Melanie Martin

Quantitative MRI and ultrastructural examination of the cuprizone mouse model of demyelination

The cuprizone mouse model of demyelination was used to investigate the influence that white matter changes have on different magnetic resonance imaging results. In vivo T2‐weighted and magnetization transfer images (MTIs) were acquired weekly in control (n = 5) and cuprizone‐fed (n = 5) mice, with significant increases in signal intensity in T2‐weighted images (p < 0.001) and lower magnetization transfer ratio (p < 0.001) in the corpus callosum of the cuprizone‐fed mice starting at 3 weeks and peaking at 4 and 5 weeks, respectively. Diffusion tensor imaging (DTI), quantitative MTI (qMTI), and T1/T2 measurements were used to analyze freshly excised tissue after 6 weeks of cuprizone administration. In multicomponent T2 analysis with 10 ms echo spacing, there was no visible myelin water component associated with the short T2 value. Quantitative MTI metrics showed significant differences in the corpus callosum and external capsule of the cuprizone‐fed mice, similar to previous studies of multiple sclerosis in humans and animal models of demyelination. Fractional anisotropy was significantly lower and mean, axial, and radial diffusivity were significantly higher in the cuprizone‐fed mice. Cellular distributions measured in electron micrographs of the corpus callosum correlated strongly to several different quantitative MRI metrics. The largest Spearman correlation coefficient varied depending on cellular type: T1 versus the myelinated axon fraction (ρ = −0.90), the bound pool fraction (ƒ) versus the myelin sheath fraction (ρ = 0.93), and axial diffusivity versus the non‐myelinated cell fraction (ρ = 0.92). Using Pearsons correlation coefficient, ƒ was strongly correlated to the myelin sheath fraction (r = 0.98) with a linear equation predicting myelin content (5.37ƒ − 0.25). Of the calculated MRI metrics, ƒ was the strongest indicator of myelin content, while longitudinal relaxation rates and diffusivity measurements were the strongest indicators of changes in tissue structure. Copyright

Cerebellar cortical atrophy in experimental autoimmune encephalomyelitis

Brain atrophy measured by MRI is an important correlate with clinical disability and disease duration in multiple sclerosis (MS). Unfortunately, neuropathologic mechanisms which lead to this grey matter atrophy remain unknown. The objective of this study was to determine whether brain atrophy occurs in the mouse model, experimental autoimmune encephalomyelitis (EAE). Postmortem high-resolution T2-weighted magnetic resonance microscopy (MRM) images from 32 mouse brains (21 EAE and 11 control) were collected. A minimum deformation atlas was constructed and a deformable atlas approach was used to quantify volumetric changes in neuroanatomical structures. A significant decrease in the mean cerebellar cortex volume in mice with late EAE (48-56 days after disease induction) as compared to normal strain, gender, and age-matched controls was observed. There was a direct correlation between cerebellar cortical atrophy and disease duration. At an early time point in disease, 15 days after disease induction, cerebellar white matter lesions were detected by both histology and MRM. These data demonstrate that myelin-specific autoimmune responses can lead to grey matter atrophy in an otherwise normal CNS. The model described herein can now be used to investigate neuropathologic mechanisms that lead to the development of gray matter atrophy in this setting.

Purkinje Cell Loss in Experimental Autoimmune Encephalomyelitis

Gray matter atrophy observed by brain MRI is an important correlate to clinical disability and disease duration in multiple sclerosis. The objective of this study was to link brain atrophy visualized by neuroimaging to its underlying neuropathology using the MS model, experimental autoimmune encephalomyelitis (EAE). Volumetric changes in brains of EAE mice, as well as matched healthy normal controls, were quantified by collecting post-mortem high-resolution T2-weighted magnetic resonance microscopy and actively stained magnetic resonance histology images. Anatomical delineations demonstrated a significant decrease in the volume of the whole cerebellum, cerebellar cortex, and molecular layer of the cerebellar cortex in EAE as compared to normal controls. The pro-apoptotic marker caspase-3 was detected in Purkinje cells and a significant decrease in Purkinje cell number was found in EAE. Cross modality and temporal correlations revealed a significant association between Purkinje cell loss on neuropathology and atrophy of the molecular layer of the cerebellar cortex by neuroimaging. These results demonstrate the power of using combined population atlasing and neuropathology approaches to discern novel insights underlying gray matter atrophy in animal models of neurodegenerative disease.

Region-Specific Myelin Pathology in Mice Lacking the Golli Products of the Myelin Basic Protein Gene

The myelin basic protein (MBP) gene encodes two families of proteins, the classic MBP constituents of myelin and the golli-MBPs, the function of which is less well understood. In this study, targeted ablation of the golli-MBPs, but not the classic MBPs, resulted in a distinct phenotype unlike that of knock-outs (KOs) of the classic MBPs or other myelin proteins. Although the golli KO animals did not display an overt dysmyelinating phenotype, they did exhibit delayed and/or hypomyelination in selected areas of the brain, such as the visual cortex and the optic nerve, as determined by Northern and Western blots and immunohistochemical analysis with myelin protein markers. Hypomyelination in some areas, such as the visual cortex, persisted into adulthood. Ultrastructural analysis of the KOs confirmed both the delay and hypomyelination and revealed abnormalities in myelin structure and in some oligodendrocytes. Abnormal visual-evoked potentials indicated that the hypomyelination in the visual cortex had functional consequences in the golli KO brain. Evidence that the abnormal myelination in these animals was a consequence of intrinsic problems with the oligodendrocyte was indicated by an impaired ability of oligodendrocytes to form myelin sheets in culture and by the presence of abnormal Ca2+ transients in purified cortical oligodendrocytes studied in vitro. The Ca2+ results reported in this study complement previous results implicating golli proteins in modulating intracellular signaling in T-cells. Together, all these findings suggest a role for golli proteins in oligodendrocyte differentiation, migration, and/or myelin elaboration in the brain.

Digital Three-Dimensional Atlas of Quail Development Using High-Resolution MRI

We present an archetypal set of three-dimensional digital atlases of the quail embryo based on microscopic magnetic resonance imaging (μMRI). The atlases are composed of three modules: (1) images of fixed ex ovo quail, ranging in age from embryonic day 5 to 10 (e05 to e10); (2) a coarsely delineated anatomical atlas of the μMRI data; and (3) an organ system-based hierarchical graph linked to the anatomical delineations. The atlas is designed to be accessed using SHIVA, a free Java application. The atlas is extensible and can contain other types of information including anatomical, physiological, and functional descriptors. It can also be linked to online resources and references. This digital atlas provides a framework to place various data types, such as gene expression and cell migration data, within the normal three-dimensional anatomy of the developing quail embryo. This provides a method for the analysis and examination of the spatial relationships among the different types of information within the context of the entire embryo.

A Comparison of Rapid-Scanning X-Ray Fluorescence Mapping And Magnetic Resonance Imaging to Localize Brain Iron Distribution

The clinical diagnosis of many neurodegenerative disorders relies primarily or exclusively on observed behaviors rather than measurable physical tests. One of the hallmarks of Alzheimer disease (AD) is the presence of amyloid-containing plaques associated with deposits of iron, copper and/or zinc. Work in other laboratories has shown that iron-rich plaques can be seen in the mouse brain in vivo with magnetic resonance imaging (MRI) using a high-field strength magnet but this iron cannot be visualized in humans using clinical magnets. To improve the interpretation of MRI, we correlated iron accumulation visualized by X-ray fluorescence spectroscopy, an element-specific technique with T1, T2, and susceptibility weighted MR (SWI) in a mouse model of AD. We show that SWI best shows areas of increased iron accumulation when compared to standard sequences.

Evidence for the involvement of calbindin D28k in the presenilin 1 model of Alzheimer's disease.

Pathological hallmarks of Alzheimers disease include memory deficits, accumulation of amyloid beta (Abeta) plaques, the appearance of neurofibrillary tangles, and dysregulation of calcium homeostasis, which has been linked to mutations in the presenilin gene that code for presenilin (PS) proteins. PSs are a family of multi-pass transmembrane proteins where normal presenilins (PS1 and PS2) are highly localized in the endoplasmic reticulum (ER). Several past studies have explored alterations in long-term potentiation (LTP), a proposed molecular correlate of memory, and in behavioral tests of spatial memory in a variety of PS1 models. These reports suggest that calcium plays a role in these alterations, but mechanistic explanations for changes in LTP and in behavioral tests of memory are still lacking. To test the hypothesis that calcium-related mechanisms, such as changes in calcium buffering, are associated with alterations in LTP and memory, we utilized in vitro experimental paradigms of LTP in hippocampal slices obtained from the PS1-M146V transgenic mouse model of Alzheimers disease (AD). We also used the in vivo Morris water maze (MWM), a test for hippocampal dependent spatial memory. In addition, we used cellular assays to explore molecular mechanisms. We confirm that PS1 mutations (M146V) enhance LTP. We also find increases in some parameters of the MWM, and alterations in other parameters, such as path length indicating impairment in cognitive functioning in PS1-M146V mice. In addition, these findings are observed in association with increased calbindin D28K expression in the CA1 hippocampus of PS1-M146V mice.

Neurofibrillary tangles and plaques are not accompanied by white matter pathology in aged triple transgenic-Alzheimer disease mice

Alzheimers disease (AD) is a progressive neurodegenerative disorder that is the most common cause of dementia in aging populations. Although senile plaques and neurofibrillary tangles are well-established hallmarks of AD, changes in cerebral white matter correlate with cognitive decline and may increase the risk of the development of dementia. We used the triple transgenic (3xTg)-AD mouse model of AD, previously used to show that white matter changes precede plaque formation, to test the hypothesis that MRI detectable changes occur in the corpus callosum, external capsule and the fornix. T2-weighted and diffusion tensor magnetic resonance imaging and histological stains were employed to assess white matter in older (11-17months) 3xTg-AD mice and controls. We found no statistically significant changes in white matter between 3xTg-AD mice and controls, despite well-developed neurofibrillary tangles and beta amyloid immunoreactive plaques. Myelin staining was normal in affected mice. These data suggest that the 3xTg-AD mouse model does not develop MRI detectable white matter changes at the ages we examined.

Intracortical injection of endothelin-1 induces cortical infarcts in mice: effect of neuronal expression of an adenosine transporter

BackgroundActivation of adenosine A1 receptors has neuroprotective effects in animal stroke models. Adenosine levels are regulated by nucleoside transporters. In vitro studies showed that neuron-specific expression of human equilibrative nucleoside transporter 1 (hENT1) decreases extracellular adenosine levels and adenosine A1 receptor activity. In this study, we tested the effect of hENT1 expression on cortical infarct size following intracerebral injection of the vasoconstrictor endothelin-1 (ET-1) or saline.MethodsMice underwent stereotaxic intracortical injection of ET-1 (1 μl; 400 pmol) or saline (1 μl). Some mice received the adenosine receptor antagonist caffeine (25 mg/kg, intraperitoneal) 30 minutes prior to ET-1. Perfusion and T2-weighted magnetic resonance imaging (MRI) were used to measure cerebral blood flow (CBF) and subsequent infarct size, respectively.ResultsET-1 reduced CBF at the injection site to 7.3 ± 1.3% (n = 12) in hENT1 transgenic (Tg) and 12.5 ± 2.0% (n = 13) in wild type (Wt) mice. At 48 hours following ET-1 injection, CBF was partially restored to 35.8 ± 4.5% in Tg and to 45.2 ± 6.3% in Wt mice; infarct sizes were significantly greater in Tg (9 ± 1.1 mm3) than Wt (5.4 ± 0.8 mm3) mice. Saline-treated Tg and Wt mice had modest decreases in CBF and infarcts were less than 1 mm3. For mice treated with caffeine, CBF values and infarct sizes were not significantly different between Tg and Wt mice.ConclusionsET-1 produced greater ischemic injury in hENT1 Tg than in Wt mice. This genotype difference was not observed in mice that had received caffeine. These data indicate that hENT1 Tg mice have reduced ischemia-evoked increases in adenosine receptor activity compared to Wt mice.

Comparison of manual and semi-automated segmentation methods to evaluate hippocampus volume in APP and PS1 transgenic mice obtained via in vivo magnetic resonance imaging

BACKGROUND Magnetic resonance imaging (MRI) of transgenic mouse models of Alzheimers disease is valuable to understand better the structural changes that occur in the brain and could provide a means to test drug treatments. A hallmark pathological feature of Alzheimers disease is atrophy of the hippocampus, which is an early biomarker of the disease. MRI can be used to detect and monitor this biomarker. METHOD Repeated measurements using in vivo 3D T2-weighted imaging of mice were used to assess the methods. Each mouse was imaged twice in one week and twice the following week and no changes in volume were expected. The hippocampus was segmented both manually and semi-automatically. Registration was done to gain information on shape changes. The volumes from each mouse were compared intra-mouse, between mice and to hippocampus volume values in the literature. RESULTS A reliable method was developed which was able to detect difference in volumes of hippocampus between mice when performed by a single individual. The semi-automated segmentation was unable to detect the same level of differences. The semi-automated segmentation method gave larger hippocampus volumes, with 78-87% reliability between the manual and semi-automated segmentation. Although more accurate, the manual segmentation is laborious and suffers from inter- and intra-variability. CONCLUSION These results suggest that manual segmentation is still considered the most reliable segmentation method for small structures. However, if performing longitudinal studies, where there is at least one year between imaging sessions, the segmentation should be done all at once at the end of all the imaging sessions. If segmentation is done after each imaging session, with at least a year passing between segmentations, very small variations in volumes can be missed. This method provides a means to quantify the volume of the hippocampus in a live mouse using manual segmentation, which is the first step toward studying hippocampus atrophy in a mouse model of Alzheimers disease.