Single-chain fluorescent integrators for mapping G-protein-coupled receptor agonists
Kayla Kroning1,2, Noam Gannot1,3, Xingyu Li1,3,4
1Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109.
Summary
Scientists developed novel protein sensors to track G protein-coupled receptor (GPCR) agonists in the brain. This technology, SPOTall, enables precise localization of neuromodulators like dopamine and opioids, advancing neuroscience research.
Area of Science:
- Neuroscience
- Molecular Biology
- Biochemistry
Background:
- G protein-coupled receptors (GPCRs) are crucial for mediating the effects of diverse neuromodulators, including dopamine, serotonin, acetylcholine, and opioids.
- Understanding the precise localization of GPCR agonists is vital for deciphering their impact on specific neuronal pathways.
- Existing methods may lack the specificity or modularity to comprehensively map agonist activity across the brain.
Purpose of the Study:
- To engineer a versatile platform for creating single-protein chain integrator sensors capable of detecting GPCR agonist localization.
- To develop red fluorescent variants of existing opioid receptor sensors (SPOTIT) for multiplexed imaging.
- To adapt the SPOTIT platform into a broader system (SPOTall) for sensing agonists of various GPCRs.
Main Methods:
- Engineering of modular, single-protein chain integrator sensors.
- Development of red fluorescent protein variants for enhanced imaging capabilities.
- Adaptation of the SPOTIT platform to create the SPOTall design for broad GPCR applicability.
- Testing sensor functionality in vivo using viral delivery in mouse models.
Main Results:
- Successful engineering of red fluorescent SPOTIT sensors for multiplexed imaging.
- Creation of the SPOTall platform and development of sensors for beta 2-adrenergic receptor (B2AR), dopamine D1, and muscarinic 2 receptors.
- Demonstration of M-SPOTIT and B2AR-SPOTall in detecting exogenously administered morphine, isoproterenol, and epinephrine in the mouse brain.
- Validation of sensor efficacy in detecting specific agonists in vivo.
Conclusions:
- The SPOTIT and SPOTall sensor design platform offers a powerful tool for unbiased detection of synthetic and endogenous neuromodulator agonists.
- This technology facilitates the study of GPCR signaling and agonist localization across the brain.
- The modular nature of the platform holds significant potential for broad application in neuroscience research.
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