Our study provides evidence supporting the potential repositioning of varenicline as a treatment for cognitive impairment.

Reduction of nicotine and ethanol intake in alcohol-preferring (UChB) female rats by the α4β2 nicotinic acetylcholine receptor partial agonists 5-bromocytisine and cytisine

RationaleNeuronal nicotinic acetylcholine receptors (nAChRs) are implicated in the reinforcing effects of nicotine and ethanol. Previous studies have shown that cytisine and its 5-bromo derivative are partial agonists at the α4β2 nAChRs and that the parent molecule cytisine is effective in reducing both nicotine- and ethanol-self-administration in rats. However, whether 5-bromocytisine affects nicotine or ethanol self-administration was unknown. ObjectivesThe present study compared the effects of 5-bromocytisine and cytisine on nicotine self-administration and further assessed the effect of daily drug injection on voluntary ethanol consumption in alcohol-preferring female rats. Animals were administered a 1.5mg/kg i.p. dose of 5-bromocytisine or cytisine every day for 15–16 days. ResultsThe initial efficacy of 5-bromocytisine and cytisine in reducing nicotine intake was similar (−80%) while for voluntary ethanol intake 5-bromocytisine was a superior inhibitor over cytisine (−78% and −40% respectively). The efficacy of cytisine began to diminish after 10 days of daily administration, which was attributed to tolerance development to its inhibitory effects both on nicotine and ethanol self-administration. Tolerance did not develop for 5-bromocytisine. Conclusion5-Bromocytisine, a weaker α4β2 nAChR partial agonist than cytisine, also produces a sustained inhibition of both nicotine and ethanol self-administration, and unlike cytisine, it does not develop tolerance.

Cytisinicline for Smoking Cessation

Cytisinicline (cytisine) is a plant-based alkaloid that, like varenicline, binds selectively to α4β2 nicotinic acetylcholine receptors, which mediate nicotine dependence. Although not licensed in the US, cytisinicline is used in some European countries to aid smoking cessation, but its traditional dosing regimen and treatment duration may not be optimal. To evaluate the efficacy and tolerability of cytisinicline for smoking cessation when administered in a novel pharmacokinetically based dosing regimen for 6 or 12 weeks vs placebo. A 3-group, double-blind, placebo-controlled, randomized trial (ORCA-2) compared 2 durations of cytisinicline treatment (6 or 12 weeks) vs placebo, with follow-up to 24 weeks, among 810 adults who smoked cigarettes daily and wanted to quit. It was conducted at 17 US sites from October 2020 to December 2021. Participants were randomized (1:1:1) to cytisinicline, 3 mg, 3 times daily for 12 weeks (n = 270); cytisinicline, 3 mg, 3 times daily for 6 weeks then placebo 3 times daily for 6 weeks (n = 269); or placebo 3 times daily for 12 weeks (n = 271). All participants received behavioral support. Biochemically verified continuous smoking abstinence for the last 4 weeks of cytisinicline treatment vs placebo (primary) and from end of treatment to 24 weeks (secondary). Of 810 randomized participants (mean age, 52.5 years; 54.6% female; mean of 19.4 cigarettes smoked daily), 618 (76.3%) completed the trial. For the 6-week course of cytisinicline vs placebo, continuous abstinence rates were 25.3% vs 4.4% during weeks 3 to 6 (odds ratio [OR], 8.0 [95% CI, 3.9-16.3]; P < .001) and 8.9% vs 2.6% during weeks 3 to 24 (OR, 3.7 [95% CI, 1.5-10.2]; P = .002). For the 12-week course of cytisinicline vs placebo, continuous abstinence rates were 32.6% vs 7.0% for weeks 9 to 12 (OR, 6.3 [95% CI, 3.7-11.6]; P < .001) and 21.1% vs 4.8% during weeks 9 to 24 (OR, 5.3 [95% CI, 2.8-11.1]; P < .001). Nausea, abnormal dreams, and insomnia occurred in less than 10% of each group. Sixteen participants (2.9%) discontinued cytisinicline due to an adverse event. No drug-related serious adverse events occurred. Both 6- and 12-week cytisinicline schedules, with behavioral support, demonstrated smoking cessation efficacy and excellent tolerability, offering new nicotine dependence treatment options. ClinicalTrials.gov Identifier: NCT04576949.

Effects of varenicline on the serum levels of olanzapine in male patients with Schizophrenia: a randomized controlled trial.

Smoking in patients with Schizophrenia is more common than in the general population. Varenicline, a partial agonist at α4β2 nicotinic acetylcholine receptors, is an effective smoking cessation pharmacotherapy in patients with Schizophrenia. However, its effects on the serum levels of antipsychotics in Schizophrenia are understudied. This study investigated the impact of smoking cessation with varenicline on the serum concentration of olanzapine in patients with Schizophrenia. Adult smokers with Schizophrenia were enrolled in a 12-week course of varenicline and placebo for smoking cessation. The serum concentration of olanzapine was measured at baseline and weeks 1, 2, 4, 8, and 12. Data were analyzed with the generalized additive mixed model. During the 12-week study, the results indicated that olanzapine concentrations increased nonlinearly in the varenicline and placebo groups. Threshold effect analysis suggested that the olanzapine concentrations increased over time until the turning point (week 4). However, there was no significant difference between the two treatment groups. Varenicline showed safety and efficacy in smoking cessation in people with Schizophrenia.

Selective α3β4 Nicotinic Acetylcholine Receptor Ligand as a Potential Tracer for Drug Addiction.

α3β4 Nicotinic acetylcholine receptor (nAChR) has been recognized as an emerging biomarker for the early detection of drug addiction. Herein, α3β4 nAChR ligands were designed and synthesized to improve the binding affinity and selectivity of two lead compounds, (S)-QND8 and (S)-T2, for the development of an α3β4 nAChR tracer. The structural modification was achieved by retaining the key features and expanding the molecular structure with a benzyloxy group to increase the lipophilicity for blood-brain barrier penetration and to extend the ligand-receptor interaction. The preserved key features are a fluorine atom for radiotracer development and a p-hydroxyl motif for ligand-receptor binding affinity. Four (R)- and (S)-quinuclidine-triazole (AK1-AK4) were synthesized and the binding affinity, together with selectivity to α3β4 nAChR subtype, were determined by competitive radioligand binding assay using [3H]epibatidine as a radioligand. Among all modified compounds, AK3 showed the highest binding affinity and selectivity to α3β4 nAChR with a Ki value of 3.18 nM, comparable to (S)-QND8 and (S)-T2 and 3069-fold higher affinity to α3β4 nAChR in comparison to α7 nAChR. The α3β4 nAChR selectivity of AK3 was considerably higher than those of (S)-QND8 (11.8-fold) and (S)-T2 (294-fold). AK3 was shown to be a promising α3β4 nAChR tracer for further development as a radiotracer for drug addiction.

Examining the effect of a substituted carbamate positive allosteric modulator on the (α4)3(β2)2 and (α4)2(β2)3 nicotinic acetylcholine receptors.

The α4β2 nicotinic acetylcholine receptors (nAChR) are the most abundant nAChR subtypes in the brain and exist in two distinct isoforms: (α4)2(β2)3 nAChR, the high sensitivity (HS) isoform, and (α4)3(β2)2 nAChR, the low sensitivity (LS) isoform. Given the pathophysiological and pharmacological importance of these brain receptors, there are ongoing efforts to develop pharmacophores that selectively potentiate α4β2 nAChRs (positive allosteric modulators, PAMs). PAMs selective to the α4β2 nAChRs would be beneficial for neuropsychiatric conditions such as nicotine addiction and Alzheimer's disease. A chemical synthesis and high throughput screens at Amgen laboratories have identified series of compounds that preferentially potentiate the α4β2 nAChRs (Albrecht, Berry et al. 2008, Springer, Woodin et al. 2008). Notable PAMs identified in these screens include CMPI, which selectively potentiates the (α4)3(β2)2 nAChR isoform (Wang, Deba et al. 2017). In this study, we started to pharmacologically characterize additional derivatives (C9M) from these screens, focusing on its effects on the LS and HS nAChR isoforms. Unlike CMPI, C9M potentiated current response of Xenopus oocytes expressing LS and HS nAChR isoforms elicited by submaximal and saturating concentrations of ACh. This potentiation indicates that C9M enhances the agonist's efficacy at both α4β2 nAChR isoforms. Furthermore, mutations known to alter CMPI potentiation of LS nAChR (e.g. α4G41M) did not abolish the potentiating effects of C9M. Additionally, C9M enhancement of agonist's efficacy at HS (α4)2(β2)3 nAChR was notably greater than that for a panel of known nAChR PAMs including dFBr and LY2087101 (all co-applied at 1 µM with 10 µM ACh). Experiments to determine the concentration dependent effect of C9M at both LS and HS nAChR isoforms and to determine the extent of C9M effect on the agonist's potency are ongoing.

Kinetic modelling of ligand trapping inside acidic vesicles containing alpha-4 beta-2 nicotinic receptors as an explanation for long-term cessation effect of varenicline (Chantix).

Govind's et. al. 2017 paper proposes that the smoking cessation effect of varenicline (or epibatidine) is caused by these molecules being trapped inside acidic vesicles containing α4β2 nicotinic receptors. These ligands are weak bases with high affinity to these receptors. The trapping would lead to their slower release and longer desensitization of the receptors, thereby reducing nicotine upregulation. Meanwhile, nicotine, with lower pKa and lower binding affinity is not trapped within the vesicles. It is unclear whether the acidity of the vesicles and the presence of receptors is sufficient to explain the biphasic trapping behavior observed empirically. To better understand this phenomenon, we model flux of ligands between cellular compartments using Flick's first law of diffusion and the Nernst-Plank Equation. The ratio of neutral to charged ligands within each compartment is calculated via Henderson-Hasselbach equation. Main parameters in the model include the number of vesicles, receptors per vesicle, ratio of vesicles to non-vesicle receptors, pH, membrane permeability, pKa and dissociation constants of the ligand. The system is solved numerically using MATLAB and optimized against empirical data. The results show the experimentally observed biphasic release of epibatidine, affirming the hypothesis of trapping inside the vesicles. Our model also shows that the main trapping effect is due to the presence of receptors inside vesicles, not those located on the cell surface and the high affinity of these ligands to the receptors. The exact acidity level inside the vesicles is not as important. Our model is also able to predict the much faster nicotine outward flow compared with epibatidine. This model can be used to predict the kinetics of other weak basic ligands like those used in positron emission tomography.

Interaction of Varenicline with Classic Antiseizure Medications in the Mouse Maximal Electroshock-Induced Seizure Model.

Varenicline (VAR) is a partial agonist of brain α4β2 nicotinic acetylcholine receptors recommended as a first line pharmacotherapy for smoking cessation. The aim of this study was to examine whether VAR affects the protective activity of four classic antiseizure medications, i.e., carbamazepine (CBZ), phenobarbital (PB), phenytoin (PHT), and valproate (VPA) on maximal electroshock (MES)-induced seizures, which may serve as an experimental model of human-generalized tonic-clonic seizures in mice. VAR administered intraperitoneally (i.p.) at a subthreshold dose of 0.5 mg/kg decreased the protective activity of CBZ against MES-induced convulsions, increasing its median effective dose (ED50) from 10.92 ± 1.0 to 18.15 ± 1.73 mg/kg (p < 0.01). The effect of VAR was dose-dependent because a lower dose of VAR (0.25 mg/kg) failed to antagonize the protective activity of CBZ. VAR administered at the subthreshold dose of 0.5 mg/kg had no impact on the protective activity of PB, PHT, and VPA in the mouse MES model. The inhibitory effect of VAR on the protective activity of CBZ against tonic-clonic convulsions most likely resulted from the pharmacodynamic mechanism(s) and was not associated with the changes in total brain concentrations of CBZ. VAR-evoked alterations in the anticonvulsive activity of CBZ may be of serious concern for epileptic tobacco smokers.

Rare Missense Variants of the Human β4 Subunit Alter Nicotinic α3β4 Receptor Plasma Membrane Localisation.

α3β4 nicotinic acetylcholine receptors (nARs) are pentameric ligand-gated cation channels that function in peripheral tissue and in the peripheral and central nervous systems, where they are critical mediators of ganglionic synaptic transmission and modulators of reward-related behaviours. In the pentamer, two α3β4 subunit couples provide ligand-binding sites, and the fifth single (accessory) subunit (α3 or β4) regulates receptor trafficking from the endoplasmic reticulum to the cell surface. A number of rare missense variants of the human β4 subunit have recently been linked to nicotine dependence and/or sporadic amyotrophic lateral sclerosis, and altered responses to nicotine have been reported for these variants; however, it is unknown whether the effects of mutations depend on the subunit within the ligand-binding couples and/or on the fifth subunit. Here, by expressing single populations of pentameric receptors with fixed stoichiometry in cultured cells, we investigated the effect of β4 variants in the fifth position on the assembly and surface exposure of α3β4 nAChRs. The results demonstrate that the missense mutations in the accessory subunit alone, despite not affecting the assembly of α3β4 receptors, alter their trafficking and surface localisation. Thus, altered trafficking of an otherwise functional nAChR may underlie the pathogenic effects of these mutations.

Nicotine suppresses central post-stroke pain via facilitation of descending noradrenergic neuron through activation of orexinergic neuron

Central post-stroke pain (CPSP) is a type of central neuropathic pain, whose underlying mechanisms remain unknown. We previously reported that bilateral carotid artery occlusion (BCAO)-induced CPSP model mice showed mechanical hypersensitivity and decreased mRNA levels of preproorexin, an orexin precursor, in the hypothalamus. Recently, nicotine was shown to regulate the neuronal activity of orexin in the lateral hypothalamus (LH) and suppress inflammatory and neuropathic pain. In this study, we evaluated whether nicotine could suppress BCAO-induced mechanical allodynia through the activation of orexinergic neurons. Mice were subjected to BCAO for 30 min. Mechanical hypersensitivity was assessed by the von Frey test. BCAO mice showed hypersensitivity to mechanical stimuli three days after BCAO surgery. The intracerebroventricular injection of nicotine suppressed BCAO-induced mechanical hypersensitivity in a dose-dependent manner. These effects were inhibited by α7 or α4β2-nicotinic receptor antagonists. After nicotine injection, the level of c-fos, a neuronal activity marker, increased in the LH and locus coeruleus (LC) of Sham and BCAO mice. Increased number of c-Fos-positive cells partly colocalized with orexin A-positive cells in the LH, as well as tyrosine hydroxylase-positive cells in the LC.Orexinergic neurons project to the LC area. Nicotine-induced antinociception tended to cancel by the pretreatment of SB334867, an orexin receptor1 antagonist into the LC. Intra-LH microinjection of nicotine attenuated BCAO-induced mechanical hypersensitivity. Nicotine-induced antinociception was inhibited by intrathecal pre-treatment with yohimbine, an α2 adrenergic receptor antagonist. These results indicated that nicotine may suppress BCAO-induced mechanical hypersensitivity through the activation of the descending pain control system via orexin neurons.

Detection of Neonicotinoid Insecticides and Their Metabolites in Human Cerebrospinal Fluid.

Detection of Neonicotinoid Insecticides and Their Metabolites in Human Cerebrospinal Fluid.

What We Have Gained from Ibogaine: α3β4 Nicotinic Acetylcholine Receptor Inhibitors as Treatments for Substance Use Disorders.

For decades, ibogaine─the main psychoactive alkaloid found in Tabernanthe iboga─has been investigated as a possible treatment for substance use disorders (SUDs) due to its purported ability to interrupt the addictive properties of multiple drugs of abuse. Of the numerous pharmacological actions of ibogaine and its derivatives, the inhibition of α3β4 nicotinic acetylcholine receptors (nAChRs), represents a probable mechanism of action for their apparent anti-addictive activity. In this Perspective, we examine several classes of compounds that have been discovered and developed to target α3β4 nAChRs. Specifically, by focusing on compounds that have proven efficacious in pre-clinical models of drug abuse and have been evaluated clinically, we highlight the promising potential of the α3β4 nAChRs as viable targets to treat a wide array of SUDs. Additionally, we discuss the challenges faced by the existing classes of α3β4 nAChR ligands that must be overcome to develop them into therapeutic treatments.

Pharmacophore Mapping Combined with dbCICA Reveal New Structural Features for the Development of Novel Ligands Targeting α4β2 and α7 Nicotinic Acetylcholine Receptors

The neuronal nicotinic acetylcholine receptors (nAChRs) belong to the ligand-gated ion channel (GLIC) group, presenting a crucial role in several biological processes and neuronal disorders. The α4β2 and α7 nAChRs are the most abundant in the central nervous system (CNS), being involved in challenging diseases such as epilepsy, Alzheimer's disease, schizophrenia, and anxiety disorder, as well as alcohol and nicotine dependencies. In addition, in silico-based strategies may contribute to revealing new insights into drug design and virtual screening to find new drug candidates to treat CNS disorders. In this context, the pharmacophore maps were constructed and validated for the orthosteric sites of α4β2 and α7 nAChRs, through a docking-based Comparative Intermolecular Contacts Analysis (dbCICA). In this sense, bioactive ligands were retrieved from the literature for each receptor. A molecular docking protocol was developed for all ligands in both receptors by using GOLD software, considering GoldScore, ChemScore, ASP, and ChemPLP scoring functions. Output GOLD results were post-processed through dbCICA to identify critical contacts involved in protein-ligand interactions. Moreover, Crossminer software was used to construct a pharmacophoric map based on the most well-behaved ligands and negative contacts from the dbCICA model for each receptor. Both pharmacophore maps were validated by using a ROC curve. The results revealed important features for the ligands, such as the presence of hydrophobic regions, a planar ring, and hydrogen bond donor and acceptor atoms for α4β2. Parallelly, a non-planar ring region was identified for α7. These results can enable fragment-based drug design (FBDD) strategies, such as fragment growing, linking, and merging, allowing an increase in the activity of known fragments. Thus, our results can contribute to a further understanding of structural subunits presenting the potential for key ligand-receptor interactions, favoring the search in molecular databases and the design of novel ligands.

Molecular docking and dynamic simulation studies of α4β2 and α7 nicotinic acetylcholine receptors with tobacco smoke constituents nicotine, NNK and NNN

Smoking constitutes a major global health problem. As it triggers various health hazards including cancers, cardiac and pulmonary illness, it is imperative to understand the mechanism of action of various smoke constituents on our cellular processes. Various in vitro studies have compiled the affinity of cigarette smoke constituents on various nicotinic acetylcholine receptors (nAChRs). But the nature of the intermolecular interactions contributing to this affinity and the key amino acids in the receptor active sites involved in this are not investigated so far. Here, we are examining the interaction of α7nAChR and α4β2nAChR on nicotine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N-nitrosornicotine (NNN), the physiologically significant constituents in smoke, through molecular docking and dynamics simulations study. The docking of α4β2nAChR structure with the ligands nicotine, NNK and NNN yielded docking scores of −41.45 kcal/mol, −59.28 kcal/mol and −54.60 kcal/mol, respectively, and that of α7nAChR receptor molecule with the ligands yielded docking scores of −59.54 kcal/mol, −71.06 kcal/mol and −70.86 kcal/mol, respectively. The study showed that NNK exhibited the highest affinity with the ligands which was confirmed by dynamics simulation. But higher stability of interactions as surmised from Molecular dynamics simulations was found for nicotine with α4β2nAChR and NNN with α7nAChR. The findings validate the in vitro studies comparing the affinities of these compounds. The study will be useful in formulating effective nAChR agonists to treat neurological disorders and antagonists for smoke deaddiction and improve health standards. Communicated by Ramaswamy H. Sarma

Challenge exposure to whole cigarette smoke condensate upregulates locomotor sensitization by stimulating α4β2 nicotinic acetylcholine receptors in the nucleus accumbens of rats

Nicotine, the primary addictive substance in tobacco, produces the psychomotor, rewarding, and reinforcing effects of tobacco dependence by stimulating nicotinic acetylcholine receptors (nAChRs) in the brain. The present study determined that α4β2 nAChRs regulate locomotor sensitization by altering dopamine concentration in the nucleus accumbens (NAc) after systemic challenge exposure to whole cigarette smoke condensate (WCSC). Rats were administered subcutaneous injection of WCSC (0.2 mg/kg nicotine/day) for 7 consecutive days and then re-exposed to WCSC after 3 days of withdrawal. Challenge exposure to WCSC significantly increased locomotor activity. This increase was decreased by the subcutaneous injection of the α4β2 nAChR antagonist, DHβE (3 mg/kg), but not by the intraperitoneal injection of the α7 nAChR antagonist, MLA (5 mg/kg). In parallel with a decrease in locomotor activity, blockade of α4β2 nAChRs with DHβE decreased dopamine concentration in the NAc which was elevated by challenge exposure to WCSC. These findings suggest that challenge WCSC leads to the expression of locomotor sensitization by elevating dopamine concentration via stimulation of α4β2 nAChRs expressed in neurons of the NAc in rats.

Desformylflustrabromine (dFBr), a positive allosteric modulator of α4β2 nicotinic acetylcholine receptors decreases voluntary ethanol consumption and preference in male and female Sprague-Dawley rats.

Alcohol use disorder is a medical condition that impacts millions of individuals worldwide. Although there are a few pharmacotherapeutic options for alcohol-dependent individuals; there is a need for the development of novel and more effective therapeutic approaches. Alcohol and nicotine are commonly co-abused, and there is evidence that neuronal nicotinic acetylcholine receptors (nAChRs) play a role in both alcohol and nicotine dependence. Desformylflustrabromine (dFBr), a positive allosteric modulator of the α4β2 nAChRs has been shown to reduce nicotine intake, compulsive-like behavior and neuropathic pain in animal models. dFBr has also been previously shown to cross the blood-brain-barrier. We have recently shown that dFBr can attenuate the response to an acute, hypnotic dose of ethanol, via β2 nAchR. Here, we have investigated the effect of dFBr in modulating ethanol consumption using the intermittent access two-bottle choice (IA2BC) model of voluntary ethanol consumption in male and female Sprague Dawley rats. We show that dFBr selectively reduced ethanol but not sucrose consumption in the IA2BC model. Furthermore, dFBr decreased preference for ethanol in both male and female rats. No rebound increase in ethanol intake was observed after the washout period after dFBr treatment. The ability of dFBr to decrease ethanol consumption, along with its previously demonstrated ability to decrease nicotine self-administration in rodents, suggest that dFBr is an attractive therapeutic candidate to target both nicotine and alcohol abuse.