Unraveling the structure and function of the brain requires a detailed knowledge about the neuronal connections, i.e., the spatial architecture of the nerve fibers. One of the most powerful histological methods to reconstruct the three-dimensional nerve fiber pathways is 3D-polarized light imaging (3D-PLI). The technique measures the birefringence of histological brain sections and derives the spatial fiber orientations of whole human brain sections with micrometer resolution. However, the technique yields only a single fiber orientation for each measured tissue voxel even if it is composed of fibers with different orientations, so that in-plane crossing fibers are misinterpreted as out-of-plane fibers. When generating a detailed model of the three-dimensional nerve fiber architecture in the brain, a correct detection and interpretation of nerve fiber crossings is crucial. Here, we show how light scattering in transmission microscopy measurements can be leveraged to identify nerve fiber crossings in 3D-PLI data and demonstrate that measurements of the scattering pattern can resolve the substructure of brain tissue like the crossing angles of the nerve fibers. For this purpose, we develop a simulation framework that permits the study of transmission microscopy measurements - in particular, light scattering - on large-scale complex fiber structures like brain tissue, using finite-difference time-domain (FDTD) simulations and high-performance computing. The simulations are used not only to model and explain experimental observations, but also to develop new analysis methods and measurement techniques. We demonstrate in various experimental studies on brain sections from different species (rodent, monkey, and human) and in FDTD simulations that the polarization-independent transmitted light intensity (transmittance) decreases significantly (by more than 50%) with an increasing out-of-plane angle of the nerve fibers and that it is mostly independent of the in-plane crossing angle. Hence, the transmittance can be used to distinguish regions with low fiber density and in-plane crossing fibers from regions with out-of-plane fibers, solving a major problem in 3D-PLI and allowing for a much better reconstruction of the complex nerve fiber architecture in the brain. Finally, we show that light scattering (oblique illumination) in the visible spectrum reveals the underlying structure of brain tissue like the crossing angle of the nerve fibers with micrometer resolution, enabling an even more detailed reconstruction of nerve fiber crossings in the brain and opening up new fields of research.

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We explore the polarization-(in)dependent transmitted light intensity of histological brain sections. Using experimental and simulation studies, we demonstrate that it contains valuable information about nerve fiber architecture and tissue structure.

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When transmitting polarised light through histological brain sections, different types of diattenuation (polarisation-dependent attenuation of light) can be observed: In some brain regions, the light is minimally attenuated when it is polarised parallel to the nerve fibres (referred to as D⁺), in others, it is maximally attenuated (referred to as D⁻). The underlying mechanisms of these effects and their relationship to tissue properties were so far unknown. Here, we demonstrate in experimental studies that diattenuation of both types D⁺ and D⁻ can be observed in brain tissue samples from different species (rodent, monkey, and human) and that the strength and type of diattenuation depend on the nerve fibre orientations. By combining finite-difference time-domain simulations and analytical modelling, we explain the observed diattenuation effects and show that they are caused both by anisotropic absorption (dichroism) and by anisotropic light scattering. Our studies demonstrate that the diattenuation signal depends not only on the nerve fibre orientations but also on other brain tissue properties like tissue homogeneity, fibre size, and myelin sheath thickness. This allows to use the diattenuation signal to distinguish between brain regions with different tissue properties and establishes Diattenuation Imaging as a valuable imaging technique.

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Three-dimensional Polarized Light Imaging (3D-PLI) is a promising technique to reconstruct the nerve fiber architecture of human post-mortem brains from birefringence measurements of histological brain sections with micrometer resolution. To better understand how the reconstructed fiber orientations are related to the underlying fiber structure, numerical simulations are employed. Here, we present two complementary simulation approaches that reproduce the entire 3D-PLI analysis: First, we give a short review on a simulation approach that uses the Jones matrix calculus to model the birefringent myelin sheaths. Afterwards, we introduce a more sophisticated simulation tool: a 3D Maxwell solver based on a Finite-Difference Time-Domain algorithm that simulates the propagation of the electromagnetic light wave through the brain tissue. We demonstrate that the Maxwell solver is a valuable tool to better understand the interaction of polarized light with brain tissue and to enhance the accuracy of the fiber orientations extracted by 3D-PLI.

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3D Polarized Light Imaging is a neuroimaging technique that provides a high-resolution reconstruction of nerve fiber pathways in human postmortem brains. The spatial fiber orientations are derived from birefringence measurements of histological brain sections which are interpreted by a macroscopic model of uniaxial birefringence. In order to validate the macroscopic model and to investigate possible limitations, numerical simulations have been used. Simulations of a fiber bundle with different inclination angles and optical resolutions have shown that the macroscopic model ensures a reliable estimation of the fiber orientations as long as the polarimeter does not resolve structures smaller than the fiber radius.@en