ENDOSOMAL SIGNALING OF PROTEASE-ACTIVATED RECEPTOR-2 AMPLIFIES HISTAMINE-INDUCED PAIN OF IRRITABLE BOWEL SYNDROME

Background Proteases and histamine, co-secreted by mast cells and bacteria, sensitize colonic nociceptors and contribute to irritable bowel syndrome (IBS) pain. Objective To determine whether irreversible proteolytic cleavage of protease-activated receptor-2 (PAR 2 ) and its continued activity in endosomes amplify and sustain otherwise transient pronociceptive actions of histamine receptors (HRs) to cause recurrent pain, the defining symptom of IBS. Design We investigated the coexpression of PAR 2 and H 1 R in nociceptors using RNAscope in situ hybridization and assessed the consequences of coactivation using electrophysiological assays of nociceptor sensitization and biophysical measurements of receptor and effector activity. Results PAR 2 and H 1 R were coexpressed by human and mouse dorsal root ganglion nociceptors. Intracolonic infusion of fecal supernatants from IBS patients enhanced mechanosensitivity of colonic nociceptors in mice. Antagonists of PAR 2 or H 1-4 R abolished this response. Combined administration of subthreshold concentrations of trypsin and histamine replicated the effects of fecal supernatant and caused hyperexcitability of isolated nociceptors. Pre-activation of PAR 2 sensitized histamine-induced hyperexcitability. Endocytosis inhibitors prevented this hypersensitivity, consistent with sustained endosomal signaling of PAR 2 and persistent nociceptor hyperexcitability. Trypsin amplified histamine-induced activation of H 1 R and β-arrestin2 and Gαq effectors at the plasmalemma and in endosomes. Conversely, histamine did not sensitize trypsin-induced hyperexcitability of neurons, in line with the inability of histamine to induce sustained nociceptor hypersensitivity. Conclusions By amplifying and maintaining the otherwise transient actions of H 1 R and possibly other pain receptors, persistent PAR 2 endosomal signaling makes a dominant contribution to IBS-related colonic pain. Summary box What is already known on this topic Proteases and histamine are increased in IBS patients and cause visceral pain. What this study adds Prolonged intracellular PAR 2 signaling sensitizes and maintains H 1 R activity to amplify and maintain pain. How this might affect research, practice or policy Although neuroactive factors can act synergistically to amplify and maintain IBS pain, antagonists of dominant receptors ( e . g ., PAR 2 ) can provide effective treatment.