Clearing the way for optical mapping of the brain with light-sheet fluorescence microscopy

Light-sheet fluorescence microscopy (LSFM) has emerged as a powerful optical imaging technique for studying complex biological systems, particularly in neuroscience. By selectively illuminating a thin optical plane and combining it with camera-based widefield detection, LSFM enables rapid volumetric imaging with reduced photobleaching and phototoxicity compared to point-scanning approaches, such as confocal and multiphoton microscopy. Originally developed for imaging small, transparent organisms, LSFM has evolved into a versatile platform capable of probing large specimens, including whole mouse brains and human tissue samples, through the integration of advanced optical clearing techniques and sophisticated data-analysis tools. This review is focused on the optical principles underlying LSFM, the recent technological developments in light-sheet generation and system configurations, and their impact on imaging performances in highly-scattering tissues. It also discusses practical considerations related to the multiple optical clearing methodologies developed in recent decades and highlights representative applications in brain structural mapping. Finally, it outlines current challenges in data handling and imaging artifacts, and briefly considers future directions for integrating LSFM with complementary imaging modalities.