Rationale: Intravital optical imaging is a significant method for investigating cerebrovascular structure and function. However, its imaging contrast and depth are limited by the turbid skull. Tissue optical clearing has a great potential for solving this problem. Our goal was to develop a transparent skull window, without performing a craniotomy, for use in assessing cerebrovascular structure and function.Methods: Skull optical clearing agents were topically applied to the skulls of mice to create a transparent window within 15 min. The clearing efficacy, repeatability, and safety of the skull window were then investigated.Results: Imaging through the optical clearing skull window enhanced both the contrast and the depth of intravital imaging. The skull window could be used on 2-8-month-old mice and could be expanded from regional to bi-hemispheric. In addition, the window could be repeatedly established without inducing observable inflammation and metabolic toxicity.Conclusion: We successfully developed an easy-to-handle, large, switchable, and safe optical clearing skull window. Combined with various optical imaging techniques, cerebrovascular structure and function can be observed through this optical clearing skull window. Thus, it has the potential for use in basic research on the physiopathologic processes of cortical vessels.

The photodynamic (PD) effect has been reported to be efficient for the opening of the blood-brain barrier (BBB), which provides a new informative platform for developing perspective strategies towards brain disease therapy and drug delivery. However, this method is usually performed via craniotomy due to high scattering of the turbid skull. In this work, we employed a newly-developed optical clearing skull window for investigating non-invasive PD-induced BBB opening to high weight molecules and 100-nm fluid-phase liposomes containing ganglioside GM1. The results demonstrated that the BBB permeability to the Evans blue albumin complex is related to laser doses. By in vivo two-photon imaging and ex vivo confocal imaging with specific markers of the BBB, we noticed PD-related extravasation of rhodamine-dextran and liposomes from the vessels into the brain parenchyma. The PD induced an increase in oxidative stress associated with mild hypoxia and changes in the expression of tight junction (CLND-5 and ZO-1) and adherens junction (VE-cadherin) proteins, which might be one of the mechanisms underlying the PD-related BBB opening for liposomes. Our experiments indicate that optical clearing skull window will be a promising tool for non-invasive PD-related BBB opening for high weight molecules and liposomes that provides a novel useful tool for brain drug delivery and treatment of brain diseases.

Skin optical clearing can significantly enhance the ability of biomedical optical imaging. Some alcohols and sugars have been selected to be optical clearing agents (OCAs). In this work, we paid attention to the optical clearing potential of disaccharides. Sucrose and maltose were chosen as typical disaccharides to compare with fructose, an excellent monosaccharide-OCA, by using molecular dynamics simulation and an ex vivo experiment. The experimental results indicated that the optical clearing efficacy of skin increases linearly with the concentration for each OCA. Both the theoretical predication and experimental results revealed that the two disaccharides exerted a better optical clearing potential than fructose at the same concentration, and sucrose is optimal. Since maltose has an extremely low saturation concentration, the other two OCAs with saturation concentrations were treated topically on rat skin in vivo, and optical coherence tomography imaging was applied to monitor the optical clearing process. The results demonstrated that sucrose could cause a more significant increase in imaging depth and signal intensity than fructose.