Alpha9alpha10 knockout mice show altered physiological and behavioral responses to signals in masking noise.

Medial olivocochlear (MOC) efferents modulate outer hair cell motility through specialized nicotinic acetylcholine receptors to support encoding of signals in noise. Transgenic mice lacking the alpha9 subunits of these receptors (α9KOs) have normal hearing in quiet and noise, but lack classic cochlear suppression effects and show abnormal temporal, spectral, and spatial processing. Mice deficient for both the alpha9 and alpha10 receptor subunits (α9α10KOs) may exhibit more severe MOC-related phenotypes. Like α9KOs, α9α10KOs have normal auditory brainstem response (ABR) thresholds and weak MOC reflexes. Here, we further characterized auditory function in α9α10KO mice. Wild-type (WT) and α9α10KO mice had similar ABR thresholds and acoustic startle response amplitudes in quiet and noise, and similar frequency and intensity difference sensitivity. α9α10KO mice had larger ABR Wave I amplitudes than WTs in quiet and noise. Other ABR metrics of hearing-in-noise function yielded conflicting findings regarding α9α10KO susceptibility to masking effects. α9α10KO mice also had larger startle amplitudes in tone backgrounds than WTs. Overall, α9α10KO mice had grossly normal auditory function in quiet and noise, although their larger ABR amplitudes and hyperreactive startles suggest some auditory processing abnormalities. These findings contribute to the growing literature showing mixed effects of MOC dysfunction on hearing.

Single-cell quantitative expression of nicotinic acetylcholine receptor mRNA in rat hippocampal interneurons.

Hippocampal interneurons are a very diverse population of cells. Using single-cell quantitative PCR to analyze rat CA1 hippocampal interneurons, we quantified neuronal nicotinic acetylcholine receptor (nAChR) mRNA subunit expression and detailed possible nAChR subtype combinations for the α2, α3, α4, α5, α7, ß2, ß3, and ß4 subunits. We also compared the expression detected in the stratum oriens and the stratum radiatum hippocampal layers. We show that the majority of interneurons in the CA1 of the rat hippocampus contain detectable levels of nAChR subunit mRNA. Our results highlight the complexity of the CA1 nAChR population. Interestingly, the α3 nAChR subunit is one of the highest expressed subunit mRNAs in this population, while the α4 is one of the least likely subunits to be detected in CA1 interneurons. The ß2 nAChR subunit is the highest expressed beta subunit mRNA in these cells. In addition, Pearson's correlation coefficient values are calculated to identify significant differences between the nAChR subunit combinations expressed in the CA1 stratum oriens and the stratum radiatum. Statistical analysis also indicates that there are likely over 100 different nAChR subunit mRNA combinations expressed in rat CA1 interneurons. These results provide a valid avenue for identifying nAChR subtype targets that may be effective hippocampus-specific pharmacological targets.

The Cloning and Characterization of a Three-Finger Toxin Homolog (NXH8) from the Coralsnake Micrurus corallinus That Interacts with Skeletal Muscle Nicotinic Acetylcholine Receptors.

Coralsnakes (Micrurus spp.) are the only elapids found throughout the Americas. They are recognized for their highly neurotoxic venom, which is comprised of a wide variety of toxins, including the stable, low-mass toxins known as three-finger toxins (3FTx). Due to difficulties in venom extraction and availability, research on coralsnake venoms is still very limited when compared to that of other Elapidae snakes like cobras, kraits, and mambas. In this study, two previously described 3FTx from the venom of M. corallinus, NXH1 (3SOC1_MICCO), and NXH8 (3NO48_MICCO) were characterized. Using in silico, in vitro, and ex vivo experiments, the biological activities of these toxins were predicted and evaluated. The results showed that only NXH8 was capable of binding to skeletal muscle cells and modulating the activity of nAChRs in nerve-diaphragm preparations. These effects were antagonized by anti-rNXH8 or antielapidic sera. Sequence analysis revealed that the NXH1 toxin possesses eight cysteine residues and four disulfide bonds, while the NXH8 toxin has a primary structure similar to that of non-conventional 3FTx, with an additional disulfide bond on the first loop. These findings add more information related to the structural diversity present within the 3FTx class, while expanding our understanding of the mechanisms of the toxicity of this coralsnake venom and opening new perspectives for developing more effective therapeutic interventions.

High-Voltage Toxin'Roll: Electrostatic Charge Repulsion as a Dynamic Venom Resistance Trait in Pythonid Snakes.

The evolutionary interplay between predator and prey has significantly shaped the development of snake venom, a critical adaptation for subduing prey. This arms race has spurred the diversification of the components of venom and the corresponding emergence of resistance mechanisms in the prey and predators of venomous snakes. Our study investigates the molecular basis of venom resistance in pythons, focusing on electrostatic charge repulsion as a defense against α-neurotoxins binding to the alpha-1 subunit of the postsynaptic nicotinic acetylcholine receptor. Through phylogenetic and bioactivity analyses of orthosteric site sequences from various python species, we explore the prevalence and evolution of amino acid substitutions that confer resistance by electrostatic repulsion, which initially evolved in response to predatory pressure by Naja (cobra) species (which occurs across Africa and Asia). The small African species Python regius retains the two resistance-conferring lysines (positions 189 and 191) of the ancestral Python genus, conferring resistance to sympatric Naja venoms. This differed from the giant African species Python sebae, which has secondarily lost one of these lysines, potentially due to its rapid growth out of the prey size range of sympatric Naja species. In contrast, the two Asian species Python brongersmai (small) and Python bivittatus (giant) share an identical orthosteric site, which exhibits the highest degree of resistance, attributed to three lysine residues in the orthosteric sites. One of these lysines (at orthosteric position 195) evolved in the last common ancestor of these two species, which may reflect an adaptive response to increased predation pressures from the sympatric α-neurotoxic snake-eating genus Ophiophagus (King Cobras) in Asia. All these terrestrial Python species, however, were less neurotoxin-susceptible than pythons in other genera which have evolved under different predatory pressure as: the Asian species Malayopython reticulatus which is arboreal as neonates and juveniles before rapidly reaching sizes as terrestrial adults too large for sympatric Ophiophagus species to consider as prey; and the terrestrial Australian species Aspidites melanocephalus which occupies a niche, devoid of selection pressure from α-neurotoxic predatory snakes. Our findings underline the importance of positive selection in the evolution of venom resistance and suggest a complex evolutionary history involving both conserved traits and secondary evolution. This study enhances our understanding of the molecular adaptations that enable pythons to survive in environments laden with venomous threats and offers insights into the ongoing co-evolution between venomous snakes and their prey.

Dopamine and Norepinephrine Tissue Levels in the Developing Limbic Brain Are Impacted by the Human CHRNA6 3'-UTR Single-Nucleotide Polymorphism (rs2304297) in Rats.

We previously demonstrated that a genetic single-nucleotide polymorphism (SNP, rs2304297) in the 3' untranslated region (UTR) of the human CHRNA6 gene has sex- and genotype-dependent effects on nicotine-induced locomotion, anxiety, and nicotine + cue-induced reinstatement in adolescent rats. This study aims to investigate how the CHRNA6 3'-UTR SNP influences dopaminergic and noradrenergic tissue levels in brain reward regions during baseline and after the reinstatement of drug-seeking behavior. Naïve adolescent and adult rats, along with those undergoing nicotine + cue reinstatement and carrying the CHRNA6 3'-UTR SNP, were assessed for dopamine (DA), norepinephrine (NE), and metabolites in reward pathway regions. The results reveal age-, sex-, and genotype-dependent baseline DA, NE, and DA turnover levels. Post-reinstatement, male α6GG rats show suppressed DA levels in the Nucleus Accumbens (NAc) Shell compared to the baseline, while nicotine+ cue-induced reinstatement behavior correlates with neurotransmitter levels in specific brain regions. This study emphasizes the role of CHRNA6 3'-UTR SNP in the developmental maturation of the dopaminergic and noradrenergic system in the adolescent rat brain, with tissue levels acting as predictors of nicotine + cue-induced reinstatement.

Vagus nerve stimulation modulates distinct acetylcholine receptors on B cells and limits the germinal center response.

Acetylcholine is produced in the spleen in response to vagus nerve activation; however, the effects on antibody production have been largely unexplored. Here, we use a chronic vagus nerve stimulation (VNS) mouse model to study the effect of VNS on T-dependent B cell responses. We observed lower titers of high-affinity IgG and fewer antigen-specific germinal center (GC) B cells. GC B cells from chronic VNS mice exhibited altered mRNA and protein expression suggesting increased apoptosis and impaired plasma cell differentiation. Follicular dendritic cell (FDC) cluster dispersal and altered gene expression suggested poor function. The absence of acetylcholine-producing CD4+ T cells diminished these alterations. In vitro studies revealed that α7 and α9 nicotinic acetylcholine receptors (nAChRs) directly regulated B cell production of TNF, a cytokine crucial to FDC clustering. α4 nAChR inhibited coligation of CD19 to the B cell receptor, presumably decreasing B cell survival. Thus, VNS-induced GC impairment can be attributed to distinct effects of nAChRs on B cells.

Ion transport in muscle acetylcholine receptor maintained by conserved salt bridges between the pore and lipid membrane.

Pores through ion channels rapidly transport small inorganic ions along their electrochemical gradients. Here, applying single-channel electrophysiology and mutagenesis to the archetypal muscle nicotinic acetylcholine receptor (AChR) channel, we show that a conserved pore-peripheral salt bridge partners with those in the other subunits to regulate ion transport. Disrupting the salt bridges in all five receptor subunits greatly decreases the amplitude of the unitary current and increases its fluctuations. However, disrupting individual salt bridges has unequal effects that depend on the structural status of the other salt bridges. The AChR ε- and δ-subunits are structurally unique in harboring a putative palmitoylation site near each salt bridge and bordering the lipid membrane. The effects of disrupting the palmitoylation sites mirror those of disrupting the salt bridges, but the effect of disrupting either of these structures depends on the structural status of the other. Thus, rapid ion transport through the AChR channel is maintained by functionally interdependent salt bridges linking the pore to the lipid membrane.

Nicotinic Acetylcholine Receptor Alpha6 Contributes to Antiviral Immunity via IMD Pathway in Drosophila melanogaster.

Currently, insecticides that target nicotinic acetylcholine receptors (nAChR) are widely used. Studies on the sublethal effects of insecticides have found that they can affect the amount of virus in insects. The mechanism by which insecticides affect insect virus load remain unclear. Here, we show that nAChR targeting insecticide can affect viral replication through the immune deficiency (IMD) pathway. We demonstrate that a low dose of spinosad (6.8 ng/mL), acting as an antagonist to Drosophila melanogaster nicotinic acetylcholine receptor α6 (Dα6), significantly elevates Drosophila melanogaster sigmavirus (DMelSV) virus titers in adults of Drosophila melanogaster. Conversely, a high dose of spinosad (50 ng/mL), acting as an agonist to Dα6, substantially decreases viral load. This bidirectional regulation of virus levels is absent in Dα6-knockout flies, signifying the specificity of spinosad's action through Dα6. Furthermore, the knockdown of Dα6 results in decreased expression of genes in the IMD pathway, including dredd, imd, relish, and downstream antimicrobial peptide genes AttA and AttB, indicating a reduced innate immune response. Subsequent investigations reveal no significant difference in viral titers between relish mutant flies and Dα6-relish double mutants, suggesting that the IMD pathway's role in antiviral defense is dependent on Dα6. Collectively, our findings shed light on the intricate interplay between nAChR signaling and the IMD pathway in mediating antiviral immunity, highlighting the potential for nAChR-targeting compounds to inadvertently influence viral dynamics in insect hosts. This knowledge may inform the development of integrated pest management strategies that consider the broader ecological impact of insecticide use.

Aberrant splicing of a nicotinic acetylcholine receptor alpha 6 subunit is associated with spinosad tolerance in the thrips predator Orius laevigatus.

Susceptibility to insecticides is one of the limiting factors preventing wider adoption of natural enemies to control insect pest populations. Identification and selective breeding of insecticide tolerant strains of commercially used biological control agents (BCAs) is one of the approaches to overcome this constraint. Although a number of beneficial insects have been selected for increased tolerance to insecticides the molecular mechanisms underpinning these shifts in tolerance are not well characterised. Here we investigated the molecular mechanisms of enhanced tolerance of a lab selected strain of Orius laevigatus (Fieber) to the commonly used biopesticide spinosad. Transcriptomic analysis showed that spinosad tolerance is not a result of overexpressed detoxification genes. Molecular analysis of the target site for spinosyns, the nicotinic acetylcholine receptor (nAChR), revealed increased expression of truncated transcripts of the nAChR α6 subunit in the spinosad selected strain, a mechanism of resistance which was described previously in insect pest species. Collectively, our results demonstrate the mechanisms by which some beneficial biological control agents can evolve insecticide tolerance and will inform the development and deployment of insecticide-tolerant natural enemies in integrated pest management strategies.

Investigating the role of nicotinic acetylcholine receptors in menthol's effects in mice.

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 ß2 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 ß2 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.

Mutation V65I in the ß1 Subunit of the Nicotinic Acetylcholine Receptor Confers Neonicotinoid and Sulfoxaflor Resistance in Insects.

Neonicotinoids have been widely used to control pests with remarkable effectiveness. Excessive insecticides have led to serious insect resistance. Mutations of the nicotinic acetylcholine receptor (nAChR) are one of the reasons for neonicotinoid resistance conferred in various agricultural pests. Two mutations, V65I and V104I, were found in the nAChR ß1 subunit of two neonicotinoid-resistant aphid populations. However, the specific functions of the two mutations remain unclear. In this study, we cloned and identified four nAChR subunits (α1, α2, α8, and ß1) of thrips and found them to be highly homologous to the nAChR subunits of other insects. Subsequently, we successfully expressed two subtypes nAChR (α1/α2/α8/ß1 and α1/α8/ß1) by coinjecting three cofactors for the first time in thrips, and α1/α8/ß1 showed abundant current rapidly. Acetylcholine, neonicotinoids, and sulfoxaflor exhibited different activation capacities for the two subtypes of nAChRs. Finally, V65I was found to significantly reduce the binding ability of nAChR to neonicotinoids and sulfoxaflor through electrophysiology and computer simulations. V104I caused a decrease in agonist affinity (pEC50) but an increase in the efficacy (Imax) of nAChR against neonicotinoids and reduced the binding ability of nAChR to sulfoxaflor. This study provides theoretical and technical support for studying the molecular mechanisms of neonicotinoid resistance in pests.

Unraveling the molecular interactions between α7 nicotinic receptor and a RIC3 variant associated with backward speech.

Recent work putatively linked a rare genetic variant of the chaperone Resistant to Inhibitors of acetylcholinesterase (RIC3) (NM_024557.4:c.262G > A, NP_078833.3:p.G88R) to a unique ability to speak backwards, a language skill that is associated with exceptional working memory capacity. RIC3 is important for the folding, maturation, and functional expression of α7 nicotinic acetylcholine receptors (nAChR). We compared and contrasted the effects of RIC3G88R on assembly, cell surface expression, and function of human α7 receptors using fluorescent protein tagged α7 nAChR and Förster resonance energy transfer (FRET) microscopy imaging in combination with functional assays and 125I-α-bungarotoxin binding. As expected, the wild-type RIC3 protein was found to increase both cell surface and functional expression of α7 receptors. In contrast, the variant form of RIC3 decreased both. FRET analysis showed that RICG88R increased the interactions between RIC3 and α7 protein in the endoplasmic reticulum. These results provide interesting and novel data to show that a RIC3 variant alters the interaction of RIC3 and α7, which translates to decreased cell surface and functional expression of α7 nAChR.

Subtype-Selective Peptide and Protein Neurotoxic Inhibitors of Nicotinic Acetylcholine Receptors Enhance Proliferation of Patient-Derived Glioblastoma Cell Lines.

Glioblastoma multiforme (GBM) is the most aggressive type of brain cancer, with a poor prognosis. GBM cells, which develop in the environment of neural tissue, often exploit neurotransmitters and their receptors to promote their own growth and invasion. Nicotinic acetylcholine receptors (nAChRs), which play a crucial role in central nervous system signal transmission, are widely represented in the brain, and GBM cells express several subtypes of nAChRs that are suggested to transmit signals from neurons, promoting tumor invasion and growth. Analysis of published GBM transcriptomes revealed spatial heterogeneity in nAChR subtype expression, and functional nAChRs of α1*, α7, and α9 subtypes are demonstrated in our work on several patient-derived GBM microsphere cultures and on the U87MG GBM cell line using subtype-selective neurotoxins and fluorescent calcium mobilization assay. The U87MG cell line shows reactions to nicotinic agonists similar to those of GBM patient-derived culture. Selective α1*, α7, and α9 nAChR neurotoxins stimulated cell growth in the presence of nicotinic agonists. Several cultivating conditions with varying growth factor content have been proposed and tested. The use of selective neurotoxins confirmed that cell cultures obtained from patients are representative GBM models, but the use of media containing fetal bovine serum can lead to alterations in nAChR expression and functioning.

Dual mutations in the whitefly nicotinic acetylcholine receptor ß1 subunit confer target-site resistance to multiple neonicotinoid insecticides.

Neonicotinoid insecticides, which target insect nicotinic acetylcholine receptors (nAChRs), have been widely and intensively used to control the whitefly, Bemisia tabaci, a highly damaging, globally distributed, crop pest. This has inevitably led to the emergence of populations with resistance to neonicotinoids. However, to date, there have been no reports of target-site resistance involving mutation of B. tabaci nAChR genes. Here we characterize the nAChR subunit gene family of B. tabaci and identify dual mutations (A58T&R79E) in one of these genes (BTß1) that confer resistance to multiple neonicotinoids. Transgenic D. melanogaster, where the native nAChR Dß1 was replaced with BTß1A58T&R79E, were significantly more resistant to neonicotinoids than flies where Dß1 were replaced with the wildtype BTß1 sequence, demonstrating the causal role of the mutations in resistance. The two mutations identified in this study replace two amino acids that are highly conserved in >200 insect species. Three-dimensional modelling suggests a molecular mechanism for this resistance, whereby A58T forms a hydrogen bond with the R79E side chain, which positions its negatively-charged carboxylate group to electrostatically repulse a neonicotinoid at the orthosteric site. Together these findings describe the first case of target-site resistance to neonicotinoids in B. tabaci and provide insight into the molecular determinants of neonicotinoid binding and selectivity.

A release of local subunit conformational heterogeneity underlies gating in a muscle nicotinic acetylcholine receptor.

Synaptic receptors respond to neurotransmitters by opening an ion channel across the post-synaptic membrane to elicit a cellular response. Here we use recent Torpedo acetylcholine receptor structures and functional measurements to delineate a key feature underlying allosteric communication between the agonist-binding extracellular and channel-gating transmembrane domains. Extensive mutagenesis at this inter-domain interface re-affirms a critical energetically coupled role for the principal α subunit ß1-ß2 and M2-M3 loops, with agonist binding re-positioning a key ß1-ß2 glutamate/valine to facilitate the outward motions of a conserved M2-M3 proline to open the channel gate. Notably, the analogous structures in non-α subunits adopt a locally active-like conformation in the apo state even though each L9' hydrophobic gate residue in each pore-lining M2 α-helix is closed. Agonist binding releases local conformational heterogeneity transitioning all five subunits into a conformationally symmetric open state. A release of conformational heterogeneity provides a framework for understanding allosteric communication in pentameric ligand-gated ion channels.

CHRNA5 links chandelier cells to severity of amyloid pathology in aging and Alzheimer's disease.

Changes in high-affinity nicotinic acetylcholine receptors are intricately connected to neuropathology in Alzheimer's Disease (AD). Protective and cognitive-enhancing roles for the nicotinic α5 subunit have been identified, but this gene has not been closely examined in the context of human aging and dementia. Therefore, we investigate the nicotinic α5 gene CHRNA5 and the impact of relevant single nucleotide polymorphisms (SNPs) in prefrontal cortex from 922 individuals with matched genotypic and post-mortem RNA sequencing in the Religious Orders Study and Memory and Aging Project (ROS/MAP). We find that a genotype robustly linked to increased expression of CHRNA5 (rs1979905A2) predicts significantly reduced cortical ß-amyloid load. Intriguingly, co-expression analysis suggests CHRNA5 has a distinct cellular expression profile compared to other nicotinic receptor genes. Consistent with this prediction, single nucleus RNA sequencing from 22 individuals reveals CHRNA5 expression is disproportionately elevated in chandelier neurons, a distinct subtype of inhibitory neuron known for its role in excitatory/inhibitory (E/I) balance. We show that chandelier neurons are enriched in amyloid-binding proteins compared to basket cells, the other major subtype of PVALB-positive interneurons. Consistent with the hypothesis that nicotinic receptors in chandelier cells normally protect against ß-amyloid, cell-type proportion analysis from 549 individuals reveals these neurons show amyloid-associated vulnerability only in individuals with impaired function/trafficking of nicotinic α5-containing receptors due to homozygosity of the missense CHRNA5 SNP (rs16969968A2). Taken together, these findings suggest that CHRNA5 and its nicotinic α5 subunit exert a neuroprotective role in aging and Alzheimer's disease centered on chandelier interneurons.

Nicotine Motivated Behavior in C. elegans.

To maximize the advantages offered by Caenorhabditis elegans as a high-throughput (HTP) model for nicotine dependence studies, utilizing its well-defined neuroconnectome as a robust platform, and to unravel the genetic basis of nicotine-motivated behaviors, we established the nicotine conditioned cue preference (CCP) paradigm. Nicotine CCP enables the assessment of nicotine preference and seeking, revealing a parallel to fundamental aspects of nicotine-dependent behaviors observed in mammals. We demonstrated that nicotine-elicited cue preference in worms is mediated by nicotinic acetylcholine receptors and requires dopamine for CCP development. Subsequently, we pinpointed nAChR subunits associated with nicotine preference and validated human GWAS candidates linked to nicotine dependence involved in nAChRs. Functional validation involves assessing the loss-of-function strain of the CACNA2D3 ortholog and the knock-out (KO) strain of the CACNA2D2 ortholog, closely related to CACNA2D3 and sharing human smoking phenotypes. Our orthogonal approach substantiates the functional conservation of the α2δ subunit of the calcium channel in nicotine-motivated behavior. Nicotine CCP in C. elegans serves as a potent affirmation of the cross-species functional relevance of GWAS candidate genes involved in nicotine seeking associated with tobacco abuse, providing a streamlined yet comprehensive system for investigating intricate behavioral paradigms within a simplified and reliable framework.

Novel CHRNA3 variants identified in a patient with bladder dysfunction, dysautonomia, and gastrointestinal dysmotility.

Congenital anomalies of the kidney and urinary tract (CAKUT) are estimated to be responsible for 20%-50% of congenital anomalies and are also a leading etiology of early-onset renal disease. Primary CAKUT are caused by genetic factors that impair proper in-utero genitourinary tract development and secondary CAKUT result from the influence of environmental factors. The CHRNA3 gene, which encodes the Alpha-3 subunit of the nicotinic acetylcholine receptor, is hypothesized to be associated with Megacystis-microcolon-intestinal hyperperistalsis syndrome. More recently, pathogenic variants in CHRNA3 have been identified in individuals with CAKUT as well as individuals with panautonomic failure. Here we present a patient with neurogenic bladder, vesicoureteral reflux, mydriasis, and gastrointestinal dysmotility found to have novel compound heterozygous variants in CHRNA3. These findings support the consideration of CHRNA3 disruption in the differential for CAKUT with dysautonomia and gastrointestinal dysmotility.

Cell signaling and epigenetic regulation of nicotine-induced carcinogenesis.

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.

Acetylcholine receptor based chemogenetics engineered for neuronal inhibition and seizure control assessed in mice.

Epilepsy is a prevalent disorder involving neuronal network hyperexcitability, yet existing therapeutic strategies often fail to provide optimal patient outcomes. Chemogenetic approaches, where exogenous receptors are expressed in defined brain areas and specifically activated by selective agonists, are appealing methods to constrain overactive neuronal activity. We developed BARNI (Bradanicline- and Acetylcholine-activated Receptor for Neuronal Inhibition), an engineered channel comprised of the α7 nicotinic acetylcholine receptor ligand-binding domain coupled to an α1 glycine receptor anion pore domain. Here we demonstrate that BARNI activation by the clinical stage α7 nicotinic acetylcholine receptor-selective agonist bradanicline effectively suppressed targeted neuronal activity, and controlled both acute and chronic seizures in male mice. Our results provide evidence for the use of an inhibitory acetylcholine-based engineered channel activatable by both exogenous and endogenous agonists as a potential therapeutic approach to treating epilepsy.