Inhibition Of Acetylcholine Receptors Research Articles

Structure-Activity Relationships of Alanine Scan Mutants αO-Conotoxins GeXIVA[1,2] and GeXIVA[1,4

αO-Conotoxin GeXIVA is a selective α9α10 nicotinic acetylcholine receptor (nAChR) inhibitor displaying two disulfide bonds that can form three isomers. The bead (GeXIVA[1,2]) and ribbon (GeXIVA[1,4]) isomers possess the highest activity on rat and human α9α10 nAChRs. However, the molecular mechanism by which they inhibit the α9α10 nAChR is unknown. Here, an alanine scan of GeXIVA was used to elucidate key interactions between the peptides and the α9α10 nAChR. The majority of GeXIVA[1,2] analogues preserved affinity at α9α10 nAChR, but [R17A]GeXIVA[1,2] enhanced selectivity on the α9α10 nAChR. The I23A replacement of GeXIVA[1,4] increased activity at both rat and human α9α10 nAChRs by 10-fold. Surprisingly, these results do not support the molecular model of an interaction in the orthosteric binding site proposed previously, but rather may involve allosteric coupling with the voltage-sensitive domain of the α9α10 nAChR. These results could help to guide further development of GeXIVA analogues as analgesics.

Alzheimer’s Disease: Treatment Today and Tomorrow

Abstract Background and Aims: The scope of treatment in Alzheimer’s Disease has widened in recent times with FDA approval of new drugs. This review looks at established treatments in AD as well as critically analyses the newer drugs available. Methods: Data in this review was gathered from PubMed; Google Scholar and MEDLINE from January-March 2023. Search words used were ‘Alzheimer’s Disease treatment’ and ‘Dementia treatment’. Results: Older time tested drugs like Acetyl Choline Receptor Inhibitors and NMDA Receptor antagonists remain the mainstay of pharmacological treatment in AD. Despite a lot of excitement about newer FDA approved drugs; we have to be cautious in their use. Aducanumab showed good reduction in CSF amyloid levels (biomarker of AD); but this did not necessarily translate into better clinical outcomes of patients. Conclusion: Despite the recent advances and approval of drugs in treatment of AD, we have to exhibit caution while prescribing these drugs. Even with a sound mechanism of action, these drugs do not always show improvement in clinical outcomes. More clinical trials are required for development of drugs in treatment of AD which explore various different mechanisms of action.

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.

Oligo-basic amino acids, potential nicotinic acetylcholine receptor inhibitors.

Oligo-basic amino acids have been extensively studied in molecular biology and pharmacology, but the inhibitory activity on nicotinic acetylcholine receptors (nAChRs) was unknown. In this study, the inhibitory activity of 8 oligopeptides, including both basic and acidic amino acids, was evaluated on 9 nAChR subtypes by a two-electrode voltage clamp (TEVC). Among them, the oligo-lysine K9, K12, d-K9, d-K9F, and oligo-arginine R9 showed nanomolar inhibitory activity on various nAChRs, especially for α7 and α9α10 nAChRs. d-K9 containing N-Fmoc protecting group (d-K9F) has an enhanced inhibitory activity on most of the nAChRs, including 47-fold promotion on α1β1δε nAChR. However, H9 and H12 only showed weak inhibitory activity on α9α10 and α1β1δε nAChRs, and the acidic oligopeptide D9 has no inhibitory activity on nAChRs. Flexible docking of K9 in α10(+) α9(-) and α7(+) α7(-) binding pockets showed particularly strong dipole-dipole interactions, which may be responsible for the inhibition of nAChRs. These results demonstrated that oligo-basic amino acids have the potential to be the lead compounds as selective nAChR subtype inhibitors, and oligo-lysines deserved to be modified for further exploitation and utilization. On the other hand, the toxicity and side effects of these nAChR inhibitory peptides should be contemplated in the application.

The Effects of Structural Alterations in the Polyamine and Amino Acid Moieties of Philanthotoxins on Nicotinic Acetylcholine Receptor Inhibition in the Locust, Schistocerca gregaria.

Alterations in the polyamine and amino acid (tyrosine) moieties of philanthotoxin-343 (PhTX-343) were investigated for their effects on the antagonism of nicotinic acetylcholine receptors (nAChRs) isolated from the locust (Schistocerca gregaria) mushroom body. Through whole-cell patch-clamp recordings, the philanthotoxin analogues in this study were shown to cause inhibition of the inward current when co-applied with acetylcholine (ACh). PhTX-343 (IC50 = 0.80 μM at −75 mV) antagonised locust nAChRs in a use-dependent manner, suggesting that it acts as an open-channel blocker. The analogue in which both the secondary amine functionalities were replaced with methylene groups (i.e., PhTX-12) was ~6-fold more potent (IC50 (half-maximal inhibitory concentration) = 0.13 μM at −75 mV) than PhTX-343. The analogue containing cyclohexylalanine as a substitute for the tyrosine moiety of PhTX-343 (i.e., Cha-PhTX-343) was also more potent (IC50 = 0.44 μM at −75 mV). A combination of both alterations to PhTX-343 generated the most potent analogue, i.e., Cha-PhTX-12 (IC50 = 1.71 nM at −75 mV). Modulation by PhTX-343 and Cha-PhTX-343 fell into two distinct groups, indicating the presence of two pharmacologically distinct nAChR groups in the locust mushroom body. In the first group, all concentrations of PhTX-343 and Cha-PhTX-343 inhibited responses to ACh. In the second group, application of PhTX-343 or Cha-PhTX-343 at concentrations ≤100 nM caused potentiation, while concentrations ≥ 1 μM inhibited responses to ACh. Cha-PhTX-12 may have potential to be developed into insecticidal compounds with a novel mode of action.

Effects of Acrylamide-Induced Vasorelaxation and Neuromuscular Blockage: A Rodent Study.

Acrylamide (ACR), which is formed during the Maillard reaction, is used in various industrial processes. ACR accumulation in humans and laboratory animals results in genotoxicity, carcinogenicity, neurotoxicity, and reproductive toxicity. In this study, we investigated the mechanisms by which ACR may induce vasorelaxation and neuromuscular toxicity. Vasorelaxation was studied using an isolated rat aortic ring model. The aortic rings were divided into the following groups: with or without endothelium, with nitric oxide synthase (NOS) inhibition, with acetylcholine receptor inhibition, and with extracellular calcium inhibition. Changes in tension were used to indicate vasorelaxation. Neuromuscular toxicity was assessed using a phrenic nerve–diaphragm model. Changes in muscle contraction stimulated by the phrenic nerve were used to indicate neuromuscular toxicity. ACR induced the vasorelaxation of phenylephrine-precontracted aortic rings, which could be significantly attenuated by NOS inhibitors. The results of the phrenic nerve–diaphragm experiments revealed that ACR reduced muscle stimulation and contraction through nicotinic acetylcholine receptor (AChR). ACR-induced vasotoxicity was regulated by NOS through the aortic endothelium. Nicotinic AChR regulated ACR-induced neuromuscular blockage.

Nicotine and its metabolite cotinine target MD2 and inhibit TLR4 signaling

Nicotine is the principal alkaloid of tobacco often manufactured into cigarettes and belongs to a highly addictive class of drugs. Nicotine attenuates the neuroinflammation induced by microglial activation. However, the molecular target(s) underlying anti-inflammatory action of nicotine has not been fully understood. Considering the psychoactive substances morphine, cocaine, and methamphetamine act as xenobiotic-associated molecular patterns and can be specifically sensed by the innate immune receptor Toll-like receptor 4 (TLR4), here we sought to delineate whether nicotine and/or its metabolite cotinine may be recognized by the innate immune system via myeloid differentiation protein 2 (MD2), an accessory protein of TLR4 that is responsible for ligand recognition. MD2-intrinsic fluorescence titrations, surface plasmon resonance, and competitive displacement binding assays with curcumin (MD2 probe) demonstrated that both nicotine and cotinine targeted the lipopolysaccharide (LPS; TLR4 agonist) binding pocket of MD2 with similar affinities. The cellular thermal shift assay indicated that nicotine binding increased, while cotinine binding decreased, MD2 stability. These biophysical binding results were further supported by in silico simulations. In keeping with targeting MD2, both nicotine and cotinine inhibited LPS-induced production of nitric oxide and tumor necrosis factor alpha (TNF-α) and blocked microglial activation. Neither a pan nicotinic acetylcholine receptor (nAChR) inhibitor nor RNAi for nAChRs abolished the suppressive effect of nicotine- and cotinine-induced neuroinflammation. These data indicate that TLR4 inhibition by nicotine and cotinine at the concentrations tested in BV-2 cells is independent of classic neuronal nAChRs and validate that MD2 is a direct target of nicotine and cotinine in the inhibition of innate immunity.

Methyl palmitate modulates the nicotine-induced increase in basilar arterial blood flow.

Methyl palmitate (MP) is a fatty acid methyl ester. Our recent study indicated that adrenergic nerve-dependent functional sympathetic-sensory nerve interactions were abolished by MP in mesenteric arteries. However, the effect of MP on perivascular nerves and cerebral blood flow remains unclear. In this study, the increase in basilar arterial blood flow (BABF) after the topical application of nicotinic acetylcholine receptor agonists was measured using laser Doppler flowmetry in anesthetized rats. The choline (a selective α7-nicotinic acetylcholine receptor agonist)-induced increase in BABF was abolished by tetrodotoxin (a neurotoxin), NG -nitro-L-arginine (a nonselective NO synthase inhibitor), α-bungarotoxin (a selective α7-nicotinic acetylcholine receptor inhibitor), and chronic sympathetic denervation. In addition, the nicotine (a nicotinic acetylcholine receptor agonist)-induced increase in BABF was inhibited by MP in a concentration-dependent manner. The acetylcholine-induced increase in BABF was not affected by MP. The myography results revealed that nicotine-induced vasorelaxation was significantly inhibited by MP, but was reversed by chelerythrine (a protein kinase C inhibitor). MP-induced vasodilation was significantly greater in BA rings without endothelium compared to those with endothelium. Meanwhile, MP did not affect baseline BABF. Our results indicate that MP acts as a neuromodulator in the cerebral circulation where it activates the PKC pathway and causes a diminished nicotine-induced increase in blood flow in the brainstem, and that the vasorelaxation effect of MP may play a minor role.

In vitro Neuromuscular Junction Induced from Human Induced Pluripotent Stem Cells.

The neuromuscular junction (NMJ) is a specialized synapse that transmits action potentials from the motor neuron to skeletal muscle for mechanical movement. The architecture of the NMJ structure influences the functions of the neuron, the muscle and the mutual interaction. Previous studies have reported many strategies by co-culturing the motor neurons and myotubes to generate NMJ in vitro with complex induction process and long culture period but have struggled to recapitulate mature NMJ morphology and function. Our in vitro NMJ induction system is constructed by differentiating human iPSC in a single culture dish. By switching the myogenic and neurogenic induction medium for induction, the resulting NMJ contained pre- and post- synaptic components, including motor neurons, skeletal muscle and Schwann cells in the one month culture. The functional assay of NMJ also showed that the myotubes contraction can be triggered by Ca++ then inhibited by curare, an acetylcholine receptor (AChR) inhibitor, in which the stimulating signal is transmitted through NMJ. This simple and robust approach successfully derived the complex structure of NMJ with functional connectivity. This in vitro human NMJ, with its integrated structures and function, has promising potential for studying pathological mechanisms and compound screening.

Structure-Function of Neuronal Nicotinic Acetylcholine Receptor Inhibitors Derived From Natural Toxins.

Neuronal nicotinic acetylcholine receptors (nAChRs) are prototypical cation-selective, ligand-gated ion channels that mediate fast neurotransmission in the central and peripheral nervous systems. nAChRs are involved in a range of physiological and pathological functions and hence are important therapeutic targets. Their subunit homology and diverse pentameric assembly contribute to their challenging pharmacology and limit their drug development potential. Toxins produced by an extensive range of algae, plants and animals target nAChRs, with many proving pivotal in elucidating receptor pharmacology and biochemistry, as well as providing templates for structure-based drug design. The crystal structures of these toxins with diverse chemical profiles in complex with acetylcholine binding protein (AChBP), a soluble homolog of the extracellular ligand-binding domain of the nAChRs and more recently the extracellular domain of human α9 nAChRs, have been reported. These studies have shed light on the diverse molecular mechanisms of ligand-binding at neuronal nAChR subtypes and uncovered critical insights useful for rational drug design. This review provides a comprehensive overview and perspectives obtained from structure and function studies of diverse plant and animal toxins and their associated inhibitory mechanisms at neuronal nAChRs.

Recent Research Progress in and Perspectives of Mesoionic Insecticides: Nicotinic Acetylcholine Receptor Inhibitors.

Triflumezopyrim exemplifies a new class of mesoionic insecticides and has attracted increasing attention as a result of its unique structure, high level of insecticidal activity, new mechanisms of action, low toxicity toward non-target organisms, and environmental friendliness. It inhibits the nicotinic acetylcholine receptor and has high potency against sucking pests, including the brown planthopper (Nilaparvata lugens), which has developed serious resistance to conventional neonicotinoids and low cross-resistance to some newly developed neonicotinoids. This review focuses on the discovery, synthesis, structure-activity relationships, and mechanism of action of mesoionic insecticides. Finally, potential directions for the development of mesoionic insecticides are discussed.

Robust Sampling of Defective Pathways in Alzheimer's Disease. Implications in Drug Repositioning.

We present the analysis of the defective genetic pathways of the Late-Onset Alzheimer’s Disease (LOAD) compared to the Mild Cognitive Impairment (MCI) and Healthy Controls (HC) using different sampling methodologies. These algorithms sample the uncertainty space that is intrinsic to any kind of highly underdetermined phenotype prediction problem, by looking for the minimum-scale signatures (header genes) corresponding to different random holdouts. The biological pathways can be identified performing posterior analysis of these signatures established via cross-validation holdouts and plugging the set of most frequently sampled genes into different ontological platforms. That way, the effect of helper genes, whose presence might be due to the high degree of under determinacy of these experiments and data noise, is reduced. Our results suggest that common pathways for Alzheimer’s disease and MCI are mainly related to viral mRNA translation, influenza viral RNA transcription and replication, gene expression, mitochondrial translation, and metabolism, with these results being highly consistent regardless of the comparative methods. The cross-validated predictive accuracies achieved for the LOAD and MCI discriminations were 84% and 81.5%, respectively. The difference between LOAD and MCI could not be clearly established (74% accuracy). The most discriminatory genes of the LOAD-MCI discrimination are associated with proteasome mediated degradation and G-protein signaling. Based on these findings we have also performed drug repositioning using Dr. Insight package, proposing the following different typologies of drugs: isoquinoline alkaloids, antitumor antibiotics, phosphoinositide 3-kinase PI3K, autophagy inhibitors, antagonists of the muscarinic acetylcholine receptor and histone deacetylase inhibitors. We believe that the potential clinical relevance of these findings should be further investigated and confirmed with other independent studies.

Myasthenia gravis AChR antibodies inhibit function of rapsyn-clustered AChRs

ObjectiveDirect inhibition of acetylcholine receptor (AChR) function by autoantibodies (Abs) is considered a rare pathogenic mechanism in myasthenia gravis (MG), but is usually studied on AChRs expressed in cell lines,...

Anti-cholinergics mecamylamine and scopolamine alleviate motion sickness-induced gastrointestinal symptoms through both peripheral and central actions

Enhanced cholinergic activity contributes to the production of complex autonomic manifestations of motion sickness (MS). However, whether anti-cholinergics exert their anti-MS effects through central or peripheral actions remained unclarified. In the present study, we investigated the effects of mecamylamine (MEC) and scopolamine (SCOP) on rotation-induced gastrointestinal symptoms (conditioned gaping and defecation), locomotion disturbances (hypoactivity and impaired balance performance), hypothermia as well as Fos expression in vestibulo-autonomic regions in rats. We also observed the effects of hexamethonium (HEX) and methyl scopolamine (MSCP) on those MS behavioral responses. The efficacy of all these drugs on rotation-induced emesis and other MS symptoms in cats was also examined. We found that intragastric administration of MEC and SCOP inhibited rotation-induced gaping and defecation in rats, but only MEC showed a dose-dependent manner. MEC aggravated rotation-induced balance disorder and failed to attenuate rotation-induced hypothermia as the SCOP did. MEC was more effective for inhibiting Fos expression in the caudal vestibular nucleus and nucleus of solitary tract than SCOP. Intraperitoneal injection of HEX and MSCP also significantly alleviated rotation-induced gastrointestinal symptoms, and showed benefit to balance performance in rats. In cats, MEC, SCOP and HEX had prophylactic effects against rotation-induced emesis and salivation, and deceased non-retching/vomiting symptoms, but MSCP only attenuated emesis. It suggested that MEC and SCOP might alleviate gastrointestinal symptoms of MS via inhibiting peripheral autonomic nervous system and central vestibulo-autonomic pathways. The nicotinic acetylcholine receptor inhibitors like MEC might be new candidates against gastrointestinal symptoms induced by MS or other vestibular disorders.

Abstract 1958: Benztropine enhances temozolomide sensitivity by restricting the growth of drug-resistant glioma stem cells

Abstract Glioblastomas (GBMs) are the most aggressive type of primary brain tumor in adults, the second leading cause of cancer-related death in children, and are incurable using currently available treatment modalities. GBMs are characterized by the presence of primitive glioma stem-like cell subpopulations that resist chemotherapy and sustain disease progression. In this study, we found that Benztropine, a muscarinic acetylcholine receptor (mAChR) inhibitor, suppresses glioma cell proliferation and sensitizes these cells to treatment with Temozolomide. Modeling the development of Temozolomide resistance in vitro, we identified a subpopulation of glioma cells, defined by the co-expression of CD44, CD15, and PDGFRa that was resistant to Temozolomide. This chemo-resistant subpopulation was phenotypically similar to oligodendroglial progenitor cells (OPC-like); a stem-like cell population important in the initiation and progression of glioblastoma known to be modulated by mAChR signaling. The addition of the mAChR inhibitor, benztropine, dramatically reduced the emergence of this Temozolomide-resistant sub-population. To begin understanding the mechanism of action of benztropine on GBM cells, we analyzed the effect of this drug on a panel of mitotic drivers that have been implicated in glioma initiation and progression. Sub-lethal doses of Benztropine alone or in combination with temozolomide repressed the PDGF-signaling axis as characterized by a significant reduction of PDGFA secretion and PDGFRA activation. We also developed an in vivo model of Temozolomide resistance to determine whether Benztropine could improve the survival of animals harboring glioma xenografts that had recurred following treatment with Temozolomide. These in vivo studies demonstrated that Benztropine alone improves animal survival as compared with vehicle controls and that Benztropine combined with Temozolomide reduces glioma growth following initial Temozolomide treatment and improves survival in mice xenografted with human GBM cells. Taken together, our results suggest that Benztropine might be targeting glioma stem cells by altering the glioma stem cell microenvironment. These findings support a potential role for the use of adjuvant Benztropine for the clinical management of GBM. Citation Format: Damian A. Almiron Bonnin, Myung Chang Lee, Joseph M. Howard, Alison L. Young, Matthew C. Havrda. Benztropine enhances temozolomide sensitivity by restricting the growth of drug-resistant glioma stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1958.

Histone hypo-acetylation of Sox9 mediates nicotine-induced weak cartilage repair by suppressing BMSC chondrogenic differentiation

BackgroundNicotine has negative effects on tissue repair, little research concerns its effect on the cartilage repair of tissue engineering stem cells. The present study aimed to investigate the effects of nicotine on the bone marrow-derived mesenchymal stem cells’ (BMSCs) chondrogenic repair function of cartilage defects and explored the molecular mechanism.MethodsA cartilage defect model of rat was repaired by BMSC transplantation, and treated with nicotine or saline at 2.0 mg/kg/d in 12 weeks. Nicotine’s effect on chondrogenic differentiation was studied by exposing BMSCs to nicotine at 0.1, 1, 10, and 100 μM, and methyllycaconitine (MLA), which is a selective α7-nicotinic acetylcholine receptor (nAChR) inhibitor and si-RNA of nuclear factor of activated T cells 2 (NFATc2), were used to verify the molecular mechanism of nicotine’s effect.ResultsData showed that nicotine inhibited cartilage repair function by suppressing SRY-type high-mobility group box 9 (Sox9) in regenerated tissues. Further in vitro study demonstrated that nicotine enhanced intracellular Ca2+ and activity of calcineurin (CaN) through α7-nAChR, increased the nucleic expressions of NFATc2 and the bindings to SOX9 promoter, and thus reduced the acetylation of H3K9 and H3K14 in SOX9 promoter.ConclusionsFindings from this study demonstrated that nicotine suppressed the chondrogenic differentiation of BMSCs in vivo and in vitro, which offers insight into the risk assessment of cartilage defect repair in a nicotine exposure population and its therapeutic target.

Recipient treatment with acetylcholinesterase inhibitor donepezil attenuates primary graft failure in rats through inhibiting post-transplantational donor heart ischaemia/reperfusion injury.

Ischaemia/reperfusion injury may have deleterious consequences on heart transplantation. The underlying causes such as inflammation may also contribute to the pathogenesis of primary or chronic graft failure. We hypothesize that donepezil (DO), a reversible acetylcholinesterase inhibitor that increases cholinergic receptor activation, may protect the transplanted heart through increasing the level of acetylcholine, which in turn inhibits systemic inflammation in the recipients. First, Lewis-Lewis heart transplantation model was successfully established, and 75 rats were randomly assigned into 3 groups: the DO group (received single dose of intragastric DO treatment), the donepezil + methyllycaconitine group (α7 nicotinic acetylcholine receptor inhibitor) and the control group. Ten rats per group were sacrificed at 24 h after drug administration, whereas the rest of the groups were observed 1 month after surgery. The status of inflammation, survival of the graft and function of the graft were examined. Serum tumour necrosis factor α level was significantly lower in the donepezil group when compared with the control group 24 h after first drug administration; this trend was maintained for 1 month (P < 0.001). Furthermore, DO inhibited CD11b/18-positive cell infiltration (P < 0.001) and myocardiocyte apoptosis (as shown by the percentage of terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate-biotin nick-end labelling-positive nuclei, P = 0.039) in the recipient rats at 24 h after the first drug administration. The percentage of cardiac grafts that survived for 1 month in rats given DO alone was significantly higher (80.0%, 33.3% and 26.7% in the DO, donepezil + methyllycaconitine and control groups, respectively, P = 0.014); the fractional shortening value of the DO group was significantly higher than that in the other 2 groups (29.25 ± 1.84%, 17.92 ± 3.69% and 17.07 ± 2.99% in the DO, donepezil + methyllycaconitine and control group, respectively, P < 0.001). The collagen volume fraction was lower in the DO group than in the other 2 groups (P < 0.001). Our results reveal that treatment of the recipient with DO protects the donor hearts for 1 month after transplantation through suppressing the signalling pathway of inflammation. These results suggest that DO is a novel and clinically feasible strategy to protect the donor heart in transplantation in the long term.

Nicotine induces EP4 receptor expression in lung carcinoma cells by acting on AP-2α: The intersection between cholinergic and prostanoid signaling.

It was demonstrated that nicotine increased non-small cell lung cancer cell proliferation through nicotinic acetylcholine receptor -mediated signals. However, the detailed mechanism remains incompletely understood. We evaluated whether nicotine increased EP4 receptor expression in lung carcinoma cells by activating on AP-2α. Methods: The non-small cell lung cancer cells of A549 and H1838 were cultured and treated with EP4 inhibitor AH23848, also with EP4 and control siRNAs. The extracellular signal-regulated kinases inhibitor PD98059, the p38 mitogen-activated protein kinase inhibitor SB239063, the α7 nicotinic acetylcholine receptor inhibitor α-bungarotoxin, the α4 nicotinic acetylcholine receptor inhibitor dihydro-β-erythroidine, the PI3K inhibitor wortmannin, the PKC inhibitor calphostin C, and the PKA inhibitor H89 have been used to evaluate the effects on proliferations. It indicates that nicotine increases EP4 expression through α7 nicotinic acetylcholine receptor-dependent activations of PI3-K, JNK and PKC pathways that leads to reduction of AP-2α-DNA binding. This, together with the elevated secretion of PGE2, further enhances the tumor promoting effects of nicotine. These studies suggest a novel molecular mechanism by which nicotine increases non-small cell lung cancer cell proliferation.

α7-Nicotinic acetylcholine receptor inhibition by indinavir: implications for cognitive dysfunction in treated HIV disease.

The study set out to determine if the HIV protease inhibitor, indinavir, alters responsiveness of α7-nicotinic acetylcholine receptors to acetylcholine. Treatment with HAART has dramatically reduced development of HIV-associated dementia and more severe forms of cognitive impairment. However, many individuals continue to experience cognitive decline of uncertain cause. Previous studies have failed to demonstrate significant alterations of functional brain connectivity, structural brain changes, or changes in cerebral blood flow sufficient to explain cognitive decline in virally suppressed individuals. This suggests that the mechanisms underlying development and progression of cognitive problems likely occurs at a micro rather than macro level, such as disruptions in neurotransmitter system signaling. Indinavir's effects on α7-nicotinic acetylcholine receptor activity was tested using a ScreenPatch IonWorks Barracuda-based assay in a mammalian cell model. At low concentrations (0.0003-10 μmol/l) indinavir acts as a positive allosteric modulator (EC50 = 0.021 μmol/l), whereas at concentrations greater than 10 μmol/l (30-100 μmol/l) indinavir acts as an inhibitor of the α7-nicotinic acetylcholine receptor. At concentrations greater than 10 μmol/l indinavir reduces synaptic transmission in the acetylcholine neurotransmitter system, which could possibly contribute to cognitive dysfunction. These results suggest that further experiments should be considered to assess whether patients might benefit from treatment with cholinesterase inhibitors that counteract the effects of indinavir.

Nicotine may promote tongue squamous cell carcinoma progression by activating the Wnt/β-catenin and Wnt/PCP signaling pathways.

To investigate the effects and the possible underlying mechanisms of nicotine stimulation on tongue squamous cell carcinoma (TSCC) progression, a TSCC cell line Cal27 and 34 samples of paraffin-embedded TSCC were examined. Immunofluorescence, western blot analysis, and TOP/FOP flash, CCK-8, wound healing and Transwell invasion assays were used to evaluate Cal27 in response to nicotine stimulation. We also investigated expression levels of related proteins of Wnt/β-catenin and Wnt/PCP pathways in paraffin-embedded TSCC samples with or without a history of smoking by immunohistochemistry. Nicotine stimulation can promote proliferation, migration, and invasion of TSCC cells in vitro, downregulate E-cadherin, and activate the Wnt/β-catenin and Wnt/PCP pathways, which could be antagonized by the α7 nicotine acetylcholine receptor (α7 nAChR) inhibitor α-BTX. Moreover, the expression levels of β-catenin, Wnt5a and Ror2 were higher in TSCC patients with a history of smoking than those without a history of smoking. Our results suggest nicotine may promote tongue squamous carcinoma cells progression by activating the Wnt/β-catenin and Wnt/PCP signaling pathways and may play a significant role in the progression and metastasis of smoking-related TSCC.