The powerful optogenetic pharmacology method allows the optical control of neuronal activity by photoswitchable ligands tethered to channels and receptors. However, this approach is technically demanding, as it requires the design of pharmacologically active ligands. The development of versatile technologies therefore represents a challenging issue. Here, we present optogating, a method in which the gating machinery of an ATP-activated P2X channel was reprogrammed to respond to light. We found that channels covalently modified by azobenzene-containing reagents at the transmembrane segments could be reversibly turned on and off by light, without the need of ATP, thus revealing an agonist-independent, light-induced gating mechanism. We demonstrate photocontrol of neuronal activity by a light-gated, ATP-insensitive P2X receptor, providing an original tool devoid of endogenous sensitivity to delineate P2X signaling in normal and pathological states. These findings open new avenues to specifically activate other ion channels independently of their natural stimulus.azobenzene photoswitch | purinergic receptors O ptogenetic approaches in neuroscience rely on the heterologous expression of engineered light-gated ion channels or pumps to trigger or inhibit electrical activity of selected neurons (1, 2). This powerful and revolutionizing technique provides an exquisite remote control of neuronal circuits that drive behavior in animals. Of special interest is the optogenetic pharmacology (also known as optochemical genetic) (3, 4), which allows the control of an ion channel or receptor function by a photoswitchable ligand that is irreversibly tethered to the genetically modified protein through cysteine substitution (3, 4). Ligands are pharmacologically active substances targeting either competitive (5-7), or noncompetitive binding sites (8-10), and light sensitivity is mostly conferred by substituting a photoisomerizable azobenzene derivative, which interconverts reversibly between a long trans-isomer and a short cis-isomer (11). However, this approach is technically demanding, as it requires the design of site-specific ligands for each target. The development of versatile methods with generic photoswitchable molecules would thus improve the optochemical strategy.Here, we present optogating, a unique method for the optical control of ATP-activated P2X channel gating. P2X receptors are a family of trimeric, cation-selective channels (12, 13) comprising seven mammalian subtypes that mediate a variety of physiological responses, including fast synaptic transmission, contraction of smooth muscle, modulation of neurotransmitter release, and pain sensation (12,14,15). P2X receptors are considered as emerging therapeutic targets because of their link to cancer (16), inflammatory (17), and neuronal diseases, including neuropathic pain (18). Inspired by previous works showing that chemical modification of the transmembrane (TM) pore region of the P2X2 receptor affects channel gating through labeling of single cysteine mutants by p...
