Immune responses and microbial loads vary drastically along the length of the intestine, but how environmental perturbations locally modulate enteric neuron function remains poorly understood. Here, we regionally profile enteric neurons across gnotobiotic, allergic and parasite-infected mice. We demonstrate that microbial perturbations and type 2 inflammation shift the transcriptional state of inhibitory motor neurons, and establish an in vivo AAV-Perturb-seq approach to identify regulators of this differentiation program. We also show that Nmu-hi sensory neurons are remarkably plastic, and actively respond to IL-13 and leukotrienes, while Grp-hi sensory neurons expand in germ-free colon interacting with Grpr+ interstitial cells of Cajal. These insights deepen our understanding of the ENS and provide a molecular basis for the mechanistic study of its impact on intestinal pathophysiology.

Peng Tan1,2,8,#, Alok Jaiswal2,#, Shane P. Murphy1, Eric M. Brown2,4, Hailey Wheeler2, Chien-Wen Su5, Emily P. Finan1, Guadalupe J. Jasso2, Hai Ning Shi5, Daniel B. Graham2,3,4, Toni M. Delorey1, Jacques Deguine1,2, Ramnik J. Xavier1,2,3,4,6,7,*

1Klarman Cell Observatory, Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.

2Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.

3Department of Molecular Biology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.

4Center for Computational and Integrative Biology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.

5Mucosal Immunology and Biology Research Center, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA 02129, USA

6The Gene Lay Institute of Immunology and Inflammation, Brigham and Women's Hospital, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02115, USA

7Center for the Study of Inflammatory Bowel Disease, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA

8Center for Drug Discovery and Translational Medicine, Hainan Academy of Medical Sciences, Hainan Medical University, Haikou, 571199, PRC

# These authors contributed equally.

*Corresponding author: xavier@molbio.mgh.harvard.edu (R.J.X.)