Endosomal Signaling Of Protease-Activated Receptor-2 Amplifies Histamine-Induced Pain Of Irritable Bowel Syndrome

BackgroundProteases and histamine, co-secreted by mast cells and bacteria, sensitize colonic nociceptors and contribute to irritable bowel syndrome (IBS) pain. ObjectiveTo determine whether irreversible proteolytic cleavage of protease-activated receptor-2 (PAR2) 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. DesignWe investigated the coexpression of PAR2 and H1R 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. ResultsPAR2 and H1R 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 PAR2 or H1-4R 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 PAR2 sensitized histamine-induced hyperexcitability. Endocytosis inhibitors prevented this hypersensitivity, consistent with sustained endosomal signaling of PAR2 and persistent nociceptor hyperexcitability. Trypsin amplified histamine-induced activation of H1R and {beta}-arrestin2 and Gq 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. ConclusionsBy amplifying and maintaining the otherwise transient actions of H1R and possibly other pain receptors, persistent PAR2 endosomal signaling makes a dominant contribution to IBS-related colonic pain. Summary boxO_ST_ABSWhat is already known on this topicC_ST_ABSProteases and histamine are increased in IBS patients and cause visceral pain. What this study addsProlonged intracellular PAR2 signaling sensitizes and maintains H1R activity to amplify and maintain pain. How this might affect research, practice or policyAlthough neuroactive factors can act synergistically to amplify and maintain IBS pain, antagonists of dominant receptors (e.g., PAR2) can provide effective treatment.