Extended Data Fig. 2: Intersectional genetic strategy targets gene expression to gut EC cells.
From: Gut enterochromaffin cells drive visceral pain and anxiety
a, Histologic sections of ECPFTox (Pet1Flp;Tac1Cre;RC::PFTox) colon demonstrating colocalization of PFTox allele GFP reporter (green) and 5-HT (magenta); representative of 93 fields from at least 2 animals Scale bars = 50 μm (upper) and 10 μm (lower). Colon EC cell density is unchanged by expression of PFTox allele (n = 6, 6 independent fields). b, Histologic sections of EChM3Dq (Pet1Flp;Tac1Cre;RC::FL-hM3Dq) colon and duodenum demonstrating colocalization (white arrowheads) of the hM3Dq allele reporter (red) and 5-HT (magenta); representative of 129 fields from at least 2 animals. Scale bars=50 μm (upper) and 10 μm (lower). Colon EC cell density is unchanged following three weeks of DREADD agonist treatment (n = 6, 6 independent fields). c, Whole-mount small intestine (jejunum) of EChM3Dq demonstrating expression of single-(Pet1Flp-GFP, green) and double-recombination (Pet1Flp::Tac1Cre, mCherry, red) hM3Dq allele reporters and 5-HT (magenta); representative of 10 fields from at least 2 animals. Scale bar 50 μm. d, Whole-mount DRG from spinal segments L6/S1 of EChM3Dq demonstrating the absence of mCherry (red) expression; representative of 8 fields from 1 animal. Scale bars = 50 μm. e, Images of EChM3Dq dorsal raphe (DR) and median raphe (MnR) nuclei showing 5-HT-expressing neurons (yellow) and lack of mCherry (red) expression; representative of 6-8 fields from each of 3 animals. Scale bar = 100 μm. f, Images of EChM3Dq lumbosacral spinal cord (L6-S1) showing the absence of mCherry (red) and 5-HT (yellow) expression; representative of 10 fields from each of 3 animals. Scale bar = 100 µm. g, Quantitation of specificity and penetrance of intersectional genetic approach demonstrating ≥ 95% (Tac1Cre, light grey) and 80% (Vil1Cre, dark grey) EC cell specificity and ~60% penetrance in the colon from male mice (data collected from 20–30 random fields). Student’s t-test (unpaired, 2-tailed) in panels a, b; ns = not significant, error bars represent mean ± SEM.
