Extended Data Fig. 8: Silencing nociceptors and targeting NP3 cells as controls for strategies used to functionally determine nociceptor contribution to allodynia.

(a) Triple label ISH validating our approach for silencing a large subset of nociceptors (Trpv1 expressing PEP, NP3 and NP2A neurons). In DRGs, >96% of Trpv1 expressing neurons were TeNT positive (624/643 cells in sections from 3 mice) and >97% of TeNT positive cells were nociceptors i.e., co-expressed Trpv1 and/or Scn10a. (b) Sst-Cre was used to target NP3 cells; Cre-recombination (green) occurred in 63% of NP3 neurons expressing Sst/Nppb (red, 246/389) with variable (50-80%) recombination across 3 animals. Moreover, NP3 cells accounted for only 29% of the recombined neurons (246/847). Recombination also labeled S100b-positive neurons (blue), which do not express Sst, these cells were large-diameter and accounted for the other 71% of recombined cells. (c) Functional responses (magenta) were aligned to gene expression (green) showing that small diameter Sst/Nppb-NP3 cells never respond to brush but are heat sensitive consistent with data in Fig. 1. Brush but not heat activates the large diameter S100b-expressing neurons; scale bars, (a, c) 50 µm. (d) Heatmaps of DRG imaging for mice expressing GCaMP under the control of Sst-Cre. Responses from 174 neurons from 7 mice show that Sst-Cre labeled neurons are spontaneously active after injection of PGE2 into the paw, but brush cells were not activated by inflammation.