A chemogenetic screen for neuro-immune interactions in the gut: Trpv1+ nociceptor neurons control T regulatory cells

Neuroimmune crosstalk is critical for intestinal and tissue homeostasis. Yet the role of molecularly distinct subsets of gut-innervating neurons in regulating the activity of gut immunocytes, and the mechanisms of this neuroimmune signaling remain unclear. Here, we performed a chemogenetic and flow cytometry-based analysis of mice targeting eight different peripheral neuron subsets to assess how gut immunocytes are altered following neural activation. We found that distinct neurons modulated discreet anatomical populations of immunocytes in the gut in ileum, cecum, and colon. Nos1+ neuron activation decreased the percentage of RORg+ ileal conventional CD4+ T cells, whereas ChAT+ neuron activation decreased ileal neutrophils. Trpv1+ neuron activation displayed the most robust immunomodulatory phenotype, causing downregulation of RORg+ T regulatory cells in the colon and cecum. The immune cells exhibited decreased proliferation, enhanced cell stress, and altered cell activation markers. Further genetic and pharmacological approaches showed that spinal afferent Trpv1+ neurons specifically decreased Treg cells by signaling via the neuropeptide CGRP. Our study provided a comprehensive understanding of neuro-immune interactions, revealing a role for mechanisms by which Trpv1+ neurons regulate gut Treg cells. Overall design: We investigated the transcriptional status of cecum immunocytes upon Trpv1+ neuronal activation. To do so we purified CD45+ cecum immunocytes from Trpv1-ADC mice and littermate CTRL mice (2 replicates for each group). Each sample was tagged with DNA-coded anti-CD45 hashtag antibodies (BioLegend). Cells were encapsulated using the Chromium Single Cell 5' v3 platform (10X Genomics).