Abstract

This project tests two hypotheses: (A) Most of the acute effects of nicotine, and most of the addictive effects of chronic application, arise from actions on nicotinic receptors containing the alpha4 subunit (termed alpha4* receptors). (B). Addiction arises in part from changes in the number and distribution of alpha4* receptors. A series of knock-in mouse strains serve as appropriate tools to test these hypotheses. Some strains harbor hypersensitive alpha4 receptor subunits, which isolate and amplify nicotine responses evoked by alpha4* receptors. The most promising strain is the neo-deleted Leu9'Ala strain. This project characterizes cellular and circuit aspects of responses to nicotine, primarily in the VTA-nucleus accumbens system and forebrain of the hypersensitive mice. Most experiments are performed with acute application of nicotine as well as with tissue from animals chronically exposed to nicotine. Leu9'Ala and WT strains are characterized neurochemically, including epibatidine binding in brain homogenates, nicotine-induced release of dopamine, GABA, and glutamate, and tyrosine hydroxylase staining in midbrain. Time-resolved cellular aspects of nicotinic responses and of synaptic transmission are studied, in midbrain, forebrain, and hindbrain. These studies are enhanced by crossing the hypersensitive mice with strains that have GFP-labeled interneurons and dopaminergic neurons: whole-cell patch in transfected cells and cultures; electrode arrays in slices; single-unit and field potential recording in awake, behaving animals; and multi-tetrode single-unit recording. The project characterizes nicotine-induced immediate early genes, primarily fos, in baseline and nicotine-dependent states, using unbiased stereology. Gene expression is analyzed with microarrays, to determine differences between mutant WT mice and nicotine-induced changes. The project also characterizes knock-in mice that have fluorescently labeled alpha4 receptors, both normally sensitive and hypersensitive. Receptor assembly and localization is studied, including FRET, in naive and nicotine-treated states.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Project (R01)
Project #
5R01DA017279-02
Application #
6920049
Study Section
Neurotransporters, Receptors, and Calcium Signaling Study Section (NTRC)
Program Officer
Lin, Yu
Project Start
2004-07-15
Project End
2009-06-30
Budget Start
2005-07-01
Budget End
2006-06-30
Support Year
2
Fiscal Year
2005
Total Cost
$364,500
Indirect Cost

  Institution

Name
California Institute of Technology
Department
Type
Schools of Arts and Sciences
DUNS #
009584210
City
Pasadena
State
CA
Country
United States
Zip Code
91125

  Publications

Henley, Beverley M; Cohen, Bruce N; Kim, Charlene H et al. (2017) Reliable Identification of Living Dopaminergic Neurons in Midbrain Cultures Using RNA Sequencing and TH-promoter-driven eGFP Expression. J Vis Exp :
Henderson, Brandon J; Wall, Teagan R; Henley, Beverley M et al. (2016) Menthol Alone Upregulates Midbrain nAChRs, Alters nAChR Subtype Stoichiometry, Alters Dopamine Neuron Firing Frequency, and Prevents Nicotine Reward. J Neurosci 36:2957-74
Xiao, Cheng; Miwa, Julie M; Henderson, Brandon J et al. (2015) Nicotinic receptor subtype-selective circuit patterns in the subthalamic nucleus. J Neurosci 35:3734-46
Henderson, Brandon J; Lester, Henry A (2015) Inside-out neuropharmacology of nicotinic drugs. Neuropharmacology 96:178-93
Henderson, Brandon J; Srinivasan, Rahul; Nichols, Weston A et al. (2014) Nicotine exploits a COPI-mediated process for chaperone-mediated up-regulation of its receptors. J Gen Physiol 143:51-66
Marotta, Christopher B; Dilworth, Crystal N; Lester, Henry A et al. (2014) Probing the non-canonical interface for agonist interaction with an ?5 containing nicotinic acetylcholine receptor. Neuropharmacology 77:342-9
Marotta, Christopher B; Rreza, Iva; Lester, Henry A et al. (2014) Selective ligand behaviors provide new insights into agonist activation of nicotinic acetylcholine receptors. ACS Chem Biol 9:1153-9
Frazier, Shawnalea J; Cohen, Bruce N; Lester, Henry A (2013) An engineered glutamate-gated chloride (GluCl) channel for sensitive, consistent neuronal silencing by ivermectin. J Biol Chem 288:21029-42
O'Neill, Heidi C; Laverty, Duncan C; Patzlaff, Natalie E et al. (2013) Mice expressing the ADNFLE valine 287 leucine mutation of the ?2 nicotinic acetylcholine receptor subunit display increased sensitivity to acute nicotine administration and altered presynaptic nicotinic receptor function. Pharmacol Biochem Behav 103:603-21
Henley, Beverley M; Williams, Brian A; Srinivasan, Rahul et al. (2013) Transcriptional regulation by nicotine in dopaminergic neurons. Biochem Pharmacol 86:1074-83

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