Fig. 2

TRPA1 and IL1R1 co-localize on nodose ganglia sensory neurons, and TRPA1 is required for IL-1β mediated calcium signaling. a Representative in situ image of proximity ligation assay showing close proximity of TRPA1 and IL1R1 on nodose ganglion neurons (Scale bar = 50 µm). Nodose sensory neurons were isolated from wildtype and TRPA1 KO mice and subjected to in situ proximity ligation assay (“Methods”), using primary antibody pairs of anti-TRPA1 and anti-IL1R1 antibodies. The cells were counterstained with DAPI (blue) to visualize the nucleus. b Quantification of nodose ganglion neurons showing TRPA1 and IL1R1 co-localization. c–f IL-1β-induces sensory neuron activation in a TRPA1-dependent manner. Nodose ganglion sensory neurons harvested from c wildtype (B6.129PF) or e TRPA1 KO mice were stimulated in culture with IL-1β (20 µg/mL), AITC (100 µM; TRPA1-agonist) and capsaicin (10 µM; TRPV1-agonist) indicated by the labeled black line segments. c, d Sensory neurons from wildtype (B6.129PF) mice respond to IL-1β. c Representative examples of calcium traces of wildtype (B6.129PF) sensory neurons responding to IL-1β, AITC, and capsaicin (blue line); to IL-1β and capsaicin (red line); to AITC and capsaicin (green line) and only capsaicin (black line) are shown. d Percentage breakdown of wildtype calcium responses. Neurons responding to IL-1β, AITC, and capsaicin (blue); to IL-1β and capsaicin (red); AITC and capsaicin (green); and only capsaicin (yellow), are shown. These data were obtained from 17 coverslips cultured using nodose neurons harvested from 16 to 20 mice. e, f Sensory neurons from TRPA1 KO mice fail to respond to IL-1β. e Representative calcium traces of sensory neurons isolated from TRPA1 KO in response to IL-1β, AITC and capsaicin. f Percentage breakdown of TRPA1 KO calcium responses. Neurons responding to IL-1β, AITC, and capsaicin (blue); to IL-1β and capsaicin (red); AITC and capsaicin (green); and only capsaicin (yellow)