Intestinal neuropod cell GUCY2C regulates visceral pain
Joshua R. Barton, … , Manuel Covarrubias, Scott A. Waldman
Joshua R. Barton, … , Manuel Covarrubias, Scott A. Waldman
Published December 22, 2022
Citation Information: J Clin Invest. 2023;133(4):e165578. https://doi.org/10.1172/JCI165578.
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Research Article Gastroenterology Neuroscience

Intestinal neuropod cell GUCY2C regulates visceral pain

Abstract

Visceral pain (VP) is a global problem with complex etiologies and limited therapeutic options. Guanylyl cyclase C (GUCY2C), an intestinal receptor producing cyclic GMP(cGMP), which regulates luminal fluid secretion, has emerged as a therapeutic target for VP. Indeed, FDA-approved GUCY2C agonists ameliorate VP in patients with chronic constipation syndromes, although analgesic mechanisms remain obscure. Here, we revealed that intestinal GUCY2C was selectively enriched in neuropod cells, a type of enteroendocrine cell that synapses with submucosal neurons in mice and humans. GUCY2Chi neuropod cells associated with cocultured dorsal root ganglia neurons and induced hyperexcitability, reducing the rheobase and increasing the resulting number of evoked action potentials. Conversely, the GUCY2C agonist linaclotide eliminated neuronal hyperexcitability produced by GUCY2C-sufficient — but not GUCY2C-deficient — neuropod cells, an effect independent of bulk epithelial cells or extracellular cGMP. Genetic elimination of intestinal GUCY2C amplified nociceptive signaling in VP that was comparable with chemically induced VP but refractory to linaclotide. Importantly, eliminating GUCY2C selectively in neuropod cells also increased nociceptive signaling and VP that was refractory to linaclotide. In the context of loss of GUCY2C hormones in patients with VP, these observations suggest a specific role for neuropod GUCY2C signaling in the pathophysiology and treatment of these pain syndromes.

Authors

Joshua R. Barton, Annie K. Londregan, Tyler D. Alexander, Ariana A. Entezari, Shely Bar-Ad, Lan Cheng, Angelo C. Lepore, Adam E. Snook, Manuel Covarrubias, Scott A. Waldman

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Figure 5

Neuropod cells regulate DRG neuron excitability through GUCY2C.

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Neuropod cells regulate DRG neuron excitability through GUCY2C. (A) Immu...
(A) Immunofluorescence of E15.5 DRG neurons (Pgp9.5+) plated with small intestinal crypts from GUCY2C-GFP mice exhibit outgrowth directed specifically toward GUCY2Chi neuropod cells 24 hours after plating. Scale bar: 50 μm. (B) Representative AP traces of stimulated DRG neurons in coculture with intestinal crypts. DRG neurons exhibit (blue) 1 AP when not near a neuropod (DRG Alone) or (red) repetitive firing when near a neuropod. (C) Summary of DRG neuron rheobase and number of APs fired under conditions B above. (D) Representative AP traces of DRG neurons + Neuropod before (blue) and after addition of 1 μM linaclotide (green), revealing loss of repetitive AP firing after linaclotide treatment. (E) Summary of DRG neuron rheobase and number of APs fired under conditions E above. (F) Representative AP traces of DRG neurons + Neuropod cells from Gucy2c–/– crypts before (blue) and after 1 μM linaclotide (green), reveal no change in repetitive firing. (G) Summary of DRG neuron rheobase and number of APs fired under conditions F above. (H) Representative AP traces of DRG cells + Neuropod cells showing repetitive AP firing before (black) and after 10 μM cGMP treatment (pink). (I) Representative AP traces of DRG neurons showing repetitive AP firing after 10 μM extracellular cGMP and decreased AP firing after subsequent 1 μM linaclotide. (J) Summary of DRG neuron rheobase and number of APs fired under conditions from H above. (K) Summary of DRG neuron rheobase and number of APs fired under conditions from I above. Representative traces represent 1 recording of the DRG neurons plotted in the subsequent rheobase and AP graphs. Statistics for C, E, G, J, and K were calculated using 2-tailed unpaired t test with an f test for comparisons of variances.