Figure 3.
Ventral but not dorsal RGC axons are retarded after Sema3D ubiquitious misexpression. A-C, Lateral views (anterior is to the left) of 48 hpf embryos labeled with DiI injections into ventral retina. Eye on opposite side of embryo is labeled, and axons (white) are shown extending up the contralateral diencephalon. Ventral RGC axons in a wild-type (WT), heat-treated embryo (A) or an hsp70:sema3Dmyc transgenic, nonheated embryo (C) have extended to a position in the diencephalon dorsal to the contralateral eye. Ventral axons in a transgenic, heat-induced embryo (B) have only extended to a position in the diencephalon at the level of the ventral one-third of the contralateral eye. Arrows indicate position of axon tips. L, Lens of contralateral eye. D-F, Ventral views (anterior is to the top) of chiasm region of 51 hpf embryos labeled with DiI injected into dorsal retina. Dorsal axons extend later than ventral and thus have not advanced as far. There is no difference in length of dorsal RGC axons in controls (wild-type heat-treated in D or transgenic nonheat induced in F) versus transgenic heat-induced embryos (E). Scale bar, 50 μm. G, Bar graphs of mean axon position. Error bars are SEM; values of n are indicated in parentheses. y-Axis is axon position as defined in Table 1. Different populations of embryos are distributed along the x-axis. Ventral inj 37 hr, Ventral retinal injections into wild type (WT) or transgenic (3D) embryos heat induced at 30 hpf and fixed at 37 hpf (hs30-37). ** denotes groups significantly different from others. Groups 1 and 2 are significantly different; t test; p < 0.0008. At these younger ages, heat induction causes a temporary developmental delay. Thus, nonheated transgenic embryos are not included as a control. Ventral inj 48 hr, Ventral retinal injections into wild-type (WT) or transgenic (3D) embryos heat induced at 41 hpf and fixed at 48 hpf. 3D no hs 48, Nonheat induced transgenic embryos fixed at 48 hpf. Groups 1 and 2 are significantly different; t test; p < 0.0178. Groups 2 and 3 are significantly different; t test; p < 0.0057. Dorsal inj 51 hr, Dorsal retinal injections into wild-type (WT) or transgenic (3D) embryos heat induced at 44 hpf and fixed at 51 hpf. 3D no hs 51, Nonheat induced transgenic embryos fixed at 51 hpf. There is no significant difference between any condition. t tests between groups 1 and 2, p < 0.3545; between 2 and 3, p < 0.1880. Dorsal inj 55 hr, Dorsal retinal injections into wild-type (WT) or transgenic (3D) embryos heat induced at 48 hpf and fixed at 55 hpf. 3D no hs 55, Nonheat induced transgenic embryos fixed at 55 hpf. There is no significant difference between any condition. t tests between groups 1 and 2, p < 0.3996; between 2 and 3, p < 0.9473.