Article Figures & Data
Figures
- Figure 1.
A tool for the flexible atlas registration of biological images and brain-wide mapping of a label of interest. When running the analysis pipeline, the user is presented with the opportunity to limit the range of images and to define whether to count labels of interest or apply a densitometry-based approach (A). The user will then be prompted to select which atlas plate most closely aligns with the image they are currently registering (B, thumbnail reference atlas images credited to the Allen Institute and the Allen Mouse Brain Atlas; Lein et al., 2007). The selected atlas plate then loads over the target image and individual regions resize sequentially to suit size of the target image (C). Regions are manually moved and adjusted to align with the target image (D). Once the alignment is correct, the registration is then applied to a binarized label of interest, in this case c-Fos-expressing cells (E). The regional density of c-Fos-expressing cells (F), the number of c-Fos-expressing cells (G), and the area of each region (H) can all be obtained as outputs using this analysis type. For program source code and user guides, see Extended Data 1.
Figure Contributions: Dylan J. Terstege prepared the tissue, collected photomicrographs, and conducted the analysis.
- Figure 2.
DAPI provides sufficient contrast for accurate and reliable neuroanatomical registration. A, Representative photomicrograph of parvalbumin and DAPI staining from a coronal section of adult mouse brain. Overlay shows traced regions for each channel (parvalbumin = cyan, DAPI = magenta) B, The thalamic reticular nucleus was traced across multiple sections based on parvalbumin (cyan) and DAPI (magenta) labeling. C, There were no significant differences in area measurements of thalamic reticular nuclei traced from each imaging channel. D, Areas traced using DAPI labeling as a reference overlapped with areas traced based on parvalbumin labeling by an average of 96.66%, with a range of 6.172% (92.84–99.012%). E, Using a DAPI channel, independent raters (n = 4) used FASTMAP to record the areas of the amygdalar areas (AMY), hippocampal formation (HPF), hypothalamus (HY), isocortex (ISO), midbrain (MB), olfactory cortex (OLF), pallidum (PAL), striatum (STR), and thalamus (TH) from a subset of images (n = 2–5 per area) across an adult mouse brain. The extent to which the individual tracings overlapped with a summed composite trace was calculated to have median values of 93.83% for AMY, 92.85% for HPF, 92.07% for HY, 94.41% for ISO, 95.46% for MB, 91.82% for OLF, 89.41% for PAL, 92.34% for STR, and 93.89% for TH. Areas recorded from these tracings, summed across images for each region (F–N), did not differ from areas collected using the commercial registration tool NeuroInfo. Representative NeuroInfo registrations (colored outlines) and FASTMAP registrations (white outlines) are provided for each region. Representative FASTMAP registrations were selected for visualization based on which of the independent raters produced a summed area measurement (across all regions) closest to the median value among independent raters. The measurement that corresponds to the sample trace is indicated in each plot with an octothorpe (#). Data presented as individual matched datapoints (C), median ± max/min (D, E), and mean ± 95%CI (F–N).
Figure Contributions: Dylan J. Terstege prepared the tissue, collected photomicrographs, and conducted the analysis.
- Figure 3.
Assessing regional density of brain vasculature. Representative photomicrographs of vasculature in the adult mouse brain (A) and the dentate gyrus (B). FITC-perfused vasculature is labeled in cyan, while propidium iodide is displayed in magenta. Vasculature was segmented from background and binarized using Ilastik (C, D). The image was registered to a custom atlas plate consisting of the isocortex (ISO), hippocampus (HPF), amygdala (AMY), olfactory bulbs (OLF), pallidum (PAL), cerebellum (CB), midbrain (MB), striatum (STR), thalamus (TH), hypothalamus (HY), medulla (MY), and hindbrain (HB; E). Using a densitometry-based analysis approach, the regional density of vasculature was determined as a percentage of the overall region area (F).
Figure Contributions: Dylan J. Terstege prepared the tissue, collected photomicrographs, and conducted the analysis.
- Figure 4.
Mapping of age-related changes in amyloid plaque deposition density. Representative photomicrographs of amyloid pathology in 3-month-old (A), 5-month-old (B), and 10-month-old (C) CRND8 mice. Propidium iodide is displayed in magenta while the fluorescently labeled β-amyloid is labeled in cyan. D, Amyloid density across the isocortex (ISO), hippocampus (HPF), amygdala (AMY), olfactory bulbs (OLF), pallidum (PAL), cerebellum (CB), midbrain (MB), striatum (STR), thalamus (TH), hypothalamus (HY), medulla (MY), and hindbrain (HB). High-resolution photomicrographs of the somatosensory cortex (E–G) and the reticular nucleus of the thalamus (H–J) further illustrate differences in amyloid pathology progression at 3 (E, H), 5 (F, I), and 10 (G, J) months. Data presented as mean ± SEM.
Figure Contributions: Dylan J. Terstege prepared the tissue, collected photomicrographs, and conducted the analysis.
- Figure 5.
Compatibility of the registration tool with different tissue types, sample preparations, and imaging planes. Application of the registration tool to coronally sectioned adult mouse tissue (A), sagittally sectioned adult mouse tissue (B), horizontally sectioned adult mouse tissue (C), coronally sectioned tissue blocks of adult mouse tissue (D), and uDISCO-cleared e16 mouse embryo (E). Figure Contributions: Dylan J. Terstege prepared the tissue and collected the photomicrographs. Daniela O. Oboh prepared the custom atlas plates.
Tables
Abbreviation Region Abbreviation Region AMY Amygdala MB Midbrain CB Cerebellum MY Medulla HB Hindbrain OLF Olfactory bulbs HPF Hippocampal
formationPAL Pallidum HY Hypothalamus STR Striatum ISO Isocortex TH Thalamus List of the 12 regions and their associated abbreviations which were included in the custom lower-level neuroanatomical atlases.
Abbreviation Region Abbreviation Region ACA Anterior cingulate area PAG Periaqueductal gray ACB Nucleus accumbens PALc Caudal pallidum AHN Anterior hypothalamic nucleus PALm Medial pallidum AON Anterior olfactory nucleus PALv Ventral pallidum ATN Anterior dorsal thalamus PCG Pontine central gray CA1 Field CA1 PG Pontine gray CA2 Field CA2 PH Posterior hypothalamus CA3 Field CA3 PHY Perihypoglossal nucleus DG Dentate gyrus PMd Dorsal premammilary DMH Dorsomedial hypothalamus PMv Ventral premammilary DMX Dorsal motor nucleus of the vagus nerve PRN Pontine reticular nucleus EPI Epithalamus PRT Pretectal region FN Fastigial nucleus PVH Paraventricular hypothalamus FRP Frontal pole RCH Retrochiasmatic area GRN Gigantocellular reticular nucleus RN Red nucleus IC Inferior colliculus RSP Retrosplenial cortex ILM Intralaminar nucleus RT Reticular nucleus of the thalamus IO Inferior olivary complex SCm Motor superior colliculus IRN Intermediate reticular nucleus SCs Sensory superior colliculus LDT Laterodorsal tegmental nucleus SF Septofimbrial nucleus LS Lateral septum SOC Superior olivary complex MARN Magnocellular reticular nucleus SPF Subparafascicular nucleus MDRN Medullary reticular nucleus SUB Subiculum MED Medial dorsal thalamus TRN Tegmental reticular nucleus MO Somatomotor areas TT Taenia tecta MOB Main olfactory bulb VENT Ventral group of the dorsal thalamus MPN Medial preoptic nucleus VERM Vermal regions MPO Medial preoptic area VI Abducens nucleus MRN Midbrain reticular nucleus VMH Ventromedial hypothalamic nucleus NTS Nucleus of the solitary tract VNC Vestibular nuclei ORB Orbital area VTA Ventral tegmental area OT Olfactory tubercle List of the 63 regions and their associated abbreviations which were included in the custom higher-level neuroanatomical atlases.
Extended Data 1
FASTMAP user guide. Full FASTMAP user guide posted to the GitHub repository, at https://github.com/dterstege/FASTMAP. Download Extended Data 1. Download Extended Data 1, ZIP file.
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