Bioorthogonal chemical labeling of endogenous receptors in the live mouse brain

Selective chemical labeling of endogenous proteins in living organisms is an important approach for analyzing intact protein function without genetic manipulation. Recently, we achieved the labeling of neurotransmitter receptors in the living brain by employing ligand-directed chemistry that enables covalent modification of endogenous proteins. By designing labeling reagents usable in the living brain and investigating their administration methods, we successfully labeled multiple major neurotransmitter receptors, including AMPA, NMDA, mGlu1, and GABA_A receptors. The covalent labeling properties enabled analysis of fixed brain samples, achieving imaging of endogenous receptors under co-immunolabeling and tissue clearing conditions. Furthermore, by utilizing the pulse-tagging of target receptors, we analyzed the degradation kinetics and translocation dynamics of active receptors. Here, we describe in detail a protocol for the selective covalent labeling of neurotransmitter receptors in the brains of living mice. This protocol provides neurobiologists with a powerful method for visualizing and quantitatively analyzing the function and dynamics of endogenous receptors without genetic manipulation.