High-throughput light-sheet 3D microscopy of whole rodent brain microstructure for integration with MRI

Whole-brain cellular mapping is essential for understanding how cellular architecture shapes brain organisation and function. Magnetic Resonance Imaging (MRI) provides whole-brain coverage but is only indirectly sensitive to cellular features, motivating integration with microscopy to improve the interpretation and specificity of microstructural MRI. Here, we present a scalable cross-resolution framework integrating 3D light-sheet microscopy (LSM) of intact rodent brains with diffusion MRI (dMRI) to bridge cellular and macroscopic brain imaging. By combining active tissue clearing and immunolabelling with accelerated LSM acquisition, we achieve multiplexed mapping of neuronal and glial populations across the whole brain on timescales comparable to conventional histology. We apply this framework across eight genetically diverse mouse strains to develop predictive models of microscopy-derived cellular features from multi-shell dMRI contrasts. Data-driven combinations of dMRI contrasts predicted neuronal and astrocytic features better than individual contrasts across animals. This framework enables scalable integration of MRI and 3D LSM, providing a means to identify cellular determinants of MRI contrast and guide the development and interpretation of microstructural MRI measures ultimately translatable to the living human brain.