Figure 4

Effects of exposure to embryonic ethanol (0.5% v/v, 22–24 hpf) and injection of cxcl12a MO^sp (850 ng/µL) on the number of Hcrt neurons in the aAH, POA and pAH subpopulations of 6 dpf transgenic Hcrt:EGFP zebrafish brains. (A) Bar graphs (n = 4–6/group) show the number of Hcrt neurons in the aAH, POA and pAH. (Two-way ANOVA, Condition main effect: (F (3, 45) = 7.765, p = 0.0003), Brain area main effect: (F (2, 45) = 209.4, p < 0.0001), Condition ×Brain area interaction: (F (6, 45) = 4.381, p = 0.014), followed by Holm–Sidak post-hoc analysis). (B) Photomicrographs (25×, dorsal view) illustrate Hcrt neurons (green) that are located in the aAH and pAH of Control + Control MO (left), Ethanol + Control MO (middle) and Ethanol + cxcl12a MO^sp (right). Scale bar: 10 µm. All results are shown as means ± standard errors. ** p < 0.01, *** p < 0.001. Abbreviations: MO: morpholino, aAH: anterior part of the anterior hypothalamus, POA: preoptic area, pAH: posterior part of the anterior hypothalamus, Hcrt: hypocretin, hpf: hours post fertilization, dpf: days post fertilization.