An optical clearing imaging window: Realization of mouse brain imaging and manipulation through scalp and skull

Cortical visualization is essential to understand the dynamic changes in brain microenvironment under physiopathological conditions. However, the turbid scalp and skull severely limit the imaging depth and resolution. Existing cranial windows require invasive scalp excision and various subsequent skull treatments. Non-invasive in vivo imaging of skull bone marrow, meninges, and cortex through scalp and skull with high resolution yet remains a challenge. In this work, a non-invasive trans-scalp/skull optical clearing imaging window is proposed for cortical and calvarial imaging, which is achieved by applying a novel skin optical clearing reagent. The imaging depth and resolution are greatly enhanced in near infrared imaging and optical coherence tomography imaging. Combining this imaging window with adaptive optics, we achieve the visualization and manipulation of the calvarial and cortical microenvironment through the scalp and skull using two-photon imaging for the first time. Our method provides a well-performed imaging window and paves the way for intravital brain studies with the advantages of easy-operation, convenience and non-invasiveness.