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Updated: Jun 24, 2026
A Polished and Reinforced Thinned-skull Window for Long-term Imaging of the Mouse Brain
Published on: March 7, 2012
A Head-Mounted Optically Transparent Skull (HOTS) Window For Deep Transcranial Imaging of the Mouse Cortex
Lina Liu1, Yufeng Gao1, Yuezhi He2
1Research Center for Biomedical Optics and Molecular Imaging, Shenzhen Key Laboratory for Molecular Imaging, Guangdong Provincial Key Laboratory of Biomedical Optical Imaging Technology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences; University of Chinese Academy of Sciences.
We developed a head-mounted optically transparent skull (HOTS) window for improved in vivo brain imaging in mice. This method enhances skull transparency, enabling deeper two-photon imaging through the intact skull.
Area of Science:
- Neuroscience
- Biomedical Engineering
- Optical Imaging
Background:
- High-resolution in vivo two-photon imaging of the mouse cerebral cortex is hindered by light scattering from the skull.
- Existing skull-clearing methods have limitations in clearing time and cocktail efficacy.
Purpose of the Study:
- To present a detailed protocol for a head-mounted optically transparent skull (HOTS) window.
- To enable extended skull clearing in awake, freely behaving mice for improved optical access.
Main Methods:
- A head-mounted cap was used to maintain clearing solutions over the skull for several hours.
- A two-step clearing procedure employed optimized reagents (HOTS-S1 and HOTS-S2) developed through chemical screening.
- A step-by-step protocol covers skull preparation, cap mounting, reagent delivery, and imaging preparation.
Main Results:
- The HOTS protocol achieved a highly transparent skull in mice, supporting two-photon imaging up to ~800 µm below the pia.
- This performance approaches that of open-skull windows.
- Enabled structural and functional calcium imaging in transgenic mouse models.
Conclusions:
- The HOTS window is a convenient and minimally invasive approach for deep transcranial imaging.
- It facilitates various in vivo light-based manipulations, including optogenetics and photopharmacology.
- This technique significantly advances deep brain imaging capabilities in freely behaving animals.
