A conformational transition based fluorescent probe for mapping lysosomal viscosity fluctuations by fluorescence lifetime imaging

The local viscosity within lysosomes fluctuates dynamically under various physiological and pathological processes (e.g., autophagy and apoptosis), and has long been regarded as a key indicator to reflect lysosomal status and functionality. Therefore, accurate monitoring of lysosomal viscosity fluctuations is essential for a better understanding of lysosomal function and pathology. Herein, we present a distorted-conformation-induced viscosity-sensitive fluorescent molecular rotor (FMR), BODIPY–P, for quantitative monitoring of lysosomal viscosity. BODIPY–P exhibited outstanding performance, including high sensitivity and selectivity to viscosity, dual response mode of fluorescence intensity and fluorescence lifetime, specific lysosome targeting, quick washing-free imaging and excellent intracellular photostability. Utilizing BODIPY–P, we successfully achieved specific mapping and quantifying the fluctuations of lysosomal viscosity during autophagy and apoptosis through dual-mode imaging of fluorescence intensity and fluorescence lifetime. Density functional theory revealed that the intrinsic viscosity-sensitive mechanism of BODIPY–Pwas ascribed to the conformational transition between planar and distortion caused by the rotation of the meso-benzene ring. Our study will spur additional research on FMRs, including the in-depth exploration of their underlying viscosity-responsive mechanism and potential applications in viscosity-related physiology and pathology investigations.