G protein-coupled receptor (GPCR) signaling pathways are known contributors to a variety of human diseases. Within the central nervous system, diseases such as depression, anxiety, schizophrenia, and Parkinson's disease are but a few of the many pathologies in which GPCR signaling plays a role. Importantly, GPCRs represent the largest class of drug targets for these and other disorders, and developing new therapeutic approaches requires an understanding of the role these receptors play in signal transduction within specific neuronal populations. Traditional pharmacological approaches to investigating these roles are limited by off-target drug actions and constitutive receptor activity. To overcome this barrier, the Roth lab has engineered a family of GPCRs, derived from the human muscarinic receptors, which have lost affinity for their native ligand, acetylcholine, lack any detectable intrinsic activity, and are selectively activated by the bioavailable inert synthetic ligand, clozapine-N-oxide (CNO). We refer to these receptors as Designer Receptors Exclusively Activated by Designer Drug, or DREADDs. This research proposal investigates the hypothesis that the use of DREADDs will allow for unprecedented spatiotemporal control of neuronal GPCR signaling and provide a novel system to probe the contributions of GPCR signaling to neuronal pathologies.
The specific aims are (1) a proof-of-concept study to characterize the effects of expression and activation of a Gq-coupled DREADD when selectively expressed in the forebrain of transgenic mice;and (2) to determine the in vivo biochemical and signaling effects of selectively activating and silencing serotonergic neurons via DREADDs. To achieve these aims, a variety of techniques will be used, including immunohistochemistry, in vivo microdialysis, measurement of inositol phosphate accumulation, immunoblotting, and stereotaxic viral infusion. Follow-up studies will incorporate mouse behavioral testing using models of psychiatric disease. This novel system has the potential to revolutionize the approach to understanding and. manipulating neuronal GPCR signaling in vivo. Additionally, the particular signaling pathways this proposal will investigate could reveal critical information regarding the etiology and treatment of mental illness.
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|Krashes, Michael J; Koda, Shuichi; Ye, ChianPing et al. (2011) Rapid, reversible activation of AgRP neurons drives feeding behavior in mice. J Clin Invest 121:1424-8|
|Dong, Shuyun; Rogan, Sarah C; Roth, Bryan L (2010) Directed molecular evolution of DREADDs: a generic approach to creating next-generation RASSLs. Nat Protoc 5:561-73|