Attenuation of Nicotine Effects on A549 Lung Cancer Cells by Synthetic α7 nAChR Antagonists APS7-2 and APS8-2.

Nicotine binds to nicotinic acetylcholine receptors (nAChRs) that are overexpressed in different cancer cells, promoting tumor growth and resistance to chemotherapy. In this study, we aimed to investigate the potential of APS7-2 and APS8-2, synthetic analogs of a marine sponge toxin, to inhibit nicotine-mediated effects on A549 human lung cancer cells. Our electrophysiological measurements confirmed that APS7-2 and APS8-2 act as α7 nAChR antagonists. APS8-2 showed no cytotoxicity in A549 cells, while APS7-2 showed concentration-dependent cytotoxicity in A549 cells. The different cytotoxic responses of APS7-2 and APS8-2 emphasize the importance of the chemical structure in determining their cytotoxicity on cancer cells. Nicotine-mediated effects include increased cell viability and proliferation, elevated intracellular calcium levels, and reduced cisplatin-induced cytotoxicity and reactive oxygen species production (ROS) in A549 cells. These effects of nicotine were effectively attenuated by APS8-2, whereas APS7-2 was less effective. Our results suggest that APS8-2 is a promising new therapeutic agent in the chemotherapy of lung cancer.

The Cyclic Imine Core Common to the Marine Macrocyclic Toxins Is Sufficient to Dictate Nicotinic Acetylcholine Receptor Antagonism.

Macrocyclic imine phycotoxins are an emerging class of chemical compounds associated with harmful algal blooms and shellfish toxicity. Earlier binding and electrophysiology experiments on nAChR subtypes and their soluble AChBP surrogates evidenced common trends for substantial antagonism, binding affinities, and receptor-subtype selectivity. Earlier, complementary crystal structures of AChBP complexes showed that common determinants within the binding nest at each subunit interface confer high-affinity toxin binding, while distinctive determinants from the flexible loop C, and either capping the nest or extending toward peripheral subsites, dictate broad versus narrow receptor subtype selectivity. From these data, small spiroimine enantiomers mimicking the functional core motif of phycotoxins were chemically synthesized and characterized. Voltage-clamp analyses involving three nAChR subtypes revealed preserved antagonism for both enantiomers, despite lower subtype specificity and binding affinities associated with faster reversibility compared with their macrocyclic relatives. Binding and structural analyses involving two AChBPs pointed to modest affinities and positional variability of the spiroimines, along with a range of AChBP loop-C conformations denoting a prevalence of antagonistic properties. These data highlight the major contribution of the spiroimine core to binding within the nAChR nest and confirm the need for an extended interaction network as established by the macrocyclic toxins to define high affinities and marked subtype specificity. This study identifies a minimal set of functional pharmacophores and binding determinants as templates for designing new antagonists targeting disease-associated nAChR subtypes.

Catharanthine Modulates Mesolimbic Dopamine Transmission and Nicotine Psychomotor Effects via Inhibition of α6-Nicotinic Receptors and Dopamine Transporters.

Iboga alkaloids, also known as coronaridine congeners, have shown promise in the treatment of alcohol and opioid use disorders. The objective of this study was to evaluate the effects of catharanthine and 18-methoxycoronaridine (18-MC) on dopamine (DA) transmission and cholinergic interneurons in the mesolimbic DA system, nicotine-induced locomotor activity, and nicotine-taking behavior. Utilizing ex vivo fast-scan cyclic voltammetry (FSCV) in the nucleus accumbens core of male mice, we found that catharanthine or 18-MC differentially inhibited evoked DA release. Catharanthine inhibition of evoked DA release was significantly reduced by both α4 and α6 nicotinic acetylcholine receptors (nAChRs) antagonists. Additionally, catharanthine substantially increased DA release more than vehicle during high-frequency stimulation, although less potently than an α4 nAChR antagonist, which confirms previous work with nAChR antagonists. Interestingly, while catharanthine slowed DA reuptake measured via FSCV ex vivo, it also increased extracellular DA in striatal dialysate from anesthetized mice in vivo in a dose-dependent manner. Superfusion of catharanthine or 18-MC inhibited the firing rate of striatal cholinergic interneurons in a concentration dependent manner, which are known to potently modulate presynaptic DA release. Catharanthine or 18-MC suppressed acetylcholine currents in oocytes expressing recombinant rat α6/α3ß2ß3 or α6/α3ß4 nAChRs. In behavioral experiments using male Sprague-Dawley rats, systemic administration of catharanthine or 18-MC blocked nicotine enhancement of locomotor activity. Importantly, catharanthine attenuated nicotine self-administration in a dose-dependent manner while having no effect on food reinforcement. Lastly, administration of catharanthine and nicotine together greatly increased head twitch responses, indicating a potential synergistic hallucinogenic effect. These findings demonstrate that catharanthine and 18-MC have similar, but not identical effects on striatal DA dynamics, striatal cholinergic interneuron activity and nicotine psychomotor effects.

Unlocking the role of dorsal hippocampal α4ß2 nicotinic acetylcholine receptors in Ethanol-Induced conditioned place preference in mice.

Alcohol Use Disorder (AUD) presents a significant and challenging public health concern, marked by a dearth of effective pharmacological treatments. Understanding the neurobiological underpinnings of AUD is of paramount importance for the development of efficacious interventions. The process of addiction entails the acquisition of associative behaviors, prominently engaging the dorsal region of the hippocampus for encoding these associative memories. Nicotinic receptor systems have been implicated in mediating the rewarding effects of ethanol, as well as memory and learning processes. In our current investigation, we delved into the role of α4ß2 nicotinic acetylcholine receptors (nAChRs) within the dorsal hippocampus in the context of ethanol-induced conditioned place preference (CPP), a robust model for scrutinizing the rewarding properties and drug-associated behaviors. To establish CPP, ethanol (2 g/kg) was administered intraperitoneally during a 8-day conditioning phase. Fos immunohistochemistry was employed to assess the involvement of discrete subregions within the dorsal hippocampus in ethanol-induced CPP. Additionally, we probed the influence of α4ß2 nAChRs on CPP via microinjections of a selective nAChR antagonist, dihydro-ß-erythroidine (DHBE, at dosages of 6, 12, and 18 µg/0.5 µL per hemisphere) within the hippocampus. Our results unveiled that ethanol-induced CPP was associated with an increase Fos -positive cells in various subregions of the dorsal hippocampus, including CA1, CA2, CA3, and the dentate gyrus. Intrahippocampal administration of DHBE (at doses of 6 and 18 µg/0.50 µL per hemisphere) effectively blocked ethanol-induced CPP, while leaving locomotor activity unaffected. These findings underscore the critical involvement of the dorsal hippocampus and α4ß2 nAChRs in the acquisition of ethanol-associated learning and reward.

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.

Behavioral characterization of early nicotine withdrawal in the mouse: a potential model of acute dependence.

BACKGROUND: Clinical and preclinical research have demonstrated that short-term exposure to nicotine during the initial experimentation stage can lead to early manifestation of withdrawal-like signs, indicating the state of "acute dependence". As drug withdrawal is a major factor driving the progression toward regular drug intake, characterizing and understanding the features of early nicotine withdrawal may be important for the prevention and treatment of drug addiction. In this study, we corroborate the previous studies by showing that withdrawal-like signs can be precipitated after short-term nicotine exposure in mice, providing a potential animal model of acute dependence on nicotine. RESULTS: To model nicotine exposure from light tobacco use during the initial experimentation stage, mice were treated with 0.5 mg/kg (-)-nicotine ditartrate once daily for 3 days. On the following day, the behavioral tests were conducted after implementing spontaneous or mecamylamine-precipitated withdrawal. In the open field test, precipitated nicotine withdrawal reduced locomotor activity and time spent in the center zone. In the elevated plus maze test, the mecamylamine challenge increased the time spent in the closed arm and reduced the number of entries irrespective of nicotine experience. In the examination of the somatic aspect, precipitated nicotine withdrawal enhanced the number of somatic signs. Finally, nicotine withdrawal did not affect cognitive functioning or social behavior in the passive avoidance, spatial object recognition, or social interaction test. CONCLUSIONS: Collectively, our data demonstrate that early nicotine withdrawal-like signs could be precipitated by the nicotinic antagonist mecamylamine in mice, and that early withdrawal from nicotine primarily causes physical symptoms.

Dual Antagonism of α9α10 nAChR and GABAB Receptor-Coupled CaV2.2 Channels by an Analgesic αO-Conotoxin Analogue.

Pain severely affects the physical and mental health of patients. The need to develop nonopioid analgesic drugs to meet medical demands is urgent. In this study, we designed a truncated analogue of αO-conotoxin, named GeX-2, based on disulfide-bond deletion and sequence truncation. GeX-2 retained the potency of its parent peptide at the human α9α10 nAChR and exhibited potent inhibitory activity at CaV2.2 channels via activation of the GABAB receptor (GABABR). Importantly, GeX-2 significantly alleviated pain in the rat model of chronic constriction injury. The dual inhibition of GeX-2 at both α9α10 nAChRs and CaV2.2 channels is speculated to synergistically mediate the potent analgesic effects. Results from site-directed mutagenesis assay and computational modeling suggest that GeX-2 preferentially interacts with the α10(+)α10(-) binding site of α9α10 nAChR and favorably binds to the top region of the GABABR2 subunit. The study offers vital insights into the molecular action mechanism of GeX-2, demonstrating its potential as a novel nonopioid analgesic.

Analogs of α-conotoxin PnIC selectively inhibit α7ß2- over α7-only subtype nicotinic acetylcholine receptors via a novel allosteric mechanism.

This study was undertaken to identify and characterize the first ligands capable of selectively identifying nicotinic acetylcholine receptors containing α7 and ß2 subunits (α7ß2-nAChR subtype). Basal forebrain cholinergic neurons express α7ß2-nAChR. Here, they appear to mediate neuronal dysfunction induced by the elevated levels of oligomeric amyloid-ß associated with early Alzheimer's disease. Additional work indicates that α7ß2-nAChR are expressed across several further critically important cholinergic and GABAergic neuronal circuits within the central nervous system. Further studies, however, are significantly hindered by the inability of currently available ligands to distinguish heteromeric α7ß2-nAChR from the closely related and more widespread homomeric α7-only-nAChR subtype. Functional screening using two-electrode voltage-clamp electrophysiology identified a family of α7ß2-nAChR-selective analogs of α-conotoxin PnIC (α-CtxPnIC). A combined electrophysiology, functional kinetics, site-directed mutagenesis, and molecular dynamics approach was used to further characterize the α7ß2-nAChR selectivity and site of action of these α-CtxPnIC analogs. We determined that α7ß2-nAChR selectivity of α-CtxPnIC analogs arises from interactions at a site distinct from the orthosteric agonist-binding site shared between α7ß2- and α7-only-nAChR. As numerous previously identified α-Ctx ligands are competitive antagonists of orthosteric agonist-binding sites, this study profoundly expands the scope of use of α-Ctx ligands (which have already provided important nAChR research and translational breakthroughs). More immediately, analogs of α-CtxPnIC promise to enable, for the first time, both comprehensive mapping of the distribution of α7ß2-nAChR and detailed investigations of their physiological roles.

αO-Conotoxin GeXIVA[1,2] Reduced Neuropathic Pain and Changed Gene Expression in Chronic Oxaliplatin-Induced Neuropathy Mice Model.

Chemotherapy-induced peripheral neuropathy (CIPN) is a dose-limiting painful neuropathy that occurs commonly during cancer management, which often leads to the discontinuation of medication. Previous studies suggest that the α9α10 nicotinic acetylcholine receptor (nAChR)-specific antagonist αO-conotoxin GeXIVA[1,2] is effective in CIPN models; however, the related mechanisms remain unclear. Here, we analyzed the preventive effect of GeXIVA[1,2] on neuropathic pain in the long-term oxaliplatin injection-induced CIPN model. At the end of treatment, lumbar (L4-L6) spinal cord was extracted, and RNA sequencing and bioinformatic analysis were performed to investigate the potential genes and pathways related to CIPN and GeXIVA[1,2]. GeXIVA[1,2] inhibited the development of mechanical allodynia induced by chronic oxaliplatin treatment. Repeated injections of GeXIVA[1,2] for 3 weeks had no effect on the mice's normal pain threshold or locomotor activity and anxiety-like behavior, as evaluated in the open field test (OFT) and elevated plus maze (EPM). Our RNA sequencing results identified 209 differentially expressed genes (DEGs) in the CIPN model, and simultaneously injecting GeXIVA[1,2] with oxaliplatin altered 53 of the identified DEGs. These reverted genes were significantly enriched in immune-related pathways represented by the cytokine-cytokine receptor interaction pathway. Our findings suggest that GeXIVA[1,2] could be a potential therapeutic compound for chronic oxaliplatin-induced CIPN management.

Synthesis of α3ß4 Nicotinic Acetylcholine Receptor Modulators Derived from Aristoquinoline That Reduce Reinstatement of Cocaine-Seeking Behavior.

Growing evidence suggests that inhibition of the α3ß4 nicotinic acetylcholine receptor (nAChR) represents a promising therapeutic strategy to treat cocaine use disorder. Recently, aristoquinoline (1), an alkaloid from Aristotelia chilensis, was identified as an α3ß4-selective nAChR inhibitor. Here, we prepared 22 derivatives of 1 and evaluated their ability to inhibit the α3ß4 nAChR. These studies revealed structure-activity trends and several compounds with increased potency compared to 1 with few off-target liabilities. Additional mechanistic studies indicated that these compounds inhibit the α3ß4 nAChR noncompetitively, but do not act as channel blockers, suggesting they are negative allosteric modulators. Finally, using a cocaine-primed reinstatement paradigm, we demonstrated that 1 significantly attenuates drug-seeking behavior in an animal model of cocaine relapse. The results from these studies further support a role for the α3ß4 nAChR in the addictive properties of cocaine and highlight the possible utility of aristoquinoline derivatives in treating cocaine use disorder.

Basic Residues at Position 11 of α-Conotoxin LvIA Influence Subtype Selectivity between α3ß2 and α3ß4 Nicotinic Receptors via an Electrostatic Mechanism.

Understanding the determinants of α-conotoxin (α-CTX) selectivity for different nicotinic acetylcholine receptor (nAChR) subtypes is a prerequisite for the design of tool compounds to study nAChRs. However, selectivity optimization of these small, disulfide-rich peptides is difficult not only because of an absence of α-CTX/nAChR co-structures but also because it is challenging to predict how a mutation to an α-CTX will alter its potency and selectivity. As a prototypical system to investigate selectivity, we employed the α-CTX LvIA that is 25-fold selective for the α3ß2 nAChR over the related α3ß4 nAChR subtype, which is a target for nicotine addiction. Using two-electrode voltage clamp electrophysiology, we identified LvIA[D11R] that is 2-fold selective for the α3ß4 nAChR, reversing the subtype preference. This effect is specifically due to the change in charge and not shape of LvIA[D11R], as substitution of D11 with citrulline retains selectivity for the α3ß2 nAChR. Furthermore, LvIA[D11K] shows a stronger reversal, with 4-fold selectivity for the α3ß4 nAChR. Motivated by these findings, using site-directed mutagenesis, we found that ß2[K79A] (I79 on ß4), but not ß2[K78A] (N78 on ß4), largely restores the potency of basic mutants at position 11. Finally, to understand the structural basis of this effect, we used AlphaFold2 to generate models of LvIA in complex with both nAChR subtypes. Both models confirm the plausibility of an electrostatic mechanism to explain the data and also reproduce a broad range of potency and selectivity structure-activity relationships for LvIA mutants, as measured using free energy perturbation simulations. Our work highlights how electrostatic interactions can drive α-CTX selectivity and may serve as a strategy for optimizing the selectivity of LvIA and other α-CTXs.

Addressing Motor Dysfunction by a Selective α6-Containing Nicotinic Receptor Antagonist.

In the striatum, presynaptic α6-containig nicotinic receptors are crucially involved in the modulation of dopamine release. CVN417, a novel selective antagonist at this receptor subtype, attenuates motor dysfunction in a Parkinson's disease-relevant animal model, suggesting, for this pathology, a therapeutic strategy that could greatly profit from the restricted localization of α6* nicotinic receptors in the brain.

CDP-choline modulates cholinergic signaling and gut microbiota to alleviate DSS-induced inflammatory bowel disease.

Inflammatory bowel diseases (IBD) represent chronic gastrointestinal inflammatory disorders characterized by a complex and underexplored pathogenic mechanism. Previous research has revealed that IBD patients often have a deficiency of choline and its metabolites, including acetylcholine (ACh) and phosphatidylcholine (PC), within the colon. However, a comprehensive study linking these three substances and their mechanistic implications in IBD remains lacking. This study aimed to investigate the efficacy and underlying mechanism of cytidine diphosphate (CDP)-choline (citicoline), an intermediate product of choline metabolism, in a mouse model of IBD induced by dextran sulfate sodium salt (DSS). The results demonstrated that CDP-choline effectively alleviated colonic inflammation and deficiencies in choline, ACh, and PC by increasing the raw material. Further detection showed that CDP-choline also increased the ACh content by altering the expression of high-affinity choline transporter (ChT1) and acetylcholinesterase (AChE) in DSS-induced mice colon. Moreover, CDP-choline increased the expression of alpha7 nicotinic acetylcholine receptor (α7 nAChR) and activated the cholinergic anti-inflammatory pathway (CAP), leading to reduced colon macrophage activation and proinflammatory M1 polarization in IBD mice, thus reducing the levels of TNF-α and IL-6. In addition, CDP-choline reduced intestinal ecological imbalance and increased the content of hexanoic acid in short-chain fatty acids (SCFAs) in mice. In conclusion, this study elucidates the ability of CDP-choline to mitigate DSS-induced colon inflammation by addressing choline and its metabolites deficiencies, activating the CAP, and regulating the composition of the intestinal microbiome and SCFAs content, providing a potential prophylactic and therapeutic approach for IBD.

Discovery of CVN417, a Novel Brain-Penetrant α6-Containing Nicotinic Receptor Antagonist for the Modulation of Motor Dysfunction.

Nicotinic acetylcholine receptor (nAChR) α6 subunit RNA expression is relatively restricted to midbrain regions and is located presynaptically on dopaminergic neurons projecting to the striatum. This subunit modulates dopamine neurotransmission and may have therapeutic potential in movement disorders. We aimed to develop potent and selective α6-containing nAChR antagonists to explore modulation of dopamine release and regulation of motor function in vivo. High-throughput screening (HTS) identified novel α6-containing nAChR antagonists and led to the development of CVN417. This molecule blocks α6-containing nAChR activity in recombinant cells and reduces firing frequency of noradrenergic neurons in the rodent locus coeruleus. CVN417 modulated phasic dopaminergic neurotransmission in an impulse-dependent manner. In a rodent model of resting tremor, CVN417 attenuated this behavioral phenotype. These data suggest that selective antagonism of α6-containing nAChR, with molecules such as CVN417, may have therapeutic utility in treating the movement dysfunctions observed in conditions such as Parkinson's disease.

Advances in small molecule selective ligands for heteromeric nicotinic acetylcholine receptors.

The study of nicotinic acetylcholine receptors (nAChRs) has significantly progressed in the last decade, due to a) the improved techniques available for structural studies; b) the identification of ligands interacting at orthosteric and allosteric recognition sites on the nAChR proteins, able to tune channel conformational states; c) the better functional characterization of receptor subtypes/subunits and their therapeutic potential; d) the availability of novel pharmacological agents able to activate or block nicotinic-mediated cholinergic responses with subtype or stoichiometry selectivity. The copious literature on nAChRs is related to the pharmacological profile of new, promising subtype selective derivatives as well as the encouraging preclinical and early clinical evaluation of known ligands. However, recently approved therapeutic derivatives are still missing, and examples of ligands discontinued in advanced CNS clinical trials include drug candidates acting at both neuronal homomeric and heteromeric receptors. In this review, we have selected heteromeric nAChRs as the target and comment on literature reports of the past five years dealing with the discovery of new small molecule ligands or the advanced pharmacological/preclinical investigation of more promising compounds. The results obtained with bifunctional nicotinic ligands and a light-activated ligand as well as the applications of promising radiopharmaceuticals for heteromeric subtypes are also discussed.

Synthesis and Biological Activity of Novel α-Conotoxins Derived from Endemic Polynesian Cone Snails.

α-Conotoxins are well-known probes for the characterization of the various subtypes of nicotinic acetylcholine receptors (nAChRs). Identifying new α-conotoxins with different pharmacological profiles can provide further insights into the physiological or pathological roles of the numerous nAChR isoforms found at the neuromuscular junction, the central and peripheral nervous systems, and other cells such as immune cells. This study focuses on the synthesis and characterization of two novel α-conotoxins obtained from two species endemic to the Marquesas Islands, namely Conus gauguini and Conus adamsonii. Both species prey on fish, and their venom is considered a rich source of bioactive peptides that can target a wide range of pharmacological receptors in vertebrates. Here, we demonstrate the versatile use of a one-pot disulfide bond synthesis to achieve the α-conotoxin fold [Cys 1-3; 2-4] for GaIA and AdIA, using the 2-nitrobenzyl (NBzl) protecting group of cysteines for effective regioselective oxidation. The potency and selectivity of GaIA and AdIA against rat nicotinic acetylcholine receptors were investigated electrophysiologically and revealed potent inhibitory activities. GaIA was most active at the muscle nAChR (IC50 = 38 nM), whereas AdIA was most potent at the neuronal α6/3 ß2ß3 subtype (IC50 = 177 nM). Overall, this study contributes to a better understanding of the structure-activity relationships of α-conotoxins, which may help in the design of more selective tools.

Loop2 Size Modification Reveals Significant Impacts on the Potency of α-Conotoxin TxID.

α4/6-conotoxin TxID, which was identified from Conus textile, simultaneously blocks rat (r) α3ß4 and rα6/α3ß4 nicotinic acetylcholine receptors (nAChRs) with IC50 values of 3.6 nM and 33.9 nM, respectively. In order to identify the effects of loop2 size on the potency of TxID, alanine (Ala) insertion and truncation mutants were designed and synthesized in this study. An electrophysiological assay was used to evaluate the activity of TxID and its loop2-modified mutants. The results showed that the inhibition of 4/7-subfamily mutants [+9A]TxID, [+10A]TxID, [+14A]TxID, and all the 4/5-subfamily mutants against rα3ß4 and rα6/α3ß4 nAChRs decreased. Overall, ala-insertion or truncation of the 9th, 10th, and 11th amino acid results in a loss of inhibition and the truncation of loop2 has more obvious impacts on its functions. Our findings have strengthened the understanding of α-conotoxin, provided guidance for further modifications, and offered a perspective for future studies on the molecular mechanism of the interaction between α-conotoxins and nAChRs.

L-theanine attenuates nicotine reward and withdrawal signs in mice.

BACKGROUND: L-theanine, 2-amino-4-(ethylcarbamoyl) butyric acid, an amino acid detected in green tea leaves, is used as a dietary supplement to attenuate stress and enhance mood and cognition. Furthermore, L-theanine induces anxiolytic effects in humans. Recently, L-theanine was reported to reduce morphine physical dependence in primates, suggesting the potential usefulness of L-theanine for drug dependence intervention. OBJECTIVE: The aim of this study is to determine whether L-theanine attenuates nicotine-withdrawal (somatic and affective signs) and nicotine reward in mice. We also investigated the effects of L-theanine on nicotinic receptors binding and function. METHODS: ICR male mice rendered dependent to nicotine through implanted subcutaneous osmotic minipumps for 14 days undertook precipitated nicotine withdrawal by mecamylamine on day 15. Anxiety-like behaviors using LDB, somatic signs observation and hot plate latency were assessed consecutively after treatment with L-theanine. Furthermore, we examined the effect of L-theanine on acute nicotine responses and nicotine conditioned reward in mice and on expressed nicotinic receptors in oocytes. KEY FINDINGS: L-theanine reduced in a dose-dependent manner anxiety-like behavior, hyperalgesia and somatic signs during nicotine withdrawal. Also, L-theanine decreased the nicotine CPP, but it did not affect the acute responses of nicotine. Finally, L-theanine did not alter the binding or the function of expressed α4ß2 and α7 nAChRs. CONCLUSION: Our results support the potential of L-theanine as a promising candidate for treating nicotine dependence.

Nicotinic acetylcholine receptor subtype expression, function, and pharmacology: Therapeutic potential of α-conotoxins.

The pentameric nicotinic acetylcholine receptors (nAChRs) are typically classed as muscle- or neuronal-type, however, the latter has also been reported in non-neuronal cells. Given their broad distribution, nAChRs mediate numerous physiological and pathological processes including synaptic transmission, presynaptic modulation of transmitter release, neuropathic pain, inflammation, and cancer. There are 17 different nAChR subunits and combinations of these subunits produce subtypes with diverse pharmacological properties. The expression and role of some nAChR subtypes have been extensively deciphered with the aid of knock-out models. Many nAChR subtypes expressed in heterologous systems are selectively targeted by the disulfide-rich α-conotoxins. α-Conotoxins are small peptides isolated from the venom of cone snails, and a number of them have potential pharmaceutical value.

The role of nicotinic receptors in alcohol consumption.

The use of alcohol causes significant morbidity and mortality across the globe. Alcohol use disorder (AUD) is defined by the excessive use of this drug despite a negative impact on the individual's life. While there are currently medications available to treat AUD, they have limited efficacy and several side effects. As such, it is essential to continue to look for novel therapeutics. One target for novel therapeutics is nicotinic acetylcholine receptors (nAChRs). Here we systematically review the literature on the involvement of nAChRs in alcohol consumption. Data from both genetic and pharmacology studies provide evidence that nAChRs modulate alcohol intake. Interestingly, pharmacological modulation of all nAChR subtypes examined can decrease alcohol consumption. The reviewed literature demonstrates that nAChRs should continue to be investigated as novel therapeutics for AUD.