Morphological and molecular analysis of PKCε expression in dorsal root ganglia in a rat model of fibromyalgia

Fibromyalgia is a chronic pain syndrome characterized by widespread musculoskeletal pain, fatigue, and altered nociceptive processing . Although both central and peripheral mechanisms have been implicated, the molecular pathways underlying sensory neuron sensitization remain incompletely defined. Protein kinase C epsilon (PKCε), a serine/threonine kinase involved in several cellular processes, is highly expressed in sensory neurons and has been associated with hyperalgesia, ion channel modulation, and neuroinflammatory signaling. This study aimed to investigate PKCε expression and activation in dorsal root ganglia (DRG) in a reserpine-induced rat model of fibromyalgia. Experiments were conducted on Sprague-Dawley rats of both sexes, which were randomized to receive vehicle (acetic acid 0.5%) or reserpine (1 mg/kg). To monitor the development of the fibromyalgia-like phenotype, mechanical allodynia and thermal hyperalgesia were assessed by von Frey and hot plate tests, respectively. Behavioural evaluations were performed before the first treatment on day 1, after three consecutive daily injections of reserpine or vehicle on day 4, and weekly thereafter up to day 28. Animals were euthanized on days 4 and 28, and cervical, thoracic, and lumbar DRG were collected for molecular and morphological analyses. PKCε gene expression was evaluated by RT-PCR, while total and phosphorylated PKCε protein levels were assessed by Western blot. Immunohistochemical analyses were performed to localize PKCε and phosphorylated PKCε in DRG tissue and to evaluate their colocalization with neuronal markers. Since behavioural testing, particularly the hot plate assay, showed heterogeneous results, molecular analysis was performed based on the von Frey-derived mechanical sensitivity profile of reserpine-treated animals. In reserpine-treated and responsive animals, RT-PCR showed an increase in PKCε expression in lumbar DRG compared with controls, whereas cervical and thoracic DRG displayed comparable gene expression levels. Interestingly, western blot analysis revealed increased levels of PKCε and phosphorylated PKCε in cervical and thoracic DRG of reserpine-treated von Frey-responsive rats, suggesting enhanced PKCε activation in animals displaying mechanical allodynia. At later time points, PKCε and phosphorylated PKCε levels returned to values comparable to controls, indicating that these changes may represent an early and transient response. Immunohistochemistry confirmed the quality of DRG samples, as shown by preserved sensory neuron morphology and NeuN positivity, and supported increased PKCε and phosphorylated PKCε expression in DRG from reserpine-treated and responsive rats. Overall, these preliminary findings suggest that PKCε activation may be involved in early peripheral sensory alterations associated with mechanical allodynia in a reserpine-induced fibromyalgia model. Although further studies are required to validate these results and clarify the functional role of PKCε, the present morphological and molecular evidence supports PKCε as a potential contributor to pain-processing pathways and as a candidate target for future mechanism-based studies in fibromyalgia