Rapid and reversible optogenetic silencing of synaptic transmission by clustering of synaptic vesicles

Abstract

Acutely silencing specific neurons informs about their functional roles in circuits and behavior. Existing optogenetic silencers include ion pumps, channels, metabotropic receptors, and tools that damage the neurotransmitter release machinery. While the former hyperpolarize the cell, alter ionic gradients or cellular biochemistry, the latter allow only slow recovery, requiring de novo synthesis. Thus, tools combining fast activation and reversibility are needed. Here, we use light-evoked homo-oligomerization of cryptochrome CRY2 to silence synaptic transmission, by clustering synaptic vesicles (SVs). We benchmark this tool, optoSynC, in Caenorhabditis elegans, zebrafish, and murine hippocampal neurons. optoSynC clusters SVs, observable by electron microscopy. Locomotion silencing occurs with tauon ~7.2 s and recovers with tauoff ~6.5 min after light-off. optoSynC can inhibit exocytosis for several hours, at very low light intensities, does not affect ion currents, biochemistry or synaptic proteins, and may further allow manipulating different SV pools and the transfer of SVs between them.

A bistable inhibitory optoGPCR for multiplexed optogenetic control of neural circuits

“…Taken together, our results demonstrate that PdCO is a rapid, reversible and versatile optoGPCR that mediates efficient silencing of glutamatergic and neuromodulatory synaptic transmission in diverse cell types in vitro and in vivo that expands and complements the collection of presynaptic optogenetic tools 16,59 . For manipulating the presynapse, PdCO could potentially serve as a suitable template to create optoGPCR chimeras with altered signaling specificity by exchanging the intracellular GPCR interface as previously demonstrated for other rhodopsin GPCRs [60][61][62][63][64][65][66][67][68][69][70] .…”

A bistable inhibitory OptoGPCR for multiplexed optogenetic control of neural circuits

“…Taken together, we demonstrated that PdCO is a rapid, reversible, and versatile optoGPCR that mediates inhibition of synaptic transmission efficiently in diverse cell types in vitro and in vivo. With activation time constants in the sub 100-ms and inactivation switching times <10 s, PdCO serves as a fast inhibitory optoGPCR for precise presynaptic inhibition that expands and complements the collection of established and recently developed presynaptic optogenetic tools 16,57 . For manipulating the presynapse, PdCO could potentially serve as a suitable template to create optoGPCR chimeras with altered signaling specificity by exchanging the intracellular GPCR interface as previously demonstrated for other rhodopsin GPCRs [58][59][60][61][62][63][64][65][66][67][68] .…”

Closed-loop experiments and brain machine interfaces with multiphoton microscopy

“…Light-sensitive Gi proteins, such as eOPN3 127 and PPO, 128 can suppress synaptic transmission when expressed presynaptically and are potentially usable with two-photon excitation, although they may cause effects via activation of Kir channels at presynapses. In this respect, opto-SynC, 129 which uses light-evoked homo-oligomerization of cryptochrome CRY2 to cluster synaptic vesicles and inhibit exocytosis, would be completely independent of membrane potential. If the opposite is also possible [i.e., photoactivation of Gs and Gq proteins (opto-XR 130 ), photoactivation of cAMP (bPAC), 131 or light-evoked clustering of synaptic proteins] and exocytosis can be facilitated with two-photon targeted photostimulation, it would be possible to stimulate targeted synapses.…”