Journal: Biology Open

Article Title: Lateral line placodes of aquatic vertebrates are evolutionarily conserved in mammals

doi: 10.1242/bio.031815

Figure Lengend Snippet: Neurogenesis in the lateral line placodes of Q-VD-OPh-treated mice. (A,C,F,G) Reconstructions of serially sectioned mouse embryos (section interval evaluated=10 µm) show ectoderm (light grey), otic vesicle with detachment site (dark grey), epibranchial placodes (orange), lateral line placodes (blue), immunopositive cells (brown dots), developing cranial nerves (partly numbered brown contours in G). (B,D,E) Micrographs taken from serially sectioned mouse embryos. (A) Sox2 + cells are present in epibranchial placodes and, to a much lesser extent, in lateral line placodes (embryo #021). (B) Sox2 + mantle (m) and support (s) cells in a neuromast primordium of a middle lateral line placode. Note the apical cavity (arrowhead). (C) Ngn1 + cells are present in epibranchial placodes and, to a much lesser extent, in lateral line placodes (embryo #039, mirror-imaged right side). (D) Ngn1 + neuroblasts in an anterodorsal lateral line placode. (E,F) Ngn2 + neuroblasts are present in epibranchial placodes, but absent from lateral line placodes (embryo #001). (G) Tubb3 + neurons constitute the anlagen of the cranial nerves 7 and 8 (VII/VIII), 9 (IX), 10 (X), 11 (XI) and 12 (XII). Tubb3 + candidates for vestigial lateralis ganglia (asterisks) reside in close proximity to the middle lateral line placode (embryo #014). All micrographs were adjusted for brightness (including slight gamma changes), colour balance, and sharpness. Scale bars: 5 µm. ad, m, p, anterodorsal, middle, and posterior lateral line placode, respectively; e1, e2, e3, epibranchial placodes 1, 2, 3, respectively; ot, otic anlage; ov, optic vesicle; Q-VD-OPh, pan-caspase inhibitor; wec, whole embryo culture.

Article Snippet: Primary antibodies included: mouse anti-Atonal homolog 1 (Atoh1; Atoh1 supernatant, Developmental Studies Hybridoma Bank, Iowa City, USA, lot 6/20/13, RRID: AB_10805299; 1:50, overnight, 4°C) ( ; ), mouse anti-β-Tubulin-III (Tubb3; clone SDL.3D10, T8660, Sigma-Aldrich, lot 073K4835, RRID: AB_477590; 1:8000, overnight, 4°C) , rabbit anti-cleaved caspase-3 (9661; Cell Signaling Technology, lot 37, RRID: AB_2341188; 1:8000, overnight, 4°C) ( , ), mouse anti-Islet 1 (Isl1; 39.4D5 ascites fluid, Developmental Studies Hybridoma Bank, lot 2/12/09, RRID: AB_2314683; 1:2000, overnight, 4°C) , goat anti-Neurogenin1 (Ngn1; sc-19231, Santa Cruz Biotechnology, lot C1215, RRID: AB_2298242; 1:100, 4 h, 37°C) , mouse anti-Neurogenin2 (Ngn2; clone 7G4, MAB3314, R&D Systems, Minneapolis, USA, lot WWI01, RRID: AB_2149520; 1:20,000, overnight, 4°C) , goat anti-Neurogenic differentiation 1 (NeuroD1; sc-1084, Santa Cruz Biotechnology, lot A0616, RRID: AB_630922; 1:800, 2 h, 37°C) , rabbit anti-Paired homeobox (Pax) 2 (71-6000; Thermo Fisher Scientific, lot 1117672A, RRID: AB_2533990; 1:4000, overnight, 4°C) , mouse anti-Pax8 (ACI 438; Biocare Medical, Pacheco, USA, lot 051712, RRID: AB_10922964; 1:100, overnight, 4°C) , rabbit anti-phospho-Histone H3 (pHH3; 9701, Cell Signaling Technology, lot 13, RRID: AB_331535; 1:1000, 2 h, 37°C) , rabbit anti-Sine oculis homeobox 1 (Six1; HPA001893, Atlas Antibodies, Stockholm, Sweden, lot A00939, RRID: AB_1079991; 1:800, overnight, 4°C) , mouse anti-Six1 (clone CLO185, AMAb90544, Atlas Antibodies, lot 02852, RRID: AB_2665581; 1:100, overnight, 4°C) that, compared to rabbit anti-Six1, was raised against the same immunogen and shows identical reactivities (present study), mouse anti-Sex determining region Y-box (Sox) 2 (clone 245610, MAB2018, R&D Systems, lots KGQ0311111, KGQ0315062, RRID: AB_358009; 1:200, overnight, 4°C) , goat anti-Sox10 (sc-17342, Santa Cruz Biotechnology, lot L2908, RRID: AB_2195374; 1:800, overnight, 4°C) , mouse anti-Sox10 (sc-365692, Santa Cruz Biotechnology, lot I0516, RRID: AB_10844002; 1:1600, overnight, 4°C) that, compared to goat anti-Sox10, produced identical staining patterns in mice (present study) and rats , and rabbit anti-T-box transcription factor (Tbx) 3 (42-4800, Thermo Fisher Scientific, lot SI256120, RRID: AB_2533526; 1:200, 2 h, 37°C) ( ).

Techniques: Embryo Culture