Developmental expression of ldlra, ldlrb and pcsk9 and cerebral expression during adulthood. (a?c) Transcript quantification of ldlra, ldlrb and pcsk9 during zebrafish development between zygote stage (one cell) to 120 h post-fertilization (120 hpf or larval day 5). Note that these data were obtained from the reanalysis of an RNA seq dataset performed by (White et al., 2017). (d) Transcript quantification of ldlra, ldlrb and pcsk9 in the telencephalon of adult zebrafish (male + female). Note that these data were obtained from the reanalysis of an RNA seq dataset performed by (Gourain et al., 2021; Rodriguez Viales et al., 2015). RPKM, reads per kilobase million.

ldlra, ldlrb and pcsk9 in situ hybridization in the brain of adult zebrafish. (a?f) The schemes (left columns) provide the localization of the transversal brain sections with the name of the different brain nuclei and/or domains. (a?c) ldlra, ldlrb and pcsk9 in situ hybridization in the telencephalon (a,b), diencephalon with the anterior (b; PPa) and posterior (c; PPp) parts of the preoptic area. (d?f) ldlra, ldlrb and pcsk9 in situ hybridization in transversal brain section through the anterior part of the hypothalamus (D, Hv), the mediobasal hypothalamus (e; Hv LR) and the caudal hypothalamus (f; at the level of LR PR). Note that staining for each gene was detected in the anterior part of the hypothalamus, the periventricular nucleus of the posterior tuberculum (TPp), the central posterior thalamic nucleus (CP), the torus semi-circularis (TS) and midbrain parenchyma. The general overall pattern of expression for ldlra, ldlrb and pcsk9 was almost similar between the different genes and was almost ubiquitous within the brain. Bar: 400 ?m (c?f), 100 ?m (b) and 70 ?m (a).

ldlra, ldlrb and pcsk9 are expressed in AroB-positive neural stem cells in the telencephalon. (a?l) Fluorescence in situ hybridization (red) for ldlra, ldlrb and pcsk9 (a,e,i), followed by AroB immunodetection (b,f,j) and DAPI counterstaining (blue). The merged pictures demonstrated the general expression of ldlra, ldlrb and pcsk9 in AroB-positive neural stem cells without (c,g,k) or with DAPI staining (d,h,l) (see arrows). Bar: 15 ?m.

ldlra, ldlrb and pcsk9 are expressed in BLBP-positive neural stem cells in the telencephalon. (a?l) Fluorescence in situ hybridization (red) for ldlra, ldlrb and pcsk9 (b,f,j), followed by BLBP immunodetection (green) (a,e,i). The images shown in (c), (g) and (k) are the respective merged images of ab, ef and ij. (d,h,l) High magnifications of the corresponding white squares. The merged images showed the general expression of ldlra, ldlrb and pcsk9 in BLBP-positive neural stem cells (see arrows). Bar: 70 ?m (a,b,c,e,f,g,i,j,k), 20 ?m (d,h,l) and 7 ?m for high magnifications (right panels).

ldlra, ldlrb and pcsk9 are mainly expressed by neurons in the telencephalon. (a?l) Fluorescence in situ hybridization for ldlra, ldlrb and pcsk9 (b,f,j), followed by HuC/D immunodetection (c,g,k) with DAPI counterstaining (a,e,i). The merged pictures demonstrated the general expression of ldlra, ldlrb and pcsk9 in HuC/D-positive neurons (d,h,l; see arrows). Bar: 10 ?m.

ldlra, ldlrb and pcsk9 gene expression in endothelial cells. (a?l) Fluorescence in situ hybridization (red) for ldlra, ldlrb and pcsk9 (B, F and J) in the telencephalon of adult zebrafish with DAPI counterstaining (blue, a,e,i). (c,g,k,d,h,l) Merged images with high magnifications of the corresponding white squares. The arrows indicate the detection of transcripts in some cells exhibiting an elongated and flat nucleus localized along the blood vessels, corresponding to endothelial cells. Note that a weak expression is shown in endothelial cells for ldlra and pcsk9, whereas ldlrb appears qualitatively more expressed in these cells. Asterisks show DAPI nuclei without ISH staining. Bar: 21 ?m (a,b,c,e,f,g,i,j,k) and 7 ?m (d,h,l).

LDL are taken up by cerebral endothelial cells. (a?h?) intraperitoneal injection of Dil-labelled LDL in Tg(fli1:EGFP) fish (green) allowing to show the vascular distribution of LDLs (in red) in the telencephalon. White squares in d and h identify the high magnifications provided in D?, D?, H? and H?. The arrows indicate the colocalization of GFP (green) and stained LDL (red), demonstrating that a subset of plasma LDL are taken up by endothelial cells (yellow colour). Bar: 40 ?m (a?d), 16 ?m (e?h). 10 ?m (D?, D?, H? and H?).

LDL are not taken up by neural stem cells in homeostatic and regenerative conditions. (a?h) Intracerebroventricular injection of Dil-labelled LDLs in Tg (GFAP::GFP) fish allowing to show that LDLs (in red) are not widely taken up by neural stem cells in homeostatic conditions. Arrows show little LDL detection in GFP positive neural stem cells. (i?p) Intraventricular injection of fluorescent LDLs followed by stab wound injury allows the diffusion of LDL particles within the brain parenchyma but not their increased uptake by neural stem cells. In this context, LDLs were intraperitoneally injected 30 min prior the stab wound and euthanized 1 h30 after this injury. (m?p) Magnification of above pictures (i?k). Bar: 200 ?m (i?l), 100 ?m (m?p), 50 ?m (a?d) and 20 ?m (e?h).

Downregulation of ldlr and pcsk9 gene expression after telencephalic injury. Quantification of ldlra, ldlrb and pcsk9 transcripts in the contralateral hemisphere (CTRL) and the stab wounded hemisphere (SW) at 5 days post lesion (dpl). Note that these data were obtained from a re-analysis of an RNA seq dataset performed by Gourain et al. (2021) and Rodriguez Viales et al. (2015). FPKM, fragments per kilobase million; RPKM, reads per kilobase million.