Brain virtual histology with X-ray phase-contrast tomography Part I: whole-brain myelin mapping in white-matter injury models

White-matter injury leads to severe functional loss in many neurological diseases. Myelin staining on histological samples is the most common technique to investigate white-matter fibers. However, tissue processing and sectioning may affect the reliability of 3D volumetric assessments. The purpose of this study was to propose an approach that enables myelin fibers to be mapped in the whole rodent brain with microscopic resolution and without the need for strenuous staining. With this aim, we coupled in-line (propagation-based) X-ray phase-contrast tomography (XPCT) to ethanol-induced brain sample dehydration. We here provide the proof-of-concept that this approach enhances myelinated axons in rodent and human brain tissue. In addition, we demonstrated that white-matter injuries could be detected and quantified with this approach, using three animal models: ischemic stroke, premature birth and multiple sclerosis. Furthermore, in analogy to diffusion tensor imaging (DTI), we retrieved fiber directions and DTI-like diffusion metrics from our XPCT data to quantitatively characterize white-matter microstructure. Finally, we showed that this non-destructive approach was compatible with subsequent complementary brain sample analysis by conventional histology. In-line XPCT might thus become a novel gold-standard for investigating white-matter injury in the intact brain. This is Part I of a series of two articles reporting the value of in-line XPCT for virtual histology of the brain; Part II shows how in-line XPCT enables the whole-brain 3D morphometric analysis of amyloid-β (Aβ) plaques.HighlightsX-ray phase-contrast tomography (XPCT) enables myelin mapping of the whole brainXPCT detects and quantifies white-matter injuries in a range of diseasesFiber directions and anisotropy metrics can be retrieved from XPCT dataXPCT is compatible with subsequent conventional histology of brain samplesXPCT is a powerful virtual histology tool that requires minimal sample preparationGraphical Abstract

Proteomics analysis of a human brain sample from a mucolipidosis type IV patient reveals pathophysiological pathways

Background Mucolipidosis type IV (MLIV), an ultra-rare neurodevelopmental and neurodegenerative disorder, is caused by mutations in the MCOLN1 gene, which encodes the late endosomal/lysosomal transient receptor potential channel TRPML1 (mucolipin 1). The precise pathophysiogical pathways that cause neurological disease in MLIV are poorly understood. Recently, the first post-mortem brain sample became available from a single MLIV patient, and in the current study we performed mass spectrometry (MS)-based proteomics on this tissue with a view to delineating pathological pathways, and to compare with previously-published data on MLIV, including studies using the Mcoln1−/− mouse. Results A number of pathways were altered in two brain regions from the MLIV patient, including those related to the lysosome, lipid metabolism, myelination, cellular trafficking and autophagy, mTOR and calmodulin, the complement system and interferon signaling. Of these, levels of some proteins not known previously to be associated with MLIV were altered, including APOD, PLIN4, ATG and proteins related to interferon signaling. Moreover, when proteins detected by proteomics in the human brain were compared with their orthologs detected in the Mcoln1−/− mouse by RNAseq, the results were remarkably similar. Finally, analysis of proteins in human and mouse CSF suggest that calbindin 1 and calbindin 2 might be useful as biomarkers to help chart the course of disease development. Conclusions Despite the sample size limitations, our findings are consistent with the relatively general changes in lysosomal function previously reported in MLIV, and shed light on new pathways of disease pathophysiology, which is required in order to understand the course of disease development and to determine the efficacy of therapies when they become available for this devastating disease.

Confirmation of Rabies in a Stray Dog in Umuahia, Nigeria – A Case Report.

Rabies is an important zoonotic disease and the increased numbers of stray dogs constitute a constraint on the prevention of human cases in Nigeria. This report reaffirms that rabies is endemic in Nigeria and stray dogs constitute a risk to humans. A stray Nigeria indigenous dog was presented to the Veterinary Teaching Hospital, Michael Okpara University of Agriculture, Umudike Nigeria with clinical manifestations of salivation, delirium, maniac and attack displays. It was quarantined for a period of 14 days, clinical manifestations persisted and a presumptive diagnosis of end stage furious form of rabies was made. The dog was then humanely euthanized, brain sample was collected and taken to the National Veterinary Research Institute (NVRI), Vom, Nigeria for rabies virus detection using direct fluorescent antibody test (DFAT). Result revealed that brain sample was positive for canine rabies depicted by apple green fuorescence. In conclusion, the stray dog was confirmed to be positive for rabies. To the authors’ knowledge, this is the first report of confirmed rabies case in a stray dog in Umuahia, Nigeria. Hence, there should be intense public awareness on the danger of rabies since the disease remains a public health concern in Nigeria.

Neuroanatomical underpinning of diffusion kurtosis measurements in the cerebral cortex of healthy macaque brains

PurposeTo investigate the neuroanatomical underpinning of healthy macaque brain cortical microstructure measured by diffusion kurtosis imaging (DKI) which characterizes non-Gaussian water diffusion.MethodsHigh-resolution DKI was acquired from 6 postmortem macaque brains. Neurofilament density (ND) was quantified based on structure tensor from neurofilament histological images of a different macaque brain sample. After alignment of DKI-derived mean kurtosis (MK) maps to the histological images, MK and histology-based ND were measured at corresponding regions of interests characterized by distinguished cortical MK values in the prefrontal/precentral-postcentral and temporal cortices. Pearson correlation was performed to test significant correlation between these cortical MK and ND measurements.ResultsHeterogeneity of cortical MK across different cortical regions was revealed, with significantly and consistently higher MK measurements in the prefrontal/precentral-postcentral cortex compared to those in the temporal cortex across all 6 scanned macaque brains. Corresponding higher ND measurements in the prefrontal/precentral-postcentral cortex than in the temporal cortex were also found. The heterogeneity of cortical MK is associated with heterogeneity of histology-based ND measurements, with significant correlation between cortical MK and corresponding ND measurements (P <0.005).ConclusionThese findings suggested that DKI-derived MK can potentially be an effective noninvasive biomarker quantifying underlying neuroanatomical complexity inside the cerebral cortical mantle for clinical and neuroscientific research.

Relationship between age and brainstem allometry in the African grasscutter (Thryonomys swinderianus Temminck, 1827)

Allometric values of brainstem structures were evaluated in African grasscutters Thryonomys swinderianus (n = 27). Brain samples were extracted from 9 animals each of 3 days (neonates), 72 days (juveniles) and 450 days of age (adults). The midbrain, pons and medulla oblongata were separated from each brain sample and dimensions and weights obtained. The weights of the midbrain in the neonate, juvenile and adult African grasscutters were 0.33 g ± 0.01 g, 0.47 g ± 0.01 g and 0.93 g ± 0.02 g, respectively. The increase from neonate to juvenile (p = 0.002) and adult (p = 0.003) was significant. The pons lengths in the neonate, juvenile and adult were 2.05 mm ± 0.05 mm, 3.86 mm ± 0.05 mm and 4.16 mm ± 0.22 mm, respectively. There was a significant increase in the length of the pons from the neonate to the juvenile (p = 0.002), but the increase from the juvenile to the adult period was not significant (p = 0.263). There was also a significant (p < 0.05) increase in the weights and lengths of the medulla oblongata from neonate to juvenile and adult periods. In adults, the nose-rump length and the length of the medulla were significantly negatively correlated (r² = 0.47; p = 0.043). The present study concluded that the postnatal development of some brainstem structures in the African grasscutter varies with age.

Quantitative mRNA Imaging Throughout the Entire Drosophila Brain

We describe a fluorescence in situ hybridization method that permits detection of the localization and abundance of single mRNAs (smFISH) in cleared whole-mount adult Drosophila brains. The approach is rapid, multiplexable and does not require molecular amplification; it allows facile mRNA expression quantification with subcellular resolution on a standard confocal microscope. Using a custom Bessel Beam-Structured Illumination microscope (BB-SIM), we further demonstrate single-mRNA detection across the entire brain sample.