Mapping Mesoscale Cardiac Architecture with Axially Swept Dithered Light-Sheet Microscopy

Comprehensive characterization of cardiac microstructure and vascular networks is critical for assessing cardiovascular development, disease progression, and therapeutic interventions. While volumetric imaging combined with tissue clearing enables high-resolution organ-level analysis, the densely organized cardiac microenvironment, characterized by compacted and trabeculated myocardium, remains technically challenging to resolve in its entirety. To address these limitations, we exploit the temporal dynamics of illumination-detection interplay to create a time-averaged beam profile, thereby maximizing system throughput across multi-scale specimens. Our dithered light-sheet (DiLS) approach extends the confocal region by over 40% and improves the space-bandwidth product while largely preserving optical sectioning capability. Building on this, the axially swept dithered light-sheet (AS-DiLS) employs a dual-actuator strategy that combines rapid beam dithering and axial sweeping to further accelerate imaging speed while achieving micrometer-scale resolution across centimeter-scale scanning ranges, enabling detailed cardiovascular morphological and topological assessment from embryonic to adult stages. We anticipate that this scalable imaging scheme will advance in-depth investigations of organ morphogenesis and disease pathogenesis across a broad spectrum of models.