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Structural determinants and regulation of spontaneous activity in GABAA receptors

Sexton, Craig Alexander; (2020) Structural determinants and regulation of spontaneous activity in GABAA receptors. Doctoral thesis (Ph.D), UCL (University College London). Green open access

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Abstract

Rapid inhibition in the central nervous system predominantly occurs through the activation of GABAA receptors. These are anion-selective ligand-gated ion channels permeable to chloride ions, and activation typically results in the inhibition of neuronal activity through phasic and tonic mechanisms. Some GABAA receptor subtypes are capable of opening in the absence of GABA to contribute towards tonic inhibition, but the mechanism by which this occurs and the effect on neuronal activity is not well understood. A variety of recombinant GABAA receptor isoforms were examined for spontaneous activity by expression in HEK cells. Only those receptors containing the β3 subunit showed demonstrable levels of spontaneous activity, and key residues underpinning this were identified within the β3 extracellular domain. To investigate how spontaneous activity may be regulated in vivo, we examined phosphorylation of the β3 subunit using mutagenesis and by manipulating kinase activity, demonstrating a role for phosphorylation in regulating spontaneous current. Furthermore, the effect of various allosteric modulators, including neurosteroids, general anaesthetics and benzodiazepines, may exert a substantive part of their modulatory action through potentiation of spontaneous GABAA receptor activity. We demonstrate that the β3 subunit is also important for spontaneous activity of receptors expressed in hippocampal neurons in culture by both expressing subunits via transfection and by knock-down of native subunits using β3-selective shRNAs. Examination of hippocampal and thalamic areas in adolescent rat brain slices identified varying levels of spontaneous current contributing to tonic inhibition, consistent with variable expression of the β3 subunit. Finally, mutations of the β3 subunit linked with epilepsy were studied and found to impact on spontaneous currents. Overall, these data demonstrate the profound importance of the β3 subunit in controlling neuronal excitability, including the mechanisms by which neurons may alter levels of tonic inhibition through regulation of spontaneous GABAA receptor currents.

Type: Thesis (Doctoral)
Qualification: Ph.D
Title: Structural determinants and regulation of spontaneous activity in GABAA receptors
Event: UCL (University College London)
Open access status: An open access version is available from UCL Discovery
Language: English
Additional information: Copyright © The Author 2020. Original content in this thesis is licensed under the terms of the Creative Commons Attribution 4.0 International (CC BY 4.0) Licence (https://creativecommons.org/licenses/by/4.0/). Any third-party copyright material present remains the property of its respective owner(s) and is licensed under its existing terms. Access may initially be restricted at the author’s request.
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Life Sciences > Div of Biosciences > Neuro, Physiology and Pharmacology
URI: https://discovery.ucl.ac.uk/id/eprint/10115488
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