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Title: Photoactive antagonists for the GABAA receptor
Author: Huckvale, R.
ISNI:       0000 0004 7659 9291
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2016
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GABAA receptors are an integral part of regulating neuronal transmission. Through the action of their endogenous agonist, γ-aminobutyric acid, they control most neuronal signal inhibition. GABAA receptors constantly move through the membrane, readily adapting to different stimuli, aided by transient interactions with scaffolding proteins. This thesis attempts to design and synthesise small, novel chemical tracking devices in order to gain insight into the driving forces of GABAA receptor diffusion. To this aim, photoaffinity labelled probe 1, based on a GABAA receptor antagonist gabazine, was synthesised. A benzophenone photoaffinity label was prepared, and installed onto a pyridazinyl-phenol core. A Sonogashira reaction, to install a carboxylic acid handle for the biotinylated linker, initially proved unsuccessful until the replacement of aryl bromide with an aryl iodide (Figure 1). Biotinylated probe 1 was associated with a quantum dot coated in streptavidin, and placed in live neurons. Real-time recording of the quantum dot trajectories show receptors moving in synaptic and extrasynaptic sites. Figure 1: Photoaffinity probe 1 for tracking GABAA receptors in cells. This thesis also describes a modular synthetic strategy to build additional chemical tools. A "clickable" alkyne gabazine fragment 2 was synthesised and underwent copper catalysed alkyne-azide couplings to give probes 3 and 4. These covalently bind to the receptor through an acrylamide electrophilic moiety, and cleavage of the directing gabazine fragment after UV light leaves a silently-tagged receptor. Further generations of these probes could be used to track unblocked, native receptors in order to compare the effects of antagonism on receptor movement (Figure 2). Figure 2: Photocleavable affinity antagonists 3 and 4 In addition, this thesis discusses a novel photoswitchable antagonist, azo-gabazine 5, which uses the cis-trans photoisomerisation of azobenzene to tune its potency under different wavelengths of light. It is synthesised and used to confer a photoswitching ability onto a native neuronal GABAA receptors in a biological setting (Figure 3). Figure 3: Development of a photoswitchable antagonist to the GABAA receptor, azo-gabazine 5.
Supervisor: Baker, J. R. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available