Protocol for using serial two-photon tomography to map cell types and cerebrovasculature at single-cell resolution in the whole adult mouse brain

Liwang, Josephine; Bennett, Hannah C.; Pi, Hyun Jae; Kim, Yongsoo

doi:10.1016/j.xpro.2023.102048

Cited by 5 publications

(9 citation statements)

References 12 publications

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“…We labeled the brain vasculature by cardiac perfusion of fluorescein isothiocyanate (FITC)-conjugated albumin gel 27,29,31,32 . Then, we utilized serial two-photon tomography (STPT) imaging to image the whole mouse brain at 1x1x5 µm resolution ( x,y,z; media-lateral, dorsal-ventral, rostral-caudal) followed by computational analysis for vasculature tracing and quantification 29,33 . All signals were registered to the Allen Common Coordinate Framework (AllenCCF) as a reference brain 34 (Figure 1).…”

Section: Resultsmentioning

confidence: 99%

“…To quantitatively determine changes of pericytes, we compared capillary pericyte densities in 2-month-old and 18-month-old PDGFRβ-Cre;Ai14 mice 37 , 38 , where tdTomato is expressed in pericytes and other mural cells. We used STPT imaging of PDGFRβ-Cre;Ai14 mice with previously developed computational analyses to image, identify, and quantify changes of capillary pericytes upon aging across the whole mouse brain 28 , 31 ( Figure 1 bottom and Figure 3A ).…”

Section: Resultsmentioning

confidence: 99%

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Aging drives cerebrovascular network remodeling and functional changes in the mouse brain

Bennett

Zhang

et al. 2023

Preprint

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Aging is the largest risk factor for neurodegenerative disorders, and commonly associated with compromised cerebrovasculature and pericytes. However, we do not know how normal aging differentially impacts the vascular structure and function in different brain areas. Here we utilize mesoscale microscopy methods (serial two-photon tomography and light sheet microscopy) and in vivo imaging (wide field optical spectroscopy and two-photon imaging) to determine detailed changes in aged cerebrovascular networks. Whole-brain vascular tracing showed an overall ~10% decrease in vascular length and branching density, and light sheet imaging with 3D immunolabeling revealed increased arteriole tortuosity in aged brains. Vasculature and pericyte densities showed significant reductions in the deep cortical layers, hippocampal network, and basal forebrain areas. Moreover, in vivo imaging in awake mice identified delays in neurovascular coupling and disrupted blood oxygenation. Collectively, we uncover regional vulnerabilities of cerebrovascular network and physiological changes that can mediate cognitive decline in normal aging.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Aging drives cerebrovascular network remodeling and functional changes in the mouse brain

Bennett

Zhang

et al. 2023

Preprint

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“…Sampling rate (pixel size) was 1 × 1 μm (xy) and image acquisition occurred at intervals of 50 μm (z). We utilized custom-built algorithms to reconstruct STPT images into 3D volumes 45,82 .…”

Section: Methodsmentioning

confidence: 99%

“…Animals with incomplete perfusion or dissection were excluded from imaging and analysis. At PSUCOM, precise STPT vibratome cutting was achieved by embedding fixed brains in 4% oxidized agarose in custom-built molds, ensuring consistent 3D orientation 82 . Cross-linking was achieved by incubating samples in 0.05 M sodium borohydride solution at 4°C overnight before imaging.…”

Section: Brain Collection Embedding Stpt Imaging and 3d Reconstructionmentioning

confidence: 99%

epDevAtlas: Mapping GABAergic cells and microglia in postnatal mouse brains

Liwang,

Kronman,

Minteer

et al. 2023

Preprint

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During development, brain regions follow encoded growth trajectories. Compared to classical brain growth charts, high-definition growth charts could quantify regional volumetric growth and constituent cell types, improving our understanding of typical and pathological brain development. Here, we create high-resolution 3D atlases of the early postnatal mouse brain, using Allen CCFv3 anatomical labels, at postnatal days (P) 4, 6, 8, 10, 12, and 14, and determine the volumetric growth of different brain regions. We utilize 11 different cell type-specific transgenic animals to validate and refine anatomical labels. Moreover, we reveal region-specific density changes in γ-aminobutyric acid-producing (GABAergic), cortical layer-specific cell types, and microglia as key players in shaping early postnatal brain development. We find contrasting changes in GABAergic neuronal densities between cortical and striatal areas, stabilizing at P12. Moreover, somatostatin-expressing cortical interneurons undergo regionally distinct density reductions, while vasoactive intestinal peptide-expressing interneurons show no significant changes. Remarkably, microglia transition from high density in white matter tracks to gray matter at P10, and show selective density increases in sensory processing areas that correlate with the emergence of individual sensory modalities. Lastly, we create an open-access web-visualization (https://kimlab.io/brain-map/epDevAtlas) for cell-type growth charts and developmental atlases for all postnatal time points.

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“…Our mouse brain preparation is based on an existing protocol for whole mouse brain imaging using serial 2-photon tomography. 2 The protocol will be briefly summarized. For the full details, refer to the original publication.…”

Section: Sample Preparationmentioning

confidence: 99%

Multiorientation mapping of white matter fiber microstructures in whole mouse brains using serial optical coherence tomography

Lefebvre,

Pragassam,

Reynaud

et al. 2024

Neural Imaging and Sensing 2024

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Previous studies have shown that the optical coherence tomography (OCT) signal in white matter (WM) is affected by the WM fiber bundles orientation with respect to the microscope's optical axis. In this paper, we aim to exploit this contrast mechanism to generate a multi-orientation representation of WM microstructure in whole mouse brains. To achieve this, a serial blockface histology set-up has been developed combined with spectral domain OCT equipped with a long-range 10x magnification objective, achieving a near isotropic resolution of 3 micron laterally (xy) and 3.5 micron axially (z). With this imaging system, a map of WM structures can be generated for an entire agarose embedded mouse brain. To precisely control the mouse brain orientation within the agarose, we designed a multi-part 3D printed mold, which allows us to choose the vibratome's slicing plane (e.g., coronal, axial, sagittal, etc.). After the serial OCT acquisition, every slice is reconstructed as 2D images and stacked to obtain a 2.5D volume. The reconstruction process uses a nextflow computational pipeline, allowing us to parallelize the calculations. Our proposed imaging method emphasizes different WM structures according to their orientation, which we illustrated in the mouse's anterior commissure olfactory limb. This structure is very bright when observed in axial slices, whereas it has a darker appearance in the coronal slices. Using this method, we plan to acquire whole mouse brains oriented in multiple directions and to create a multi-orientation mouse brain template, which we believe will prove useful to get a better understanding of complex WM microstructure geometries, such as fiber crossing areas.

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Protocol for using serial two-photon tomography to map cell types and cerebrovasculature at single-cell resolution in the whole adult mouse brain

Cited by 5 publications

References 12 publications

Aging drives cerebrovascular network remodeling and functional changes in the mouse brain

Aging drives cerebrovascular network remodeling and functional changes in the mouse brain

epDevAtlas: Mapping GABAergic cells and microglia in postnatal mouse brains

Multiorientation mapping of white matter fiber microstructures in whole mouse brains using serial optical coherence tomography

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