Next Generation Opto-GPCRs for Neuromodulatory Control

Project Summary/Abstract: The field of optogenetics — utilizing light to engage biological systems — is
widely used for the dissection of neural circuits, cellular signaling and manipulating neurophysiological systems
in awake, behaving animals. However, while many new opsins have been developed and are actively used,
challenges still remain, and the current technology lacks a full toolbox for sub-cellular, spatiotemporal control of
signaling — the predominant means for neuromodulator communication in the brain. Here we propose, an
innovative effort combining neuroscience with structural biology and high-throughput pharmacology for the
development of a series of cutting-edge novel Opto-GPCRs that will allow spatiotemporally precise and
pathway-selective control of neuromodulator signaling in vitro and in freely moving animals. In four aims across
five leading laboratories, we will develop and test these novel tools in vitro and in vivo. Specifically, we will
work to 1) Develop and fully optimize OptoGPCR-v3.0 (Gi coupled) receptors for enhanced spectral
multiplexing and altered sensitivity using structure-function analysis together with mutant-library HTS landing
pad system; 2) Develop and fully optimize OptoGPCR-v3.0-Gq receptors for selective coupling to Gq signaling
pathways using structure-guidance and the HTS landing pad system; 3) Utilize databases of less-explored
naturally-occurring opsin-GPCRs to test, screen and further develop new optical tools with unique profiles; and
4) Assess the spectral compatibility for simultaneous use of OptoGPCR-v3.0 constructs in vivo, together with
biosensors using photometry, 2p imaging and concurrent behavioral measures in vivo. Successful completion
of the proposal will provide the wide neuroscience community with the long awaited capabilities of
spatiotemporal manipulation of GPCR – neuromodulator signaling within neural circuits in vitro and in vivo, in
awake freely behaving animals, and could be used for a wide variety of applications. This new technology will
also further widen the field for unique optical approaches that allow discrete control and optodynamic
simulation of neuromodulator function in brain tissue. We therefore believe that by the fulfilment of these goals
we directly address the central purpose of this RFA-NS-21-027 call.

Grant Number: 5U01NS128537-04
NIH Institute/Center: NIH
Principal Investigator: Michael Bruchas