Skip to main content
SpringerLink
Log in

Nicotinic Acetylcholine Receptors as Targets for Tobacco Cessation Therapeutics: Cutting-Edge Methodologies to Understand Receptor Assembly and Trafficking

  • Protocol
  • First Online:
Nicotinic Acetylcholine Receptor Technologies

Part of the book series: Neuromethods ((NM,volume 117))

Abstract

Tobacco dependence is a chronic relapsing disorder and nicotine, the primary alkaloid in tobacco, acts at nicotinic receptors to stimulate dopamine release in brain, which is responsible for the reinforcing properties of nicotine, leading to addiction. Although the majority of tobacco users express the desire to quit, only a small percentage of those attempting to quit are successful using the currently available pharmacotherapies. Nicotine upregulates the number of specific nicotinic receptors on the neuronal cell surface. An increase in receptor trafficking or preferential stoichiometric assembly of receptor subunits involves changes in assembly, endoplasmic reticulum export, vesicle transport, decreased degradation, desensitization, enhanced maturation of functional pentamers, and pharmacological chaperoning. Understanding these changes on a mechanistic level is important to the development of nicotinic receptors as drug targets. For this reason, cutting-edge methodologies are being developed and employed to pinpoint distinct changes in localization, assembly, export, vesicle trafficking, and stoichiometry in order to further understand the physiology of these receptors and to evaluate the action of novel therapeutics for smoking cessation.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. George TP, O’Malley SS (2004) Current pharmacological treatments for nicotine dependence. Trends Pharmacol Sci 25:42–48

    Article  CAS  PubMed  Google Scholar 

  2. Le Foll B, Goldberg SR (2009) Effects of nicotine in experimental animals and humans: an update on addictive properties. Handb Exp Pharmacol 192:335–367

    Article  PubMed  Google Scholar 

  3. Rose JE (2008) Disrupting nicotine reinforcement from cigarette to brain. Ann N Y Acad Sci 1141:233–256

    Article  PubMed  Google Scholar 

  4. Benowitz NL (2010) Nicotine addiction. N Engl J Med 362:2295–2303

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Hajek P, McRobbie H, Myers K (2013) Efficacy of cytisine in helping smokers quit: systematic review and meta-analysis. Thorax 68:1037–1042

    Article  PubMed  Google Scholar 

  6. Hajek P, Stead LF, West R, Jarvis M, Hartmann-Boyce J, Lancaster T (2013) Relapse prevention interventions for smoking cessation. Cochrane Database Syst Rev (8):CD003999.

    Google Scholar 

  7. Albuquerque EX, Pereira EF, Alkondon M, Rogers SW (2009) Mammalian nicotinic acetylcholine receptors: from structure to function. Physiol Rev 89:73–120

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Taly A, Corringer PJ, Guedin D, Lestage P, Changeux JP (2009) Nicotinic receptors: allosteric transitions and therapeutic targets in the nervous system. Nat Rev Drug Discov 8:733–750

    Article  CAS  PubMed  Google Scholar 

  9. Dani JA, Bertrand D (2007) Nicotinic acetylcholine receptors and nicotinic cholinergic mechanisms of the central nervous system. Annu Rev Pharmacol Toxicol 47:699–729

    Article  CAS  PubMed  Google Scholar 

  10. De Biasi M (2002) Nicotinic mechanisms in the autonomic control of organ systems. J Neurobiol 53:568–579

    Article  PubMed  Google Scholar 

  11. Govind AP, Vezina P, Green WN (2009) Nicotine-induced upregulation of nicotinic receptors: underlying mechanisms and relevance to nicotine addiction. Biochem Pharmacol 78:756–765

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Millar NS, Harkness PC (2008) Assembly and trafficking of nicotinic acetylcholine receptors. Mol Membr Biol 25:279–292

    Article  CAS  PubMed  Google Scholar 

  13. Flores CM, Rogers SW, Pabreza LA, Wolfe BB, Kellar KJ (1992) A subtype of nicotinic cholinergic receptor in rat brain is composed of α4 and β2 subunits and is up-regulated by chronic nicotine treatment. Mol Pharmacol 41:31–37

    CAS  PubMed  Google Scholar 

  14. Gotti C, Zoli M, Clementi F (2006) Brain nicotinic acetylcholine receptors: native subtypes and their relevance. Trends Pharmacol Sci 27:482–491

    Article  CAS  PubMed  Google Scholar 

  15. McIntosh JM, Plazas PV, Watkins M, Gomez-Casati ME, Olivera BM, Elgoyhen AB (2005) A novel α-conotoxin, PeIA, cloned from Conus pergrandis, discriminates between rat α9α10 and α7 nicotinic cholinergic receptors. J Biol Chem 28:30107–30112

    Article  Google Scholar 

  16. Wada E, Wada K, Boulter J, Deneris E, Heinemann S, Patrick J, Swanson LW (1989) Distribution of alpha2, alpha3, alpha4, and beta2 neuronal nicotinic receptor subunit mRNAs in the central nervous system: A hybridization histochemical study in the rat. J Comp Neurol 284:314–335

    Article  CAS  PubMed  Google Scholar 

  17. Srinivasan R, Henderson BJ, Lester HA, Richards CI (2014) Pharmacological chaperoning of nAChRs: a therapeutic target for Parkinson's disease. Pharmacol Res 83:20–29

    Article  CAS  PubMed  Google Scholar 

  18. Tapia L, Kuryatov A, Lindstrom J (2007) Ca2+ permeability of the (α4)3(β2)2 stoichiometry greatly exceeds that of (α4)2(β2)3 human acetylcholine receptors. Mol Pharmacol 71:769–776

    Article  CAS  PubMed  Google Scholar 

  19. Fox AM, Moonschi FH, Richards CI (2015) The nicotine metabolite, cotinine, alters the assembly and trafficking of a subset of nicotinic acetylcholine receptors. J Biol Chem 290:24403–24412

    Article  CAS  PubMed  Google Scholar 

  20. Henderson BJ, Srinivasan R, Nichols WA, Dilworth CN, Gutierrez DF, Mackey EDW, McKinney S, Drenan RM, Richards CI, Lester HA (2014) Nicotine exploits a COPI-mediated process for chaperone-mediated up-regulation of its receptors. J Gen Physiol 143:51–66

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Srinivasan R, Richards CI, Dilworth C, Moss FJ, Dougherty DA, Lester HA (2012) Forster resonance energy transfer (FRET) correlates of altered subunit stoichiometry in Cys-loop receptors, exemplified by nicotinic alpha 4 beta 2. Int J Mol Sci 13:10022–10040

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Richards CI, Luong K, Srinivasan R, Turner SW, Dougherty DA, Korlach J, Lester HA (2012) Live-cell imaging of single receptor composition using zero-mode waveguide nanostructures. Nano Lett 12:3690–3694

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Richards CI, Srinivasan R, Xiao C, Mackey EDW, Miwa JM, Lester HA (2011) Trafficking of α4* nicotinic receptors revealed by superecliptic phluorin. J Biol Chem 286:31241–31249

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Colombo SF, Mazzo F, Pistillo F, Gotti C (2013) Biogenesis, trafficking and up-regulation of nicotinic ACh receptors. Biochem Pharmacol 86:1063–1073

    Article  CAS  PubMed  Google Scholar 

  25. Marks MJ, Burch JB, Collins AC (1983) Effects of chronic nicotine infusion on tolerance development and nicotinic receptors. J Pharmacol Exp Ther 226:817–825

    CAS  PubMed  Google Scholar 

  26. Marks MJ, Grady SR, Salminen O, Paley MA, Wageman CR, McIntosh JM, Whiteaker P (2014) alpha6beta2*-subtype nicotinic acetylcholine receptors are more sensitive than alpha4beta2*-subtype receptors to regulation by chronic nicotine administration. J Neurochem 130:185–198

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Lester HA, Xiao C, Srinivasan R, Son CD, Miwa J, Pantoja R, Banghart MR, Dougherty DA, Goate AM, Wang JC (2009) Nicotine is a selective pharmacological chaperone of acetylcholine receptor number and stoichiometry. Implications for drug discovery. Am Assoc Pharm Scient J 11:167–177

    CAS  Google Scholar 

  28. Miwa JM, Freedman R, Lester HA (2011) Neural systems governed by nicotinic acetylcholine receptors: emerging hypotheses. Neuron 70:20–33

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Stolerman IP, Jarvis MJ (1995) The scientific case that nicotine is addictive. Psychopharmacology (Berl) 117:2–10

    Article  CAS  Google Scholar 

  30. Pauly JR, Marks MJ, Robinson SF, van de Kamp JL, Collins AC (1996) Chronic nicotine and mecamylamine treatment increase brain nicotinic receptor binding without changing α4 or β2 mRNA levels. J Pharmacol Exp Ther 278:361–369

    CAS  PubMed  Google Scholar 

  31. Changeux JP (2010) Nicotine addiction and nicotinic receptors: lessons from genetically modified mice. Nat Rev Neurosci 11:389–401

    Article  CAS  PubMed  Google Scholar 

  32. Turner JR, Castellano LM, Blendy JA (2011) Parallel anxiolytic-like effects and upregulation of neuronal nicotinic acetylcholine receptors following chronic nicotine and varenicline. Nicotine Tob Res 13:41–46

    Article  CAS  PubMed  Google Scholar 

  33. Whiting P, Lindstrom J (1987) Purification and characterization of a nicotinic acetylcholine receptor from rat brain. Proc Natl Acad Sci U S A 84:595–599

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Zoli M, Lena C, Picciotto MR, Changeux JP (1998) Identification of four classes of brain nicotinic receptors using beta2 mutant mice. J Neurosci 18:4461–4472

    CAS  PubMed  Google Scholar 

  35. Lukas R (2007) Phamacological effects of nicotine and nicotinic receptor subtype pharmacological profiles. CRC Press LLC, Boca Raton, FL

    Google Scholar 

  36. Etter JF (2006) Cytisine for smoking cessation: a literature review and a meta-analysis. Arch Intern Med 166:1553–1559

    Article  CAS  PubMed  Google Scholar 

  37. Zatonski WCM, Tutka P, West R (2006) An uncontrolled trial of cytisine (Tabex) for smoking cessation. Tob Control 15:481–484

    Article  PubMed  PubMed Central  Google Scholar 

  38. Mihalak KB, Carroll FI, Luetje CW (2006) Varenicline is a partial agonist at alpha4beta2 and a full agonist at alpha7 neuronal nicotinic receptors. Mol Pharmacol 70:801–805

    Article  CAS  PubMed  Google Scholar 

  39. Cohen C, Bergis OE, Galli F, Lochead AW, Jegham S, Biton B, Leonardon J, Avenet P, Sgard F, Besnard F, Graham D, Coste A, Oblin A, Curet O, Voltz C, Gardes A, Caille D, Perrault G, George P, Soubrie P, Scatton B (2003) SSR591813, a novel selective and partial alpha4beta2 nicotinic receptor agonist with potential as an aid to smoking cessation. J Pharmacol Exp Ther 306:407–420

    Article  CAS  PubMed  Google Scholar 

  40. Fagerström K, Balfour DJ (2006) Neuropharmacology and potential efficacy of new treatments for tobacco dependence. Expert Opin Investig Drugs 15:107–116

    Article  PubMed  Google Scholar 

  41. Xiao Y, Fan H, Musachio JL, Wei ZL, Chellappan SK, Kozikowski AP, Kellar KJ (2006) Sazetidine-A, a novel ligand that desensitizes alpha4beta2 nicotinic acetylcholine receptors without activating them. Mol Pharmacol 70:1454–1460

    Article  CAS  PubMed  Google Scholar 

  42. Zwart RCA, Moroni M, Bermudez I, Mogg AJ, Folly EA, Broad LM, Williams AC, Zhang D, Ding C, Heinz BA, Sher E (2008) Sazetidine-A is a potent and selective agonist at native and recombinant alpha 4 beta 2 nicotinic acetylcholine receptors. Mol Pharmacol 73:1838–1843

    Article  CAS  PubMed  Google Scholar 

  43. Nelson ME, Kuryatov A, Choi CH, Zhou Y, Lindstrom J (2003) Alternate stoichiometries of α4β2 nicotinic acetylcholine receptors. Mol Pharmacol 63:332–341

    Article  CAS  PubMed  Google Scholar 

  44. Marotta CB, Rreza I, Lester HA, Dougherty DA (2014) Selective ligand behaviors provide new insights into agonist activation of nicotinic acetylcholine receptors. ACS Chem Biol 9:1153–1159

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Mazzaferro S, Benallegue N, Carbone A, Gasparri F, Vijayan R, Biggin PC, Moroni M, Bermudez I (2011) Additional acetylcholine (ACh) binding site at α4/α4 interface of (α4β2)2α4 nicotinic receptor influences agonist sensitivity. J Biol Chem 286:31043–31054

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Moroni M, Zwart R, Sher E, Cassels BK, Bermudez I (2006) Alpha4beta2 nicotinic receptors with high and low acetylcholine sensitivity: pharmacology, stoichiometry, and sensitivity to long-term exposure to nicotine. Mol Pharmacol 70:755–768

    Article  CAS  PubMed  Google Scholar 

  47. Tumkosit P, Kuryatov A, Luo J, Lindstrom J (2006) Beta3 subunits promote expression and nicotine-induced up-regulation of human nicotinic alpha6* nicotinic acetylcholine receptors expressed in transfected cell lines. Mol Pharmacol 70:1358–1368

    Article  CAS  PubMed  Google Scholar 

  48. Perez XA, Bordia T, McIntosh JM, Grady SR, Quik M (2008) Long-term nicotine treatment differentially regulates striatal alpha6alpha4beta2* and alpha6(nonalpha4)beta2* nAChR expression and function. Mol Pharmacol 74:844–853

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Walsh H, Govind AP, Mastro R, Hoda JC, Bertrand D, Vallejo Y, Green WN (2008) Up-regulation of nicotinic receptors by nicotine varies with receptor subtype. J Biol Chem 283:6022–6032

    Article  CAS  PubMed  Google Scholar 

  50. Perry DC, Mao D, Gold AB, McIntosh JM, Pezzullo JC, Kellar KJ (2007) Chronic nicotine differentially regulates alpha6- and beta3-containing nicotinic cholinergic receptors in rat brain. J Pharmacol Exp Ther 322:306–315

    Article  CAS  PubMed  Google Scholar 

  51. Picciotto MR, Corrigall WA (2002) Neuronal systems underlying behaviors related to nicotine addiction: neural circuits and molecular genetics. J Neurosci 22:3338–3341

    CAS  PubMed  Google Scholar 

  52. Wise RA, Rompre PP (1989) Brain dopamine and reward. Annu Rev Psychol 40:191–225

    Article  CAS  PubMed  Google Scholar 

  53. Salminen O, Murphy KL, McIntosh JM, Drago J, Marks MJ, Collins AC, Grady SR (2004) Subunit composition and pharmacology of two classes of striatal presynaptic nicotinic acetylcholine receptors mediating dopamine release in mice. Mol Pharmacol 65:1526–1535

    Article  CAS  PubMed  Google Scholar 

  54. Gotti C, Moretti M, Clementi F, Riganti L, McIntosh JM, Collins AC, Marks MJ, Whiteaker P (2005) Expression of nigrostriatal alpha 6-containing nicotinic acetylcholine receptors is selectively reduced, but not eliminated, by beta 3 subunit gene deletion. Mol Pharmacol 67:2007–2015

    Article  CAS  PubMed  Google Scholar 

  55. De Biasi M, McLaughlin I, Perez EE, Crooks PA, Dwoskin LP, Bardo MT, Pentel PR, Hatsukami D (2014) Scientific overview: 2013 BBC plenary symposium on tobacco addiction. Drug Alcohol Depend 141:107–117

    Article  PubMed  PubMed Central  Google Scholar 

  56. Patterson GH, Knobel SM, Sharif WD, Kain SR, Piston DW (1997) Use of the green fluorescent protein and its mutants in quantitative fluorescence microscopy. Biophys J 73:2782–2790

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Shih PY, Engle SE, Oh G, Deshpande P, Puskar NL, Lester HA, Drenan RM (2014) Differential expression and function of nicotinic acetylcholine receptors in subdivisions of medial habenula. J Neurosci 34:9789–9802

    Article  PubMed  PubMed Central  Google Scholar 

  58. Yudowski GA, Puthenveedu MA, Leonoudakis D, Panicker S, Thorn KS, Beattie EC, von Zastrow M (2007) Real-time imaging of discrete exocytic events mediating surface delivery of AMPA receptors. J Neurosci 27:11112–11121

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Lin DT, Makino Y, Sharma K, Hayashi T, Neve R, Takamiya K, Huganir RL (2009) Regulation of AMPA receptor extrasynaptic insertion by 4.1N, phosphorylation and palmitoylation. Nat Neurosci 12:879–887

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Mattheyses AL, Simon SM, Rappoport JZ (2010) Imaging with total internal reflection fluorescence microscopy for the cell biologist. J Cell Sci 123:3621–3628

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Paroutis P, Touret N, Grinstein S (2004) The pH of the secretory pathway: measurement, determinants, and regulation. Physiology 19:207–215

    Article  CAS  PubMed  Google Scholar 

  62. Khiroug S, Pryazhnikov E, Coleman S, Jeromin A, Keinanen K, Khiroug L (2009) Dynamic visualization of membrane-inserted fraction of pHluorin-tagged channels using repetitive acidification technique. BMC Neurosci 10:141

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This research was supported by NIH grants DA03881, DA16176, and TR000117.

Author information

Authors and Affiliations

  1. Department of Chemistry, College of Pharmacy, University of Kentucky, 789 South Limestone, 465 Bio Pharm, Lexington, KY, 40536-0596, USA

    Ashley M. Fox-Loe & Christopher I. Richards

  2. Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone, 465 Bio Pharm, Lexington, KY, 40536-0596, USA

    Linda P. Dwoskin Ph.D.

Authors
  1. Ashley M. Fox-Loe
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Linda P. Dwoskin Ph.D.
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Christopher I. Richards
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Linda P. Dwoskin Ph.D. .

Editor information

Editors and Affiliations

  1. University of Virginia, Charlottesville, Virginia, USA

    Ming D. Li

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Fox-Loe, A.M., Dwoskin, L.P., Richards, C.I. (2016). Nicotinic Acetylcholine Receptors as Targets for Tobacco Cessation Therapeutics: Cutting-Edge Methodologies to Understand Receptor Assembly and Trafficking. In: Li, M. (eds) Nicotinic Acetylcholine Receptor Technologies. Neuromethods, vol 117. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3768-4_7

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3768-4_7

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3766-0

  • Online ISBN: 978-1-4939-3768-4

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation