Quantitative and volumetric analysis of mouse hippocampal region using optical coherence microscopy and tissue clearing technique

Abstract

The hippocampus is a complex structure of the limbic system, which plays a vital role in regulating learning and memory encoding, emotion, and spatial navigation. The observation of the architecture of the hippocampus is particularly important because its morphological abnormalities are directly associated with a number of neurological illnesses such as Alzheimer’s disease, epilepsy, and schizophrenia. Therefore, the volumetric quantification of the hippocampal region in mouse is required not only for understanding the brain structure but also for identifying various neurological diseases. Recently, various optical imaging methods have been introduced to create volumetric anatomy data of ex vivo tissues using physical tissue sectioning or optical clearing. Even though these new approaches present the distinguished volumetric anatomy on various scales, they are still not suitable for use in statistical studies with multiple brains.Here, we introduce a novel label-free and quantitative imaging modality based on serial optical coherence microscopy (OCM) and tissue clearing technique. We developed the home-built spectral-domain OCM system using 840 nm light source, and its axial and lateral resolution has shown as 1.3 μm and 5.2 μm, respectively. For imaging the entire hippocampal region, we conducted the serial block-face imaging technique, which repeatedly cut out and images the brain tissue, which is cleared by OptiMuS clearing solution for deeper imaging. OptiMuS is an aqueous-based single-step solution with fast clearing and high retention ability in volume. After acquiring a high-resolution brain image, we manually segmented cornu ammonis (CA) and dentate gyrus (DG) regions based on the Allen brain atlas map using AMIRA software. In our preliminary experiment, we were successfully able to obtain 3D hippocampal images within ten sliced tissues without any labeling or contrast agents. Our results also showed that the new imaging platform could accurately measure regional volumes such as CA and DG while providing statistical information in multiple hippocampal tissues. Our technique would be widely utilized in various neuronal studies, including the effects of environmental enrichment.