Fig. 3. Basal constriction at the MHBC is laminin-dependent and not dependent on ventricle inflation.

(A–C′) Live laser-confocal imaging of wild type, snk and nok embryos at 24 hpf after injection with memGFP. Boxed regions in A–C are enlarged in panels A′–C′. Cells at the MHBC in (A–A′) wild type, (B–B′) snk and (C–C′) nok undergo basal constriction (see cell outlines). (D–F′) Imaging of sly mutants injected with memGFP mRNA at the one cell stage and imaged at 17, 21, and 24 hpf. Boxed regions in D–F are enlarged for panels D′–F′. (D′) At 17 hpf, MHB and surrounding cells are similar in length (see outlined cells). (E′) At 21 hpf, cells at the MHB are shorter than those surrounding. One cell at and one cell outside the MHB are outlined in yellow. Some cells are visible outside the neural tube. (F′) At 24 hpf, cells at the MHBC fail to basally constrict. For panels A–C and D–F asterisks indicate the cell that is outlined in the image below. Cells with two asterisks are outside MHB. Arrowheads indicate the MHBC. Dotted lines delineate the outside of the neural tube. Some green fluorescence is visible within outlined cells since the plane of section contains the surface of the cell membrane. Anterior is to the left in all images. Scale bars: A–C = 22 μM, A′–C′ = 12 μM, D–F = 18 μM, D′–F′ = 9 μM. (G) Length of cells at the MHB relative to those outside the MHB in WT and sly mutants. At 21 hpf, cells at the MHB (at MHB) in sly mutants are 0.76 (+/− 0.094 s.d.) the length of those outside the MHB (outside MHB), as in WT embryos (WT data is the same as from Figure 2) (n = 6 embryos, 2 cells at MHB, 3 cells outside MHB were measured per embryo). (H) Graph compares the apical width of cells outside MHBC to cells at MHBC in WT, sly, snk, and nok embryos at 24 hpf. Basally constricted cells at the MHBC do not apparently show corresponding apical expansion snk and nok (n = 3 embryos each mutant, 2 cells at MHBC, 3 cells outside MHBC were measured per embryo). Cells at the MHBC in sly mutants that have shortened, but are not constricted basally, are also not expanded apically (n = 6 embryos, 3 cells at MHBC, 4 cells outside MHBC were measured per embryo).