cb200360z_si_001.avi (3.56 MB)
Visualization and Quantitative Analysis of G Protein-Coupled Receptor−β-Arrestin Interaction in Single Cells and Specific Organs of Living Mice Using Split Luciferase Complementation
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posted on 2012-05-18, 00:00 authored by Hideo Takakura, Mitsuru Hattori, Masaki Takeuchi, Takeaki OzawaMethods used to assess the efficacy of potentially therapeutic
reagents for G protein-coupled receptors (GPCRs) have been developed.
Previously, we demonstrated sensitive detection of the interaction
of GPCRs and β-arrestin2 (ARRB2) using 96-well microtiter plates
and a bioluminescence microscope based on split click beetle luciferase
complementation. Herein, using firefly luciferase emitting longer
wavelength light, we demonstrate quantitative analysis of the interaction
of β2-adrenergic receptor (ADRB2), a kind of GPCR, and ARRB2
in a 96-well plate assay with single-cell imaging. Additionally, we
showed bioluminescence in vivo imaging of the ADRB2–ARRB2
interaction in two systems: cell implantation and hydrodynamic tail
vein (HTV) methods. Specifically, in the HTV method, the luminescence
signal from the liver upon stimulation of an agonist for ADRB2 was
obtained in the intact systems of mice. The results demonstrate that
this method enables noninvasive screening of the efficacy of chemicals
at the specific organ in in vivo testing. This in vivo system can contribute to effective evaluation in
pharmacokinetics and pharmacodynamics and expedite the development
of new drugs for GPCRs.
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vivo systemvivo testingQuantitative AnalysisLiving Micenoninvasive screeningSpecific Organshydrodynamic tail veininteractionluminescence signalHTV methodwavelength lightvivo imagingreceptorcell implantationsplit click beetle luciferase complementationefficacyADRB 2Single Cellsbioluminescence microscopeGPCRplate assayfirefly luciferasemicrotiter platesLuciferase ComplementationMethodsARRB 2
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