Cheng et al., 2016 - Activation and inhibition of tph2 serotonergic neurons operate in tandem to influence larval zebrafish preference for light over darkness. Scientific Reports   6:20788 Full text @ Sci. Rep.

Fig. 2

Response of dorsal raphe serotonergic neurons to light and darkness.

(a) Serotonergic neurons (red arrow) in the dorsal raphe of a 5 day-old zebrafish, as labelled by Kaede (coloured yellow) in a Tg(tph2:GAL4, UAS:Kaede) line. Axons extend broadly in the brain, which is pan-neurally labelled (blue). (b) Activity in the raphe of a 9 day-old Tg(tph2:GCaMP6s) fish, in response to four pulses of light. Pixels have the same color scheme as the activity profiles shown in panel c (see Methods). (c) Time series of three clusters obtained by running k-means clustering following principal component analysis (PCA) with two components on the data in panel (b). Blue light was delivered during the periods indicated by the blue bars. (d) Average percentage of tph2 neurons in the raphe excited (ON neurons) or inhibited (OFF neurons) by light (n = 11 fish; squares are individual fish, circles indicate mean and error bar indicates 95% c.i. around the mean). Anterior is to the left in panels (a,b) Pa: pallium; scale bar = 25 μm.

Fig. 4

Effect of optogenetic manipulation of tph2 neurons on preference.

(a) Dorsal view of a Tg(tph2:GAL4, UAS:ChR2(H124R)-eYFP) larva, with anterior to the left. Channelrhodopsin is expressed in the dorsal raphe (red arrow). The image is a maximal projection of optical sections covering 122 μm. (b,c) The percentage of time spent in the light side of the tank, with either blue (b) or white (c) light. All control sibling fish preferred the side with blue or white light. However, fish expressing ChR2 (H134R)-eYFP under the tph2 driver had a significant preference for the dark side when blue light was used. (d) The number of crossovers between the blue light and the dark side as a function of their percentage of time spent on the blue light side. (e) Preference of eNpHR3.0-expressing fish and their siblings for red over green (n = 12 each). There is a significant change towards red in fish expressing halorhodopsin (p = 0.0007). Fb: forebrain; OT: optic tectum; scale bar = 25 μm. Gamma = 0.65 was applied to panel (a). In panels c and e, each square represents one fish (n = 11 fish in each group). Circles are the median and error bar indicates 95% c.i. around the median. p-value is obtained from Wilcoxon Rank-sum test. Z is the test statistic; r is the non-parametric effect size.

Fig. S1

Response of pretectal tph2 neurons to light and darkness.

(a) Dorsal view of the pre-tectal area of a 11 dpf Tg(tph2:GAL4, UAS:GCaMP6s) fish, with anterior at the top. The purple colour indicates cells that are excited by light (ON cells), whereas the blue colour indicates cells that are excited by darkness (OFF cells). (b) Response of pretectal tph2 neurons to light and darkness, determined using k-means clustering of a time-series recorded at 1 Hz. The series was analysed using the Thunder platform. Scale bar = 25 μm.

Acknowledgments:
ZFIN wishes to thank the journal Scientific Reports for permission to reproduce figures from this article. Please note that this material may be protected by copyright. Full text @ Sci. Rep.