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Fig. 4
th2+ Cells Are Replaced after Ablation and Mediate Functional Recovery of Swimming Behavior
(A–D) Representative brains from th2:Gal4-VP16; UAS:NTR-mCherry larvae that were treated with 0.5% DMSO or 5 mM MTZ from 5 to 7 dpf, followed by 10 mM BrdU from 8 to 10 dpf and allowed to recover for 2 weeks (10–25 dpf) before acute treatments with 0.5% DMSO or 5 mM MTZ.
(E) Recovery of th2+ cells measured by total cell number. Analysis of the four groups by two-way ANOVA indicates that the 25 dpf treatment has a significant effect on the number of th2:mCherry cells at 28 dpf (p = 0.0001), whereas the 7 dpf treatment does not (p = 0.082); nor is there a significant interaction between two successive treatments (p = 0.20). Adjusted p values shown for pairwise comparisons are based on Bonferroni multiple comparison test. n = 3 brains for larvae untreated at 7 dpf; n = 4 brains for larvae treated with MTZ at 7 dpf.
(F) Percent labeled by BrdU at 8 dpf. Error bars represent SEM; n = 5 brains. p value based on Student’s t test.
(G and H) Recovery of behavior 3 weeks after ablation as measured by swimming frequency (G) and time spent swimming (H); n = 5 groups for each condition except n = 4 for wild-type larvae treated with MTZ plus DMSO after exclusion of an outlier (p < 0.001; Grubbs’ test). Statistical analyses were performed separately for the wild-type and NTR+ larvae. Two-way ANOVA of wild-type larvae shows no significant effect of either the 7 dpf treatment (p = 0.95 for swim frequency; p = 0.89 for swim time) or the 25 dpf treatment (p = 0.13 for swim frequency; p = 0.37 for swim time) at 28 dpf, and there is no significant interaction between the successive treatments (p = 0.66 for swim frequency; p = 0.76 for swim time). For NTR+ larvae, two-way ANOVA indicates that the 7 dpf treatment does not affect swimming behavior at 28 dpf (p = 0.60 for swim frequency; p = 0.80 for swim time), whereas the 25 dpf treatment has a significant effect (p = 0.034 for swim frequency; p = 0.032 for swim time). There was no significant interaction between the successive treatments (p = 0.55 for swim frequency; p = 0.64 for swim time). Adjusted p values shown for pairwise comparisons are based on Bonferroni multiple comparison test.
Images in (A)–(D) are ventral maximum intensity confocal z projections of the hypothalamic posterior recess. The scale bar represents 10 µM. See also Figure S2 for representative plots of swimming behavior in ablated and unablated animals.