Scatterometry Measurements With Scattered Light Imaging Enable New Insights Into the Nerve Fiber Architecture of the Brain

Miriam Menzel,Jan A Reuter,David Gräßel,Markus Axer,Katrin Amunts,Marouan Ritzkowski

doi:10.3389/fnana.2021.767223

Abstract

The correct reconstruction of individual (crossing) nerve fibers is a prerequisite when constructing a detailed network model of the brain. The recently developed technique Scattered Light Imaging (SLI) allows the reconstruction of crossing nerve fiber pathways in whole brain tissue samples with micrometer resolution: the individual fiber orientations are determined by illuminating unstained histological brain sections from different directions, measuring the transmitted scattered light under normal incidence, and studying the light intensity profiles of each pixel in the resulting image series. So far, SLI measurements were performed with a fixed polar angle of illumination and a small number of illumination directions, providing only an estimate of the nerve fiber directions and limited information about the underlying tissue structure. Here, we use a display with individually controllable light-emitting diodes to measure the full distribution of scattered light behind the sample (scattering pattern) for each image pixel at once, enabling scatterometry measurements of whole brain tissue samples. We compare our results to coherent Fourier scatterometry (raster-scanning the sample with a non-focused laser beam) and previous SLI measurements with fixed polar angle of illumination, using sections from a vervet monkey brain and human optic tracts. Finally, we present SLI scatterometry measurements of a human brain section with 3 μm in-plane resolution, demonstrating that the technique is a powerful approach to gain new insights into the nerve fiber architecture of the human brain.

Highlights

Disentangling the highly complex and densely grown nerve fiber network in the brain is key to understanding its function and to developing treatments for neurodegenerative diseases
When illuminating with larger angles, the transmitted light intensity decreases significantly, especially for large illumination times and gain factors. This can be explained by the fact that the distance between the light source and sample increases with increasing illumination angle and the lightemitting diodes (LEDs) have a limited angle of radiation so that outer LEDs do not emit much light under large angles
Coherent Fourier scatterometry (Menzel and Pereira, 2020) measures the complete scattering patterns with high detail but requires mechanical rasterizing of the sample, and the objectspace resolution is limited to the minimum diameter (> 100 μm) of the laser beam

Summary

Introduction

Disentangling the highly complex and densely grown nerve fiber network in the brain is key to understanding its function and to developing treatments for neurodegenerative diseases. Diffusion MRI allows to measure the spatial orientations of crossing nerve fibers, but only with resolutions down to a few hundred micrometers in post-mortem human brains (Calabrese et al, 2018; Roebroeck et al, 2018), which is not sufficient to resolve individual nerve fibers with diameters in the order of 1 μm Scatterometry With Scattered Light Imaging (Liewald et al, 2014). Recent studies (Menzel and Pereira, 2020; Menzel et al, 2020) have shown that nerve fiber crossings can be visualized with scattered light: when shining light in the optical regime through unstained, histological brain sections and studying the spatial distribution of scattered light behind the sample (scattering pattern), we obtain valuable information about the tissue substructure of the illuminated region, such as the individual directions of crossing nerve fibers. The technique demands raster-scanning of the brain section, and the minimum diameter of the laser beam (> 100 μm) limits the spatial resolution

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Results

Conclusion