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Fig. S1 Serotonin (5-HT) promotes motor neuron development.
A,C: Lateral views of the trunk region (left column) indicate that spinal-intrinsic serotonergic cells (arrows) are present at 48, but not 33 hpf. (arrowheads indicate serotonergic cells in the periphery).
B: A dorsal view of the brain indicates only a few serotonergic cells in the region of the preoptic nucleus (Po) of the diencephalon.
D: In contrast, large populations of serotonin-labeled neurons are present in dorsal views in the hypothalamus (Hyp) and raphe region at 48 hpf.
E-G: Lateral views of the trunk indicate that treatment with pCPA from 24 hpf effectively inhibits serotonin (5-HT) accumulation in spinal neurons (E,F), but does not influence the presence of tyrosine hydroxylase (TH) expressing descending axons (arrows) in the spinal cord (G).
H: pCPA treatment does not affect numbers of motor neurons or vsx1:GFP+ interneurons at 33 hpf. I: Endogenous olig2 mRNA, as well as DsRed mRNA are enriched in FAC sorted progenitor cells, compared to the motor neuron fraction, as shown by RT-PCR in olig2:DsRed and HB9:GFP double transgenic fish. GAPDH is used for comparison.
J-M: A 5-HT1A agonist (8OH-DPAT) increases and an antagonist (WAY100645) decreases numbers of hb9:GFP+ motor neurons (ANOVA, P<0.0001). Lateral views of embryos at trunk level are shown; drug incubation form 24 to 33 hpf.
N-R: Similarly, 8-OH-DPAT promotes (Student t-test, P=0.0002) and WAY100645 inhibits (Mann-Whitney U-test, one-tailed, P=0.0383) development of islet- 1:GFP+ motor neurons.
S: Serotonin re-uptake inhibitor fluvoxamine promotes development of islet-1:GFP+ motor neurons, similar to serotonin (ANOVA with Dunn’s Multiple Comparison Test, p < 0.05).
T: Serotonin rescues the effect of blocking Drd4 receptors with L-745870 on development of islet-1:GFP+ motor neurons (Mann-Whitney U-test, one-tailed, p=0.0383), indicating that serotonin does not act through the Drd4a receptor.
U: Dopamine agonist pergolide (ANOVA with Bonferroni’s Multiple Comparison Test, p < 0.001) and serotonin (p < 0.05) increase numbers of HB9+ motor neurons, but have no additive effect. Scale bars = 50 µm.