Neuronal nicotinic acetylcholine receptors (nAChRs) are expressed in both the periperhal and central nervous systems, and are involved in pre-, post-, and non-synaptic control of neuronal activation. In the brain, these receptors play an important role in a variety of physiological processes such as cognition, development, learning, and memory formation. Malfunction of these receptors have been implicated in neurodegenerative diseases like Alzheimer's disease (AD), schizophrenia, and Parkinson's disease. To date, 17 different nAChR subunits, including α2-α7 and β2-β4, have been cloned that can form homo- and/or hetero-pentameric ionotropic receptors. The unique combinations of subunit pentamers manifest in distinct functional receptors. Using single-cell real-time quantitative RT-PCR, we identified the individual expression rates and co-expression rates of the different nAChR subunits in rat CA1 hippocampal interneurons in efforts to characterize functional receptors involved in learning and memory. The two-way combination of subunits with highest expression in hippocampal interneurons was α3β2. Moreover, this combination was expressed in ratios near 1:3 or 3:1 α3 to β2 respectively. To investigate the functionality of α3β2 receptors in different stoichiometries, we injected human α3 and rat β2 subunit mRNA in 1:3, 1:1, and 3:1 ratios into Xenopus laevis oocytes for expression. Two-electrode voltage clamp was then performed with the application of different concentrations of ACh to produce full dose-response curves and channel kinetics data. Distinct α3β2 functional channels were identified from the different expression ratios based on significant differences in channel kinetics (i.e.- peak current rise times, peak current decay times, steady state current in forced desensitization) Dose-response curves produced no significant difference in EC50 values in the different expression groups. However, there was a trend to greater agonist sensitivity with increased α3 expression relative to β2. α3β2 receptors were further characterized through forced desensitization of the receptors and generation of IV plots. The findings from this study elucidate the neuronal nAChR subunit combinations that form functional channels in hippocampal interneurons.
| Identifer | oai:union.ndltd.org:BGMYU2/oai:scholarsarchive.byu.edu:etd-2766 |
| Date | 17 July 2008 |
| Creators | Mizukawa, John Hideo |
| Publisher | BYU ScholarsArchive |
| Source Sets | Brigham Young University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Theses and Dissertations |
| Rights | http://lib.byu.edu/about/copyright/ |
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