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. Author manuscript; available in PMC: 2022 Feb 10.
Published in final edited form as: Cell Host Microbe. 2020 Dec 21;29(2):179–196.e9. doi: 10.1016/j.chom.2020.11.011

Figure 6. EEC Trpa1 signaling activates vagal sensory ganglia.

Figure 6.

(A) Working model. (B) Confocal image of zebrafish vagal sensory ganglia labelled with Tg(neurod1:EGFP) (green) and acetylated αTubulin antibody staining (AC-αTub, magenta). (C) Lightsheet projection of zebrafish stained for AC-αTub. Yellow arrow indicates vagal nerve innervation to the intestine. (D) neurod1:EGFP+ EECs (green) directly contact vagal sensory nerve fibers labelled with αTubulin (white). (E) Confocal image of the vagal sensory nucleus in zebrafish larvae hindbrain where vagal sensory neurons project. Vagal sensory nerve fibers are labeled with different fluorophores through Cre-brainbow recombination in Tg(neurod1:cre); Tg(βact2:Brainbow) zebrafish. The 3D zebrafish brain image was generated using mapzebrain (Kunst et al., 2019). (F) Confocal image of vagal sensory ganglia in Tg(neurod1:cre); Tg(βact2:Brainbow) zebrafish. Asterisk indicates posterior lateral line afferent nerve fibers. Blue arrowheads indicate three branches from vagal sensory ganglia that project to the hindbrain. (G) Confocal image demonstrates the EEC-vagal network in zebrafish intestine. EECs are labeled as magenta by neurod1:TagRFP and the vagal nerve is labeled green by isl1:EGFP. (H) EECs (neurod1+; magenta) directly contact vagal nerve fibers (isl1+; green) as indicated by yellow arrows. (I-J) In vivo calcium imaging of vagal sensory ganglia in zebrafish gavaged with PBS (I) or E. tarda (J). (K) Quantification of individual vagal sensory neuron Gcamp6f fluorescence intensity in E. tarda or PBS gavaged zebrafish. (L-N) Confocal image of vagal ganglia (neurod1+; green) stained with p-ERK antibody (activated vagal sensory neurons; magenta) in WT or EEC ablated zebrafish gavaged with PBS or Trpa1 agonist AITC. (O-Q) Confocal projection of vagal ganglia stained with p-ERK antibody in WT or EEC ablated zebrafish gavaged with PBS or E. tarda. (R) Quantification of p-ERK+ vagal sensory neurons in WT or EEC ablated zebrafish following PBS or AITC gavage. (S) Quantification of p-ERK+ vagal sensory neurons in WT or EEC ablated zebrafish following PBS or E. tarda gavage. (T) Quantification of p-ERK+ vagal sensory neurons in WT or trpa1b−/− zebrafish following E. tarda gavage. One-way ANOVA with Tukey’s post test was used in R and S and Student’s t-test was used in T. *p<0.05; **p<0.01; ***p<0.001; ****p<0.0001.