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Title: Exploring the activity of an inhibitory neurosteroid at GABAA receptors
Author: Seljeset, S.
ISNI:       0000 0004 7659 5944
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2017
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The GABAA receptor is the main mediator of inhibitory neurotransmission in the central nervous system. Its activity is regulated by various endogenous molecules that act either by directly modulating the receptor or by affecting the presynaptic release of GABA. Neurosteroids are an important class of endogenous modulators, and can either potentiate or inhibit GABAA receptor function. Whereas the binding site and physiological roles of the potentiating neurosteroids are well characterised, less is known about the role of inhibitory neurosteroids in modulating GABAA receptors. Using hippocampal cultures and recombinant GABAA receptors expressed in HEK cells, the binding and functional profile of the inhibitory neurosteroid pregnenolone sulphate (PS) were studied using whole-cell patch-clamp recordings. In HEK cells, PS inhibited steady-state GABA currents more than peak currents. Receptor subtype selectivity was minimal, except that the ρ1 receptor was largely insensitive. PS showed state-dependence but little voltage-sensitivity and did not compete with the open-channel blocker picrotoxinin for binding, suggesting that the channel pore is an unlikely binding site. By using ρ1-α1/β2/γ2L receptor chimeras and point mutations, the binding site for PS was probed. All chimeras and mutants remained sensitive to PS, raising the question as to whether modulation could be due to indirect interactions between PS and the cell membrane. In hippocampal neurones, the major postsynaptic effect of PS was to increase the IPSC decay rate. However, PS also increased GABA release by activating presynaptic TRPM3 receptors. Upon block of TRPM3, GABA release was reduced by PS due to potentiation of presynaptic Kir2 channel activity. In conclusion, PS directly modulates GABAA receptor kinetics by speeding up current decay at both neuronal and recombinant receptors. At inhibitory synapses, PS can enhance or inhibit GABA release by acting at TRPM3 or Kir2 channels respectively.
Supervisor: Smart, T. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available