Accepted

Satellite glial cells (SGCs) activation in the trigeminal ganglia (TG) is critical in various abnormal orofacial sensation in nerve injury and inflammatory conditions. SGCs express several subtypes of P2 purinergic receptors contributing to the initiation and maintenance of neuropathic pain. The P2Y receptor, a G-protein-coupled receptor activated by uridine diphosphate (UDP)-glucose and other UDP sugars, mediates various physiologic events such as immune, inflammation, and pain. However, the expression, distribution, and function of P2Y receptor in SGCs remains largely unexplored. Our study reported the expression and functional identification of P2Y receptor in SGCs. SGCs were isolated from TG of rat, and the P2Y receptor expression was examined using immunofluorescence technique. Cell proliferation and viability were examined via cell counting kit-8 experiment. Immunofluorescence demonstrated the presence of P2Y receptor in SGCs. Immunofluorescence and western blot showed that UDP-glucose treatment upregulated glial fibrillary acid protein, a common marker for glial activation. Extracellular UDP-glucose enhanced the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38, which were both abolished by the P2Y receptor inhibitor (PPTN). Furthermore, quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay demonstrated that extracellular UDP-glucose significantly enhanced interleukin-1β (IL-1β) and chemokine CCL2 (CCL2) release, which was abolished by PPTN and significantly decreased by inhibitors of MEK/ERK (U0126) and p38 (SB202190). Our findings directly proved the functional presence of P2Y receptor in SGCs. It was also verified that P2Y receptor activation was involved in activating SGCs, phosphorylating MAPKs, and promoting the secretion of IL-1β and CCL2 via ERK and p38 pathway.

Functional disorders (i.e., interstitial cystitis/painful bladder syndrome and irritable bowel syndrome) are associated with hyperexcitability of afferent nerves innervating the urinary tract and the bowel respectively. Various non-5-HT receptor mRNA transcripts are expressed in mouse urothelium and exert functional responses to 5-HT. Whilst 5-HT receptors were not detected in mouse urothelium, 5-HT receptors expressed on bladder sensory neurons plays a role in bladder afferent excitability under both normal conditions and in a mouse model of chronic visceral hypersensitivity (CVH). These data suggest that the role 5-HT receptors play in bladder afferent signaling warrants further study as a potential therapeutic target for functional bladder disorders.

Studies using rodent models of neuropathic pain employ sham surgery control procedures that cause deep tissue damage. Sham surgeries would thus be expected to induce potentially long-lasting postsurgical pain, but little evidence for such pain has been reported. Operant tests of voluntary behavior can reveal negative motivational and cognitive aspects of pain that may provide sensitive tools for detecting pain-related alterations. In a previously described operant mechanical conflict (MC) test involving lengthy familiarization and training, rodents freely choose to either escape from a brightly lit chamber by crossing sharp probes or refuse to cross. Here, we describe a brief (2-day) MC protocol that exploits rats' innate exploratory response to a novel environment in order to detect persistently enhanced pain-avoidance behavior after sham surgeries for two neural injury models: thoracic spinal cord injury (SCI) and chronic constriction injury (CCI) of the sciatic nerve. Pitting the combined motivations to avoid the bright light and to explore the novel device against pain from crossing noxious probes disclosed a conflicting, hyperalgesia-related reluctance to repeatedly cross the probes after injury. Rats receiving standard sham surgeries demonstrated enhanced pain-like avoidance behavior compared to naive controls, and this behavior was similar to that of corresponding CCI or SCI rats weeks or months after injury. In the case of sham surgery for SCI, video analysis of voluntary exploratory behavior directed at the probes revealed enhanced forepaw withdrawal responses. These findings have important implications for preclinical investigations into behavioral alterations and physiological mechanisms associated with postsurgical and neuropathic pain.