Reverse optogenetics of G protein signaling by zebrafish non-visual opsin Opn7b for synchronization of neuronal networks

Abstract

Opn7b is a non-visual G protein-coupled receptor expressed in zebrafish. Here we find that Opn7b expressed in HEK cells constitutively activates the Gi/o pathway and illumination with blue/green light inactivates G protein-coupled inwardly rectifying potassium channels. This suggests that light acts as an inverse agonist for Opn7b and can be used as an optogenetic tool to inhibit neuronal networks in the dark and interrupt constitutive inhibition in the light. Consistent with this prediction, illumination of recombinant expressed Opn7b in cortical pyramidal cells results in increased neuronal activity. In awake mice, light stimulation of Opn7b expressed in pyramidal cells of somatosensory cortex reliably induces generalized epileptiform activity within a short (<10 s) delay after onset of stimulation. Our study demonstrates a reversed mechanism for G protein-coupled receptor control and Opn7b as a tool for controlling neural circuit properties.

All Light, Everywhere? Photoreceptors at Nonconventional Sites

“…A striking example is presented in a study of zebrafish that reveals that these fish express as many as 42 opsins all over their body ( 28 ). For at least 27 of these opsins, action and/or absorbance spectra exist, coupled with additional functional and/or signaling information ( 28 , 31 – 40 ). The fish opsins, for which proven light sensitivity exists, have been shown to be expressed in the eye, brain, fin, heart, skin, gills, muscle, pineal, pituitary, and testis ( 28 ).…”

Optical Approaches for Investigating Neuromodulation and G Protein–Coupled Receptor Signaling

“…Importantly, eOPN3 shows maximal activation from green light illumination, allowing this tool to be potentially multiplexed with other opsins or biosensors with spectral properties tuned to far-shifted blue or red light. Additional nonvertebrate opsins have since been characterized for efficient manipulations of neural activity, including the zebrafish Opn7b ( Karapinar et al, 2021 ).…”

Rhodopsins: An Excitingly Versatile Protein Species for Research, Development and Creative Engineering

“…Another remarkable subset are Opn5L, peropsin and Opn7 members, which also bind all- trans retinal in the dark state, but that seems to be the active state binding the G protein. Upon illumination they generate the 11- cis chromophore, which represents the resting state that in the case of Opn5L members may even thermally revert to the active state ( Nagata et al, 2018 ; Yamashita, 2020 ; Karapinar et al, 2021 ; Sakai et al, 2022 ). An even more surprising observation is that some type-2 pigments may be involved in recognizing temperature differences or mechanical changes, or function as chemosensors or tumorigenic elements, possibly even without requiring their retinal cofactor ( Shen et al, 2011 ; Park et al, 2013 ; Baker et al, 2015 ; Pérez-Cerezales et al, 2015 ; Leung et al, 2020 ; Xu et al, 2020 ; Córdova et al, 2021 ; Moraes et al, 2021 ).…”

“…( Lin et al, 2013 ; Kushibiki et al, 2014 ; Kato et al, 2015b ; Brinks et al, 2016 ; Govorunova et al, 2017 ; Cho et al, 2019 ; Krol et al, 2019 ; Kojima et al, 2020b ; Gong et al, 2020 ). Eventually, enzyme-rhodopsins as well as bistable type-2 pigments also entered the field, being able to modulate cellular metabolic processes up to gene expression ( Mukherjee et al, 2019 ; Karapinar et al, 2021 ; Mahn et al, 2021 ; Rodgers et al, 2021 ; Tsunoda et al, 2021 ; Vierock et al, 2021 ). Bistable type-1 and -2 pigments allow further control, since their activity is triggered by illumination, but ends near the M(eta) stage, which can be photoreversed by illumination in another spectral range ( Sheves and Friedman, 1986 ; Koyanagi and Terakita, 2014 ; Mederos et al, 2019 ; Eickelbeck et al, 2020 ).…”