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Sample records for nicotinic receptor ion

  1. Nicotinic Receptors in Neurodegeneration

    PubMed Central

    Posadas, Inmaculada; López-Hernández, Beatriz; Ceña, Valentín

    2013-01-01

    Many studies have focused on expanding our knowledge of the structure and diversity of peripheral and central nicotinic receptors. Nicotinic acetylcholine receptors (nAChRs) are members of the Cys-loop superfamily of pentameric ligand-gated ion channels, which include GABA (A and C), serotonin, and glycine receptors. Currently, 9 alpha (α2-α10) and 3 beta (β2-β4) subunits have been identified in the central nervous system (CNS), and these subunits assemble to form a variety of functional nAChRs. The pentameric combination of several alpha and beta subunits leads to a great number of nicotinic receptors that vary in their properties, including their sensitivity to nicotine, permeability to calcium and propensity to desensitize. In the CNS, nAChRs play crucial roles in modulating presynaptic, postsynaptic, and extrasynaptic signaling, and have been found to be involved in a complex range of CNS disorders including Alzheimer’s disease (AD), Parkinson’s disease (PD), schizophrenia, Tourette´s syndrome, anxiety, depression and epilepsy. Therefore, there is growing interest in the development of drugs that modulate nAChR functions with optimal benefits and minimal adverse effects. The present review describes the main characteristics of nAChRs in the CNS and focuses on the various compounds that have been tested and are currently in phase I and phase II trials for the treatment of neurodegenerative diseases including PD, AD and age-associated memory and mild cognitive impairment. PMID:24179465

  2. Isotopic rubidium ion efflux assay for the functional characterization of nicotinic acetylcholine receptors on clonal cell lines

    SciTech Connect

    Lukas, R.J.; Cullen, M.J.

    1988-11-15

    An isotopic rubidium ion efflux assay has been developed for the functional characterization of nicotinic acetylcholine receptors on cultured neurons. This assay first involves the intracellular sequestration of isotopic potassium ion analog by the ouabain-sensitive action of a sodium-potassium ATPase. Subsequently, the release of isotopic rubidium ion through nicotinic acetylcholine receptor-coupled monovalent cation channels is activated by application of nicotinic agonists. Specificity of receptor-mediated efflux is demonstrated by its sensitivity to blockade by nicotinic, but not muscarinic, antagonists. The time course of agonist-mediated efflux, within the temporal limitations of the assay, indicates a slow inactivation of receptor function on prolonged exposure to agonist. Dose-response profiles (i) have characteristic shapes for different nicotinic agonists, (ii) are described by three operationally defined parameters, and (iii) reflect different affinities of agonists for binding sites that control receptor activation and functional inhibition. The rubidium ion efflux assay provides fewer hazards but greater sensitivity and resolution than isotopic sodium or rubidium ion influx assays for functional nicotinic receptors.

  3. Megacystis, mydriasis, and ion channel defect in mice lacking the α3 neuronal nicotinic acetylcholine receptor

    PubMed Central

    Xu, Wei; Gelber, Shari; Orr-Urtreger, Avi; Armstrong, Dawna; Lewis, Richard A.; Ou, Ching-Nan; Patrick, James; Role, Lorna; De Biasi, Mariella; Beaudet, Arthur L.

    1999-01-01

    The α3 subunit of the neuronal nicotinic acetylcholine receptor is widely expressed in autonomic ganglia and in some parts of the brain. The α3 subunit can form heteromultimeric ion channels with other α subunits and with β2 and β4 subunits, but its function in vivo is poorly understood. We prepared a null mutation for the α3 gene by deletion of exon 5 and found that homozygous (−/−) mice lacked detectable mRNA on Northern blotting. The −/− mice survive to birth but have impaired growth and increased mortality before and after weaning. The −/− mice have extreme bladder enlargement, dribbling urination, bladder infection, urinary stones, and widely dilated ocular pupils that do not contract in response to light. Detailed histological studies of −/− mice revealed no significant abnormalities in brain or peripheral tissues except urinary bladder, where inflammation was prominent. Ganglion cells and axons were present in bladder and bowel. Bladder strips from −/− mice failed to contract in response to 0.1 mM nicotine, but did contract in response to electrical field stimulation or carbamoylcholine. The number of acetylcholine-activated single-channel currents was severely reduced in the neurons of superior cervical ganglia in −/− mice with five physiologically distinguishable nicotinic acetylcholine receptor subtypes with different conductance and kinetic properties in wild-type mice, all of which were reduced in −/− mice. The findings in the α3-null mice suggest that this subunit is an essential component of the nicotinic receptors mediating normal function of the autonomic nervous system. The phenotype in −/− mice may be similar to the rare human genetic disorder of megacystis–microcolon–intestinal hypoperistalsis syndrome. PMID:10318955

  4. Mutations in the channel domain of a neuronal nicotinic receptor convert ion selectivity from cationic to anionic.

    PubMed

    Galzi, J L; Devillers-Thiéry, A; Hussy, N; Bertrand, S; Changeux, J P; Bertrand, D

    1992-10-01

    Introduction by site-directed mutagenesis of three amino acids from the MII segment of glycine or gamma-aminobutyric acid (GABAA) receptors into the MII segment of alpha 7 nicotinic receptor was sufficient to convert a cation-selective channel into an anion-selective channel gated by acetylcholine. A critical mutation was the insertion of an uncharged residue at the amino-terminal end of MII, stressing the importance of protein geometrical constraints on ion selectivity. PMID:1383829

  5. Nicotinic receptors in addiction pathways.

    PubMed

    Leslie, Frances M; Mojica, Celina Y; Reynaga, Daisy D

    2013-04-01

    Neuronal nicotinic acetylcholine receptors (nAChRs) are ligand-gated ion channels that consist of pentameric combinations of α and β subunits. These receptors are widely distributed throughout the brain and are highly expressed in addiction circuitry. The role of nAChRs in regulating neuronal activity and motivated behavior is complex and varies both in and among brain regions. The rich diversity of central nAChRs has hampered the characterization of their structure and function with use of classic pharmacological techniques. However, recent molecular approaches using null mutant mice with specific regional lentiviral re-expression, in combination with neuroanatomical and electrophysiological techniques, have allowed the elucidation of the influence of different nAChR types on neuronal circuit activity and behavior. This review will address the influence of nAChRs on limbic dopamine circuitry and the medial habenula-interpeduncular nucleus complex, which are critical mediators of reinforced behavior. Characterization of the mechanisms underlying regulation of addiction pathways by endogenous cholinergic transmission and by nicotine may lead to the identification of new therapeutic targets for treating tobacco dependence and other addictions. PMID:23247824

  6. Nicotinic acetylcholine receptors and cancer

    PubMed Central

    DANG, NINGNING; MENG, XIANGUANG; SONG, HAIYAN

    2016-01-01

    Nicotine, the primary addictive constituent of cigarettes, is believed to contribute to cancer promotion and progression through the activation of nicotinic acetylcholine receptors (nAChRs), which are membrane ligand-gated cation channels. nAChRs activation can be triggered by the neurotransmitter Ach, or certain other biological compounds, such as nicotine. In recent years, genome-wide association studies have indicated that allelic variation in the α5-α3-β4 nAChR cluster on chromosome 15q24-15q25.1 is associated with lung cancer risk. The role of nAChRs in other types of cancer has also been reported. The present review highlights the role of nAChRs in types of human cancer. PMID:27123240

  7. Nitrosamines as nicotinic receptor ligands.

    PubMed

    Schuller, Hildegard M

    2007-05-30

    Nitrosamines are carcinogens formed in the mammalian organism from amine precursors contained in food, beverages, cosmetics and drugs. The potent carcinogen, NNK, and the weaker carcinogen, NNN, are nitrosamines formed from nicotine. Metabolites of the nitrosamines react with DNA to form adducts responsible for genotoxic effects. We have identified NNK as a high affinity agonist for the alpha7 nicotinic acetylcholine receptor (alpha7nAChR) whereas NNN bound with high affinity to epibatidine-sensitive nAChRs. Diethylnitrosamine (DEN) bound to both receptors but with lower affinity. High levels of the alpha7nAChR were expressed in human small cell lung cancer (SCLC) cell lines and in hamster pulmonary neuroendocrine cells (PNECs), which serve as a model for the cell of origin of human SCLC. Exposure of SCLC or PNECs to NNK or nicotine increased expression of the alpha7nAChR and caused influx of Ca(2+), activation of PKC, Raf-1, ERK1/2, and c-myc, resulting in the stimulation of cell proliferation. Signaling via the alpha7nAChR was enhanced when cells were maintained in an environment of 10-15% CO(2) similar to that in the diseased lung. Hamsters with hyperoxia-induced pulmonary fibrosis developed neuroendocrine lung carcinomas similar to human SCLC when treated with NNK, DEN, or nicotine. The development of the NNK-induced tumors was prevented by green tea or theophylline. The beta-adrenergic receptor agonist, isoproterenol or theophylline blocked NNK-induced cell proliferation in vitro. NNK and nicotine-induced hyperactivity of the alpha7nAChR/RAF/ERK1/2 pathway thus appears to play a crucial role in the development of SCLC in smokers and could be targeted for cancer prevention. PMID:17459420

  8. Identification of domains influencing assembly and ion channel properties in α7 nicotinic receptor and 5-HT3 receptor subunit chimaeras

    PubMed Central

    Gee, V J; Kracun, S; Cooper, S T; Gibb, A J; Millar, N S

    2007-01-01

    Background and purpose: Nicotinic acetylcholine receptors (nAChRs) and 5-hydroxytryptamine type 3 receptors (5-HT3Rs) are members of the superfamily of neurotransmitter-gated ion channels. Both contain five subunits which assemble to form either homomeric or heteromeric subunit complexes. With the aim of identifying the influence of subunit domains upon receptor assembly and function, a series of chimaeras have been constructed containing regions of the neuronal nAChR α7 subunit and the 5-HT3 receptor 3A subunit. Experimental approach: A series of subunit chimaeras containing α7 and 5-HT3A subunit domains have been constructed and expressed in cultured mammalian cells. Properties of the expressed receptors have been examined by means of radioligand binding, agonist-induced changes in intracellular calcium and patch-clamp electrophysiology. Key results: Subunit domains which influence properties such as rectification, desensitization and conductance have been identified. In addition, the influence of subunit domains upon subunit folding, receptor assembly and cell-surface expression has been identified. Co-expression studies with the nAChR-associated protein RIC-3 revealed that, in contrast to the potentiating effect of RIC-3 on α7 nAChRs, RIC-3 caused reduced levels of cell-surface expression of some α7/5-HT3A chimaeras. Conclusions and implications: Evidence has been obtained which demonstrates that subunit transmembrane domains are critical for efficient subunit folding and assembly. In addition, functional characterization of subunit chimaeras revealed that both extracellular and cytoplasmic domains exert a dramatic and significant influence upon single-channel conductance. These data support a role for regions other than hydrophobic transmembrane domains in determining ion channel properties. PMID:17721553

  9. Differential Effects of Quercetin and Quercetin Glycosides on Human α7 Nicotinic Acetylcholine Receptor-Mediated Ion Currents.

    PubMed

    Lee, Byung-Hwan; Choi, Sun-Hye; Kim, Hyeon-Joong; Jung, Seok-Won; Hwang, Sung-Hee; Pyo, Mi-Kyung; Rhim, Hyewhon; Kim, Hyoung-Chun; Kim, Ho-Kyoung; Lee, Sang-Mok; Nah, Seung-Yeol

    2016-07-01

    Quercetin is a flavonoid usually found in fruits and vegetables. Aside from its antioxidative effects, quercetin, like other flavonoids, has a various neuropharmacological actions. Quercetin-3-O-rhamnoside (Rham1), quercetin-3-O-rutinoside (Rutin), and quercetin- 3-(2(G)-rhamnosylrutinoside (Rham2) are mono-, di-, and tri-glycosylated forms of quercetin, respectively. In a previous study, we showed that quercetin can enhance α7 nicotinic acetylcholine receptor (α7 nAChR)-mediated ion currents. However, the role of the carbohydrates attached to quercetin in the regulation of α7 nAChR channel activity has not been determined. In the present study, we investigated the effects of quercetin glycosides on the acetylcholine induced peak inward current (IACh) in Xenopus oocytes expressing the α7 nAChR. IACh was measured with a two-electrode voltage clamp technique. In oocytes injected with α7 nAChR copy RNA, quercetin enhanced IACh, whereas quercetin glycosides inhibited IACh. Quercetin glycosides mediated an inhibition of IACh, which increased when they were pre-applied and the inhibitory effects were concentration dependent. The order of IACh inhibition by quercetin glycosides was Rutin≥Rham1>Rham2. Quercetin glycosides-mediated IACh enhancement was not affected by ACh concentration and appeared voltage-independent. Furthermore, quercetin-mediated IACh inhibition can be attenuated when quercetin is co-applied with Rham1 and Rutin, indicating that quercetin glycosides could interfere with quercetin-mediated α7 nAChR regulation and that the number of carbohydrates in the quercetin glycoside plays a key role in the interruption of quercetin action. These results show that quercetin and quercetin glycosides regulate the α7 nAChR in a differential manner. PMID:27098860

  10. Differential Effects of Quercetin and Quercetin Glycosides on Human α7 Nicotinic Acetylcholine Receptor-Mediated Ion Currents

    PubMed Central

    Lee, Byung-Hwan; Choi, Sun-Hye; Kim, Hyeon-Joong; Jung, Seok-Won; Hwang, Sung-Hee; Pyo, Mi-Kyung; Rhim, Hyewhon; Kim, Hyoung-Chun; Kim, Ho-Kyoung; Lee, Sang-Mok; Nah, Seung-Yeol

    2016-01-01

    Quercetin is a flavonoid usually found in fruits and vegetables. Aside from its antioxidative effects, quercetin, like other flavonoids, has a various neuropharmacological actions. Quercetin-3-O-rhamnoside (Rham1), quercetin-3-O-rutinoside (Rutin), and quercetin-3-(2(G)-rhamnosylrutinoside (Rham2) are mono-, di-, and tri-glycosylated forms of quercetin, respectively. In a previous study, we showed that quercetin can enhance α7 nicotinic acetylcholine receptor (α7 nAChR)-mediated ion currents. However, the role of the carbohydrates attached to quercetin in the regulation of α7 nAChR channel activity has not been determined. In the present study, we investigated the effects of quercetin glycosides on the acetylcholine induced peak inward current (IACh) in Xenopus oocytes expressing the α7 nAChR. IACh was measured with a two-electrode voltage clamp technique. In oocytes injected with α7 nAChR copy RNA, quercetin enhanced IACh, whereas quercetin glycosides inhibited IACh. Quercetin glycosides mediated an inhibition of IACh, which increased when they were pre-applied and the inhibitory effects were concentration dependent. The order of IACh inhibition by quercetin glycosides was Rutin≥Rham1>Rham2. Quercetin glycosides-mediated IACh enhancement was not affected by ACh concentration and appeared voltage-independent. Furthermore, quercetin-mediated IACh inhibition can be attenuated when quercetin is co-applied with Rham1 and Rutin, indicating that quercetin glycosides could interfere with quercetin-mediated α7 nAChR regulation and that the number of carbohydrates in the quercetin glycoside plays a key role in the interruption of quercetin action. These results show that quercetin and quercetin glycosides regulate the α7 nAChR in a differential manner. PMID:27098860

  11. [Desensitization of the nicotinic acetylcholine receptor].

    PubMed

    Quiñonez, M; Rojas, L

    1994-01-01

    In biological membranes, ionic channels act speeding up ion movements. Each ionic channel is excited by a specific stimulus (i.e. electric, mechanical, chemical, etc.). Chemically activated ionic channels (CAIC), such as the nicotinic acetylcholine receptor (nAChR), suffer desensitization when the receptor site is still occupied by the agonist molecule. The desensitized CAIC is a non functional channel state regarded as a particular case of receptors rundown. CAIC desensitization only involve reduced activity and not their membrane elimination. Desensitization is important to control synaptic transmission and the development of the nervous system. In this review we discuss results related to its production, modulation and some aspects associated to models that consider it. Finally, an approach combining molecular biology and electrophysiology techniques to understand desensitization and its importance in biological systems is presented. PMID:8525756

  12. Nicotinic Receptor Antagonists as Treatments for Nicotine Abuse

    PubMed Central

    Crooks, Peter A.; Bardo, Michael T.; Dwoskin, Linda P.

    2014-01-01

    Despite the proven efficacy of current pharmacotherapies for tobacco dependence, relapse rates continue to be high, indicating that novel medications are needed. Currently, several smoking cessation agents are available, including varenicline (Chantix®), bupropion (Zyban®), and cytisine (Tabex®). Varenicline and cytisine are partial agonists at the α4β2* nicotinic acetylcholine receptor (nAChR). Bupropion is an antidepressant but is also an antagonist at α3β2* ganglionic nAChRs. The rewarding effects of nicotine are mediated, in part, by nicotine-evoked dopamine (DA) release leading to sensitization, which is associated with repeated nicotine administration and nicotine addiction. Receptor antagonists that selectivity target central nAChR subtypes mediating nicotine-evoked DA release should have efficacy as tobacco use cessation agents with the therapeutic advantage of a limited side-effect profile. While α-conotoxin MII (α-CtxMII)-insensitive nAChRs (e.g., α4β2*) contribute to nicotine-evoked DA release, these nAChRs are widely distributed in the brain, and inhibition of these receptors may lead to nonselective and untoward effects. In contrast, α-CtxMII-sensitive nAChRs mediating nicotine-evoked DA release offer an advantage as targets for smoking cessation, due to their more restricted localization primarily to dopaminergic neurons. Small drug-like molecules that are selective antagonists at α-CtxMII-sensitive nAChR subtypes that contain α6 and β2 subunits have now been identified. Early research identified a variety of quaternary ammonium analogs that were potent and selective antagonists at nAChRs mediating nicotine-evoked DA release. More recent data have shown that novel, non-quaternary bis-1,2,5,6-tetrahydropyridine analogs potently inhibit (IC50<1 nM) nicotine-evoked DA release in vitro by acting as antagonists at α-CtxMII-sensitive nAChR subtypes; these compounds also decrease NIC self-administration in rats. PMID:24484986

  13. Conotoxins Targeting Nicotinic Acetylcholine Receptors: An Overview

    PubMed Central

    Lebbe, Eline K. M.; Peigneur, Steve; Wijesekara, Isuru; Tytgat, Jan

    2014-01-01

    Marine snails of the genus Conus are a large family of predatory gastropods with an unparalleled molecular diversity of pharmacologically active compounds in their venom. Cone snail venom comprises of a rich and diverse cocktail of peptide toxins which act on a wide variety of ion channels such as voltage-gated sodium- (NaV), potassium- (KV), and calcium- (CaV) channels as well as nicotinic acetylcholine receptors (nAChRs) which are classified as ligand-gated ion channels. The mode of action of several conotoxins has been the subject of investigation, while for many others this remains unknown. This review aims to give an overview of the knowledge we have today on the molecular pharmacology of conotoxins specifically interacting with nAChRs along with the structure–function relationship data. PMID:24857959

  14. Nicotine recruits glutamate receptors to postsynaptic sites.

    PubMed

    Duan, Jing-Jing; Lozada, Adrian F; Gou, Chen-Yu; Xu, Jing; Chen, Yuan; Berg, Darwin K

    2015-09-01

    Cholinergic neurons project throughout the nervous system and activate nicotinic receptors to modulate synaptic function in ways that shape higher order brain function. The acute effects of nicotinic signaling on long-term synaptic plasticity have been well-characterized. Less well understood is how chronic exposure to low levels of nicotine, such as those encountered by habitual smokers, can alter neural connections to promote addiction and other lasting behavioral effects. We show here that chronic exposure of hippocampal neurons in culture to low levels of nicotine recruits AMPA and NMDA receptors to the cell surface and sequesters them at postsynaptic sites. The receptors include GluA2-containing AMPA receptors, which are responsible for most of the excitatory postsynaptic current mediated by AMPA receptors on the neurons, and include NMDA receptors containing GluN1 and GluN2B subunits. Moreover, we find that the nicotine treatment also increases expression of the presynaptic component synapsin 1 and arranges it in puncta juxtaposed to the additional AMPA and NMDA receptor puncta, suggestive of increases in synaptic contacts. Consistent with increased synaptic input, we find that the nicotine treatment leads to an increase in the excitatory postsynaptic currents mediated by AMPA and NMDA receptors. Further, the increases skew the ratio of excitatory-to-inhibitory input that the cell receives, and this holds both for pyramidal neurons and inhibitory neurons in the hippocampal CA1 region. The GluN2B-containing NMDA receptor redistribution at synapses is associated with a significant increase in GluN2B phosphorylation at Tyr1472, a site known to prevent GluN2B endocytosis. These results suggest that chronic exposure to low levels of nicotine not only alters functional connections but also is likely to change excitability levels across networks. Further, it may increase the propensity for synaptic plasticity, given the increase in synaptic NMDA receptors. PMID

  15. Neuronal nicotinic acetylcholine receptor modulation by general anesthetics.

    PubMed

    Flood, P; Role, L W

    1998-11-23

    1. General anesthetics have been shown to inhibit synaptic transmission in multiple areas of the central and peripheral nervous systems. 2. The mechanism of inhibition is not well understood. 3. It has become clear that general anesthetics modulate the function of members of the ligand gated ion channel superfamily, including receptors for GABA(A), glycine (Harrison et al., Mol. Pharmacol. 44(3), 1993, 628-632) and 5HT3 (Zhou and Lovinger, J. Pharmacol. Exp. Therap. 278(2), 1996, 732-740). 4. Studies of the activity of general anesthetics on recombinant neuronal nicotinic acetylcholine receptors have added this receptor family to those potently inhibited by general anesthetics (Flood et al., Anesthesiology 86(4), 1997, 859-865; Violet et al., Anesthesiology 86(4), 1997, 866-874). 5. Studies of neuronal nicotinic receptors in native neurons suggest that the inhibition of these receptors by general anesthetics at low clinical concentrations may be biologically significant (Nicoll, Science 199(4327), 1978, 451-452). 6. Recent work on neuronal nicotinic acetylcholine receptors in the central nervous system suggests that their primary role may be to modulate synaptic transmission (Role and Berg, Neuron 16(6), 1996, 1077-1085). 7. Thus, inhibition of nicotinic modulation in the central nervous system may result in inhibition of synaptic transmission and some of the behavioral consequences of general anesthesia. PMID:10049135

  16. Neuronal Nicotinic Acetylcholine Receptors: Common Molecular Substrates of Nicotine and Alcohol Dependence

    PubMed Central

    Hendrickson, Linzy M.; Guildford, Melissa J.; Tapper, Andrew R.

    2013-01-01

    Alcohol and nicotine are often co-abused. As many as 80–95% of alcoholics are also smokers, suggesting that ethanol and nicotine, the primary addictive component of tobacco smoke, may functionally interact in the central nervous system and/or share a common mechanism of action. While nicotine initiates dependence by binding to and activating neuronal nicotinic acetylcholine receptors (nAChRs), ligand-gated cation channels normally activated by endogenous acetylcholine (ACh), ethanol is much less specific with the ability to modulate multiple gene products including those encoding voltage-gated ion channels, and excitatory/inhibitory neurotransmitter receptors. However, emerging data indicate that ethanol interacts with nAChRs, both directly and indirectly, in the mesocorticolimbic dopaminergic (DAergic) reward circuitry to affect brain reward systems. Like nicotine, ethanol activates DAergic neurons of the ventral tegmental area (VTA) which project to the nucleus accumbens (NAc). Blockade of VTA nAChRs reduces ethanol-mediated activation of DAergic neurons, NAc DA release, consumption, and operant responding for ethanol in rodents. Thus, ethanol may increase ACh release into the VTA driving activation of DAergic neurons through nAChRs. In addition, ethanol potentiates distinct nAChR subtype responses to ACh and nicotine in vitro and in DAergic neurons. The smoking cessation therapeutic and nAChR partial agonist, varenicline, reduces alcohol consumption in heavy drinking smokers and rodent models of alcohol consumption. Finally, single nucleotide polymorphisms in nAChR subunit genes are associated with alcohol dependence phenotypes and smoking behaviors in human populations. Together, results from pre-clinical, clinical, and genetic studies indicate that nAChRs may have an inherent role in the abusive properties of ethanol, as well as in nicotine and alcohol co-dependence. PMID:23641218

  17. Neuronal nicotinic acetylcholine receptors: neuroplastic changes underlying alcohol and nicotine addictions

    PubMed Central

    Feduccia, Allison A.; Chatterjee, Susmita; Bartlett, Selena E.

    2012-01-01

    Addictive drugs can activate systems involved in normal reward-related learning, creating long-lasting memories of the drug's reinforcing effects and the environmental cues surrounding the experience. These memories significantly contribute to the maintenance of compulsive drug use as well as cue-induced relapse which can occur even after long periods of abstinence. Synaptic plasticity is thought to be a prominent molecular mechanism underlying drug-induced learning and memories. Ethanol and nicotine are both widely abused drugs that share a common molecular target in the brain, the neuronal nicotinic acetylcholine receptors (nAChRs). The nAChRs are ligand-gated ion channels that are vastly distributed throughout the brain and play a key role in synaptic neurotransmission. In this review, we will delineate the role of nAChRs in the development of ethanol and nicotine addiction. We will characterize both ethanol and nicotine's effects on nAChR-mediated synaptic transmission and plasticity in several key brain areas that are important for addiction. Finally, we will discuss some of the behavioral outcomes of drug-induced synaptic plasticity in animal models. An understanding of the molecular and cellular changes that occur following administration of ethanol and nicotine will lead to better therapeutic strategies. PMID:22876217

  18. Nicotinic Acetylcholine Receptors in Sensory Cortex

    ERIC Educational Resources Information Center

    Metherate, Raju

    2004-01-01

    Acetylcholine release in sensory neocortex contributes to higher-order sensory function, in part by activating nicotinic acetylcholine receptors (nAChRs). Molecular studies have revealed a bewildering array of nAChR subtypes and cellular actions; however, there is some consensus emerging about the major nAChR subtypes and their functions in…

  19. Nicotine effect on cardiovascular system and ion channels.

    PubMed

    Hanna, Salma Toma

    2006-03-01

    Smoking is a leading cause of cardiovascular disease, hypertension, myocardial infarction, and stroke. Nicotine is one of the components of cigarette smoke. Nicotine effects on the cardiovascular system reflect the activity of the nicotine receptors centrally and on peripheral autonomic ganglia. It has been found that cigarette smoke extract-induced contraction of porcine coronary arteries is related to superoxide anion-mediated degradation of nitric oxide. Treatment of rabbit aortas with an oxygen free radicals scavenger attenuated cigarette smoke impairment of arterial relaxation. Treatment of smokers with vitamin C, an antioxidant, improved impaired endothelium-dependent reactivity of large peripheral arteries. Thus it appears that chronic smoking and acute exposure to cigarette smoke extract may alter endothelium-dependent reactivity via the production of oxygen derived free radicals. This review discusses the effects of nicotine on resistance arterioles, compliance arteries, smooth muscle cells, and ion channels in the cardiovascular system. We discuss studies performed on humans, nicotine-exposed animals, and cell cultures yielding varying and inconsistent results that may be due to differences in experimental design, species, and the dose of exposure. Nicotine exposure appears to induce a combination of free radical production, vascular wall adhesion, and a reduction of fibrinolytic activity in the plasma. PMID:16633075

  20. Neuronal nicotinic receptors as brain targets for pharmacotherapy of drug addiction.

    PubMed

    Rahman, Shafiqur; López-Hernández, Gretchen Y; Corrigall, William A; Papke, Roger L

    2008-11-01

    Nicotine addiction and other forms of drug addiction continue to be significant public health problems in the United States and the rest of the world. Accumulated evidence indicates that brain nicotinic acetylcholine receptors (nAChRs) are a heterogenous family of ion channels expressed in the various parts of the brain. A growing body of preclinical studies suggests that brain nAChRs are critical targets for the development of pharmacotherapies for nicotine and other drug addictions. In this review, we will discuss the nAChR subtypes, their function in response to endogenous brain transmitters, and how their functions are regulated in the presence of nicotine. Furthermore, we will discuss the role of nAChRs in mediating nicotine-induced addictive behavior in animal models. Additionally, we will provide an overview of the effects of nicotine and nicotinic compounds on the mesolimbic dopamine system, part of the reinforcement/reward circuitry of the brain, as an example of the neurochemical basis of nicotine addiction and other drug addictions. An appreciation of the complexity of nicotinic receptors and their regulation will be necessary for the development of nicotinic receptor modulators as potential pharmacotherapy for drug addiction. PMID:19128201

  1. Regulation of nicotinic receptors in rat brain following quasi-irreversible nicotinic blockade by chlorisondamine and chronic treatment with nicotine.

    PubMed Central

    el-Bizri, H; Clarke, P B

    1994-01-01

    1. Chronic administration of nicotinic agonists in vivo increases the density of brain nicotinic binding sites. It has been proposed that this up-regulation results from agonist-induced functional blockade of nicotinic receptors. This hypothesis was tested by examining post mortem [3H]-nicotine and [125I]-alpha-bungarotoxin ([125I]-alpha BTX) binding following treatment in vivo with the quasi-irreversible and insurmountable CNS nicotinic blocker chlorisondamine, given either alone or in combination with chronic nicotine administration. 2. In rats that had not received chlorisondamine pretreatment, chronic nicotine administration (0.6 mg kg-1 s.c., twice daily for 12 days) increased [3H]-nicotine binding density (Bmax) in forebrain tissue sections by 19%, with no change in the apparent dissociation constant (KD). Chlorisondamine (10 mg kg-1, s.c.), given once prior to the chronic treatment phase, neither increased [3H]-nicotine binding by itself, nor altered the extent of nicotine-induced up-regulation. Nevertheless, chlorisondamine pretreatment resulted in a persistent blockade of CNS nicotinic receptors, as demonstrated by complete block of acute locomotor responses to nicotine. 3. In a second experiment, [3H]-nicotine and [125I]-alpha BTX binding was measured in tissue homogenates prepared from several brain regions. In the absence of chlorisondamine pretreatment, chronic nicotine administration (1 mg kg-1 s.c., twice daily for 12 days) increased the Bmax of [3H]-nicotine binding in the cerebral cortex (by 34%), striatum (by 28%), midbrain (by 16%) and hippocampus (by 36%); KD was unchanged. As before, this up-regulation was neither mimicked nor blocked by chlorisondamine pretreatment (10 mg kg-1, s.c., given twice), despite persistent blockade of acute locomotor responses to nicotine.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7858886

  2. The Drosophila nicotinic acetylcholine receptor subunits Dα5 and Dα7 form functional homomeric and heteromeric ion channels

    PubMed Central

    2012-01-01

    Background Nicotinic acetylcholine receptors (nAChRs) play an important role as excitatory neurotransmitters in vertebrate and invertebrate species. In insects, nAChRs are the site of action of commercially important insecticides and, as a consequence, there is considerable interest in examining their functional properties. However, problems have been encountered in the successful functional expression of insect nAChRs, although a number of strategies have been developed in an attempt to overcome such difficulties. Ten nAChR subunits have been identified in the model insect Drosophila melanogaster (Dα1-Dα7 and Dβ1-Dβ3) and a similar number have been identified in other insect species. The focus of the present study is the Dα5, Dα6 and Dα7 subunits, which are distinguished by their sequence similarity to one another and also by their close similarity to the vertebrate α7 nAChR subunit. Results A full-length cDNA clone encoding the Drosophila nAChR Dα5 subunit has been isolated and the properties of Dα5-, Dα6- and Dα7-containing nAChRs examined in a variety of cell expression systems. We have demonstrated the functional expression, as homomeric nAChRs, of the Dα5 and Dα7 subunits in Xenopus oocytes by their co-expression with the molecular chaperone RIC-3. Also, using a similar approach, we have demonstrated the functional expression of a heteromeric ‘triplet’ nAChR (Dα5 + Dα6 + Dα7) with substantially higher apparent affinity for acetylcholine than is seen with other subunit combinations. In addition, specific cell-surface binding of [125I]-α-bungarotoxin was detected in both Drosophila and mammalian cell lines when Dα5 was co-expressed with Dα6 and RIC-3. In contrast, co-expression of additional subunits (including Dα7) with Dα5 and Dα6 prevented specific binding of [125I]-α-bungarotoxin in cell lines, suggesting that co-assembly with other nAChR subunits can block maturation of correctly folded nAChRs in some cellular

  3. New quinoline derivatives as nicotinic receptor modulators.

    PubMed

    Manetti, Dina; Bellucci, Cristina; Dei, Silvia; Teodori, Elisabetta; Varani, Katia; Spirova, Ekaterina; Kudryavtsev, Denis; Shelukhina, Irina; Tsetlin, Victor; Romanelli, Maria Novella

    2016-03-01

    As a continuation of previous work on quinoline derivatives, which showed some preference (2-3 times) for the α7 with respect to α4β2 acetylcholine nicotinic receptors (nAChRs), we synthesized a series of novel azabicyclic or diazabicyclic compounds carrying a quinoline or isoquinoline ring, with the aim of searching for more selective α7 nAChR compounds. Radioligand binding studies on α7* and α4β2* nAChRs (rat brain homogenate) revealed one compound (7) with a 2-fold higher affinity for the α4β2*-subtype, and four compounds (11, 13, 14 and 16) with at least 3-fold higher affinity for α7* nAChR. The most promising was 11, showing Ki∼100 nM and over 10-fold selectivity for α7* nAChR. Compounds 7, 11, 13 and 16 at 50 μM suppressed ion currents induced in the rat α4β2 nAChR and the chimeric nAChR composed of the ligand-binding domain of the chick α7 and transmembrane domain of the α1 glycine receptor, expressed in Xenopus oocytes. Calcium imaging experiments on the human α7 nAChR expressed in the Neuro2a cells and potentiated by PNU-120596 confirmed the antagonistic activity for 7; on the contrary, 11, 13 and 16 were agonists with the EC50 values in the range of 1.0-1.6 μM. Thus, the introduced modifications allowed us to enhance the selectivity of quinolines towards α7 nAChR and to get novel compounds with agonistic activity. PMID:26840365

  4. Cholinergic modulation of dopamine pathways through nicotinic acetylcholine receptors.

    PubMed

    de Kloet, Sybren F; Mansvelder, Huibert D; De Vries, Taco J

    2015-10-15

    Nicotine addiction is highly prevalent in current society and is often comorbid with other diseases. In the central nervous system, nicotine acts as an agonist for nicotinic acetylcholine receptors (nAChRs) and its effects depend on location and receptor composition. Although nicotinic receptors are found in most brain regions, many studies on addiction have focused on the mesolimbic system and its reported behavioral correlates such as reward processing and reinforcement learning. Profound modulatory cholinergic input from the pedunculopontine and laterodorsal tegmentum to dopaminergic midbrain nuclei as well as local cholinergic interneuron projections to dopamine neuron axons in the striatum may play a major role in the effects of nicotine. Moreover, an indirect mesocorticolimbic feedback loop involving the medial prefrontal cortex may be involved in behavioral characteristics of nicotine addiction. Therefore, this review will highlight current understanding of the effects of nicotine on the function of mesolimbic and mesocortical dopamine projections in the mesocorticolimbic circuit. PMID:26208783

  5. Role of α5-containing nicotinic receptors in neuropathic pain and response to nicotine.

    PubMed

    Xanthos, Dimitris N; Beiersdorf, Johannes W; Thrun, Ariane; Ianosi, Bogdan; Orr-Urtreger, Avi; Huck, Sigismund; Scholze, Petra

    2015-08-01

    Nicotinic receptors in the central nervous system (nAChRs) are known to play important roles in pain processing and modulate behavioral responses to analgesic drugs, including nicotine. The presence of the α5-neuronal nicotinic accessory subunit in the nicotinic receptor complex is increasingly understood to modulate reward and aversive states, addiction, and possibly pathological pain. In the current study, using α5-knockout (KO) mice and subunit-specific antibodies, we assess the role of α5-containing neuronal nicotinic receptors in neuropathic pain and in the analgesic response to nicotine. After chronic constriction injury (CCI) or partial sciatic nerve ligation (PSNL), no differences in mechanical, heat, or cold hyperalgesia were found in wild-type (WT) versus α5-KO littermate mice. The number of α5-containing nAChRs was decreased (rather than increased) after CCI in the spinal cord and in the thalamus. Nevertheless, thermal analgesic response to nicotine was marginally reduced in CCI α5-KO mice at 4 days after CCI, but not at later timepoints or after PSNL. Interestingly, upon daily intermittent nicotine injections in unoperated mice, WT animals developed tolerance to nicotine-induced analgesia to a larger extent than α5-KO mice. Our results suggest that α5-containing nAChRs mediate analgesic tolerance to nicotine but do not play a major role in neuropathic pain. PMID:25725336

  6. Nicotinic acetylcholine receptors: upregulation, age-related effects and associations with drug use

    PubMed Central

    Melroy-Greif, W. E.; Stitzel, J. A.; Ehringer, M. A.

    2016-01-01

    Nicotinic acetylcholine receptors are ligand-gated ion channels that exogenously bind nicotine. Nicotine produces rewarding effects by interacting with these receptors in the brain’s reward system. Unlike other receptors, chronic stimulation by an agonist induces an upregulation of receptor number that is not due to increased gene expression in adults; while upregulation also occurs during development and adolescence there have been some opposing findings regarding a change in corresponding gene expression. These receptors have also been well studied with regard to human genetic associations and, based on evidence suggesting shared genetic liabilities between substance use disorders, numerous studies have pointed to a role for this system in comorbid drug use. This review will focus on upregulation of these receptors in adulthood, adolescence and development, as well as the findings from human genetic association studies which point to different roles for these receptors in risk for initiation and continuation of drug use. PMID:26351737

  7. Modal gating of muscle nicotinic acetylcholine receptors

    NASA Astrophysics Data System (ADS)

    Vij, Ridhima

    Many ion channels exhibit multiple patterns of kinetic activity in single-channel currents. This behavior is rare in WT mouse muscle nicotinic acetylcholine receptors (AChRs), where A2C↔A2O gating events are well-described by single exponentials. Also, single-channel open probability (PO) is essentially homogeneous at a given agonist concentration in the WT receptors. Here I report that perturbations of almost all the residues in loop C (alpha188-alpha199, at the agonist binding site) generate heterogeneity in PO ('modes'). Such unsettled activity was apparent with an alanine substitution at all positions in loop C (except alphaY190 and alphaY198) and with different side chain substitutions at alphaP197 for both adult- and fetal-type AChRs. I used single channel electrophysiology along with site-directed mutagenesis to study modal gating in AChRs consequent to mutations/deletions in loop C. The multiple patterns of kinetic activity arose from the difference in agonist affinity rather than in intrinsic AChR gating. Out of the four different agonists used to study the modal behavior, acetylcholine (ACh) showed a higher degree of kinetic heterogeneity compared to others. The time constant for switching between modes was long (~mins), suggesting that they arise from alternative, stable protein conformations. By studying AChRs having only 1 functional binding site, I attempted to find the source of the affinity difference, which was traced mainly to the alphadelta agonist site. Affinity at the neurotransmitter binding site is mainly determined by a core of five aromatic residues (alphaY93, alphaW149, alphaY190, alphaY198 and deltaW57). Phenylalanine substitutions at all aromatic residues except alphaY93 resulted in elimination of modes. Modes were also eliminated by alanine mutation at deltaW57 on the complementary side but not at other aromatics. Also, by substituting four gamma subunit residues into the delta subunit on the complementary beta sheet, I found that

  8. Desformylflustrabromine: A Novel Positive Allosteric Modulator for beta2 Subunit Containing Nicotinic Receptor Sub-Types.

    PubMed

    Pandya, Anshul A

    2016-01-01

    Nicotinic acetylcholine receptors are ligand-gated transmembrane ion channels that are present at the neuromuscular junction and in different locations in the nervous system. The different subtypes of neuronal nicotinic acetylcholine receptors that are found in the brain are thought to be involved in many neurological processes such as pain, cognitive function and depression, as well as in the pathophysiology of numerous neurological diseases and conditions. While the neurotransmitter acetylcholine is an endogenous agonist for all nicotinic receptors subtypes, many drugs that act as agonists and antagonists have also been identified or developed for these receptors. In addition, a novel class of compounds described as allosteric modulators have also been identified or developed for nicotinic acetylcholine receptors. Allosteric modulators are ligands that bind to nicotinic receptors at sites other than the orthosteric site where acetylcholine binds. One such allosteric modulator is desformylflustrabromine. Five chemical analogs along with desformylflustrabromine act as positive allosteric modulator for nAChRs that contain the beta2 subunit in their pentameric structure. Here the discovery and development, medicinal chemistry and pharmacological actions of desformylflustrabromine have been discussed. Desformylflustrabromine and its chemical analogs have the potential to develop into clinically used drugs for neurological diseases and conditions where nicotinic acetylcholine receptors are involved. PMID:26818864

  9. Nicotine Dependence Reveals Distinct Responses from Neurons and Their Resident Nicotinic Receptors in Medial Habenula.

    PubMed

    Shih, Pei-Yu; McIntosh, J Michael; Drenan, Ryan M

    2015-12-01

    Nicotinic acetylcholine receptors (nAChRs) are the molecular target of nicotine. nAChRs in the medial habenula (MHb) have recently been shown to play a role in nicotine dependence, but it is not clear which nAChR subtypes or MHb neuron types are most important. To identify MHb nAChRs and/or cell types that play a role in nicotine dependence, we studied these receptors and cells with brain slice electrophysiology using both acute and chronic nicotine application. Cells in the ventroinferior (MHbVI) and ventrolateral MHb (MHbVL) subregions expressed functional nAChRs with different pharmacology. Further, application of nicotine to cells in these subregions led to different action potential firing patterns. The latter result was correlated with a differing ability of nicotine to induce nAChR desensitization. Chronic nicotine caused functional upregulation of nAChRs selectively in MHbVI cells, but did not change nAChR function in MHbVL. Importantly, firing responses were also differentially altered in these subregions following chronic nicotine. MHbVI neurons treated chronically with nicotine exhibited enhanced basal pacemaker firing but a blunted nicotine-induced firing response. MHbVL neurons did not change their firing properties in response to chronic nicotine. Together, these results suggest that acute and chronic nicotine differentially affect nAChR function and output of cells in MHb subregions. Because the MHb extensively innervates the interpeduncular nucleus, an area critical for both affective and somatic signs of withdrawal, these results could reflect some of the neurophysiological changes thought to occur in the MHb to the interpeduncular nucleus circuit in human smokers. PMID:26429939

  10. Role of α5 Nicotinic Acetylcholine Receptors in Pharmacological and Behavioral Effects of Nicotine in Mice

    PubMed Central

    Marks, M. J.; Vann, R. E.; Chen, X.; Gamage, T. F.; Warner, J. A.; Damaj, M. I.

    2010-01-01

    Incorporation of the α5 nicotinic acetylcholine receptor (nAChR) subunit can greatly influence nAChR function without altering receptor number. Although few animal studies have assessed the role of the α5 nAChR in nicotine-mediated behaviors, recent evidence suggests an association between polymorphisms in the α5 nAChR gene and nicotine dependence phenotypes in humans. Thus, additional studies are imperative to elucidate the role and function of the α5 nAChR subunit in nicotine dependence. Using α5(−/−) mice, the current study aimed to examine the role of α5 nAChRs in the initial pharmacological effects of nicotine, nicotine reward using the conditioned place preference model, and the discriminative effects of nicotine using a two-lever drug discrimination model. 86Rb+ efflux and 125I-epibatidine binding assays were conducted to examine the effect of α5 nAChR subunit deletion on expression and activity of functional nAChRs. Results show that α5(−/−) mice are less sensitive to the initial effects of nicotine in antinociception, locomotor activity, and hypothermia measures and that the α5 nAChR is involved in nicotine reward. Alternatively, α5(−/−) mice did not differ from wild-type littermates in sensitivity to the discriminative stimulus effects of nicotine. Furthermore, deletion of the α5 nAChR subunit resulted in a statistically significant decrease in function in the thalamus and hindbrain, but the decreases noted in spinal cord were not statistically significant. Receptor number was unaltered in all areas tested. Taken together, results of the study suggest that α5 nAChRs are involved in nicotine-mediated behaviors relevant to development of nicotine dependence. PMID:20400469

  11. Nicotine evokes kinetic tremor by activating the inferior olive via α7 nicotinic acetylcholine receptors.

    PubMed

    Kunisawa, Naofumi; Iha, Higor A; Shimizu, Saki; Tokudome, Kentaro; Mukai, Takahiro; Kinboshi, Masato; Serikawa, Tadao; Ohno, Yukihiro

    2016-11-01

    Nicotinic acetylcholine (nACh) receptors are implicated in the pathogenesis of movement disorders (e.g., tremor) and epilepsy. Here, we performed behavioral and immunohistochemical studies using mice and rats to elucidate the mechanisms underlying nicotine-induced tremor. Treatments of animals with nicotine (0.5-2mg/kg, i.p.) elicited kinetic tremor, which was completely suppressed by the nACh receptor antagonist mecamylamine (MEC). The specific α7 nACh receptor antagonist methyllycaconitine (MLA) also inhibited nicotine-induced tremor, whereas the α4β2 nACh antagonist dihydro-β-erythroidine (DHβE) or the peripheral α3β4 nACh antagonist hexamethonium showed no effects. Mapping analysis of Fos protein expression, a biological marker of neural excitation, revealed that a tremorgenic dose (1mg/kg) of nicotine region-specifically elevated Fos expression in the piriform cortex (PirC), medial habenula, solitary nucleus and inferior olive (IO) among 44 brain regions examined. In addition, similarly to the tremor responses, nicotine-induced Fos expression in the PirC and IO was selectively antagonized by MLA, but not by DHβE. Furthermore, an electrical lesioning of the IO, but not the PirC, significantly suppressed the induction of nicotine tremor. The present results suggest that nicotine elicits kinetic tremor in rodents by activating the IO neurons via α7 nACh receptors. PMID:27506652

  12. The role of nicotinic receptor alpha 7 subunits in nicotine discrimination.

    PubMed

    Stolerman, I P; Chamberlain, S; Bizarro, L; Fernandes, C; Schalkwyk, L

    2004-03-01

    The subtypes of nicotinic receptors at which the behavioural effects of nicotine originate are not fully understood. The experiments described here use mice lacking the alpha7 subunit of nicotinic receptors to investigate the role of alpha7-containing receptors in nicotine discrimination. Wild-type and alpha7-knockout mice were trained in a two-lever nicotine discrimination procedure using a tandem schedule of food reinforcement. Mutant mice exhibited baseline rates of lever-pressing as low as 52.2% of rates in wild-type controls (n=21-24). Mutant and wild-type mice acquired discrimination of nicotine (0.4 or 0.8 mg/kg) at a similar rate (n=10-12) and reached similar final levels of accuracy (71.9 +/- 4.4% and 90.8 +/- 3.1% after 60 training sessions for 0.4 and 0.8 mg/kg training doses, respectively, in mutant mice, as compared with 75.0 +/- 6.5% and 87.6 +/- 4.8% for wild types). The genotypes exhibited similar steep dose-response curves for nicotine discrimination. In both genotypes, dose-response curves for mice trained with 0.8 mg/kg of nicotine were displaced three- to four-fold to the right as compared with those for the mice trained with the smaller dose. The predominant effect of nicotine on the overall rate of responding was a reduction at the largest doses tested and there was no difference between the genotypes. The results suggest that nicotinic receptors containing the alpha7 subunit do not contribute to the discriminative stimulus or response-rate-depressant effects of nicotine, although they may regulate baseline rates of operant responding. PMID:14975691

  13. Parazoanthoxanthin A blocks Torpedo nicotinic acetylcholine receptors.

    PubMed

    Rozman, Klara Bulc; Araoz, Romulo; Sepcić, Kristina; Molgo, Jordi; Suput, Dusan

    2010-09-01

    Nicotinic acetylcholine receptors are implicated in different nervous system-related disorders, and their modulation could improve existing therapy of these diseases. Parazoanthoxanthin A (ParaA) is a fluorescent pigment of the group of zoanthoxanthins. Since it is a potent acetylcholinesterase inhibitor, it may also bind to nicotinic acetylcholine receptors (nAChRs). For this reason its effect on Torpedo nAChR (alpha1(2)betagammadelta) transplanted to Xenopus laevis oocytes was evaluated, using the voltage-clamp technique. ParaA dose-dependently reduced the acetylcholine-induced currents. This effect was fully reversible only at lower concentrations. ParaA also reduced the Hill coefficient and the time to peak current, indicating a channel blocking mode of action. On the other hand, the combined effect of ParaA and d-tubocurarine (d-TC) on acetylcholine-induced currents exhibited only partial additivity, assuming a competitive mode of action of ParaA on nAChR. These results indicate a dual mode of action of ParaA on the Torpedo AChR. PMID:20230806

  14. Optochemical control of genetically engineered neuronal nicotinic acetylcholine receptors

    NASA Astrophysics Data System (ADS)

    Tochitsky, Ivan; Banghart, Matthew R.; Mourot, Alexandre; Yao, Jennifer Z.; Gaub, Benjamin; Kramer, Richard H.; Trauner, Dirk

    2012-02-01

    Advances in synthetic chemistry, structural biology, molecular modelling and molecular cloning have enabled the systematic functional manipulation of transmembrane proteins. By combining genetically manipulated proteins with light-sensitive ligands, innately ‘blind’ neurobiological receptors can be converted into photoreceptors, which allows them to be photoregulated with high spatiotemporal precision. Here, we present the optochemical control of neuronal nicotinic acetylcholine receptors (nAChRs) with photoswitchable tethered agonists and antagonists. Using structure-based design, we produced heteromeric α3β4 and α4β2 nAChRs that can be activated or inhibited with deep-violet light, but respond normally to acetylcholine in the dark. The generation of these engineered receptors should facilitate investigation of the physiological and pathological functions of neuronal nAChRs and open a general pathway to photosensitizing pentameric ligand-gated ion channels.

  15. Optochemical control of genetically engineered neuronal nicotinic acetylcholine receptors

    PubMed Central

    Tochitsky, Ivan; Banghart, Matthew R.; Mourot, Alexandre; Yao, Jennifer Z.; Gaub, Benjamin

    2016-01-01

    Advances in synthetic chemistry, structural biology, molecular modelling and molecular cloning have enabled the systematic functional manipulation of transmembrane proteins. By combining genetically manipulated proteins with light-sensitive ligands, innately ‘blind’ neurobiological receptors can be converted into photoreceptors, which allows them to be photoregulated with high spatiotemporal precision. Here, we present the optochemical control of neuronal nicotinic acetylcholine receptors (nAChRs) with photoswitchable tethered agonists and antagonists. Using structure-based design, we produced heteromeric α3β4 and α4β2 nAChRs that can be activated or inhibited with deep-violet light, but respond normally to acetylcholine in the dark. The generation of these engineered receptors should facilitate investigation of the physiological and pathological functions of neuronal nAChRs and open a general pathway to photosensitizing pentameric ligand-gated ion channels. PMID:22270644

  16. Nicotinic receptor modulation to treat alcohol and drug dependence

    PubMed Central

    Rahman, Shafiqur; Engleman, Eric A.; Bell, Richard L.

    2015-01-01

    Alcohol and drug dependence are serious public health problems worldwide. The prevalence of alcohol and drug dependence in the United States and other parts of the world is significant. Given the limitations in the efficacy of current pharmacotherapies to treat these disorders, research in developing alternative pharmacotherapies continues. Preclinical and clinical evidence thus far has indicated that brain nicotinic acetylcholine receptors (nAChRs) are important pharmacological targets for the development of medications to treat alcohol and drug dependence. The nAChRs are a super family of ligand gated ion channels, and are expressed throughout the brain with twelve neuronal nAChR subunits (α2–α10 and β2–β4) identified. Here, we review preclinical and clinical evidence involving a number of nAChR ligands that target different nAChR subtypes in alcohol and nicotine addiction. The important ligands include cytisine, lobeline, mecamylamine, varenicline, sazetidine A and others that target α4β2* nAChR subtypes as small molecule modulators of the brain nicotinic cholinergic system are also discussed. Taken together, both preclinical and clinical data exist that support nAChR–based ligands as promising therapeutic agents for the treatment of alcohol and drug dependence. PMID:25642160

  17. Nicotine induces fibrogenic changes in human liver via nicotinic acetylcholine receptors expressed on hepatic stellate cells

    SciTech Connect

    Soeda, Junpei; Morgan, Maelle; McKee, Chad; Mouralidarane, Angelina; Lin, ChingI; Roskams, Tania; Oben, Jude A.

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Cigarette smoke may induce liver fibrosis via nicotine receptors. Black-Right-Pointing-Pointer Nicotine induces proliferation of hepatic stellate cells (HSCs). Black-Right-Pointing-Pointer Nicotine activates hepatic fibrogenic pathways. Black-Right-Pointing-Pointer Nicotine receptor antagonists attenuate HSC proliferation. Black-Right-Pointing-Pointer Nicotinic receptor antagonists may have utility as novel anti-fibrotic agents. -- Abstract: Background and aims: Cigarette smoke (CS) may cause liver fibrosis but possible involved mechanisms are unclear. Among the many chemicals in CS is nicotine - which affects cells through nicotinic acetylcholine receptors (nAChR). We studied the effects of nicotine, and involved pathways, on human primary hepatic stellate cells (hHSCs), the principal fibrogenic cells in the liver. We then determined possible disease relevance by assaying nAChR in liver samples from human non-alcoholic steatohepatitis (NASH). Methods: hHSC were isolated from healthy human livers and nAChR expression analyzed - RT-PCR and Western blotting. Nicotine induction of hHSC proliferation, upregulation of collagen1-{alpha}2 and the pro-fibrogenic cytokine transforming growth factor beta 1 (TGF-{beta}1) was determined along with involved intracellular signaling pathways. nAChR mRNA expression was finally analyzed in whole liver biopsies obtained from patients diagnosed with non-alcoholic steatohepatitis (NASH). Results: hHSCs express muscle type ({alpha}1, {beta}1, delta and epsilon) and neuronal type ({alpha}3, {alpha}6, {alpha}7, {beta}2 and {beta}4) nAChR subunits at the mRNA level. Among these subunits, {alpha}3, {alpha}7, {beta}1 and {epsilon} were predominantly expressed as confirmed by Western blotting. Nicotine induced hHSC proliferation was attenuated by mecamylamine (p < 0.05). Additionally, collagen1-{alpha}2 and TGF-{beta}1 mRNA expression were significantly upregulated by nicotine and inhibited by

  18. CHARACTERIZATION OF NICOTINE ACETYLCHOLINE RECEPTOR SUBUNITS IN THE COCKROACH Periplaneta americana MUSHROOM BODIES REVEALS A STRONG EXPRESSION OF β1 SUBUNIT: INVOLVEMENT IN NICOTINE-INDUCED CURRENTS.

    PubMed

    Taillebois, Emiliane; Thany, Steeve H

    2016-09-01

    Nicotinic acetylcholine receptors are ligand-gated ion channels expressed in many insect structures, such as mushroom bodies, in which they play a central role. We have recently demonstrated using electrophysiological recordings that different native nicotinic receptors are expressed in cockroach mushroom bodies Kenyon cells. In the present study, we demonstrated that eight genes coding for cockroach nicotinic acetylcholine receptor subunits are expressed in the mushroom bodies. Quantitative real-time polymerase chain reaction (PCR) experiments demonstrated that β1 subunit was the most expressed in the mushroom bodies. Moreover, antisense oligonucleotides performed against β1 subunit revealed that inhibition of β1 expression strongly decreases nicotine-induced currents amplitudes. Moreover, co-application with 0.5 μM α-bungarotoxin completely inhibited nicotine currents whereas 10 μM d-tubocurarine had a partial effect demonstrating that β1-containing neuronal nicotinic acetylcholine receptor subtypes could be sensitive to the nicotinic acetylcholine receptor antagonist α-bungarotoxin. PMID:27357353

  19. Effects of prenatal nicotine on expression of nicotine receptor subunits in the fetal brain

    PubMed Central

    Lv, Juanxiu; Mao, Caiping; Zhu, Liyan; Zhang, Hong; Pengpeng, Hui; Xu, Feichao; Liu, Yujuan; Zhang, Lubo; Xu, Zhice

    2008-01-01

    Previous studies have suggested that prenatal exposure to nicotine is associated with abnormal development in fetuses, including fetal brain damage. The present study determined the effect of maternal administration of nicotine during different gestational periods on brain nicotine receptor subunits in fetal rats. Subcutaneous injections of nicotine in maternal rats from the early and middle gestation decreased fetal blood PO2, increased fetal blood PCO2 and hemoglobin, and decreased fetal brain weight. The nicotinic acetylcholine receptor (nAChRs) mRNA abundance in the fetal brain was significantly changed by prenatal treatment with nicotine during pregnancy. Fetal α2, α4, α7, and β2 units were significantly increased in the brain by prenatal exposure to nicotine in rat fetuses. However, the expression of mRNA of fetal brain α3, α5, β3, and β4 units were not changed. The results showed that prenatal nicotine can change the development of both α and β subunits of nAChRs in the fetal brain at gene level in association with restriction of fetal brain growth and in utero hypoxia. PMID:18541304

  20. Habenula cholinergic neurons regulate anxiety during nicotine withdrawal via nicotinic acetylcholine receptors.

    PubMed

    Pang, Xueyan; Liu, Liwang; Ngolab, Jennifer; Zhao-Shea, Rubing; McIntosh, J Michael; Gardner, Paul D; Tapper, Andrew R

    2016-08-01

    Cholinergic neurons in the medial habenula (MHb) modulate anxiety during nicotine withdrawal although the molecular neuroadaptation(s) within the MHb that induce affective behaviors during nicotine cessation is largely unknown. MHb cholinergic neurons are unique in that they robustly express neuronal nicotinic acetylcholine receptors (nAChRs), although their behavioral role as autoreceptors in these neurons has not been described. To test the hypothesis that nAChR signaling in MHb cholinergic neurons could modulate anxiety, we expressed novel "gain of function" nAChR subunits selectively in MHb cholinergic neurons of adult mice. Mice expressing these mutant nAChRs exhibited increased anxiety-like behavior that was alleviated by blockade with a nAChR antagonist. To test the hypothesis that anxiety induced by nicotine withdrawal may be mediated by increased MHb nicotinic receptor signaling, we infused nAChR subtype selective antagonists into the MHb of nicotine naïve and withdrawn mice. While antagonists had little effect on nicotine naïve mice, blocking α4β2 or α6β2, but not α3β4 nAChRs in the MHb alleviated anxiety in mice undergoing nicotine withdrawal. Consistent with behavioral results, there was increased functional expression of nAChRs containing the α6 subunit in MHb neurons that also expressed the α4 subunit. Together, these data indicate that MHb cholinergic neurons regulate nicotine withdrawal-induced anxiety via increased signaling through nicotinic receptors containing the α6 subunit and point toward nAChRs in MHb cholinergic neurons as molecular targets for smoking cessation therapeutics. PMID:27020042

  1. Roles of nicotinic acetylcholine receptor β subunits in function of human α4-containing nicotinic receptors

    PubMed Central

    Wu, Jie; Liu, Qiang; Yu, Kewei; Hu, Jun; Kuo, Yen-Ping; Segerberg, Marsha; St John, Paul A; Lukas, Ronald J

    2006-01-01

    Naturally expressed nicotinic acetylcholine receptors (nAChR) containing α4 subunits (α4*-nAChR) in combination with β2 subunits (α4β2-nAChR) are among the most abundant, high-affinity nicotine binding sites in the mammalian brain. β4 subunits are also richly expressed and colocalize with α4 subunits in several brain regions implicated in behavioural responses to nicotine and nicotine dependence. Thus, α4β4-nAChR also may exist and play important functional roles. In this study, properties were determined of human α4β2- and α4β4-nAChR heterologously expressed de novo in human SH-EP1 epithelial cells. Whole-cell currents mediated via human α4β4-nAChR have ∼4-fold higher amplitude than those mediated via human α4β2-nAChR and exhibit much slower acute desensitization and functional rundown. Nicotinic agonists induce peak whole-cell current responses typically with higher functional potency at α4β4-nAChR than at α4β2-nAChR. Cytisine and lobeline serve as full agonists at α4β4-nAChR but are only partial agonists at α4β2-nAChR. However, nicotinic antagonists, except hexamethonium, have comparable affinities for functional α4β2- and α4β4-nAChR. Whole-cell current responses show stronger inward rectification for α4β2-nAChR than for α4β4-nAChR at a positive holding potential. Collectively, these findings demonstrate that human nAChR β2 or β4 subunits can combine with α4 subunits to generate two forms of α4*-nAChR with distinctive physiological and pharmacological features. Diversity in α4*-nAChR is of potential relevance to nervous system function, disease, and nicotine dependence. PMID:16825297

  2. α4β2 nicotinic acetylcholine receptors on dopaminergic neurons mediate nicotine reward and anxiety relief.

    PubMed

    McGranahan, Tresa M; Patzlaff, Natalie E; Grady, Sharon R; Heinemann, Stephen F; Booker, T K

    2011-07-27

    Nicotine is the primary psychoactive substance in tobacco, and it exerts its effects by interaction with various subtypes of nicotinic acetylcholine receptors (nAChRs) in the brain. One of the major subtypes expressed in brain, the α4β2-nAChR, endogenously modulates neuronal excitability and thereby, modifies certain normal as well as nicotine-induced behaviors. Although α4-containing nAChRs are widely expressed across the brain, a major focus has been on their roles within midbrain dopaminergic regions involved in drug addiction, mental illness, and movement control in humans. We developed a unique model system to examine the role of α4-nAChRs within dopaminergic neurons by a targeted genetic deletion of the α4 subunit from dopaminergic neurons in mice. The loss α4 mRNA and α4β2-nAChRs from dopaminergic neurons was confirmed, as well as selective loss of α4β2-nAChR function from dopaminergic but not GABAergic neurons. Two behaviors central to nicotine dependence, reward and anxiety relief, were examined. α4-nAChRs specifically on dopaminergic neurons were demonstrated to be necessary for nicotine reward as measured by nicotine place preference, but not for another drug of addiction, cocaine. α4-nAChRs are necessary for the anxiolytic effects of nicotine in the elevated plus maze, and elimination of α4β2-nAChRs specifically from dopaminergic neurons decreased sensitivity to the anxiolytic effects of nicotine. Deletion of α4-nAChRs specifically from dopaminergic neurons also increased sensitivity to nicotine-induced locomotor depression; however, nicotine-induced hypothermia was unaffected. This is the first work to develop a dopaminergic specific deletion of a nAChR subunit and examine resulting changes in nicotine-related behaviors. PMID:21795541

  3. alpha4beta2 nicotinic acetylcholine receptors on dopaminergic neurons mediate nicotine reward and anxiety relief

    PubMed Central

    McGranahan, Tresa M.; Patzlaff, Natalie E.; Grady, Sharon R.; Heinemann, Stephen F.; Booker, T.K.

    2012-01-01

    Nicotine is the primary psychoactive substance in tobacco and it exerts its effects by interaction with various subtypes of nicotinic acetylcholine receptors (nAChRs) in the brain. One of the major subtypes expressed in brain, the alpha4beta2-nAChR, endogenously modulates neuronal excitability and thereby, modifies certain normal, as well as nicotine-induced, behaviors. Although alpha4-containing nAChRs are widely expressed across the brain, a major focus has been on their roles within midbrain dopaminergic regions involved in drug addition, mental illness and movement control in humans. We developed a unique model system to examine the role of alpha4-nAChRs within dopaminergic neurons by a targeted genetic deletion of the alpha4 subunit from dopaminergic neurons in mice. The loss alpha4 mRNA and alpha4beta2-nAChRs from dopaminergic neurons was confirmed, as well as selective loss of alpha4beta2-nAChR function from dopaminergic but not GABAergic neurons. Two behaviors central to nicotine dependence, reward and anxiety relief, were examined. Alpha4-nAChRs specifically on dopaminergic neurons were demonstrated to be necessary for nicotine reward as measured by nicotine place preference, but not for another drug of addiction, cocaine. Alpha4-nAChRs are necessary for the anxiolytic effects of nicotine in the elevated plus maze and elimination of alpha4-beta2-nAChRs specifically from dopaminergic neurons decreased sensitivity to the anxiolytic effects of nicotine. Deletion of alpha4-nAChRs specifically from dopaminergic neurons also increased sensitivity to nicotine-induced locomotor depression, however nicotine-induced hypothermia was unaffected. This is the first work to develop a dopaminergic specific deletion of a nAChR subunit and examine resulting changes in nicotine behaviors. PMID:21795541

  4. Schizophrenia and the alpha7 nicotinic acetylcholine receptor.

    PubMed

    Martin, Laura F; Freedman, Robert

    2007-01-01

    In addition to the devastating symptoms of psychosis, many people with schizophrenia also suffer from cognitive impairment. These cognitive symptoms lead to marked dysfunction and can impact employability, treatment adherence, and social skills. Deficits in P50 auditory gating are associated with attentional impairment and may contribute to cognitive symptoms and perceptual disturbances. This nicotinic cholinergic-mediated inhibitory process represents a potential new target for therapeutic intervention in schizophrenia. This chapter will review evidence implicating the nicotinic cholinergic, and specifically, the alpha7 nicotinic receptor system in the pathology of schizophrenia. Impaired auditory sensory gating has been linked to the alpha7 nicotinic receptor gene on the chromosome 15q14 locus. A majority of persons with schizophrenia are heavy smokers. Although nicotine can acutely reverse diminished auditory sensory gating in people with schizophrenia, this effect is lost on a chronic basis due to receptor desensitization. The alpha7 nicotinic agonist 3-(2,4 dimethoxy)benzylidene-anabaseine (DMXBA) can also enhance auditory sensory gating in animal models. DMXBA is well tolerated in humans and a new study in persons with schizophrenia has found that DMXBA enhances both P50 auditory gating and cognition. alpha7 Nicotinic acetylcholine receptor agonists appear to be viable candidates for the treatment of cognitive disturbances in schizophrenia. PMID:17349863

  5. Photoaffinity labeling of insect nicotinic acetylcholine receptors with a novel [(3)H]azidoneonicotinoid.

    PubMed

    Tomizawa, M; Wen, Z; Chin, H L; Morimoto, H; Kayser, H; Casida, J E

    2001-09-01

    The nicotinic acetylcholine receptor (nAChR) is a ligand-gated ion channel in the insect CNS and a target for major insecticides. Here we use photoaffinity labeling to approach the functional architecture of insect nAChRs. Two candidate 5-azido-6-chloropyridin-3-yl photoaffinity probes are evaluated for their receptor potencies: azidoneonicotinoid (AzNN) with an acyclic nitroguanidine moiety; azidodehydrothiacloprid. Compared to their non-azido parents, both probes are of decreased potencies at Drosophila (fruit fly) and Musca (housefly) receptors but AzNN retains full potency at the Myzus (aphid) receptor. [(3)H]AzNN was therefore radiosynthesized at high specific activity (84 Ci/mmol) as a novel photoaffinity probe. [(3)H]AzNN binds to a single high-affinity site in Myzus that is competitively inhibited by imidacloprid and nicotine and further characterized as to its pharmacological profile with various nicotinic ligands. [(3)H]AzNN photoaffinity labeling of Myzus and Homalodisca (leafhopper) detects a single radiolabeled peak in each case displaceable with imidacloprid and nicotine and with molecular masses corresponding to approximately 45 and approximately 56 kDa, respectively. The photoaffinity-labeled receptor in both Drosophila and Musca has imidacloprid- and nicotine-sensitive profiles and migrates at approximately 66 kDa. These photoaffinity-labeled polypeptides are considered to be the insecticide-binding subunits of native insect nAChRs. PMID:11579144

  6. Decreased nicotinic receptor availability in smokers with slow rates of nicotine metabolism

    PubMed Central

    Dubroff, Jacob G.; Doot, Robert K.; Falcone, Mary; R, Robert A. Schnoll; Ray, Riju; Tyndale, Rachel F.; Brody, Arthur L.; Hou, Catherine; Schmitz, Alexander; Lerman, Caryn

    2015-01-01

    The nicotine metabolite ratio (NMR), a stable measure of hepatic nicotine metabolism via the CYP2A6 pathway and total nicotine clearance, is a predictive biomarker of response to nicotine replacement therapy, with increased quit rates in slower metabolizers. Nicotine binds directly to nicotinic acetylcholine receptors (nAChRs) to exert its psychoactive effects. This study examined the relationship between NMR and nAChR availability (α4β2* subtype) using positron emission tomography (PET) imaging of the radiotracer 2-18F-FA-85380 (2-18F-FA). Methods Twenty four smokers, 12 slow metabolizers (NMR <0.26) and 12 normal metabolizers (NMR ≥0.26), underwent 2-18F-FA-PET brain imaging following overnight nicotine abstinence (18 hours prior to scanning), using a validated bolus plus infusion protocol. Availability of nAChRs was compared between NMR groups in a priori volumes of interest (VOIs), with total distribution volume (VT/fP) being the measure of nAChR availability. Cravings to smoke were assessed prior to and following the scans. Results Thalamic nAChR α4β2* availability was significantly reduced in slow (versus normal) nicotine metabolizers (P=0.04). Slow metabolizers exhibited greater reductions in craving than normal metabolizers from pre- to post-scanning; however, craving was unrelated to availability. Conclusion The rate of nicotine metabolism is associated with thalamic nAChR availability. Additional studies could examine whether altered nAChR availability underlies differences in treatment response between slow and normal metabolizers of nicotine. PMID:26272810

  7. Role of the D3 dopamine receptor in nicotine sensitization.

    PubMed

    Smith, Laura N; Bachus, Susan E; McDonald, Craig G; Smith, Robert F

    2015-08-01

    Adolescent cigarette use is associated with reduced quitting success and continued smoking in adulthood. Interestingly, polymorphisms of the dopamine D3 receptor (DRD3) gene have been associated with smoking behavior, and the receptor is expressed in an age- and brain region-dependent manner that suggests relevance to addiction. Here, we investigate the possible role of dopamine-related receptors, including DRD3 and an intriguing splice variant known as D3nf, in nicotine-induced sensitization. In adolescent and adult male rats, we examined (1) alterations occurring in dopamine receptor-related mRNAs (DRD1, DRD2, DRD3 and D3nf) at two time points during a sensitizing regimen of nicotine and (2) whether DRD3 antagonism either during the initial treatment (induction) or at a later challenge exposure (expression) is able to block nicotine sensitization. Nicotine-induced changes were seen for DRD3 and D3nf mRNAs in the nucleus accumbens shell early in repeated exposure in both age groups. DRD3 antagonism only blocked the induction of sensitization in adolescents and did not block the expression of sensitization in either age group. Adolescents and adults showed opposite DRD1 mRNA responses to nicotine treatment, while no age- and nicotine-related changes in DRD2 mRNA were observed. These data reveal important age-dependent regulation of DRD1- and DRD3-related mRNAs during the course of nicotine exposure. Furthermore, they highlight a requirement for DRD3 signaling in the development of adolescent nicotine sensitization, suggesting it may represent an appropriate target in the prevention of nicotine dependence initiated at this age. PMID:25907750

  8. Recent developments in the synthesis of nicotinic acetylcholine receptor ligands.

    PubMed

    Breining, Scott R

    2004-01-01

    The extraordinary pharmacology of nicotine and epibatidine have indicated the potential for nicotinic acetylcholine receptor (nAChR) ligands to serve as a new therapeutic class for a host of CNS disorders. Many such ligands are natural products, or analogs thereof, which represent a significant challenge to the synthetic chemist. Synthesis of such molecules often serves as a showcase to demonstrate the potential of newly developed methodology. This synthetic challenge coupled with the promise of pharmacological activity in compounds possessing the nicotinic pharmacophore has stimulated a great deal of synthetic activity over the last five years. The present report provides an overview of novel synthetic methodology occurring during this period directed toward the synthesis of compounds with presumed affinity for the neuronal nAChR. Syntheses chosen for review here represent the major efforts toward molecules such as epibatidine analogs, anatoxin-a, nicotine and related alkaloids, conformationally constrained nicotine derivatives, cytisine and methyllycaconitine (MLA). PMID:14965298

  9. Disruption of nicotine conditioning by dopamine D(3) receptor ligands.

    PubMed

    Le Foll, B; Schwartz, J-C; Sokoloff, P

    2003-02-01

    Tobacco smoking is the first cause of preventable death in modern countries. Nicotine replacement therapy or sustained release bupropion helps smoking cessation, but relapse rates are still very high. Nicotine, like other drugs of abuse, activates the dopamine mesolimbic system, which originates in the ventral tegmental area and projects notably to the nucleus accumbens. Situations or environmental stimuli previously associated with cigarette smoking, for example, smell of cigarette smoke, can elicit craving in abstinent smokers and promote relapse. Reducing the effects of nicotine-associated cues might therefore have potential therapeutic utility for smoking cessation. Such an approach has been validated for cocaine in animals, by using the dopamine D(3) receptor-selective partial agonist BP 897, which inhibits cocaine cue-induced drug-seeking behavior. Here we show that rats repeatedly injected with nicotine in a particular environment develop nicotine-conditioned locomotor responses, accompanied by an increase in D(3) receptor expression in the nucleus accumbens. This conditioned behavior was inhibited by BP 897 or a selective D(3) receptor antagonist, suggesting that antagonizing dopamine selectively at the D(3) receptor disrupts nicotine-conditioned effects and might represent a novel therapeutic approach for smoking cessation. PMID:12610655

  10. Photolabeling reveals the proximity of the alpha-neurotoxin binding site to the M2 helix of the ion channel in the nicotinic acetylcholine receptor.

    PubMed Central

    Machold, J; Utkin, Y; Kirsch, D; Kaufmann, R; Tsetlin, V; Hucho, F

    1995-01-01

    A photoactivatable derivative of neurotoxin II from Naja naja oxiana containing a 125I-labeled p-azidosalicylamidoethyl-1,3'-dithiopropyl label at Lys-25 forms a photo-induced cross-link with the delta subunit of the membrane-bound Torpedo californica nicotinic acetylcholine receptor (AChR). The cross-linked radioactive receptor peptide was isolated by reverse-phase HPLC after tryptic digestion of the labeled delta subunit. The sequence of this peptide, delta-(260-277), and the position of the label at Ala-268 were established by matrix-assisted laser-desorption-ionization mass spectrometry based on the molecular mass and on post-source decay fragment analysis. With the known dimensions of the AChR molecule, of the photolabel, and of alpha-neurotoxin, finding the cross-link at delta Ala-268 (located in the upper part of the channel-forming transmembrane helix M2) means that the center of the alpha-neurotoxin binding site is situated at least approximately 40 A from the extracellular surface of the AChR, proximal to the channel axis. Images Fig. 2 PMID:7543679

  11. Functional interaction between Lypd6 and nicotinic acetylcholine receptors.

    PubMed

    Arvaniti, Maria; Jensen, Majbrit M; Soni, Neeraj; Wang, Hong; Klein, Anders B; Thiriet, Nathalie; Pinborg, Lars H; Muldoon, Pretal P; Wienecke, Jacob; Imad Damaj, M; Kohlmeier, Kristi A; Gondré-Lewis, Marjorie C; Mikkelsen, Jens D; Thomsen, Morten S

    2016-09-01

    Nicotinic acetylcholine receptors (nAChRs) affect multiple physiological functions in the brain and their functions are modulated by regulatory proteins of the Lynx family. Here, we report for the first time a direct interaction of the Lynx protein LY6/PLAUR domain-containing 6 (Lypd6) with nAChRs in human brain extracts, identifying Lypd6 as a novel regulator of nAChR function. Using protein cross-linking and affinity purification from human temporal cortical extracts, we demonstrate that Lypd6 is a synaptically enriched membrane-bound protein that binds to multiple nAChR subtypes in the human brain. Additionally, soluble recombinant Lypd6 protein attenuates nicotine-induced hippocampal inward currents in rat brain slices and decreases nicotine-induced extracellular signal-regulated kinase phosphorylation in PC12 cells, suggesting that binding of Lypd6 is sufficient to inhibit nAChR-mediated intracellular signaling. We further show that perinatal nicotine exposure in rats (4 mg/kg/day through minipumps to dams from embryonic day 7 to post-natal day 21) significantly increases Lypd6 protein levels in the hippocampus in adulthood, which did not occur after exposure to nicotine in adulthood only. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain, and that Lypd6 is dysregulated by nicotine exposure during early development. Regulatory proteins of the Lynx family modulate the function of nicotinic receptors (nAChRs). We report for the first time that the Lynx protein Lypd6 binds to nAChRs in human brain extracts, and that recombinant Lypd6 decreases nicotine-induced ERK phosphorylation and attenuates nicotine-induced hippocampal inward currents. Our findings suggest that Lypd6 is a versatile inhibitor of cholinergic signaling in the brain. PMID:27344019

  12. Inhibitory Learning is Modulated by Nicotinic Acetylcholine Receptors

    PubMed Central

    Meyer, Heidi C.; Putney, Rachel B.; Bucci, David J.

    2014-01-01

    Prior research has established that stimulating nicotinic acetylcholine receptors can facilitate learning and memory. However, most studies have focused on learning to emit a particular behavior, while little is known about the effects of nicotine on learning to withhold a behavioral response. The present study consisted of a dose response analysis of the effects of nicotine on negative occasion setting, a form of learned inhibition. In this paradigm, rats received one type of training trial in which presentation of a tone by itself was followed immediately by food reward. During the other type of trials, the tone was preceded by presentation of a light and no food was delivered after the tone. Rats gradually learned to approach the cup in anticipation of receiving food reward during presentations of the tone alone, but withheld that behavior when the tone was preceded by the light. Nicotine (0.35mg/kg) facilitated negative occasion setting by reducing the number of sessions needed to learn the discrimination between trial types and by reducing the rate of responding on non-reinforced trials. Nicotine also increased the orienting response to the light, suggesting that nicotine may have affected the ability to withhold food cup behavior on non-reinforced trials by increasing attention to the light. In contrast to the effects of nicotine, rats treated with mecamylamine (0.125, 0.5, or 2 mg/kg) needed more training sessions to discriminate between reinforced and non-reinforced trials compared to saline-treated rats. The findings indicate that nicotinic acetylcholine receptors may be active during negative occasion setting and that nicotine can potentiate learned inhibition. PMID:25445487

  13. Spectral Confocal Imaging of Fluorescently tagged Nicotinic Receptors in Knock-in Mice with Chronic Nicotine Administration

    PubMed Central

    Renda, Anthony; Nashmi, Raad

    2012-01-01

    Ligand-gated ion channels in the central nervous system (CNS) are implicated in numerous conditions with serious medical and social consequences. For instance, addiction to nicotine via tobacco smoking is a leading cause of premature death worldwide (World Health Organization) and is likely caused by an alteration of ion channel distribution in the brain1. Chronic nicotine exposure in both rodents and humans results in increased numbers of nicotinic acetylcholine receptors (nAChRs) in brain tissue1-3. Similarly, alterations in the glutamatergic GluN1 or GluA1 channels have been implicated in triggering sensitization to other addictive drugs such as cocaine, amphetamines and opiates4-6. Consequently, the ability to map and quantify distribution and expression patterns of specific ion channels is critically important to understanding the mechanisms of addiction. The study of brain region-specific effects of individual drugs was advanced by the advent of techniques such as radioactive ligands. However, the low spatial resolution of radioactive ligand binding prevents the ability to quantify ligand-gated ion channels in specific subtypes of neurons. Genetically encoded fluorescent reporters, such as green fluorescent protein (GFP) and its many color variants, have revolutionized the field of biology7.By genetically tagging a fluorescent reporter to an endogenous protein one can visualize proteins in vivo7-10. One advantage of fluorescently tagging proteins with a probe is the elimination of antibody use, which have issues of nonspecificity and accessibility to the target protein. We have used this strategy to fluorescently label nAChRs, which enabled the study of receptor assembly using Förster Resonance Energy Transfer (FRET) in transfected cultured cells11.More recently, we have used the knock-in approach to engineer mice with yellow fluorescent protein tagged α4 nAChR subunits (α4YFP), enabling precise quantification of the receptor ex vivo at submicrometer

  14. The α3β4* nicotinic acetylcholine receptor subtype mediates nicotine reward and physical nicotine withdrawal signs independently of the α5 subunit in the mouse

    PubMed Central

    Jackson, Kia J.; Sanjakdar, Sarah S.; Muldoon, Pretal P.; McIntosh, J. Michael; Damaj, M. Imad

    2013-01-01

    The 15q25 gene cluster contains genes that code for the α5, α3, and β4 nicotinic acetylcholine receptor (nAChRs) subunits, and in human genetic studies, has shown the most robust association with smoking behavior and nicotine dependence to date. The limited available animal studies implicate a role for the α5 and β4 nAChR subunits in nicotine dependence and withdrawal; however studies focusing on the behavioral role of the α3β4* nAChR receptor subtype in nicotine dependence are lacking. Because of the apparent role of the α3β4* nAChR subtype in nicotine dependence, the goal of the current study was to better evaluate the involvement of this subtype in nicotine mediated behavioral responses. Using the selective α3β4* nAChR antagonist, α-conotoxin AuIB, we assessed the role of α3β4* nAChRs in acute nicotine, nicotine reward, and physical and affective nicotine withdrawal. Because α5 has also been implicated in nicotine dependence behaviors in mice and can form functional receptors with α3β4*, we also evaluated the role of the α3β4α5* nAChR subtype in nicotine reward and somatic nicotine withdrawal signs by blocking the α3β4* nAChR subtype in α5 nAChR knockout mice with AuIB. AuIB had no significant effect on acute nicotine behaviors, but dose-dependently attenuated nicotine reward and physical withdrawal signs, with no significant effect in affective withdrawal measures. Interestingly, AuIB also attenuated nicotine reward and somatic signs in α5 nAChR knockout mice. This study shows that α3β4* nAChRs mediate nicotine reward and physical nicotine withdrawal, but not acute nicotine behaviors or affective nicotine withdrawal signs in mice. The α5 subunit is not required in the receptor assembly to mediate these effects. Our findings suggest an important role for the α3β4* nAChR subtype in nicotine reward and physical aspects of the nicotine withdrawal syndrome. PMID:23416040

  15. Nicotine increases GABAergic input on rat dorsal raphe serotonergic neurons through alpha7 nicotinic acetylcholine receptor.

    PubMed

    Hernández-Vázquez, F; Chavarría, K; Garduño, J; Hernández-López, S; Mihailescu, S P

    2014-12-15

    The dorsal raphe nucleus (DRN) contains large populations of serotonergic (5-HT) neurons. This nucleus receives GABAergic inhibitory afferents from many brain areas and from DRN interneurons. Both GABAergic and 5-HT DRN neurons express functional nicotinic acetylcholine receptors (nAChRs). Previous studies have demonstrated that nicotine increases 5-HT release and 5-HT DRN neuron discharge rate by stimulating postsynaptic nAChRs and by increasing glutamate and norepinephrine release inside DRN. However, the influence of nicotine on the GABAergic input to 5-HT DRN neurons was poorly investigated. Therefore, the aim of this work was to determine the effect of nicotine on GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) of 5-HT DRN neurons and the subtype of nAChR(s) involved in this response. Experiments were performed in coronal slices obtained from young Wistar rats. GABAergic sIPSCs were recorded from post hoc-identified 5-HT DRN neurons with the whole cell voltage patch-clamp technique. Administration of nicotine (1 μM) increased sIPSC frequency in 72% of identified 5-HT DRN neurons. This effect was not reproduced by the α4β2 nAChR agonist RJR-2403 and was not influenced by TTX (1 μM). It was mimicked by the selective agonist for α7 nAChR, PNU-282987, and exacerbated by the positive allosteric modulator of the same receptor, PNU-120596. The nicotine-induced increase in sIPSC frequency was independent on voltage-gated calcium channels and dependent on Ca(2+)-induced Ca(2+) release (CICR). These results demonstrate that nicotine increases the GABAergic input to most 5-HT DRN neurons, by activating α7 nAChRs and producing CICR in DRN GABAergic terminals. PMID:25231613

  16. Multiple cholinergic nicotinic receptor genes affect nicotine dependence risk in African and European Americans

    PubMed Central

    Saccone, Nancy L.; Schwantes-An, Tae-Hwi; Wang, Jen C.; Grucza, Richard A.; Breslau, Naomi; Hatsukami, Dorothy; Johnson, Eric O.; Rice, John P.; Goate, Alison M.; Bierut, Laura J.

    2010-01-01

    Several independent studies show that the chromosome 15q25.1 region, which contains the CHRNA5-CHRNA3-CHRNB4 gene cluster, harbors variants strongly associated with nicotine dependence, other smoking behaviors, lung cancer, and chronic obstructive pulmonary disease. We investigated whether variants in other cholinergic nicotinic receptor subunit (CHRN) genes affect risk for nicotine dependence in a new sample of African-Americans (N = 710). We also analyzed this African-American sample together with a European-American sample (N=2062, 1608 of which have been previously studied), allowing for differing effects in the two populations. Cases are current nicotine-dependent smokers and controls are non-dependent smokers. Variants in or near CHRND-CHRNG, CHRNA7, and CHRNA10 show modest association with nicotine dependence risk in the African-American sample. In addition, CHRNA4, CHRNB3-CHRNA6, and CHRNB1 show association in at least one population. CHRNG and CHRNA4 harbor SNPs that have opposite directions of effect in the two populations. In each of the population samples, these loci substantially increase the trait variation explained, although no loci meet Bonferroni-corrected significance in the African-American sample alone. The trait variation explained by three key associated SNPs in CHRNA5-CHRNA3-CHRNB4 is 1.9% in European-Americans and also 1.9% in African-Americans; this increases to 4.5% in EAs and 7.3% in AAs when we add six variants representing associations at other CHRN genes. Multiple nicotinic receptor subunit genes outside of chromosome 15q25 are likely to be important in the biological processes and development of nicotine dependence, and some of these risks may be shared across diverse populations. PMID:20584212

  17. Activation of endplate nicotinic acetylcholine receptors by agonists.

    PubMed

    Auerbach, Anthony

    2015-10-15

    The interaction of a small molecule made in one cell with a large receptor made in another is the signature event of cell signaling. Understanding the structure and energy changes associated with agonist activation is important for engineering drugs, receptors and synapses. The nicotinic acetylcholine receptor (AChR) is a ∼300kD ion channel that binds the neurotransmitter acetylcholine (ACh) and other cholinergic agonists to elicit electrical responses in the central and peripheral nervous systems. This mini-review is in two sections. First, general concepts of skeletal muscle AChR operation are discussed in terms of energy landscapes for conformational change. Second, adult vs. fetal AChRs are compared with regard to interaction energies between ACh and agonist-site side chains, measured by single-channel electrophysiology and molecular dynamics simulations. The five aromatic residues that form the core of each agonist binding site can be divided into two working groups, a triad (led by αY190) that behaves similarly at all sites and a coupled pair (led by γW55) that has a large influence on affinity only in fetal AChRs. Each endplate AChR has 5 homologous subunits, two of α(1) and one each of β, δ, and either γ (fetal) or ϵ (adult). These nicotinic AChRs have only 2 functional agonist binding sites located in the extracellular domain, at αδ and either αγ or αϵ subunit interfaces. The receptor undergoes a reversible, global isomerization between structures called C and O. The C shape does not conduct ions and has a relatively low affinity for ACh, whereas O conducts cations and has a higher affinity. When both agonist sites are empty (filled only with water) the probability of taking on the O conformation (PO) is low, <10(-6). When ACh molecules occupy the agonist sites the C→O opening rate constant and C↔O gating equilibrium constant increase dramatically. Following a pulse of ACh at the nerve-muscle synapse, the endplate current rises rapidly

  18. Agonist and antagonist effects of nicotine on chick neuronal nicotinic receptors are defined by alpha and beta subunits.

    PubMed

    Hussy, N; Ballivet, M; Bertrand, D

    1994-09-01

    1. Functional neuronal nicotinic receptors were reconstituted in Xenopus oocytes by the nuclear injection of different combinations of chick and rat cDNAs encoding alpha and beta subunits. The pharmacology of these nicotinic receptors was investigated using two-electrode voltage clamp. 2. The sensitivity of the chick alpha 3/beta 2, alpha 3/beta 4, and alpha 4/beta 2 receptors to acetylcholine (ACh) and neuronal bungarotoxin differed markedly, indicating that both subunits contribute to the pharmacological properties of the receptors. 3. Nicotine acted as an agonist on the chick alpha 3/beta 4 and alpha 4/beta 2 receptors and rat alpha 3/beta 2 receptor. In contrast, nicotine (at concentrations > 3 microM) was only a weak partial agonist of the chick alpha 3/beta 2 receptor. Moreover, nicotine coapplied with 3 microM ACh on the chick alpha 3/beta 2 receptor acted as a potent competitive antagonist, with an IC50 of 0.43 microM. No antagonist effect of nicotine could be revealed on the other nicotinic receptors. 4. The effect of nicotine was tested on hybrid receptors obtained by coinjection of chick and rat cDNAs encoding the alpha 3 and beta 2 subunits (yielding the rat alpha 3/chick beta 2 and chick alpha 3/rat beta 2 receptors). Nicotine (10 microM) strongly inhibited both hybrid receptors. 5. Chimeric subunits were constructed by exchanging a segment located in the extracellular N-termini of chick alpha 3 and alpha 4 subunits and chick alpha 3 and rat alpha 3 subunits. These subunits were coexpressed in oocytes with chick or rat beta 2 subunits. The effect of nicotine on these receptors pointed to the importance of a 15 amino acid stretch located 3' of the first transmembrane segment in the determination of the agonist and antagonist action of nicotine. 6. Within this 15 amino acid segment, a single residue differs in chick and rat alpha 3 subunits, at position 198, within the ligand binding site of alpha subunits. Gln198 of the rat alpha 3 subunit was replaced

  19. Nicotinic Receptors: Role in Addiction and Other Disorders of the Brain

    PubMed Central

    Sharma, Geeta; Vijayaraghavan, Sukumar

    2008-01-01

    Nicotine, the addictive component of cigarette smoke has profound effects on the brain. Activation of its receptors by nicotine has complex consequences for network activity throughout the brain, potentially contributing to the addictive property of the drug. Nicotinic receptors have been implicated in psychiatric illnesses like schizophrenia and are also neuroprotective, potentially beneficial for neurodegenerative diseases. These effects of nicotine serve to emphasize the multifarious roles the drug, acting through multiple nicotinic acetylcholine receptor subtypes. The findings also remind us of the complexity of signaling mechanisms and stress the risks of unintended consequences of drugs designed to combat nicotine addiction. PMID:20148179

  20. Nicotine improves the functional activity of late endothelial progenitor cells via nicotinic acetylcholine receptors.

    PubMed

    Yu, Min; Liu, Qian; Sun, Jing; Yi, Kaihong; Wu, Libiao; Tan, Xuerui

    2011-08-01

    The aim of this study is to investigate whether nicotinic acetylcholine receptors (nAChRs) are involved in the modulation of functional activity of late endothelial progenitor cells (EPCs) induced by nicotine. Total mononuclear cells (MNCs) were isolated from human umbilical cord blood by Ficoll density gradient centrifugation, and then the cells were plated on fibronectin-coated culture plates. Late EPCs were positive for 1,1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine-labeled acetylated low-density lipoprotein (DiI-acLDL) uptake and fluorescein-isothiocyanate-conjugated Ulex europaeus agglutinin lectin (UEA-1) binding. Expression of von Willbrand factor (vWF), kinase insert domain receptor (KDR), and α7 nAChR was detected by indirect immunofluorescence staining. Late EPCs of 3-5 passages were treated for 32 h with either vehicle or nicotine with or without pre-incubation of nAChR antagonism, mecamylamine, or α-bungarotoxin. The viability, migration, and in vitro vasculogenesis activity of late EPCs were assayed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, modified Boyden chamber assay, and in vitro angiogenesis assay, respectively. Late EPCs adhesion assay was performed by replating cells on fibronectin-coated plates, and then adherent cells were counted. Incubation with 10 nmol/L nicotine enhanced viable, migratory, adhesive, and in vitro vasculogenesis capacity of late EPCs. The effect of nicotine on late EPCs can be attenuated by mecamylamine or α-bungarotoxin. In conclusion, nicotine improves the functional activity of late EPCs via nAChRs. PMID:21774635

  1. Cannabinoid Receptor 1 Gene Association With Nicotine Dependence

    PubMed Central

    Chen, Xiangning; Williamson, Vernell S.; An, Seon-Sook; Hettema, John M.; Aggen, Steven H.; Neale, Michael C.; Kendler, Kenneth S.

    2009-01-01

    Context The endogenous cannabinoid system has been implicated in drug addiction in animal models. The cannabinoid receptor 1 (CNR1) gene is 1 of the 2 receptors expressed in the brain. It has been reported to be associated with alcoholism and multiple drug abuse and dependence. Objective To test the hypothesis that the CNR1 gene is associated with nicotine dependence. Design Genotype-phenotype association study. Ten single-nucleotide polymorphisms were genotyped in the CNR1 gene in 2 independent samples. For the first sample (n=688), a 3-group case-control design was used to test allele association with smoking initiation and nicotine dependence. For the second sample (n = 961), association was assessed with scores from the Fagerström Test for Nicotine Dependence (FTND). Settings Population samples selected from the Mid-Atlantic Twin Registry. Participants White patients aged 18 to 65 years who met the criteria of inclusion. Main Outcome Measures Fagerström Tolerance Questionnaire and FTND scores. Results Significant single-marker and haplotype associations were found in both samples, and the associations were female specific. Haplotype 1-1-2 of markers rs2023239-rs12720071-rs806368 was associated with nicotine dependence and FTND score in the 2 samples (P<.001 and P = .009, respectively). Conclusion Variants and haplotypes in the CNR1 gene may alter the risk for nicotine dependence, and the associations are likely sex specific. PMID:18606954

  2. RIC-3 differentially modulates α4β2 and α7 nicotinic receptor assembly, expression, and nicotine-induced receptor upregulation

    PubMed Central

    2013-01-01

    Background Recent work has shown that the chaperone resistant to inhibitors of acetylcholinesterase (RIC-3) is critical for the folding, maturation and functional expression of a variety of neuronal nicotinic acetylcholine receptors. α7 nicotinic receptors can only assemble and functionally express in select lines of cells, provided that RIC-3 is present. In contrast, α4β2 nicotinic receptors can functionally express in many cell lines even without the presence of RIC-3. Depending on the cell line, RIC-3 has differential effects on α4β2 receptor function – enhancement in mammalian cells but inhibition in Xenopus oocytes. Other differences between the two receptor types include nicotine-induced upregulation. When expressed in cell lines, α4β2 receptors readily and robustly upregulate with chronic nicotine exposure. However, α7 nicotinic receptors appear more resistant and require higher concentrations of nicotine to induce upregulation. Could the coexpression of RIC-3 modulate the extent of nicotine-induced upregulation not only for α7 receptors but also α4β2 receptors? We compared and contrasted the effects of RIC-3 on assembly, trafficking, protein expression and nicotine-induced upregulation on both α7 and α4β2 receptors using fluorescent protein tagged nicotinic receptors and Förster resonance energy transfer (FRET) microscopy imaging. Results RIC-3 increases assembly and cell surface trafficking of α7 receptors but does not alter α7 protein expression in transfected HEK293T cells. In contrast, RIC-3 does not affect assembly of α4β2 receptors but increases α4 and β2 subunit protein expression. Acute nicotine (30 min exposure) was sufficient to upregulate FRET between α4 and β2 subunits. Surprisingly, when RIC-3 was coexpressed with α4β2 receptors nicotine-induced upregulation was prevented. α7 receptors did not upregulate with acute nicotine in the presence or absence of RIC-3. Conclusions These results provide interesting novel data

  3. Nicotinic receptor blockade decreases fos immunoreactivity within orexin/hypocretin-expressing neurons of nicotine-exposed rats.

    PubMed

    Simmons, Steven J; Gentile, Taylor A; Mo, Lili; Tran, Fionya H; Ma, Sisi; Muschamp, John W

    2016-11-01

    Tobacco smoking is the leading cause of preventable death in the United States. Nicotine is the principal psychoactive ingredient in tobacco that causes addiction. The structures governing nicotine addiction, including those underlying withdrawal, are still being explored. Nicotine withdrawal is characterized by negative affective and cognitive symptoms that enhance relapse susceptibility, and suppressed dopaminergic transmission from ventral tegmental area (VTA) to target structures underlies behavioral symptoms of nicotine withdrawal. Agonist and partial agonist therapies help 1 in 4 treatment-seeking smokers at one-year post-cessation, and new targets are needed to more effectively aid smokers attempting to quit. Hypothalamic orexin/hypocretin neurons send excitatory projections to dopamine (DA)-producing neurons of VTA and modulate mesoaccumbal DA release. The effects of nicotinic receptor blockade, which is commonly used to precipitate withdrawal, on orexin neurons remain poorly investigated and present an attractive target for intervention. The present study sought to investigate the effects of nicotinic receptor blockade on hypothalamic orexin neurons using mecamylamine to precipitate withdrawal in rats. Separate groups of rats were treated with either chronic nicotine or saline for 7-days at which point effects of mecamylamine or saline on somatic signs and anxiety-like behavior were assessed. Finally, tissue from rats was harvested for immunofluorescent analysis of Fos within orexin neurons. Results demonstrate that nicotinic receptor blockade leads to reduced orexin cell activity, as indicated by lowered Fos-immunoreactivity, and suggest that this underlying cellular activity may be associated with symptoms of nicotine withdrawal as effects were most prominently observed in rats given chronic nicotine. We conclude from this study that orexin transmission becomes suppressed in rats upon nicotinic receptor blockade, and that behavioral symptoms associated

  4. Topographical studies of the nicotinic acetylcholine receptor. [Torpedo californica

    SciTech Connect

    Middlemas, D.S.

    1987-01-01

    All four subunits of the nicotinic acetylcholine receptor in membrane vesicles isolated from Torpedo californica have been labeled with the photoactivated hydrophobic probe, (/sup 3/H)adamantanediazirine, which selectively labels regions of integral membrane proteins in contact with the hydrocarbon core of the lipid bilayer. All four subunits of the acetylcholine receptor in membrane vesicles isolated from Torpedo californica have been labeled with (/sup 3/H)cholesteryl diazoacetate. As this probe incorporates into lipid bilayers analogously to cholesterol, this result indicates that acetylcholine receptor interacts with cholesterol. Since the photogenerated carbene is situated near the lipid-water interface, this probe has potential as a topographic tool for mapping membrane protein structure. The labeling studies with both (/sup 3/H)adamantanediazirine and (/sup 3/H)cholesteryl diazoacetate support the concept that the acetylcholine receptor is a pseudosymmetric complex of homologous subunits, all of which interact with and span the membrane. The synthesis of the fluorine-containing agonists for the Torpedo californica nicotinic acetylcholine receptor, fluoroacetylcholine bromide and p-fluorophenyltrimethylammonium iodide, are described. It is demonstrated that both are agonists using a cation flux assay with acetylcholine receptor enriched membrane vesicles. The affinity cleavage reagent, p-thiocyanophenyltrimethylammonium iodide, specifically cleaves a peptide bond of the nicotinic acetylcholine receptor in membrane vesicles isolated from Torpedo californica. It is demonstrated that this reagent is an agonist using a cation flux assay. The cleavage is blocked by stoichiometric quantities of ..cap alpha..-bungarotoxin.

  5. Presynaptic P2X1-3 and α3-containing nicotinic receptors assemble into functionally interacting ion channels in the rat hippocampus.

    PubMed

    Rodrigues, Ricardo J; Almeida, Teresa; Díaz-Hernández, Miguel; Marques, Joana M; Franco, Rafael; Solsona, Carles; Miras-Portugal, María Teresa; Ciruela, Francisco; Cunha, Rodrigo A

    2016-06-01

    Previous studies documented a cross-talk between purinergic P2X (P2XR) and nicotinic acetylcholine receptors (nAChR) in heterologous expression systems and peripheral preparations. We now investigated if this occurred in native brain preparations and probed its physiological function. We found that P2XR and nAChR were enriched in hippocampal terminals, where both P2X1-3R and α3, but not α4, nAChR subunits were located in the active zone and in dopamine-β-hydroxylase-positive hippocampal terminals. Notably, P2XR ligands displaced nAChR binding and nAChR ligands displaced P2XR binding to hippocampal synaptosomes. In addition, a negative P2XR/nAChR cross-talk was observed in the control of the evoked release of noradrenaline from rat hippocampal synaptosomes, characterized by a less-than-additive facilitatory effect upon co-activation of both receptors. This activity-dependent cross-inhibition was confirmed in Xenopus oocytes transfected with P2X1-3Rs and α3β2 (but not α4β2) nAChR. Besides, P2X2 co-immunoprecipitated α3β2 (but not α4β2) nAChR, both in HEK cells and rat hippocampal membranes indicating that this functional interaction is supported by a physical association between P2XR and nAChR. Moreover, eliminating extracellular ATP with apyrase in hippocampal slices promoted the inhibitory effect of the nAChR antagonist tubocurarine on noradrenaline release induced by high- but not low-frequency stimulation. Overall, these results provide integrated biochemical, pharmacological and functional evidence showing that P2X1-3R and α3β2 nAChR are physically and functionally interconnected at the presynaptic level to control excessive noradrenergic terminal activation upon intense synaptic firing in the hippocampus. PMID:26801076

  6. Nicotinic receptor Alpha7 expression during mouse adrenal gland development.

    PubMed

    Gahring, Lorise C; Myers, Elizabeth; Palumbos, Sierra; Rogers, Scott W

    2014-01-01

    The nicotinic acetylcholine receptor alpha 7 (α7) is a ligand-activated ion channel that contributes to a diversity of cellular processes involved in development, neurotransmission and inflammation. In this report the expression of α7 was examined in the mouse developing and adult adrenal gland that expresses a green fluorescent protein (GFP) reporter as a bi-cistronic extension of the endogenous α7 transcript (α7(G)). At embryonic day 12.5 (E12.5) α7(G) expression was associated with the suprarenal ganglion and precursor cells of the adrenal gland. The α7(G) cells are catecholaminergic chromaffin cells as reflected by their progressive increase in the co-expression of tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH) that is complete by E18.5. In the adult, α7(G) expression is limited to a subset of chromaffin cells in the adrenal medulla that cluster near the border with the adrenal cortex. These chromaffin cells co-express α7(G), TH and DBH, but they lack phenylethanolamine N-methyltransferase (PNMT) consistent with only norepinephrine (NE) synthesis. These cell groups appear to be preferentially innervated by pre-ganglionic afferents identified by the neurotrophin receptor p75. No afferents identified by beta-III tubulin, neurofilament proteins or p75 co-expressed α7(G). Occasional α7(G) cells in the pre-E14.5 embryos express neuronal markers consistent with intrinsic ganglion cells and in the adult some α7(G) cells co-express glutamic acid decarboxylase. The transient expression of α7 during adrenal gland development and its prominent co-expression by a subset of NE chromaffin cells in the adult suggests that the α7 receptor contributes to multiple aspects of adrenal gland development and function that persist into adulthood. PMID:25093893

  7. Role of β4* Nicotinic Acetylcholine Receptors in the Habenulo-Interpeduncular Pathway in Nicotine Reinforcement in Mice.

    PubMed

    Harrington, Lauriane; Viñals, Xavier; Herrera-Solís, Andrea; Flores, Africa; Morel, Carole; Tolu, Stefania; Faure, Philippe; Maldonado, Rafael; Maskos, Uwe; Robledo, Patricia

    2016-06-01

    Nicotine exerts its psychopharmacological effects by activating the nicotinic acetylcholine receptor (nAChR), composed of alpha and/or beta subunits, giving rise to a diverse population of receptors with a distinct pharmacology. β4-containing (β4*) nAChRs are located almost exclusively in the habenulo-interpeduncular pathway. We examined the role of β4* nAChRs in the medial habenula (MHb) and the interpeduncular nucleus (IPN) in nicotine reinforcement using behavioral, electrophysiological, and molecular techniques in transgenic mice. Nicotine intravenous self-administration (IVSA) was lower in constitutive β4 knockout (KO) mice at all doses tested (7.5, 15, 30, and 60 μg/kg/infusion) compared with wild-type (WT) mice. In vivo microdialysis showed that β4KO mice have higher extracellular dopamine (DA) levels in the nucleus accumbens than in WT mice, and exhibit a differential sensitivity to nicotine-induced DA outflow. Furthermore, electrophysiological recordings in the ventral tegmental area (VTA) demonstrated that DA neurons of β4KO mice are more sensitive to lower doses of nicotine than that of WT mice. Re-expression of β4* nAChRs in IPN neurons fully restored nicotine IVSA, and attenuated the increased sensitivity of VTA DA neurons to nicotine. These findings suggest that β4* nAChRs in the IPN have a role in maintaining nicotine IVSA. PMID:26585290

  8. A Multi-Route Model of Nicotine-Cotinine Pharmacokinetics, Pharmacodynamics and Brain Nicotinic Acetylcholine Receptor Binding in Humans

    SciTech Connect

    Teeguarden, Justin G.; Housand, Conrad; Smith, Jordan N.; Hinderliter, Paul M.; Gunawan, Rudy; Timchalk, Charles

    2013-02-01

    The pharmacokinetics of nicotine, the pharmacologically active alkaloid in tobacco responsible for addiction, are well characterized in humans. We developed a physiologically based pharmacokinetic/pharmacodynamic model of nicotine pharmacokinetics, brain dosimetry and brain nicotinic acetylcholine receptor (nAChRs) occupancy. A Bayesian framework was applied to optimize model parameters against multiple human data sets. The resulting model was consistent with both calibration and test data sets, but in general underestimated variability. A pharmacodynamic model relating nicotine levels to increases in heart rate as a proxy for the pharmacological effects of nicotine accurately described the nicotine related changes in heart rate and the development and decay of tolerance to nicotine. The PBPK model was utilized to quantitatively capture the combined impact of variation in physiological and metabolic parameters, nicotine availability and smoking compensation on the change in number of cigarettes smoked and toxicant exposure in a population of 10,000 people presented with a reduced toxicant (50%), reduced nicotine (50%) cigarette Across the population, toxicant exposure is reduced in some but not all smokers. Reductions are not in proportion to reductions in toxicant yields, largely due to partial compensation in response to reduced nicotine yields. This framework can be used as a key element of a dosimetry-driven risk assessment strategy for cigarette smoke constituents.

  9. Caenorhabditis elegans nicotinic acetylcholine receptors are required for nociception

    PubMed Central

    Cohen, Emiliano; Chatzigeorgiou, Marios; Husson, Steven J.; Steuer-Costa, Wagner; Gottschalk, Alexander; Schafer, William R.; Treinin, Millet

    2014-01-01

    Polymodal nociceptors sense and integrate information on injurious mechanical, thermal, and chemical stimuli. Chemical signals either activate nociceptors or modulate their responses to other stimuli. One chemical known to activate or modulate responses of nociceptors is acetylcholine (ACh). Across evolution nociceptors express subunits of the nicotinic acetylcholine receptor (nAChR) family, a family of ACh-gated ion channels. The roles of ACh and nAChRs in nociceptor function are, however, poorly understood. Caenorhabditis elegans polymodal nociceptors, PVD, express nAChR subunits on their sensory arbor. Here we show that mutations reducing ACh synthesis and mutations in nAChR subunits lead to defects in PVD function and morphology. A likely cause for these defects is a reduction in cytosolic calcium measured in ACh and nAChR mutants. Indeed, overexpression of a calcium pump in PVD mimics defects in PVD function and morphology found in nAChR mutants. Our results demonstrate, for the first time, a central role for nAChRs and ACh in nociceptor function and suggest that calcium permeating via nAChRs facilitates activity of several signaling pathways within this neuron. PMID:24518198

  10. Theoretical investigation of interaction between the set of ligands and α7 nicotinic acetylcholine receptor

    NASA Astrophysics Data System (ADS)

    Glukhova, O. E.; Prytkova, T. R.; Shmygin, D. S.

    2016-03-01

    Nicotinic acetylcholine receptors (nAChRs) are neuron receptor proteins that provide a transmission of nerve impulse through the synapses. They are composed of a pentametric assembly of five homologous subunits (5 α7 subunits for α7nAChR, for example), oriented around the central pore. These receptors might be found in the chemical synapses of central and peripheral nervous system, and also in the neuromuscular synapses. Transmembrane domain of the one of such receptors constitutes ion channel. The conductive properties of ion channel strongly depend on the receptor conformation changes in the response of binding with some molecule, f.e. acetylcholine. Investigation of interaction between ligands and acetylcholine receptor is important for drug design. In this work we investigate theoretically the interaction between the set of different ligands (such as vanillin, thymoquinone, etc.) and the nicotinic acetylcholine receptor (primarily with subunit of the α7nAChR) by different methods and packages (AutodockVina, GROMACS, KVAZAR, HARLEM, VMD). We calculate interaction energy between different ligands in the subunit using molecular dynamics. On the base of obtained calculation results and using molecular docking we found an optimal location of different ligands in the subunit.

  11. Does chronic nicotine alter neurotransmitter receptors involved in Parkinson's disease

    SciTech Connect

    Reilly, M.A.; Lapin, E.P.; Lajtha, A.; Maker, H.S.

    1986-03-05

    Cigarette smokers are fewer in number among Parkinson's Disease (PD) patients than among groups of persons who do not have PD. Several hypotheses have been proposed to explain this observation. One which must be tested is the possibility that some pharmacologic agent present in cigarette smoke may interact with some central nervous system component involved in PD. To this end, they have investigated the effect of chronic nicotine administration on receptors for some of the neurotransmitters that are affected in PD. Rats were injected for six weeks with saline or nicotine 0.8 mg/kg S.C., then killed and brains removed and dissected. The binding of (/sup 3/H)-ketanserin to serotonin receptors in frontal cortex and of (/sup 3/H)-domperidone to dopamine receptors in caudate was not affected. However, the binding of (/sup 3/H)-domperidone in nucleus accumbens was altered: the K/sub d/ increased from 0.16 +/- 0.02 nM to 0.61 +/- 0.07 nM, and the B/sub max/ increased from 507 +/- 47 fmol/mg protein to 910 +/- 43 fmol/mg (p < 0.001 for both comparisons). These values are based on three ligand concentrations. Additional studies are in progress to substantiate the data. It is concluded that chronic nicotine administration may alter dopamine receptors in nucleus accumbens.

  12. Nicotine receptors mediating sensorimotor gating and its enhancement by systemic nicotine

    PubMed Central

    Pinnock, Farena; Bosch, Daniel; Brown, Tyler; Simons, Nadine; Yeomans, John R.; DeOliveira, Cleusa; Schmid, Susanne

    2015-01-01

    Prepulse inhibition (PPI) of startle occurs when intensity stimuli precede stronger startle-inducing stimuli by 10–1000 ms. PPI deficits are found in individuals with schizophrenia and other psychiatric disorders, and they correlate with other cognitive impairments. Animal research and clinical studies have demonstrated that both PPI and cognitive function can be enhanced by nicotine. PPI has been shown to be mediated, at least in part, by mesopontine cholinergic neurons that project to pontine startle neurons and activate muscarinic and potentially nicotine receptors (nAChRs). The subtypes and anatomical location of nAChRs involved in mediating and modulating PPI remain unresolved. We tested the hypothesis that nAChRs that are expressed by pontine startle neurons contribute to PPI. We also explored whether or not these pontine receptors are responsible for the nicotine enhancement of PPI. While systemic administration of nAChR antagonists had limited effects on PPI, PnC microinfusions of the non-α7nAChR preferring antagonist TMPH, but not of the α7nAChR antagonist MLA, into the PnC significantly reduced PPI. Electrophysiological recordings from startle-mediating PnC neurons confirmed that nicotine affects excitability of PnC neurons, which could be antagonized by TMPH, but not by MLA, indicating the expression of non-α7nAChR. In contrast, systemic nicotine enhancement of PPI was only reversed by systemic MLA and not by TMPH or local microinfusions of MLA into the PnC. In summary, our data indicate that non-α7nAChRs in the PnC contribute to PPI at stimulus intervals of 100 ms or less, whereas activation of α7nAChRs in other brain areas is responsible for the systemic nicotine enhancement of PPI. This is important knowledge for the correct interpretation of behavioral, preclinical, and clinical data as well as for developing drugs for the amelioration of PPI deficits and the enhancement of cognitive function. PMID:25717295

  13. Electrophysiology-Based Assays to Detect Subtype-Selective Modulation of Human Nicotinic Acetylcholine Receptors

    PubMed Central

    Kirsch, Glenn E.; Fedorov, Nikolai B.; Kuryshev, Yuri A.; Liu, Zhiqi; Orr, Michael S.

    2016-01-01

    Abstract The Family Smoking Prevention and Tobacco Control Act of 2009 (Public Law 111-31) gave the US Food and Drug Administration (FDA) the responsibility for regulating tobacco products. Nicotine is the primary addictive component of tobacco and its effects can be modulated by additional ingredients in manufactured products. Nicotine acts by mimicking the neurotransmitter acetylcholine on neuronal nicotinic acetylcholine receptors (nAChRs), which function as ion channels in cholinergic modulation of neurotransmission. Subtypes within the family of neuronal nAChRs are defined by their α- and β-subunit composition. The subtype-selective profiles of tobacco constituents are largely unknown, but could be essential for understanding the physiological effects of tobacco products. In this report, we report the development and validation of electrophysiology-based high-throughput screens (e-HTS) for human nicotinic subtypes, α3β4, α3β4α5, α4β2, and α7 stably expressed in Chinese Hamster Ovary cells. Assessment of agonist sensitivity and acute desensitization gave results comparable to those obtained by conventional manual patch clamp electrophysiology assays. The potency of reference antagonists for inhibition of the receptor channels and selectivity of positive allosteric modulators also were very similar between e-HTS and conventional manual patch voltage clamp data. Further validation was obtained in pilot screening of a library of FDA-approved drugs that identified α7 subtype-selective positive allosteric modulation by novel compounds. These assays provide new tools for profiling of nicotinic receptor selectivity. PMID:27505073

  14. Electrophysiology-Based Assays to Detect Subtype-Selective Modulation of Human Nicotinic Acetylcholine Receptors.

    PubMed

    Kirsch, Glenn E; Fedorov, Nikolai B; Kuryshev, Yuri A; Liu, Zhiqi; Armstrong, Lucas C; Orr, Michael S

    2016-08-01

    The Family Smoking Prevention and Tobacco Control Act of 2009 (Public Law 111-31) gave the US Food and Drug Administration (FDA) the responsibility for regulating tobacco products. Nicotine is the primary addictive component of tobacco and its effects can be modulated by additional ingredients in manufactured products. Nicotine acts by mimicking the neurotransmitter acetylcholine on neuronal nicotinic acetylcholine receptors (nAChRs), which function as ion channels in cholinergic modulation of neurotransmission. Subtypes within the family of neuronal nAChRs are defined by their α- and β-subunit composition. The subtype-selective profiles of tobacco constituents are largely unknown, but could be essential for understanding the physiological effects of tobacco products. In this report, we report the development and validation of electrophysiology-based high-throughput screens (e-HTS) for human nicotinic subtypes, α3β4, α3β4α5, α4β2, and α7 stably expressed in Chinese Hamster Ovary cells. Assessment of agonist sensitivity and acute desensitization gave results comparable to those obtained by conventional manual patch clamp electrophysiology assays. The potency of reference antagonists for inhibition of the receptor channels and selectivity of positive allosteric modulators also were very similar between e-HTS and conventional manual patch voltage clamp data. Further validation was obtained in pilot screening of a library of FDA-approved drugs that identified α7 subtype-selective positive allosteric modulation by novel compounds. These assays provide new tools for profiling of nicotinic receptor selectivity. PMID:27505073

  15. L-theanine inhibits nicotine-induced dependence via regulation of the nicotine acetylcholine receptor-dopamine reward pathway.

    PubMed

    Di, Xiaojing; Yan, Jingqi; Zhao, Yan; Chang, Yanzhong; Zhao, Baolu

    2012-12-01

    In this study, the inhibitory effect of L-theanine, an amino acid derivative of tea, on the rewarding effects of nicotine and its underlying mechanisms of action were studied. We found that L-theanine inhibited the rewarding effects of nicotine in a conditioned place preference (CPP) model of the mouse and reduced the excitatory status induced by nicotine in SH-SY5Y cells to the same extent as the nicotine receptor inhibitor dihydro-beta-erythroidine (DHβE). Further studies using high performance liquid chromatography, western blotting and immunofluorescence staining analyses showed that L-theanine significantly inhibited nicotine-induced tyrosine hydroxylase (TH) expression and dopamine production in the midbrain of mice. L-theanine treatment also reduced the upregulation of the α(4), β(2) and α(7) nicotine acetylcholine receptor (nAChR) subunits induced by nicotine in mouse brain regions that related to the dopamine reward pathway, thus decreasing the number of cells that could react to nicotine. In addition, L-theanine treatment inhibited nicotine-induced c-Fos expression in the reward circuit related areas of the mouse brain. Knockdown of c-Fos by siRNA inhibited the excitatory status of cells but not the upregulation of TH induced by nicotine in SH-SY5Y cells. Overall, the present study showed that L-theanine reduced the nicotine-induced reward effects via inhibition of the nAChR-dopamine reward pathway. These results may offer new therapeutic strategies for treatment of tobacco addiction. PMID:23233221

  16. Diversity of native nicotinic receptor subtypes in mammalian brain.

    PubMed

    Zoli, Michele; Pistillo, Francesco; Gotti, Cecilia

    2015-09-01

    Neuronal nicotinic acetylcholine receptors (nAChRs) are a heterogeneous family of pentameric ligand-gated cation channels that are expressed throughout the brain and involved in a wide range of physiological and pathophysiological processes. The nAChR subtypes share a common basic structure, but their biophysical and pharmacological properties depend on their subunit composition, which is therefore central to understanding their function in the nervous system and discovering new subtype selective drugs. The development of new technologies and the generation of mice carrying deletions or the expression of gain-of-function nAChR subunits, or GFP-tagged receptor genes has allowed the in vivo identification of complex subtypes and to study the role of individual subtypes in specific cells and complex neurobiological systems but much less is known about which native nAChR subtypes are involved in specific physiological functions and pathophysiological conditions in human brain. We briefly review some recent findings concerning the structure and function of native nAChRs, focussing on the subtypes identified in the rodent habenulo-interpeduncular pathway, a pathway involved in nicotine reinforcement and withdrawal. We also discuss recent findings concerning the expression of native subtypes in primate brain. This article is part of the Special Issue entitled 'The Nicotinic Acetylcholine Receptor: From Molecular Biology to Cognition'. PMID:25460185

  17. Tracking the Molecular Evolution of Calcium Permeability in a Nicotinic Acetylcholine Receptor

    PubMed Central

    Lipovsek, Marcela; Fierro, Angélica; Pérez, Edwin G.; Boffi, Juan C.; Millar, Neil S.; Fuchs, Paul A.; Katz, Eleonora; Elgoyhen, Ana Belén

    2014-01-01

    Nicotinic acetylcholine receptors are a family of ligand-gated nonselective cationic channels that participate in fundamental physiological processes at both the central and the peripheral nervous system. The extent of calcium entry through ligand-gated ion channels defines their distinct functions. The α9α10 nicotinic cholinergic receptor, expressed in cochlear hair cells, is a peculiar member of the family as it shows differences in the extent of calcium permeability across species. In particular, mammalian α9α10 receptors are among the ligand-gated ion channels which exhibit the highest calcium selectivity. This acquired differential property provides the unique opportunity of studying how protein function was shaped along evolutionary history, by tracking its evolutionary record and experimentally defining the amino acid changes involved. We have applied a molecular evolution approach of ancestral sequence reconstruction, together with molecular dynamics simulations and an evolutionary-based mutagenesis strategy, in order to trace the molecular events that yielded a high calcium permeable nicotinic α9α10 mammalian receptor. Only three specific amino acid substitutions in the α9 subunit were directly involved. These are located at the extracellular vestibule and at the exit of the channel pore and not at the transmembrane region 2 of the protein as previously thought. Moreover, we show that these three critical substitutions only increase calcium permeability in the context of the mammalian but not the avian receptor, stressing the relevance of overall protein structure on defining functional properties. These results highlight the importance of tracking evolutionarily acquired changes in protein sequence underlying fundamental functional properties of ligand-gated ion channels. PMID:25193338

  18. Tracking the molecular evolution of calcium permeability in a nicotinic acetylcholine receptor.

    PubMed

    Lipovsek, Marcela; Fierro, Angélica; Pérez, Edwin G; Boffi, Juan C; Millar, Neil S; Fuchs, Paul A; Katz, Eleonora; Elgoyhen, Ana Belén

    2014-12-01

    Nicotinic acetylcholine receptors are a family of ligand-gated nonselective cationic channels that participate in fundamental physiological processes at both the central and the peripheral nervous system. The extent of calcium entry through ligand-gated ion channels defines their distinct functions. The α9α10 nicotinic cholinergic receptor, expressed in cochlear hair cells, is a peculiar member of the family as it shows differences in the extent of calcium permeability across species. In particular, mammalian α9α10 receptors are among the ligand-gated ion channels which exhibit the highest calcium selectivity. This acquired differential property provides the unique opportunity of studying how protein function was shaped along evolutionary history, by tracking its evolutionary record and experimentally defining the amino acid changes involved. We have applied a molecular evolution approach of ancestral sequence reconstruction, together with molecular dynamics simulations and an evolutionary-based mutagenesis strategy, in order to trace the molecular events that yielded a high calcium permeable nicotinic α9α10 mammalian receptor. Only three specific amino acid substitutions in the α9 subunit were directly involved. These are located at the extracellular vestibule and at the exit of the channel pore and not at the transmembrane region 2 of the protein as previously thought. Moreover, we show that these three critical substitutions only increase calcium permeability in the context of the mammalian but not the avian receptor, stressing the relevance of overall protein structure on defining functional properties. These results highlight the importance of tracking evolutionarily acquired changes in protein sequence underlying fundamental functional properties of ligand-gated ion channels. PMID:25193338

  19. ICV STZ induced impairment in memory and neuronal mitochondrial function: A protective role of nicotinic receptor.

    PubMed

    Saxena, Gunjan; Patro, Ishan K; Nath, Chandishwar

    2011-10-10

    The present study was planned to evaluate the cholinergic influence on mitochondrial activity and neurodegeneration associated with impaired memory in intracerebroventricular (ICV) streptozotocin (STZ) treated rats. STZ (3mg/kg), administered ICV twice with an interval of 48h between the two doses, showed significant impairment in spatial memory tested by water maze test 14 days after first dose without altering blood glucose level and locomotor activity. Animals were sacrificed on 21st day of ICV administration. STZ significantly increased malondialdehyde (MDA), reactive oxygen species (ROS), Ca(2+) ion influx, caspase-3 activity and decreased glutathione (GSH) level. Acetylcholinesterase inhibitors tacrine and donepezil (5mg/kg, PO) pretreatment significantly prevented STZ induced memory deficit, oxidative stress, Ca(2+) influx and caspase-3 activity. Carbachol, a muscarinic cholinergic agonist (0.01mg/kg, SC) did not show any significant effect on ROS generation, Ca(2+) ion influx and caspase-3 activity. While nicotinic cholinergic agonist, nicotine, significantly attenuated ICV STZ induced mitochondrial dysfunction and caspase-3 activity. The results indicate that instead of muscarinic receptors nicotinic receptors may be involved in neuroprotection by maintaining mitochondrial functions. PMID:21620901

  20. Vulnerability to nicotine self-administration in adolescent mice correlates with age-specific expression of α4* nicotinic receptors.

    PubMed

    Renda, Anthony; Penty, Nora; Komal, Pragya; Nashmi, Raad

    2016-09-01

    The majority of smokers begin during adolescence, a developmental period with a high susceptibility to substance abuse. Adolescents are affected differently by nicotine compared to adults, with adolescents being more vulnerable to nicotine's rewarding properties. It is unknown if the age-dependent molecular composition of a younger brain contributes to a heightened susceptibility to nicotine addiction. Nicotine, the principle pharmacological component of tobacco, binds and activates nicotinic acetylcholine receptors (nAChRs) in the brain. The most prevalent is the widely expressed α4-containing (α4*) subtype which mediates reward and is strongly implicated in nicotine dependence. Exposing different age groups of mice, postnatal day (P) 44-86 days old, to a two bottle-choice oral nicotine self-administration paradigm for five days yielded age-specific consumption levels. Nicotine self-administration was elevated in the P44 group, peaked at P54-60 and was drastically lower in the P66 through P86 groups. We also quantified α4* nAChR expression via spectral confocal imaging of brain slices from α4YFP knock-in mice, in which the α4 nAChR subunit is tagged with a yellow fluorescent protein. Quantitative fluorescence revealed age-specific α4* nAChR expression in dopaminergic and GABAergic neurons of the ventral tegmental area. Receptor expression showed a strong positive correlation with daily nicotine dose, suggesting that α4* nAChR expression levels are age-specific and may contribute to the propensity to self-administer nicotine. PMID:27102349

  1. Functional expression of human α7 nicotinic acetylcholine receptor in human embryonic kidney 293 cells.

    PubMed

    Gong, Yuan; Jiang, Ji-Hong; Li, Shi-Tong

    2016-09-01

    The functional expression of recombinant α7 nicotinic acetylcholine receptors in human embryonic kidney (HEK) 293 cells has presented a challenge. Resistance to inhibitors of cholinesterase 3 (RIC‑3) has been confirmed to act as a molecular chaperone of nicotinic acetylcholine receptors. The primary objectives of the present study were to investigate whether the co‑expression of human (h)RIC‑3 with human α7 nicotinic acetylcholine receptor in HEK 293 cells facilitates functional expression of the α7 nicotinic acetylcholine receptor. Subsequent to transfection, western blotting and polymerase chain reaction were used to test the expression of α7 nicotinic acetylcholine receptor and RIC-3. The α7 nicotinic acetylcholine receptor was expressed alone or co‑expressed with hRIC‑3 in the HEK 293 cells. Drug‑containing solution was then applied to the cells via a gravity‑driven perfusion system. Calcium influx in the cells was analyzed using calcium imaging. Nicotine did not induce calcium influx in the HEK 293 cells expressing human α7 nicotinic acetylcholine receptor only. However, in the cells co‑expressing human RIC‑3 and α7 nicotinic acetylcholine receptor, nicotine induced calcium influx via the α7 nicotinic acetylcholine receptor in a concentration‑dependent manner (concentration required to elicit 50% of the maximal effect=29.21 µM). Taken together, the results of the present study suggested that the co‑expression of RIC‑3 in HEK 293 cells facilitated the functional expression of the α7 nicotinic acetylcholine receptor. PMID:27430244

  2. An autoradiographic analysis of cholinergic receptors in mouse brain after chronic nicotine treatment

    SciTech Connect

    Pauly, J.R.; Marks, M.J.; Gross, S.D.; Collins, A.C. )

    1991-09-01

    Quantitative autoradiographic procedures were used to examine the effects of chronic nicotine infusion on the number of central nervous system nicotinic cholinergic receptors. Female DBA mice were implanted with jugular cannulas and infused with saline or various doses of nicotine (0.25, 0.5, 1.0 or 2.0 mg/kg/hr) for 10 days. The animals were then sacrificed and the brains were removed and frozen in isopentane. Cryostat sections were collected and prepared for autoradiographic procedures as previously described. Nicotinic cholinergic receptors were labeled with L-(3H)nicotine or alpha-(125I)bungarotoxin; (3H)quinuclidinyl benzilate was used to measure muscarinic cholinergic receptor binding. Chronic nicotine infusion increased the number of sites labeled by (3H)nicotine in most brain areas. However, the extent of the increase in binding as well as the dose-response curves for the increase were widely different among brain regions. After the highest treatment dose, binding was increased in 67 of 86 regions measured. Septal and thalamic regions were most resistant to change. Nicotinic binding measured by alpha-(125I)bungarotoxin also increased after chronic treatment, but in a less robust fashion. At the highest treatment dose, only 26 of 80 regions were significantly changes. Muscarinic binding was not altered after chronic nicotine treatment. These data suggest that brain regions are not equivalent in the mechanisms that regulate alterations in nicotinic cholinergic receptor binding after chronic nicotine treatment.

  3. α3β4 nicotinic acetylcholine receptors in the medial habenula modulate the mesolimbic dopaminergic response to acute nicotine in vivo.

    PubMed

    McCallum, Sarah E; Cowe, Matthew A; Lewis, Samuel W; Glick, Stanley D

    2012-09-01

    Habenulo-interpeduncular nicotinic receptors, particularly those containing α3, β4 and α5 subunits, have recently been implicated in the reinforcing effects of nicotine. Our laboratory has shown that injection of α3β4 nicotinic receptor antagonists into the medial habenula (MHb) decreases self-administration of multiple abused drugs, including nicotine (Glick et al., 2006, 2008; 2011). However, it is unclear whether blockade of MHb nicotinic receptors has a direct effect on mesolimbic dopamine. Here, we performed in vivo microdialysis in female rats. Microdialysis probes were implanted into the nucleus accumbens (NAcc) and α3β4 nicotinic receptor antagonists (18-methoxycoronaridine; 18-MC or α-conotoxin AuIB; AuIB), were injected into the ipsilateral MHb, just prior to systemic nicotine (0.4 mg/kg, s.c.). Dialysate samples were collected before and after drug administration and levels of extracellular dopamine and its metabolites were measured using HPLC. Acute nicotine administration increased levels of extracellular dopamine and its metabolites in the NAcc. Pre-treatment with intra-habenular AuIB or 18-MC prevented nicotine-induced increases in accumbal dopamine. Neither drug had an effect on nicotine-induced increases in dopamine metabolites, suggesting that α3β4 receptors do not play a role in dopamine metabolism. The effect of intra-habenular blockade of α3β4 receptors on NAcc dopamine was selective for acute nicotine: neither AuIB nor 18-MC prevented increases in NAcc dopamine stimulated by acute d-amphetamine or morphine. These results suggest the mesolimbic response to acute nicotine, but not to acute administration of other drugs of abuse, is directly modulated by α3β4 nicotinic receptors in the MHb, and emphasize a critical role for habenular nicotinic receptors in nicotine's reinforcing effects. PMID:22561751

  4. Nicotine reinforcement is reduced by cannabinoid CB1 receptor blockade in the ventral tegmental area.

    PubMed

    Simonnet, Amelie; Cador, Martine; Caille, Stephanie

    2013-11-01

    Cannabinoid type 1 (CB1) receptors control the motivational properties and reinforcing effects of nicotine. Indeed, peripheral administration of a CB1 receptor antagonist dramatically decreases both nicotine taking and seeking. However, the neural substrates through which the cannabinoid CB1 receptors regulate the voluntary intake of nicotine remain to be elucidated. In the present study, we sought to determine whether central injections of a CB1 receptor antagonist delivered either into the ventral tegmental area (VTA) or the nucleus accumbens (NAC) may alter nicotine intravenous self-administration (IVSA). Rats were first trained to self-administer nicotine (30 μg/kg/0.1 ml). The effect of central infusions of the CB1 antagonist AM 251 (0, 1 and 10 μg/0.5 μl/side) on nicotine-taking behavior was then tested. Intra-VTA infusions of AM 251 dose dependently reduced IVSA with a significant decrease for the dose 10 μg/0.5 μl/side. Moreover, operant responding for water was unaltered by intra-VTA AM 251 at the same dose. Surprisingly, intra-NAC delivery of AM 251 did not alter nicotine behavior at all. These data suggest that in rats chronically exposed to nicotine IVSA, the cannabinoid CB1 receptors located in the VTA rather than in the NAC specifically control nicotine reinforcement and, subsequently, nicotine-taking behavior. PMID:22784230

  5. A key role for the N/OFQ-NOP receptor system in modulating nicotine taking in a model of nicotine and alcohol co-administration.

    PubMed

    Cippitelli, Andrea; Schoch, Jennifer; Debevec, Ginamarie; Brunori, Gloria; Zaveri, Nurulain T; Toll, Lawrence

    2016-01-01

    Alcohol and nicotine are often co-abused. Although the N/OFQ-NOP receptor system is considered a potential target for development of drug abuse pharmacotherapies, especially for alcoholism, little is known about the role of this system in nicotine dependence. Furthermore, the effect of prior history of nicotine dependence on subsequent nicotine and alcohol taking is understudied. Using an operant co-administration paradigm, in which rats concurrently self-administer nicotine and alcohol, we found that nicotine dependent rats increased nicotine self-administration over time as compared to non-dependent animals, while patterns of alcohol lever pressing did not change between groups. Pretreatment with the potent NOP receptor agonist AT-202 (0.3-3 mg/kg) increased nicotine lever pressing of both dependent and non-dependent groups, whereas the selective antagonist SB612111 (1-10 mg/kg) elicited a clear reduction of nicotine responses, in both dependent and non-dependent rats. In parallel, AT-202 only produced minor changes on alcohol responses and SB612111 reduced alcohol taking at a dose that also reduced locomotor behavior. Results indicate that a history of nicotine dependence affects subsequent nicotine- but not alcohol-maintained responding, and that NOP receptor antagonism, rather than agonism, blocks nicotine self-administration, which strongly suggests a critical role for the endogenous N/OFQ in the modulation of nicotine reinforcement processes. PMID:27199205

  6. A key role for the N/OFQ-NOP receptor system in modulating nicotine taking in a model of nicotine and alcohol co-administration

    PubMed Central

    Cippitelli, Andrea; Schoch, Jennifer; Debevec, Ginamarie; Brunori, Gloria; Zaveri, Nurulain T.; Toll, Lawrence

    2016-01-01

    Alcohol and nicotine are often co-abused. Although the N/OFQ-NOP receptor system is considered a potential target for development of drug abuse pharmacotherapies, especially for alcoholism, little is known about the role of this system in nicotine dependence. Furthermore, the effect of prior history of nicotine dependence on subsequent nicotine and alcohol taking is understudied. Using an operant co-administration paradigm, in which rats concurrently self-administer nicotine and alcohol, we found that nicotine dependent rats increased nicotine self-administration over time as compared to non-dependent animals, while patterns of alcohol lever pressing did not change between groups. Pretreatment with the potent NOP receptor agonist AT-202 (0.3–3 mg/kg) increased nicotine lever pressing of both dependent and non-dependent groups, whereas the selective antagonist SB612111 (1–10 mg/kg) elicited a clear reduction of nicotine responses, in both dependent and non-dependent rats. In parallel, AT-202 only produced minor changes on alcohol responses and SB612111 reduced alcohol taking at a dose that also reduced locomotor behavior. Results indicate that a history of nicotine dependence affects subsequent nicotine- but not alcohol-maintained responding, and that NOP receptor antagonism, rather than agonism, blocks nicotine self-administration, which strongly suggests a critical role for the endogenous N/OFQ in the modulation of nicotine reinforcement processes. PMID:27199205

  7. Functional Upregulation of α4* Nicotinic Acetylcholine Receptors in VTA GABAergic Neurons Increases Sensitivity to Nicotine Reward

    PubMed Central

    Ngolab, Jennifer; Liu, Liwang; Zhao-Shea, Rubing; Gao, Guangping; Gardner, Paul D.

    2015-01-01

    Chronic nicotine exposure increases sensitivity to nicotine reward during a withdrawal period, which may facilitate relapse in abstinent smokers, yet the molecular neuroadaptation(s) that contribute to this phenomenon are unknown. Interestingly, chronic nicotine use induces functional upregulation of nicotinic acetylcholine receptors (nAChRs) in the mesocorticolimbic reward pathway potentially linking upregulation to increased drug sensitivity. In the ventral tegmental area (VTA), functional upregulation of nAChRs containing the α4 subunit (α4* nAChRs) is restricted to GABAergic neurons. To test the hypothesis that increased functional expression of α4* nAChRs in these neurons modulates nicotine reward behaviors, we engineered a Cre recombinase-dependent gene expression system to selectively express α4 nAChR subunits harboring a “gain-of-function” mutation [a leucine mutated to a serine residue at the 9′ position (Leu9′Ser)] in VTA GABAergic neurons of adult mice. In mice expressing Leu9′Ser α4 nAChR subunits in VTA GABAergic neurons (Gad2VTA:Leu9′Ser mice), subreward threshold doses of nicotine were sufficient to selectively activate VTA GABAergic neurons and elicit acute hypolocomotion, with subsequent nicotine exposures eliciting tolerance to this effect, compared to control animals. In the conditioned place preference procedure, nicotine was sufficient to condition a significant place preference in Gad2VTA:Leu9′Ser mice at low nicotine doses that failed to condition control animals. Together, these data indicate that functional upregulation of α4* nAChRs in VTA GABAergic neurons confers increased sensitivity to nicotine reward and points to nAChR subtypes specifically expressed in GABAergic VTA neurons as molecular targets for smoking cessation therapeutics. PMID:26041923

  8. Single-Channel Current Through Nicotinic Receptor Produced by Closure of Binding Site C-Loop

    SciTech Connect

    Wang, Hailong; Cheng, Xiaolin; McCammon, Jonathan

    2009-01-01

    We investigated the initial coupling of agonist binding to channel gating of the nicotinic acetylcholine receptor using targeted molecular-dynamics (TMD) simulation. After TMD simulation to accelerate closure of the C-loops at the agonist binding sites, the region of the pore that passes through the cell membrane expands. To determine whether the structural changes in the pore result in ion conduction, we used a coarse-grained ion conduction simulator, Biology Boltzmann transport Monte Carlo, and applied it to two structural frames taken before and after TMD simulation. The structural model before TMD simulation represents the channel in the proposed resting state, whereas the model after TMD simulation represents the channel in the proposed active state. Under external voltage biases, the channel in the active state was permeable to cations. Our simulated ion conductance approaches that obtained experimentally and recapitulates several functional properties characteristic of the nicotinic acetylcholine receptor. Thus, closure of the C-loop triggers a structural change in the channel sufficient to account for the open channel current. This approach of applying Biology Boltzmann transport Monte Carlo simulation can be used to further investigate the binding to gating transduction mechanism and the structural bases for ion selection and translocation.

  9. Blockade of Dopamine D4 Receptors Attenuates Reinstatement of Extinguished Nicotine-Seeking Behavior in Rats

    PubMed Central

    Yan, Yijin; Pushparaj, Abhiram; Le Strat, Yann; Gamaleddin, Islam; Barnes, Chanel; Justinova, Zuzana; Goldberg, Steven R; Le Foll, Bernard

    2012-01-01

    Since cloning of the dopamine receptor D4 (DRD4), its role in the brain has remained unclear. It has been reported that polymorphism of the DRD4 gene in humans is associated with reactivity to cues related to tobacco smoking. However, the role of DRD4 in animal models of nicotine addiction has seldom been explored. In our study, male Long-Evans rats learned to intravenously self-administer nicotine under a fixed-ratio (FR) schedule of reinforcement. Effects of the selective DRD4 antagonist L-745,870 were evaluated on nicotine self-administration behavior and on reinstatement of extinguished nicotine-seeking behavior induced by nicotine-associated cues or by priming injections of nicotine. L-745,870 was also tested on reinstatement of extinguished food-seeking behavior as a control. In addition, the selective DRD4 agonist PD 168,077 was tested for its ability to reinstate extinguished nicotine-seeking behavior. Finally, L-745,870 was tested in Sprague Dawley rats trained to discriminate administration of 0.4 mg/kg nicotine from vehicle under an FR schedule of food delivery. L-745,870 significantly attenuated reinstatement of nicotine-seeking induced by both nicotine-associated cues and nicotine priming. In contrast, L-745,870 did not affect established nicotine self-administration behavior or reinstatement of food-seeking behavior induced by food cues or food priming. L-745,870 did not produce nicotine-like discriminative-stimulus effects and did not alter discriminative-stimulus effects of nicotine. PD 168,077 did not reinstate extinguished nicotine-seeking behavior. As DRD4 blockade by L-745,870 selectively attenuated both cue- and nicotine-induced reinstatement of nicotine-seeking behavior, without affecting cue- or food-induced reinstatement of food-seeking behavior, DRD4 antagonists are potential therapeutic agents against tobacco smoking relapse. PMID:22030716

  10. Unconventional pharmacology of a neuronal nicotinic receptor mutated in the channel domain.

    PubMed

    Bertrand, D; Devillers-Thiéry, A; Revah, F; Galzi, J L; Hussy, N; Mulle, C; Bertrand, S; Ballivet, M; Changeux, J P

    1992-02-15

    The putative channel-forming MII domains of the nicotinic, gamma-aminobutyric acid type A, and glycine receptors contain a highly conserved leucine residue. Mutation of this hydrophobic amino acid in the neuronal nicotinic receptor alpha 7 (Leu-247), reconstituted in Xenopus oocytes, modifies the ionic response to acetylcholine and alters desensitization. Furthermore, the Leu----Thr (L247T) mutant has two conducting states (46 pS and 80 pS), in contrast with the wild-type (WT) receptor, which has only one (45 pS). We now show that this mutant possesses a rather paradoxical pharmacology: antagonists of the WT receptor such as dihydro-beta-erythroidin, hexamethonium, or (+)-tubocurarine elicit ionic currents when applied to the L247T alpha 7 mutant and these responses are blocked by alpha-bungarotoxin. Furthermore, prolonged application of acetylcholine causes desensitization in the WT but leads to a potentiation of the responses to acetylcholine or dihydro-beta-erythroidin in the mutant. These data are consistent with a scheme in which mutation of Leu-247 renders a desensitized state in the WT channel a conducting state. They also strengthen the proposal that, in the WT, some competitive antagonists may stabilize desensitized states. Finally, these observations may shed light on properties of other ion channels, in particular the glutamate receptors, which display multiple conductance levels associated with various pharmacological agents. PMID:1741378

  11. Multiple binding sites in the nicotinic acetylcholine receptors: An opportunity for polypharmacolgy.

    PubMed

    Iturriaga-Vásquez, Patricio; Alzate-Morales, Jans; Bermudez, Isabel; Varas, Rodrigo; Reyes-Parada, Miguel

    2015-11-01

    For decades, the development of selective compounds has been the main goal for chemists and biologists involved in drug discovery. However, diverse lines of evidence indicate that polypharmacological agents, i.e. those that act simultaneously at various protein targets, might show better profiles than selective ligands, regarding both efficacy and side effects. On the other hand, the availability of the crystal structure of different receptors allows a detailed analysis of the main interactions between drugs and receptors in a specific binding site. Neuronal nicotinic acetylcholine receptors (nAChRs) constitute a large and diverse family of ligand-gated ion channels (LGICs) that, as a product of its modulation, regulate neurotransmitter release, which in turns produce a global neuromodulation of the central nervous system. nAChRs are pentameric protein complexes in such a way that expression of compatible subunits can lead to various receptor assemblies or subtypes. The agonist binding site, located at the extracellular region, exhibits different properties depending on the subunits that conform the receptor. In the last years, it has been recognized that nAChRs could also contain one or more allosteric sites which could bind non-classical nicotinic ligands including several therapeutically useful drugs. The presence of multiple binding sites in nAChRs offers an interesting possibility for the development of novel polypharmacological agents with a wide spectrum of actions. PMID:26318763

  12. Promoter polymorphisms and transcript levels of nicotinic receptor CHRNA5.

    PubMed

    Falvella, Felicia S; Galvan, Antonella; Colombo, Francesca; Frullanti, Elisa; Pastorino, Ugo; Dragani, Tommaso A

    2010-09-01

    Chromosomal locus 15q25, implicated in lung cancer risk and nicotine dependence, shows extensive linkage disequilibrium that complicates identification of causal variation. Cholinergic receptor nicotinic alpha5 (CHRNA5) has been identified as a lung cancer risk factor. We identified by sequence analysis three haplotypes (delTTC, insATC, and insTGG) in the 5' promoter region and three at the 3'-untranslated region of CHRNA5. Linkage disequilibrium analysis of the 5' variants showed that the insTGG haplotype is associated with three tightly linked risk alleles (nicotine dependence, lung cancer, and chronic obstructive pulmonary disease). The three CHRNA5 promoter haplotypes were statistically significantly associated with lung CHRNA5 transcript levels, determined by real-time polymerase chain reaction. In nontumor lung parenchyma from 68 patients who underwent lung lobectomy, the delTTC haplotype was associated with the highest CHRNA5 transcript levels (relative quantification units = 1.82), whereas the insTGG haplotype was associated with the lowest (0.88 units, P(diff) < .001, Welch t test; all statistical tests were two-sided). Luciferase reporter assays in human lung cancer cell lines A549, H460, H520, and H596 also showed that the 5' region haplotypes were statistically significantly associated with changes in CHRNA5 promoter activity, whereas the 3'-untranslated region variants were not. PMID:20733116

  13. NMDA receptors regulate nicotine-enhanced brain reward function and intravenous nicotine self-administration: role of the ventral tegmental area and central nucleus of the amygdala.

    PubMed

    Kenny, Paul J; Chartoff, Elena; Roberto, Marisa; Carlezon, William A; Markou, Athina

    2009-01-01

    Nicotine is considered an important component of tobacco responsible for the smoking habit in humans. Nicotine increases glutamate-mediated transmission throughout brain reward circuitries. This action of nicotine could potentially contribute to its intrinsic rewarding and reward-enhancing properties, which motivate consumption of the drug. Here we show that the competitive N-methyl-D-aspartate (NMDA) receptor antagonist LY235959 (0.5-2.5 mg per kg) abolished nicotine-enhanced brain reward function, reflected in blockade of the lowering of intracranial self-stimulation (ICSS) thresholds usually observed after experimenter-administered (0.25 mg per kg) or intravenously self-administered (0.03 mg per kg per infusion) nicotine injections. The highest LY235959 dose (5 mg per kg) tested reversed the hedonic valence of nicotine from positive to negative, reflected in nicotine-induced elevations of ICSS thresholds. LY235959 doses that reversed nicotine-induced lowering of ICSS thresholds also markedly decreased nicotine self-administration without altering responding for food reinforcement, whereas the alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptor antagonist NBQX had no effects on nicotine intake. In addition, nicotine self-administration upregulated NMDA receptor subunit expression in the central nucleus of the amygdala (CeA) and ventral tegmental area (VTA), suggesting important interactions between nicotine and the NMDA receptor. Furthermore, nicotine (1 microM) increased NMDA receptor-mediated excitatory postsynaptic currents in rat CeA slices, similar to its previously described effects in the VTA. Finally, infusion of LY235959 (0.1-10 ng per side) into the CeA or VTA decreased nicotine self-administration. Taken together, these data suggest that NMDA receptors, including those in the CeA and VTA, gate the magnitude and valence of the effects of nicotine on brain reward systems, thereby regulating motivation to consume the drug. PMID:18418357

  14. Pharmacological characterisation of strychnine and brucine analogues at glycine and alpha7 nicotinic acetylcholine receptors.

    PubMed

    Jensen, Anders A; Gharagozloo, Parviz; Birdsall, Nigel J M; Zlotos, Darius P

    2006-06-01

    Strychnine and brucine from the plant Strychnos nux vomica have been shown to have interesting pharmacological effects on several neurotransmitter receptors, including some members of the superfamily of ligand-gated ion channels. In this study, we have characterised the pharmacological properties of tertiary and quaternary analogues as well as bisquaternary dimers of strychnine and brucine at human alpha1 and alpha1beta glycine receptors and at a chimera consisting of the amino-terminal domain of the alpha7 nicotinic receptor (containing the orthosteric ligand binding site) and the ion channel domain of the 5-HT3A serotonin receptor. Although the majority of the analogues displayed significantly increased Ki values at the glycine receptors compared to strychnine and brucine, a few retained the high antagonist potencies of the parent compounds. However, mirroring the pharmacological profiles of strychnine and brucine, none of the analogues displayed significant selectivity between the alpha1 and alpha1beta subtypes. The structure-activity relationships for the compounds at the alpha7/5-HT3 chimera were significantly different from those at the glycine receptors. Most strikingly, quaternization of strychnine and brucine with substituents possessing different steric and electronic properties completely eliminated the activity at the glycine receptors, whereas binding affinity to the alpha7/5-HT3 chimera was retained for the majority of the quaternary analogues. This study provides an insight into the structure-activity relationships for strychnine and brucine analogues at these ligand-gated ion channels. PMID:16687139

  15. Insect Nicotinic Receptor Agonists as Flea Adulticides in Small Animals

    PubMed Central

    Vo, Dai Tan; Hsu, Walter H.; Martin, Richard J.

    2013-01-01

    Fleas are significant ectoparasites of small animals. They can be a severe irritant to animals and serve as a vector for a number of infectious diseases. In this article, we discuss the pharmacological characteristics of four insect nicotinic acetylcholine receptor (nAChR) agonists used as fleacides in dogs and cats, which include three neonicotinoids (imidacloprid, nitenpyram, and dinotefuran) and spinosad. Insect nAChR agonists are one of the most important new classes of insecticides, which are used to control sucking insects both on plants and on companion animals. These new compounds provide a new approach for practitioners to safely and effectively eliminate fleas. PMID:20646191

  16. Positive allosteric modulators of α7 nicotinic acetylcholine receptors affect neither the function of other ligand- and voltage-gated ion channels and acetylcholinesterase, nor β-amyloid content.

    PubMed

    Arias, Hugo R; Ravazzini, Federica; Targowska-Duda, Katarzyna M; Kaczor, Agnieszka A; Feuerbach, Dominik; Boffi, Juan C; Draczkowski, Piotr; Montag, Dirk; Brown, Brandon M; Elgoyhen, Ana Belén; Jozwiak, Krzysztof; Puia, Giulia

    2016-07-01

    The activity of positive allosteric modulators (PAMs) of α7 nicotinic acetylcholine receptors (AChRs), including 3-furan-2-yl-N-p-tolyl-acrylamide (PAM-2), 3-furan-2-yl-N-o-tolylacrylamide (PAM-3), and 3-furan-2-yl-N-phenylacrylamide (PAM-4), was tested on a variety of ligand- [i.e., human (h) α7, rat (r) α9α10, hα3-containing AChRs, mouse (m) 5-HT3AR, and several glutamate receptors (GluRs)] and voltage-gated (i.e., sodium and potassium) ion channels, as well as on acetylcholinesterase (AChE) and β-amyloid (Aβ) content. The functional results indicate that PAM-2 inhibits hα3-containing AChRs (IC50=26±6μM) with higher potency than that for NR1aNR2B and NR1aNR2A, two NMDA-sensitive GluRs. PAM-2 affects neither the activity of m5-HT3ARs, GluR5/KA2 (a kainate-sensitive GluR), nor AChE, and PAM-4 does not affect agonist-activated rα9α10 AChRs. Relevant clinical concentrations of PAM-2-4 do not inhibit Nav1.2 and Kv3.1 ion channels. These PAMs slightly enhance the activity of GluR1 and GluR2, two AMPA-sensitive GluRs. PAM-2 does not change the levels of Aβ42 in an Alzheimer's disease mouse model (i.e., 5XFAD). The molecular docking and dynamics results using the hα7 model suggest that the active sites for PAM-2 include the intrasubunit (i.e., PNU-120596 locus) and intersubunit sites. These results support our previous study showing that these PAMs are selective for the α7 AChR, and clarify that the procognitive/promnesic/antidepressant activity of PAM-2 is not mediated by other targets. PMID:27129924

  17. Effects of antihistamines on the function of human α7-nicotinic acetylcholine receptors.

    PubMed

    Sadek, Bassem; Khanian, Seyedeh Soha; Ashoor, Abrar; Prytkova, Tatiana; Ghattas, Mohammad A; Atatreh, Noor; Nurulain, Syed M; Yang, Keun-Hang Susan; Howarth, Frank Christopher; Oz, Murat

    2015-01-01

    Effects of the histamine H₁ receptor (H1R) antagonists (antihistamines), promethazine (PMZ), orphenadrine (ORP), chlorpheniramine (CLP), pyrilamine (PYR), diphenhydramine (DPH), citerizine (CTZ), and triprolidine (TRP) on the functional properties of the cloned α7 subunit of the human nicotinic acetylcholine receptor expressed in Xenopus oocytes were investigated. Antihistamines inhibited the α7-nicotinic acetylcholine receptor in the order PYR>CLP>TRP>PMZ>ORP≥DPH≥CTZ. Among the antihistamines, PYR showed the highest reversible inhibition of acetylcholine (100 µM)-induced responses with IC₅₀ of 6.2 µM. PYR-induced inhibition was independent of the membrane potential and could not be reversed by increasing the concentration of acetylcholine. Specific binding of [¹²⁵I] α-bungarotoxin, a selective antagonist for α7-nicotinic acetylcholine receptor, was not changed in the presence of PYR suggesting a non-competitive inhibition of nicotinic receptors. In line with functional experiments, docking studies indicated that PYR can potentially bind allosterically with the α7 transmembrane domain. Our results indicate that the H₂-H₄ receptor antagonists tested in this study (10 µM) showed negligible inhibition of α7-nicotinic acetylcholine receptors. On the other hand, H₁ receptor antagonists inhibited the function of human α7-nicotinic acetylcholine receptor, with varying potencies. These results emphasize the importance of α7-nicotinic acetylcholine receptor for future pharmacological/toxicological profiling. PMID:25445036

  18. Dextromethorphan and its metabolite dextrorphan block alpha3beta4 neuronal nicotinic receptors.

    PubMed

    Hernandez, S C; Bertolino, M; Xiao, Y; Pringle, K E; Caruso, F S; Kellar, K J

    2000-06-01

    Dextromethorphan (DM), a structural analog of morphine and codeine, has been widely used as a cough suppressant for more than 40 years. DM is not itself a potent analgesic, but it has been reported to enhance analgesia produced by morphine and nonsteroidal anti-inflammatory drugs. Although DM is considered to be nonaddictive, it has been reported to reduce morphine tolerance in rats and to be useful in helping addicted subjects to withdraw from heroin. Here we studied the effects of DM on neuronal nicotinic receptors stably expressed in human embryonic kidney cells. Studies were carried out to examine the effects of DM on nicotine-stimulated whole cell currents and nicotine-stimulated (86)Rb(+) efflux. We found that both DM and its metabolite dextrorphan block nicotinic receptor function in a noncompetitive but reversible manner, suggesting that both drugs block the receptor channel. Consistent with blockade of the receptor channel, neither drug competed for the nicotinic agonist binding sites labeled by [(3)H]epibatidine. Although DM is approximately 9-fold less potent than the widely used noncompetitive nicotinic antagonist mecamylamine in blocking nicotinic receptor function, the block by DM appears to reverse more slowly than that by mecamylamine. These data indicate that DM is a useful antagonist for studying nicotinic receptor function and suggest that it might prove to be a clinically useful neuronal nicotinic receptor antagonist, possibly helpful as an aid for helping people addicted to nicotine to refrain from smoking, as well as in other conditions where blockade of neuronal nicotinic receptors would be helpful. PMID:10869398

  19. The nicotinic acetylcholine receptor and its prokaryotic homologues: Structure, conformational transitions & allosteric modulation.

    PubMed

    Cecchini, Marco; Changeux, Jean-Pierre

    2015-09-01

    Pentameric ligand-gated ion channels (pLGICs) play a central role in intercellular communications in the nervous system by converting the binding of a chemical messenger - a neurotransmitter - into an ion flux through the postsynaptic membrane. Here, we present an overview of the most recent advances on the signal transduction mechanism boosted by X-ray crystallography of both prokaryotic and eukaryotic homologues of the nicotinic acetylcholine receptor (nAChR) in conjunction with time-resolved analyses based on single-channel electrophysiology and Molecular Dynamics simulations. The available data consistently point to a global mechanism of gating that involves a large reorganization of the receptor mediated by two distinct quaternary transitions: a global twisting and a radial expansion/contraction of the extracellular domain. These transitions profoundly modify the organization of the interface between subunits, which host several sites for orthosteric and allosteric modulatory ligands. The same mechanism may thus mediate both positive and negative allosteric modulations of pLGICs ligand binding at topographically distinct sites. The emerging picture of signal transduction is expected to pave the way to new pharmacological strategies for the development of allosteric modulators of nAChR and pLGICs in general. This article is part of the Special Issue entitled 'The Nicotinic Acetylcholine Receptor: From Molecular Biology to Cognition'. PMID:25529272

  20. Comparison of nicotinic receptor binding and biotransformation of coniine in the rat and chick.

    PubMed

    Forsyth, C S; Speth, R C; Wecker, L; Galey, F D; Frank, A A

    1996-12-31

    Coniine, an alkaloid from Conium maculatum (poison hemlock), is a known teratogen in many domestic species with maternal ingestion resulting in arthrogryposis of the offspring. We have previously shown that rats are not susceptible and rabbits only weakly susceptible to coniine-induced arthrogryposis. However, the chick embryo does provide a reproducible laboratory animal model of coniine-induced teratogenesis. The reason for this cross-species variation is unknown. The purpose of this study was to evaluate coniine binding to nicotinic receptors and to measure coniine metabolism in vitro between susceptible and non-susceptible species. Using the chick model, neither the peripheral nicotinic receptor antagonist d-tubocurarine chloride nor the central nicotinic receptor antagonist trimethaphan camsylate blocked the teratogenesis or lethality of 1.5% coniine (50 microliters/egg). Trimethaphan camsylate enhanced coniine-induced lethality in a dose-dependent manner. Neither nicotinic receptor blocker prevented nicotine sulfate-induced malformations but d-tubocurarine chloride did block lethality in a dose-dependent manner. Competition by coniine for [125I]-alpha-bungarotoxin to nicotinic receptors isolated from adult rat diaphragm and chick thigh muscle and competition by coniine for [3H]-cytisine to receptors from rat and chick brain were used to assess coniine binding to nicotinic receptors. The IC50 for coniine in rat diaphragm was 314 microM while that for chick leg muscle was 70 microM. For neuronal nicotinic receptors, the IC50s of coniine for maternal rat brain, fetal rat brain, and chick brain were 1100 microM, 820 microM, and 270 microM, respectively. There were no differences in coniine biotransformation in vitro by microsomes from rat or chick livers. Differences in apparent affinity of coniine for nicotinic receptors or differences in the quantity of the nicotinic receptor between the rat and chick may explain, in part, the differences in susceptibility of

  1. Differential effects of subtype-specific nicotinic acetylcholine receptor agonists on early and late hippocampal LTP.

    PubMed

    Kroker, Katja S; Rast, Georg; Rosenbrock, Holger

    2011-12-01

    Brain nicotinic acetylcholine receptors are involved in several neuropsychiatric disorders, e.g. Alzheimer's and Parkinson's diseases, Tourette's syndrome, schizophrenia, depression, autism, attention deficit hyperactivity disorder, and anxiety. Currently, approaches selectively targeting the activation of specific nicotinic acetylcholine receptors are in clinical development for treatment of memory impairment of Alzheimer's disease patients. These are α4β2 and α7 nicotinic acetylcholine receptor agonists which are believed to enhance cholinergic and glutamatergic neurotransmission, respectively. In order to gain a better insight into the mechanistic role of these two nicotinic acetylcholine receptors in learning and memory, we investigated the effects of the α4β2 nicotinic acetylcholine receptor agonist TC-1827 and the α7 nicotinic acetylcholine receptor partial agonist SSR180711 on hippocampal long-term potentiation (LTP), a widely accepted cellular experimental model of memory formation. Generally, LTP is distinguished in an early and a late form, the former being protein-synthesis independent and the latter being protein-synthesis dependent. TC-1827 was found to increase early LTP in a bell-shaped dose dependent manner, but did not affect late LTP. In contrast, the α7 nicotinic acetylcholine receptor partial agonist SSR180711 showed enhancing effects on both early and late LTP in a bell-shaped manner. Furthermore, SSR180711 not only increased early LTP, but also transformed it into late LTP, which was not observed with the α4β2 nicotinic acetylcholine receptor agonist. Therefore, based on these findings α7 nicotinic acetylcholine receptor (partial) agonists appear to exhibit stronger efficacy on memory improvement than α4β2 nicotinic acetylcholine receptor agonists. PMID:21968142

  2. Nicotinic Acetylcholine Receptor (nAChR) Dependent Chorda Tympani Taste Nerve Responses to Nicotine, Ethanol and Acetylcholine

    PubMed Central

    Ren, Zuo Jun; Mummalaneni, Shobha; Qian, Jie; Baumgarten, Clive M.; DeSimone, John A.; Lyall, Vijay

    2015-01-01

    Nicotine elicits bitter taste by activating TRPM5-dependent and TRPM5-independent but neuronal nAChR-dependent pathways. The nAChRs represent common targets at which acetylcholine, nicotine and ethanol functionally interact in the central nervous system. Here, we investigated if the nAChRs also represent a common pathway through which the bitter taste of nicotine, ethanol and acetylcholine is transduced. To this end, chorda tympani (CT) taste nerve responses were monitored in rats, wild-type mice and TRPM5 knockout (KO) mice following lingual stimulation with nicotine free base, ethanol, and acetylcholine, in the absence and presence of nAChR agonists and antagonists. The nAChR modulators: mecamylamine, dihydro-β-erythroidine, and CP-601932 (a partial agonist of the α3β4* nAChR), inhibited CT responses to nicotine, ethanol, and acetylcholine. CT responses to nicotine and ethanol were also inhibited by topical lingual application of 8-chlorophenylthio (CPT)-cAMP and loading taste cells with [Ca2+]i by topical lingual application of ionomycin + CaCl2. In contrast, CT responses to nicotine were enhanced when TRC [Ca2+]i was reduced by topical lingual application of BAPTA-AM. In patch-clamp experiments, only a subset of isolated rat fungiform taste cells exposed to nicotine responded with an increase in mecamylamine-sensitive inward currents. We conclude that nAChRs expressed in a subset of taste cells serve as common receptors for the detection of the TRPM5-independent bitter taste of nicotine, acetylcholine and ethanol. PMID:26039516

  3. Cell-surface translational dynamics of nicotinic acetylcholine receptors

    PubMed Central

    Barrantes, Francisco J.

    2014-01-01

    Synapse efficacy heavily relies on the number of neurotransmitter receptors available at a given time. In addition to the equilibrium between the biosynthetic production, exocytic delivery and recycling of receptors on the one hand, and the endocytic internalization on the other, lateral diffusion and clustering of receptors at the cell membrane play key roles in determining the amount of active receptors at the synapse. Mobile receptors traffic between reservoir compartments and the synapse by thermally driven Brownian motion, and become immobilized at the peri-synaptic region or the synapse by: (a) clustering mediated by homotropic inter-molecular receptor–receptor associations; (b) heterotropic associations with non-receptor scaffolding proteins or the subjacent cytoskeletal meshwork, leading to diffusional “trapping,” and (c) protein-lipid interactions, particularly with the neutral lipid cholesterol. This review assesses the contribution of some of these mechanisms to the supramolecular organization and dynamics of the paradigm neurotransmitter receptor of muscle and neuronal cells -the nicotinic acetylcholine receptor (nAChR). Currently available information stemming from various complementary biophysical techniques commonly used to interrogate the dynamics of cell-surface components is critically discussed. The translational mobility of nAChRs at the cell surface differs between muscle and neuronal receptors in terms of diffusion coefficients and residence intervals at the synapse, which cover an ample range of time regimes. A peculiar feature of brain α7 nAChR is its ability to spend much of its time confined peri-synaptically, vicinal to glutamatergic (excitatory) and GABAergic (inhibitory) synapses. An important function of the α7 nAChR may thus be visiting the territories of other neurotransmitter receptors, differentially regulating the dynamic equilibrium between excitation and inhibition, depending on its residence time in each domain. PMID

  4. Nicotinic ACh Receptors as Therapeutic Targets in CNS Disorders

    PubMed Central

    Dineley, Kelly T.; Pandya, Anshul A.; Yakel, Jerrel L.

    2015-01-01

    The neurotransmitter acetylcholine (ACh) can regulate neuronal excitability by acting on the cys-loop cation-conducting ligand-gated nicotinic ACh receptor channels (nAChRs). These receptors are widely distributed throughout the central nervous system, being expressed on neurons and non-neuronal cells, where they participate in a variety of physiological responses such as anxiety, the central processing of pain, food intake, nicotine seeking behavior, and cognitive functions. In the mammalian brain, nine different subunits have been found thus far, which assemble into pentameric complexes with much subunit diversity; however the α7 and α4β2 subtypes predominate in the CNS. Neuronal nAChR dysfunction is involved in the pathophysiology of many neurological disorders. Here we will briefly discuss the functional makeup and expression of the nAChRs in the mammalian brain, and their role as targets in neurodegenerative diseases (in particular Alzheimer’s disease), neurodevelopmental disorders (in particular autism and schizophrenia), and neuropathic pain. PMID:25639674

  5. Nicotinic ACh receptors as therapeutic targets in CNS disorders.

    PubMed

    Dineley, Kelly T; Pandya, Anshul A; Yakel, Jerrel L

    2015-02-01

    The neurotransmitter acetylcholine (ACh) can regulate neuronal excitability by acting on the cys-loop cation-conducting ligand-gated nicotinic ACh receptor (nAChR) channels. These receptors are widely distributed throughout the central nervous system (CNS), being expressed on neurons and non-neuronal cells, where they participate in a variety of physiological responses such as anxiety, the central processing of pain, food intake, nicotine seeking behavior, and cognitive functions. In the mammalian brain, nine different subunits have been found thus far, which assemble into pentameric complexes with much subunit diversity; however, the α7 and α4β2 subtypes predominate in the CNS. Neuronal nAChR dysfunction is involved in the pathophysiology of many neurological disorders. Here we will briefly discuss the functional makeup and expression of the nAChRs in mammalian brain, and their role as targets in neurodegenerative diseases (in particular Alzheimer's disease, AD), neurodevelopmental disorders (in particular autism and schizophrenia), and neuropathic pain. PMID:25639674

  6. Mixed nicotinic and muscarinic features of cholinergic receptor coupled to secretion in bovine chromaffin cells.

    PubMed Central

    Shirvan, M H; Pollard, H B; Heldman, E

    1991-01-01

    Acetylcholine evokes release from cultured bovine chromaffin cells by a mechanism that is believed to be classically nicotinic. However, we found that the full muscarinic agonist oxotremorine-M (Oxo-M) induced a robust catecholamine (CA) secretion. By contrast, muscarine, pilocarpine, bethanechol, and McN-A-343 did not elicit any secretory response. Desensitization of the response to nicotine by Oxo-M and desensitization of the response to Oxo-M by nicotine suggest that both nicotine and Oxo-M were acting at the same receptor. Additional experiments supporting this conclusion show that nicotine-induced secretion and Oxo-M-induced secretion were similarly blocked by various muscarinic and nicotinic antagonists. Moreover, secretion induced by nicotine and Oxo-M were Ca2+ dependent, and both agonists induced 45Ca2+ uptake. Equilibrium binding studies showed that [3H]Oxo-M bound to chromaffin cell membranes with a Kd value of 3.08 x 10(-8) M and a Hill coefficient of 1.00, suggesting one binding site for this ligand. Nicotine inhibited Oxo-M binding in a noncompetitive manner, suggesting that both ligands bind at two different sites on the same receptor. We propose that the receptor on bovine chromaffin cells that is coupled to secretion represents an unusual cholinergic receptor that has both nicotinic and muscarinic features. Images PMID:2052567

  7. Mixed nicotinic and muscarinic features of cholinergic receptor coupled to secretion in bovine chromaffin cells

    SciTech Connect

    Shirvan, M.H.; Pollard, H.B.; Heldman, E. )

    1991-06-01

    Acetylcholine evokes release from cultured bovine chromaffin cells by a mechanism that is believed to be classically nicotinic. However, the authors found that the full muscarinic agonist oxotremorine-M (Oxo-M) induced a robust catecholamine (CA) secretion. By contrast, muscarine, pilocarpine, bethanechol, and McN-A-343 did not elicit any secretory response. Desensitization of the response to nicotine by Oxo-M and desensitization of the response to Oxo-M by nicotine suggest that both nicotine and Oxo-M were acting at the same receptor. Additional experiments supporting this conclusion show that nicotine-induced secretion and Oxo-M-induced secretion were similarly blocked by various muscarinic and nicotinic antagonists. Moreover, secretion induced by nicotine and Oxo-M were Ca{sup 2+} dependent, and both agonists induced {sup 45}Ca{sup 2+} uptake. Equilibrium binding studies showed that ({sup 3}H)Oxo-M bound to chromaffin cell membranes with a K{sub d} value of 3.08 {times} 10{sup {minus}8}M and a Hill coefficient of 1.00, suggesting one binding site for this ligand. Nicotine inhibited Oxo-M binding in a noncompetitive manner, suggesting that both ligands bind at two different sites on the same receptor. They propose that the receptor on bovine chromaffin cells that is coupled to secretion represents an unusual cholinergic receptor that has both nicotinic and muscarinic features.

  8. PRENATAL NICOTINE EXPOSURE SELECTIVELY AFFECTS NICOTINIC RECEPTOR EXPRESSION IN PRIMARY AND ASSOCIATIVE VISUAL CORTICES OF THE FETAL BABOON

    PubMed Central

    Duncan, Jhodie R.; Garland, Marianne; Stark, Raymond I.; Myers, Michael M.; Fifer, William P.; Mokler, David J.; Kinney, Hannah C.

    2014-01-01

    Exposure to nicotine during pregnancy via maternal cigarette smoking is associated with visual deficits in children. This is possibly due to activation of nicotinic acetylcholine receptors (nAChRs) in the occipital cortex which are important in the development of visual mapping. Using a baboon model we explored the effects of prenatal nicotine on parameters in the primary and associated visual cortices. Pregnant baboons were infused with nicotine (0.5 mg/hr, i.v.) or saline from 86 days gestation. At 161 days gestation fetal brains were collected (n=5/group) and the occipital lobe assessed for nAChRs and markers of the serotonergic and catecholaminergic systems using tissue autoradiography and/or high performance liquid chromatography. Neuronal nAChRs and serotonergic markers were expressed in a region and subunit dependent manner. Prenatal nicotine exposure was associated with increased binding for 3H-epibatidine sensitive nAChRs in the primary visual cortex (BA 17) and BA 18, but not BA 19, of the associative visual cortex (p<0.05). Markers of the serotonergic or catecholaminergic systems were not significantly altered. Thus, prenatal nicotine exposure is associated with alterations in the cholinergic system in the occipital lobe which may aid in the explanation of the appearance of visual deficits in children from mothers who smoke during pregnancy. PMID:24903536

  9. Nicotinic acetylcholine receptors mediate donepezil-induced oligodendrocyte differentiation.

    PubMed

    Imamura, Osamu; Arai, Masaaki; Dateki, Minori; Ogata, Toru; Uchida, Ryuji; Tomoda, Hiroshi; Takishima, Kunio

    2015-12-01

    Oligodendrocytes are the myelin-forming cells of the central nervous system (CNS). Failure of myelin development and oligodendrocyte loss results in serious human disorders, including multiple sclerosis. Here, we show that donepezil, an acetlycholinesterase inhibitor developed for the treatment of Alzheimer's disease, can stimulate oligodendrocyte differentiation and maturation of neural stem cell-derived oligodendrocyte progenitor cells without affecting proliferation or cell viability. Transcripts for essential myelin-associated genes, such as PLP, MAG, MBP, CNPase, and MOG, in addition to transcription factors that regulate oligodendrocyte differentiation and myelination, were rapidly increased after treatment with donepezil. Furthermore, luciferase assays confirmed that both MAG and MBP promoters display increased activity upon donepezil-induced oligodendrocytes differentiation, suggesting that donepezil increases myelin gene expression mainly through enhanced transcription. We also found that the increase in the number of oligodendrocytes observed following donepezil treatment was significantly inhibited by the nicotinic acetylcholine receptor (nAChR) antagonist mecamylamine, but not by the muscarinic acetylcholine receptor antagonist scopolamine. Moreover, donepezil-induced myelin-related gene expression was suppressed by mecamylamine at both the mRNA and protein level. These results suggest that donepezil stimulates oligodendrocyte differentiation and myelin-related gene expression via nAChRs in neural stem cell-derived oligodendrocyte progenitor cells. We show that donepezil, a drug for the treatment of Alzheimer disease, can stimulate oligodendrocyte differentiation and maturation of oligodendrocyte progenitor cells. Transcripts for essential myelin-associated genes, such as PLP, MAG, MBP, CNPase and MOG in addition to transcripton factors that regulate oligodendrocyte differentiation and myelination were rapidly increased after treatment with donepezil

  10. Chalcones as positive allosteric modulators of α7 nicotinic acetylcholine receptors: a new target for a privileged structure.

    PubMed

    Balsera, Beatriz; Mulet, José; Fernández-Carvajal, Asia; de la Torre-Martínez, Roberto; Ferrer-Montiel, Antonio; Hernández-Jiménez, José G; Estévez-Herrera, Judith; Borges, Ricardo; Freitas, Andiara E; López, Manuela G; García-López, M Teresa; González-Muñiz, Rosario; Pérez de Vega, María Jesús; Valor, Luis M; Svobodová, Lucie; Sala, Salvador; Sala, Francisco; Criado, Manuel

    2014-10-30

    The α7 acetylcholine nicotine receptor is a ligand-gated ion channel that is involved in cognition disorders, schizophrenia, pain and inflammation among other diseases. Therefore, the development of new agents that target this receptor has great significance. Positive allosteric modulators might be advantageous, since they facilitate receptor responses without directly interacting with the agonist binding site. Here we report the search for and further design of new positive allosteric modulators having the relatively simple chalcone structure. From the natural product isoliquiritigenin as starting point, chalcones substituted with hydroxyl groups at defined locations were identified as optimal and specific promoters of α7 nicotinic function. The most potent compound (2,4,2',5'-tetrahydroxychalcone, 111) was further characterized showing its potential as neuroprotective, analgesic and cognitive enhancer, opening the way for future developments around the chalcone structure. PMID:25232969

  11. Aporphine metho salts as neuronal nicotinic acetylcholine receptor blockers.

    PubMed

    Iturriaga-Vásquez, Patricio; Pérez, Edwin G; Slater, E Yvonne; Bermúdez, Isabel; Cassels, Bruce K

    2007-05-15

    (S)-Aporphine metho salts with the 1,2,9,10 oxygenation pattern displaced radioligands from recombinant human alpha7 and alpha4beta2 neuronal nicotinic acetylcholine receptors (nAChR) at low micromolar concentrations. The affinity of the nonphenolic glaucine methiodide (4) (vs [(3)H]cytisine) was the lowest at alpha4beta2 nAChR (K(i)=10 microM), and predicentrine methiodide (2) and xanthoplanine iodide (3), with free hydroxyl groups at C-2 or C-9, respectively, had the highest affinity at these receptors (K(i) approximately 1 microM), while the affinity of the diphenolic boldine methiodide (1) was intermediate between these values. At homomeric alpha7 nAChR, xanthoplanine had the highest affinity (K(i)=10 microM) vs [(125)I]alpha-bungarotoxin while the other three compounds displaced the radioligand with K(i) values between 15 and 21 microM. At 100 microM, all four compounds inhibited the responses of these receptors to EC(50) concentrations of ACh. The effects of xanthoplanine iodide (3) were studied in more detail. Xanthoplanine fully inhibited the EC(50) ACh responses of both alpha7 and alpha4beta2 nACh receptors with estimated IC(50) values of 9+/-3 microM (alpha7) and 5+/-0.8 microM (alpha4beta2). PMID:17391965

  12. Menthol Binding and Inhibition of α7-Nicotinic Acetylcholine Receptors

    PubMed Central

    Ashoor, Abrar; Nordman, Jacob C.; Veltri, Daniel; Yang, Keun-Hang Susan; Al Kury, Lina; Shuba, Yaroslav; Mahgoub, Mohamed; Howarth, Frank C.; Sadek, Bassem; Shehu, Amarda; Kabbani, Nadine; Oz, Murat

    2013-01-01

    Menthol is a common compound in pharmaceutical and commercial products and a popular additive to cigarettes. The molecular targets of menthol remain poorly defined. In this study we show an effect of menthol on the α7 subunit of the nicotinic acetylcholine (nACh) receptor function. Using a two-electrode voltage-clamp technique, menthol was found to reversibly inhibit α7-nACh receptors heterologously expressed in Xenopus oocytes. Inhibition by menthol was not dependent on the membrane potential and did not involve endogenous Ca2+-dependent Cl− channels, since menthol inhibition remained unchanged by intracellular injection of the Ca2+ chelator BAPTA and perfusion with Ca2+-free bathing solution containing Ba2+. Furthermore, increasing ACh concentrations did not reverse menthol inhibition and the specific binding of [125I] α-bungarotoxin was not attenuated by menthol. Studies of α7- nACh receptors endogenously expressed in neural cells demonstrate that menthol attenuates α7 mediated Ca2+ transients in the cell body and neurite. In conclusion, our results suggest that menthol inhibits α7-nACh receptors in a noncompetitive manner. PMID:23935840

  13. The selective dopamine D3 receptor antagonist SB-277011A reduces nicotine-enhanced brain reward and nicotine-paired environmental cue functions.

    PubMed

    Pak, Arlene C; Ashby, Charles R; Heidbreder, Christian A; Pilla, Maria; Gilbert, Jeremy; Xi, Zheng-Xiong; Gardner, Eliot L

    2006-10-01

    Increasing evidence suggests that enhanced dopamine (DA) neurotransmission in the nucleus accumbens (NAc) may play a role in mediating the reward and reinforcement produced by addictive drugs and in the attentional processing of drug-associated environmental cues. The meso-accumbens DA system is selectively enriched with DA D3 receptors, a DA receptor subtype increasingly implicated in reward-related brain and behavioural processes. From a variety of evidence, it has been suggested that selective DA D3 receptor antagonism may be a useful pharmacotherapeutic approach for treating addiction. The present experiments tested the efficacy of SB-277011A, a selective DA D3 receptor antagonist, in rat models of nicotine-enhanced electrical brain-stimulation reward (BSR), nicotine-induced conditioned locomotor activity (LMA), and nicotine-induced conditioned place preference (CPP). Nicotine was given subcutaneously within the dose range of 0.25-0.6 mg/kg (nicotine-free base). SB-277011A, given intraperitoneally within the dose range of 1-12 mg/kg, dose-dependently reduced nicotine-enhanced BSR, nicotine-induced conditioned LMA, and nicotine-induced CPP. The results suggest that selective D3 receptor antagonism constitutes a new and promising pharmacotherapeutic approach to the treatment of nicotine dependence. PMID:16942635

  14. Role of α4- and α6-containing nicotinic receptors in the acquisition and maintenance of nicotine self-administration.

    PubMed

    Madsen, Heather B; Koghar, Harcharan S; Pooters, Tine; Massalas, Jim S; Drago, John; Lawrence, Andrew J

    2015-05-01

    Tobacco smoking is a major cause of death and disease and as such there is a critical need for the development of new therapeutic approaches to treat nicotine addiction. Here, we utilize genetic and pharmacological tools to further investigate the nicotinic acetylcholine receptor (nAChR) subtypes that support intravenous self-administration of nicotine. α4-S248F mice contain a point mutation within the α4 nAChR subunit which confers increased sensitivity to nicotine and resistance to mecamylamine. Here, we show that acute administration of mecamylamine (2 mg/kg, i.p.) reduces established nicotine self-administration (0.05 mg/kg/infusion) in wild-type (WT), but not in α4-S248F heterozygous mice, demonstrating a role for α4* nAChRs in the modulation of ongoing nicotine self-administration. Administration of N,N-decane-1,10-diyl-bis-3-picolinium diiodide (bPiDI), a selective α6β2* nAChR antagonist, dose dependently (5 and 10 mg/kg, i.p.) impairs the acquisition of nicotine self-administration and reduces established nicotine self-administration in WT mice when administered acutely (10 mg/kg, i.p.). This was not due to a general reduction in locomotor activity and the same dose of bPiDI did not affect operant responding for sucrose. bPiDI treatment (10 mg/kg, i.p.) also impaired both the acquisition and maintenance of nicotine self-administration in α4-S248F heterozygous mice. This provides further evidence for the involvement of α6β2* nAChRs in the reinforcing effects of nicotine that underlies its ability to support ongoing self-administration. Taken together, selective targeting of α6β2* or α4α6β2* nAChRs may prove to be an effective strategy for the development of smoking cessation therapies. PMID:24750355

  15. Agonist self-inhibition at the nicotinic acetylcholine receptor a nonspecific action

    SciTech Connect

    Forman, S.A.; Firestone, L.L.; Miller, K.W.

    1987-05-19

    Agonist concentration-response relationships at nicotinic postsynaptic receptors were established by measuring /sup 86/Rb/sup +/ efflux from acetylcholine receptor rich native Torpedo membrane vesicles under three different conditions: (1) integrated net ion efflux (in 10 s) from untreated vesicles, (2) integrated net efflux from vesicles in which most acetylcholine sites were irreversibly blocked with ..cap alpha..-bungarotoxin, and (3) initial rates of efflux (5-100 ms) from vesicles that were partially blocked with ..cap alpha..-bungarotoxin. Exposure to acetylcholine, carbamylcholine, suberyldicholine, phenyltrimethylammonium, or (-)-nicotine over 10/sup 8/-fold concentration ranges results in bell-shaped ion flux response curves due to stimulation of acetylcholine receptor channel opening at low concentrations and inhibition of channel function at 60-2000 times higher concentrations. Concentrations of agonists that inhibit their own maximum /sup 86/Rb/sup +/ efflux by 50% (K/sub B/ values) are 110, 211, 3.0, 39, and 8.9 mM, respectively, for the agonists listed above. For acetylcholine and carbamylcholine, K/sub B/ values determined from both 10-s and 15-ms efflux measurements are the same, indicating that the rate of agonist-induced desensitization increases to maximum at concentrations lower than those causing self-inhibition. For all partial and full agonists studied, Hill coefficients for self-inhibition are close to 1.0. Concentrations of agonists up to 8 times K/sub B/ did not change the order parameter reported by a spin-labeled fatty acid incorporated in Torpedo membranes. The authors conclude that agonist self-inhibition cannot be attributed to a general nonspecific membrane perturbation. Instead, these results are consistent with a saturable site of action either at the lipid-protein interface or on the acetylcholine receptor protein itself.

  16. Coantagonism of Glutamate Receptors and Nicotinic Acetylcholinergic Receptors Disrupts Fear Conditioning and Latent Inhibition of Fear Conditioning

    ERIC Educational Resources Information Center

    Gould, Thomas J.; Lewis, Michael C.

    2005-01-01

    The present study investigated the hypothesis that both nicotinic acetylcholinergic receptors (nAChRs) and glutamate receptors ([alpha]-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors (AMPARs) and N-methyl-D-aspartate glutamate receptors (NMDARs)) are involved in fear conditioning, and may modulate similar processes. The effects of the…

  17. Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations

    SciTech Connect

    Xu, Yuan Cardell, Lars-Olaf

    2014-02-15

    Introduction: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. Methods: Segments were cultured for 24 h in the presence of vehicle, nicotine (10 μM) and/or dexamethasone (1 μM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 μM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. Results: The organ culture procedure markedly increased bradykinin- (selective B{sub 2} receptor agonist) and des-Arg{sup 9}-bradykinin- (selective B{sub 1} receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE{sub 2}. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg{sup 9}-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B{sub 2} receptors, but not those on B{sub 1}. Dexamethasone completely abolished kinin-induced relaxations. Conclusion: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in

  18. The neuronal nicotinic acetylcholine receptor {alpha}7 subunit gene: Cloning, mapping, structure, and targeting in mouse

    SciTech Connect

    Orr-Urtreger, A.; Baldini, A.; Beaudet, A.L.

    1994-09-01

    The neuronal nicotinic acetylcholine receptor {alpha}7 subunit is a member of a family of ligand-gated ion channels, and is the only subunit know to bind {alpha}-bungarotoxin in mammalian brain. {alpha}-Bungarotoxin binding sites are known to be more abundant in the hippocampus of mouse strains that are particularly sensitive to nicotine-induced seizures. The {alpha}7 receptor is highly permeable to calcium, which could suggest a role in synaptic plasticity in the nervous system. Auditory gating deficiency, an abnormal response to a second auditory stimulus, is characteristic of schizophrenia. Mouse strains that exhibit a similar gating deficit have reduced hippocampal expression of the {alpha}7 subunit. We have cloned and sequenced the full length cDNA for the mouse {alpha}7 gene (Acra-7) and characterized its gene structure. The murine {alpha}7 shares amino acid identity of 99% and 93% with the rat and human {alpha}7 subunits, respectively. Using an interspecies backcross panel, the murine gene was mapped to chromosome 7 near the p locus, a region syntenic with human chromosome 15; the human gene (CHRNA7) was confirmed to map to 15q13-q14 by FISH. To generate a mouse {alpha}7 mutant by homologous recombination, we have constructed a replacement vector which will delete transmembrane domains II-IV and the cytoplasmic domain from the gene product. Recombinant embryonic stem (ES) cell clones were selected and used to develop mouse chimeras that are currently being bred to obtain germline transmission.

  19. Positive allosteric modulation of α4β2 nicotinic acetylcholine receptors as a new approach to smoking reduction: evidence from a rat model of nicotine self-administration

    PubMed Central

    Liu, Xiu

    2013-01-01

    Rationale The α4β2 subtype of nicotinic acetylcholine receptors (nAChRs) plays a central role in the mediation of nicotine reinforcement. Positive allosteric modulators (PAMs) at α4β2 nAChRs facilitate the intrinsic efficiency of these receptors although they do not directly activate the receptors. α4β2 PAMs are hypothesized to reduce nicotine self-administration in subjects engaged in routine nicotine consumption. The present study tested this hypothesis using a rat model of nicotine self-administration. Methods Male Sprague-Dawley rats were trained in daily 1 h sessions to intravenously self-administer nicotine (0.03 mg/kg/infusion, free base) on a fixed-ratio 5 schedule. Effects of the α4β2 PAM desformylflustrabromine (dFBr), α4β2 agonist 5-iodo-A-85380, and acetylcholinesterase inhibitor galantamine on nicotine intake were examined. The ability of dFBr and 5-iodo-A-85380 to substitute for nicotine was also assessed. Results dFBr and 5-iodo-A-85380 dose-dependently reduced nicotine self-administration without changing lever responses for food. Galantamine decreased self-administration of nicotine and food at high doses. Unlike 5-iodo-A-85380, dFBr failed to substitute for nicotine in supporting self-administration behavior. Conclusions These results demonstrated the effectiveness of dFBr in reducing nicotine intake and the inability of dFBr to support self-administration behavior. These findings suggest that positive allosteric modulation of α4β2 nAChRs may be a promising target for the treatment of nicotine addiction. Moreover, α4β2 PAMs, in contrast to agonist medications, may have clinical advantages because they may have little liability for abuse because of their lack of reinforcing actions on their own. PMID:23712602

  20. Minimum number of lipids are required to support the functional properties of the nicotinic acetylcholine receptor

    SciTech Connect

    Jones, O.T.; Eubanks, J.H.; Earnest, J.P.; McNamee, M.G.

    1988-05-17

    The detergent sodium cholate was used to both solubilize and partially delipidate the nicotinic acetylcholine receptor from Torpedo californica. Using both native membranes and reconstituted membranes, it is shown that the detergent to lipid molar ratio is the most important parameter in determining the effect of the detergent on the functional properties of the receptor. Receptor-lipid complexes were quantitatively separated from detergent and excess lipids by centrifugation through detergent-free sucrose gradients. The lipid to protein molar ratio of the complexes could be precisely controlled by adjusting the cholate and lipid concentrations of the starting membranes. Analyses of both ion influx activity and ligand binding revealed that a minimum of 45 lipids per receptor was required for stabilization of the receptor in a fully functional state. Progressive irreversible inactivation occurred as the lipid to protein mole ratio was decreased below 45, and complete inactivation occurred below a ratio of 20. The results are consistent with a functional requirement for a single shell of lipids around the perimeter of the receptor.

  1. ( sup 3 H)cytisine binding to nicotinic cholinergic receptors in brain

    SciTech Connect

    Pabreza, L.A.; Dhawan, S.; Kellar, K.J. )

    1991-01-01

    Cytisine, a ganglionic agonist, competes with high affinity for brain nicotinic cholinergic receptors labeled by any of several nicotinic {sup 3}H-agonist ligands. Here we have examined the binding of ({sup 3}H)cytisine in rat brain homogenates. ({sup 3}H)Cytisine binds with high affinity (Kd less than 1 nM), and specific binding represented 60-90% of total binding at all concentrations examined up to 15 nM. The nicotinic cholinergic agonists nicotine, acetylcholine, and carbachol compete with high affinity for ({sup 3}H)cytisine binding sites, whereas among nicotinic receptor antagonists only dihydro-beta-erythroidine competes with high affinity (in the nanomolar range). Comparison of binding in several brain regions showed that ({sup 3}H)cytisine binding is higher in the thalamus, striatum, and cortex than in the hippocampus, cerebellum, or hypothalamus. The pharmacology and brain regional distribution of ({sup 3}H)cytisine binding sites are those predicted for neuronal nicotinic receptor agonist recognition sites. The high affinity and low nonspecific binding of ({sup 3}H)cytisine should make it a very useful ligand for studying neuronal nicotinic receptors.

  2. Evidence for thymopoietin and thymopoietin/. alpha. -bungarotoxin/nicotinic receptors within the brain

    SciTech Connect

    Quik, M. ); Babu, U.; Audhya, T.; Goldstein, G. )

    1991-03-15

    Thymopoietin, a polypeptide hormone of the thymus that has pleiotropic actions on the immune, endocrine, and nervous systems, potently interacts with the neuromuscular nicotinic acetylcholine receptor. Thymopoietin binds to the nicotinic {alpha}-bungarotoxin ({alpha}-BGT) receptor in muscle and, like {alpha}BGT, inhibits cholinergic transmission at this site. Evidence is given that radiolabeled thymopoietin similarly binds to a nicotinic {alpha}-BGT-binding site within the brain and does so with the characteristics of a specific receptor ligand. Thus specific binding to neuronal membranes was saturable, of high affinity linear with increased tissue concentration, and readily reversible; half-time was {approximately}5 min for association and 10 min for dissociation. Binding of {sup 125}I-labeled thymopoietin was displaced not only by unlabeled thymopoietin but also by {alpha}-BGT and the nicotinic receptor ligands d-tubocurarine and nicotine; various other receptor ligands (muscarinic, adrenergic, and dopaminergic) did not affect binding of {sup 125}I-labeled thymopoietin. Thymopoietin was shown by ELISA to be present in brain extracts, displacement curves of thymus and brain extracts being parallel to the standard thymopoietin curve, and Western (immuno) blot identified in brain and thymus extracts a thymopoietin-immunoreactive polypeptide of the same molecular mass as purified thymopoietin polypeptide. The authors conclude that thymopoietin and thymopoietin-binding sites are present within the brain and that the receptor for thymopoietin is the previously identified nicotinic {alpha}-BGT-binding site of neuronal tissue.

  3. α6β2*-subtype nicotinic acetylcholine receptors are more sensitive than α4β2*-subtype receptors to regulation by chronic nicotine administration

    PubMed Central

    Marks, MJ; Grady, SR; Salminen, O; Paley, MA; Wageman, CR; McIntosh, JM; Whiteaker, P

    2014-01-01

    Nicotinic acetylcholine receptors (nAChR) of the α6β2* subtype (where * indicates the possible presence of additional subunits) are prominently expressed on dopaminergic neurons. Because of this, their role in tobacco use and nicotine dependence has received much attention. Previous studies have demonstrated that α6β2*-nAChR are downregulated following chronic nicotine exposure (unlike other subtypes that have been investigated – most prominently α4β2* nAChR). This study examines, for the first time, effects across a comprehensive chronic nicotine dose range. Chronic nicotine dose-responses and quantitative ligand-binding autoradiography were used to define nicotine sensitivity of changes in α4β2*-nAChR and α6β2*-nAChR expression. α6β2*-nAChR downregulation by chronic nicotine exposure in dopaminergic and optic-tract nuclei was ≈three-fold more sensitive than upregulation of α4β2*-nAChR. In contrast, nAChR-mediated [3H]-dopamine release from dopamine-terminal region synaptosomal preparations changed only in response to chronic treatment with high nicotine doses, while dopaminergic parameters (transporter expression and activity, dopamine receptor expression) were largely unchanged. Functional measures in olfactory tubercle preparations were made for the first time; both nAChR expression levels and nAChR-mediated functional measures changed differently between striatum and olfactory tubercles. These results show that functional changes measured using synaptosomal [3H]-DA release are primarily due to changes in nAChR, rather than in dopaminergic, function. PMID:24661093

  4. Structure, oligosaccharide structures, and posttranslationally modified sites of the nicotinic acetylcholine receptor.

    PubMed Central

    Poulter, L; Earnest, J P; Stroud, R M; Burlingame, A L

    1989-01-01

    Using mass spectrometry, we have examined the transmembrane topography of the nicotinic acetylcholine receptor, a five-subunit glycosylated protein complex that forms a gated ion channel in the neuromuscular junction. The primary sequences of the four polypeptide chains making up the acetylcholine receptor from Torpedo californica contain many possible sites for glycosylation or phosphorylation. We have used liquid secondary ion mass spectrometry to identify posttranslationally modified residues and to determine the intact oligosaccharide structures of the carbohydrate present on the acetylcholine receptor. Asparagine-143 of the alpha subunit (in consensus numbering) is shown to be glycosylated with high-mannose oligosaccharide. Asparagine-453 of the gamma subunit is not glycosylated, a fact that bears on the question of the orientations of putative transmembranous helices M3, MA, and M4. The structures of the six major acetylcholine receptor oligosaccharides are determined: the major components (70%) are of the high-mannose type, with bi-, tri-, and tetraantennary complex oligosaccharides making up approximately equal to 22 mol% of the total carbohydrate. This application of a multichannel array detector mass spectrometer provided a breakthrough in sensitivity that allowed us to identify the site of attachment of, and the sequence of, oligosaccharides on a 300-kDa membrane protein from only 5 pmol of the isolated oligosaccharide. Images PMID:2771948

  5. Positive allosteric modulators as an approach to nicotinic acetylcholine receptor- targeted therapeutics: advantages and limitations

    PubMed Central

    Williams, Dustin K.; Wang, Jingyi; Papke, Roger L.

    2011-01-01

    Neuronal nicotinic acetylcholine receptors (nAChR), recognized targets for drug development in cognitive and neuro-degenerative disorders, are allosteric proteins with dynamic interconversions between multiple functional states. Activation of the nAChR ion channel is primarily controlled by the binding of ligands (agonists, partial agonists, competitive antagonists) at conventional agonist binding sites, but is also regulated in either negative or positive ways by the binding of ligands to other modulatory sites. In this review, we discuss models for the activation and desensitization of nAChR, and the discovery of multiple types of ligands that influence those processes in both heteromeric nAChR, such as the high affinity nicotine receptors of the brain, and homomeric α7-type receptors. In recent years, α7 nAChRs have been identified as a potential target for therapeutic indications leading to the development of α7-selective agonists and partial agonists. However, unique properties of α7 nAChR, including low probability of channel opening and rapid desensitization, may limit the therapeutic usefulness of ligands binding exclusively to conventional agonist binding sites. New enthusiasm for the therapeutic targeting of α7 has come from the identification of α7-selective positive allosteric modulators (PAMs) that work effectively on the intrinsic factors that limit α7 ion channel activation. While these new drugs appear promising for therapeutic development, we also consider potential caveats and possible limitations for their use, including PAM-insensitive forms of desensitization and cytotoxicity issues. PMID:21575610

  6. Alpha9 nicotinic acetylcholine receptors and the treatment of pain.

    PubMed

    McIntosh, J Michael; Absalom, Nathan; Chebib, Mary; Elgoyhen, Ana Belén; Vincler, Michelle

    2009-10-01

    Chronic pain is a vexing worldwide problem that causes substantial disability and consumes significant medical resources. Although there are numerous analgesic medications, these work through a small set of molecular mechanisms. Even when these medications are used in combination, substantial amounts of pain often remain. It is therefore highly desirable to develop treatments that work through distinct mechanisms of action. While agonists of nicotinic acetylcholine receptors (nAChRs) have been intensively studied, new data suggest a role for selective antagonists of nAChRs. alpha-Conotoxins are small peptides used offensively by carnivorous marine snails known as Conus. A subset of these peptides known as alpha-conotoxins RgIA and Vc1.1 produces both acute and long lasting analgesia. In addition, these peptides appear to accelerate the recovery of function after nerve injury, possibly through immune mediated mechanisms. Pharmacological analysis indicates that RgIA and Vc1.1 are selective antagonists of alpha9alpha10 nAChRs. A recent study also reported that these alpha9alpha10 antagonists are also potent GABA-B agonists. In the current study, we were unable to detect RgIA or Vc1.1 binding to or action on cloned GABA-B receptors expressed in HEK cells or Xenopus oocytes. We review the background, findings and implications of use of compounds that act on alpha9* nAChRs.(1). PMID:19477168

  7. Therapeutic Potential of α7 Nicotinic Acetylcholine Receptors.

    PubMed

    Bertrand, Daniel; Lee, Chih-Hung L; Flood, Dorothy; Marger, Fabrice; Donnelly-Roberts, Diana

    2015-10-01

    Progress in the fields of neuroscience and molecular biology has identified the forebrain cholinergic system as being important in many higher order brain functions. Further analysis of the genes encoding the nicotinic acetylcholine receptors (nAChRs) has highlighted, in particular, the role of α7 nAChRs in these higher order brain functions as evidenced by their peculiar physiologic and pharmacological properties. As this receptor has gained the attention of scientists from academia and industry, our knowledge of its roles in various brain and bodily functions has increased immensely. We have also seen the development of small molecules that have further refined our understanding of the roles of α7 nAChRs, and these molecules have begun to be tested in clinical trials for several indications. Although a large body of data has confirmed a role of α7 nAChRs in cognition, the translation of small molecules affecting α7 nAChRs into therapeutics has to date only progressed to the stage of testing in clinical trials. Notably, however, most recent human genetic and biochemical studies are further underscoring the crucial role of α7 nAChRs and associated genes in multiple organ systems and disease states. The aim of this review is to discuss our current knowledge of α7 nAChRs and their relevance as a target in specific functional systems and disease states. PMID:26419447

  8. Increased expression of the nicotinic acetylcholine receptor in stimulated muscle.

    PubMed

    O'Reilly, Clare; Pette, Dirk; Ohlendieck, Kay

    2003-01-10

    Chronic low-frequency stimulation has been used as a model for investigating responses of skeletal muscle fibres to enhanced neuromuscular activity under conditions of maximum activation. Fast-to-slow isoform shifting of markers of the sarcoplasmic reticulum and the contractile apparatus demonstrated successful fibre transitions prior to studying the effect of chronic electro-stimulation on the expression of the nicotinic acetylcholine receptor. Comparative immunoblotting revealed that the alpha- and delta-subunits of the receptor were increased in 10-78 day stimulated specimens, while an associated component of the surface utrophin-glycoprotein complex, beta-dystroglycan, was not drastically changed in stimulated fast skeletal muscle. Previous studies have shown that electro-stimulation induces degeneration of fast glycolytic fibres, trans-differentiation leading to fast-to-slow fibre transitions and activation of muscle precursor cells. In analogy, our results indicate a molecular modification of the central functional unit of the post-synaptic muscle surface within existing neuromuscular junctions and/or during remodelling of nerve-muscle contacts. PMID:12504123

  9. Prior nicotine self-administration attenuates subsequent dopaminergic deficits of methamphetamine in rats: role of nicotinic acetylcholine receptors.

    PubMed

    Baladi, Michelle G; Nielsen, Shannon M; McIntosh, J Michael; Hanson, Glen R; Fleckenstein, Annette E

    2016-08-01

    Preclinical studies have demonstrated that oral nicotine exposure attenuates long-term dopaminergic damage induced by toxins, including repeated, high doses of methamphetamine. It is suggested that alterations in nicotinic acetylcholine receptor (nAChR) expression, including α4β2* and α6β2* subtypes, likely contribute to this protection. The current study extended these findings by investigating whether nicotine self-administration in male, Sprague-Dawley rats (a) attenuates short-term dopaminergic damage induced by methamphetamine and (b) causes alterations in levels of α4β2* and α6β2* nAChR subtypes. The findings indicate that nicotine self-administration (0.032 mg/kg/infusion for 14 days) per se did not alter α4β2* and α6β2* nAChR expression or dopamine transporter (DAT) expression and function. Interestingly, prior nicotine self-administration attenuated methamphetamine-induced decreases in DAT function when assessed 24 h, but not 1 h, after methamphetamine treatment (4×7.5 mg/kg/injection). The ability of nicotine to attenuate the effects of methamphetamine on DAT function corresponded with increases in α4β2*, but not α6β2*, nAChR binding density. Understanding the role of nAChRs in methamphetamine-induced damage has the potential to elucidate mechanisms underlying the etiology of disorders involving dopaminergic dysfunction, as well as to highlight potential new therapeutic strategies for prevention or reduction of dopaminergic neurodegeneration. PMID:26871405

  10. Nicotine and sympathetic neurotransmission.

    PubMed

    Haass, M; Kübler, W

    1997-01-01

    Nicotine increases heart rate, myocardial contractility, and blood pressure. These nicotine-induced cardiovascular effects are mainly due to stimulation of sympathetic neurotransmission, as nicotine stimulates catecholamine release by an activation of nicotine acetylcholine receptors localized on peripheral postganglionic sympathetic nerve endings and the adrenal medulla. The nicotinic acetylcholine receptor is a ligand-gated cation channel with a pentameric structure and a central pore with a cation gate, which is essential for ion selectivity and permeability. Binding of nicotine to its extracellular binding site leads to a conformational change of the central pore, which results in the influx of sodium and calcium ions. The resulting depolarization of the sympathetic nerve ending stimulates calcium influx through voltage-dependent N-type calcium channels, which triggers the nicotine-evoked exocytotic catecholamine release. In the isolated perfused guinea-pig heart, cardiac energy depletion sensitizes cardiac sympathetic nerves to the norepinephrine-releasing effect of nicotine, as indicated by a leftward shift of the concentration-response curve, a potentiation of maximum transmitter release, and a delay of the tachyphylaxis of nicotine-evoked catecholamine release. This sensitization was also shown to occur in the human heart under in vitro conditions. Through the intracardiac release of norepinephrine, nicotine induces a beta-adrenoceptor-mediated increase in heart rate and contractility, and an alpha-adrenoceptor-mediated increase in coronary vasomotor tone. The resulting simultaneous increase in oxygen demand and coronary resistance has a detrimental effect on the oxygen balance of the heart, especially in patients with coronary artery disease. Sensitization of the ischemic heart to the norepinephrine-releasing effect of nicotine may be a trigger for acute cardiovascular events in humans, such as acute myocardial infarction and/or life

  11. Targeting the interaction between fatty acid ethanolamides and nicotinic receptors: therapeutic perspectives.

    PubMed

    Melis, Miriam; Pistis, Marco

    2014-08-01

    Nicotine is one of the drugs of abuse that frequently causes addiction and relapse during abstinence. Nicotine's strong addicting properties reside in its ability to enhance dopamine transmission, and to induce specific changes in synaptic plasticity. Currently, approved therapies for smoking cessation increase the chances of remaining abstinent, but lack high levels of efficacy and are associated with significant adverse side effects. As a result, there is an urgent need for more effective antismoking medications. Studies have revealed that drugs targeting the peroxisome proliferator-activated-receptor-α (PPARα) show promise for the treatment of nicotine addiction. These drugs include synthetic PPARα ligands, such as the clinically available hypolipidemic fibrates, and drugs that increase levels of endogenous endocannabinoid-like fatty acid ethanolamides (FAEs) that act as PPARα agonists. In this review, we will discuss the specific interaction between PPARα and nicotine, and the molecular mechanisms whereby these intracellular receptors regulate nicotinic acetylcholine receptor functions in neurons. Modulation of neurophysiological, neurochemical and behavioral effects of nicotine by PPARα will be also reviewed. Indeed, a picture is emerging where FAEs are endogenous regulators of acetylcholine transmission. Notably, the implications of this specific cross talk extend beyond nicotine addiction, and might bear relevance for psychiatric disorders and epilepsy. PMID:24704146

  12. Effects of the Sazetidine-a Family of Compounds on the Body Temperature in Wildtype, Nicotinic Receptor B2(-/-) and a7(-/-) Mice

    EPA Science Inventory

    Nicotine elicits hypothermic responses in rodents. This effect appears to be related to nicotinic receptor desensitization because sazetidine-A, an a4B2 nicotinic receptor desensitizing agent, produces marked hypothermia and potentiates nicotine-induced hypothermia in mice. To de...

  13. N-Benzylpiperidine Derivatives as α7 Nicotinic Receptor Antagonists.

    PubMed

    Criado, Manuel; Mulet, José; Sala, Francisco; Sala, Salvador; Colmena, Inés; Gandía, Luis; Bautista-Aguilera, Oscar M; Samadi, Abdelouahid; Chioua, Mourad; Marco-Contelles, José

    2016-08-17

    A series of multitarget directed propargylamines, as well as other differently susbstituted piperidines have been screened as potential modulators of neuronal nicotinic acetylcholine receptors (nAChRs). Most of them showed antagonist actions on α7 nAChRs. Especially, compounds 13, 26, and 38 displayed submicromolar IC50 values on homomeric α7 nAChRs, whereas they were less effective on heteromeric α3β4 and α4β2 nAChRs (up to 20-fold higher IC50 values in the case of 13). Antagonism was concentration dependent and noncompetitive, suggesting that these compounds behave as negative allosteric modulators of nAChRs. Upon the study of a series of less complex derivatives, the N-benzylpiperidine motif, common to these compounds, was found to be the main pharmacophoric group. Thus, 2-(1-benzylpiperidin-4-yl)-ethylamine (48) showed an inhibitory potency comparable to the one of the previous compounds and also a clear preference for α7 nAChRs. In a neuroblastoma cell line, representative compounds 13 and 48 also inhibited, in a concentration-dependent manner, cytosolic Ca(2+) signals mediated by nAChRs. Finally, compounds 38 and 13 inhibited 5-HT3A serotonin receptors whereas they had no effect on α1 glycine receptors. Given the multifactorial nature of many pathologies in which nAChRs are involved, these piperidine antagonists could have a therapeutic potential in cases where cholinergic activity has to be negatively modulated. PMID:27254782

  14. Varenicline, a Partial Agonist at Neuronal Nicotinic Receptors, Reduces Nicotine-Induced Increases in 20% Ethanol Operant Self-Administration in Sprague-Dawley Rats

    PubMed Central

    Bito-Onon, Jade J.; Simms, Jeffrey A.; Chatterjee, Susmita; Holgate, Joan; Bartlett, Selena E.

    2010-01-01

    Alcohol and nicotine use disorders are often treated as separate diseases, despite evidence that approximately 80–90% of alcohol dependent individuals are also heavy smokers. Both nicotine and ethanol have been shown to interact with neuronal nicotinic acetylcholine receptors (nAChRs), suggesting these receptors are a common biological target for the effects of nicotine and ethanol in the brain. There are few studies that have examined the effects of co-administered nicotine and ethanol on the activity of nAChRs in rodents. In the present study, we show that Sprague-Dawley rats, a strain often used for nicotine studies but not as often for voluntary ethanol intake studies, will consume 20% ethanol using both the intermittent-access two-bottle-choice and operant self-administration models without the need for sucrose fading. We show that nicotine (0.2mg/kg and 0.8mg/kg, s.c.) significantly increases operant 20% ethanol self-administration and varenicline (2mg/kg, s.c), a partial agonist at nAChRs, significantly decreases operant ethanol self-administration and nicotine-induced increases in ethanol self-administration. This suggests that nAChRs play an important role in increasing ethanol self-administration and that varenicline may be an efficacious treatment for alcohol and nicotine co-dependencies. PMID:21392178

  15. Functional organization and conformational dynamics of the nicotinic receptor: a plausible structural interpretation of myasthenic mutations.

    PubMed

    Taly, Antoine; Changeux, Jean-Pierre

    2008-01-01

    To understand the structural causes of myasthenic mutations, molecular models of the nicotinic acetycholine receptor were first constructed. Then, the gating transition between resting and open conformation was investigated in silico by normal mode analysis using a physically meaningful description of protein flexibility. This analysis revealed a global quaternary twist motion that opens the ion pore. Second, it was found that most (24/27) of the spontaneous mutations known to cause congenital myasthenia (and autosomal dominant nocturnal frontal lobe epilepsy) are located either at the interface between subunits or, within a given subunit, at the interface between rigid blocks. These interfaces are modified significantly by the twist mode together with significant changes of the tertiary organization of the subunits. These data provide a qualitative interpretation of the molecular phenotype of pathological mutations responsible for congenital myasthenia. PMID:18567852

  16. Molecular environment of the phencyclidine binding site in the nicotinic acetylcholine receptor membrane

    SciTech Connect

    Palma, A.L.; Wang, H.H. )

    1991-06-01

    Phencyclidine is a highly specific noncompetitive inhibitor of the nicotinic acetylcholine receptor. In a novel approach to study this site, a spin-labeled analogue of phencyclidine, 4-phenyl-4-(1-piperidinyl)-2,2,6,6-tetramethylpiperidinoxyl (PPT) was synthesized. The binding of PPT inhibits 86Rb flux (IC50 = 6.6 microM), and (3H)phencyclidine binding to both resting and desensitized acetylcholine receptor (IC50 = 17 microM and 0.22 microM, respectively). From an indirect Hill plot of the inhibition of (3H)phencyclidine binding by PPT, a Hill coefficient of approximately one was obtained in the presence of carbamylcholine and 0.8 in alpha-bungarotoxin-treated preparations. Taken together, these results indicate that PPT mimics phencyclidine in its ability to bind to the noncompetitive inhibitor site and is functionally active in blocking ion flux across the acetylcholine receptor channel. Analysis of the electron spin resonance signal of the bound PPT suggests that the environment surrounding the probe within the ion channel is hydrophobic, with a hydrophobicity parameter of 1.09. A dielectric constant for the binding site was estimated to be in the range of 2-3 units.

  17. Alpha-5 and -3 nicotinic receptor gene variants predict nicotine dependence but not cessation: Findings from the COMMIT cohort

    PubMed Central

    Bousman, Chad A.; Rivard, Cheryl; Den Haese, Jason; Ambrosone, Christine; Hyland, Andrew

    2012-01-01

    Smoking many cigarettes per day (CPD) and short interval to first cigarette (TTF) after waking are two of the most heritable smoking phenotypes and comprise the Heavy Smoking Index (HSI). These phenotypes are often used as proxies for nicotine dependence (ND) and are associated with smoking cessation outcomes. Case-control and genome-wide association studies have reported links between single nucleotide polymorphisms (SNPs) in the alpha-5 and -3 nicotinic receptor subunit (CHRNA5 and CHRNA3) genes and CPD but few have examined TTF or cessation outcomes. In this study we longitudinally assessed 1301 European-American smokers at four time-points from 1988 to 2005. One CHRNA5 (rs16969968) and two CHRNA3 (rs1051703, rs6495308) SNPs were examined for their ability to predict smokers who ‘ever’ reported ND based on three phenotypic classifications: 1) 25+ CPD, 2) TTF < 10 minutes, and 3) HSI ≥ 4. In a subsample of 1157 quit attempters, we also examined each SNP’s ability to predict ‘ever’ quitting for a period of >6 months. Demographically adjusted logistic regressions showed significant allelic and genotypic associations between all three SNPs and CPD but not TTF, HSI, or smoking cessation. Carriers of both the rs16969968-AA and rs6495308-TT genotypes had approximately two-fold greater odds for ND defined using CPD or TTF. Results suggest nicotinic receptor variants are associated with greater odds of ND according to CPD and to a lesser extent TTF. Research examining the effect of nicotinic receptor genetic variation on ND phenotypes beyond CPD is warranted. PMID:22223462

  18. α-Conotoxin dendrimers have enhanced potency and selectivity for homomeric nicotinic acetylcholine receptors.

    PubMed

    Wan, Jingjing; Huang, Johnny X; Vetter, Irina; Mobli, Mehdi; Lawson, Joshua; Tae, Han-Shen; Abraham, Nikita; Paul, Blessy; Cooper, Matthew A; Adams, David J; Lewis, Richard J; Alewood, Paul F

    2015-03-11

    Covalently attached peptide dendrimers can enhance binding affinity and functional activity. Homogenous di- and tetravalent dendrimers incorporating the α7-nicotinic receptor blocker α-conotoxin ImI (α-ImI) with polyethylene glycol spacers were designed and synthesized via a copper-catalyzed azide-alkyne cycloaddition of azide-modified α-ImI to an alkyne-modified polylysine dendron. NMR and CD structural analysis confirmed that each α-ImI moiety in the dendrimers had the same 3D structure as native α-ImI. The binding of the α-ImI dendrimers to binding protein Ac-AChBP was measured by surface plasmon resonance and revealed enhanced affinity. Quantitative electrophysiology showed that α-ImI dendrimers had ∼100-fold enhanced potency at hα7 nAChRs (IC50 = 4 nM) compared to native α-ImI (IC50 = 440 nM). In contrast, no significant potency enhancement was observed at heteromeric hα3β2 and hα9α10 nAChRs. These findings indicate that multimeric ligands can significantly enhance conotoxin potency and selectivity at homomeric nicotinic ion channels. PMID:25710197

  19. Nicotinic cholinergic receptors in rat brain. Annual report No. 2

    SciTech Connect

    Kellar, K.J.

    1985-05-13

    We have conducted experiments to determine if 3H acetylcholine (3Hach) nicotinic recognition sites are located presynaptically on catecholamine and/or serotonin axons. Lesions of these axons by intraventricular injections of neurotoxins resulted in marked decreases in 3Hach binding sites in the striatum and hypothalamus, but not in the cortex or thalamus. These results indicate that 3Hach nicotinic binding sites are located on catecholamine and serotonin axons in specific areas of the brain. In other experiments, we determined that repeated administration of nicotine results in enhanced behavioral responses to a subsequent injection of nicotine, and that there appears to be a correlation between the enhanced response to nicotine and increased 3Hach binding sites in cerebral cortex.

  20. Transmembrane topography of the nicotinic acetylcholine receptor delta subunit.

    PubMed Central

    McCrea, P D; Popot, J L; Engelman, D M

    1987-01-01

    Current folding models for the nicotinic acetylcholine receptor (AChR) predict either four or five transmembrane segments per subunit. The N-terminus of each subunit is almost certainly extracellular. We have tested folding models by determining biochemically the cellular location of an intermolecular disulfide bridge thought to lie at the delta subunit C-terminus. Dimers of AChR linked through the delta-delta bridge were prepared from Torpedo marmorata and T.californica electric organ. The disulfide's accessibility to hydrophilic reductants was tested in a reconstituted vesicle system. In right-side-out vesicles (greater than 95% ACh binding sites outwards), the bridge was equally accessible whether or not vesicles had been disrupted by freeze--thawing or by detergents. Control experiments based on the rate of reduction of entrapped diphtheria toxin and measurements of radioactive reductant efflux demonstrated that the vesicles provide an adequate permeability barrier. In reconstituted vesicles containing AChR dimers in scrambled orientations, right-side-out dimers were reduced to monomers three times more rapidly than inside-out dimers, consistent with the measured rate of reductant permeation. These observations indicate that in reconstituted vesicles the delta-delta disulfide bridge lies in the same aqueous space as the ACh binding sites. They are most easily reconciled with folding models that propose an even number of transmembrane crossing per subunit. PMID:3428268

  1. Early Life Stress, Nicotinic Acetylcholine Receptors and Alcohol Use Disorders

    PubMed Central

    Holgate, Joan Y.; Bartlett, Selena E.

    2015-01-01

    Stress is a major driving force in alcohol use disorders (AUDs). It influences how much one consumes, craving intensity and whether an abstinent individual will return to harmful alcohol consumption. We are most vulnerable to the effects of stress during early development, and exposure to multiple traumatic early life events dramatically increases the risk for AUDs. However, not everyone exposed to early life stress will develop an AUD. The mechanisms determining whether an individual’s brain adapts and becomes resilient to the effects of stress or succumbs and is unable to cope with stress remain elusive. Emerging evidence suggests that neuroplastic changes in the nucleus accumbens (NAc) following early life stress underlie the development of AUDs. This review discusses the impact of early life stress on NAc structure and function, how these changes affect cholinergic signaling within the mesolimbic reward pathway and the role nicotinic acetylcholine receptors (nAChRs) play in this process. Understanding the neural pathways and mechanism determining stress resilience or susceptibility will improve our ability to identify individuals susceptible to developing AUDs, formulate cognitive interventions to prevent AUDs in susceptible individuals and to elucidate and enhance potential therapeutic targets, such as the nAChRs, for those struggling to overcome an AUD. PMID:26136145

  2. Cycloxaprid insecticide: nicotinic acetylcholine receptor binding site and metabolism.

    PubMed

    Shao, Xusheng; Swenson, Tami L; Casida, John E

    2013-08-21

    Cycloxaprid (CYC) is a novel neonicotinoid prepared from the (nitromethylene)imidazole (NMI) analogue of imidacloprid. In this study we consider whether CYC is active per se or only as a proinsecticide for NMI. The IC50 values (nM) for displacing [(3)H]NMI binding are 43-49 for CYC and 2.3-3.2 for NMI in house fly and honeybee head membranes and 302 and 7.2, respectively, in mouse brain membranes, potency relationships interpreted as partial conversion of some CYC to NMI under the assay conditions. The 6-8-fold difference in toxicity of injected CYC and NMI to house flies is consistent with their relative potencies as in vivo nicotinic acetylcholine receptor (nAChR) inhibitors in brain measured with [(3)H]NMI binding assays. CYC metabolism in mice largely involves cytochrome P450 pathways without NMI as a major intermediate. Metabolites of CYC tentatively assigned are five monohydroxy derivatives and one each of dihydroxy, nitroso, and amino modifications. CYC appears be a proinsecticide, serving as a slow-release reservoir for NMI with selective activity for insect versus mammalian nAChRs. PMID:23889077

  3. Transmembrane topography of the nicotinic acetylcholine receptor delta subunit.

    PubMed

    McCrea, P D; Popot, J L; Engelman, D M

    1987-12-01

    Current folding models for the nicotinic acetylcholine receptor (AChR) predict either four or five transmembrane segments per subunit. The N-terminus of each subunit is almost certainly extracellular. We have tested folding models by determining biochemically the cellular location of an intermolecular disulfide bridge thought to lie at the delta subunit C-terminus. Dimers of AChR linked through the delta-delta bridge were prepared from Torpedo marmorata and T.californica electric organ. The disulfide's accessibility to hydrophilic reductants was tested in a reconstituted vesicle system. In right-side-out vesicles (greater than 95% ACh binding sites outwards), the bridge was equally accessible whether or not vesicles had been disrupted by freeze--thawing or by detergents. Control experiments based on the rate of reduction of entrapped diphtheria toxin and measurements of radioactive reductant efflux demonstrated that the vesicles provide an adequate permeability barrier. In reconstituted vesicles containing AChR dimers in scrambled orientations, right-side-out dimers were reduced to monomers three times more rapidly than inside-out dimers, consistent with the measured rate of reductant permeation. These observations indicate that in reconstituted vesicles the delta-delta disulfide bridge lies in the same aqueous space as the ACh binding sites. They are most easily reconciled with folding models that propose an even number of transmembrane crossing per subunit. PMID:3428268

  4. The twin drug approach for novel nicotinic acetylcholine receptor ligands.

    PubMed

    Tomassoli, Isabelle; Gündisch, Daniela

    2015-08-01

    The association of two pharmacophoric entities generates so-called 'twin drugs' or dimer derivatives. We applied this approach for the design of a small compound library for the interaction with α4β2(∗) nicotinic acetylcholine receptors (nAChRs). In this compound series, the nAChR ligand N,N-dimethyl-2-(pyridin-3-yloxy)ethan-1-amine 9 served as one pharmacological entity and it was initially kept constant as one part of the 'twin' compound. 'Twin' compounds with identical or non-identical entities using the 'no linker mode' or 'overlap' mode were synthesized and evaluated for their nAChR affinities. Compound 17a showed the highest affinity for the α4β2(∗) nAChR subtype (Ki=0.188 nM) and its (di)fluoro analogs could retain nanomolar affinities, when replacing pyridine as the hydrogen bond acceptor system by mono- or difluoro-phenyls. The 'twin drug' approach proved to provide compounds with high affinity and subtype selectivity for α4β2(∗) nAChRs. PMID:26142318

  5. Neuronal Nicotinic Acetylcholine Receptor Modulators Reduce Sugar Intake

    PubMed Central

    Shariff, Masroor; Quik, Maryka; Holgate, Joan; Morgan, Michael; Patkar, Omkar L.; Tam, Vincent; Belmer, Arnauld; Bartlett, Selena E.

    2016-01-01

    Excess sugar consumption has been shown to contribute directly to weight gain, thus contributing to the growing worldwide obesity epidemic. Interestingly, increased sugar consumption has been shown to repeatedly elevate dopamine levels in the nucleus accumbens (NAc), in the mesolimbic reward pathway of the brain similar to many drugs of abuse. We report that varenicline, an FDA-approved nicotinic acetylcholine receptor (nAChR) partial agonist that modulates dopamine in the mesolimbic reward pathway of the brain, significantly reduces sucrose consumption, especially in a long-term consumption paradigm. Similar results were observed with other nAChR drugs, namely mecamylamine and cytisine. Furthermore, we show that long-term sucrose consumption increases α4β2 * and decreases α6β2* nAChRs in the nucleus accumbens, a key brain region associated with reward. Taken together, our results suggest that nAChR drugs such as varenicline may represent a novel treatment strategy for reducing sugar consumption. PMID:27028298

  6. Early Life Stress, Nicotinic Acetylcholine Receptors and Alcohol Use Disorders.

    PubMed

    Holgate, Joan Y; Bartlett, Selena E

    2015-01-01

    Stress is a major driving force in alcohol use disorders (AUDs). It influences how much one consumes, craving intensity and whether an abstinent individual will return to harmful alcohol consumption. We are most vulnerable to the effects of stress during early development, and exposure to multiple traumatic early life events dramatically increases the risk for AUDs. However, not everyone exposed to early life stress will develop an AUD. The mechanisms determining whether an individual's brain adapts and becomes resilient to the effects of stress or succumbs and is unable to cope with stress remain elusive. Emerging evidence suggests that neuroplastic changes in the nucleus accumbens (NAc) following early life stress underlie the development of AUDs. This review discusses the impact of early life stress on NAc structure and function, how these changes affect cholinergic signaling within the mesolimbic reward pathway and the role nicotinic acetylcholine receptors (nAChRs) play in this process. Understanding the neural pathways and mechanism determining stress resilience or susceptibility will improve our ability to identify individuals susceptible to developing AUDs, formulate cognitive interventions to prevent AUDs in susceptible individuals and to elucidate and enhance potential therapeutic targets, such as the nAChRs, for those struggling to overcome an AUD. PMID:26136145

  7. Nicotine Effects in Adolescence and Adulthood on Cognition and α4β2-Nicotinic Receptors in the Neonatal Ventral Hippocampal Lesion Rat Model of Schizophrenia

    PubMed Central

    Berg, Sarah A.; Sentir, Alena M.; Bell, Richard L.; Engleman, Eric A.; Chambers, R. Andrew

    2014-01-01

    Rational Nicotine use in schizophrenia has traditionally been explained as ‘self-medication’ of cognitive and/or nicotinic acetylcholinergic receptor (nAChR) abnormalities. Objectives We test this hypothesis in a neurodevelopmental rat model of schizophrenia that shows increased addiction behaviors including enhanced nicotine reinforcement and drug-seeking. Methods Nicotine transdermal patch (5 mg/kg/day vs. placebo × 10 days in adolescence or adulthood) effects on subsequent radial-arm maze learning (15 sessions) and frontal-cortical-striatal nAChR densities (α4β2; [3H]-epibatidine binding) were examined in neonatal ventral hippocampal lesion (NVHL) and SHAM-operated rats. Results NVHL cognitive deficits were not differentially affected by nicotine history compared to SHAMs. Nicotine history produced minimal cognitive effects while increasing food–reward consumption on the maze, compounding with NVHL-induced overconsumption. Acute nicotine (0.5 mg/kg) delivered before the final maze sessions produced modest improvements in maze performance in rats with nicotine patch histories only, but not differentially so in NVHLs. Consistent with in vivo neuroimaging of β2 nAChR binding in schizophrenia smokers vs. non-smokers and healthy controls, adult NVHLs showed 12% reductions in nAChR binding in MPFC (p<0.05) but not ventral striatum (<5% changes, p>.40), whereas nicotine history elevated nAChRs across both regions (>30%, p<0.001) without interacting with NVHLs. Adolescent vs. adult nicotine exposure did not alter nAChRs differentially. Conclusions Although replicating nicotine-induced up-regulation of nAChRs in human smokers and demonstrating NVHL validity in terms of schizophrenia-associated nAChR density patterns, these findings do not support hypotheses explaining increased nicotine use in schizophrenia as reflecting illness-specific effects of nicotine to therapeutically alter cognition or nAChR densities. PMID:25388292

  8. Opioid and nicotine receptors affect growth regulation of human lung cancer cell lines

    SciTech Connect

    Maneckjee, R.; Minna, J.D. Uniformed Services Univ. of the Health Sciences, Bethesda, MD )

    1990-05-01

    Using specific radioactively-labeled ligands, the authors find that lung cancer cell lines of diverse histologic types express multiple, high-affinity membrane receptors for {mu}, {delta}, and {kappa} opioid agonists and for nicotine and {alpha}-bungarotoxin. These receptors are biologically active because cAMP levels decreased in lung cancer cells after opioid and nicotine application. Nicotine at concentrations found in the blood of smokers had no effect on in vitro lung cancer cell growth, whereas {mu}, {delta}, and {kappa} opioid agonists at low concentrations inhibited lung cancer growth in vitro. They also found that lung cancer cells expressed various combinations of immunoreactive opioid peptides ({beta}-endorphin, enkephalin, or dynorphin), suggesting the participation of opioids in a negative autocrine loop or tumor-suppressing system. Due to the almost universal exposure of patients with lung cancer to nicotine, they tested whether nicotine affected the response of lung cancer cell growth to opioids and found that nicotine at concentrations of 100-200 nM partially or totally reversed opioid-induced growth inhibition in 9/14 lung cancer cell lines. These in vitro results for lung cancer cells suggest that opioids could function as part of a tumor suppressor system and that nicotine can function to circumvent this system in the pathogenesis of lung cancer.

  9. Recurrent exposure to nicotine differentiates human bronchial epithelial cells via epidermal growth factor receptor activation

    SciTech Connect

    Martinez-Garcia, Eva; Irigoyen, Marta; Anso, Elena; Martinez-Irujo, Juan Jose; Rouzaut, Ana

    2008-05-01

    Cigarette smoking is the major preventable cause of lung cancer in developed countries. Nicotine (3-(1-methyl-2-pyrrolidinyl)-pyridine) is one of the major alkaloids present in tobacco. Besides its addictive properties, its effects have been described in panoply of cell types. In fact, recent studies have shown that nicotine behaves as a tumor promoter in transformed epithelial cells. This research focuses on the effects of acute repetitive nicotine exposure on normal human bronchial epithelial cells (NHBE cells). Here we show that treatment of NHBE cells with recurrent doses of nicotine up to 500 {mu}M triggered cell differentiation towards a neuronal-like phenotype: cells emitted filopodia and expressed neuronal markers such as neuronal cell adhesion molecule, neurofilament-M and the transcription factors neuronal N and Pax-3. We also demonstrate that nicotine treatment induced NF-kB translocation to the nucleus, phosphorylation of the epidermal growth factor receptor (EGFR), and accumulation of heparin binding-EGF in the extracellular medium. Moreover, addition of AG1478, an inhibitor of EGFR tyrosine phosphorylation, or cetuximab, a monoclonal antibody that precludes ligand binding to the same receptor, prevented cell differentiation by nicotine. Lastly, we show that differentiated cells increased their adhesion to the extracellular matrix and their protease activity. Given that several lung pathologies are strongly related to tobacco consumption, these results may help to better understand the damaging consequences of nicotine exposure.

  10. In vivo labeling of nicotinic cholinergic receptors in brain with [3H]cytisine.

    PubMed

    Flesher, J E; Scheffel, U; London, E D; Frost, J J

    1994-01-01

    [3H]Cytisine was evaluated as an in vivo ligand for the nicotinic cholinergic receptor (nAchR) in mouse brain. The tracer was injected intravenously, and radioactivity in brain regions was analyzed. Radioactivity peaked in the brain at 30 minutes. It was highest in the thalamus, intermediate in the superior colliculi, prefrontal cortex and hippocampus, and low in the cerebellum. Pretreatment with unlabeled cytisine inhibited binding in the thalamus, but not in the cerebellum. Binding was displaced by l-nicotine, but not by d-nicotine or dexetimide. The results suggest that cytisine, appropriately labeled with a positron emitting radionuclide, may be useful for study of nicotinic cholinergic receptors in humans by emission computed tomography. PMID:8196506

  11. Structural Studies of Nicotinic Acetylcholine Receptors: Using Acetylcholine-Binding Protein as a Structural Surrogate.

    PubMed

    Shahsavar, Azadeh; Gajhede, Michael; Kastrup, Jette S; Balle, Thomas

    2016-06-01

    Nicotinic acetylcholine receptors (nAChRs) are members of the pentameric ligand-gated ion channel superfamily that play important roles in the control of neurotransmitter release in the central and peripheral nervous system. These receptors are important therapeutic targets for the development of drugs against a number of mental health disorders and for marketed smoking cessation aids. Unfortunately, drug discovery has been hampered by difficulties in obtaining sufficiently selective compounds. Together with functional complexity of the receptors, this has made it difficult to obtain drugs with sufficiently high-target to off-target affinity ratios. The recent and ongoing progress in structural studies holds promise to help understand structure-function relationships of nAChR drugs at the atomic level. This will undoubtedly lead to the design of more efficient drugs with fewer side effects. As a high-resolution structure of a nAChR is yet to be determined, structural studies are to a large extent based on acetylcholine-binding proteins (AChBPs) that despite low overall sequence identity display a high degree of conservation of overall structure and amino acids at the ligand-binding site. Further, AChBPs reproduce relative binding affinities of ligands at nAChRs. Over the past decade, AChBPs have been used extensively as models for nAChRs and have aided the understanding of drug receptor interactions at nAChRs significantly. PMID:26572235

  12. Evaluation of the Nicotinic Acetylcholine Receptor-Associated Proteome at Baseline and Following Nicotine Exposure in Human and Mouse Cortex

    PubMed Central

    Esterlis, Irina; Stone, Kathryn L.; Grady, Sharon R.; Lindstrom, Jon M.; Marks, Michael J.

    2016-01-01

    Abstract Nicotinic acetylcholine receptors (nAChRs) support the initiation and maintenance of smoking, but the long-term changes occurring in the protein complex as a result of smoking and the nicotine in tobacco are not known. Human studies and animal models have also demonstrated that increasing cholinergic tone increases behaviors related to depression, suggesting that the nAChR-associated proteome could be altered in individuals with mood disorders. We therefore immunopurified nAChRs and associated proteins for quantitative proteomic assessment of changes in protein–protein interactions of high-affinity nAChRs containing the β2 subunit (β2*-nAChRs) from either cortex of mice treated with saline or nicotine, or postmortem human temporal cortex tissue from tobacco-exposed and nonexposed individuals, with a further comparison of diagnosed mood disorder to control subjects. We observed significant effects of nicotine exposure on the β2*-nAChR-associated proteome in human and mouse cortex, particularly in the abundance of the nAChR subunits themselves, as well as putative interacting proteins that make up core components of neuronal excitability (Na/K ATPase subunits), presynaptic neurotransmitter release (syntaxins, SNAP25, synaptotagmin), and a member of a known nAChR protein chaperone family (14-3-3ζ). These findings identify candidate-signaling proteins that could mediate changes in cholinergic signaling via nicotine or tobacco use. Further analysis of identified proteins will determine whether these interactions are essential for primary function of nAChRs at presynaptic terminals. The identification of differences in the nAChR-associated proteome and downstream signaling in subjects with various mood disorders may also identify novel etiological mechanisms and reveal new treatment targets. PMID:27559543

  13. Evaluation of the Nicotinic Acetylcholine Receptor-Associated Proteome at Baseline and Following Nicotine Exposure in Human and Mouse Cortex.

    PubMed

    McClure-Begley, Tristan D; Esterlis, Irina; Stone, Kathryn L; Lam, TuKiet T; Grady, Sharon R; Colangelo, Christopher M; Lindstrom, Jon M; Marks, Michael J; Picciotto, Marina R

    2016-01-01

    Nicotinic acetylcholine receptors (nAChRs) support the initiation and maintenance of smoking, but the long-term changes occurring in the protein complex as a result of smoking and the nicotine in tobacco are not known. Human studies and animal models have also demonstrated that increasing cholinergic tone increases behaviors related to depression, suggesting that the nAChR-associated proteome could be altered in individuals with mood disorders. We therefore immunopurified nAChRs and associated proteins for quantitative proteomic assessment of changes in protein-protein interactions of high-affinity nAChRs containing the β2 subunit (β2*-nAChRs) from either cortex of mice treated with saline or nicotine, or postmortem human temporal cortex tissue from tobacco-exposed and nonexposed individuals, with a further comparison of diagnosed mood disorder to control subjects. We observed significant effects of nicotine exposure on the β2*-nAChR-associated proteome in human and mouse cortex, particularly in the abundance of the nAChR subunits themselves, as well as putative interacting proteins that make up core components of neuronal excitability (Na/K ATPase subunits), presynaptic neurotransmitter release (syntaxins, SNAP25, synaptotagmin), and a member of a known nAChR protein chaperone family (14-3-3ζ). These findings identify candidate-signaling proteins that could mediate changes in cholinergic signaling via nicotine or tobacco use. Further analysis of identified proteins will determine whether these interactions are essential for primary function of nAChRs at presynaptic terminals. The identification of differences in the nAChR-associated proteome and downstream signaling in subjects with various mood disorders may also identify novel etiological mechanisms and reveal new treatment targets. PMID:27559543

  14. Synaptic modulation of excitatory synaptic transmission by nicotinic acetylcholine receptors in spinal ventral horn neurons.

    PubMed

    Mine, N; Taniguchi, W; Nishio, N; Izumi, N; Miyazaki, N; Yamada, H; Nakatsuka, T; Yoshida, M

    2015-04-01

    Nicotinic acetylcholine receptors (nAChRs) are distributed widely in the central nervous system and play important roles in higher brain functions, including learning, memory, and recognition. However, functions of the cholinergic system in spinal motoneurons remain poorly understood. In this study, we investigated the actions of presynaptic and postsynaptic nAChRs in spinal ventral horn neurons by performing whole-cell patch-clamp recordings on lumbar slices from male rats. The application of nicotine or acetylcholine generated slow inward currents and increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). Slow inward currents by acetylcholine or nicotine were not inhibited by tetrodotoxin (TTX) or glutamate receptor antagonists. In the presence of TTX, the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) were also increased by acetylcholine or nicotine. A selective α4β2 nicotinic receptor antagonist, dihydro-β-erythroidine hydrobromide (DhβE), significantly decreased nicotine-induced inward currents without affecting the enhancement of sEPSCs and mEPSCs. In addition, a selective α7 nicotinic receptor antagonist, methyllycaconitine, did not affect either nicotine-induced inward currents or the enhancement of sEPSCs and mEPSCs. These results suggest that α4β2 AChRs are localized at postsynaptic sites in the spinal ventral horn, non-α4β2 and non-α7 nAChRs are located presynaptically, and nAChRs enhance excitatory synaptic transmission in the spinal ventral horn. PMID:25613686

  15. Ineffectiveness of nicotinic acetylcholine receptor antagonists for treatment-resistant depression: a meta-analysis.

    PubMed

    Wang, Hee Ryung; Woo, Young Sup; Bahk, Won-Myong

    2016-09-01

    Emerging preclinical and clinical evidences suggest a potential role of nicotinic acetylcholine receptors in the pathophysiology of depression. Several clinical trials have investigated the efficacy of nicotinic acetylcholine receptor antagonists in treatment-resistant depression. We carried out this meta-analysis to investigate whether nicotinic acetylcholine receptor antagonists significantly improve symptoms in patients with major depressive disorder who have an inadequate response to standard antidepressant therapy. A comprehensive literature search identified six randomized-controlled trials. These six trials, which included 2067 participants, were pooled for this meta-analysis using a random-effects model. Nicotinic acetylcholine receptor antagonists failed to show superior efficacy compared with placebo in terms of the mean change in the Montgomery-Asberg Depression Rating Scale score [mean difference=-0.12 (95% confidence interval (CI)=-0.96 to 0.71]; response rate [risk ratio=0.92 (95% CI=0.83-1.02)]; and remission rate [risk ratio=1.01 (95% CI=0.83-1.23)]. This meta-analysis failed to confirm preliminary positive evidence for the efficacy of nicotinic acetylcholine receptor antagonists in treatment-resistant depression. Further studies investigating the efficacy of various alternative treatment strategies for treatment-resistant depression will help clinicians to better understand and choose better treatment options for these populations. PMID:26982579

  16. Sequence and functional expression of a single alpha subunit of an insect nicotinic acetylcholine receptor.

    PubMed Central

    Marshall, J; Buckingham, S D; Shingai, R; Lunt, G G; Goosey, M W; Darlison, M G; Sattelle, D B; Barnard, E A

    1990-01-01

    We report the isolation and sequence of a cDNA clone that encodes a locust (Schistocerca gregaria) nervous system nicotinic acetylcholine receptor (AChR) subunit (alpha L1). The calculated molecular weight of the unglycosylated polypeptide, which contains in the proposed extracellular domain two adjacent cysteine residues which are characteristic of alpha (ligand binding) subunits, is 60,641 daltons. Injection into Xenopus oocytes, of RNA synthesized from this clone in vitro, results in expression of functional nicotinic receptors in the oocyte membrane. In these, nicotine opens a cation channel; the receptors are blocked by both alpha-bungarotoxin (alpha-Bgt) and kappa-bungarotoxin (kappa-Bgt). Reversible block of the expressed insect AChR by mecamylamine, d-tubocurarine, tetraethylammonium, bicuculline and strychnine has also been observed. These data are entirely consistent with previously reported electrophysiological studies on in vivo insect nicotinic receptors and also with biochemical studies on an alpha-Bgt affinity purified locust AChR. Thus, a functional receptor exhibiting the characteristic pharmacology of an in vivo insect nicotinic AChR can be expressed in Xenopus oocytes by injection with a single subunit RNA. PMID:1702381

  17. Perinatal Nicotine Exposure Increases Angiotensin II Receptor-Mediated Vascular Contractility in Adult Offspring

    PubMed Central

    Xiao, DaLiao; Dasgupta, Chiranjib; Li, Yong; Huang, Xiaohui; Zhang, Lubo

    2014-01-01

    Previous studies have reported that perinatal nicotine exposure causes development of hypertensive phenotype in adult offspring. Aims The present study was to determine whether perinatal nicotine exposure causes an epigenetic programming of vascular Angiotensin II receptors (ATRs) and their-mediated signaling pathway leading to heightened vascular contraction in adult offspring. Main methods Nicotine was administered to pregnant rats via subcutaneous osmotic minipumps from day 4 of gestation to day 10 after birth. The experiments were conducted at 5 months of age of male offspring. Key Findings Nicotine treatment enhanced Angitension II (Ang II)-induced vasoconstriction and 20-kDa myosin light chain phosphorylation (MLC20-P) levels. In addition, the ratio of Ang II-induced tension/MLC-P was also significantly increased in nicotine-treated group compared with the saline group. Nicotine-mediated enhanced constrictions were not directly dependent on the changes of [Ca2+]i concentrations but dependent on Ca2+ sensitivity. Perinatal nicotine treatment significantly enhanced vascular ATR type 1a (AT1aR) but not AT1bR mRNA levels in adult rat offspring, which was associated with selective decreases in DNA methylation at AT1aR promoter. Contrast to the effect on AT1aR, nicotine decreased the mRNA levels of vascular AT2R gene, which was associated with selective increases in DNA methylation at AT2R promoter. Significance Our results indicated that perinatal nicotine exposure caused an epigenetic programming of vascular ATRs and their-mediated signaling pathways, and suggested that differential regulation of AT1R/AT2R gene expression through DNA methylation mechanism may be involved in nicotine-induced heightened vasoconstriction and development of hypertensive phenotype in adulthood. PMID:25265052

  18. Genetic association of bipolar disorder with the β3 nicotinic receptor subunit gene

    PubMed Central

    Hartz, Sarah M.; Lin, Peng; Edenberg, Howard J.; Xuei, Xiaoling; Rochberg, Nanette; Saccone, Scott; Berrettini, Wade; Nelson, Elliot; Nurnberger, John; Bierut, Laura J.; Rice, John P.

    2011-01-01

    Objective Owing to the clinical relationship between bipolar disorder and nicotine dependence, we investigated two research questions: (i) are genetic associations with nicotine dependence different in individuals with bipolar disorder as compared with individuals without bipolar disorder, and (ii) do loci earlier associated with nicotine dependence have pleiotropic effects on these two diseases. Method Our study consisted of 916 cases with bipolar disorder and 1028 controls. On the basis of known associations with nicotine dependence, we genotyped eight single-nucleotide polymorphisms (SNPs) on chromosome 8 (three bins) in the regions of CHRNB3 and CHRNA6, and six SNPs on chromosome 15 (three bins) in the regions of CHRNA5 and CHRNA3. Results To determine whether the genetic associations with nicotine dependence are different in bipolar disorder than in the general population, we compared allele frequencies of candidate SNPs between individuals with nicotine dependence only and individuals with both nicotine dependence and bipolar disorder. There were no statistical differences between these frequencies, indicating that genetic association with nicotine dependence is similar in individuals with bipolar disorder as in the general population. In the investigation of pleiotropic effects of these SNPs on bipolar disorder, two highly correlated synonymous SNPs in CHRNB3, rs4952 and rs4953, were significantly associated with bipolar disorder (odds ratio 1.7, 95% confidence interval: 1.2–2.4, P = 0.001). This association remained significant both after adjusting for a smoking covariate and analyzing the association in nonsmokers only. Conclusion Our results suggest that (i) bipolar disorder does not modify the association between nicotine dependence and nicotinic receptor subunit genes, and (ii) variants in CHRNB3/CHRNA6 are independently associated with bipolar disorder. Psychiatr Genet 00:000–000. PMID:21191315

  19. ROLE OF NMDA, NICOTINIC, AND GABA RECEPTORS IN THE STEADY STATE VISUAL EVOKED POTENTIAL IN RATS.

    EPA Science Inventory

    This manuscript characterizes the receptor pathways involved in pattern-evoked potential generation in rats

    " NMDA and nicotinic acetylcholine receptors appear to be involved in the generation of the steady-state pattern evoked response in vivo.

    " The pattern evok...

  20. Serotoninergic dorsal raphe neurons possess functional postsynaptic nicotinic acetylcholine receptors.

    PubMed

    Galindo-Charles, Luis; Hernandez-Lopez, Salvador; Galarraga, Elvira; Tapia, Dagoberto; Bargas, José; Garduño, Julieta; Frías-Dominguez, Carmen; Drucker-Colin, René; Mihailescu, Stefan

    2008-08-01

    Very few neurons in the telencephalon have been shown to express functional postsynaptic nicotinic acetylcholine receptors (nAChRs), among them, the noradrenergic and dopaminergic neurons. However, there is no evidence for postsynaptic nAChRs on serotonergic neurons. In this study, we asked if functional nAChRs are present in serotonergic (5-HT) and nonserotonergic (non-5-HT) neurons of the dorsal raphe nucleus (DRN). In rat midbrain slices, field stimulation at the tegmental pedunculopontine (PPT) nucleus evoked postsynaptic currents (eEPSCs) with different components in DRN neurons. After blocking the glutamatergic and GABAergic components, the remaining eEPSCs were blocked by mecamylamine and reduced by either the selective alpha7 nAChR antagonist methyllycaconitine (MLA) or the selective alpha4beta2 nAChR antagonist dihydro-beta-eritroidine (DHbetaE). Simultaneous addition of MLA and DHbetaE blocked all eEPSCs. Integrity of the PPT-DRN pathway was assessed by both anterograde biocytin tracing and antidromic stimulation from the DRN. Inward currents evoked by the direct application of acetylcholine (ACh), in the presence of atropine and tetrodotoxin, consisted of two kinetically different currents: one was blocked by MLA and the other by DHbetaE; in both 5-HT and non-5-HT DR neurons. Analysis of spontaneous (sEPSCs) and evoked (eEPSCs) synaptic events led to the conclusion that nAChRs were located at the postsynaptic membrane. The possible implications of these newly described nAChRs in various physiological processes and behavioral events, such as the wake-sleep cycle, are discussed. PMID:18512214

  1. Nicotinic acetylcholine receptor expression in human airway correlates with lung function.

    PubMed

    Lam, David Chi-Leung; Luo, Susan Yang; Fu, Kin-Hang; Lui, Macy Mei-Sze; Chan, Koon-Ho; Wistuba, Ignacio Ivans; Gao, Boning; Tsao, Sai-Wah; Ip, Mary Sau-Man; Minna, John Dorrance

    2016-02-01

    Nicotine and its derivatives, by binding to nicotinic acetylcholine receptors (nAChRs) on bronchial epithelial cells, can regulate cellular signaling and inflammatory processes. Delineation of nAChR subtypes and their responses to nicotine stimulation in bronchial epithelium may provide information for therapeutic targeting in smoking-related inflammation in the airway. Expression of nAChR subunit genes in 60 bronchial epithelial biopsies and immunohistochemical staining for the subcellular locations of nAChR subunit expression were evaluated. Seven human bronchial epithelial cell lines (HBECs) were exposed to nicotine in vitro for their response in nAChR subunit gene expression to nicotine exposure and removal. The relative normalized amount of expression of nAChR α4, α5, and α7 and immunohistochemical staining intensity of nAChR α4, α5, and β3 expression showed significant correlation with lung function parameters. Nicotine stimulation in HBECs resulted in transient increase in the levels of nAChR α5 and α6 but more sustained increase in nAChR α7 expression. nAChR expression in bronchial epithelium was found to correlate with lung function. Nicotine exposure in HBECs resulted in both short and longer term responses in nAChR subunit gene expression. These results gave insight into the potential of targeting nAChRs for therapy in smoking-related inflammation in the airway. PMID:26608528

  2. Decreasing nicotinic receptor activity and the spatial learning impairment caused by the NMDA glutamate antagonist dizocilpine in rats

    PubMed Central

    Burke, Dennis A.; Heshmati, Pooneh; Kholdebarin, Ehsan; Levin, Edward D.

    2014-01-01

    Nicotinic systems have been shown by a variety of studies to be involved in cognitive function. Nicotinic receptors have an inherent property to become desensitized after activation. The relative role of nicotinic receptor activation vs. net receptor inactivation by desensitization in the cognitive effects of nicotinic drugs remains to be fully understood. In these studies, we tested the effects of the α7 nicotinic receptor antagonist methyllycaconitine (MLA), the α4β2 nicotinic receptor antagonist dihydro-β-erythroidine (DHβE), the nonspecific nicotinic channel blocker mecamylamine and the α4β2 nicotinic receptor desensitizing agent sazetidine-A on learning in a repeated acquisition test. Adult female Sprague-Dawley rats were trained on a repeated acquisition learning procedure in an 8-arm radial maze. MLA (1–4 mg/kg), DHβE (1–4 mg/kg), mecamylamine (0.125–0.5 mg/kg) or sazetidine-A (1 and 3 mg/kg) were administered in four different studies either alone or together with the NMDA glutamate antagonist dizocilpine (0.05 and 0.10 mg/kg). MLA significantly counteracted the learning impairment caused by dizocilpine. The overall choice accuracy impairment caused by dizocilpine was significantly attenuated by co-administration of DHβE. Low doses of the non-specific nicotinic antagonist mecamylamine also reduced dizocilpine-induced repeated acquisition impairment. Sazetidine-A reversed the accuracy impairment caused by dizocilpine. These studies provide evidence that a net decrease in nicotinic receptor activity can improve learning by attenuating learning impairment induced by NMDA glutamate blockade. This adds to evidence in cognitive tests that nicotinic antagonists can improve cognitive function. Further research characterizing the efficacy and mechanisms underlying nicotinic antagonist and desensitization induced cognitive improvement is warranted. PMID:25064338

  3. Fixation of allosteric states of the nicotinic acetylcholine receptor by chemical cross-linking

    PubMed Central

    Watty, Anke; Methfessel, Christoph; Hucho, Ferdinand

    1997-01-01

    Receptor activity can be described in terms of ligand-induced transitions between functional states. The nicotinic acetylcholine receptor (nAChR), a prototypic ligand-gated ion channel, is an “unconventional allosteric protein” which exists in at least three interconvertible conformations, referred to as resting (low agonist affinity, closed channel), activated (open channel), and desensitized (high agonist affinity, closed channel). Here we show that 3,3′-dimethyl suberimidate (DMS) is an agonistic bifunctional cross-linking reagent, which irreversibly “freezes” the nAChR in a high agonist affinity/closed-channel state. The monofunctional homologue methyl acetoimidate, which is also a weak cholinergic agonist, has no such irreversible effect. Glutardialdehyde, a cross-linker that is not a cholinergic effector, fixes the receptor in a low-affinity state in the absence of carbamoylcholine, but, like DMS, in a high-affinity state in its presence. Covalent cross-linking thus allows us to arrest the nAChR in defined conformational states. PMID:9223339

  4. Determinants of zinc potentiation on the alpha4 subunit of neuronal nicotinic receptors.

    PubMed

    Hsiao, Bernard; Mihalak, Karla B; Repicky, Sarah E; Everhart, Drew; Mederos, Ana H; Malhotra, Arun; Luetje, Charles W

    2006-01-01

    We have shown previously that the function of neuronal nicotinic acetylcholine receptors can be modulated by zinc. This modulation varies from potentiation to inhibition, depending on receptor subunit composition and zinc concentration, with the alpha4beta2 and alpha4beta4 receptors displaying the most dramatic potentiation. In this study, we used site-directed mutagenesis to identify glutamate 59 and histidine 162 on the rat alpha4 subunit as potential mediators of zinc potentiation. By modeling the extracellular domain of the receptor pentamer, we locate these residues to two subunit-subunit interfaces that alternate with the two acetylcholine-binding interfaces. Substitution of a cysteine at either position allows additional reduction of zinc potentiation upon treatment with the methanethiosulfonate reagents N-biotinoylaminoethyl methanethiosulfonate (MTSEA-biotin) and [2-(trimethylammonium)ethyl] methanethiosulfonate. Mutagenesis and methanethiosulfonate treatment are most effective at position 162, and the presence of zinc hinders the reaction of MTSEA-biotin with the substituted cysteine at this position, suggesting that alpha4His162 participates in forming a coordination site for zinc. Mutagenesis and methanethiosulfonate treatment are less effective at position 59, suggesting that whereas alpha4Glu59 may be near the zinc coordination site, it may not be participating in coordination of the zinc ion. It is noteworthy that the position of alpha4Glu59 within the neuronal nAChR is identical to that of a residue that lines the benzodiazepine-binding site on GABA(A) receptors. We suggest that the zinc potentiation sites on neuronal nAChRs are structurally and functionally similar to the benzodiazepine-binding sites on GABA(A) receptors. PMID:16189299

  5. Association of a nicotinic receptor gene polymorphism with spontaneous eyeblink rates

    PubMed Central

    Nakano, Tamami; Kuriyama, Chiho; Himichi, Toshiyuki; Nomura, Michio

    2015-01-01

    Spontaneous eyeblink rates greatly vary among individuals from several blinks to a few dozen blinks per minute. Because dopamine agonists immediately increase the blink rate, individual differences in blink rate are used as a behavioral index of central dopamine functioning. However, an association of the blink rate with polymorphisms in dopamine-related genes has yet not been found. In this study, we demonstrated that a genetic variation of the nicotinic acetylcholine receptor CHRNA4 (rs1044396) increased the blink rate while watching a video. A receiver operating characteristic analysis revealed that the blink rate predicts a genetic variation in the nicotinic receptor gene with a significant discrimination level (0.66, p < 0.004). The present study suggests that differences in sensitivity to acetylcholine because of the genetic variation of the nicotinic receptor are associated with individual differences in spontaneous eye blink rate. PMID:25729002

  6. Mu Opioid Receptor Binding Correlates with Nicotine Dependence and Reward in Smokers

    PubMed Central

    Brasic, James R.; Contoreggi, Carlo; Cascella, Nicola; Mackowick, Kristen M.; Taylor, Richard; Rousset, Olivier; Willis, William; Huestis, Marilyn A.; Concheiro, Marta; Wand, Gary; Wong, Dean F.; Volkow, Nora D.

    2014-01-01

    The rewarding effects of nicotine are associated with activation of nicotine receptors. However, there is increasing evidence that the endogenous opioid system is involved in nicotine's rewarding effects. We employed PET imaging with [11C]carfentanil to test the hypotheses that acute cigarette smoking increases release of endogenous opioids in the human brain and that smokers have an upregulation of mu opioid receptors (MORs) when compared to nonsmokers. We found no significant changes in binding potential (BPND) of [11C]carfentanil between the placebo and the active cigarette sessions, nor did we observe differences in MOR binding between smokers and nonsmokers. Interestingly, we showed that in smokers MOR availability in bilateral superior temporal cortices during the placebo condition was negatively correlated with scores on the Fagerström Test for Nicotine Dependence (FTND). Also in smokers, smoking-induced decreases in [11C]carfentanil binding in frontal cortical regions were associated with self-reports of cigarette liking and wanting. Although we did not show differences between smokers and nonsmokers, the negative correlation with FTND corroborates the role of MORs in superior temporal cortices in nicotine addiction and provides preliminary evidence of a role of endogenous opioid signaling in frontal cortex in nicotine reward. PMID:25493427

  7. Nicotine ameliorates NMDA receptor antagonist-induced deficits in contextual fear conditioning through high-affinity nicotinic acetylcholine receptors in the hippocampus.

    PubMed

    André, Jessica M; Leach, Prescott T; Gould, Thomas J

    2011-03-01

    NMDA glutamate receptors (NMDARs) and nicotinic acetylcholine receptors (nAChRs) are both involved in learning and synaptic plasticity. Increasing evidence suggests processes mediated by these receptors may interact to modulate learning; however, little is known about the neural substrates involved in these interactive processes. The present studies investigated the effects of nicotine on MK-801 hydrogen maleate (MK-801) and DL-2-Amino-5-phosphonovaleric acid (APV)-induced disruption of contextual fear conditioning in male C57BL/6J mice, using direct drug infusion and selective nAChR antagonists to define the brain regions and the nAChR subtypes involved. Mice treated with MK-801 showed a deficit in contextual fear conditioning that was ameliorated by nicotine. Direct drug infusion demonstrated that the NMDAR antagonists disrupted hippocampal function and that nicotine acted in the dorsal hippocampus to ameliorate the deficit in learning. The high-affinity nAChR antagonist Dihydro-β-erythroidine hydrobromide (DhβE) blocked the effects of nicotine on MK-801-induced deficits while the α7 nAChR antagonist methyllycaconitine citrate salt hydrate (MLA) did not. These results suggest that NMDARs and nAChRs may mediate similar hippocampal processes involved in contextual fear conditioning. Furthermore, these results may have implications for developing effective therapeutics for the cognitive deficits associated with schizophrenia because a large subset of patients with schizophrenia exhibit cognitive deficits that may be related to NMDAR dysfunction and smoke at much higher rates than the healthy population, which may be an attempt to ameliorate cognitive deficits. PMID:21167848

  8. Nicotinic receptor activation negatively modulates pro-inflammatory cytokine production in multiple sclerosis patients.

    PubMed

    Reale, Marcella; Di Bari, Maria; Di Nicola, Marta; D'Angelo, Chiara; De Angelis, Federica; Velluto, Lucia; Tata, Ada Maria

    2015-11-01

    Acetylcholine (ACh) and its receptors of muscarinic and nicotinic types are involved in the modulation of immune and inflammatory responses. In present work we have characterized the nicotinic receptors expression in PBMC of RR-MS patients and healthy donors (HD) and their ability to modulate pro-inflammatory cytokines. Here we report that the IL-1β e IL-17 levels are significantly increased in serum of RR-MS patients in respect to HD and that the PBMC stimulation with PHA caused a significant increase in pro-inflammatory cytokine levels both in RR-MS and HD subjects, with higher increase of protein release in RR-MS patients than in HD. The PBMC treatment with PHA plus nicotine produced a significant decrease of IL-1β e IL-17 both as transcript and as protein, confirming that the PBMC of the patients respond to the cholinergic stimulation more than PBMC of HD. By real time PCR and western blot analysis we have also demonstrated that in particular α7 receptor subtype appeared expressed at comparable levels both in RR-MS patients and HD. The PHA stimulation results to inhibit the α7 subunit expression while the nicotine causes a significant increase in α7 transcripts but only in MS patients. The data obtained highlight the role of α7 receptor subtype in the modulation of anti-inflammatory cytokines also in MS. Moreover the ability of nicotine to up-regulate the expression of α7 receptor subtype in RR-MS patients, indicates that nicotinic receptor stimulation may contribute to down-modulate the inflammation occurred in MS by a positive feedback control of its expression. PMID:26209886

  9. Understanding the Bases of Function and Modulation of α7 Nicotinic Receptors: Implications for Drug Discovery.

    PubMed

    Corradi, Jeremías; Bouzat, Cecilia

    2016-09-01

    The nicotinic acetylcholine receptor (nAChR) belongs to a superfamily of pentameric ligand-gated ion channels involved in many physiologic and pathologic processes. Among nAChRs, receptors comprising the α7 subunit are unique because of their high Ca(2+) permeability and fast desensitization. nAChR agonists elicit a transient ion flux response that is further sustained by the release of calcium from intracellular sources. Owing to the dual ionotropic/metabotropic nature of α7 receptors, signaling pathways are activated. The α7 subunit is highly expressed in the nervous system, mostly in regions implicated in cognition and memory and has therefore attracted attention as a novel drug target. Additionally, its dysfunction is associated with several neuropsychiatric and neurologic disorders, such as schizophrenia and Alzheimer's disease. α7 is also expressed in non-neuronal cells, particularly immune cells, where it plays a role in immunity, inflammation, and neuroprotection. Thus, α7 potentiation has emerged as a therapeutic strategy for several neurologic and inflammatory disorders. With unique activation properties, the receptor is a sensitive drug target carrying different potential binding sites for chemical modulators, particularly agonists and positive allosteric modulators. Although macroscopic and single-channel recordings have provided significant information about the underlying molecular mechanisms and binding sites of modulatory compounds, we know just the tip of the iceberg. Further concerted efforts are necessary to effectively exploit α7 as a drug target for each pathologic situation. In this article, we focus mainly on the molecular basis of activation and drug modulation of α7, key pillars for rational drug design. PMID:27190210

  10. Nicotinic Acid Activates the Capsaicin Receptor TRPV1 – A Potential Mechanism for Cutaneous Flushing

    PubMed Central

    Ma, Linlin; Lee, Bo Hyun; Mao, Rongrong; Cai, Anping; Jia, Yunfang; Clifton, Heather; Schaefer, Saul; Xu, Lin; Zheng, Jie

    2014-01-01

    Objective Nicotinic acid (a.k.a. niacin or vitamin B3), widely used to treat dyslipidemias, represents an effective and safe means to reduce the risk of mortality from cardiovascular disease. Nonetheless, a substantial fraction of patients discontinue treatment due to a strong side effect of cutaneous vasodilation, commonly termed flushing. In the present study we tested the hypothesis that nicotinic acid causes flushing partially by activating the capsaicin receptor TRPV1, a polymodal cellular sensor that mediates the flushing response upon consumption of spicy food. Approach and Results We observed that the nicotinic acid-induced increase in blood flow was substantially reduced in Trpv1−/− knockout mice, indicating involvement of the channel in flushing response. Using exogenously expressed TRPV1, we confirmed that nicotinic acid at sub-millimolar to millimolar concentrations directly and potently activates TRPV1 from the intracellular side. Binding of nicotinic acid to TRPV1 lowers its activation threshold for heat, causing channel opening at physiological temperatures. Activation of TRPV1 by voltage or ligands (capsaicin and 2-APB) is also potentiated by nicotinic acid. We further demonstrated that nicotinic acid does not compete directly with capsaicin but may activate TRPV1 through the 2-APB activation pathway. Using live-cell fluorescence imaging, we observed that nicotinic acid can quickly enter the cell through a transporter-mediated pathway to activate TRPV1. Conclusions Direct activation of TRPV1 by nicotinic acid may lead to cutaneous vasodilation that contributes to flushing, suggesting a potential novel pathway to inhibit flushing and improve compliance. PMID:24675661

  11. Activation of α2A-Containing Nicotinic Acetylcholine Receptors Mediates Nicotine-Induced Motor Output in Embryonic Zebrafish

    PubMed Central

    Menelaou, Evdokia; Udvadia, Ava J.; Tanguay, Robert L.; Svoboda, Kurt R.

    2014-01-01

    It is well established that cholinergic signaling has critical roles during central nervous system development. In physiological and behavioral studies, activation of nicotinic acetylcholine receptors has been implicated in mediating cholinergic signaling. In developing spinal cord, cholinergic transmission is associated with neural circuits responsible for producing locomotor behaviors. In this study, we investigated the expression pattern of the α2A nAChR subunit as evidence from others suggested it could be expressed by spinal neurons. In situ hybridization and immunohistochemistry revealed that the α2A nAChR subunits are expressed in spinal Rohon-Beard (RB) neurons and olfactory sensory neurons in young embryos. In order to examine the functional role of the α2A nAChR subunit during embryogenesis, we blocked its expression using antisense modified oligonucleotides. Blocking the expression of α2A nAChR subunits had no effect on spontaneous motor activity. However, it did alter the embryonic nicotine-induced motor output. This reduction in motor activity was not accompanied by defects in neuronal and muscle elements associated with the motor output. Moreover, the anatomy and functionality of RB neurons was normal even in the absence of the α2A nAChR subunit. Thus, we propose that α2A-containing nAChR are dispensable for normal RB development. However, in the context of nicotine-induced motor output, α2A-containing nAChRs on RB neurons provide the substrate that nicotine acts upon to induce the motor output. These findings also indicate that functional neuronal nAChRs are present within spinal cord at the time when locomotor output in zebrafish first begins to manifest itself. PMID:24738729

  12. Peer Smoking and the Nicotinic Receptor Genes: An Examination of Genetic and Environmental Risks for Nicotine Dependence

    PubMed Central

    Johnson, Eric O.; Chen, Li-Shiun; Breslau, Naomi; Hatsukami, Dorothy; Robbins, Tania; Saccone, Nancy L.; Grucza, Richard A.; Bierut, Laura J.

    2010-01-01

    Background Peer smoking provides a socially reinforcing context of friends’ encouragement and approval that contributes to smoking behavior. Twin studies show correlations and interactions between peer substance use and genetic liability for substance use. However, none examined specific genes. Here we test the hypothesis that the nicotinic receptor genes CHRNA5 (rs16969968), CHRNA3 (rs578776), CHRNB3 (rs13277254), and CHRND (rs12466358) modify the risk for nicotine dependence (ND) associated with peer smoking. Methods Cases of current nicotine dependence (FTND ≥ 4) and smoking-exposed (smoked 100+ cigarettes lifetime), but non-dependent controls (lifetime FTND = 0) came from the Collaborative Genetic Study of Nicotine Dependence (n=2,038). Peer smoking was retrospectively assessed for grades 9–12. Results Peer smoking and the four SNPs were associated with ND. A statistically significant interaction was found between peer smoking and rs16969968 (p = 0.0077). Overall risk of ND was highest for the rs16969968 AA genotype. However, variance in ND attributable to peer smoking was substantially lower among those with the AA genotype at rs16969968 than the lower risk genotypes: AA = 2.5%, GA/AG = 11.2%, GG = 14.2%; p ≤ 0.004. Conclusions Peer smoking had a substantially lower effect on ND among those with the high risk AA genotype at the functional SNP rs16969968 (CHRNA5) than among those with lower risk genotypes. Such results highlight the possibility that given drug exposure those with specific genetic risks may be less affected by social contexts and intervention strategies focused on social factors could have less influence on those at highest genetic risk. PMID:20840187

  13. Functional nicotinic acetylcholine receptor reconstitution in Au(111)-supported thiolipid monolayers

    NASA Astrophysics Data System (ADS)

    Pissinis, Diego E.; Diaz, Carolina; Maza, Eliana; Bonini, Ida C.; Barrantes, Francisco J.; Salvarezza, Roberto C.; Schilardi, Patricia L.

    2015-09-01

    The insertion and function of the muscle-type nicotinic acetylcholine receptor (nAChR) in Au(111)-supported thiolipid self-assembled monolayers have been studied by atomic force microscopy (AFM), surface plasmon resonance (SPR), and electrochemical techniques. It was possible for the first time to resolve the supramolecular arrangement of the protein spontaneously inserted in a thiolipid monolayer in an aqueous solution. Geometric supramolecular arrays of nAChRs were observed, most commonly in a triangular form compatible with three nAChR dimers of ~20 nm each. Addition of the full agonist carbamoylcholine activated and opened the nAChR ion channel, as revealed by the increase in capacitance relative to that of the nAChR-thiolipid system under basal conditions. Thus, the self-assembled system appears to be a viable biomimetic model to measure ionic conductance mediated by ion-gated ion channels under different experimental conditions, with potential applications in biotechnology and pharmacology.

  14. Discovery of a novel nicotinic receptor antagonist for the treatment of nicotine addiction: 1-(3-picolinium)-12-triethylammonium-dodecane dibromide (TMPD)

    PubMed Central

    Dwoskin, Linda P.; Joyce, B. Matthew; Zheng, Guangrong; Neugebauer, Nichole M.; Manda, Vamshi K.; Lockman, Paul; Papke, Roger L.; Bardo, Michael T.; Crooks, Peter A.

    2007-01-01

    Limitations in efficacy and high relapse rates of currently available smoking cessation agents reveal the need for more efficacious pharmacotherapies. One strategy is to develop subtype-selective nicotinic receptor (nAChR) antagonists that inhibit nicotine-evoked dopamine (DA) release, the primary neurotransmitter involved in nicotine reward. Simple alkylation of the pyridino N-atom converts nicotine from a potent agonist into a potent antagonist. The classical antagonists, hexamethonium and decamethonium, differentiate between peripheral nAChR subtypes. Using a similar approach, we interconnected varying quaternary ammonium moieties with a lipophilic linker to provide N,N′- bis-nicotinium analogs, affording a lead compound, N,N′-dodecyl-1,12-diyl-bis-3-picolinium dibromide (bPiDDB), which inhibited nicotine-evoked DA release and decreased nicotine self-administration. The current work describes a novel compound, 1-(3-picolinium)-12-triethylammonium-dodecane dibromide (TMPD), a hybrid of bPiDDB and decamethonium. TMPD completely inhibited (IC50 = 500 nM) nicotine-evoked DA release from superfused rat striatal slices, suggesting that TMPD acts as a nAChR antagonist at more than one subtype. TMPD (1 μM) inhibited the response to acetylcholine at α3β4, α4β4, α4β2, and α1β1εδ receptors expressed in Xenopus oocytes. TMPD had a 2-fold higher affinity for the blood-brain barrier choline transporter, suggesting that is brain bioavailable. TMPD did not inhibit the hyperactivity in nicotine sensitized rats, but significantly and specifically decreased nicotine self-administration. Together, the results suggest that TMPD may have the ability to reduce the rewarding effect of nicotine with minimal side effects, a pharmacological profile indicative of potential clinical utility for the treatment of tobacco dependence. PMID:17727820

  15. Mood and anxiety regulation by nicotinic acetylcholine receptors: A potential pathway to modulate aggression and related behavioral states.

    PubMed

    Picciotto, Marina R; Lewis, Alan S; van Schalkwyk, Gerrit I; Mineur, Yann S

    2015-09-01

    The co-morbidity between smoking and mood disorders is striking. Preclinical and clinical studies of nicotinic effects on mood, anxiety, aggression, and related behaviors, such as irritability and agitation, suggest that smokers may use the nicotine in tobacco products as an attempt to self-medicate symptoms of affective disorders. The role of nicotinic acetylcholine receptors (nAChRs) in circuits regulating mood and anxiety is beginning to be elucidated in animal models, but the mechanisms underlying the effects of nicotine on aggression-related behavioral states (ARBS) are still not understood. Clinical trials of nicotine or nicotinic medications for neurological and psychiatric disorders have often found effects of nicotinic medications on ARBS, but few trials have studied these outcomes systematically. Similarly, the increase in ARBS resulting from smoking cessation can be resolved by nicotinic agents, but the effects of nicotinic medications on these types of mental states and behaviors in non-smokers are less well understood. Here we review the literature on the role of nAChRs in regulating mood and anxiety, and subsequently on the closely related construct of ARBS. We suggest avenues for future study to identify how nAChRs and nicotinic agents may play a role in these clinically important areas. This article is part of the Special Issue entitled 'The Nicotinic Acetylcholine Receptor: From Molecular Biology to Cognition'. PMID:25582289

  16. Characterization of a purified nicotinic receptor from rat brain by using idiotypic and anti-idiotypic antibodies

    SciTech Connect

    Abood, L.G.; Langone, J.J.; Bjercke, R.; Lu, X.; Banerjee, S.

    1987-09-01

    The availability of an anti-nicotine monoclonal antibody has made it possible to further establish the nature of the nicotine recognition proteins purified from rat brain by affinity chromatography and to provide a highly sensitive assay for determining (/sup 3/H)nicotine binding to the purified material. An enantiomeric analogue of nicotine. (-)-6-hydroxymethylnicotine, was used to prepare the affinity column. In addition, with the use of an anti-idiotypic monoclonal antibody, it was confirmed that the recognition site for nicotine resides on a protein complex composed of two components with molecular masses of 62 and 57 kDa. It was also demonstrated that the same two proteins could be purified by immunoaffinity chromatography with the use of an anti-idiotypic monoclonal antibody. With the use of the anti-nicotine antibody to measure (/sup 3/H)nicotine binding, the purified material was shown to bind 250 pmol/mg of protein. By utilizing a procedure in which the purified receptor protein was conjugated to membranes by disulfide bonds, a binding activity of 80 pmol/mg was obtained. With the availability of sterospecific monoclonal antibodies to (-)-nicotine as well as monoclonal anti-idiotypic antibodies derived when the anti-nicotine antibodies were used as immunogens, additional procedures became available for the further characterization of the purified nicotine receptor and examining its (-)-(/sup 3/H)nicotine-binding characteristics.

  17. Localized low-level re-expression of high-affinity mesolimbic nicotinic acetylcholine receptors restores nicotine-induced locomotion but not place conditioning.

    PubMed

    Mineur, Y S; Brunzell, D H; Grady, S R; Lindstrom, J M; McIntosh, J M; Marks, M J; King, S L; Picciotto, M R

    2009-04-01

    High-affinity, beta2-subunit-containing (beta2*) nicotinic acetylcholine receptors (nAChRs) are essential for nicotine reinforcement; however, these nAChRs are found on both gamma-aminobutyric acid (GABA) and dopaminergic (DA) neurons in the ventral tegmental area (VTA) and also on terminals of glutamatergic and cholinergic neurons projecting from the pedunculopontine tegmental area and the laterodorsal tegmental nucleus. Thus, systemic nicotine administration stimulates many different neuronal subtypes in various brain nuclei. To identify neurons in which nAChRs must be expressed to mediate effects of systemic nicotine, we investigated responses in mice with low-level, localized expression of beta2* nAChRs in the midbrain/VTA. Nicotine-induced GABA and DA release were partially rescued in striatal synaptosomes from transgenic mice compared with tissue from beta2 knockout mice. Nicotine-induced locomotor activation, but not place preference, was rescued in mice with low-level VTA expression, suggesting that low-level expression of beta2* nAChRs in DA neurons is not sufficient to support nicotine reward. In contrast to control mice, transgenic mice with low-level beta2* nAChR expression in the VTA showed no increase in overall levels of cyclic AMP response element-binding protein (CREB) but did show an increase in CREB phosphorylation in response to exposure to a nicotine-paired chamber. Thus, CREB activation in the absence of regulation of total CREB levels during place preference testing was not sufficient to support nicotine place preference in beta2 trangenic mice. This suggests that partial activation of high-affinity nAChRs in VTA might block the rewarding effects of nicotine, providing a potential mechanism for the ability of nicotinic partial agonists to aid in smoking cessation. PMID:19077117

  18. Pesticide exposure during pregnancy, like nicotine, affects the brainstem α7 nicotinic acetylcholine receptor expression, increasing the risk of sudden unexplained perinatal death.

    PubMed

    Lavezzi, Anna Maria; Cappiello, Achille; Pusiol, Teresa; Corna, Melissa Felicita; Termopoli, Veronica; Matturri, Luigi

    2015-01-15

    This study indicates the impact of nicotine and pesticides (organochlorine and organophosphate insecticides used in agriculture) on neuronal α7-nicotinic acetylcholine receptor expression in brainstem regions receiving cholinergic projections in human perinatal life. An in-depth anatomopathological examination of the autonomic nervous system and immunohistochemistry to analyze the α7-nicotinic acetylcholine receptor expression in the brainstem from 44 fetuses and newborns were performed. In addition, the presence of selected agricultural pesticides in cerebral cortex samples of the victims was determined by specific analytical procedures. Hypodevelopment of brainstem structures checking the vital functions, frequently associated with α7-nicotinic acetylcholine receptor immunopositivity and smoke absorption in pregnancy, was observed in high percentages of victims of sudden unexpected perinatal death. In nearly 30% of cases however the mothers never smoked, but lived in rural areas. The search for pesticides highlighted in many of these cases traces of both organochlorine and organophosphate pesticides. We detain that exposition to pesticides in pregnancy produces homologous actions to those of nicotine on neuronal α7-nicotinic acetylcholine receptor, allowing to developmental alterations of brainstem vital centers in victims of sudden unexplained death. PMID:25433450

  19. Nicotine promotes cell proliferation via {alpha}7-nicotinic acetylcholine receptor and catecholamine-synthesizing enzymes-mediated pathway in human colon adenocarcinoma HT-29 cells

    SciTech Connect

    Wong, Helen Pui Shan; Yu Le; Lam, Emily Kai Yee; Tai, Emily Kin Ki; Wu, William Ka Kei; Cho, Chi Hin . E-mail: chcho@cuhk.edu.hk

    2007-06-15

    Cigarette smoking has been implicated in colon cancer. Nicotine is a major alkaloid in cigarette smoke. In the present study, we showed that nicotine stimulated HT-29 cell proliferation and adrenaline production in a dose-dependent manner. The stimulatory action of nicotine was reversed by atenolol and ICI 118,551, a {beta}{sub 1}- and {beta}{sub 2}-selective antagonist, respectively, suggesting the role of {beta}-adrenoceptors in mediating the action. Nicotine also significantly upregulated the expression of the catecholamine-synthesizing enzymes [tyrosine hydroxylase (TH), dopamine-{beta}-hydroxylase (D{beta}H) and phenylethanolamine N-methyltransferase]. Inhibitor of TH, a rate-limiting enzyme in the catecholamine-biosynthesis pathway, reduced the actions of nicotine on cell proliferation and adrenaline production. Expression of {alpha}7-nicotinic acetylcholine receptor ({alpha}7-nAChR) was demonstrated in HT-29 cells. Methyllycaconitine, an {alpha}7-nAChR antagonist, reversed the stimulatory actions of nicotine on cell proliferation, TH and D{beta}H expression as well as adrenaline production. Taken together, through the action on {alpha}7-nAChR nicotine stimulates HT-29 cell proliferation via the upregulation of the catecholamine-synthesis pathway and ultimately adrenaline production and {beta}-adrenergic activation. These data reveal the contributory role {alpha}7-nAChR and {beta}-adrenoceptors in the tumorigenesis of colon cancer cells and partly elucidate the carcinogenic action of cigarette smoke on colon cancer.

  20. Direct muscarinic and nicotinic receptor-mediated excitation of rat medial vestibular nucleus neurons in vitro

    NASA Technical Reports Server (NTRS)

    Phelan, K. D.; Gallagher, J. P.

    1992-01-01

    We have utilized intracellular recording techniques to investigate the cholinoceptivity of rat medial vestibular nucleus (MVN) neurons in a submerged brain slice preparation. Exogenous application of the mixed cholinergic agonists, acetylcholine (ACh) or carbachol (CCh), produced predominantly membrane depolarization, induction of action potential firing, and decreased input resistance. Application of the selective muscarinic receptor agonist muscarine (MUSC), or the selective nicotinic receptor agonists nicotine (NIC) or 1,1-dimethyl-4-phenylpiperazinium (DMPP) also produced membrane depolarizations. The MUSC-induced depolarization was accompanied by decreased conductance, while an increase in conductance appeared to underlie the NIC- and DMPP-induced depolarizations. The muscarinic and nicotinic receptor mediated depolarizations persisted in tetrodotoxin and/or low Ca2+/high Mg2+ containing media, suggesting direct postsynaptic receptor activation. The MUSC-induced depolarization could be reversibly blocked by the selective muscarinic-receptor antagonist, atropine, while the DMPP-induced depolarization could be reversibly suppressed by the selective ganglionic nicotinic-receptor antagonist, mecamylamine. Some neurons exhibited a transient membrane hyperpolarization during the depolarizing response to CCh or MUSC application. This transient inhibition could be reversibly blocked by the gamma-aminobutyric acid (GABA) antagonist, bicuculline, suggesting that the underlying hyperpolarization results indirectly from the endogenous release of GABA acting at GABA receptors. This study confirms the cholinoceptivity of MVN neurons and establishes that individual MVN cells possess muscarinic as well as nicotinic receptors. The data provide support for a prominent role of cholinergic mechanisms in the direct and indirect regulation of the excitability of MVN neurons.

  1. Differential regulation of nicotinic receptor-mediated neurotransmitter release following chronic (-)-nicotine administration.

    PubMed

    Jacobs, Iris; Anderson, David J; Surowy, Carol S; Puttfarcken, Pamela S

    2002-10-01

    The objective of this study was to compare nAChR-mediated neurotransmitter release from slices of rat striatum, frontal cortex and hippocampus following chronic (-)-nicotine (Nic) administration (tartrate salt, 2 mg/kg twice daily for 10 days). Binding studies were also conducted to measure changes in receptor density. Relative to saline-treated animals, the number of nAChRs measured by [(3)H]-cytisine (CYT) binding was significantly increased in all brain regions examined by 15% to 25% following chronic Nic administration. Using a relatively high throughput method to measure neurotransmitter release, we found that Nic, CYT, and (+/-)-epibatidine (EB) evoked similar concentration-dependent striatal [(3)H]-dopamine (DA) and hippocampal [(3)H]-norepinephrine (NE) release from both saline (rank order of potency for [(3)H]-DA: EB>CYT>Nic; pEC(50) values, EB (9 +/- 0.1), CYT (8 +/- 0.13), Nic (7.3 +/- 0.19); rank order potency for [(3)H]-NE: EB>Nic=CYT; pEC(50) values, EB (8 +/- 0.18), Nic (5.5 +/- 0.09), CYT (5.12 +/- 0.1)) -and Nic-treated animals (pEC(50) values [(3)H]-DA, EB (9.5 +/- 0.15), Nic (8 +/- 0.16, CYT (6.6 +/- 0.52); [(3)H]-NE, EB (8.4 +/- 0.23), Nic (5.19 +/- 0.1), CYT (5.18 +/- 0.29)). Although no change in potency was detected between the two treatment groups, the agonist efficacies in both tissues were significantly reduced by approximately 17-54% following chronic Nic administration. In contrast to striatum, treatment with Nic did not affect the maximal [(3)H]-DA response (efficacy) in the frontal cortex. However, as observed in the striatum, no change in agonist potency was observed in the frontal cortex following chronic Nic administration (pEC(50) values, saline; EB (9.2 +/- 0.2), >CYT (6.95 +/- 0.75) = Nic (6.9 +/- 0.16); Nic-treated, EB (9 +/- 0.42)>CYT (6.88 +/- 0.27) = Nic (7.1 +/- 0.17)). Chronic Nic treatment did not significantly affect KCl-evoked [(3)H]-NE release from hippocampus or [(3)H]-DA release from frontal cortex or striatum. Since

  2. Nicotinic acetylcholine receptors regulate type 1 inositol 1,4,5-trisphosphate receptor expression via calmodulin kinase IV activation.

    PubMed

    Mizuno, Koji; Kurokawa, Kazuhiro; Ohkuma, Seitaro

    2015-04-01

    Type 1 inositol 1,4,5-trisphosphate receptors (IP3 R-1) are among the important calcium channels regulating intracellular Ca(2+) concentration in the central nervous system. In a previous study, we showed that drugs of abuse, such as cocaine, methamphetamine, and ethanol, induced IP3 R-1 upregulation via the calcium signal transduction pathway in psychological dependence. Although nicotine, a major component in tobacco smoke, participates in psychological and/or physical dependence, it has not yet been clarified how nicotine alters IP3 R-1 expression. The present study, therefore, seeks to clarify the mechanism bgy which nicotine modifies IP3 R-1 expression by using mouse cerebral cortical neurons in primary culture. Nicotine induced dose- and time-dependent upregulation of IP3 R-1 protein following its mRNA increase, and the latter was significantly suppressed by a nonselective nicotinic acetylcholine receptors (nAChR) antagonist, mecamylamine. Both cFos and phosphorylated-cJun (p-cJun) were immediately increased in the nucleus, together with an increase of calmodulin kinase (CaMK) IV but not CaMKII expression after nicotine exposure. A nonselective inhibitor of CaMKs, KN-93, and a calcium chelating regent, BAPTA-AM, completely suppressed the expression of cFos and p-cJun in the nucleus as well as the nicotine-induced IP3 R-1 upregulation. These results indicate that nAChR activation by nicotine upregulates IP3 R-1 via increase of activator protein-1, which is a cFos and cJun dimmer, in the nucleus, with activation of Ca(2+) signaling transduction processes. PMID:25430056

  3. Functional Expression of Two Neuronal Nicotinic Acetylcholine Receptors from cDNA Clones Identifies a Gene Family

    NASA Astrophysics Data System (ADS)

    Boulter, Jim; Connolly, John; Deneris, Evan; Goldman, Dan; Heinemann, Steven; Patrick, Jim

    1987-11-01

    A family of genes coding for proteins homologous to the α subunit of the muscle nicotinic acetylcholine receptor has been identified in the rat genome. These genes are transcribed in the central and peripheral nervous systems in areas known to contain functional nicotinic receptors. In this paper, we demonstrate that three of these genes, which we call alpha3, alpha4, and beta2, encode proteins that form functional nicotinic acetylcholine receptors when expressed in Xenopus oocytes. Oocytes expressing either alpha3 or alpha4 protein in combination with the beta2 protein produced a strong response to acetylcholine. Oocytes expressing only the alpha4 protein gave a weak response to acetylcholine. These receptors are activated by acetylcholine and nicotine and are blocked by Bungarus toxin 3.1. They are not blocked by α -bungarotoxin, which blocks the muscle nicotinic acetylcholine receptor. Thus, the receptors formed by the alpha3, alpha4, and beta2 subunits are pharmacologically similar to the ganglionic-type neuronal nicotinic acetylcholine receptor. These results indicate that the alpha3, alpha4, and beta2 genes encode functional nicotinic acetylcholine receptor subunits that are expressed in the brain and peripheral nervous system.

  4. Local Application of Drugs to Study Nicotinic Acetylcholine Receptor Function in Mouse Brain Slices

    PubMed Central

    Engle, Staci E.; Broderick, Hilary J.; Drenan, Ryan M.

    2012-01-01

    Tobacco use leads to numerous health problems, including cancer, heart disease, emphysema, and stroke. Addiction to cigarette smoking is a prevalent neuropsychiatric disorder that stems from the biophysical and cellular actions of nicotine on nicotinic acetylcholine receptors (nAChRs) throughout the central nervous system. Understanding the various nAChR subtypes that exist in brain areas relevant to nicotine addiction is a major priority. Experiments that employ electrophysiology techniques such as whole-cell patch clamp or two-electrode voltage clamp recordings are useful for pharmacological characterization of nAChRs of interest. Cells expressing nAChRs, such as mammalian tissue culture cells or Xenopus laevis oocytes, are physically isolated and are therefore easily studied using the tools of modern pharmacology. Much progress has been made using these techniques, particularly when the target receptor was already known and ectopic expression was easily achieved. Often, however, it is necessary to study nAChRs in their native environment: in neurons within brain slices acutely harvested from laboratory mice or rats. For example, mice expressing "hypersensitive" nAChR subunits such as α4 L9′A mice 1 and α6 L9′S mice 2, allow for unambiguous identification of neurons based on their functional expression of a specific nAChR subunit. Although whole-cell patch clamp recordings from neurons in brain slices is routinely done by the skilled electrophysiologist, it is challenging to locally apply drugs such as acetylcholine or nicotine to the recorded cell within a brain slice. Dilution of drugs into the superfusate (bath application) is not rapidly reversible, and U-tube systems are not easily adapted to work with brain slices. In this paper, we describe a method for rapidly applying nAChR-activating drugs to neurons recorded in adult mouse brain slices. Standard whole-cell recordings are made from neurons in slices, and a second micropipette filled with a drug of

  5. Nicotinic Acetylcholine Receptors Modulate Bone Marrow-Derived Pro-Inflammatory Monocyte Production and Survival

    PubMed Central

    St-Pierre, Stéphanie; Jiang, Wei; Roy, Patrick; Champigny, Camille; LeBlanc, Éric; Morley, Barbara J.; Hao, Junwei; Simard, Alain R.

    2016-01-01

    It is increasingly clear that nicotinic acetylcholine receptors (nAChRs) are involved in immune regulation, and that their activation can protect against inflammatory diseases. Previous data have shown that nicotine diminishes the numbers of peripheral monocytes and macrophages, especially those of the pro-inflammatory phenotype. The goal of the present study was to determine if nicotine modulates the production of bone marrow -derived monocytes/macrophages. In this study, we first found that murine bone marrow cells express multiple nAChR subunits, and that the α7 and α9 nAChRs most predominant subtypes found in immune cells and their precursors. Using primary cultures of murine bone marrow cells, we then determined the effect of nicotine on monocyte colony-stimulating factor and interferon gamma (IFNγ)-induced monocyte production. We found that nicotine lowered the overall number of monocytes, and more specifically, inhibited the IFNγ-induced increase in pro-inflammatory monocytes by reducing cell proliferation and viability. These data suggested that nicotine diminishes the ratio of pro-inflammatory versus anti-inflammatory monocyte produced in the bone marrow. We thus confirmed this hypothesis by measuring cytokine expression, where we found that nicotine inhibited the production of the pro-inflammatory cytokines TNFα, IL-1β and IL-12, while stimulating the secretion of IL-10, an anti-inflammatory cytokine. Finally, nicotine also reduced the number of pro-inflammatory monocytes in the bone marrow of LPS-challenged mice. Overall, our data demonstrate that both α7 and α9 nAChRs are involved in the regulation of pro-inflammatory M1 monocyte numbers. PMID:26925951

  6. Activation of nicotinic acetylcholine receptors increases the rate of fusion of cultured human myoblasts.

    PubMed Central

    Krause, R M; Hamann, M; Bader, C R; Liu, J H; Baroffio, A; Bernheim, L

    1995-01-01

    1. Fusion of myogenic cells is important for muscle growth and repair. The aim of this study was to examine the possible involvement of nicotinic acetylcholine receptors (nAChR) in the fusion process of myoblasts derived from postnatal human satellite cells. 2. Acetylcholine-activated currents (ACh currents) were characterized in pure preparations of freshly isolated satellite cells, proliferating myoblasts, myoblasts triggered to fuse and myotubes, using whole-cell and single-channel voltage clamp recordings. Also, the effect of cholinergic agonists on myoblast fusion was tested. 3. No nAChR were observed in freshly isolated satellite cells. nAChR were first observed in proliferating myoblasts, but ACh current densities increased markedly only just before fusion. At that time most mononucleated myoblasts had ACh current densities similar to those of myotubes. ACh channels had similar properties at all stages of myoblast maturation. 4. The fraction of myoblasts that did not fuse under fusion-promoting conditions had no ACh current and thus resembled freshly isolated satellite cells. 5. The rate of myoblast fusion was increased by carbachol, an effect antagonized by alpha-bungarotoxin, curare and decamethonium, but not by atropine, indicating that nAChR were involved. Even though a prolonged exposure to carbachol led to desensitization, a residual ACh current persisted after several days of exposure to the nicotinic agonist. 6. Our observations suggest that nAChR play a role in myoblast fusion and that part of this role is mediated by the flow of ions through open ACh channels. Images Figure 1 Figure 2 Figure 3 PMID:8788942

  7. Catharanthine alkaloids are noncompetitive antagonists of muscle-type nicotinic acetylcholine receptors.

    PubMed

    Arias, Hugo R; Feuerbach, Dominik; Targowska-Duda, Katarzyna M; Jozwiak, Krzysztof

    2010-09-01

    We compared the interaction of several catharanthine alkaloids including, ibogaine, vincristine, and vinblastine, with that for the noncompetitive antagonist phencyclidine (PCP) at muscle nicotinic acetylcholine receptors (AChRs) in different conformational states. The results established that catharanthine alkaloids: (a) inhibit, in a noncompetitive manner, (+/-)-epibatidine-induced Ca(2+) influx in TE671-halpha1beta1gammadelta cells with similar potencies (IC(50)=17-25microM), (b) inhibit [(3)H]TCP binding to the desensitized Torpedo AChR with higher affinity compared to the resting AChR, and (c) enhance [(3)H]cytisine binding to resting but activatable Torpedo AChRs, suggesting desensitizing properties. Interestingly, PCP inhibits [(3)H]ibogaine binding to the AChR in a steric fashion. This is corroborated by additional docking experiments indicating that the amino groups of neutral ibogaine form hydrogen bonds with the serine ring (position 6'), a location shared with PCP. Since protonated ibogaine forms a salt bridge with one of the acidic residues at the outer ring (position 20'), this ligand could be first attracted to the entrance of the channel by electrostatic interactions. Our data indicate that the catharanthine moiety is a minimum structural requirement for AChR inhibition including, ion channel blocking and desensitization, and that ibogaine and PCP bind to overlapping sites in the desensitized AChR ion channel. PMID:20493225

  8. Nicotinic receptors in non-human primates: analysis of genetic and functional conservation with humans

    PubMed Central

    Shorey-Kendrick, Lyndsey E.; Ford, Matthew M.; Allen, Daicia C.; Kuryatov, Alexander; Lindstrom, Jon; Wilhelm, Larry; Grant, Kathleen A.; Spindel, Eliot R.

    2015-01-01

    Nicotinic acetylcholine receptors (nAChRs) are highly conserved between humans and non-human primates. Conservation exists at the level of genomic structure, protein structure and epigenetics. Overall homology of nAChRs at the protein level is 98% in macaques versus 89% in mice, which is highly relevant for evaluating subtype-specific ligands that have different affinities in humans versus rodents. In addition to conservation at the protein level, there is high conservation of genomic structure in terms of intron and exon size and placement of CpG sites that play a key role in epigenetic regulation. Analysis of single nucleotide polymorphisms (SNPs) shows that while the majority of SNPs are not conserved between humans and macaques, some functional polymorphisms are. Most significantly, cynomolgus monkeys express a similar α5 nAChR Asp398Asn polymorphism to the human α5 Asp398Asn polymorphism that has been linked to greater nicotine addiction and smoking related disease. Monkeys can be trained to readily self-administer nicotine, and in an initial study we have demonstrated that cynomolgus monkeys bearing the α5 D398N polymorphism show a reduced behavioral sensitivity to oral nicotine and tend to consume it in a different pattern when compared to wild-type monkeys. Thus the combination of highly homologous nAChR, higher cortical functions and capacity for complex training makes non-human primates a unique model to study in vivo functions of nicotinic receptors. In particular, primate studies on nicotine addiction and evaluation of therapies to prevent or overcome nicotine addiction are likely to be highly predictive of treatment outcomes in humans. PMID:25661700

  9. Nicotinic acid-adenine dinucleotide phosphate activates the skeletal muscle ryanodine receptor.

    PubMed Central

    Hohenegger, Martin; Suko, Josef; Gscheidlinger, Regina; Drobny, Helmut; Zidar, Andreas

    2002-01-01

    Calcium is a universal second messenger. The temporal and spatial information that is encoded in Ca(2+)-transients drives processes as diverse as neurotransmitter secretion, axonal outgrowth, immune responses and muscle contraction. Ca(2+)-release from intracellular Ca(2+) stores can be triggered by diffusible second messengers like Ins P (3), cyclic ADP-ribose or nicotinic acid-adenine dinucleotide phosphate (NAADP). A target has not yet been identified for the latter messenger. In the present study we show that nanomolar concentrations of NAADP trigger Ca(2+)-release from skeletal muscle sarcoplasmic reticulum. This was due to a direct action on the Ca(2+)-release channel/ryanodine receptor type-1, since in single channel recordings, NAADP increased the open probability of the purified channel protein. The effects of NAADP on Ca(2+)-release and open probability of the ryanodine receptor occurred over a similar concentration range (EC(50) approximately 30 nM) and were specific because (i) they were blocked by Ruthenium Red and ryanodine, (ii) the precursor of NAADP, NADP, was ineffective at equimolar concentrations, (iii) NAADP did not affect the conductance and reversal potential of the ryanodine receptor. Finally, we also detected an ADP-ribosyl cyclase activity in the sarcoplasmic reticulum fraction of skeletal muscle. This enzyme was not only capable of synthesizing cyclic GDP-ribose but also NAADP, with an activity of 0.25 nmol/mg/min. Thus, we conclude that NAADP is generated in the vicinity of type 1 ryanodine receptor and leads to activation of this ion channel. PMID:12102654

  10. INHIBITORY EFFECTS OF VOLATILE ORGANIC COMPOUNDS ON NEURONAL NICOTINIC ACETYLCHOLINE RECEPTORS.

    EPA Science Inventory

    INHIBITORY EFFECTS OF VOLATILE ORGANIC COMPOUNDS ON NEURONAL NICOTINIC ACETYLCHOLINE RECEPTORS.
    A.S. Bale*; P.J. Bushnell; C.A. Meacham; T.J. Shafer
    Neurotoxicology Division, NHEERL, ORD, US Environmental Protection Agency, Research Triangle Park, NC, USA
    Toluene (TOL...

  11. Effect of a nicotinic acetylcholine receptor agonists and antagonists on motor function in mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nicotinic acetylcholine receptors (nAChR) are ligand-gated cation channels found throughout the body, and serve to mediate diverse physiological functions. Muscle-type nAChR located in the motor endplate region of muscle fibers play an integral role in muscle contraction and thus motor function. The...

  12. Positive Modulation of GABAB Receptors Decreased Nicotine Self-administration and Counteracted Nicotine-induced Enhancement of Brain Reward Function in Rats

    PubMed Central

    Paterson, Neil E.; Vlachou, Styliani; Guery, Sebastien; Kaupmann, Klemens; Froestl, Wolfgang; Markou, Athina

    2008-01-01

    Acute administration of γ-aminobutyric acid (GABA)-B receptor agonists decreases nicotine, cocaine, ethanol, and heroin self-administration, and also decreases food-maintained responding and suppresses locomotor activity at high doses. GABAB receptor positive modulators may represent potentially improved therapeutic compounds because of their fewer side-effects than receptor agonists. The present study investigated the effects of administration of the GABAB receptor positive modulators 2,6-Di-tert-butyl-4-(3-hydroxy-2,2-dimethyl-propyl)-phenol (CGP7930) and N-[(1R,2R,4S)-bicyclo[2.2.1]hept-2-yl]-2-methyl-5-[4-(trifluoromethyl)phenyl]-4-pyrimidinamine (BHF177), and co-administration of the GABAB receptor positive modulator N,N'-dicyclopentyl-2-methylsulfanyl-5-nitro-pyrimidine-4,6-diamine (GS39783) with the GABAB receptor agonist (3-amino-2[S]-hydroxypropyl)-methylphosphinic acid (CGP44532) on nicotine- and food-maintained responding under fixed-ratio 5 (FR5) and progressive-ratio schedules of reinforcement. Furthermore, the effects of BHF177 and CGP44532 on nicotine-induced enhancement of brain reward function were evaluated. The results indicated that administration of CGP7930 decreased nicotine self-administration under an FR5 schedule. Administration of either GS39783 or CGP44532 selectively decreased nicotine self-administration, while co-administration of these compounds had additive effects. BHF177 administration selectively decreased nicotine-, but not food-, maintained responding under FR5 and progressive-ratio schedules. The nicotine-induced enhancement of brain reward function was blocked by BHF177 or CGP44532, although the highest doses of both compounds, particularly CGP44532, decreased brain reward function when administered alone, suggesting an additive, rather than interactive, effect. Overall, the present results indicate that GABAB receptor positive modulators, similarly to GABAB receptor agonists, attenuated the reinforcing and reward

  13. Prenatal nicotine-exposure alters fetal autonomic activity and medullary neurotransmitter receptors: implications for sudden infant death syndrome.

    PubMed

    Duncan, Jhodie R; Garland, Marianne; Myers, Michael M; Fifer, William P; Yang, May; Kinney, Hannah C; Stark, Raymond I

    2009-11-01

    During pregnancy, exposure to nicotine and other compounds in cigarette smoke increases the risk of the sudden infant death syndrome (SIDS) two- to fivefold. Serotonergic (5-HT) abnormalities are found, in infants who die of SIDS, in regions of the medulla oblongata known to modulate cardiorespiratory function. Using a baboon model, we tested the hypothesis that prenatal exposure to nicotine alters 5-HT receptor and/or transporter binding in the fetal medullary 5-HT system in association with cardiorespiratory dysfunction. At 87 (mean) days gestation (dg), mothers were continuously infused with saline (n = 5) or nicotine (n = 5) at 0.5 mg/h. Fetuses were surgically instrumented at 129 dg for cardiorespiratory monitoring. Cesarean section delivery and retrieval of fetal medulla were performed at 161 (mean) dg for autoradiographic analyses of nicotinic and 5-HT receptor and transporter binding. In nicotine-exposed fetuses, high-frequency heart rate variability was increased 55%, possibly reflecting increases in the parasympathetic control of heart rate. This effect was more pronounced with greater levels of fetal breathing and age. These changes in heart rate variability were associated with increased 5-HT(1A) receptor binding in the raphé obscurus (P = 0.04) and increased nicotinic receptor binding in the raphé obscurus and vagal complex (P < 0.05) in the nicotine-exposed animals compared with controls (n = 6). The shift in autonomic balance in the fetal primate toward parasympathetic predominance with chronic exposure to nicotine may be related, in part, to abnormal 5-HT-nicotine alterations in the raphé obscurus. Thus increased risk for SIDS due to maternal smoking may be partly related to the effects of nicotine on 5-HT and/or nicotinic receptors. PMID:19729586

  14. What is the effect of nicotinic acetylcholine receptor stimulation on osteoarthritis in a rodent animal model?

    PubMed Central

    Bock, Kilian; Plaass, Christian; Coger, Vincent; Peck, Claas-Tido; Reimers, Kerstin; Stukenborg-Colsman, Christina; Claassen, Leif

    2016-01-01

    Objectives: Despite the rising number of patients with osteoarthritis, no sufficient chondroprotective and prophylactic therapy for osteoarthritis has been established yet. The purpose of this study was to verify whether stimulation of the nicotinic acetylcholine receptor via nicotine has a beneficial effect on cartilage degeneration in the development of osteoarthritis and is capable of reducing the expression of proinflammatory cytokines and cartilage degrading enzymes in synovial membranes after osteoarthritis induction. Methods: Experimental osteoarthritis was induced in Lewis rats using a standardized osteoarthritis model with monoiodoacetate. A total of 16 Lewis rats were randomized into four groups: control, sham + nicotine application, osteoarthritis, and osteoarthritis + nicotine application. Nicotine (0.625 mg/kg twice daily) was administered intraperitoneally for 42 days. We analyzed histological sections, radiological images and the expression of the proinflammatory cytokines, such as interleukin-1β, tumor necrosis factor-α and interleukin-6, and of matrix metalloproteases 3, 9 and 13 and tissue inhibitors of metalloprotease-1 in synovial membranes via quantitative polymerase chain reaction. Results: Histological and x-ray examination revealed cartilage degeneration in the osteoarthritis group compared to control or sham + nicotine groups (histological control vs osteoarthritis: p = 0.002 and x-ray control vs osteoarthritis: p = 0.004). Nicotine treatment reduced the cartilage degeneration without significant differences. Osteoarthritis induction led to a higher expression of proinflammatory cytokines and matrix metalloproteases as compared to control groups. This effect was attenuated after nicotine administration. The differences of proinflammatory cytokines and matrix metalloproteases did not reach statistical significance. Conclusion: With the present small-scale study, we could not prove a positive effect of nicotinic

  15. Nicotine alters limbic function in adolescent rat by a 5-HT1A receptor mechanism.

    PubMed

    Dao, Jasmin M; McQuown, Susan C; Loughlin, Sandra E; Belluzzi, James D; Leslie, Frances M

    2011-06-01

    Epidemiological studies have shown that adolescent smoking is associated with health risk behaviors, including high-risk sexual activity and illicit drug use. Using rat as an animal model, we evaluated the behavioral and biochemical effects of a 4-day, low-dose nicotine pretreatment (60 μg/kg; intravenous) during adolescence and adulthood. Nicotine pretreatment significantly increased initial acquisition of cocaine self-administration, quinpirole-induced locomotor activity, and penile erection in adolescent rats, aged postnatal day (P)32. These effects were long lasting, remaining evident 10 days after the last nicotine treatment, and were observed when nicotine pretreatment was administered during early adolescence (P28-31), but not late adolescence (P38-41) or adulthood (P86-89). Neurochemical analyses of c-fos mRNA expression, and of monoamine transmitter and transporter levels, showed that forebrain limbic systems are continuing to develop during early adolescence, and that this maturation is critically altered by brief nicotine exposure. Nicotine selectively increased c-fos mRNA expression in the nucleus accumbens shell and basolateral amygdala in adolescent, but not adult animals, and altered serotonin markers in these regions as well as the prefrontal cortex. Nicotine enhancement of cocaine self-administration and quinpirole-induced locomotor activity was blocked by co-administration of WAY 100 635 (N-{2-[4-(2-methoxyphenyl)-1-piperazinyl] ethyl}-N-(2-pyridinyl)cyclohexanecarboxamide), a selective serotonin 1A (5-HT1A) receptor antagonist. Early adolescent pretreatment with the mixed autoreceptor/heteroceptor 5-HT1A receptor agonist, 8-OH-DPAT, but not the autoreceptor-selective agonist, S-15535, also enhanced quinpirole-induced locomotor activation. Nicotine enhancement of quinpirole-induced penile erection was not blocked by WAY 100 635 nor mimicked by 8-OH-DPAT. These findings indicate that early adolescent nicotine exposure uniquely alters limbic

  16. Nicotine enhances murine airway contractile responses to kinin receptor agonists via activation of JNK- and PDE4-related intracellular pathways

    PubMed Central

    2010-01-01

    Background Nicotine plays an important role in cigarette-smoke-associated airway disease. The present study was designed to examine if nicotine could induce airway hyperresponsiveness through kinin receptors, and if so, explore the underlying mechanisms involved. Methods Murine tracheal segments were cultured for 1, 2 or 4 days in serum-free DMEM medium in presence of nicotine (1 and 10 μM) or vehicle (DMSO). Contractile responses induced by kinin B1 receptor agonist, des-Arg9-bradykinin, and B2 receptor agonist, bradykinin, were monitored with myographs. The B1 and B2 receptor mRNA expressions were semi-quantified using real-time PCR and their corresponding protein expressions assessed with confocal-microscopy-based immunohistochemistry. Various pharmacological inhibitors were used for studying intracellular signaling pathways. Results Four days of organ culture with nicotine concentration-dependently increased kinin B1 and B2 receptor-mediated airway contractions, without altering the kinin receptor-mediated relaxations. No such increase was seen at day 1 or day 2. The airway contractile responses to 5-HT, acetylcholine and endothelin receptor agonists remained unaffected by nicotine. Two different neuronal nicotinic receptor antagonists MG624 and hexamethonium blocked the nicotine-induced effects. The enhanced contractile responses were accompanied by increased mRNA and protein expression for both kinin receptors, suggesting the involvement of transcriptional mechanisms. Confocal-microscopy-based immunohistochemistry showed that 4 days of nicotine treatment induced activation (phosphorylation) of c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase 1 and 2 (ERK1/2) and p38. Inhibition of JNK with its specific inhibitor SP600125 abolished the nicotine-induced effects on kinin receptor-mediated contractions and reverted the enhanced receptor mRNA expression. Administration of phosphodiesterase inhibitors (YM976 and theophylline

  17. Exposure to nicotine increases nicotinic acetylcholine receptor density in the reward pathway and binge ethanol consumption in C57BL/6J adolescent female mice.

    PubMed

    Locker, Alicia R; Marks, Michael J; Kamens, Helen M; Klein, Laura Cousino

    2016-05-01

    Nearly 80% of adult smokers begin smoking during adolescence. Binge alcohol consumption is also common during adolescence. Past studies report that nicotine and ethanol activate dopamine neurons in the reward pathway and may increase synaptic levels of dopamine in the nucleus accumbens through nicotinic acetylcholine receptor (nAChR) stimulation. Activation of the reward pathway during adolescence through drug use may produce neural alterations affecting subsequent drug consumption. Consequently, the effect of nicotine exposure on binge alcohol consumption was examined along with an assessment of the neurobiological underpinnings that drive adolescent use of these drugs. Adolescent C57BL/6J mice (postnatal days 35-44) were exposed to either water or nicotine (200μg/ml) for ten days. On the final four days, ethanol intake was examined using the drinking-in-the-dark paradigm. Nicotine-exposed mice consumed significantly more ethanol and displayed higher blood ethanol concentrations than did control mice. Autoradiographic analysis of nAChR density revealed higher epibatidine binding in frontal cortical regions in mice exposed to nicotine and ethanol compared to mice exposed to ethanol only. These data show that nicotine exposure during adolescence increases subsequent binge ethanol consumption, and may affect the number of nAChRs in regions of the brain reward pathway, specifically the frontal cortex. PMID:26428091

  18. Pharmacological Beta-Adrenergic Receptor Activation Attenuates Neutrophil Recruitment by a Mechanism Dependent on Nicotinic Receptor and the Spleen.

    PubMed

    Silva, Rangel L; Castanheira, Fernanda V; Figueiredo, Jozi G; Bassi, Gabriel S; Ferreira, Sérgio H; Cunha, Fernando Q; Cunha, Thiago M; Kanashiro, Alexandre

    2016-08-01

    The aim of this study was to identify the effect of beta-adrenergic receptor activation on neutrophil migration in experimental peritonitis elucidating the neuroimmune components involved such as nicotinic receptors and the spleen. Mice pre-treated with mecamylamine (nicotinic antagonist) and propranolol (beta-adrenergic antagonist) or splenectomized animals were treated with isoproterenol (beta-adrenergic agonist) prior to intraperitoneal injection of carrageenan. After 4 h, the infiltrating neutrophils and the local cytokine/chemokine levels were evaluated in the peritoneal lavage. The effect of isoproterenol on neutrophil chemotaxis was investigated in a Boyden chamber. Isoproterenol inhibited neutrophil trafficking, reducing the cytokine/chemokine release and neutrophil chemotaxis. Surprisingly, the isoproterenol effect on neutrophil migration was totally reverted by splenectomy and mecamylamine pre-treatment. In contrast, the inhibitory effect of nicotine on neutrophil migration was abrogated only by splenectomy but not by propranolol pre-treatment. Collectively, our data show that beta-adrenergic receptor activation regulates the acute neutrophil recruitment via splenic nicotinic receptor. PMID:27262431

  19. Structural features of phenoxycarbonylimino neonicotinoids acting at the insect nicotinic receptor.

    PubMed

    Ohno, Ikuya; Tomizawa, Motohiro; Miyazu, Nozomi; Kushibiki, Gohito; Noda, Kumiko; Hasebe, Yasunori; Durkin, Kathleen A; Miyake, Taiji; Kagabu, Shinzo

    2010-10-01

    Substituted-phenoxycarbonylimino neonicotinoid ligands with an electron-donating group showed significantly higher affinity to the insect nicotinic receptor relative to that of the analogue with an electron-withdrawing substituent, thereby establishing in silico binding site interaction model featuring that the phenoxy ring of neonicotinoids and the receptor loop D tryptophan indole plane form a face-to-edge aromatic interaction. PMID:20729079

  20. Interaction of ibogaine with human alpha3beta4-nicotinic acetylcholine receptors in different conformational states.

    PubMed

    Arias, Hugo R; Rosenberg, Avraham; Targowska-Duda, Katarzyna M; Feuerbach, Dominik; Yuan, Xiao Juan; Jozwiak, Krzysztof; Moaddel, Ruin; Wainer, Irving W

    2010-09-01

    The interaction of ibogaine and phencyclidine (PCP) with human (h) alpha3beta4-nicotinic acetylcholine receptors (AChRs) in different conformational states was determined by functional and structural approaches including, radioligand binding assays, Ca2+ influx detections, and thermodynamic and kinetics measurements. The results established that (a) ibogaine inhibits (+/-)-epibatidine-induced Ca2+ influx in h(alpha)3beta4 AChRs with approximately 9-fold higher potency than that for PCP, (b) [3H]ibogaine binds to a single site in the h(alpha)3beta4 AChR ion channel with relatively high affinity (Kd = 0.46 +/- 0.06 microM), and ibogaine inhibits [3H]ibogaine binding to the desensitized h(alpha)3beta4 AChR with slightly higher affinity compared to the resting AChR. This is explained by a slower dissociation rate from the desensitized ion channel compared to the resting ion channel, and (c) PCP inhibits [3H]ibogaine binding to the h(alpha)3beta4 AChR, suggesting overlapping sites. The experimental results correlate with the docking simulations suggesting that ibogaine and PCP interact with a binding domain located between the serine (position 6') and valine/phenylalanine (position 13') rings. This interaction is mediated mainly by van der Waals contacts, which is in agreement with the observed enthalpic contribution determined by non-linear chromatography. However, the calculated entropic contribution also indicates local conformational changes. Collectively our data suggest that ibogaine and PCP bind to overlapping sites located between the serine and valine/phenylalanine rings, to finally block the AChR ion channel, and in the case of ibogaine, to probably maintain the AChR in the desensitized state for longer time. PMID:20684041

  1. Alteration in contractile G-protein coupled receptor expression by moist snus and nicotine in rat cerebral arteries

    SciTech Connect

    Sandhu, Hardip; Xu Cangbao; Edvinsson, Lars

    2011-04-15

    The cardiovascular risk for users of use of Swedish snus/American snuff (moist tobacco) has been debated for a long time. The present study was designed to examine the effects of water- or lipid-soluble (DMSO-soluble) snus and nicotine, the most important substance in tobacco, on the expression of vasocontractile G-protein coupled receptors (GPCR), such as endothelin ET{sub B}, serotonin 5-HT{sub 1B}, and thromboxane A{sub 2} TP receptors, in rat cerebral arteries. Studies show that these vasocontractile GPCR show alterations by lipid-soluble cigarette smoke particles via activation of mitogen-activated protein kinases (MAPK). However, the effects of moist tobacco on the expression of GPCR are less studied. Rat middle cerebral arteries were isolated and organ cultured in serum-free medium for 24 h in the presence of water-soluble snus (WSS), DMSO-soluble snus (DSS), or nicotine. The dose of snus and nicotine was kept at plasma level of snus users (25 ng nicotine/ml). A high dose (250 ng nicotine/ml) was also included due to the previous results showing alteration in the GPCR expression by nicotine at this concentration. Contractile responses to the ET{sub B} receptor agonist sarafotoxin 6c, 5-HT{sub 1B} receptor agonist 5-carboxamidotryptamine, and TP receptor agonist U46619 were investigated by a sensitive myograph. The expression of ET{sub B}, 5-HT{sub 1B}, and TP receptors was studied at mRNA and protein levels using quantitative real-time PCR and immunohistochemistry, respectively. Organ culture with WSS or DSS (25 ng nicotine/ml) lowered the 5-HT{sub 1B} receptor-mediated contraction. Furthermore, DSS shifted the TP receptor-mediated contraction curve left-wards with a stronger contraction. High dose of nicotine (250 ng nicotine/ml) increased the ET{sub B} receptor-mediated contraction. The combined 5-HT{sub 1B} and 5-HT{sub 2A} receptor-mediated contraction was increased, and both the 5-CT and TxA2 induced contractions were left-ward shifted by WSS, DSS, or

  2. Subunit composition of α5-containing nicotinic receptors in the rodent habenula

    PubMed Central

    Scholze, Petra; Koth, Gabriele; Orr-Urtreger, Avi; Huck, Sigismund

    2012-01-01

    Gene association studies in humans have linked the α5 subunit gene CHRNA5 to an increased risk for nicotine dependence. In the CNS, nicotinic acetylcholine receptors (nAChRs) that contain the α5 subunit are expressed at relatively high levels in the habenulo-interpeduncular system. Recent experimental evidence furthermore suggests that α5-containing receptors in the habenula play a key role in controlling the intake of nicotine in rodents. We have now analysed the subunit composition of hetero-oligomeric nAChRs in the habenula of postnatal day 18 (P18) C57Bl/6J control mice and of mice with deletions of the α5, the β2, or the β4 subunit genes. Receptors consisting of α3β4*1 clearly outnumbered α4β2*-containing receptors not only in P18 but also in adult mice. We found low levels of α5-containing receptors in both mice (6%) and rats (2.5% of overall nAChRs). Observations in β2 and β4 null mice indicate that although α5 requires the presence of the β4 subunit for assembling (but not of β2), α5 in wild-type mice assembles into receptors that also contain the subunits α3, β2, and β4. PMID:22380605

  3. Primary structure of nicotinic acetylcholine receptor. Final report, 9 April 1989-6 April 1992

    SciTech Connect

    Patrick, J.W.

    1992-05-06

    Signals are transmitted between cells in the brain using neurotransmitters and neurotransmitter receptors. Poisons that interfere with this process stop normal brain function and often kill nerve cells. One of the neurotransmitters used in the mammalian brain is acetylcholine. We discovered that there is a large number of different nicotinic receptors for the neurotransmitter acetylcholine, each with its different properties. We used recombinant DNA technology to clone and sequence the gene transcripts that encode the subunits of these receptors. From these sequences we deduced the primary structures of the nicotinic receptor subunits. We also used the cDNA clones to determine which brain loci express the respective genes. We have expressed the clones in the Xenopus oocyte and have demonstrated that each functional combination of subunits has a unique pharmacology Unlike their homologs at the neuromuscular junction, the nicotinic acetylcholine receptors in the brain are exceptionally permeable to calcium. This property suggests that these receptors may play an important role in regulating calcium-dependent cytoplasmic processes and that they may be important contributors to use-dependent cell death.

  4. Interaction of nicotinic receptor affinity reagents with central nervous system. cap alpha. -bungarotoxin-binding entities

    SciTech Connect

    Lukas, R.J.; Bennett, E.L.

    1980-01-01

    Membrane-bound ..cap alpha..-bungarotoxin-binding entities derived from rat brain are found to interact specifically with the affinity reagents maleimidobenzyltrimethylammonium (MBTA) and bromoacetylcholine (BAC), originally designed to label nicotinic acetylcholine receptors from electroplax and skeletal muscle. Following treatment of membranes with dithiothreitol, all specific toxin binding sites are irreversibly blocked by reaction with MBTA or BAC. Affinity reagent labeling of dithiothreitol-reduced membranes is prevented (toxin binding sites are not blocked) by prior alkylaction with N-ethylmaleimide, by prior oxidation with dithiobis(2-nitrobenzoic acid), or by incubation with neurotoxin. Reversibly associating cholinergic agonists and antagonists retard the rate of affinity reagent interaction with toxin receptors. The apparent rates of affinity reagent alkylation of toxin receptors, and the influences of other sulfhydryl/disulfide reagents on affinity labeling are comparable to those observed for reaction with nicotinic acetylcholine receptors in the periphery. The results provide further evidence that central nervous system ..cap alpha..-bungarotoxin receptors share a remarkable number of biochemical properties with nicotinic receptors from the periphery.

  5. Functional Contribution of α3L8′ to the Neuronal Nicotinic α3 Receptor

    PubMed Central

    Nieves-Cintrón, Madeline; Caballero-Rivera, Daniel; Silva, Walter I.; Navedo, Manuel F.; Lasalde-Dominicci, José A.

    2015-01-01

    The role of position L8′, located in transmembrane domain 1 of the neuronal nicotinic α3 subunit, was characterized by using two-electrode voltage clamp in Xenopus oocytes. Four amino acids (Ala, Ser, Phe, and Tyr) were inserted at this conserved position, and the mutant subunit was coexpressed with either wild-type β2 or β4 subunits. These substitutions led to significant alterations in the pharmacodynamic parameters of cholinergic agents, resulting in loss of function. Ala and Ser substitutions resulted in losses in agonist (ACh, nicotine, and DMPP) potency and intrinsic activity at both α3β2 and α3β4 receptors. Similarly, significant changes in antagonist potency were produced by the Ala and Ser substitutions. Phe and Tyr mutations did not alter the receptor's EC50 for ACh or nicotine but reduced the EC50 for DMPP at both receptors. The Phe mutation also reduced the intrinsic activity of all agonists tested at both receptors. The Tyr mutation, though, led to a decrease in intrinsic activity for all agonists at the α3β2 receptor, yet resulted in no changes for DMPP, a decrease for nicotine, and an increase for ACh at the α3β4 receptor. The most dramatic changes in the receptor's functional properties were produced by substitutions that introduced the largest changes in amino acid volume. Additional replacements (Gly, Thr, and Val) suggested an inverse correlation between amino acid volume at position α3L8′ and EC50 for α3β4 nAChRs; however, α3β2 nAChRs displayed a nonlinear correlation. These data demonstrate that structural alterations at position α3L8′ could propagate to the agonist-binding site. PMID:18615639

  6. Nicotinic receptor involvement in regulation of functions of mouse neutrophils from inflammatory site.

    PubMed

    Safronova, Valentina G; Vulfius, Catherine A; Shelukhina, Irina V; Mal'tseva, Valentina N; Berezhnov, Alexey V; Fedotova, Eugeniya I; Miftahova, Regina G; Kryukova, Elena V; Grinevich, Andrey A; Tsetlin, Victor I

    2016-07-01

    Participation of nicotinic acetylcholine receptors (nAChRs) in functioning of polymorphonuclear neutrophils (PMNs) isolated from inflammatory site of mice and expression of different nAChR subunits were studied. Nicotine and acetylcholine (ACh) modified respiratory burst induced by a chemotactic peptide N-formyl-MLF in neutrophils of male (but not female) mice. Antagonists of nAChRs α-cobratoxin (αCTX), α-conotoxins MII and [A10L]PnIA at concentrations of 0.01-5μM, 0.2μM and 1μM, respectively, eliminated nAChR agonist effects. ACh also affected adhesion of PMNs, this effect was also prevented by αCTX (100nM) and MII (1nM). Neutrophils of female mice after chronic nicotine consumption acquired sensitivity to nAChR agonists. Changes of free intracellular Ca(2+) concentration in neutrophils under the action of nAChR ligands were analyzed. In cells with no Ca(2+) oscillations and relatively low resting level of intracellular Ca(2+), nicotine triggered Ca(2+)-spikes, the lag of the response shortened with increasing nicotine concentration. A nicotinic antagonist caramiphen strongly decreased the effect of nicotine. RT-PCR analysis revealed mRNAs of α2, α3, α4, α5, α6, α7, α9, β2, β3, and β4 nAChR subunits. Specific binding of [(125)I]-α-bungarotoxin was demonstrated. Thus in view of the effects and binding characteristics the results obtained suggest a regulatory role of α7, α3β2 or α6* nAChR types in specific functions of PMNs. PMID:26965141

  7. Suitability of Nicotinic Acetylcholine Receptor α7 and Muscarinic Acetylcholine Receptor 3 Antibodies for Immune Detection

    PubMed Central

    Rommel, Frank R.; Raghavan, Badrinarayanan; Paddenberg, Renate; Kummer, Wolfgang; Tumala, Susanne; Lochnit, Günter; Gieler, Uwe

    2015-01-01

    Recent evidence reveals a crucial role for acetylcholine and its receptors in the regulation of inflammation, particularly of nicotinic acetylcholine receptor α7 (Chrna7) and muscarinic acetylcholine receptor 3 (Chrm3). Immunohistochemistry is a key tool for their cellular localization in functional tissues. We evaluated nine different commercially available antibodies on back skin tissue from wild-type (Wt) and gene-deficient (KO) mice. In the immunohistochemical analysis, we focused on key AChR-ligand sensitive skin cells (mast cells, nerve fibers and keratinocytes). All five antibodies tested for Chrm3 and the first three Chrna7 antibodies stained positive in both Wt and respective KO skin. With the 4th antibody (ab23832) nerve fibers were unlabeled in the KO mice. By western blot analysis, this antibody detected bands in both Wt and Chrna7 KO skin and brain. qRT-PCR revealed mRNA amplification with a primer set for the undeleted region in both Wt and KO mice, but none with a primer set for the deleted region in KO mice. By 2D electrophoresis, we found β-actin and β-enolase cross reactivity, which was confirmed by double immunolabeling. In view of the present results, the tested antibodies are not suitable for immunolocalization in skin and suggest thorough control of antibody specificity is required if histomorphometry is intended. PMID:25673288

  8. The Nicotinic Receptor Alpha7 Impacts the Mouse Lung Response to LPS through Multiple Mechanisms

    PubMed Central

    Enioutina, Elena Y.; Myers, Elizabeth J.; Tvrdik, Petr; Hoidal, John R.; Rogers, Scott W.; Gahring, Lorise C.

    2015-01-01

    The nicotinic acetylcholine receptor alpha7 (α7) is expressed by neuronal and non-neuronal cells throughout the body. We examined the mechanisms of the lung inflammatory response to intranasal (i.n.) lipopolysaccharide (LPS) regulated by α7. This was done in mice using homologous recombination to introduce a point mutation in the α7 receptor that replaces the glutamate residue 260 that lines the pore with alanine (α7E260A), which has been implicated in controlling the exceptional calcium ion conductance of this receptor. The α7E260A mice exhibit normal inflammatory cell recruitment to the blood in response to i.n. LPS administration. This differs from the α7knock-out (α7KO) in which upstream signaling to initiate the recruitment to the blood following i.n. LPS is significantly impaired. While hematopoietic cells are recruited to the bloodstream in the α7E260A mouse, they fail to be recruited efficiently into both the interstitium and alveolar spaces of the lung. Bone marrow reconstitution experiments demonstrate that the responsiveness of both CD45+ and CD45- cells of the α7E260A mouse are impaired. The expression of several pro-inflammatory cytokine and chemokine RNAs including TNFα, IL-1α, Ccl2 and Cxcl10 are decreased in the α7E260A mouse. However, there is a substantial increase in IL-13 expression by CD45- lung interstitial cells in the α7E260A mouse. Our results support the conclusion that α7 functional pleiotropy contributes to modulating the tissue response to an inflammatory insult through impacting upon a variety of mechanisms reflecting the individual cell composition of the lung. PMID:25803612

  9. Phosphocholine - an agonist of metabotropic but not of ionotropic functions of α9-containing nicotinic acetylcholine receptors.

    PubMed

    Richter, K; Mathes, V; Fronius, M; Althaus, M; Hecker, A; Krasteva-Christ, G; Padberg, W; Hone, A J; McIntosh, J M; Zakrzewicz, A; Grau, V

    2016-01-01

    We demonstrated previously that phosphocholine and phosphocholine-modified macromolecules efficiently inhibit ATP-dependent release of interleukin-1β from human and murine monocytes by a mechanism involving nicotinic acetylcholine receptors (nAChR). Interleukin-1β is a potent pro-inflammatory cytokine of innate immunity that plays pivotal roles in host defence. Control of interleukin-1β release is vital as excessively high systemic levels cause life threatening inflammatory diseases. In spite of its structural similarity to acetylcholine, there are no other reports on interactions of phosphocholine with nAChR. In this study, we demonstrate that phosphocholine inhibits ion-channel function of ATP receptor P2X7 in monocytic cells via nAChR containing α9 and α10 subunits. In stark contrast to choline, phosphocholine does not evoke ion current responses in Xenopus laevis oocytes, which heterologously express functional homomeric nAChR composed of α9 subunits or heteromeric receptors containing α9 and α10 subunits. Preincubation of these oocytes with phosphocholine, however, attenuated choline-induced ion current changes, suggesting that phosphocholine may act as a silent agonist. We conclude that phophocholine activates immuno-modulatory nAChR expressed by monocytes but does not stimulate canonical ionotropic receptor functions. PMID:27349288

  10. Phosphocholine – an agonist of metabotropic but not of ionotropic functions of α9-containing nicotinic acetylcholine receptors

    PubMed Central

    Richter, K.; Mathes, V.; Fronius, M.; Althaus, M.; Hecker, A.; Krasteva-Christ, G.; Padberg, W.; Hone, A. J.; McIntosh, J. M.; Zakrzewicz, A.; Grau, V.

    2016-01-01

    We demonstrated previously that phosphocholine and phosphocholine-modified macromolecules efficiently inhibit ATP-dependent release of interleukin-1β from human and murine monocytes by a mechanism involving nicotinic acetylcholine receptors (nAChR). Interleukin-1β is a potent pro-inflammatory cytokine of innate immunity that plays pivotal roles in host defence. Control of interleukin-1β release is vital as excessively high systemic levels cause life threatening inflammatory diseases. In spite of its structural similarity to acetylcholine, there are no other reports on interactions of phosphocholine with nAChR. In this study, we demonstrate that phosphocholine inhibits ion-channel function of ATP receptor P2X7 in monocytic cells via nAChR containing α9 and α10 subunits. In stark contrast to choline, phosphocholine does not evoke ion current responses in Xenopus laevis oocytes, which heterologously express functional homomeric nAChR composed of α9 subunits or heteromeric receptors containing α9 and α10 subunits. Preincubation of these oocytes with phosphocholine, however, attenuated choline-induced ion current changes, suggesting that phosphocholine may act as a silent agonist. We conclude that phophocholine activates immuno-modulatory nAChR expressed by monocytes but does not stimulate canonical ionotropic receptor functions. PMID:27349288

  11. Nicotine modulation of adolescent dopamine receptor signaling and hypothalamic peptide response.

    PubMed

    Mojica, Celina Y; Dao, Jasmin M; Yuan, Menglu; Loughlin, Sandra E; Leslie, Frances M

    2014-02-01

    Adolescence is a sensitive developmental period for limbic and dopamine systems that coincides with the typical age for onset of tobacco use. We have previously shown that a 4-day, low-dose nicotine (0.06 mg/kg) pretreatment enhances locomotor and penile response to the D2-like agonist, quinpirole (0.4 mg/kg), in adolescent but not adult rats. The present study is designed to determine mechanisms underlying this effect. Nicotine enhancement of adolescent quinpirole-induced locomotion was mediated by D2 receptors (D2Rs) since it was blocked by the D2R antagonist, L-741,626, but not by the D3R and D4R antagonists, NGB 2904 and L-745,870. Enhancement of quinpirole-induced erectile response was blocked by both L-741,626 and NGB 2904, indicating involvement of D3Rs. Whereas D2R binding was unaffected by adolescent nicotine pretreatment, effector coupling in the striatum was increased, as determined by GTPγS binding. Nicotine pretreatment enhanced quinpirole-induced c-fos mRNA expression in the hypothalamic paraventricular and supraoptic nuclei in adolescents only. Adolescent nicotine pretreatment enhanced c-fos mRNA expression in corticotropin releasing factor (CRF) cells of the paraventricular nucleus, and enhancement of penile erection was blocked by the CRF-1 receptor antagonist, CP 376,396. These findings suggest that adolescent dopamine and CRF systems are vulnerable to alteration by nicotine. This is the first evidence for a role of CRF in adolescent erectile response. PMID:24157491

  12. Neuronal Acetylcholine Nicotinic Receptors as New Targets for Lung Cancer Treatment.

    PubMed

    Mucchietto, Vanessa; Crespi, Arianna; Fasoli, Francesca; Clementi, Francesco; Gotti, Cecilia

    2016-01-01

    Lung cancer is the leading cause of cancer-related deaths worldwide. Smoking accounts for approximately 70% of the cases of non- small cell lung cancer (NSCLC) and 90% of the cases of small-cell lung cancer (SCLC), although some patients develop lung cancer without a history of smoking. Nicotine is the most active addictive component of tobacco smoke. It does not initiate tumorigenesis in humans and rodents, but it alters the pathophysiology of lung cells by inducing the secretion of growth factors, neurotransmitters and cytokines, and promotes tumour growth and metastases by inducing cell cycle progression, migration, invasion, angiogenesis and the evasion of apoptosis. Most of these effects are a result of nicotine binding and activation of cell-surface neuronal nicotinic acetylcholine receptors (nAChRs) and downstream intracellular signalling cascades, and many are blocked by nAChR subtype-selective antagonists. Recent genome-wide association studies have revealed single nucleotide polymorphisms of nAChR subunits that influence nicotine dependence and lung cancer. This review describes the molecular basis of nAChR structural and functional diversity in normal and cancer lung cells, and the genetic alterations facilitating smoking-induced lung cancers. It also summarises current knowledge concerning the intracellular pathways activated by nicotine and other compounds present in tobacco smoke. PMID:26845123

  13. Tritiated-nicotine- and /sup 125/I-alpha-bungarotoxin-labeled nicotinic receptors in the interpeduncular nucleus of rats. II. Effects of habenular destruction

    SciTech Connect

    Clarke, P.B.; Hamill, G.S.; Nadi, N.S.; Jacobowitz, D.M.; Pert, A.

    1986-09-15

    The cholinergic innervation of the interpeduncular nucleus (IPN) is wholly extrinsic and is greatly attenuated by bilateral habenular destruction. We describe changes in the labeling of putative nicotinic receptors within this nucleus at 3, 5, or 11 days after bilateral habenular lesions. Adjacent tissue sections of the rat IPN were utilized for /sup 3/H-nicotine and /sup 125/I-alpha-bungarotoxin (/sup 125/I-BTX) receptor autoradiography. Compared to sham-operated controls, habenular destruction significantly reduced autoradiographic /sup 3/H-nicotine labeling in rostral (-25%), intermediate (-13%), and lateral subnuclei (-36%). Labeling in the central subnucleus was unchanged. Loss of labeling was maximal at the shortest survival time (3 days) and did not change thereafter. In order to establish whether this loss was due to a reduction in the number or the affinity of /sup 3/H-nicotine-binding sites, a membrane assay was performed on microdissected IPN tissue from rats that had received surgery 3 days previously. Bilateral habenular lesions produced a 35% reduction of high-affinity /sup 3/H-nicotine-binding sites, with no change in binding affinity. Bilateral habenular lesions reduced /sup 125/I-BTX labeling in the intermediate subnuclei, and a slight increase occurred in the rostral subnucleus. In the lateral subnuclei, /sup 125/I-BTX labeling was significantly reduced (27%) at 3 days but not at later survival times. In view of the known synaptic morphology of the habenulointerpeduncular tract, it is concluded that a subpopulation of /sup 3/H-nicotine binding sites within the IPN is located on afferent axons and/or terminals. This subpopulation, located within rostral, intermediate, and lateral subnuclei, may correspond to presynaptic nicotinic cholinergic receptors. Sites that bind /sup 125/I-BTX may include a presynaptic subpopulation located in the lateral and possibly the intermediate subnuclei.

  14. Transcription factor assembly on the nicotinic receptor beta4 subunit gene promoter.

    PubMed

    Scofield, Michael D; Brüschweiler-Li, Lei; Mou, Zhongming; Gardner, Paul D

    2008-04-16

    Nicotinic acetylcholine receptors are involved in a plethora of fundamental biological processes ranging from muscle contraction to formation of memories. The receptors are pentameric proteins whose subunits are encoded by distinct genes. Subunit composition of a mature nicotinic receptor is governed in part by the transcriptional regulation of each subunit gene. Here, using chromatin immunoprecipitation assays, we report the interaction of the transcription factors Sp1, Sp3, c-Jun and Sox10 with the beta4 subunit gene promoter in neuronal-like cell lines and rodent brain tissue. Our results corroborate previous in-vitro data demonstrating that these transcription factors interact with the beta4 promoter. Taken together, these data suggest that Sp1, Sp3, c-Jun and Sox10 regulate expression of the beta4 subunit gene in the mammalian brain. PMID:18382288

  15. Neuronal-type alpha-bungarotoxin receptors and the alpha 5-nicotinic receptor subunit gene are expressed in neuronal and nonneuronal human cell lines.

    PubMed Central

    Chini, B; Clementi, F; Hukovic, N; Sher, E

    1992-01-01

    alpha-Bungarotoxin (alpha Bgtx) is a toxin known to interact with muscle nicotinic receptors and with some neuronal nicotinic receptors. We show that alpha Bgtx binding sites are also expressed in nonmuscle and nonneuronal human cells, including small cell lung carcinoma and several epithelial cell lines. These receptors are immunologically related to the alpha Bgtx receptors of unknown function described in the nervous system and in the IMR32 neuroblastoma cell line and are distinct from muscle nicotinic receptors. We have also cloned from IMR32 cells the human alpha 5-nicotinic receptor subunit, which is supposed to participate in the formation of alpha Bgtx receptors. Transcripts corresponding to the alpha 5-subunit gene were found not only in neuroblastoma cells but also in all the cell lines expressing alpha Bgtx receptors, with the exception of the TE671 cell line, whose nicotinic receptor subunits are of the muscle type. We conclude that both alpha Bgtx receptors and the alpha 5-nicotinic subunit gene are not neuron-specific, as previously thought, but are expressed in a number of human cell lines of various origin. Images PMID:1542648

  16. The Role of Nicotinic Acetylcholine Receptors in the Medial Prefrontal Cortex and Hippocampus in Trace Fear Conditioning

    PubMed Central

    Raybuck, J. D.; Gould, T. J.

    2010-01-01

    Acute nicotine enhances multiple types of learning including trace fear conditioning but the underlying neural substrates of these effects are not well understood. Trace fear conditioning critically involves the medial prefrontal cortex and hippocampus, which both express nicotinic acetylcholine receptors (nAChRs). Therefore, nicotine could act in either or both areas to enhance trace fear conditioning. To identify the underlying neural areas and nAChR subtypes, we examined the effects of infusion of nicotine, or nicotinic antagonists dihydro-beta-erythroidine (DHβE: high-affinity nAChRs) or methyllycaconitine (MLA: low-affinity nAChRs) into the dorsal hippocampus, ventral hippocampus, and medial prefrontal cortex (mPFC) on trace and contextual fear conditioning. We found that the effects of nicotine on trace and contextual fear conditioning vary by brain region and nAChR subtype. The dorsal hippocampus was involved in the effects of nicotine on both trace and contextual fear conditioning but each task was sensitive to different doses of nicotine. Additionally, dorsal hippocampal infusion of the antagonist DHβE produced deficits in trace but not contextual fear conditioning. Nicotine infusion into the ventral hippocampus produced deficits in both trace and contextual fear conditioning. In the mPFC, nicotine enhanced trace but not contextual fear conditioning. Interestingly, infusion of the antagonists MLA or DHβE in the mPFC also enhanced trace fear conditioning. These findings suggest that nicotine acts on different substrates to enhance trace versus contextual fear conditioning, and that nicotine-induced desensitization of nAChRs in the mPFC may contribute to the effects of nicotine on trace fear conditioning. PMID:20727979

  17. Nicotinic cholinergic receptors in rat brain. Annual report No. 3, 1 May 85-30 Apr 86

    SciTech Connect

    Kellar, K.J.

    1986-05-01

    We have compared the characteristics of the recognition sites for 3(H)acetylcholine and 3H(-)nicotine in rat brain and found that the pharmacology, distribution, disulfide bond requirement, and regulation by chronic administration of nicotine and soman are identical. From these studies we conclude that 3Hacetylcholine and 3H(-)nicotine recognize the same recognition site which has the characteristics expected of a nicotinic cholinergic receptor. We have also determined that 3Hacetylcholine of high specific radioactivity (80 Ci/mmol) is an excellent ligand with which to study muscarinic receptors that have high affinity for agonists. These receptors may represent a subtype of muscarinic receptors found in brain, heart, glands, an some smooth muscle. (JS)

  18. In vivo chronic nicotine exposure differentially and reversibly affects upregulation and stoichiometry of α4β2 nicotinic receptors in cortex and thalamus.

    PubMed

    Fasoli, F; Moretti, M; Zoli, M; Pistillo, F; Crespi, A; Clementi, F; Mc Clure-Begley, T; Marks, M J; Gotti, C

    2016-09-01

    Studies with heterologous expression systems have shown that the α4β2 nicotinic acetylcholine receptor (nAChR) subtype can exist in two stoichiometries (with two [(α4)2(β2)3] or three [(α4)3(β2)2] copies of the α subunit in the receptor pentamer) which have different pharmacological and functional properties and are differently regulated by chronic nicotine treatment. However, the effects of nicotine treatment in vivo on native α4β2 nAChR stoichiometry are not well known. We investigated in C57BL/6 mice the in vivo effect of 14-day chronic nicotine treatment and subsequent withdrawal, on the subunit expression and β2/α4 subunit ratio of (3)H-epibatidine labeled α4β2*-nAChR in total homogenates of cortex and thalamus. We found that in basal conditions the ratio of the β2/α4 subunit in the cortex and thalamus is different indicating a higher proportion in receptors with (α4)2(β2)3 subunit stoichiometry in the thalamus. For cortex exposure to chronic nicotine elicited an increase in receptor density measured by (3)H-epibatidine binding, an increase in the α4 and β2 protein levels, and an increase in β2/α4 subunit ratio, that indicates an increased proportion of receptors with the (α4)2(β2)3 stoichiometry. For thalamus we did not find a significant increase in receptor density, α4 and β2 protein levels, or changes in β2/α4 subunit ratio. All the changes elicited by chronic nicotine in cortex were transient and returned to basal levels with an average half-life of 2.8 days following nicotine withdrawal. These data suggest that chronic nicotine exposure in vivo favors increased assembly of α4β2 nAChR containing three β2 subunits. A greater change in stoichiometry was observed for cortex (which has relatively low basal expression of (α4)2(β2)3 nAChR) than in thalamus (which has a relatively high basal expression of (α4)2(β2)3 nAChR). PMID:27157710

  19. Mapping of the acetylcholine binding site of the nicotinic acetylcholine receptor: ( sup 3 H)nicotine as an agonist photoaffinity label

    SciTech Connect

    Middleton, R.E.; Cohen, J.B. )

    1991-07-16

    The agonist ({sup 3}H)nicotine was used as a photoaffinity label for the acetylcholine binding sties on the Torpedo nicotinic acetylcholine receptor (AChR). ({sup 3}H)Nicotine binds at equilibrium with K{sub eq} = 0.6 {mu}M to the agonist binding sites. Irradiation with 254-nm light of AChR-rich membranes equilibrated with ({sup 3}H)nicotine resulted in covalent incorporation into the {alpha}- and {gamma}-subunits, which was inhibited by agonists and competitive antagonists but not by noncompetitive antagonists. Inhibition of labeling by d-tubocurarine demonstrated that the {alpha}-subunit was labeled via both agonist sites but the {gamma}-subunit was labeled only via the site that binds d-tubocurarine with high affinity. Chymotryptic digestion of the {alpha}-subunit confirmed that Try-198 was the principal amino acid labeled by ({sup 3}H)nicotine. This confirmation required a novel radiosequencing strategy employing o-phthalaldehyde ({sup 3}H)Nicotine, which is the first photoaffinity agonist used, labels primarily Tyr-198 in contrast to competitive antagonist affinity labels, which label primarily Tyr-190 and Cys-192/Cys-193.

  20. Contributions from Caenorhabditis elegans functional genetics to antiparasitic drug target identification and validation: nicotinic acetylcholine receptors, a case study.

    PubMed

    Brown, L A; Jones, A K; Buckingham, S D; Mee, C J; Sattelle, D B

    2006-05-31

    Following the complete sequencing of the genome of the free-living nematode, Caenorhabditis elegans, in 1998, rapid advances have been made in assigning functions to many genes. Forward and reverse genetics have been used to identify novel components of synaptic transmission as well as determine the key components of antiparasitic drug targets. The nicotinic acetylcholine receptors (nAChRs) are prototypical ligand-gated ion channels. The functions of these transmembrane proteins and the roles of the different members of their extensive subunit families are increasingly well characterised. The simple nervous system of C. elegans possesses one of the largest nicotinic acetylcholine receptor gene families known for any organism and a combination of genetic, microarray, physiological and reporter gene expression studies have added greatly to our understanding of the components of nematode muscle and neuronal nAChR subtypes. Chemistry-to-gene screens have identified five subunits that are components of nAChRs sensitive to the antiparasitic drug, levamisole. A novel, validated target acting downstream of the levamisole-sensitive nAChR has also been identified in such screens. Physiology and molecular biology studies on nAChRs of parasitic nematodes have also identified levamisole-sensitive and insensitive subtypes and further subdivisions are under investigation. PMID:16620825

  1. Nicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone induce cyclooxygenase-2 activity in human gastric cancer cells: Involvement of nicotinic acetylcholine receptor (nAChR) and {beta}-adrenergic receptor signaling pathways

    SciTech Connect

    Shin, Vivian Yvonne; Jin, H.C.; Ng, Enders K.O.; Yu Jun; Leung, W.K.; Cho, C.H.; Sung, J.J.Y.

    2008-12-01

    Induction of cyclooxygenase-2 (COX-2) associates with cigarette smoke exposure in many malignancies. Nicotine and its derivative, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), are the two important components in cigarette smoke that contributes to cancer development. However, the molecular mechanism(s) by which nicotine or NNK promotes gastric carcinogenesis remains largely unknown. We found that nicotine and NNK significantly enhanced cell proliferation in AGS cells that expressed both alpha7 nicotinic acetylcholine receptor ({alpha}7 nAChR) and {beta}-adrenergic receptors. Treatment of cells with {alpha}-bungarotoxin ({alpha}-BTX, {alpha}7nAChR antagonist) or propranolol ({beta}-adrenergic receptor antagonist) blocked NNK-induced COX-2/PGE{sub 2} and cell proliferation, while nicotine-mediated cell growth and COX-2/PGE{sub 2} induction can only be suppressed by propranolol, but not {alpha}-BTX. Moreover, in contrast to the dependence of growth promoting effect of nicotine on Erk activation, inhibitor of p38 mitogen-activated protein kinase (MAPK) repressed NNK-induced COX-2 upregulation and resulted in suppression of cell growth. In addition, nicotine and NNK mediated COX-2 induction via different receptors to modulate several G1/S transition regulatory proteins and promote gastric cancer cell growth. Selective COX-2 inhibitor (SC-236) caused G1 arrest and abrogated nicotine/NNK-induced cell proliferation. Aberrant expression of cyclin D1 and other G1 regulatory proteins are reversed by blockade of COX-2. These results pointed to the importance of adrenergic and nicotinic receptors in gastric tumor growth through MAPK/COX-2 activation, which may perhaps provide a chemoprevention strategy for cigarette smoke-related gastric carcinogenesis.

  2. Opioid receptor types involved in the development of nicotine physical dependence in an invertebrate (Planaria) model.

    PubMed

    Raffa, Robert B; Baron, Steve; Bhandal, Jaspreet S; Brown, Tevin; Song, Kevin; Tallarida, Christopher S; Rawls, Scott M

    2013-11-01

    Recent data suggest that opioid receptors are involved in the development of nicotine physical dependence in mammals. Evidence in support of a similar involvement in an invertebrate (Planaria) is presented using the selective opioid receptor antagonist naloxone, and the more receptor subtype-selective antagonists CTAP (D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2) (μ, MOR), naltrindole (δ, DOR), and nor-BNI (norbinaltorphimine) (κ, KOR). Induction of physical dependence was achieved by 60-min pre-exposure of planarians to nicotine and was quantified by abstinence-induced withdrawal (reduction in spontaneous locomotor activity). Known MOR and DOR subtype-selective opioid receptor antagonists attenuated the withdrawal, as did the non-selective antagonist naloxone, but a KOR subtype-selective antagonist did not. An involvement of MOR and DOR, but not KOR, in the development of nicotine physical dependence or in abstinence-induced withdrawal was thus demonstrated in a sensitive and facile invertebrate model. PMID:24084318

  3. α4 nicotinic acetylcholine receptor modulated by galantamine on nigrostriatal terminals regulates dopamine receptor-mediated rotational behavior.

    PubMed

    Inden, Masatoshi; Takata, Kazuyuki; Yanagisawa, Daijiro; Ashihara, Eishi; Tooyama, Ikuo; Shimohama, Shun; Kitamura, Yoshihisa

    2016-03-01

    Galantamine, an acetylcholine esterase (AChE) inhibitor used to treat dementia symptoms, also acts as an allosteric potentiating ligand (APL) at nicotinic acetylcholine receptors (nAChRs). This study was designed to evaluate the allosteric effect of galantamine on nAChR regulation of nigrostrial dopaminergic neuronal function in the hemiparkinsonian rat model established by unilateral nigral 6-hydroxydopamine (6-OHDA) injection. Methamphetamine, a dopamine releaser, induced ipsilateral rotation, whereas dopamine agonists apomorphine (a non-selective dopamine receptor agonist), SKF38393 (a selective dopamine D1 receptor agonist), and quinpirole (a selective dopamine D2 receptor agonist) induced contralateral rotation. When 6-OHDA-injected rats were co-treated with nomifensine, a dopamine transporter inhibitor, a more pronounced and a remarkable effect of nicotine and galantamine was observed. Under these conditions, the combination of nomifensine with nicotine or galantamine induced the ipsilateral rotation similar to the methamphetamine-induced rotational behavior, indicating that nicotine and galantamine also induce dopamine release from striatal terminals. Both nicotine- and galantamine-induced rotations were significantly blocked by flupenthixol (an antagonist of both D1 and D2 dopamine receptors) and mecamylamine (an antagonist of nAChRs), suggesting that galantamine modulation of nAChRs on striatal dopaminergic terminals regulates dopamine receptor-mediated movement. Immunohistochemical staining showed that α4 nAChRs were highly expressed on striatal dopaminergic terminals, while no α7 nAChRs were detected. Pretreatment with the α4 nAChR antagonist dihydroxy-β-erythroidine significantly inhibited nicotine- and galantamine-induced rotational behaviors, whereas pretreatment with the α7 nAChR antagonist methyllycaconitine was ineffective. Moreover, the α4 nAChR agonist ABT-418 induced ipsilateral rotation, while the α7 nAChR agonist PNU282987 had no

  4. Nicotinic Acid Adenine Dinucleotide Phosphate Analogs Substituted on the Nicotinic Acid and Adenine Ribosides. Effects on Receptor-Mediated Ca2+ release

    PubMed Central

    Trabbic, Christopher J.; Zhang, Fan; Walseth, Timothy F.; Slama, James T.

    2015-01-01

    Nicotinic acid adenine dinucleotide phosphate (NAADP) is a Ca2+ releasing intracellular second messenger in both mammals and echinoderms. We report that large functionalized substituents introduced at the nicotinic acid 5-position are recognized by the sea urchin receptor, albeit with a 20–500 fold loss in agonist potency. 5-(3-Azidopropyl)-NAADP was shown to release Ca2+ with an EC50 of 31 µM and to compete with NAADP for receptor binding with an IC50 of 56 nM. Attachment of charged groups to the nicotinic acid of NAADP is associated with loss of activity, suggesting that the nicotinate riboside moiety is recognized as a neutral zwitterion. Substituents (Br- and N3-) can be introduced at the 8-adenosyl position of NAADP while preserving high potency and agonist efficacy and an NAADP derivative substituted at both the 5-position of the nicotinic acid and at the 8-adenosyl position was also recognized although the agonist potency was significantly reduced. PMID:25826221

  5. Anxiolytic-like and anxiogenic-like effects of nicotine are regulated via diverse action at β2*nicotinic acetylcholine receptors

    PubMed Central

    Anderson, S M; Brunzell, D H

    2015-01-01

    Background and Purpose Nicotine dose-dependently activates or preferentially desensitizes β2 subunit containing nicotinic ACh receptors (β2*nAChRs). Genetic and pharmacological manipulations assessed effects of stimulation versus inhibition of β2*nAChRs on nicotine-associated anxiety-like phenotype. Experimental Approach Using a range of doses of nicotine in β2*nAChR subunit null mutant mice (β2KO; backcrossed to C57BL/6J) and their wild-type (WT) littermates, administration of the selective β2*nAChR agonist, 5I-A85380, and the selective β2*nAChR antagonist dihydro-β-erythroidine (DHβE), we determined the behavioural effects of stimulation and inhibition of β2*nAChRs in the light–dark and elevated plus maze (EPM) assays. Key Results Low-dose i.p. nicotine (0.05 mg·kg−1) supported anxiolysis-like behaviour independent of genotype whereas the highest dose (0.5 mg·kg−1) promoted anxiogenic-like phenotype in WT mice, but was blunted in β2KO mice for the measure of latency. Administration of 5I-A85380 had similar dose-dependent effects in C57BL/6J WT mice; 0.001 mg·kg−1 5I-A85380 reduced anxiety on an EPM, whereas 0.032 mg·kg−1 5I-A85380 promoted anxiogenic-like behaviour in both the light–dark and EPM assays. DHβE pretreatment blocked anxiogenic-like effects of 0.5 mg·kg−1 nicotine. Similarly to DHβE, pretreatment with low-dose 0.05 mg·kg−1 nicotine did not accumulate with 0.5 mg·kg−1 nicotine, but rather blocked anxiogenic-like effects of high-dose nicotine in the light–dark and EPM assays. Conclusions and Implications These studies provide direct evidence that low-dose nicotine inhibits nAChRs and demonstrate that inhibition or stimulation of β2*nAChRs supports the corresponding anxiolytic-like or anxiogenic-like effects of nicotine. Inhibition of β2*nAChRs may relieve anxiety in smokers and non-smokers alike. PMID:25625469

  6. Effect of a D3 receptor antagonist on context-induced reinstatement of nicotine seeking.

    PubMed

    Sabioni, Pamela; Di Ciano, Patricia; Le Foll, Bernard

    2016-01-01

    Despite the existence of several treatment options for smoking cessation, the rate of relapse after treatment is very high. We and others have proposed that targeting the dopamine D3 receptor (DRD3) may be a good strategy for treatment of nicotine dependence. In human participants, reintroduction to an environment previously associated with drug-taking may induce relapse. In animals, such phenomenon can be studied using the context-induced reinstatement paradigm. As the role of DRD3 in context-induced reinstatement of nicotine-seeking has not yet been explored, we investigated the effects of different doses of the selective DRD3 antagonist SB-277011-A on this reinstatement. Sprague-Dawley adult rats were first trained to self-administer nicotine and subsequently underwent extinction in a second context for 5-7 days. We evaluated the effect of 1, 3 or 10mg/kg of SB-277011-A administered prior to the reintroduction to the training context. We used two different designs: 1) a between-subjects design with a unique reinstatement test; and 2) a counterbalanced within-subjects design, with 4 reinstatement tests. Our findings indicate that, in the within-subjects design, the magnitude of responding induced by the context-induced reinstatement of nicotine seeking was robust during the first reinstatement test, but significantly decreased with repeated testing. SB-277011-A (10mg/kg) blocked context-induced reinstatement of nicotine-seeking at first exposure to the context (between-subjects design), but not after repeated context exposure which produced weaker reinstatement over days. Our results support a role for DRD3 mediating context-induced reinstatement of nicotine seeking, but these effects may not be sustained over time. Further studies should explore this in human participants for validation. PMID:26279138

  7. Evidence for functional atypical nicotinic receptors that activate K+-dependent Cl- secretion in mouse tracheal epithelium.

    PubMed

    Hollenhorst, Monika I; Lips, Katrin S; Weitz, Ariane; Krasteva, Gabriela; Kummer, Wolfgang; Fronius, Martin

    2012-01-01

    The present study focused on the influence of nicotinic acetylcholine receptors (nAChR) on ion transport processes in mouse tracheal epithelium. RT-PCR experiments revealed expression of the α3, α4, α5, α7, α9, α10, β2, and β4 nAChR subunits in mouse tracheal epithelium. In Ussing chamber recordings of mouse tracheae, apically applied nicotine (100 μM) induced a dose-dependent increase of the transepithelial short-circuit current (EC(50): 14.6 μM). The nicotine-induced effect (I(NIC)) was attenuated by mecamylamine (25 μM, apical) and methyllycaconitine (1 μM, apical). The nAChR agonist 1.1-dimethyl-4-phenylpiperatinium iodide (DMPP) (100 μM) revealed apical and basolateral location of the receptors. I(NIC) was not affected by the sodium channel inhibitor amiloride (10 μM, apical) or the cystic fibrosis transmembrane conductance regulator inhibitor CFTR(inh)-172 (20 μM, apical) but was reduced by the chloride channel inhibitor 5-nitro-2-(3-phenylpropylamino)benzoic acid (100 μM, apical), the Na(+)/K(+)/2Cl(-) cotransporter inhibitor bumetanide (200 μM, basolateral), the potassium channel inhibitor Ba(2+) (5 mM, basolateral), and 4.4'-diisothiocyanatostilbene-2.2'-disulfonate (100 μM, apical), indicating a contribution of Ca(2+)-activated chloride channels and potassium channels. Removal of extracellular Na(+) (apical) or Ca(2+) (apical) did not influence I(NIC) but reduced the DMPP effect. Experiments with the Ca(2+)-ionophore A23187, a mix of 3-isobutyl-1-methylxanthine and forskolin, or the inositol-1,4,5-triphospate (IP(3)) receptor inhibitor 2-aminoethyl-diphenyl-borinate (75 μM, apical) decreased I(NIC), indicating a nicotine-mediated increase of intracellular Ca(2+) and cAMP levels involving the IP(3) signaling pathway. These findings indicate the activity of Ca(2+)-permeable nAChRs and alternative metabotropic pathways by nAChR activation that mediate Cl(-) and K(+) transport in tracheal epithelium. PMID:21852683

  8. Effect of the selective kappa-opioid receptor antagonist JDTic on nicotine antinociception, reward, and withdrawal in the mouse

    PubMed Central

    Jackson, K. J.; Negus, S. S.; Damaj, M. I.

    2010-01-01

    Rationale Several lines of evidence support a role for the endogenous opioid system in mediating behaviors associated with drug dependence. Specifically, recent findings suggest that the kappa-opioid receptor (KOR) may play a role in aspects of nicotine dependence, which contribute to relapse and continued tobacco smoking. Objective The objective of this study is to determine the involvement of the KOR in the initial behavioral responses of nicotine, nicotine reward, and nicotine withdrawal using the highly selective KOR antagonist JDTic. JDTic doses of 1, 4, 8, or 16 mg/kg were administered subcutaneously (s.c.) 18 h prior to nicotine treatment. Results JDTic dose-dependently blocked acute nicotine-induced antinociception in the tail-flick but not the hot-plate test and did not significantly attenuate morphine’s antinociceptive effect in either the tail-flick or hot-plate test. Furthermore, JDTic (8 and 16 mg/kg, s.c.) failed to block the expression of nicotine reward as measured by the conditioned place preference model. In contrast, JDTic and the KOR antagonist norBNI attenuated the expression of both the physical (somatic signs and hyperalgesia) and affective (anxiety-related behavior and conditioned place aversion) nicotine withdrawal signs. Conclusions Our findings clearly show that the KOR is involved in mediating the withdrawal aspects of nicotine dependence. The results from this study suggest that blockade of the KOR by selective KOR antagonists may be useful smoking cessation pharmacotherapies. PMID:20232057

  9. Heterogeneity of Drosophila nicotinic acetylcholine receptors: SAD, a novel developmentally regulated alpha-subunit.

    PubMed Central

    Sawruk, E; Schloss, P; Betz, H; Schmitt, B

    1990-01-01

    Two genes, ard and als, are known to encode subunits of the nicotinic acetylcholine receptor (nAChR) in Drosophila. Here we describe the isolation of cDNA clones encoding a novel member (SAD, or alpha 2) of this receptor protein family. The deduced amino acid sequence displays high homology to the ALS protein and shares structural features with ligand binding nAChR alpha-subunits. Sad transcripts accumulate during major periods of neuronal differentiation and, in embryos, are localized in the central nervous system. Expression of SAD cRNA in Xenopus oocytes generates cation channels that are gated by nicotine. These data indicate heterogeneity of nAChRs in Drosophila. Images Fig. 3. Fig. 4. PMID:1697262

  10. Molecular-Dynamics Simulations of ELIC a Prokaryotic Homologue of the Nicotinic Acetylcholine Receptor

    SciTech Connect

    Cheng, Xiaolin; Ivanov, Ivaylo N; Wang, Hailong; McCammon, Jonathan

    2009-01-01

    The ligand-gated ion channel from Erwinia chrysanthemi (ELIC) is a prokaryotic homolog of the eukaryotic nicotinic acetylcholine receptor (nAChR) that responds to the binding of neurotransmitter acetylcholine and mediates fast signal transmission. ELIC is similar to the nAChR in its primary sequence and overall subunit organization, but despite their structural similarity, it is not clear whether these two ligand-gated ion channels operate in a similar manner. Further, it is not known to what extent mechanistic insights gleaned from the ELIC structure translate to eukaryotic counterparts such as the nAChR. Here we use molecular-dynamics simulations to probe the conformational dynamics and hydration of the transmembrane pore of ELIC. The results are compared with those from our previous simulation of the human ?7 nAChR. Overall, ELIC displays increased stability compared to the nAChR, whereas the two proteins exhibit remarkable similarity in their global motion and flexibility patterns. The majority of the increased stability of ELIC does not stem from the deficiency of the models used in the simulations, and but rather seems to have a structural basis. Slightly altered dynamical correlation features are also observed among several loops within the membrane region. In sharp contrast to the nAChR, ELIC is completely dehydrated from the pore center to the extracellular end throughout the simulation. Finally, the simulation of an ELIC mutant substantiates the important role of F246 on the stability, hydration and possibly function of the ELIC channel.

  11. Tritiated-nicotine and /sup 125/I-alpha-bungarotoxin-labeled nicotinic receptors in the interpeduncular nucleus of rats. I. Subnuclear distribution

    SciTech Connect

    Hamill, G.S.; Clarke, P.B.; Pert, A.; Jacobowitz, D.M.

    1986-09-15

    The distribution of nicotinic receptors within the interpeduncular nucleus (IPN) was determined in male rats following in vitro labeling with the cholinergic ligands /sup 3/H-nicotine and /sup 125/I-alpha-bungarotoxin (BTX). Autoradiographic images of two rostrocaudal levels of IPN were analyzed by computer-assisted densitometry and the optical density contributed by displaceable labeling was determined in the rostral, central, intermediate, and lateral subnuclei. /sup 3/H-nicotine labeling density within the four subnuclei differs significantly at both levels of IPN. The greatest density of labeling is localized in the rostral subnucleus, followed in order of diminishing density by the central, intermediate, and lateral subnuclei. Labeling within the rostral subnucleus is prominently localized within its central zone. In the central subnucleus, a dense concentration of binding sites is apparent in the middle region, adjacent to less dense vertically oriented columns; /sup 3/H-nicotine binding sites in the lateral subnuclei appear to be most concentrated medially, adjacent to the intermediate subnuclei. /sup 125/I-BTX labeling density within the four subnuclei also differs significantly at both levels of IPN. The greatest density of labeling is found in the rostral subnucleus, followed in order of decreasing density by the lateral, central, and intermediate subnuclei. The ovoid regions of the rostral subnucleus contain dense /sup 125/I-BTX labeling. In the lateral subnuclei, /sup 125/I-BTX binding appears to be predominantly along the lateral margins of the subnucleus. The present data indicate that the IPN contains two distinct populations of putative cholinergic nicotinic receptors identified, respectively, by /sup 3/H-nicotine and /sup 125/I-BTX labeling. Each population of labeled receptors is uniquely localized in patterns that suggest differences in density within and across subnuclei.

  12. Nicotinic acid is a common regulator of heat-sensing TRPV1-4 ion channels.

    PubMed

    Ma, Linlin; Lee, Bo Hyun; Clifton, Heather; Schaefer, Saul; Zheng, Jie

    2015-01-01

    Nicotinic acid (NA, a.k.a. vitamin B3 or niacin) can reduce blood cholesterol and low-density lipoproteins whereas increase high-density lipoproteins. However, when NA is used to treat dyslipidemias, it causes a strong side effect of cutaneous vasodilation, commonly called flushing. A recent study showed that NA may cause flushing by lowering activation threshold temperature of the heat-sensitive capsaicin receptor TRPV1 ion channel, leading to its activation at body temperature. The finding calls into question whether NA might also interact with the homologous heat-sensitive TRPV2-4 channels, particularly given that TRPV3 and TRPV4 are abundantly expressed in keratinocytes of the skin where much of the flushing response occurs. We found that NA indeed potentiated TRPV3 while inhibited TRPV2 and TRPV4. Consistent with these gating effects, NA lowered the heat-activation threshold of TRPV3 but elevated that of TRPV4. We further found that activity of TRPV1 was substantially prolonged by extracellular NA, which may further enhance the direct activation effect. Consistent with the broad gating effect on TRPV1-4 channels, evidence from the present study hints that NA may share the same activation pathway as 2-aminoethoxydiphenyl borate (2-APB), a common agonist for these TRPV channels. These findings shed new light on the molecular mechanism underlying NA regulation of TRPV channels. PMID:25752528

  13. Nicotinic Acid is a Common Regulator of Heat-Sensing TRPV1-4 Ion Channels

    PubMed Central

    Ma, Linlin; Lee, Bo Hyun; Clifton, Heather; Schaefer, Saul; Zheng, Jie

    2015-01-01

    Nicotinic acid (NA, a.k.a. vitamin B3 or niacin) can reduce blood cholesterol and low-density lipoproteins whereas increase high-density lipoproteins. However, when NA is used to treat dyslipidemias, it causes a strong side effect of cutaneous vasodilation, commonly called flushing. A recent study showed that NA may cause flushing by lowering activation threshold temperature of the heat-sensitive capsaicin receptor TRPV1 ion channel, leading to its activation at body temperature. The finding calls into question whether NA might also interact with the homologous heat-sensitive TRPV2–4 channels, particularly given that TRPV3 and TRPV4 are abundantly expressed in keratinocytes of the skin where much of the flushing response occurs. We found that NA indeed potentiated TRPV3 while inhibited TRPV2 and TRPV4. Consistent with these gating effects, NA lowered the heat-activation threshold of TRPV3 but elevated that of TRPV4. We further found that activity of TRPV1 was substantially prolonged by extracellular NA, which may further enhance the direct activation effect. Consistent with the broad gating effect on TRPV1–4 channels, evidence from the present study hints that NA may share the same activation pathway as 2-aminoethoxydiphenyl borate (2-APB), a common agonist for these TRPV channels. These findings shed new light on the molecular mechanism underlying NA regulation of TRPV channels. PMID:25752528

  14. The α6 nicotinic acetylcholine receptor subunit influences ethanol-induced sedation.

    PubMed

    Kamens, Helen M; Hoft, Nicole R; Cox, Ryan J; Miyamoto, Jill H; Ehringer, Marissa A

    2012-08-01

    Alcohol and nicotine are often co-used and data from human and animals studies have demonstrated that common genes underlie responses to these two drugs. Recently, the genes that code for the subunits of the nicotinic acetylcholine receptors have been implicated as a common genetic mediator for alcohol and nicotine responses. The mammalian genes that code for the α6 and β3 subunits of the nicotinic acetylcholine receptor (Chrna6 and Chrnb3, respectively) are located adjacent to each other on human and mouse chromosome 8. These subunits have gained attention as potential regulators of drug behaviors because of their expression in the striatum where they have been shown to modulate dopamine release. Human genetic studies have shown that variation in these genes is associated with alcohol phenotypes. In the current experiments, mice lacking the Chrna6 or Chrnb3 gene were tested for three ethanol behaviors: choice ethanol consumption, ataxia, and sedation. Wildtype (WT), heterozygous (HET), and knockout (KO) mice of each strain went through a standard 2-bottle choice drinking paradigm, the balance beam, and the Loss of Righting Reflex (LORR) paradigm. No genotypic effects on any of the 3 behavioral tasks were observed in Chrnb3 animals. While the Chrna6 gene did not significantly influence ethanol consumption (g/kg) or ataxia, mice lacking the α6 subunit took significantly longer to recover their righting reflex than WT animals. These data provide evidence that receptors containing this subunit modulate the sedative effects of ethanol. Further work examining other models of ethanol consumption and behavioral responses to ethanol is needed to fully characterize the role of these receptor subunits in modulating ethanol responses. PMID:22572056

  15. The α6 nicotinic acetylcholine receptor subunit influences ethanol-induced sedation

    PubMed Central

    Kamens, Helen M.; Hoft, Nicole R.; Cox, Ryan J.; Miyamoto, Jill; Ehringer, Marissa A.

    2012-01-01

    Alcohol and nicotine are often co-used and data from human and animals studies have demonstrated that common genes underlie responses to these two drugs. Recently, the genes that code for the subunits of the nicotinic acetylcholine receptors have been implicated as a common genetic mediator for alcohol and nicotine responses. The mammalian genes that code for the α6 and β3 subunits of the nicotinic acetylcholine receptor (Chrna6 and Chrnb3, respectively) are located adjacent to each other on human and mouse chromosome 8. These subunits have gained attention as potential regulators of drug behaviors because of their expression in the striatum where they have been shown to modulate dopamine release. Human genetic studies have shown that variation in these genes is associated with alcohol phenotypes. In the current experiments, mice lacking the Chrna6 or Chrnb3 gene were tested for three ethanol behaviors: choice ethanol consumption, ataxia, and sedation. Wildtype (WT), heterozygous (HET), and knockout (KO) mice of each strain went through a standard 2-bottle choice drinking paradigm, the balance beam, and the Loss of Righting Reflex (LORR) paradigm. No genotypic effects on any of the 3 behavioral tasks were observed in Chrnb3 animals. While the Chrna6 gene did not significantly influence ethanol consumption (g/kg) or ataxia, mice lacking the α6 subunit took significantly longer to recover their righting reflex than WT animals. These data provide evidence that receptors containing this subunit modulate the sedative effects of ethanol. Further work examining other models of ethanol consumption and behavioral responses to ethanol is needed to fully characterize the role of these receptor subunits in modulating ethanol responses. PMID:22572056

  16. N-glycosylation sites on the nicotinic ACh receptor subunits regulate receptor channel desensitization and conductance.

    PubMed

    Nishizaki, Tomoyuki

    2003-06-10

    The present study investigated the effects of N-glycosylation sites on Torpedo acetylcholine (ACh) receptors expressed in Xenopus oocytes by monitoring whole-cell membrane currents and single-channel currents from excised patches. Receptors with the mutant subunit at the asparagine residue on the conserved N-glycosylation site (mbetaN141D, mgammaN141D, or mdeltaN143D) or the serine/threonine residue (mbetaT143A, mgammaS143A, or mdeltaS145A) delayed the rate of current decay as compared with wild-type receptors, and the most striking effect was found with receptors with mbetaT143A or mgammaS143A. For wild-type receptors, the lectin concanavalin A, that binds to glycosylated membrane proteins with high affinity, mimicked this effect. Receptors with mbetaN141D or mdeltaN143D exhibited lower single-channel conductance, but those with mbetaT143A, mgammaS143A, or mdeltaS145A otherwise revealed higher conductance than wild-type receptors. Mean opening time of single-channel currents was little affected by the mutation. N-glycosylation sites, thus, appear to play a role in the regulation of ACh receptor desensitization and ion permeability. PMID:12829329

  17. A New IRAK-M-Mediated Mechanism Implicated in the Anti-Inflammatory Effect of Nicotine via α7 Nicotinic Receptors in Human Macrophages

    PubMed Central

    Maldifassi, Maria C.; Atienza, Gema; Arnalich, Francisco; López-Collazo, Eduardo; Cedillo, Jose L.; Martín-Sánchez, Carolina; Bordas, Anna; Renart, Jaime; Montiel, Carmen

    2014-01-01

    Nicotine stimulation of α7 nicotinic acetylcholine receptor (α7 nAChR) powerfully inhibits pro-inflammatory cytokine production in lipopolysaccharide (LPS)-stimulated macrophages and in experimental models of endotoxemia. A signaling pathway downstream from the α7 nAChRs, which involves the collaboration of JAK2/STAT3 and NF-κB to interfere with signaling by Toll-like receptors (TLRs), has been implicated in this anti-inflammatory effect of nicotine. Here, we identifiy an alternative mechanism involving interleukin-1 receptor-associated kinase M (IRAK-M), a negative regulator of innate TLR-mediated immune responses. Our data show that nicotine up-regulates IRAK-M expression at the mRNA and protein level in human macrophages, and that this effect is secondary to α7 nAChR activation. By using selective inhibitors of different signaling molecules downstream from the receptor, we provide evidence that activation of STAT3, via either JAK2 and/or PI3K, through a single (JAK2/PI3K/STAT3) or two convergent cascades (JAK2/STAT3 and PI3K/STAT3), is necessary for nicotine-induced IRAK-M expression. Moreover, down-regulation of this expression by small interfering RNAs specific to the IRAK-M gene significantly reverses the anti-inflammatory effect of nicotine on LPS-induced TNF-α production. Interestingly, macrophages pre-exposed to nicotine exhibit higher IRAK-M levels and reduced TNF-α response to an additional LPS challenge, a behavior reminiscent of the ‘endotoxin tolerant’ phenotype identified in monocytes either pre-exposed to LPS or from immunocompromised septic patients. Since nicotine is a major component of tobacco smoke and increased IRAK-M expression has been considered one of the molecular determinants for the induction of the tolerant phenotype, our findings showing IRAK-M overexpression could partially explain the known influence of smoking on the onset and progression of inflammatory and infectious diseases. PMID:25259522

  18. Dopamine Receptor Blockade Modulates the Rewarding and Aversive Properties of Nicotine via Dissociable Neuronal Activity Patterns in the Nucleus Accumbens

    PubMed Central

    Sun, Ninglei; Laviolette, Steven R

    2014-01-01

    The mesolimbic pathway comprising the ventral tegmental area (VTA) and projection terminals in the nucleus accumbens (NAc) has been identified as a critical neural system involved in processing both the rewarding and aversive behavioral effects of nicotine. Transmission through dopamine (DA) receptors functionally modulates these effects directly within the NAc. Nevertheless, the neuronal mechanisms within the NAc responsible for these bivalent behavioral effects are presently not known. Using an unbiased conditioned place preference procedure combined with in vivo neuronal recordings, we examined the effects of nicotine reward and aversion conditioning on intra-NAc neuronal sub-population activity patterns. We report that intra-VTA doses of nicotine that differentially produce rewarding or aversive behavioral effects produce opposite effects on sub-populations of fast-spiking interneurons (FSIs) or medium spiny neurons (MSNs) within the shell region of the NAc (NAshell). Thus, while the rewarding effects of intra-VTA nicotine were associated with inhibition of FSI and activation of MSNs, the aversive effects of nicotine produced the opposite pattern of NAshell neuronal population activity. Blockade of DA transmission with a broad-spectrum DA receptor antagonist, α-flupenthixol, strongly inhibited the spontaneous activity of NAshell FSIs, and reversed the conditioning properties of intra-VTA nicotine, switching nicotine-conditioned responses from aversive to rewarding. Remarkably, DA receptor blockade switched intra-NAshell neuronal population activity from an aversion to a reward pattern, concomitant with the observed switch in behavioral conditioning effects. PMID:24896614

  19. Inhibition of human α7 nicotinic acetylcholine receptors by cyclic monoterpene carveol.

    PubMed

    Lozon, Yosra; Sultan, Ahmed; Lansdell, Stuart J; Prytkova, Tatiana; Sadek, Bassem; Yang, Keun-Hang Susan; Howarth, Frank Christopher; Millar, Neil S; Oz, Murat

    2016-04-01

    Cyclic monoterpenes are a group of phytochemicals with antinociceptive, local anesthetic, and anti-inflammatory actions. Effects of cyclic monoterpenes including vanilin, pulegone, eugenole, carvone, carvacrol, carveol, thymol, thymoquinone, menthone, and limonene were investigated on the functional properties of the cloned α7 subunit of the human nicotinic acetylcholine receptor expressed in Xenopus oocytes. Monoterpenes inhibited the α7 nicotinic acetylcholine receptor in the order carveol>thymoquinone>carvacrol>menthone>thymol>limonene>eugenole>pulegone≥carvone≥vanilin. Among the monoterpenes, carveol showed the highest potency on acetylcholine-induced responses, with IC50 of 8.3µM. Carveol-induced inhibition was independent of the membrane potential and could not be reversed by increasing the concentration of acetylcholine. In line with functional experiments, docking studies indicated that cyclic monoterpenes such as carveol may interact with an allosteric site located in the α7 transmembrane domain. Our results indicate that cyclic monoterpenes inhibit the function of human α7 nicotinic acetylcholine receptors, with varying potencies. PMID:26849939

  20. Association of α4β2 nicotinic receptor and heavy smoking in schizophrenia

    PubMed Central

    Voineskos, Sophocles; Luca, Vincenzo De; Mensah, Albert; Vincent, John B.; Potapova, Natalia; Kennedy, James L.

    2007-01-01

    Introduction Previously we suggested that the CHRNA7 polymorphism in nicotinic receptor genes, in particular the D15S1360 in CHRNA7, is associated with smoking in schizophrenia. Schizophrenia patients are usually heavy smokers. In this study we hypothesized that high-affinity nicotinic receptors are associated with smoking in such patients. Objective To investigate the role of α4 (Ch 20) and β2 (Ch 1) genes in conferring a risk for smoking and for smoking a large number of cigarettes daily in subjects with schizophrenia. Methods Our study sample consisted of 241 white European schizophrenia patients (157 smokers and 84 nonsmokers) from the Toronto area. Current smoking status was assessed by the medical history. We investigated 4 markers located in the CHRNA4 gene and 3 markers located in the CHRNB2 gene. Results There was no difference in age or ethnicity between the 2 groups and the population was not stratified (λ = 0.4527). We found a significant association between the CHRNA4 rs3746372 allele 1 and a large number of cigarettes smoked daily (p = 0.0203). The intragenic interaction between rs3787116 and rs3746372 (p = 0.0050) in CHRNA4 showed a significant interaction for the number of cigarettes smoked. Conclusion Although our findings suggest an association between rs3746372 allele 1 and heavy smoking, further study is warranted to investigate the relation between smoking and high-affinity nicotinic receptor genes in schizophrenia. PMID:18043764

  1. Identification, characterization, and regulation of a nicotinic acetylcholine receptor on bovine adrenal chromaffin cells in culture

    SciTech Connect

    Higgins, L.S.

    1988-01-01

    Synaptic input to bovine adrenal chromaffin cells is mediated by nicotinic acetylcholine receptors (AChRs) and results in secretion of catecholamines. Three probes previously shown to recognize AChRs on neurons were used to identify the AChR on bovine adrenal chromaffin cells in culture: monoclonal antibody mAb 35, a toxin that blocks receptor function, and the agonist nicotine. Competition for {sup 3}H-nicotine binding was used to measure the affinity of cholinergic ligands, and revealed the pharmacological profile expected for a neuronal-type AChR. At steady state the rate both of receptor insertion into and loss from the plasma membrane is about 3%/hour, resulting in a half-life in the surface of about 24 hours. Exposure to the anti-AChR antibody results in a loss of AChRs from the surface of the cells through a process that has the characteristics of antigenic modulation. The number of AChRs on the surface of the chromaffin cells can also be modulated by agonists and hormones, including glucocotricoids. Catecholamines, three peptides that may be secreted by chromaffin cells, and K{sup +}-induced secretion reduce agonist-induced catecholamine release by decreasing the number of AChRs, providing a mechanism for autoregulation.

  2. Structural and functional changes induced in the nicotinic acetylcholine receptor by membrane phospholipids.

    PubMed

    Fernández-Carvajal, Asia M; Encinar, José A; Poveda, José Antonio; de Juan, Entilio; Martínez-Pinna, Juan; Ivorra, Isabel; Ferragut, José Antonio; Morales, Andrés; González-Ros, José Manuel

    2006-01-01

    Ligand-gated ion channels (LGICs) constitute an important family of complex membrane proteins acting as receptors for neurotransmitters (Barnard, 1992; Ortells and Lunt, 1995). The nicotinic acetylcholine receptor (nAChR) from Torpedo is the most extensively studied member of the LGIC family and consists of a pentameric transmembrane glycoprotein composed of four different polypeptide subunits (alpha, beta, gamma, and delta) in a 2:1:1:1 stoichiometry (Galzi and Changeux, 1995; Hucho et al., 1996) that are arranged pseudosymmetrically around a central cation-selective ion channel. Conformational transitions, from the closed (nonconducting), to agonist-induced open (ion-conducting), to desensitized (nonconducting) states, are critical for functioning of the nAChR (Karlin, 2002). The ability of the nAChR to undergo these transitions is profoundly influenced by the lipid composition of the bilayer (Barrantes, 2004). Despite existing information on lipid dependence of AChR function, no satisfactory explanation has been given on the molecular events by which specific lipids exert such effects on the activity of an integral membrane protein. To date, several hypotheses have been entertained, including (1) indirect effects of lipids through the alteration of properties of the bilayer, such as fluidity (an optimal fluidity hypothesis [Fong and McNamee, 1986]) or membrane curvature and lateral pressure (Cantor, 1997; de Kruijff, 1997), or (2) direct effects through binding of lipids to defined sites on the transmembrane portion of the protein (Jones and McNamee, 1988; Blanton and Wang, 1990; Fernández et al., 1993; Fernández-Ballester et al., 1994), which has led to the postulation of a possible role of certain lipids as peculiar allosteric ligands of the protein. In this paper we have reconstituted purified AChRs from Torpedo into complex multicomponent lipid vesicles in which the phospholipid composition has been systematically altered. Stopped-flow rapid kinetics of

  3. Autoradiographic localization of putative nicotinic receptors in the rat brain using sup 125 I-neuronal bungarotoxin

    SciTech Connect

    Schulz, D.W.; Loring, R.H.; Aizenman, E.; Zigmond, R.E. )

    1991-01-01

    Neuronal bungarotoxin (NBT), a snake venom neurotoxin, selectively blocks nicotinic receptors in many peripheral and central neuronal preparations. alpha-Bungarotoxin (alpha BT), on the other hand, a second toxin isolated from the venom of the same snake, is an ineffective nicotinic antagonist in most vertebrate neuronal preparations studied thus far. To examine central nicotinic receptors recognized by NBT, we have characterized the binding of 125I-labeled NBT (125I-NBT) to rat brain membranes and have mapped the distribution of 125I-NBT binding in brain sections using quantitative light microscopic autoradiography. The binding of 125I-NBT was found to be saturable, of high affinity, and heterogeneously distributed in the brain. Pharmacological studies suggested that more than one population of sites is labeled by 125I-NBT. For example, one component of 125I-NBT binding was also recognized by alpha BT, while a second component, not recognized by alpha BT, was recognized by the nicotinic agonist nicotine. The highest densities of these alpha BT-insensitive, nicotine-sensitive sites were found in the fasciculus retroflexus, the lateral geniculate nucleus, the medial terminal nucleus of the accessory optic tract, and the olivary pretectal nucleus. alpha BT-sensitive NBT binding sites were found in highest density in the lateral geniculate nucleus, the subthalamic nucleus, the dorsal tegmental nucleus, and the medial mammillary nucleus (lateral part). The number of brain regions with a high density of 125I-NBT binding sites, blocked either by alpha BT or by nicotine, is low when compared with results obtained using other approaches to studying the central distribution of nicotinic receptors, such as labeling with 3H-nicotine or labeling with cDNA probes to mRNAs coding for putative receptor subunits.

  4. Nicotine enhances invasion and metastasis of human colorectal cancer cells through the nicotinic acetylcholine receptor downstream p38 MAPK signaling pathway.

    PubMed

    Xiang, Tao; Fei, Rushan; Wang, Zhe; Shen, Zhonglei; Qian, Jing; Chen, Wenbin

    2016-01-01

    Nicotine as a cigarette component is an established risk factor for colorectal cancer tumorigenesis. The downstream signaling pathways of nicotinic acetylcholine receptors (nAchRs) are believed to be responsible for the cellular effects. In the present study, we evaluated the effects and novel mechanisms for nicotine on the capacity for colorectal cancer cell invasion and metastasis. LOVO and SW620 colorectal cancer cells were stimulated with nicotine in vitro. A Transwell chamber model was applied to detect the capacity for tumor cell invasion. Assays for gelatin zymography and western blotting were applied to detect the activity and expression of metastasis-related matrix metalloproteinases (MMPs), respectively. Signal transduction was assessed by immunoblotting for the phosphorylation of relevant signal molecules and the application of pharmaceutical inhibitors. We showed that nicotine increased LOVO and SW620 colorectal cancer cell invasion along with enhanced activity and expression of MMP-1, -2 and -9. Nicotine increased phosphorylation of p38, ERK, Akt and PI3K p85 but had no effect on phosphorylation of JNK, or NF-κB. Of the pharmaceutical inhibitors of U0126 (ERK1/2 inhibitor), LY294002 (Akt activation inhibitor), SB239063 (p38 MAPK activation inhibitor) and hexamethonium (Hex) (nAchRs inhibitor), the cellular and molecular effects were reduced by the applications of SB239063 and Hex. We concluded that nicotine stimulates the invasion and metastasis of colon cancer cells in vitro via activation of the nAchRs and the p38 MAPK downstream signaling pathway. Therefore, p38 MAPK may have potential as a therapeutic target for smoking-related human colorectal cancer metastasis. PMID:26530054

  5. In vivo pharmacological interactions between a type II positive allosteric modulator of α7 nicotinic ACh receptors and nicotinic agonists in a murine tonic pain model

    PubMed Central

    Freitas, K; Negus, SS; Carroll, FI; Damaj, MI

    2013-01-01

    Background and Purpose The α7 nicotinic ACh receptor subtype is abundantly expressed in the CNS and in the periphery. Recent evidence suggests that α7 nicotinic ACh receptor (nAChR) subtypes, which can be activated by an endogenous cholinergic tone comprising ACh and the α7 agonist choline, play an important role in chronic pain and inflammation. In this study, we evaluated whether type II α7 positive allosteric modulator PNU-120596 induces antinociception on its own and in combination with choline in the formalin pain model. Experimental Approach We assessed the effects of PNU-120596 and choline and the nature of their interactions in the formalin test using an isobolographic analysis. In addition, we evaluated the interaction of PNU-120596 with PHA-54613, an exogenous selective α7 nAChR agonist, in the formalin test. Finally, we assessed the interaction between PNU-120596 and nicotine using acute thermal pain, locomotor activity, body temperature and convulsing activity tests in mice. Key Results We found that PNU-120596 dose-dependently attenuated nociceptive behaviour in the formalin test after systemic administration in mice. In addition, mixtures of PNU-120596 and choline synergistically reduced formalin-induced pain. PNU-120596 enhanced the effects of nicotine and α7 agonist PHA-543613 in the same test. In contrast, PNU-120596 failed to enhance nicotine-induced convulsions, hypomotility and antinociception in acute pain models. Surprisingly, it enhanced nicotine-induced hypothermia via activation of α7 nAChRs. Conclusions and Implications Our results demonstrate that type II α7 positive allosteric modulators produce antinociceptive effects in the formalin test through a synergistic interaction with the endogenous α7 agonist choline. PMID:23004024

  6. A comparative study of the effects of the intravenous self-administration or subcutaneous minipump infusion of nicotine on the expression of brain neuronal nicotinic receptor subtypes.

    PubMed

    Moretti, Milena; Mugnaini, Manolo; Tessari, Michela; Zoli, Michele; Gaimarri, Annalisa; Manfredi, Irene; Pistillo, Francesco; Clementi, Francesco; Gotti, Cecilia

    2010-08-01

    Long-term nicotine exposure changes neuronal acetylcholine nicotinic receptor (nAChR) subtype expression in the brains of smokers and experimental animals. The aim of this study was to investigate nicotine-induced changes in nAChR expression in two models commonly used to describe the effects of nicotine in animals: operant (two-lever presses) intravenous self-administration (SA) and passive subcutaneous nicotine administration via an osmotic minipump (MP). In the MP group, alpha4beta2 nAChRs were up-regulated in all brain regions, alpha6beta2* nAChRs were down-regulated in the nucleus accumbens (NAc) and caudate-putamen, and alpha7 nAChRs were up-regulated in the caudal cerebral cortex (CCx); the up-regulation of alpha4beta2alpha5 nAChRs in the CCx was also suggested. In the SA group, alpha4beta2 up-regulation was lower and limited to the CCx and NAc; there were no detectable changes in alpha6beta2* or alpha7 nACRs. In the CCx of the MP rats, there was a close correlation between the increase in alpha4beta2 binding and alpha4 and beta2 subunit levels measured by means of Western blotting, demonstrating that the up-regulation was due to an increase in alpha4beta2 proteins. Western blotting also showed that the increase in the beta2 subunit exceeded that of the alpha4 subunit, suggesting that a change in alpha4beta2 stoichiometry may occur in vivo as has been shown in vitro. These results show that nicotine has an area-specific effect on receptor subtypes, regardless of its administration route, but the effect is quantitatively greater in the case of MP administration. PMID:20439469

  7. NeuroD1 mediates nicotine-induced migration and invasion via regulation of the nicotinic acetylcholine receptor subunits in a subset of neural and neuroendocrine carcinomas

    PubMed Central

    Osborne, Jihan K.; Guerra, Marcy L.; Gonzales, Joshua X.; McMillan, Elizabeth A.; Minna, John D.; Cobb, Melanie H.

    2014-01-01

    Cigarette smoking is a major risk factor for acquisition of small cell lung cancer (SCLC). A role has been demonstrated for the basic helix-loop-helix transcription factor NeuroD1 in the pathogenesis of neural and neuroendocrine lung cancer, including SCLC. In the present study we investigate the possible function of NeuroD1 in established tumors, as well as actions early on in pathogenesis, in response to nicotine. We demonstrate that nicotine up-regulates NeuroD1 in immortalized normal bronchial epithelial cells and a subset of undifferentiated carcinomas. Increased expression of NeuroD1 subsequently leads to regulation of expression and function of the nicotinic acetylcholine receptor subunit cluster of α3, α5, and β4. In addition, we find that coordinated expression of these subunits by NeuroD1 leads to enhanced nicotine-induced migration and invasion, likely through changes in intracellular calcium. These findings suggest that aspects of the pathogenesis of neural and neuroendocrine lung cancers may be affected by a nicotine- and NeuroD1-induced positive feedback loop. PMID:24719457

  8. Adolescent nicotine-induced dendrite remodeling in the nucleus accumbens is rapid, persistent, and D1-dopamine receptor dependent.

    PubMed

    Ehlinger, D G; Bergstrom, H C; Burke, J C; Fernandez, G M; McDonald, C G; Smith, R F

    2016-01-01

    Chronic nicotine exposure during adolescence induces dendritic remodeling of medium spiny neurons (MSNs) in the nucleus accumbens (NAcc) shell. While nicotine-induced dendritic remodeling has frequently been described as persistent, the trajectory of dendrite remodeling is unknown. Specifically, no study to date has characterized the structural plasticity of dendrites in the NAcc immediately following chronic nicotine, leaving open the possibility that dendrite remodeling emerges gradually over time. Further, the neuropharmacological mechanisms through which nicotine induces dendrite remodeling are not well understood. To address these questions, rats were co-administered chronic nicotine (0.5 mg/kg) and the D1-dopamine receptor (D1DR) antagonist SCH-23390 (0.05 mg/kg) subcutaneously every other day during adolescence. Brains were then processed for Golgi-Cox staining either 1 day or 21 days following drug exposure and dendrites from MSNs in the NAcc shell digitally reconstructed in 3D. Spine density was also measured at both time points. Our morphometric results show (1) the formation of new dendritic branches and spines 1 day following nicotine exposure, (2) new dendritic branches, but not spine density, remains relatively stable for at least 21 days, (3) the co-administration of SCH-23390 completely blocked nicotine-induced dendritic remodeling of MSNs at both early and late time points, suggesting the formation of new dendritic branches in response to nicotine is D1DR-dependent, and (4) SCH-23390 failed to block nicotine-induced increases in spine density. Overall this study provides new insight into how nicotine influences the normal trajectory of adolescent brain development and demonstrates a persistent form of nicotine-induced neuroplasticity in the NAcc shell that develops rapidly and is D1DR dependent. PMID:25257604

  9. Presynaptic α4β2 nicotinic acetylcholine receptors increase glutamate release and serotonin neuron excitability in the dorsal raphe nucleus.

    PubMed

    Garduño, Julieta; Galindo-Charles, Luis; Jiménez-Rodríguez, Javier; Galarraga, Elvira; Tapia, Dagoberto; Mihailescu, Stefan; Hernandez-Lopez, Salvador

    2012-10-24

    Several behavioral effects of nicotine are mediated by changes in serotonin (5-HT) release in brain areas that receive serotonergic afferents from the dorsal raphe nucleus (DRN). In vitro experiments have demonstrated that nicotine increases the firing activity in the majority of DRN 5-HT neurons and that DRN contains nicotinic acetylcholine receptors (nAChRs) located at both somata and presynaptic elements. One of the most common presynaptic effects of nicotine is to increase glutamate release. Although DRN receives profuse glutamatergic afferents, the effect of nicotine on glutamate release in the DRN has not been studied in detail. Using whole-cell recording techniques, we investigated the effects of nicotine on the glutamatergic input to 5-HT DRN neurons in rat midbrain slices. Low nicotine concentrations, in the presence of bicuculline and tetrodotoxin (TTX), increased the frequency but did not change the amplitude of glutamate-induced EPSCs, recorded from identified 5-HT neurons. Nicotine-induced increase of glutamatergic EPSC frequency persisted 10-20 min after drug withdrawal. This nicotinic effect was mimicked by exogenous administration of acetylcholine (ACh) or inhibition of ACh metabolism. In addition, the nicotine-induced increase in EPSC frequency was abolished by blockade of α4β2 nAChRs, voltage-gated calcium channels, or intracellular calcium signaling but not by α7 nAChR antagonists. These data suggest that both nicotine and endogenous ACh can increase glutamate release through activation of presynaptic α4β2 but not α7 nAChRs in the DRN. The effect involves long-term changes in synaptic function, and it is dependent on voltage-gated calcium channels and presynaptic calcium stores. PMID:23100436

  10. Adenosine (A)(2A)receptor modulation of nicotine-induced locomotor sensitization. A pharmacological and transgenic approach.

    PubMed

    Jastrzębska, Joanna; Nowak, Ewa; Smaga, Irena; Bystrowska, Beata; Frankowska, Małgorzata; Bader, Michael; Filip, Małgorzata; Fuxe, Kjell

    2014-06-01

    Preclinical evidence indicates an important role of adenosine (A)(2A) receptors in drug addiction while their therapeutic relevance is still a matter of debate. We examined the influence of the A(2A) receptor agonist CGS 21680 and the antagonist KW 6002 on nicotine sensitization and conditioned locomotor activity in adult (8-week old) male Sprague-Dawley rats (WT). Moreover, behavioral responses to nicotine were studied in rats overexpressing A(2A) receptors under the control of the neuronal specific enolase (NSE) promotor. Changes in the levels of dopamine, glutamate and γ-aminobutyric acid in wild type (WT) and NSEA(2A) rats were determined with using LC-MS. KW 6002 significantly enhanced expression of nicotine sensitization and conditioned locomotion, while CGS 21680 reduced all these effects in WT rats. A reduction of the expression of nicotine-evoked conditioned locomotor activity was also observed in the NSEA(2A) animals. The transgenic rats displayed a reduced basal tissue level of glutamate in the prefrontal cortex and hippocampus while dopamine basal levels in the nucleus accumbens were raised. Chronic nicotine treatment caused a significant reduction in the glutamate tissue level in the dorsal and ventral striatum, prefrontal cortex and cerebellum in wild type rats. In NSEA(2A) animals the same drug treatment instead produced a rise of glutamate levels in the hippocampus and dorsal striatum. Taken together, A(2A) receptor signaling in the rat brain can counteract locomotor sensitization and conditioned locomotion to nicotine which are related to nicotine reward-learning. It is suggested that treatment with A(2A) receptor agonists can help counteract the abuse actions of nicotine. PMID:24632528