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Fluorescence microscopy of nicotinic acetylcholine receptors

Citation

Dilworth, Crystal Noelle (2014) Fluorescence microscopy of nicotinic acetylcholine receptors. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z96D5QZB. http://resolver.caltech.edu/CaltechTHESIS:05282014-200718606

Abstract

Neuronal nicotinic acetylcholine receptors (nAChRs) are pentameric ligand gated ion channels abundantly expressed in the central nervous system. Changes in the assembly and trafficking of nAChRs are pertinent to disease states including nicotine dependence, autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE), and Parkinson’s disease (PD). Here we investigate the application of high resolution fluorescence techniques for the study of nAChR assembly and trafficking. We also describe the construction and validation of a fluorescent α5 subunit and subsequent experiments to elucidate the cellular mechanisms through which α5 subunits are expressed, assembled into mature receptors, and trafficked to the cell surface. The effects of a known single nucleotide polymorphism (D398N) in the intracellular loop of α5 are also examined.

Additionally, this report describes the development of a combined total internal reflection fluorescence (TIRF) and lifetime imaging (FLIM) technique and the first application of this methodology for elucidation of stochiometric composition of nAChRs. Many distinct subunit combinations can form functional receptors. Receptor composition and stoichiometry confers unique biophysical and pharmacological properties to each receptor sub-type. Understanding the nature of assembly and expression of each receptor subtype yields important information about the molecular processes that may underlie the mechanisms through which nAChR contribute to disease and addiction states.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:nicotine, neuroscience, addiction, fluorescence, microscopy
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Chemistry
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Lester, Henry A.
Thesis Committee:
  • Dougherty, Dennis A. (chair)
  • Lester, Henry A.
  • Rees, Douglas C.
  • Dervan, Peter B.
Defense Date:21 November 2013
Funders:
Funding AgencyGrant Number
NIH1F31DA028038
Record Number:CaltechTHESIS:05282014-200718606
Persistent URL:http://resolver.caltech.edu/CaltechTHESIS:05282014-200718606
DOI:10.7907/Z96D5QZB
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:8415
Collection:CaltechTHESIS
Deposited By: Crystal Dilworth
Deposited On:29 May 2014 23:34
Last Modified:17 Jun 2016 23:09

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