Fig. 2

MHBC formation requires cell shortening and basal constriction. (A–C′) Live laser-scanning confocal imaging of wild type embryos injected with memGFP mRNA at the one cell stage and imaged at 17, 21, and 24 hpf. Boxed regions from (A) to (C) are enlarged for (A′–C′). Individual cells in the MHB are outlined, and a dividing cell is indicated by an arrow in (B). Asterisks in (A–C) mark cells outlined in (A′–C′). Cells with two asterisks are outside the MHB. (A′) At 17 hpf, cells at the MHB are similar in length to the cells in the surrounding tissue. (B′) At 21 hpf, cells at the MHB are shorter than the cells in the surrounding tissue. (C′) At 24 hpf, cells at the MHBC are constricted basally and expanded apically. (A–C′) Some green fluorescence is visible within outlined cells since the plane of section contains the surface of the cell membrane. (D–I) Time-course of MHB morphogenesis beginning at 17 hpf. A single cell is outlined and followed through the time course. Cells at the MHB shorten relative to those surrounding (n = 6 embryos). (J) Relative cell lengths at and outside the MHB in 21 hpf wild type embryos. Cells at the MHB were 0.76 times the length of those outside the MHB (± 0.06 s.d.) (n = 8 embryos, 3 cells at the MHB and 4 cells outside the MHB were measured per embryo). (K) Relative apical width of unwedged cells (those outside the MHBC) and basally constricted cells (at the MHBC) in wild type embryos at 24 hpf. Cells at the MHBC had 1.6 times the apical width of those outside the MHBC (± 0.29 s.d.) (n = 6 embryos, 2 cells at the MHBC and 3 cells outside the MHBC were measured per embryo). (L) Numbers of basally constricted cells at the MHBC in wild type embryos at 24 hpf (n = 9 embryos). Arrowheads indicate the MHBC. M, midbrain. Scale bars: A–C = 20 μM, A′–C′ = 9 μM, D–I = 30 μM.