Miconazole suppresses brain hemorrhages in fn40a−/− mutants. (See also Figs S2-S4 and Table S1). (A-D′) O-dianisidine staining for erythrocytes showing hemorrhages in the dorsal part of the brain (C′) that were suppressed by miconazole (mic) treatment (D′) in homozygous fn40a mutants whereas no brain hemorrhages occur in heterozygous fn40a mutants in the absence or presence of miconazole (mic) (A-B′) at 2 dpf. A-D, ventral view; A′-D′, dorsal view. The numbers on the lower right show the number of hemorrhagic (C,C′) and non-hemorrhagic (A-B′,D,D′) embryos out of the total embryos scored. (E-L) CtA between BA and PHBC labeled in Tg(flk1:eGFP) zebrafish are disrupted (G,K, arrows) in homozygous mutants. This is rescued by mic treatment (H,L) in homozygous mutants compared with heterozygous mutants with either DMSO or mic treatment (E-F,I-J). Images are representative of more than 10 embryos per group (n>10). E-H, 2 dpf; I-L, 3 dpf. Scale bars: 50 µm. (M-T) Transmission electron microscopy shows the invasion of blood cells (red arrows) and disrupted neurons (O) and the leakage of high-density blood plasma (red asterisks) into the surrounding brain tissue (S) in fn40a homozygous mutants. This is rescued by mic treatment (P,T) compared with heterozygous mutants with or without mic treatment (M-N, Q-R) at 2.5 dpf. Images are representative of more than 8 embryos per group (n>8). In Q-T, dashed lines mark blood vessel border; red arrows indicate invaded blood cells; yellow arrows indicate vacuoles after cell death. BA, basilar artery; CtA, central artery; PHBC, primordial hindbrain channel; RB, red blood cell; Ne, neuron. Scale bars: 5 μm.
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