Double-helix optical point spread function enables real-time mesoscopic 3D functional microangiography in the living mouse brain and skull
- Baoyuan Zhang
- Shiyao Guo
- Lin Tang
- Yi Chen
- Lukas Glandorf
- Etienne Jessen
- Xuyang Chang
- Tian Jin
- Michael Reiss
- Shuxin Lyu
- Qiang Fu
- Hadi Amata
- Wolfgang Heidrich
- Chaim Glück
- Dominik Schillinger
- Bruno Weber
- Xosé Luís Deán-Ben
- Weibo Wang
- Xiong Dun
- Daniel Razansky
- Zhenyue Chen
- Quanyu Zhou
2026-04-13
Quantitative, volumetric imaging of cerebrovascular networks and microcirculation is essential for understanding brain function. However, rapid mesoscopic 3D imaging remains challenging because of fundamental trade-offs between spatiotemporal resolution, field of view, and sensitivity to functional parameters. Here we present a mesoscopic fluorescence imaging platform featuring a double-helix phase mask for real-time, depth-resolved measurements through the intact mouse skull. The compact phase-mask design is compatible with both laser-scanning and widefield microscopy. Using multifocal laser scanning, we demonstrate real-time volumetric in vivo imaging while discriminating calvarial from cerebral vasculature across 6.6×6.6×0.8 mm 3 volume. Beyond high-resolution structural imaging, perfusion time-to-peak values are extracted from the laser-scanning configuration while accurate flow velocity/direction information is provided via widefield tracking of fluorescently labeled cells. We demonstrate the platform’s capabilities by analyzing brain-layer-specific perfusion dynamics and vascular topology in glioma-bearing mouse brains, offering unprecedented views for probing cerebrovascular alterations in both physiological and pathological contexts.