GPR68, A Proton-sensing GPCR, Mediates Acid-induced Visceral Nociception

Localised acidification from immune cell infiltration and heightened glycolysis contributes to colitis pathology by activating acid-sensing receptors such as G protein-coupled receptor 68 (GPR68), a proton-sensing G protein-coupled receptor (GPCR) expressed on immune and stromal cells. Single-cell RNA sequencing (RNA-seq) analysis revealed GPR68 is also expressed in colonic sensory neurons, prompting us to investigate its role in acid-induced colonic nociception.

Expression of GPR68 in colonic nociceptors and tissue from people with colitis was confirmed by in silico analysis of our RNA-seq databases. Its contribution to disease activity was assessed using the acute dextran sulphate sodium (DSS) model of colitis. Acid-evoked sensory signalling was evaluated via colonic afferent recordings and Ca²⁺ imaging in DRG neurons from wild-type and GPR68^-/- mice, supported by pharmacological studies using Ogerin (a GPR68 positive allosteric modulator) and Ogremorphin (a GPR68 antagonist).

RNA-seq analysis showed GPR68 is robustly expressed in Trpv1⁺ colonic nociceptors and upregulated in tissue from people with inflammatory bowel disease, consistent with reduced disease activity in DSS-treated GPR68^-/- mice. Genetic deletion of GPR68 abolished colonic afferent responses to acid, which were also attenuated by Ogremorphin and enhanced by Ogerin. In Ca²⁺-free buffer, dorsal root ganglion neurons from GPR68^-/- mice or those pretreated with Ogremorphin showed significantly reduced acid-evoked intracellular Ca²⁺ responses. By contrast, the colonic afferent and dorsal root ganglion Ca²⁺ response (in Ca²⁺-containing buffer) to capsaicin was comparable between tissue from wild-type and GPR68^-/- mice highlighting the involvement of divergent proton-dependent cellular signaling cascades.

These findings identify GPR68 as a key mediator of acid-induced colonic nociception and highlight its potential as a therapeutic target for the treatment of pain in colitis.