Vertebrate Cone Opsins Enable Sustained and Highly Sensitive Rapid Control of G i/o Signaling in Anxiety Circuitry

Abstract

G protein-coupled receptors (GPCRs) coupling to Gi/o signaling pathways are involved in the control of important physiological functions, which are difficult to investigate because of the limitation of tools to control the signaling pathway with precise kinetics and specificity. We established two vertebrate cone opsins, short- and long-wavelength opsin, for long-lasting and repetitive activation of Gi/o signaling pathways in vitro and in vivo. We demonstrate for both opsins the repetitive fast, membrane-delimited, ultra light-sensitive, and wavelength-dependent activation of the Gi/o pathway in HEK cells. We also show repetitive control of Gi/o pathway activation in 5-HT1A receptor domains in the dorsal raphe nucleus (DRN) in brain slices and in vivo, which is sufficient to modulate anxiety behavior in mice. Thus, vertebrate cone opsins represent a class of tools for understanding the role of Gi/o-coupled GPCRs in health and disease.

Optogenetic user s guide to Opto-GPCRs

“…Importantly, the light-activation properties of tagged vRh remained unchanged. Similar results were recently presented for tagged cone opsin (38) and tagged melanopsin, containing the CT of the serotonin 2c receptor (5-HT 2c ) that were expressed in the dorsal raphe nucleus (DRN) in mice. Indeed, repetitive activation of the G i pathway in 5-HT neurons with CT-tagged cone opsin regulated anxiety behavior in mice (38).…”

“…Similar results were recently presented for tagged cone opsin (38) and tagged melanopsin, containing the CT of the serotonin 2c receptor (5-HT 2c ) that were expressed in the dorsal raphe nucleus (DRN) in mice. Indeed, repetitive activation of the G i pathway in 5-HT neurons with CT-tagged cone opsin regulated anxiety behavior in mice (38). Activation of the G q pathway in the same cells with tagged melanopsin unraveled a so far unknown autoregulatory negative feedback mechanism that also modulates anxiety behaviors (48).…”

“…Color opsins from the cone photoreceptors of the retina, i.e. short-and long-wavelength mouse opsins, are bleached less rapidly and were therefore used as alternatives to vRh (38). Vertebrate cone opsins successfully modulated anxiety behavior in mice by repetitively activating G i signaling in serotonin 1A (5-HT1A) receptor domains in the dorsal raphe nucleus (DRN) (38).…”

“…short-and long-wavelength mouse opsins, are bleached less rapidly and were therefore used as alternatives to vRh (38). Vertebrate cone opsins successfully modulated anxiety behavior in mice by repetitively activating G i signaling in serotonin 1A (5-HT1A) receptor domains in the dorsal raphe nucleus (DRN) (38). Although not shown so far, different opsins with different wavelength sensitivities (OPN1SW, OPN1MW, OPN1LW) could in principle be used to independently activate G i pathways in different target cell types.…”

Subcellular optogenetics – controlling signaling and single-cell behavior

“…For instance, the ability of color opsins to be activated repeatedly has been recruited to obtain continual optical control over G protein signaling in brain slices and regulate anxiety in mice (Masseck et al, 2014). To obtain subcellular optical triggers that are capable of activating different second messenger pathways, chimeric receptors that contain the light-sensing domains of blue opsin and appropriate cytosolic G-protein-coupling regions of a different receptor can be engineered.…”