A subtype-selective photoswitchable agonist for precise manipulation of GABA_A receptors

Neuronal inhibition is largely mediated by type-A GABA receptors (GABA_ARs), a family of ligand-gated chloride-permeable channels, which can be sub-classified by their subunit composition. Unravelling the function and distribution of each GABA_AR subtype is essential for a holistic understanding of GABAergic inhibition in health and diseases. Photopharmacology, a technique that utilises light-sensitive compounds to precisely manipulate endogenous proteins, is powerful for this purpose. To resolve the molecular complexity of neuronal inhibition, we aimed to develop subtype-selective photoswitchable agonists for GABA_ARs.

Inspired by THIP (gaboxadol), an agonist selective for δ subunit-containing GABA_ARs (δ-GABA_ARs), we merged a photoswitch moiety (azobenzene) with an analogue of THIP (isoguvacine) to construct Az-IGU. Using whole-cell voltage-clamp recording, Az-IGU was tested on 13 GABA_AR subtypes expressed in human embryonic kidney (HEK) cells. Optical activation of endogenous GABA_ARs was examined via electrophysiology in cultured cortical neurons.

In HEK cells, Az-IGU exerted reversible photo-agonism selectively for α4β3δ and α6β3δ GABA_ARs, two major mediators of tonic inhibition. Pharmacological and mutagenesis studies suggested that activation of the α4β3δ GABA_AR involves interaction between Az-IGU and the GABA-binding pocket and is strongly correlated with the spontaneous activity of the receptor. In cultured cortical neurons, photoisomerisation of Az-IGU triggered responses that enabled reversible control of action potential firing.

GABA_ARs are potential therapeutic targets for many disorders. However, their physiological and pathophysiological roles remain largely unexplored. Az-IGU may enable photopharmacological studies of α4/6β3δ GABA_ARs, providing new opportunities for biomedical and neurobiological applications.