Fig. 3: CAS treatment generates and maintains internal synchronized state and vigilance behavior through persistent and intense activation of DRN 5-HT neurons in zebrafish.
From: Serotonergic modulation of vigilance states in zebrafish and mice
a Configuration for performing two-photon calcium imaging on DRN 5-HT neurons in the tph2:GCaMP6f. b Typical images illustrating calcium change after CAS treatment. c Averaged calcium changes of 40 neurons in response to CAS treatment in one fish. d Statistical quantification of calcium activity in 179 neurons from five fish. e, f Configuration of extracellular recordings from DRN 5-HT neurons using tph2:GFP. g, h Typical extracellular recording from one neuron (g) and statistic of firing frequency of 20 neurons from 20 fish (h) before and after CAS treatment. Center of boxplots is the median, upper and lower limits of boxes are the first and third quartile respectively and the upper and lower limits are the maxima and minima. i Illustration of photo-conversion of DRN 5-HT neurons in tph2:Kaede. j, k Fluorescent images showing converted Kaede protein from 5-HT projections in Dc region transported to DRN 5-HT soma (enlarged white box) after 6 h. l Mean data of conversion percentage. m LC–MS workflow for detection of 5-HT concentration in dorsal pallium after CAS treatment. n Mass spectrometry chromatograms showing retention time of standard serotonin (blue) and elution samples at 5 min after CAS treatment (pink). o Mean concentration of 5-HT in dorsal pallium at 5 min, 1 h after CAS treatment, and in control group. p–r Two-photon calcium images and the analyzed data showing CAS-induced neuronal synchronization in Dc region in tph2+/+ HuC:H2B-GCaMP6f (p). The neuronal synchronization disappeared in tph2−/− HuC:H2B-GCaMP6f after CAS treatment (q), which is restored by exogenous application of 5-HT (r). s Illustration of the correlation coefficient for the data presented in (p–r). t Quantification of locomotor speed of tph2+/+ and tph2−/− fish in response to CAS treatment in 30-min recording. Images in b, j-k represents results from six independent experiments. All data are presented as mean ± SEM. **P < 0.01, ***P < 0.001; ns denotes no significant difference. For detailed statistics, see Supplementary Table 2. Source data are provided as a Source Data file.
