Neuropathic pain, which arises from an injury to the nervous system, affects 7–10% of the population and is marked by high severity and persistence. Despite its substantial individual and socioeconomic burden, current treatments remain inadequate and rarely address underlying mechanisms. Therapies that actively promote pain resolution would be highly beneficial, yet the biological processes that drive resolution remain poorly understood. Clinically, women report higher pain scores and chronic pain prevalence. However, sex-specific analyses on the molecular-cellular level, which might help to explain sex differences in pain, are still rare. Here, we hypothesized that pain resolution is dependent on multicellular interactions in the dorsal root ganglia (DRGs), where the somata of sensory neurons are located. We used the chronic constriction injury (CCI), a mild peripheral nerve injury model which shows a natural resolution of pain and experimentally defined pain resolution with the vonFrey assay as a 50% improvement from maximal mechanical hypersensitivity. Then, we characterized cellular and molecular phenotypes of early injury responses and ongoing pain resolution with a focus on neurons, satellite glial cells (SGC) and macrophages in male and female rats. DRG tissue analysis in both sexes combined 7,495 immunofluorescence images and 62 transcriptomes. To address challenges of scalability and subjectivity of bioimage analysis, we implemented a deep learning-based pipeline using the tool deepflash2. Image segmentation was done with model ensembles based on three expert annotators, which allows to increase validity, reproducibility and objectivity of bioimage data analysis. For transcriptome data, we compared differential gene expression after CCI and during pain resolution in both sexes. Following CCI, neuropathic pain resolved without neuronal loss or tissue shrinkage in either sex. Transcriptome and imaging data revealed activation of neuronal repair programs and plasticity of the non-peptidergic marker IB4. Non-neuronal cells also exhibited sex-specific plasticity, with the most pronounced changes in SGC and macrophages during resolution. CCI induced strong activation of immune responses, which we found on both the molecular and cellular level: After injury, macrophages invaded the space between sensory neurons and SGC; this was partially reversed within 5 weeks in male, but not female rats. Conversely, SGC activation marked by increased Gfap-immunoreactivity remained heightened in males, whereas it decreased in females. While injury responses were largely shared between sexes, the resolution phase displayed distinctly sex-specific molecular and cellular signatures. Still, this sex-specific gene expression converged on programs linked to synaptic signaling, neuronal excitation, and cell–cell interactions. This thesis provides a comprehensive, open science dataset that captures the cellular and molecular dynamics of the DRG during the transition from injury to recovery in both male and female rats. Our findings highlight that pain resolution is a coordinated, multicellular, and distinctly sex-specific biological process. The study provides extensive resources for sex-specific factors and biological processes involved in pain resolution for future hypothesis-driven research
Felicitas Schlott (Thu,) studied this question.