Extended Data Fig. 2: nTS neurons were activated only after repeated allergen challenges to the lung of B6 mice.

(a,b) Fos RNAscope from brainstem section of naïve mice 1.5 h after HDM challenge to lung showing Fos RNA signals are not found in the AP as defined by Gfral (a) or Glp1r (b). (c) FOS antibody staining from brainstem section of Chat-cre; Ai14 mice 2 h after HDM showing that there is little overlap between FOS⁺ cells and tdTom⁺ DMV and 12 N regions. (d) Quantifications of the number of Fos⁺ neurons in the AP (n = 6 mice), DMV (n = 7 mice) or 12 N (n = 7 mice). (e,h,k) Experiment scheme for the first, second or third HDM challenge in wild-type mice. (f,g,i,j,l,m) FOS antibody staining (f,i,l) and quantification (g,j,m) showing no statistically significant increase of FOS⁺ cells after the first, second or third HDM challenge. n = 6 mice each, unpaired t-test, NS. (n) Experiment scheme for single saline or HDM challenge in naive mice and tissue collection 30 min after the challenge for Fos RNAscope. (o) No significant difference in the number of Fos⁺ neurons in the nTS 30 min following a single saline (n = 3 mice) or HDM (n = 3 mice) challenge, unpaired t-test, NS. (p) Experiment scheme for HDM challenge and tissue collection in wild-type mice for Fos RNAscope. (q) The number of Fos⁺ neurons in the nTS are increased in the HDM-treated (n = 11 mice) group, compared to saline-treated controls (n = 11 mice), unpaired t-test, P = 0.0024. Outlines in a,b,c,f,i and l delineate nTS regions at indicated Bregma levels. Scale bars, 200 µm. Data are mean values ± s.e.m., statistical tests are two-sided.

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