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  • Minimizing Diffusion Encoding of Slice Selection in Stimulated Echo Imaging

    24 October 2018

    Purpose: Diffusion weighted imaging (DWI) using very high b-values and long diffusion times (Δ) typically uses the stimulated echo (STE) in order that large diffusion encodings can be produced without incurring excessive T2-related signal loss. For long Δ a significant bias can be introduced through the diffusion encoding effect of the slice selection gradients that are typically used in multi-slice implementations of the STE scheme. Compensation of this effect has recently been achieved by adjustment of the prescribed diffusion encoding gradients [1], but the nature of the adjustment is specific to the exact choice of experimental parameters. In this report we show that the effect can simply be minimized through reduction of the diffusion encoding effect of the slice selection gradients themselves, as often implemented in STEAM MRS, thus providing a more straightforward method for maintaining suitable control over the applied diffusion encoding scheme.

  • Recent advancements in diffusion MRI for investigating cortical development after preterm birth-potential and pitfalls.

    24 October 2018

    Preterm infants are born during a critical period of brain maturation, in which even subtle events can result in substantial behavioral, motor and cognitive deficits, as well as psychiatric diseases. Recent evidence shows that the main source for these devastating disabilities is not necessarily white matter (WM) damage but could also be disruptions of cortical microstructure. Animal studies showed how moderate hypoxic-ischemic conditions did not result in significant neuronal loss in the developing brain, but did cause significantly impaired dendritic growth and synapse formation alongside a disturbed development of neuronal connectivity as measured using diffusion magnetic resonance imaging (dMRI). When using more advanced acquisition settings such as high-angular resolution diffusion imaging (HARDI), more advanced reconstruction methods can be applied to investigate the cortical microstructure with higher levels of detail. Recent advances in dMRI acquisition and analysis have great potential to contribute to a better understanding of neuronal connectivity impairment in preterm birth. We will review the current understanding of abnormal preterm cortical development, novel approaches in dMRI, and the pitfalls in scanning vulnerable preterm infants.

  • Imaging fibres in the brain

    24 October 2018

  • Detailed laminar characteristics of the human neocortex revealed by NODDI

    24 October 2018

    Introduction Over the past two decades, diffusion weighted imaging (DWI) enabled detailed investigation of white matter (WM) microstructure. More recently, progress in advanced DWI methods (e.g. high-field DWI) is now allowing the cortical structure to be studied noninvasively [1]. This is a particularly exciting development, because DWI has the potential to provide the richest in vivo description of cortical cytoarchitecture. Previous work has shown that the diffusion properties of the primary visual cortex (V1) are layer-specific [2]. In particular, the stria of Gennari displays low diffusivity and anisotropy. Here, we extend these findings by fitting the recent NODDI tissue model [3] to multi-shell DWI data to support a much more fine-grained division of the V1 cortical layers. Methods DWI measurements were performed on human V1 samples (1x1x1 cm). Prior to imaging, samples were soaked in phosphate buffered saline (>72h) and mounted in a syringe with proton-free liquid under low pressure to release air bubbles from the sample ( 24h). MRI was performed on a Bruker Biospec 94/20 USR system with BGA12S HP gradient set (660 mT/m; 4570 T/m/s) equipped with a Bruker CryoProbe 1x2 coil array cooled to 20-30K. DWI was acquired with 0.2 mm isotropic resolution (read-out segmented SE-EPI with 4 segments; matrix=128x128; FOV=25.6x25.6 mm; Sample A: 45 slices; TR/TE=6750/23.8 ms; b=[0,12000,4000,12000] mm2s-1 in 768 directions; δ=7 ms; Δ=11.3 ms; TA=24h; Sample B: 40 slices; TR/TE=6000/26.7 ms; b=[0,800,3000,4000, 8000, 12000,16000,20000] mm2s-1 in 48 directions; δ=8.4 ms; Δ=12.8 ms; TA=3h). Standard DWI processing included coregistration and diffusion tensor estimation from the b=4000 shell using FSLv5.0. The datasets were fitted using NODDI, a recent multi-compartment tissue model of diffusion suitable for modeling both gray and white matter [3]. NODDI distinguishes tissue into space bounded by axons and dendrites (intra-cellular compartment) and the space surrounding neurites (extra-cellular compartment). The neurite density and orientation distribution are then quantified by the intra-cellular volume fraction and a Watson distribution (providing estimates of the dominant direction μ and the concentration of the orientations around μ, κ). NODDI includes a CSF compartment to capture CSF contamination. Here, following [4], we include an isotropic restriction compartment required for ex vivo samples. The model fit was achieved using the NODDI Matlab Toolbox. Results and Discussion In the mean diffusivity (MD), three cortical layers could be distinguished (Figs.a). These coincide with the superficial layers, the stria of Gennari (low diffusivity) and the deep layers. The neurite density (Figs.d) was found to be high in WM. In the cortex, the neurite density (Figs.d) is distinct in at least four layers: the deep layers subdivide in a inner and outer layer; the stria of Gennari is characterized by high neurite density (overlapping the hypointense layer in the MD) and the superficial layers have a very low neurite density. This reflects fiber architecture as the WM, deep layers and stria of Gennari are rich in myelinated fibers, while in the superficial cortical layers these are more sparse. These superficial layers show a more prominent isotropic fraction (Figs.f) unaffected by dense cylindrical structure. The isotropic restriction compartment (Figs.e) varies slowly over the cortex. It features a hyperintense layer at the superficial boundary of the stria of Gennari, putatively layer IVA that is rich in closely packed cells. The dispersion parameter κ (Figs.c) correlates well with fractional anisotropy (Figs.b), but κ appears to be a more sensitive measure of fiber coherence because more contrast is observed in WM as well as GM. Figure 1. Sample A (2-shell data). Figure 2. Sample B (8-shell data). Conclusion Additional detail on cortical architecture is obtained by fitting the NODDI multi-compartment model to multi-shell data when compared to diffusion-tensor metrics. Additional layers were identified not visible in the MD or FA maps. Furthermore, whereas the deep and superficial layers have comparable properties in the MD, the volume fractions from the NODDI model distinguish between these as a neurite-dense deep layer and a superficial layer sparser in neurites. The dispersion parameter κ is more sensitive than FA. As NODDI does not require extreme b-values and scan times in vivo [3], these results suggest that NODDI has more potential for capturing cortical microstructure than compared to diffusion tensor metrics. References [1] Heidemann et al. NeuroImage 2012; [2] Kleinnijenhuis et al. ISMRM 2011; [3] Zhang et al. NeuroImage 2012; [4] Alexander et al. NeuroImage 2010

  • Detailed laminar characteristics of the human neocortex revealed by NODDI and histology

    24 October 2018

    Introduction Diffusion weighted imaging (DWI) has the potential to provide the richest noninvasive description of cortical cytoarchitecture. Previous work has shown that the diffusion properties of the primary visual cortex (V1) are layer‐specific (Kleinnijenhuis et al., 2012). In particular, the stria of Gennari displays low diffusivity and anisotropy. Here, we extend these findings by fitting the recent NODDI tissue model (Zhang et al., 2012) to multi‐shell DWI data to support a more fine‐grained division of cortical layers and compare our results to histology to aid in interpretation and validation of our data. Methods Human V1 samples (1x1x1 cm) were investigated with ex vivo DWI. Prior to MR imaging, samples were fixed (>2 months), soaked in phosphate buffered saline (>72h) and mounted in a syringe with proton‐free liquid ( 24h). DWI was performed on a preclinical MR system (Bruker Biospec 94/20 USR with BGA12S HP gradient set [660 mT/m; 4570 T/m/s] equipped with a Bruker CryoProbe 1x2 coil array cooled to 20‐30K). DWI was acquired in 768 unique directions with 0.2 mm isotropic resolution (read‐out segmented SE‐EPI with 4 segments; matrix=128x128; FOV=25.6x25.6 mm; 45 slices; TR/TE=6750/23.8 ms; b=[0,12000,4000,12000] mm2s‐1 in 768 directions; δ=7 ms; Δ=11.3 ms; TA=24h). DWI volumes were coregistered using FSLv5.0 and diffusion tensors and Fiber Orientation Distributions (FODs) were estimated using MRtrix v0.2.10. The neurite density (ICVF; volume fraction occupied by cylindrical structures) and orientation dispersion (κ, concentration around the dominant direction) were fitted using the multi‐ compartment tissue model NODDI (Zhang et al., 2012). Isotropic (ISO) and isotropic restriction compartment (IRVF; required for ex vivo samples [Alexander et al., 2010]) were included. Prior to histology, the samples were bisected in the axial imaging plane and embedded in parafin for sectioning at 5 μm. Histological stains included hematoxylin and eosin (H&E) for cell bodies, Bodian for axons and Luxol Fast Blue (LFB) for myelin. The sections were digitized using a Zeiss microscope equiped with an automated table operated by Neurolucida v10 software, creating seamless virtual slices at 20x magnification. The virtual slices were processed in Matlab using structure tensor analysis described by Budde et al., 2012, i.e. coding orientation, anisotropy and staining intensity as hue, saturation and brightness. Results In the mean diffusivity map (Fig.1a), three laminar subdivisions could be distinguished in V1 cortex: the superficial layers, the stria of Gennari and the deep layers. In V1, NODDI neurite density (Fig.2a) is distinct in at least four layers: the deep layers subdivide in an inner and outer layer; the stria of Gennari is characterized by high neurite density and the superficial layers have a low neurite density. The dispersion parameter κ (Figs.2d) correlates well with fractional anisotropy (Figs.1b), but κ appears to be a more sensitive measure of fiber coherence because more contrast is observed in WM as well as GM. Fig. 3 (Bodian) and Fig.4 (LFB) show that the reconstructed FODs in Fig 1c match reasonably well with the in‐ plane orientations in the histological slice. Within the cortex, histological results show that the prominent bright layer in the κ‐map originates from increased coherence of radial fascicles of myelinated fibers penetrating layers V and IV (e.g. Fig.3f: blue fibers). The dark layer above the bright layer coincides with the stria of Gennari, as shown by the increased component of transverse fibers (e.g. Fig.4g). Discussion Additional detail on cortical architecture is obtained by fitting the NODDI multi‐compartment model to multi‐shell data when compared to diffusion‐tensor metrics. The dispersion parameter κ is more sensitive than FA. As NODDI does not require extreme b‐values and scan times in vivo (Zhang et al., 2012), these results suggest that NODDI has more potential for capturing cortical microstructure than compared to diffusion tensor metrics. The 2D FODs from the Bodian sections appear to more sensitive in detecting transverse neurite components as compared to the LFB sections.

  • A new perspective on the cortical laminar pattern of the primary visual cortex in the occipital lobes

    24 October 2018

    One of the most prominent characteristics of the human neocortex is its laminated structure. The first person to observe this was Francesco Gennari in the second half the 18th century: in the middle of the depth of primary visual cortex in the occipital lobes, myelinated fibres are so abundant that he could observe them with bare eyes as a white line. Because of its saliency, the stria of Gennari has a rich history in cyto‐ and myeloarchitectural research as well as in magnetic resonance (MR) microscopy. In the present work we show the layered structure of the human neocortex with ex vivo diffusion weighted imaging (DWI). To achieve the necessary spatial and angular resolution, primary visual cortex samples were scanned on an 11.7 T small‐animal MR system using 768 diffusion directions to characterize the diffusion properties of the cortical laminae and the stria of Gennari in particular. The results demonstrated that fractional anisotropy varied over cortical depth, showing reduced anisotropy in the stria of Gennari, the inner band of Baillarger and the deepest layer of the cortex. Orientation density functions showed multiple components in the stria of Gennari and deeper layers of the cortex. Potential applications of layer‐specific diffusion imaging include characterization of clinical abnormalities, cortical mapping and (intra)cortical tractography. We conclude that future high‐resolution in vivo cortical DWI investigations should take into account the layer‐specificity of the diffusion properties.

  • Dissectie en plastinatie van witte stof in humane hersenen voor neuroanatomisch onderwijs

    24 October 2018

    Probleemstelling Het leren van de witte stof anatomie van de humane hersenen is lastig omdat het complexe 3D netwerk van witte stof banen door de hele hersenen verspeid is in zowel de oppervlakkige als de diepe lagen van de hersenen. Door de recente ontwikkelingen in de neuroimaging met technieken zoals diffusion weighted imaging (DWI) en tractography kunnen witte stof banen worden gemeten en gevisualiseerd. Dit is van groot belang voor o.a. neuroscience en neurochirurgie. Door deze ontwikkeling is onderwijs over de 3D witte stof anatomie zeer relevant. De standaard methode in het leren van de witte stof anatomie is ontoereikend. Het doel is om leerzame en duurzame preparaten te maken van de anatomie van de witte stof banen van de humane hersenen door een combinatie van witte stof dissectie en plastinatie voor medisch onderwijs. Methode Gefixeerde humane hersenen uit gebalsemde stoffelijke overschotten zijn geprepareerd volgende de methode van Klingler. Na fixatie worden de hersenen hierbij 2x snel ingevroren. Het proces van vriezen en ontdooien vormt ijskristallen tussen de zenuwbanen van de witte stof waardoor het compacte weefsel van de witte stof losser wordt en dissectie makkelijker. Dissectie van de witte stof gebeurd met verschillende soorten spatels. Verschillende preparaten worden gemaakt om oppervlakkige en diepe witte stof structuren zichtbaar te maken. De preparaten worden vervolgend geplastineerd. Het eerste geplastineerde brein is gebruikt in een cursus van de master “Cognitive Neuro Sscience”. Resultaten Het product is een reeks breinen waarbij de witte stof anatomie 3D bestudeerd kan worden. Deze witte stof preparaten zijn veel minder kwetsbaar dan de klassieke formaline preparaten en kunnen makkelijk vervoerd worden. De eerste ervaring in het gebruik van geplastineerde witte stof breinen in een educatieve cursus is positief. Conclusie, implicaties voor de praktijk Het is mogelijk om leerzame en duurzame preparaten te maken van de anatomie van de witte stof banen van de humane hersenen door een combinatie van witte stof dissectie en plastinatie voor medisch onderwijs. Deze preparaten kunnen gebruikt worden bij het leren van de ingewikkelde 3D anatomie van de hersenen en bij het interpreteren van MRI beelden gemaakt met moderne imagingtechnieken zoals DWI –tractography. Dit is van belang voor opleidingen in neuroscience, neuroimaging en medische opleidingen bv neurochirurgen.

  • Validation of Diffusion Weighted Imaging of cortical anisotropy by means of a histological stain for myelin

    24 October 2018

    Introduction Axonal membranes and myelin are known to be the main determinant of diffusion anisotropy in the brain [1]. Myelinated axons are present not only in white matter, but extend radially into the deeper cortical layers crossing with the tangentially oriented myelinated fibers also abundant in these layers [2]. Recently, several studies have shown anisotropy of water diffusion by Diffusion Weighted Imaging (DWI) in cortical grey matter [3,4]. Cortical anisotropy has been shown e.g. ex vivo in pigs [3] and in vivo in humans [4] at high magnetic field strengths allowing for the spatial resolution necessary. In the cortex, fiber orientation was found to be predominantly radial, but more complex architectures were also observed in the deeper layers [3]. To obtain a better understanding of anatomical connectivity in vivo, it would be very interesting to track fibers also in and into the cortex. However, the high isotropic component seen in the cortex by current low resolution in vivo DWI makes the estimation of cortical connectivity by tractography very challenging. In this study we aim to examine cortical fibers and also try to validate DWI results in vitro, as there are less practical limitations and a cross-validation with histological techniques (i.e. myelin staining) as the current gold standard is possible. Methods Samples of human brain tissue ( 3 cm3) containing both cortex and underlying white matter were obtained at autopsy (15h post-mortem), fixed in 10% buffered formalin and stored (>6 months) at 4 °C. Myelin staining of the tissue blocks was performed en bloc with Luxol Fast Blue (LFB), according to a protocol modified from [5]. Sections of 2mm thickness were stained for 72 h at 56 °C in saturated LFB solution to achieve full penetration of the dye. Differentiation time in 0.05 % LiCO3 (aq) was 72 h. Samples were sectioned with a Leica vibratome at 100 µm, mounted on gelatin coated glasses and imaged at 20x magnification with a Zeiss microscope controlled by MicroBrightField software using the VirtualSlice module to create seamless high resolution images. A second sample was soaked in PBS (> 2 weeks) and imaged on a Bruker Biospec 11.7T system in Galden D05 perfluoroether. Diffusion Weighted Imaging was performed using a DW-SE sequence with segmented EPI read-out (TR=3.8s; TE=27ms; b-value=1500 s/mm2; 30 directions + 5 non-diffusion weighted; FOV = 30x30 mm, matrix size=64x64, 15 slices of 469 µm). For reconstruction of the orientation density function (ODF) Camino’s mesd function was used (filter=PAS 1.4; mepointset=30). Multi-echo gradient echo (MGE) images were also acquired (3D FLASH; TR=70ms; TE=3-38ms; ΔTE=5ms; flip angle=30°; matrix size 256x256x256; FOV 30x30x30mm). Results and Discussion The section of somatosensory cortex presented in Fig.1 shows a homogeneous staining of the white matter and excellent visualization of radii of myelinated fibers fanning out in the cortex. Moreover, tangentially oriented fibers are visible in the deeper cortical layers (inset). The DWI orientation density functions (Fig.2) seem to reflect these observations as well, with a single peak in white matter, and both a radial and a tangential peak in the cortex that decreases towards the outer layers. Conclusions We have shown successful homogeneous myelin staining of blocks of human brain tissue that can be used to validate DWI of the cortex. These data suggest good qualitative agreement in fiber architecture between DWI and histology. The myelin staining might also allow for 3D reconstructions of the fiber architecture that can be quantitatively compared to DWI ODF reconstructions as was shown in 2D rat brain slices by [6]. [1] Beaulieu, NMR in Biomed. 2002 [2] Nieuwenhuys et al., The Human CNS 4th Ed. [3] Dyrby et al., HBM 2010 [4] Heidemann et al., MRM 2010 [5] Blackwell et al., NI 2009 [6] Leergaard et al., PLoS One 2010

  • In vitro layer-specific Diffusion Weighted Imaging in human primary visual cortex

    24 October 2018

    Objective Recently, with the advent of Diffusion Weighted Imaging (DWI) at high field strength, cortical anisotropy has been shown in humans indicating a radial orientation of fibers (Heidemann et al., MRM 2010). More complex configurations have been shown in the deeper cortical layers ex vivo in pigs (Dyrby et al., HBM 2010). The variable fiber density and configurations over the layers of the human cortex (Nieuwenhuys et al., 2007) is also likely to be reflected in diffusion properties. Primary visual cortex (V1) is an excellent candidate area to investigate this, because it features the line of Gennari: the prominent layer IVb consisting of horizontal myelinated fibers. In the present study, we therefore image human V1 samples in vitro with diffusion MRI at ultra-high field strength, as this allows for the spatial resolution necessary and validation with histological techniques can be performed. Methods Human brain tissue samples of V1 including underlying white matter were fixed in 10% buffered formalin and stored at 4°C at autopsy (15 h post-mortem). Before MRI, samples were rehydrated in phosphate buffered saline (> 2weeks). MRI was performed on an 11.7T Bruker BioSpec system. Diffusion-weighted images were acquired in a DW-SE protocol using a segmented EPI readout (TR/TE=13750/26.6 ms; 61 directions + 7 non-diffusion-weighted; 14 repetitions; b-value=4000 s/mm2; FOV=28.8×28.8 mm; matrix=96×96; 55 slices of 0.3 mm). Multi-echo gradient-echo images (MGE) were acquired for anatomical reference of cortical cytoarchitecture (3D FLASH; TR=40 ms; TE=3.36-38.36 ms; ΔTE=5 ms; flip angle=30°; matrix size 256×256×256; FOV 28.8×28.8×28.8 mm). DWI and MGE volumes were realigned and coregistered. Calculation of diffusion tensors (DT), fractional anisotropy (FA) and mean diffusivity (MD) and tractography were performed with diffusion MRI toolkit Camino. MGE images were averaged over echoes. Results In the cortex FA and MD were non-uniform over layers. In particular, a layer of decreased FA (Fig.1; yellow arrow) and MD is evident throughout most of the sample that coincides well with the line of Gennari in the MGE image. A second layer of reduced FA in the deeper layers of the cortex is less pronounced but clearly visible (red arrow). DT tractography results show predominant radial fiber tracts in the cortex and many u-fibers spanning most of the gray-white matter boundary. Conclusions We have provided a clear demonstration of layer-specific diffusion parameters in the human neocortex. DT tractography results show anatomically plausible fiber reconstructions. The usefulness for connectivity research has to be investigated further, as tractography within cortical layers is challenged by a probably isotropic diffusion component within the layer (i.e. horizontal fibers are likely to be equally distributed over all within-layer directions). To elucidate this, reconstruction of the orientation density functions is a topic of active investigation, as is validation with histological methods. References Dyrby, T.B. (2010), ’An ex vivo imaging pipeline for producing high-quality and high-resolution diffusion-weighted imaging datasets’, Human Brain Mapping, Epub May 13 2010. Heidemann, R.M. (2010), ’Diffusion Imaging in Humans at 7T Using Readout-Segmented EPI and GRAPPA’, Magnetic Resonance in Medicine, vol. 64, pp. 9-14 Nieuwenhuys, R. (2007), ’The Human Central Nervous System’ 4th Ed, Springer, Berlin.