An In Vitro Microfluidic Assay for Studying G-Protein Coupled Photoreceptor Signalling as Derived from the Retinal Photoactivated Outer Segment

G-protein-coupled receptors (GPCRs) are the largest family of membrane-signalling cell surface receptors. Upon light activation, the photoreceptive GPCR Rhodopsin, found in rod cells of the vertebrate retina, triggers a signalling cascade. The cascade includes G-protein activation, followed by enzymatic cGMP degradation leading to the closure of cGMP-gated ion channels so that the membrane becomes electrically polarised, which eventually leads to a neuronal response. Here, we demonstrate that extracts from porcine photoreceptor outer segment cells, incorporated into a synthetic, free-standing lipid bilayer within a microfluidic device, reproduce the expected behaviour of the GPCR signalling network. The recorded electric current across the membrane demonstrates closure or opening of gated channels as a function of illumination. GMP analogues acting as inhibitors or activators cause the channels to remain in a closed or open state regardless of light exposure. Our work represents a first step towards in vitro studies and realisations of GPCR signalling in synthetic membranes, bringing GPCR-based functional modules for synthetic biology within reach.