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Title: Top-down and bottom-up control of drug-induced sleep and anaesthesia
Author: Uygun, David
ISNI:       0000 0004 6059 1754
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2016
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In recent decades, research has unravelled fascinating detail about the molecular mechanisms underpinning pharmacologic loss of consciousness (LOC). However, the systems-level mechanisms are far less clear. Recent genetic approaches, however, enable unprecedented dissection on neural pathways, and they are paving a way for this line of research. The focus of this thesis is to investigate the neuroanatomical substrates of commonly used drugs which reversibly render us unconscious. Zolpidem is a positive allosteric modulator (PAM) of the GABAA receptor which binds to the benzodiazepine (BZ) site. Because zolpidem binds α1-3,β,γ2 containing GABAA receptors, which are widespread, it acts virtually everywhere. We do not know if zolpidem causes sleep by enhancing GABAergic inhibition throughout the entire brain, or if the therapeutic sleep-inducing property depends upon specific brain circuitry. γ2I77 mice are devoid of zolpidem-sensitivity. But, zolpidem-sensitivity can be restored selectively in brain regions, enabling dissection of the circuitry involved in zolpidem's effect. To isolate the therapeutic effect of zolpidem we deleted GABAA-γ2I77-subunits and replaced them with GABAA-γ2F77-subunits in HDC neurons or frontal-cortex in isolation. We were able to selectively restore zolpidem-sensitivity in target neurons. This conferred zolpidem-enhanced IPSCs locally. Compared with wild-type mice and zolpidem-insensitive γ2I77lox mice, we found that GABAA-γ2F77 receptors in either HDC-neurons or frontal cortex alone were enough to rescue the majority of zolpidem-mediated sleep. The response in HDC-γ2F77 mice was similar to that of an H1-receptor antagonist. By producing a null effect in a negative-control area - the superior colliculus - we show that HDC neurons and the frontal cortex are both substrates involved in zolpidem-mediated sleep. We also investigated the role of synaptic-inhibition onto corticothalamic-neurons in anaesthetic-induced LOC and sleep-wake. To do this, we genetically ablated γ2-subunits from layer-6 corticothalamic-cells by crossing Ntsr1-Cre mice with GABAA-γ2I77lox mice. We found this reduced isoflurane sensitivity, but left sleep-wake behaviours virtually unaffected.
Supervisor: Wisden, William ; Franks, Nick Sponsor: Biotechnology and Biological Sciences Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral