Rino Saiga

Three-dimensional network of Drosophila brain hemisphere.

The first step to understanding brain function is to determine the brains network structure. We report a three-dimensional analysis of the brain network of the fruit fly Drosophila melanogaster by synchrotron-radiation tomographic microscopy. A skeletonized wire model of the left half of the brain network was built by tracing the three-dimensional distribution of X-ray absorption coefficients. The obtained models of neuronal processes were classified into groups on the basis of their three-dimensional structures. These classified groups correspond to neuronal tracts that send long-range projections or repeated structures of the optic lobe. The skeletonized model is also composed of neuronal processes that could not be classified into the groups. The distribution of these unclassified structures correlates with the distribution of contacts between neuronal processes. This suggests that neurons that cannot be classified into typical structures should play important roles in brain functions. The quantitative description of the brain network provides a basis for structural and statistical analyses of the Drosophila brain. The challenge is to establish a methodology for reconstructing the brain network in a higher-resolution image, leading to a comprehensive understanding of the brain structure.

A method for estimating spatial resolution of real image in the Fourier domain

Spatial resolution is a fundamental parameter in structural sciences. In crystallography, the resolution is determined from the detection limit of high‐angle diffraction in reciprocal space. In electron microscopy, correlation in the Fourier domain is used for estimating the resolution. In this paper, we report a method for estimating the spatial resolution of real images from a logarithmic intensity plot in the Fourier domain. The logarithmic intensity plots of test images indicated that the full width at half maximum of a Gaussian point spread function can be estimated from the images. The spatial resolution of imaging X‐ray microtomography using Fresnel zone‐plate optics was also estimated with this method. A cross section of a test object visualized with the imaging microtomography indicated that square‐wave patterns up to 120‐nm pitch were resolved. The logarithmic intensity plot was calculated from a tomographic cross section of brain tissue. The full width at half maximum of the point spread function estimated from the plot coincided with the resolution determined from the test object. These results indicated that the logarithmic intensity plot in the Fourier domain provides an alternative measure of the spatial resolution without explicitly defining a noise criterion.

Three-dimensional X-ray visualization of axonal tracts in mouse brain hemisphere.

Neurons transmit active potentials through axons, which are essential for the brain to function. In this study, the axonal networks of the murine brain were visualized with X-ray tomographic microscopy, also known as X-ray microtomography or micro-CT. Murine brain samples were freeze-dried to reconstitute the intrinsic contrast of tissue constituents and subjected to X-ray visualization. A whole brain hemisphere visualized by absorption contrast illustrated three-dimensional structures including those of the striatum, corpus callosum, and anterior commissure. Axonal tracts observed in the striatum start from the basal surface of the cerebral cortex and end at various positions in the basal ganglia. The distribution of X-ray attenuation coefficients indicated that differences in water and phospholipid content between the myelin sheath and surrounding tissue constituents account for the observed contrast. A rod-shaped cutout of brain tissue was also analyzed with a phase retrieval method, wherein tissue microstructures could be resolved with up to 2.7 μm resolution. Structures of axonal networks of the striatum were reconstructed by tracing axonal tracts. Such an analysis should be able to delineate the functional relationships of the brain regions involved in the observed network.

Method for estimating modulation transfer function from sample images

The modulation transfer function (MTF) represents the frequency domain response of imaging modalities. Here, we report a method for estimating the MTF from sample images. Test images were generated from a number of images, including those taken with an electron microscope and with an observation satellite. These original images were convolved with point spread functions (PSFs) including those of circular apertures. The resultant test images were subjected to a Fourier transformation. The logarithm of the squared norm of the Fourier transform was plotted against the squared distance from the origin. Linear correlations were observed in the logarithmic plots, indicating that the PSF of the test images can be approximated with a Gaussian. The MTF was then calculated from the Gaussian-approximated PSF. The obtained MTF closely coincided with the MTF predicted from the original PSF. The MTF of an x-ray microtomographic section of a fly brain was also estimated with this method. The obtained MTF showed good agreement with the MTF determined from an edge profile of an aluminum test object. We suggest that this approach is an alternative way of estimating the MTF, independently of the image type.

Three-dimensional structure of brain tissue at submicrometer resolution

Biological objects are composed of submicrometer structures such as cells and organelles that are essential for their functions. Here, we report on three-dimensional X-ray visualization of cells and organelles at resolutions up to 100 nm by imaging microtomography (micro-CT) equipped with Fresnel zone plate optics. Human cerebral tissue, fruit fly cephalic ganglia, and Escherichia coli bacteria labeled with high atomic-number elements were embedded in epoxy resin and subjected to X-ray microtomography at the BL37XU and BL47XU beamlines of the SPring-8 synchrotron radiation facility. The obtained results indicated that soft tissue structures can be visualized with the imaging microtomography.

Spatiotemporal development of soaked protein crystal

Crystal soaking is widely performed in biological crystallography. This paper reports time-resolved X-ray crystallographic and microtomographic analyses of tetragonal crystals of chicken egg-white lysozyme soaked in mother liquor containing potassium hexachloroplatinate. The microtomographic analysis showed that X-ray attenuation spread from the superficial layer of the crystal and then to the crystal core. The crystallographic analyses indicated that platinum sites can be classified into two groups from the temporal development of the electron densities. A soaking process consisting of binding-rate-driven and equilibrium-driven layers is proposed to describe these results. This study suggests that the composition of chemical and structural species resulting from the soaking process varies depending on the position in the crystal.

Scanning Brain Networks with Micro-CT

Ryuta Mizutani,1* Rino Saiga,1 Susumu Takekoshi,2 Makoto Arai,3 Akihisa Takeuchi,4 and Yoshio Suzuki4 1Department of Applied Biochemistry, Tokai University, Hiratsuka, Kanagawa 259-1292, Japan 2Tokai University School of Medicine, Isehara, Kanagawa 259-1193, Japan 3Tokyo Metropolitan Institute of Medical Science, Setagaya, Tokyo 156-8506, Japan 4Japan Synchrotron Radiation Research Institute (JASRI/SPring-8), Sayo, Hyogo 679-5198, Japan

Synchrotron radiation microtomography of brain hemisphere and spinal cord of a mouse model of multiple sclerosis revealed a correlation between capillary dilation and clinical score

Multiple sclerosis is a neurological disorder in which the myelin sheaths of axons are damaged by the immune response. We report here a three‐dimensional structural analysis of brain and spinal cord tissues of a mouse model of multiple sclerosis, known as experimental autoimmune encephalomyelitis (EAE). EAE‐induced mice were raised with or without administration of fingolimod, which is used in the treatment of multiple sclerosis. Brains and spinal cords dissected from the EAE mice were lyophilized so as to reconstitute the intrinsic contrast of tissue elements, such as axons, in X‐ray images. Three‐dimensional structures of the brain hemispheres and spinal cords of the EAE mice were visualized with synchrotron radiation microtomography. Microtomographic cross sections reconstructed from the X‐ray images revealed dilation of capillary vessels and vacuolation in the spinal cord of the EAE mice. Vacuolation was also observed in the cerebellum, suggesting that the neuroinflammatory response progressed in the brain. The vessel networks and vacuolation lesions in the spinal cords were modelled by automatically tracing the three‐dimensional image in order to analyze the tissue structures quantitatively. The results of the analysis indicated that the distribution of vacuolations was not uniform but three‐dimensionally localized. The mean vessel diameter showed a linear correlation with the clinical score, indicating that vasodilation is relevant to paralysis severity in the disease model. We suggest that vasodilation and vacuolation are related with neurological symptoms of multiple sclerosis.

Estimating the resolution of real images

Image resolvability is the primary concern in imaging. This paper reports an estimation of the full width at half maximum of the point spread function from a Fourier domain plot of real sample images by neither using test objects, nor defining a threshold criterion. We suggest that this method can be applied to any type of image, independently of the imaging modality.

X-ray Tomographic Microscopy of Drosophila Brain Network and Skeletonized Model Building in the Three-Dimensional Image

The brain consists of a large number of neurons that make up a three-dimensional network. The first step to understanding brain functions is to analyze the structure of this network. Although three-dimensional structures of brain tissues have been reported, their structures are difficult to comprehend. This is because of a lack of quantitative descriptions of the three-dimensional network, which should be represented with three-dimensional Cartesian coordinates, rather than a three-dimensional distribution of intensities. Here, we report on x-ray tomographic microscopy of the brain network of the fruit fly Drosophila melanogaster and its analysis by skeletonized-model building [1].