Functional Characterisation of Spontaneously Active GABAA Receptors

Centre for Clinical Brain Sciences, University of Edinburgh

GABAA receptors are the principle effectors of inhibitory neurotransmission in the central nervous system. The classical view is that activation of these receptors by the endogenous neurotransmitter γ-Aminobutyric acid (GABA) produces two distinct modes of inhibition: phasic and tonic. Phasic inhibition is mediated by post-synaptic GABAA receptors which are activated by high concentrations of GABA released from the pre-synaptic terminal. In contrast, tonic inhibition is generated by extrasynaptic GABAA receptors; the conventional view is that these receptors are persistently activated by low concentrations of ambient GABA. However, recent studies have demonstrated that in certain neuronal sub-types (dentate gyrus granule cell, CA1 pyramidal cell and the central amygdala) the majority of tonic current is produced by GABA-independent, spontaneously opening GABAA receptors (s-GABAARs). s-GABAARs do not require the binding of GABA to enter an active state and, therefore, are resistant to block by competitive antagonists – but can be blocked by open-channel blockers (e.g. picrotoxin and pentylenetetrazol).

Here, we utilised whole cell recordings from dentate gryus granule cells as a means to characterise s-GABAARs. First, we assessed the role of the δ-subunit in s-GABAAR signalling using DS2, a well characterised δ-subunit positive allosteric modulator; and DPP-4-PIOL, a novel antagonist of δ-subunit containing receptors. Whilst GABA-dependent responses were potentiated by DS2 and inhibited by DPP-4-PIOL, these compounds had no effect on the GABA-independent s-GABAARs. This suggests that either s-GABAARs are not δ-subunit containing or that the aforementioned compounds exert their effects solely on GABA-bound receptors. We next evaluated if s-GABAARs are subject to modulation by intracellular signalling pathways. Experiments using Bisindolylmaleimide ii, a protein kinase C inhibitor, showed that s-GABAARs are not regulated by PKC. However, we did evidence a clear role for the Gαi/o signalling pathway: addition of pertussis toxin, a Gαi/o inhibitor, into the patch pipette caused a significant decrease in s-GABAAR mediated tonic inhibition. This points to either a direct Gαi/o-s-GABAAR inhibitory coupling or an inverse relationship between PKA activity and s-GABAAR signalling.

References

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Funded by: Edinburgh Chancellor's Fellowship

* entered into the PhD student poster competition