3D visualization of neurovascular networks in pulp-exposed rat molars using tissue clearing techniques

To explore the spatial distribution of the neurovascular network following pulp exposure using tissue clearing technology and three-dimensional imaging. An animal model with dental pulp exposure was established in maxillary molars of Sprague-Dawley rats using a round bur under water cooling. Histological evaluation was performed at various time points, and samples at 24 h postoperatively was selected for tissue clearing and whole-mount immunofluorescence staining using markers for blood vessels, nerves, lymphatics, and mesenchymal stem cells. Three-dimensional images were captured with a light-sheet microscope. Three-dimensional imaging revealed the spatial arrangement of nerves, blood vessels, and lymphatic vessels, as well as the relative positions of sympathetic and sensory nerves in the maxilla. The neurovascular network structure in pulp-exposed molars showed significant alterations compared to healthy molars. Additionally, nerves passing through lateral canals were visualized in three-dimensional space. Numerous STRO-1-positive cells were identified in the maxilla, although their distribution lacked a defined spatial pattern. This study sheds light on the three-dimensional architecture of vascular, neural, and lymphatic networks in dental pulp and surrounding tissues under healthy and pathological conditions. Tissue clearing proves to be a valuable technique for visualizing the spatial organization of dental pulp tissue in physiological and pathological states.