Fig. 2

Biodistribution of PAP3 liposomes within the head region of zebrafish embryos. a) Schematic zebrafish larvae with key blood vessels in different planes of view. See Figure S1a,b, Supporting Information for a Z-stacks depth color-coded zebrafish vasculature. Dorsal view, showing the i) top, ii) middle, and iii) bottom layer diagram. The vasculature is as follows, targeted brain endothelial cells (bECs) in yellow and non-targeted systemic endothelium in dark gray, at ≈78 h post-fertilization (hpf). PHBC, primordial hindbrain channel; CtA, central artery; DLV, dorsal longitudinal vein; MsA, mesencephalic artery; MsV, mesencephalic vein; MMCtA, middle mesencephalic central artery. b) Biodistribution (10x magnification, dorsal view, 1.5 hpi) of PAP3 liposomes (5 mM, 0.2% mol DOPE-LR) within the head region of a Tg(kdrl:GFP) zebrafish embryo at 54, c) 78, and d) 102 hpf. e) High magnification view of c) (inset, 40x, zoom) showing PAP3 liposomes accumulating within bECs. f) Posterior part of the PHBC, g) DLV, h) CtA, i) MMCtA. j,k) Biodistribution of PAP3 liposomes in a Tg(cldn5a:GFP)⁴² zebrafish embryo (102 hpf) with GFP-labeling in blood and choroid plexus brain barrier showing colocalization of cldn5a with fluorescent liposomes. Zoom-in of the region that includes the hindbrain vasculature and colocalization with cldn5a expression is shown in Figure S1c, Supporting Information. Liposomes formulated by extrusion (5 mM, 0.2 mol% DOPE-LR). Scale bars: 100 µm (dorsal view), 50 µm (tissue level).