For the design of therapeutic drugs, G protein coupled receptors (GPCRs) are notable targets. Many screening methods have been developed to identify effective agents for GPCR signaling. However, analyses of temporal variations of GPCR activity with specific ligands remain insufficient because of monitoring method limitations and difficulties. We previously developed a high-throughput bioluminescence measuring system to detect interactions of GPCR with β-arrestin based on split luciferase fragment complementation. By newly introducing a bioluminescence imaging technique into the system, we demonstrate a method for the temporal monitoring of GPCR-β-arrestin interactions in living cells during stimulation by different ligands.