Manisha Aggarwal

Magnetic resonance imaging and micro-computed tomography combined atlas of developing and adult mouse brains for stereotaxic surgery

Stereotaxic atlases of the mouse brain are important in neuroscience research for targeting of specific internal brain structures during surgical operations. The effectiveness of stereotaxic surgery depends on accurate mapping of the brain structures relative to landmarks on the skull. During postnatal development in the mouse, rapid growth-related changes in the brain occur concurrently with growth of bony plates at the cranial sutures, therefore adult mouse brain atlases cannot be used to precisely guide stereotaxis in developing brains. In this study, three-dimensional stereotaxic atlases of C57BL/6J mouse brains at six postnatal developmental stages: postnatal day (P) 7, P14, P21, P28, P63 and in adults (P140-P160) were developed, using diffusion tensor imaging (DTI) and micro-computed tomography (CT). At present, most widely-used stereotaxic atlases of the mouse brain are based on histology, but the anatomical fidelity of ex vivo atlases to in vivo mouse brains has not been evaluated previously. To account for ex vivo tissue distortion due to fixation as well as individual variability in the brain, we developed a population-averaged in vivo magnetic resonance imaging adult mouse brain stereotaxic atlas, and a distortion-corrected DTI atlas was generated by nonlinearly warping ex vivo data to the population-averaged in vivo atlas. These atlas resources were developed and made available through a new software user-interface with the objective of improving the accuracy of targeting brain structures during stereotaxic surgery in developing and adult C57BL/6J mouse brains.

Three-dimensional diffusion tensor microimaging for anatomical characterization of the mouse brain

Diffusion tensor imaging is gaining increasing importance for anatomical imaging of the developing mouse brain. However, the application of diffusion tensor imaging to mouse brain imaging at microscopic levels is hindered by the limitation on achievable spatial resolution. In this study, fast diffusion tensor microimaging of the mouse brain, based on a diffusion‐weighted gradient and spin echo technique with twin‐navigator echo phase correction, is presented. Compared to echo planar and spin echo acquisition, the diffusion‐weighted gradient and spin echo acquisition resulted in significant reduction in scan time and had minimal image distortion, thereby allowing acquisition at higher spatial resolution. In this study, three‐dimensional diffusion tensor microimaging of the mouse brains at spatial resolutions of 50‐60 μm revealed unprecedented anatomical details. Thin fiber bundles in the adult striatum and white matter tracts in the embryonic day 12 mouse brains were visualized for the first time. The study demonstrated that data acquired using the diffusion tensor microimaging technique allow three‐dimensional mapping of gene expression data and can serve as a platform to study gene expression patterns in the context of neuroanatomy in the developing mouse brain. Magn Reson Med 64:249–261, 2010.

Probing mouse brain microstructure using oscillating gradient diffusion MRI.

High resolution diffusion tensor images of the mouse brain were acquired using the pulsed gradient spin echo sequence and the oscillating gradient spin echo sequence. The oscillating gradient spin echo tensor images demonstrated frequency‐dependent changes in diffusion measurements, including apparent diffusion coefficient and fractional anisotropy, in major brain structures. Maps of the rate of change in apparent diffusion coefficient with oscillating gradient frequency revealed novel tissue contrast in the mouse hippocampus, cerebellum, and cerebral cortex. The observed frequency‐dependent contrasts resembled neuronal soma‐specific Nissl staining and nuclei‐specific 4′,6‐diamidino‐2‐phenylindole (DAPI) staining in the mouse brain, which suggests that the contrasts might be related to key features of cytoarchitecture in the brain. In the mouse cuprizone model, oscillating gradient spin echo‐based diffusion MRI revealed significantly higher frequency‐dependence of perpendicular diffusivity (λ⊥) in the demyelinated caudal corpus callosum at 4 weeks after cuprizone treatment when compared with control mice and mice at 6 weeks after cuprizone treatment. The elevated frequency‐dependence of λ⊥ coincided with the infiltration of activated microglia/macrophages and disruption of axons during acute demyelination in the caudal corpus callosum. The results demonstrate the potential of oscillating gradient spin echo‐based diffusion MRI for providing tissue contrasts complimentary to conventional pulsed gradient spin echo‐based diffusion MRI. Magn Reson Med 67:98–109, 2012.

Structural insights into the rodent CNS via diffusion tensor imaging.

Diffusion tensor imaging (DTI) is a useful tool for studying anatomy and pathology in the rodent central nervous system (CNS).The unique tissue contrasts provided by DTI are well suited for monitoring disease progression, studying brain development, and characterizing anatomical phenotypes. Recent technical developments have vastly improved the speed and resolution of rodent DTI. Ongoing research efforts exploring the microstructural basis of DTI signals have provided useful insights into its capabilities to delineate brain structures and detect neuropathology. Significant progress has also been made in combining DTI results with data acquired using other imaging modalities to enhance our understanding of the rodent CNS.

Structural MRI detects progressive regional brain atrophy and neuroprotective effects in N171-82Q Huntington's disease mouse model

Huntingtons disease (HD) displays progressive striatal atrophy that occurs long before the onset of clinical motor symptoms. As there is no treatment for the disease once overt symptoms appear, it has been suggested that neuroprotective therapy given during this presymptomatic period might slow progression of the disease. This requires biomarkers that can reliably detect early changes and are sensitive to treatment response. In mouse models of HD, structural MRI measures have been shown to detect disease onset. To determine whether such measures could also be suitable biomarkers for following responses to treatment, we used T2-weighted MR imaging combined with automated morphological analyses and characterized changes in regional brain volumes longitudinally in the N171-82Q HD mouse model in a preclinical trial. We report here that N171-82Q HD mice exhibit adult-onset and progressive brain atrophy in the striatum and neocortex as well as in whole brain; the progressive atrophy in striatum and neocortex is positively correlated with motor deficits. Most notably, MRI also detected neuroprotective effects of sertraline treatment, a neuroprotective agent confirmed in our previous studies. Our present studies provide the first evidence that longitudinal structural MRI measures can detect the therapeutic effect in HD mice, suggesting that such measures in brain could be valuable biomarkers in HD clinical trials.

Feasibility of creating a high-resolution 3D diffusion tensor imaging based atlas of the human brainstem: a case study at 11.7 T.

A three-dimensional stereotaxic atlas of the human brainstem based on high resolution ex vivo diffusion tensor imaging (DTI) is introduced. The atlas consists of high resolution (125-255 μm isotropic) three-dimensional DT images of the formalin-fixed brainstem acquired at 11.7 T. The DTI data revealed microscopic neuroanatomical details, allowing three-dimensional visualization and reconstruction of fiber pathways including the decussation of the pyramidal tract fibers, and interdigitating fascicles of the corticospinal and transverse pontine fibers. Additionally, strong gray-white matter contrasts in the apparent diffusion coefficient (ADC) maps enabled precise delineation of gray matter nuclei in the brainstem, including the cranial nerve and the inferior olivary nuclei. Comparison with myelin-stained histology shows that at the level of resolution achieved in this study, the structural details resolved with DTI contrasts in the brainstem were comparable to anatomical delineation obtained with histological sectioning. Major neural structures delineated from DTI contrasts in the brainstem are segmented and three-dimensionally reconstructed. Further, the ex vivo DTI data are nonlinearly mapped to a widely-used in vivo human brain atlas, to construct a high-resolution atlas of the brainstem in the Montreal Neurological Institute (MNI) stereotaxic coordinate space. The results demonstrate the feasibility of developing a 3D DTI based atlas for detailed characterization of brainstem neuroanatomy with high resolution and contrasts, which will be a useful resource for research and clinical applications.

Probing region-specific microstructure of human cortical areas using high angular and spatial resolution diffusion MRI

Regional heterogeneity in cortical cyto- and myeloarchitecture forms the structural basis of mapping of cortical areas in the human brain. In this study, we investigate the potential of diffusion MRI to probe the microstructure of cortical gray matter and its region-specific heterogeneity across cortical areas in the fixed human brain. High angular resolution diffusion imaging (HARDI) data at an isotropic resolution of 92-μm and 30 diffusion-encoding directions were acquired using a 3D diffusion-weighted gradient-and-spin-echo sequence, from prefrontal (Brodmann area 9), primary motor (area 4), primary somatosensory (area 3b), and primary visual (area 17) cortical specimens (n=3 each) from three human subjects. Further, the diffusion MR findings in these cortical areas were compared with histological silver impregnation of the same specimens, in order to investigate the underlying architectonic features that constitute the microstructural basis of diffusion-driven contrasts in cortical gray matter. Our data reveal distinct and region-specific diffusion MR contrasts across the studied areas, allowing delineation of intracortical bands of tangential fibers in specific layers-layer I, layer VI, and the inner and outer bands of Baillarger. The findings of this work demonstrate unique sensitivity of diffusion MRI to differentiate region-specific cortical microstructure in the human brain, and will be useful for myeloarchitectonic mapping of cortical areas as well as to achieve an understanding of the basis of diffusion NMR contrasts in cortical gray matter.

The role of myelination in measures of white matter integrity: Combination of diffusion tensor imaging and two-photon microscopy of CLARITY intact brains

ABSTRACT Diffusion tensor imaging (DTI) is used extensively in neuroscience to noninvasively estimate white matter (WM) microarchitecture. However, the diffusion signal is inherently ambiguous because it infers WM structure from the orientation of water diffusion and cannot identify the biological sources of diffusion changes. To compare inferred WM estimates to directly labeled axonal elements, we performed a novel within‐subjects combination of high‐resolution ex vivo DTI with two‐photon laser microscopy of intact mouse brains rendered optically transparent by Clear Lipid‐exchanged, Anatomically Rigid, Imaging/immunostaining compatible, Tissue hYdrogel (CLARITY). We found that myelin basic protein (MBP) immunofluorescence significantly correlated with fractional anisotropy (FA), especially in WM regions with coherent fiber orientations and low fiber dispersion. Our results provide evidence that FA is particularly sensitive to myelination in WM regions with these characteristics. Furthermore, we found that radial diffusivity (RD) was only sensitive to myelination in a subset of WM tracts, suggesting that the association of RD with myelin should be used cautiously. This combined DTI‐CLARITY approach illustrates, for the first time, a framework for using brain‐wide immunolabeling of WM targets to elucidate the relationship between the diffusion signal and its biological underpinnings. This study also demonstrates the feasibility of a within‐subject combination of noninvasive neuroimaging and tissue clearing techniques that has broader implications for neuroscience research.

Region-specific gene expression in early postnatal mouse thalamus

Previous studies in the developing mouse thalamus have demonstrated that regional identity is established during early stages of development (Suzuki‐Hirano et al. J. Comp. Neurol. 2011;519:528–543). However, the developing thalamus often shows little resemblance to the anatomical organization of the postnatal thalamus, making it difficult to identify genes that might mediate the organization of thalamic nuclei. We therefore analyzed the expression pattern of genes that we have identified as showing regional expression in embryonic thalamus on postnatal days (P) 6–8 by using in situ hybridization. We also identified several genes expressed only in the postnatal thalamus with restricted expression in specific nuclei. We first demonstrated the selective expression of neurotransmitter‐related genes (vGlut2, vGAT, D2R, and HTR2C), identifying the neurotransmitter subtypes of cells in this region, and we also demonstrated selective expression of additional genes in the thalamus (Steel, Slitrk6, and AI852580). In addition, we demonstrated expression of genes specific to somatosensory thalamic nuclei, the ventrobasal posterior nuclei (VP); a visual thalamic nucleus, the dorsal lateral geniculate nucleus (dLGN); and an auditory thalamic nucleus, the medial geniculate body (MGB) (p57Kip, Nr1d1, and GFRα1). We also identified genes that are selectively expressed in multiple different nuclei (Foxp2, Chst2, and EphA8). Finally, we demonstrated that several bone morphogenetic proteins (BMPs) and their inhibitors are expressed in the postnatal thalamus in a nucleus‐specific fashion, suggesting that BMPs play roles in the postnatal thalamus unrelated to their known role in developmental patterning. Our findings provide important information for understanding the mechanisms of nuclear specification and connectivity during development, as well as their maintenance in adult thalamus. J. Comp. Neurol. 519:544–561, 2011.

Spatiotemporal mapping of brain atrophy in mouse models of Huntington's disease using longitudinal in vivo magnetic resonance imaging

Mouse models of Huntingtons disease (HD) that recapitulate some of the phenotypic features of human HD, play a crucial role in investigating disease mechanisms and testing potential therapeutic approaches. Longitudinal studies of these models can yield valuable insights into the temporal course of disease progression and the effect of drug treatments on the progressive phenotypes. Atrophy of the brain, particularly the striatum, is a characteristic phenotype of human HD, is known to begin long before the onset of motor symptoms, and correlates strongly with clinical features. Elucidating the spatial and temporal patterns of atrophy in HD mouse models is important to characterize the phenotypes of these models, as well as evaluate the effects of neuroprotective treatments at specific time frames during disease progression. In this study, three dimensional in vivo magnetic resonance imaging (MRI) and automated longitudinal deformation-based morphological analysis was used to elucidate the spatial and temporal patterns of brain atrophy in the R6/2 and N171-82Q mouse models of HD. Using an established MRI-based brain atlas and mixed-effects modeling of deformation-based metrics, we report the rates of progression and region-specificity of brain atrophy in the two models. Further, the longitudinal analysis approach was used to evaluate the effects of sertraline and coenzyme Q(10) (CoQ(10)) treatments on progressive atrophy in the N171-82Q model. Sertraline treatment resulted in significant slowing of atrophy, especially in the striatum and frontal cortex regions, while no significant effects of CoQ(10) treatment were observed. Progressive cortical and striatal atrophy in the N171-82Q mice showed significant positive correlations with measured functional deficits. The findings of this report can be used for future testing and comparison of potential therapeutics in mouse models of HD.