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Background and PurposeNicotinic acetylcholine receptors (nAChRs), which are expressed throughout the mammalian brain, mediate a variety of physiological functions. Despite their widespread presence, the functions of nAChRs are not yet fully understood. α-Conotoxins, which are peptides derived from the venom of marine cone snails, target different subtypes of nAChRs. Specifically, α-Conotoxins [D1G, ΔQ14] LvIC, identified from Conus lividus, have demonstrated strong activity on α6β4* nAChRs in vitro. However, the effects of [D1G, ΔQ14] LvIC have not been investigated in vivo. This study aims to examine the activities of [D1G, ΔQ14] LvIC and explore its potential mechanisms in vivo.MethodsThe study involved the injection of [D1G, ΔQ14] LvIC into the lateral cerebral ventricle (LV) of mice. Following this procedure, behavioral tests were conducted to assess changes in the mice’s behavior. To investigate the molecular alterations in the mice’s brains, untargeted metabolomics and label-free Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) were employed. Subsequently, Western blot (WB) and quantitative reverse transcription PCR (RT-qPCR) techniques were utilized to detect specific molecular changes induced by [D1G, ΔQ14] LvIC.ResultsThe injection of [D1G, ΔQ14] LvIC led to a decrease in locomotor activity in mice. This treatment also resulted in reduced expression of neuronal calcium sensor 1 (NCS-1) and neuroligin 3 (NLGN-3) in the prefrontal cortex (PFC), hippocampus (Hip), and caudate putamen (CPu). Both NCS-1 and NLGN-3 are crucial for neuronal development, synapse formation, and neuron activity, and their reduction is associated with decreased synapse strength. Despite these changes, results from the Morris water maze (MWM) indicated that [D1G, ΔQ14] LvIC did not impair the learning and memory abilities of the mice.ConclusionOur findings indicate that α-conotoxin [D1G, ΔQ14] LvIC significantly decreased locomotor activity in mice. Additionally, it altered gene expression primarily in areas related to neuronal development, synapse formation, and neuron activity, while also reducing synapse strength. This study first proposed that [D1G, ΔQ14] LvIC could modulate mice’s locomotor activity. However, further investigation is needed to understand the therapeutic effects of [D1G, ΔQ14] LvIC.

Clinical and preclinical studies have identified immunosuppressive effects of nicotine, with potential implications for treating nicotine addiction. Here we review how nicotine can regulate microglia, the resident macrophages in the brain, and corresponding effects of nicotine on neuroimmune signaling. There is significant evidence that activation of α7 nicotinic acetylcholine receptors (nAChRs) on microglia can trigger an anti-inflammatory cascade that alters microglial polarization and activity, cytokine release, and intracellular calcium concentrations, leading to neuroprotection. These anti-inflammatory effects of nicotine-dependent α7 nAChR signaling are lost during withdrawal, suggesting that neuroimmune signaling is potentiated during abstinence, and thus, heightened microglial activity may drive circuit disruption that contributes to withdrawal symptoms and hyperkatifeia. In sum, the clinical literature has highlighted immunomodulatory effects of nicotine and the potential for anti-inflammatory compounds to treat addiction. The preclinical literature investigating the underlying mechanisms points to a role of microglial engagement in the circuit dysregulation and behavioral changes that occur during nicotine addiction and withdrawal, driven, at least in part, by activation of α7 nAChRs on microglia. Specifically targeting microglial signaling may help alleviate withdrawal symptoms in people with nicotine dependence and help to promote abstinence. [ABSTRACT FROM AUTHOR]

The venom of cone snails has been proven to be a rich source of bioactive peptides that target a variety of ion channels and receptors. α-Conotoxins (αCtx) interact with nicotinic acetylcholine receptors (nAChRs) and are powerful tools for investigating the structure and function of the various nAChR subtypes. By studying how conotoxins interact with nAChRs, we can improve our understanding of these receptors, leading to new insights into neurological diseases associated with nAChRs. Here, we describe the discovery and characterization of a novel conotoxin from Conus ateralbus, αCtx-AtIA, which has an amino acid sequence homologous to the well-described αCtx-PeIA, but with a different selectivity profile towards nAChRs. We tested the synthetic αCtx-AtIA using the calcium imaging-based Constellation Pharmacology assay on mouse DRG neurons and found that αCtx-AtIA significantly inhibited ACh-induced calcium influx in the presence of an α7 positive allosteric modulator, PNU-120596 (PNU). However, αCtx-AtIA did not display any activity in the absence of PNU. These findings were further validated using two-electrode voltage clamp electrophysiology performed on oocytes overexpressing mouse α3β4, α6/α3β4 and α7 nAChRs subtypes. We observed that αCtx-AtIA displayed no or low potency in blocking α3β4 and α6/α3β4 receptors, respectively, but improved potency and selectivity to block α7 nAChRs when compared with αCtx-PeIA. Through the synthesis of two additional analogs of αCtx-AtIA and subsequent characterization using Constellation Pharmacology, we were able to identify residue Trp18 as a major contributor to the activity of the peptide. [ABSTRACT FROM AUTHOR]

Gang Chen,1 Qing Mu,1 Zhao-Ji Meng2 1Department of Respiratory and Critical Care Medicine, Henan Provincial People’s Hospital, People’s Hospital of Zhengzhou University, Zhengzhou, Henan, People’s Republic of China; 2Department of Immune Allergy, Henan Provincial People’s Hospital, People’s Hospital of Zhengzhou University, Zhengzhou, Henan, People’s Republic of ChinaCorrespondence: Gang Chen, Department of Respiratory and Critical Care Medicine, Henan Provincial People’s Hospital, People’s Hospital of Zhengzhou University, 7 Weiwu Road, Zhengzhou, Henan, 450003, People’s Republic of China, Email chengangvips@163.comBackground: Dysregulation and pyroptosis of T-helper (Th) cells and inflammatory cytokines have been implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD). However, the immune response mechanisms as a consequence of tobacco smoke exposure are not fully understood. We hypothesized that cigarette smoke-induced inflammation could be modulated through the cytokine milieu and T-cell nicotinic acetylcholine receptors (nAChRs).Methods: The proportions of peripheral blood Th1 and Th2 cells from patients with COPD, smokers without airway obstruction and healthy nonsmokers were analyzed using flow cytometry. The levels of plasma proinflammatory cytokines and their potential association with pulmonary function were also measured. The influence of cigarette smoke extract (CSE) on the conditioned differentiation of T helper cell subsets was further examined in vitro.Results: Significantly higher Th1 cell and plasma IFN-γ and IL-18 levels but lower levels of Th2 cells were found in the peripheral blood from patients with COPD. The increased plasma levels of IFN-γ and IL-18 were negatively correlated with pulmonary function (FEV1% predicted value). Pyroptosis participates in COPD development probably through the activation of the NLRP3 inflammasome upon exposure to CSE. CSE does not directly induce the differentiation of T helper cells; however, under conditioned medium, CSE promotes Th1 development through α 7 nAChR modification, while it does not substantially interfere with Th2 differentiation.Conclusion: The differences in the cytokine milieu play a key role in the effects of CSE on the immune response in patients with COPD.Keywords: chronic obstructive pulmonary disease, COPD, cigarette smoke, T helper 1, Th1 cells, Th2, nicotinic acetylcholine receptors, nAChRs

α7 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain where they promote fast cholinergic synaptic transmission and serve important neuromodulatory functions. However, their high permeability to Ca2+ also predisposes them to contribute to disease states. Here, using transfected HEK-tsa cells and primary cultured hippocampal neurons from male and female rats, we demonstrate that two proteins called Ly6h and NACHO compete for access to α7 subunits, operating together but in opposition to maintain α7 assembly and activity within a narrow range that is optimal for neuronal function and viability. Using mixed gender human temporal cortex and cultured hippocampal neurons from rats we further show that this balance is perturbed during Alzheimers disease (AD) because of amyloid β (Aβ)-driven reduction in Ly6h, with severe reduction leading to increased phosphorylated tau and α7-mediated neurotoxicity. Ly6h release into human CSF is also correlated with AD severity. Thus, Ly6h links cholinergic signaling, Aβ and phosphorylated tau and may serve as a novel marker for AD progression.SIGNIFICANCE STATEMENT One of the earliest and most persistent hypotheses regarding Alzheimers disease (AD) attributes cognitive impairment to loss of cholinergic signaling. More recently, interest has focused on crucial roles for amyloid β (Aβ) and phosphorylated tau in Alzheimers pathogenesis. Here, we demonstrate that these elements are linked by Ly6h and its counterpart, NACHO, functioning in opposition to maintain assembly of nicotinic acetylcholine receptors (nAChRs) within the physiological range. Our data suggests that Aβ shifts the balance away from Ly6h and toward NACHO, resulting in increased assembly of Ca2+-permeable nAChRs and thus a conversion of basal cholinergic to neurotoxic signaling.

Sarcopenia involves in the loss of muscle mass associated with aging, which is the major cause of progressive muscle weakness and deterioration in older adults. Muscle atrophy is a direct presentation of sarcopenia, and it greatly contributes to the decline in quality of life among older adults. Neuromuscular junction (NMJ) stability is the key link to maintain muscle function. Besides, the degenerative change of NMJ promotes the process of muscle atrophy in the elderly. Based on previous transcriptome sequencing and bioinformatics analyses of aged muscle, this study used the 18-month-old aged mouse model and the 6-month-old young mouse model to deliberate the role and underlying mechanisms of Cullin-3 (Cul3) in age-related muscle atrophy. The results of reverse transcriptase polymerase chain reaction (RT-PCR) and immunoblotting analysis showed that the expression of CUL3 increased in aged muscle tissue, while the expression level of postsynaptic membrane nicotinic acetylcholine receptors (nAChRs) decreased significantly, which manfested a negative correlation. Meanwhile, immunofluorescence demonstrated that Cul3 was highly expressed in senile muscle NMJ. The results of ubiquitin indicated that the ubiquitin level of aged muscle nAChRs was evidently increased. Co-immunoprecipitation furtherly verified the correlation between Cul3 and nAChRs. Taken together, Cul3 may mediate the ubiquitination degradation of nAChRs protein at the NMJ site in aged mice, leading to NMJ degeneration and accelerated atrophy of fast-twitch muscle fibers in aged muscle. As a prominent element to maintain the stability of NMJ, Cul3 is supposed to be one of candidate intervention targets in sarcopenia.

The cholinergic system plays an essential role in brain development, physiology, and pathophysiology. Herein, we review how specific alterations in this system, through genetic mutations or abnormal receptor function, can lead to aberrant neural circuitry that triggers disease. The review focuses on the nicotinic acetylcholine receptor (nAChR) and its role in addiction and in neurodegenerative and neuropsychiatric diseases and epilepsy. Cholinergic dysfunction is associated with inflammatory processes mainly through the involvement of α7 nAChRs expressed in brain and in peripheral immune cells. Evidence suggests that these neuroinflammatory processes trigger and aggravate pathological states. We discuss the preclinical evidence demonstrating the therapeutic potential of nAChR ligands in Alzheimer disease, Parkinson disease, schizophrenia spectrum disorders, and in autosomal dominant sleep-related hypermotor epilepsy. PubMed and Google Scholar bibliographic databases were searched with the keywords indicated below. [ABSTRACT FROM AUTHOR]

The venom of cone snails has been proven to be a rich source of bioactive peptides that target a variety of ion channels and receptors. α-Conotoxins (αCtx) interact with nicotinic acetylcholine receptors (nAChRs) and are powerful tools for investigating the structure and function of the various nAChR subtypes. By studying how conotoxins interact with nAChRs, we can improve our understanding of these receptors, leading to new insights into neurological diseases associated with nAChRs. Here, we describe the discovery and characterization of a novel conotoxin from Conus ateralbus, αCtx-AtIA, which has an amino acid sequence homologous to the well-described αCtx-PeIA, but with a different selectivity profile towards nAChRs. We tested the synthetic αCtx-AtIA using the calcium imaging-based Constellation Pharmacology assay on mouse DRG neurons and found that αCtx-AtIA significantly inhibited ACh-induced calcium influx in the presence of an α7 positive allosteric modulator, PNU-120596 (PNU). However, αCtx-AtIA did not display any activity in the absence of PNU. These findings were further validated using two-electrode voltage clamp electrophysiology performed on oocytes overexpressing mouse α3β4, α6/α3β4 and α7 nAChRs subtypes. We observed that αCtx-AtIA displayed no or low potency in blocking α3β4 and α6/α3β4 receptors, respectively, but improved potency and selectivity to block α7 nAChRs when compared with αCtx-PeIA. Through the synthesis of two additional analogs of αCtx-AtIA and subsequent characterization using Constellation Pharmacology, we were able to identify residue Trp18 as a major contributor to the activity of the peptide.

Abstract Background As a human tumor disease, head and neck squamous cell carcinoma (HNSCC) is associated with a high mortality rate worldwide. Nicotinic acetylcholine receptors (nAChRs) are transmembrane receptor proteins and exert their biological effects following activation by nicotine. We aimed to construct a prognostic signature based on the expression of nAChRs among smokers with HNSCC. Methods The transcriptome profile of nAChRs was obtained from The Cancer Genome Atlas (TCGA). Following the integration of survival information, univariate Cox regression and least absolute shrinkage and selection operator (LASSO) analyses were performed to screen the prognosis-related nAChRs and construct a prognostic signature. Kaplan–Meier (KM), receiver operating characteristic (ROC), principal component analysis (PCA), and independent prognostic analysis were utilized to verify the predictive power of the nAChR-associated prognostic signature. The expression of α5 nAChR in clinical samples was verified by quantitative reverse transcriptase PCR. Results Subunits α2, α5, α9, and β4 were related to the prognosis. The prognostic signature comprised the expression of subunits α5, α9, and β4. The nAChR-associated signature showed high sensitivity and specificity for prognostic prediction and was an independent factor for overall survival. Based on the clinical variables and expression of nAChRs, a nomogram was constructed for predicting the outcomes of HNSCC patients who were smokers in the clinical settings. In clinical specimens, α5 nAChR showed high expression in HNSCC tissues, especially among smokers. Conclusions The nAChR-associated signature constructed in this study may provide a better system for the classification of HNSCC patients and facilitate personalized treatment according to their smoking habits.

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. [ABSTRACT FROM AUTHOR]

Background: As a human tumor disease, head and neck squamous cell carcinoma (HNSCC) is associated with a high mortality rate worldwide. Nicotinic acetylcholine receptors (nAChRs) are transmembrane receptor proteins and exert their biological effects following activation by nicotine. We aimed to construct a prognostic signature based on the expression of nAChRs among smokers with HNSCC.Methods: The transcriptome profile of nAChRs was obtained from The Cancer Genome Atlas (TCGA). Following the integration of survival information, univariate Cox regression and least absolute shrinkage and selection operator (LASSO) analyses were performed to screen the prognosis-related nAChRs and construct a prognostic signature. Kaplan-Meier (KM), receiver operating characteristic (ROC), principal component analysis (PCA), and independent prognostic analysis were utilized to verify the predictive power of the nAChR-associated prognostic signature. The expression of α5 nAChR in clinical samples was verified by quantitative reverse transcriptase PCR.Results: Subunits α2, α5, α9, and β4 were related to the prognosis. The prognostic signature comprised the expression of subunits α5, α9, and β4. The nAChR-associated signature showed high sensitivity and specificity for prognostic prediction and was an independent factor for overall survival. Based on the clinical variables and expression of nAChRs, a nomogram was constructed for predicting the outcomes of HNSCC patients who were smokers in the clinical settings. In clinical specimens, α5 nAChR showed high expression in HNSCC tissues, especially among smokers.Conclusions: The nAChR-associated signature constructed in this study may provide a better system for the classification of HNSCC patients and facilitate personalized treatment according to their smoking habits. [ABSTRACT FROM AUTHOR]

Conus regius is a marine venomous mollusk of the Conus genus that captures its prey by injecting a rich cocktail of bioactive disulfide bond rich peptides called conotoxins. These peptides selectively target a broad range of ion channels, membrane receptors, transporters, and enzymes, making them valuable pharmacological tools and potential drug leads. C. regius-derived conotoxins are particularly attractive due to their marked potency and selectivity against specific nicotinic acetylcholine receptor subtypes, whose signalling is involved in pain, cognitive disorders, drug addiction, and cancer. However, the species-specific differences in sensitivity and the low stability and bioavailability of these conotoxins limit their clinical development as novel therapeutic agents for these disorders. Here, we give an overview of the main pharmacological features of the C. regius-derived conotoxins described so far, focusing on the molecular mechanisms underlying their potential therapeutic effects. Additionally, we describe adoptable chemical engineering solutions to improve their pharmacological properties for future potential clinical translation. [ABSTRACT FROM AUTHOR]

The α4β2 nAChRs are crucial ion channels that control neurotransmitter release and play a role in various physiologic and pathologic processes. CHRNA4 encodes the α4-nAChRs, while CHRNB2 encodes the β2-nAChRs. Recent studies have found different variants of α4β2-nAChRs in individuals with conditions such as AD, ADHD, ALS, PD, and brain abnormalities. We conducted a scoping review following a six-stage methodology structure and adhering to PRISMA guidelines. We systematically reviewed articles using relevant keywords up to October 2, 2023. In this summary, we cover the clinical symptoms reported, the genes and protein structure of CHRNA4 and CHRNB2, mutations in these genes, inheritance patterns, the functional impact of mutations and polymorphisms in CHRNA4 and CHRNB2, and the epidemiology of these diseases. Recent research indicates that nAChRs may play a significant role in neurodegenerative disorders, possibly impacting neuronal function through yet undiscovered regulatory pathways. Studying how nAChRs interact with disease-related aggregates in neurodegenerative conditions may lead to new treatment options for these disorders., (© 2024 International Society for Neurochemistry.)

Pesticides play an active role in modern agricultural production. However, irrational application of pesticides may cause a negative impact on important pollinating bees. This paper mainly reviewed the research progress on the acute toxicity of pesticides to bees, the sub-lethal effects of pesticides on the growth, development, and behavior of bees, the combined effect of pesticides on bees, as well as the interaction between neonicotinoid insecticides and nicotinic acetylcholine receptors of bees, mechanisms of differential toxicity of pesticide to bees, effects of pesticides on detoxificationrelated enzyme activity and other physiological and biochemical indexes of bees, and the role of phytochemicals in regulating the tolerance of bees to pesticides. The main purpose of this review is to provide a reference for the rational use of pesticides and improve the safety of pesticides to bees. [ABSTRACT FROM AUTHOR]

Molecular docking, pivotal in predicting small-molecule ligand binding modes, struggles with accurately identifying binding conformations and affinities. This is particularly true for neonicotinoids, insecticides whose impacts on ecosystems require precise molecular interaction modeling. This study scrutinizes the effectiveness of prominent docking software (Ledock, ADFR, Autodock Vina, CDOCKER) in simulating interactions of environmental chemicals, especially neonicotinoid-like molecules with nicotinic acetylcholine receptors (nAChRs) and acetylcholine binding proteins (AChBPs). We aimed to assess the accuracy and reliability of these tools in reproducing crystallographic data, focusing on semi-flexible and flexible docking approaches. Our analysis identified Ledock as the most accurate in semi-flexible docking, while Autodock Vina with Vinardo scoring function proved most reliable. However, no software consistently excelled in both accuracy and reliability. Additionally, our evaluation revealed that none of the tools could establish a clear correlation between docking scores and experimental dissociation constants (K d) for neonicotinoid-like compounds. In contrast, a strong correlation was found with drug-like compounds, bringing to light a bias in considered software towards pharmaceuticals, thus limiting their applicability to environmental chemicals. The comparison between semi-flexible and flexible docking revealed that the increased computational complexity of the latter did not result in enhanced accuracy. In fact, the higher computational cost of flexible docking with its lack of enhanced predictive accuracy, rendered this approach useless for this class of compounds. Conclusively, our findings emphasize the need for continued development of docking methodologies, particularly for environmental chemicals. This study not only illuminates current software capabilities but also underscores the urgency for advancements in computational molecular docking as it is a relevant tool to environmental sciences. [Display omitted] • Benchmarked docking tools for neonicotinoids using 54 PDB protein-ligand systems with nAChRs and AChBPs. • Flexible docking appeared to be less accurate and more computationally demanding than semi-flexible docking. • Revealed biases in current software, highlighting the need for tools tailored to insecticides. [ABSTRACT FROM AUTHOR]

Physical exercise improves motor performance in individuals with Parkinson's disease and elevates mood in those with depression. Although underlying factors have not been identified, clues arise from previous studies showing a link between cognitive benefits of exercise and increases in brain-derived neurotrophic factor (BDNF). Here, we investigated the influence of voluntary wheel-running exercise on BDNF levels in the striatum of young male wild-type (WT) mice, and on the striatal release of a key motor-system transmitter, dopamine (DA). Mice were allowed unlimited access to a freely rotating wheel (runners) or a locked wheel (controls) for 30 d. Electrically evoked DA release was quantified in ex vivo corticostriatal slices from these animals using fast-scan cyclic voltammetry. We found that exercise increased BDNF levels in dorsal striatum (dStr) and increased DA release in dStr and in nucleus accumbens core and shell. Increased DA release was independent of striatal acetylcholine (ACh), and persisted after a week of rest. We tested a role for BDNF in the influence of exercise on DA release using mice that were heterozygous for BDNF deletion (BDNF+/-). In contrast to WT mice, evoked DA release did not differ between BDNF+/- runners and controls. Complementary pharmacological studies using a tropomyosin receptor kinase B (TrkB) agonist in WT mouse slices showed that TrkB receptor activation also increased evoked DA release throughout striatum in an ACh-independent manner. Together, these data support a causal role for BDNF in exercise-enhanced striatal DA release and provide mechanistic insight into the beneficial effects of exercise in neuropsychiatric disorders, including Parkinson's, depression, and anxiety. [ABSTRACT FROM AUTHOR]

Acetylcholine is an important modulator of striatal activity, and it is vital to controlling striatal-dependent behaviors, including motor and cognitive functions. Despite this significance, the mechanisms determining how acetylcholine impacts striatal signaling are still not fully understood. In particular, little is known about the role of nAChRs expressed by striatal interneurons. In the present study, we used FISH to determine which neuronal types express the most prevalent beta2 nicotinic subunit in the mouse striatum. Our data support a common view that nAChR expression is mostly restricted to striatal interneurons. Surprisingly though, cholinergic interneurons were identified as a population with the highest expression of beta2 nicotinic subunit. To investigate the functional significance of beta2- containing nAChRs in striatal interneurons, we deleted them by injecting the AAV-Cre vector into the striatum of beta2-flox/flox male mice. The deletion led to alterations in several behavioral domains, namely, to an increased anxiety- like behavior, decrease in sociability ratio, deficit in discrimination learning, and increased amphetamine-induced hyperlocomotion and c-Fos expression in mice with beta2 deletion. Further colocalization analysis showed that the increased c-Fos expression was present in both medium spiny neurons and presumed striatal interneurons. The present study concludes that, despite being relatively rare, beta2-containing nAChRs are primarily expressed in striatal neurons by cholinergic interneurons and play a significant role in behavior. [ABSTRACT FROM AUTHOR]

As an anti-inflammatory alkaloid, nicotine plays dual roles in treating diseases. Here we reviewed the anti-inflammatory and pro-inflammatory effects of nicotine on inflammatory diseases, including inflammatory bowel disease, arthritis, multiple sclerosis, sepsis, endotoxemia, myocarditis, oral/skin/muscle inflammation, etc. , mainly concerning the administration methods, different models, therapeutic concentration and duration, and relevant organs and tissues. According to the data analysis from recent studies in the past 20 years, nicotine exerts much more anti-inflammatory effects than pro-inflammatory ones, especially in ulcerative colitis, arthritis, sepsis, and endotoxemia. On the other hand, in oral inflammation, nicotine promotes and aggravates some diseases such as periodontitis and gingivitis, especially when there are harmful microorganisms in the oral cavity. We also carefully analyzed the nicotine dosage to determine its safe and effective range. Furthermore, we summarized the molecular mechanism of nicotine in these inflammatory diseases through regulating immune cells, immune factors, and the vagus and acetylcholinergic anti-inflammatory pathways. By balancing the "beneficial" and "harmful" effects of nicotine, it is meaningful to explore the effective medical value of nicotine and open up new horizons for remedying acute and chronic inflammation in humans. [ABSTRACT FROM AUTHOR]

As an anti-inflammatory alkaloid, nicotine plays dual roles in treating diseases. Here we reviewed the anti-inflammatory and pro-inflammatory effects of nicotine on inflammatory diseases, including inflammatory bowel disease, arthritis, multiple sclerosis, sepsis, endotoxemia, myocarditis, oral/skin/muscle inflammation, etc., mainly concerning the administration methods, different models, therapeutic concentration and duration, and relevant organs and tissues. According to the data analysis from recent studies in the past 20 years, nicotine exerts much more anti-inflammatory effects than pro-inflammatory ones, especially in ulcerative colitis, arthritis, sepsis, and endotoxemia. On the other hand, in oral inflammation, nicotine promotes and aggravates some diseases such as periodontitis and gingivitis, especially when there are harmful microorganisms in the oral cavity. We also carefully analyzed the nicotine dosage to determine its safe and effective range. Furthermore, we summarized the molecular mechanism of nicotine in these inflammatory diseases through regulating immune cells, immune factors, and the vagus and acetylcholinergic anti-inflammatory pathways. By balancing the “beneficial” and “harmful” effects of nicotine, it is meaningful to explore the effective medical value of nicotine and open up new horizons for remedying acute and chronic inflammation in humans.

Conus regius is a marine venomous mollusk of the Conus genus that captures its prey by injecting a rich cocktail of bioactive disulfide bond rich peptides called conotoxins. These peptides selectively target a broad range of ion channels, membrane receptors, transporters, and enzymes, making them valuable pharmacological tools and potential drug leads. C. regius-derived conotoxins are particularly attractive due to their marked potency and selectivity against specific nicotinic acetylcholine receptor subtypes, whose signalling is involved in pain, cognitive disorders, drug addiction, and cancer. However, the species-specific differences in sensitivity and the low stability and bioavailability of these conotoxins limit their clinical development as novel therapeutic agents for these disorders. Here, we give an overview of the main pharmacological features of the C. regius-derived conotoxins described so far, focusing on the molecular mechanisms underlying their potential therapeutic effects. Additionally, we describe adoptable chemical engineering solutions to improve their pharmacological properties for future potential clinical translation.

α7 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain where they promote fast cholinergic synaptic transmission and serve important neuromodulatory functions. However, their high permeability to Ca2+ also predisposes them to contribute to disease states. Here, using transfected HEK-tsa cells and primary cultured hippocampal neurons from male and female rats, we demonstrate that two proteins called Ly6h and NACHO compete for access to α7 subunits, operating together but in opposition to maintain α7 assembly and activity within a narrow range that is optimal for neuronal function and viability. Using mixed gender human temporal cortex and cultured hippocampal neurons from rats we further show that this balance is perturbed during Alzheimer's disease (AD) because of amyloid β (Aβ)-driven reduction in Ly6h, with severe reduction leading to increased phosphorylated tau and α7-mediated neurotoxicity. Ly6h release into human CSF is also correlated with AD severity. Thus, Ly6h links cholinergic signaling, Aβ and phosphorylated tau and may serve as a novel marker for AD progression. [ABSTRACT FROM AUTHOR]

Acetylcholine is an abundant neurotransmitter in all animals. Effects of acetylcholine are excitatory, inhibitory, or modulatory depending on the receptor and cell type. Research using the nematode C. elegans has made ground‐breaking contributions to the mechanistic understanding of cholinergic transmission. Powerful genetic screens for behavioral mutants or for responses to pharmacological reagents identified the core cellular machinery for synaptic transmission. Pharmacological reagents that perturb acetylcholine‐mediated processes led to the discovery and also uncovered the composition and regulators of acetylcholine‐activated channels and receptors. From a combination of electrophysiological and molecular cellular studies, we have gained a profound understanding of cholinergic signaling at the levels of synapses, neural circuits, and animal behaviors. This review will begin with a historical overview, then cover in‐depth current knowledge on acetylcholine‐activated ionotropic receptors, mechanisms regulating their functional expression and their functions in regulating locomotion. [ABSTRACT FROM AUTHOR]

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.

The medial nucleus of trapezoid body (MNTB) is a major source of inhibition in auditory brainstem circuitry. The MNTB projects well-timed inhibitory output to principal sound-localization nuclei in the superior olive (SOC) as well as other computationally important centers. Acoustic information is conveyed to MNTB neurons through a single calyx of Held excitatory synapse arising from the cochlear nucleus. The encoding efficacy of this large synapse depends on its activity rate, which is primarily determined by sound intensity and stimulus frequency. However, MNTB activity rate is additionally influenced by inhibition and possibly neuromodulatory inputs, albeit their functional role is unclear. Happe and Morley (2004) discovered prominent expression of a7 nAChRs in rat SOC, suggesting possible engagement of ACh-mediated modulation of neural activity in the MNTB. However, the existence and nature of this putative modulation have never been physiologically demonstrated. We probed nicotinic cholinergic influences on acoustic responses of MNTB neurons from adult gerbils (Meriones unguiculatus) of either sex. We recorded tone-evoked MNTB single-neuron activity in vivo using extracellular single-unit recording. Piggyback multibarrel electrodes enabled pharmacological manipulation of nAChRs by reversibly applying antagonists to two receptor types, a7 and a4b2. We observed that tone-evoked responses are dependent on ACh modulation by both nAChR subtypes. Spontaneous activity was not affected by antagonist application. Functionally, we demonstrate that ACh contributes to sustaining high discharge rates and enhances signal encoding efficacy. Additionally, we report anatomic evidence revealing novel cholinergic projections to MNTB arising from pontine and superior olivary nuclei. [ABSTRACT FROM AUTHOR]

Nicotinic acetylcholine receptors (nAChRs) have gained much attention in the scientific community since they play a significant role in multiple physiological and pathophysiological processes. Multiple approaches to study the receptors exist, with characterization of the receptors' functionality at a single cellular level using cell culturing being one of them. Derived from an adrenal medulla tumor, PC12 cells express nicotinic receptor subunits and form functional nicotinic receptors. Thus, the cells offer a convenient environment to address questions related to the functionality of the receptors. The review summarizes the findings on nicotinic receptors' expression and functions which were conducted using PC12 cells. Specific focus is given to α3-containing receptors as well as α7 receptor. Critical evaluation of findings is provided alongside insights into what can still be learned about nAChRs, using PC12 cells. [ABSTRACT FROM AUTHOR]

Smoking is the leading preventable cause of death worldwide and tobacco addiction has become a serious public health problem. Nicotine is the main addictive component of tobacco, and the majority of people that smoke regularly develop nicotine dependence. Nicotine addiction is deemed to be a chronic mental disorder. Although it is well known that nicotine binds to the nicotinic acetylcholine receptors (nAChRs) and activates the mesolimbic dopaminergic system (MDS) to generate the pleasant and rewarding effects, the molecular mechanisms of nicotine addiction are not fully understood. Brain-derived neurotrophic factor (BDNF) is the most prevalent growth factor in the brain, which regulates neuron survival, differentiation, and synaptic plasticity, mainly through binding to the high affinity receptor tyrosine kinase receptor B (TrkB). BDNF gene polymorphisms are associated with nicotine dependence and blood BDNF levels are altered in smokers. In this review, we discussed the effects of nicotine on BDNF expression in the brain and summarized the underlying signaling pathways, which further indicated BDNF as a key regulator in nicotine dependence. Further studies that aim to understand the neurobiological mechanism of BDNF in nicotine addcition would provide a valuable reference for quitting smoking and developing the treatment of other addictive substances. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The use of menthol in tobacco products has been linked to an increased likelihood of developing nicotine dependence. The widespread use of menthol can be attributed to its unique sensory characteristics; however, emerging evidence suggests that menthol also alters sensitivity to nicotine through modulation of nicotinic acetylcholine receptors (nAChRs). Nicotinic subunits, such as β2 and α5, are of interest due to their implications in nicotine reward, reinforcement, intake regulation, and aversion. This study, therefore, examined the in vivo relevance of β2 and α5 nicotinic subunits on the pharmacological and behavioral effects of menthol. Data suggests that the α5 nicotinic subunit modulates menthol intake in mice. Overall, deletion or a reduction in function of the α5 subunit lessened aversion to menthol. α5 KO mice and mice possessing the humanized α5 SNP, a variant that confers a nicotine dependence phenotype in humans, demonstrated increased menthol intake compared to their WT counterparts and in a sex-related fashion for α5 SNP mice. We further reported that the modulatory effects of the α5 subunit do not extend to other aversive tastants like quinine, suggesting that deficits in α5* nAChR signaling may not abolish general sensitivity to the aversive effects of other noxious chemicals. Further probing into the role of α5 in other pharmacological properties of menthol revealed that the α5 subunit does not modulate the antinociceptive properties of menthol in mice and suggests that the in vivo differences observed are likely not due to the direct effects of menthol on α5-containing nAChRs in vitro. • Menthol consumption in mice is modulated by α5 but not β2 nAChR subunits. • Oral consumption of menthol is increased in mice possessing the α5 SNP risk variant. • Deletion of α5 lessens aversion to menthol at higher concentrations in female mice. • α5 nAChRs do not modulate the antinociceptive properties of menthol or oral quinine consumption in mice. [ABSTRACT FROM AUTHOR]

Nicotine, a naturally occurring tobacco alkaloid responsible for tobacco addiction, has long been considered non-carcinogenic. However, emerging evidence suggests that nicotine may possess carcinogenic properties in mice and could be a potential carcinogen in humans. This review aims to summarize the potential molecular mechanisms underlying nicotine-induced carcinogenesis, with a specific focus on epigenetic regulation and the activation of nicotinic acetylcholine receptors (nAChRs) in addition to genotoxicity and excess reactive oxygen species (ROS). Additionally, we explore a novel hypothesis regarding nicotine's carcinogenicity involving the downregulation of stem-loop binding protein (SLBP), a critical regulator of canonical histone mRNA, and the polyadenylation of canonical histone mRNA. By shedding light on these mechanisms, this review underscores the need for further research to elucidate the carcinogenic potential of nicotine and its implications for human health. [Display omitted] • Nicotine is a potential carcinogen in mice and humans. • Epigenetic regulation contributes to nicotine-induced carcinogenesis. • Nicotine downregulates stem-loop binding protein (SLBP). • Nicotine induces polyadenylation of canonical histone mRNAs. [ABSTRACT FROM AUTHOR]

The proteins of the Ly6 family have a three-finger folding as snake venom α-neurotoxins, targeting nicotinic acetylcholine receptors (nAChRs), and some of them, like mammalian secreted Ly6/uPAR protein (SLURP1) and membrane-attached Ly-6/neurotoxin (Lynx1), also interact with distinct nAChR subtypes. We believed that synthetic fragments of these endogenous proteins might open new ways for drug design because nAChRs are well-known targets for developing analgesics and drugs against neurodegenerative diseases. Since interaction with nAChRs was earlier shown for synthetic fragments of the α-neurotoxin central loop II, we synthesized a 15-membered fragment of human Lynx1, its form with two Cys residues added at the N- and C-termini and forming a disulfide, as well as similar forms of human SLURP1, SLURP2, and of Drosophila sleepless protein (SSS). The IC50 values measured in competition with radioiodinated α-bungarotoxin for binding to the membrane-bound Torpedo californica nAChR were 4.9 and 7.4 µM for Lynx1 and SSS fragments, but over 300 µM for SLURP1 or SLURP2 fragments. The affinity of these compounds for the α7 nAChR in the rat pituitary tumor-derived cell line GH4C1 was different: 13.1 and 147 µM for SSS and Lynx1 fragments, respectively. In competition for the ligand-binding domain of the α9 nAChR subunit, SSS and Lynx1 fragments had IC50 values of about 40 µM, which correlates with the value found for the latter with the rat α9α10 nAChR expressed in the Xenopus oocytes. Thus, the activity of these synthetic peptides against muscle-type and α9α10 nAChRs indicates that they may be useful in design of novel myorelaxants and analgesics.

We designed the synthesis of a small library of 3-substituted-3,6-diazabicyclo[3.1.1]heptanes whose affinity on neuronal nicotinic receptors (nAChRs) was evaluated. Among the synthesized compounds, the 5-(3,6-diazabicyclo[3.1.1]heptane-3-yl)- N -(2-fluorophenyl)nicotinamide 43 proved to be the most interesting compound with α 4 β 2 Ki value of 10 pM and a very high α 7 /α 4 β 2 selectivity. Furthermore, compounds 35 , 39 and 43 elicited a selective partial agonist activity for α 4 β 2 nAChR subtype. Finally, in this paper we also report the conclusions on the 3,6-diazabicyclo[3.1.1]heptanes as ligands for nAChRs, resulting from our consolidated structure activity relationship (SAR) studies on this template. Image 1 • Synthesis and characterization of 3,6-diazabicyclo[3.1.1]heptane heteroaryl amides. • α 4 β 2 , α 3 β 4 and α 7 ligands binding studies. • Intrinsic α 4 β 2 receptor mediated activity. [ABSTRACT FROM AUTHOR]

The proteins of the Ly6 family have a three-finger folding as snake venom α-neurotoxins, targeting nicotinic acetylcholine receptors (nAChRs), and some of them, like mammalian secreted Ly6/uPAR protein (SLURP1) and membrane-attached Ly-6/neurotoxin (Lynx1), also interact with distinct nAChR subtypes. We believed that synthetic fragments of these endogenous proteins might open new ways for drug design because nAChRs are well-known targets for developing analgesics and drugs against neurodegenerative diseases. Since interaction with nAChRs was earlier shown for synthetic fragments of the α-neurotoxin central loop II, we synthesized a 15-membered fragment of human Lynx1, its form with two Cys residues added at the N- and C-termini and forming a disulfide, as well as similar forms of human SLURP1, SLURP2, and of Drosophila sleepless protein (SSS). The IC50 values measured in competition with radioiodinated α-bungarotoxin for binding to the membrane-bound Torpedo californica nAChR were 4.9 and 7.4 µM for Lynx1 and SSS fragments, but over 300 µM for SLURP1 or SLURP2 fragments. The affinity of these compounds for the α7 nAChR in the rat pituitary tumor-derived cell line GH4C1 was different: 13.1 and 147 µM for SSS and Lynx1 fragments, respectively. In competition for the ligand-binding domain of the α9 nAChR subunit, SSS and Lynx1 fragments had IC50 values of about 40 µM, which correlates with the value found for the latter with the rat α9α10 nAChR expressed in the Xenopus oocytes. Thus, the activity of these synthetic peptides against muscle-type and α9α10 nAChRs indicates that they may be useful in design of novel myorelaxants and analgesics. [ABSTRACT FROM AUTHOR]

• Nicotine at a dose of 1.5 mg/kg relieves rapamycin-induced pain in the naïve mice. • Nicotine modulates the mTORC1/S6K/IRS1 pathway in ACC of rapamycin-treated mice. • Nicotine corrects dysregulated neuronal excitability in ACC of rapamycin-treated mice. • The above effects can be blocked by the antagonists of α4β2 or α7 nAChRs. Mammalian target of rapamycin complex 1 (mTORC1) inhibitors increase the incidence of pain in patients, and this finding has been replicated in animal models. However, reports on possible analgesics for this condition are scant. Accumulating evidence finds that nicotinic acetylcholine receptors (nAChRs) are involved in mediating pain. However, whether nicotine, a full agonist of nAChRs, alleviates mTORC1 inhibition-induced pain and its underlying mechanisms remain unknown. In this study, pain was induced in naïve male C57BL/6J mice by intraperitoneally injecting rapamycin acutely or repeatedly. Subsequently, pain thresholds, including mechanical and thermal pain, were measured. The involving signaling pathway was tested using western blot analysis and immunofluorescent assay. Changes in neuronal excitability caused by different treatments were also analyzed using whole-cell recording. Microinjection into the anterior cingulate cortex (ACC) was used to test the role of nAChRs containing the α4β2 or α7 subtype in this brain region in pain modulation. Our results showed that nicotine significantly reduced hyperalgesia in mice that received acute or repeated rapamycin injections, and reversed the effects of rapamycin on the phosphorylation of S6K, 4E-BP1, insulin receptor substrate-1 (IRS-1) at Ser636/639, AKT at Ser473, and ERK at Thr202/Tyr204. Whole-cell recording results showed that nicotine reduced the firing rates of pyramidal neurons in the ACC, and a pharmacological blockade of nAChRs containing the α4β2 or α7 subtype in ACC inhibited the antinociceptive effects of nicotine in mice with rapamycin-induced pain. Our findings indicate that analgesics targeting nAChRs can be developed to help patients with rapamycin-induced pain. [ABSTRACT FROM AUTHOR]

Highlights • Nicotine involved in reward via modulation of the mesocorticolimbic system. • Reinforcing effects of nicotine in the mesocorticolimbic pathway caused addiction. • Addictive effects of nicotine caused to improvement of learning and memory. • Nicotine showed dual effects on anxiety behaviors. Abstract Nicotine, as the main component of tobacco, plays a key role in various behaviors of both humans and animals including reward, addiction, learning and memory, anxiety, pain as well as body weight and temperature. Nicotine exerts its function by activating nicotinic acetylcholine receptors (nAChRs) which one highly expressed throughout the nervous system. The mesocorticolimbic dopamine system mediates the processing of reward, learning and memory behaviors produced by nicotine. In addition, some structures of the central nervous system, such as the hippocampus and amygdala, have a major role in mediation of nicotine properties. Furthermore, the effect of nicotine in modulation of cognitive and non-cognitive functions can produce by other neurochemical systems such as opioid, dopamine, glutamate, cannabinoids, GABA and serotonin as well as nitric oxide. The present review will explain involvement of these neurotransmitters on cognitive and non-cognitive functions induced by nicotine. [ABSTRACT FROM AUTHOR]

Diet influences dopamine transmission in motor‐ and reward‐related basal ganglia circuitry. In part, this reflects diet‐dependent regulation of circulating and brain insulin levels. Activation of striatal insulin receptors amplifies axonal dopamine release in brain slices, and regulates food preference in vivo. The effect of insulin on dopamine release is indirect, and requires striatal cholinergic interneurons that express insulin receptors. However, insulin also acts directly on dopamine axons to increase dopamine uptake by promoting dopamine transporter (DAT) surface expression, counteracting enhanced dopamine release. Here, we determined the functional consequences of acute insulin exposure and chronic diet‐induced changes in insulin on DAT activity after evoked dopamine release in striatal slices from adult ad‐libitum fed (AL) rats and mice, and food‐restricted (FR) or high‐fat/high‐sugar obesogenic (OB) diet rats. Uptake kinetics were assessed by fitting evoked dopamine transients to the Michaelis‐Menten equation and extracting Cpeak and Vmax. Insulin (30 nm) increased both parameters in the caudate putamen and nucleus accumbens core of AL rats in an insulin receptor‐ and PI3‐kinase‐dependent manner. A pure effect of insulin on uptake was unmasked using mice lacking striatal acetylcholine, in which increased Vmax caused a decrease in Cpeak. Diet also influenced Vmax, which was lower in FR vs. AL. The effects of insulin on Cpeak and Vmax were amplified by FR but blunted by OB, consistent with opposite consequences of these diets on insulin levels and insulin receptor sensitivity. Overall, these data reveal acute and chronic effects of insulin and diet on dopamine release and uptake that will influence brain reward pathways. Insulin and diet have marked effects on dopamine (DA) transmission in the brain. Insulin increases evoked striatal DA release indirectly via cholinergic interneurons (ChIs) but enhances DA transporter activity and DA uptake directly by activating insulin receptors coupled to the PI3K/Akt pathway. Food restriction (FR) or obesogenic (OB) diets that produce lower or higher plasma insulin respectively decrease DA release and uptake, relative to an ad‐libitum (AL) diet. Moreover, insulin‐induced increases in DA release and uptake are enhanced with FR but blunted with OB diets. [ABSTRACT FROM AUTHOR]

nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that play an important role in the homeostatic regulation of physiological functions. Our previous studies showed that nAChRs in the genome of pearl oyster Pinctada fucata martensii (PmnAChRs) were expanded through tandem duplication. This study aimed to analyze the function of five tandemly duplicated PmnAChRs in the transplantation immunity in P. f. martensii. Transcriptome analysis reveals that the differentially expressed genes (DEGs) shared between PmnAChR-RNAi and the control group were functionally involved in Signal transduction, Immune system et al., and most of the related genes were down-regulated in the PmnAChR-RNAi group. The different copies of PmnAChR may regulate transplantation immunity through various pathways, such as Wnt, protein digestion and absorption, Hippo, and gap junction pathway. The inflammation factor interleukin-17 (IL-17) and tumor necrosis factor-alpha (TNF-α) were down-regulated in PmnAChR-1, 4, 5-RNAi group, and the serum from the pearl oysters in the PmnAChR-1-4-RNAi group could promote the proliferation of the Vibrio harveyi , indicating the immunosuppressive function after down-regulation of PmnAChRs. The different responses of antioxidant enzymes and diverse signal pathways after down-regulation of PmnAChRs suggested that the five tandemly duplicated PmnAChRs may cooperate with different α type PmnAChRs and constitute the functional ion channel in the membrane. Results of this study not only provide insight for the effective regulation of the transplantation immunity, but also provide a theoretical reference for the study of the adaptive evolutionary mechanism of repeating genes. • Down-regulation of PmnAChRs inhibited the expression of immune-regulated genes. • Down-regulation of PmnAChR1,4,5 decreased the activity of IL-17 and TNF-α. • The serum from the PmnAChR1-4-RNAi group activated the growth of Vibrio. • Different copies of PmnAChRs involved in different signal pathways. [ABSTRACT FROM AUTHOR]

The cyanotoxin cylindrospermopsin (CYN) has been postulated to cause neurotoxicity, although the studies in this concern are very few. In addition, some studies in vitro indicate its possible effects on development. Furthermore, pesticides can be present in the same environmental samples as cyanotoxins. Therefore, chlorpyrifos (CPF) has been one of the most common pesticides used worldwide. The aim of this report was to study the effects of CYN, isolated and in combination with CPF, in a developmental neurotoxicity in vitro model. The human neuroblastoma SH-SY5Y cell line was exposed during 6 days of differentiation to both toxics to study their effects on cell viability and neurite outgrowth. To further evaluate effects of both toxicants on cholinergic signaling, their agonistic and antagonistic activities on the α7 homomeric nicotinic acetylcholine receptor (nAChR) were studied upon acute exposure. Moreover, a transcriptomic analysis by qPCR was performed after 6 days of CYN-exposure during differentiation. The results showed a concentration-dependent decrease on both cell viability and neurite outgrowth for both toxics isolated, leading to effective concentration 20 (EC 20) values of 0.35 µM and 0.097 µM for CYN on cell viability and neurite outgrowth, respectively, and 100 µM and 58 µM for CPF, while the combination demonstrated no significant variations. In addition, 95 µM and 285 µM CPF demonstrated to act as an antagonist to nicotine on the nAChR, although CYN up to 2.4 µM had no effect on the efficacy of these receptors. Additionally, the EC 20 for CYN (0.097 µM) on neurite outgrowth downregulated expression of the 5 genes NTNG2 (netrin G2) , KCNJ11 (potassium channel) , SLC18A3 (vesicular acetylcholine transporter) , APOE (apolipoprotein E) , and SEMA6B (semaphorin 6B), that are all important for neuronal development. Thus, this study points out the importance of studying the effects of CYN in terms of neurotoxicity and developmental neurotoxicity. • CYN and CPF attenuated neurite outgrowth in SH-SY5Y cells during 6-days differentiation. • There was no strong interaction between CYN and CPF on neither cell viability, nor neurite outgrowth in our model. • CYN caused a dysregulation of 6 genes involved in neurodevelopment. • CYN demonstrated no effect on α7-nAChRs, while CPF had an antagonistic effect. [ABSTRACT FROM AUTHOR]

Tobacco smoking is particularly evident in individuals experiencing chronic pain. This complex relationship is poorly understood at both molecular and behavioral levels. Here, we describe experiments aimed at understanding whether a chronic pain state induces neuroadaptations into the brain or peripheral nerves that involve nicotinic acetylcholine receptors (nAChRs) and whether these neuroadaptations directly lead to increased vulnerability to nicotine addiction or to the development of coping strategies to relieve pain symptoms. We found that ligation of the rat L5 spinal nerve led to a dramatic downregulation in the mRNA expression levels of all nAChR subunits examined in dorsal root ganglia and a time-dependent downregulation of discrete subunits, particularly in the cingulate cortex and the amygdala. Spinal nerve ligation and sham-operated rats showed minor or no changes in patterns of acquisition and motivation for nicotine taking. Spinal nerve ligation rats also showed similar vulnerability to nicotine seeking as sham animals when reinstatement was induced by nicotine-associated cues, but failed to reinstate lever pressing when relapse was induced by nicotine priming. Spinal nerve ligation and sham rats were equally sensitive to nicotine-induced anxiety-like behavior and antinociception; however, nicotine produced a potent and long-lasting antiallodynic effect in spinal nerve ligation rats. These results demonstrate that chronic pain leads to plasticity of nAChRs that do not directly facilitate nicotine addictive behaviors. Instead, nicotine potently decreases allodynia, an effect that could lead to increased nicotine consumption in chronic pain subjects. [ABSTRACT FROM AUTHOR]

Cholinergic signaling, which modulates cell activities via nicotinic and muscarinic acetylcholine receptors (n- and mAChRs) in response to internal or external stimuli, has been demonstrated in mammalian non-neuronal cells that synthesize acetylcholine (ACh). One of the major pathways of excitatory transmission in the enteric nervous system (ENS) is mediated by cholinergic transmission, with the transmitter ACh producing excitatory potentials in postsynaptic effector cells. In addition to ACh-synthesizing and ACh-metabolizing elements in the ENS, the presence of non-neuronal ACh machinery has been reported in epithelial cells of the small and large intestines of rats and humans. However, little is known about how non-neuronal ACh controls physiological function in the intestine. Here, experiments using crypt-villus organoids that lack nerve and immune cells in culture suggest that endogenous ACh is synthesized in the intestinal epithelium to drive organoid growth and differentiation through activation of nAChRs. Treatment of organoids with nicotine enhanced cell growth and the expression of marker genes for stem and epithelial cells. On the other hand, the nAChR antagonist mecamylamine strongly inhibited the growth and differentiation of organoids, suggesting the involvement of nAChRs in the regulation of proliferation and differentiation of Lgr5-positive stem cells. More specifically, RNA sequencing analysis revealed that Wnt5a expression was dramatically upregulated after nicotine treatment, and Wnt5a rescued organoid growth and differentiation in response to mecamylamine. Taken together, our results indicate that coordinated activities of nAChR and Wnt signaling maintain Lgr5-positive stem cell activity and balanced differentiation. Furthermore, we could clearly separate the two groups, neuronal ACh in the ENS and non-neuronal ACh in the intestinal epithelium. Dysfunction of the non-neuronal cholinergic system is involved in the pathogenesis of disease. The data will increase our understanding of the cholinergic properties of non-neuronal cells and lead to optimization of drug therapy. [ABSTRACT FROM AUTHOR]

A loss-of-function polymorphism in the α5 nicotinic acetylcholine receptor (nAChR) subunit gene has been linked to both drug abuse and schizophrenia. The α5 nAChR subunit is strategically positioned in the prefrontal cortex (PFC), where a loss-of-function in this subunit may contribute to cognitive disruptions in both disorders. However, the specific contribution of α5 to PFC-dependent cognitive functions has yet to be illustrated. In the present studies, we used RNA interference to knockdown the α5 nAChR subunit in the PFC of adult rats. We provide evidence that through its contribution to cholinergic modulation of cholinergic modulation of neurons in the PFC, the α5 nAChR plays a specific role in the recovery of attention task performance following distraction. Our combined data reveal the potent ability of this subunit to modulate the PFC and cognitive functions controlled by this brain region that are impaired in disease. [ABSTRACT FROM AUTHOR]