A novel perivascular cell population in the zebrafish brain
- Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, United States
- National Human Genome Research Institute, National Institutes of Health, United States
- National Institute of Genetics, Japan
- SOKENDAI (The Graduate University for Advanced Studies), Japan
Figures
Figure 1 with 1 supplement
Mrc1a-positive perivascular cells cover the zebrafish brain.
(a) Schematic diagram of an adult zebrafish brain. Dorsal view, rostral at top. Box shows region imaged in panel b. (b) Epifluorescence microscopic image of the optic lobes (top) and cerebellum of …
Figure 1—figure supplement 1
Mrc1a-positive perivascular cells cover the zebrafish brain.
(a,b) Dorsal (a) and ventral (b) confocal images of the dissected brain of a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double-transgenic adult zebrafish (eGFP and mCherry are shown in green and magenta, …
Figure 2 with 1 supplement
Mrc1a-positive perivascular cells are present in the developing zebrafish brain, and express lymphatic markers.
(a,b) Confocal images of Mrc1a:eGFP-positive cells (green) and Kdrl:mCherry-positive blood vessels (magenta) on the surface of the brain in 10 dpf (a) and 15 dpf (b) Tg(mrc1a:eGFP);Tg(kdrl:mCherry) …
Figure 2—figure supplement 1
Mrc1a-positive perivascular cells are EdU negative and fail to collect and drain dye like Mrc1a-positive lymphatics.
(a) EdU staining (magenta) of a 5 dpf Tg(mrc1a:eGFP) transgenic embryo where Mrc1a:eGFP-positive cells (green) are not EdU positive (n = 5 animals). Rostral is to the left. (b) Lymphatic vessel …
Figure 3
Mrc1a-positive perivascular cells are Fluorescent Granular Perithelial cells (FGPs).
(a,b) Yellow/green epifluorescence microscopic images of the optic lobes (top) and a portion of the cerebellum of the dissected brain of a non-transgenic wild type adult zebrafish (n = 3 adult …
Figure 4 with 2 supplements
Zebrafish FGPs do not correspond to other known perivascular cell types.
(a) Schematic diagram of cells associated with brain blood vessels, including the vascular endothelial cells, pericytes/smooth muscle cells, radial glia, macrophages/neutrophils, and FGPs. (b) …
Figure 4—figure supplement 1
Radial Glia and FGPs localize to different parts of the brain.
(a) Dorsal view of a Z-projected confocal image generated by the Zebrafish Brain Browser (Marquart et al., 2015) superimposing four 6 dpf Tg(lyve1:dsRed) (green) embryos to Tg(gfap:GFP) (magenta). …
Figure 4—figure supplement 2
FGPs are not Neural Crest derived.
(a-b) Lateral view of a 3.5 dpf Tg(sox10:dsRed); Tg(mrc1a:eGFP) double transgenic embryo showing mrc1a:eGFP-positive FGPs in green (a) and Sox10:dsRed–positive neural crest derived structures in …
Figure 5 with 3 supplements
Zebrafish FGPs are not derived from definitive hematopoietic progenitors.
(a) Schematic diagram illustrating hematopoietic stem and progenitor cells (HSPCs) differentiation from endothelial cells in the ventral floor of the dorsal aorta (DA). No, Notochord; DA, Dorsal …
Figure 5—figure supplement 1
FGPs are not derived from primitive hematopoiesis.
(a-d) Dorsal yolk (a,b) and lateral trunk (c, d) transmitted light images of 20 hpf control morpholino (a,c) or pu.1 morpholino (b,d) injected embryos subjected to whole mount in situ hybridization …
Figure 5—figure supplement 2
Zebrafish FGPs are not derived from definitive hematopoietic progenitors.
(a) Schematic diagram illustrating hematopoietic stem and progenitor cells (HSPCs) differentiation from endothelial cells in the ventral floor of the dorsal aorta (DA). No, Notochord; DA, Dorsal …
Figure 5—figure supplement 3
Inhibiting HSPCs specification does not affect FGP formation.
(a,a’,b,b’) Lateral views of 5 dpf Tg(lyve1:dsRed);Tg(mpx:eGFP) double transgenic animals showing the lymphatic vessels and FGPs (magenta) and macrophages (green punctate) in an uninjected (a) or run…
Figure 6 with 1 supplement
Global analysis of gene expression in FACS-sorted adult zebrafish FGPs.
(a) Confocal microscopic image of the optic lobes (top) and cerebellum of the dissected brain of a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double-transgenic adult zebrafish (EGFP and mCherry are shown in …
Figure 6—figure supplement 1
Analysis of gene expression in FACS-sorted adult zebrafish FGPs compared to kdrl:mCherry-positive blood endothelial cells.
(a) Plots showing comparative expression levels of genes in FGPs (GFP positive) versus blood endothelial cells (mCherry positive) FACS sorted from Tg(mrc1a:eGFP);Tg(kdrl:mCherry) adult tectal …
Figure 7 with 2 supplements
Zebrafish FGPs emerge from the endothelium of the choroidal vascular plexus.
(a), Dorsal (top) and lateral (bottom) view schematic diagrams of approximately 2.5 dpf zebrafish heads with some of their associated vasculature, especially major venous tracts. CVP, choroidal …
Figure 7—figure supplement 1
Emerging FGPs on the 3 and 4 dpf zebrafish brain.
(a-d) Confocal images of Mrc1a:eGFP-positive FGPs (green) and Kdrl:mCherry-positive blood vessels (magenta) on the surface of the brain in 3 dpf (a,b) and 4 dpf (c,d) Tg(mrc1a:eGFP);Tg(kdrl:mCherry) …
Figure 7—figure supplement 2
Zebrafish FGPs emerge from the endothelium of the choroidal vascular plexus.
(a) Dorsal (top) and lateral (bottom) view schematic diagrams of approximately 2.5 dpf zebrafish heads with some of their associated vasculature, especially major venous tracts. OCVP, choroidal …
Figure 8
Zebrafish FGPs differentiate from the CVP endothelium.
(a) Model diagram showing the location of the CVP imaged during timelapse acquisition in a dorsal (top) and lateral (bottom) orientation.( b–g) Stills from time-lapse movies of cells emerging from …
Figure 9 with 1 supplement
Zebrafish FGPs are derived from kdrl-expressing endothelium.
(a-c) Lateral view confocal micrographs of a 5 dpf Tg(−9.8actb2:LOXP-DsRED-LOXP-EGFP); Tg(kdrl:Cre) double transgenic ‘switch’ embryo, with ‘unswitched’ DsRed-positive cells (magenta, panels a and …
Figure 9—figure supplement 1
Mosaic expression of EGFP in vessel endothelial cells in the Kdrl:Cre ‘switch’ double transgenic line.
(a,b) Lateral view confocal micrograph of the head of a 5 dpf Tg(−9.8actb2:LOXP-DsRED-LOXP-EGFP); Tg(kdrl:Cre) double transgenic ‘switch’ animal, showing DsRed-positive ‘unswitched’ cells (magenta, …
Videos
Video 1
Confocal time-lapse movie of emerging Fluorescent Granular Perithelial cells in a Tg(mrc1a:eGFP);Tg(sox10:dsRed) double transgenic zebrafish embryo from 50 hpf to 70 hpf.
Tg(mrc1a:eGFP) is displayed in green and Tg(sox10:dsRed) is displayed in magenta. Stacks were acquired every 5 min using a 40X water, 1.1NA objective with on a Zeiss LSM880 confocal microscope. …
Video 2
Confocal time-lapse movie (dorsal view) of developing Fluorescent Granular Perithelial cells migrating to the surface of the brain in a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double transgenic zebrafish embryo from 52 hpf to 80 hpf.
Tg(mrc1a:eGFP) displayed in green, Tg(kdrl:mCherry) displayed in gray scale. Frames acquired every 12 min using a 40 × 1.1 NA water objective with a 4-square tile and 10% overlap on a Zeiss LSM880 …
Video 3
Confocal time-lapse movie of developing Fluorescent Granular Perithelial cells around the brain of a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double transgenic zebrafish embryo from 52 hpf to 71hpf.
Tg(mrc1a:eGFP) displayed in green, Tg(kdrl:mCherry) displayed in magenta. The movie begins with a dorsal to ventral 3D rotation and ends with a ventral to dorsal 3D rotation. Stacks were acquired …
Video 4
Confocal time-lapse movie of developing Fluorescent Granular Perithelial cells around the brain of a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double transgenic zebrafish embryo from 52 hpf to 71hpf.
Tg(mrc1a:eGFP) displayed in green, Tg(kdrl:mCherry) displayed in magenta. Stacks were acquired every 5.7 min using a 20X, 1.0NA objective on a Leica SP5II confocal microscope. Rostral to the left. …
Video 5
Confocal time-lapse movie of developing Fluorescent Granular Perithelial cells emerging from the optic choroidal vascular plexus of a Tg(mrc1a:eGFP);Tg(kdrl:mCherry) double transgenic zebrafish embryo from 52 hpf to 71 hpf.
Tg(mrc1a:eGFP) displayed in gray scale, Tg(kdrl:mCherry) displayed in magenta. Nascent FGP’s are pseudocolored in green. The red square illustrates the confocal volume. The blue arrow indicates the …
Video 6
Confocal time-lapse movie of developing Fluorescent Granular Perithelial cells emerging from the optic choroidal vascular plexus of a Tg(mrc1a:eGFP);Tg(kdrl:nlsmCherry) double transgenic zebrafish embryo from 50 hpf to 60 hpf.
Tg(mrc1a:eGFP) displayed in gray scale, Tg(kdrl:nlsmCherry) displayed in magenta, FGP nuclei are pseudo-colored in teal. The red square illustrates the confocal volume. The blue arrow indicates the …
