Pain is a common debilitating symptom reported in people with endometriosis. Yet, we do not have a good understanding of factors that may contribute to higher pain and worse psychosocial outcomes in this population. One key construct linked to higher pain and distress in people with endometriosis is Fear of disease Recurrence or Progression (FORP). Accordingly, in the context of endometriosis, we aimed to identify the prevalence of severe FORP; test whether theoretical constructs from models of fear of cancer recurrence (interpretation bias, metacognitive beliefs, existential concerns) relate to FORP; and better understand the nature of relationships between FORP, theoretical and emotional distress, and pain-related constructs. Participants with a confirmed (n = 274) or provisional (n = 43) diagnosis of endometriosis were recruited through Endometriosis Australia. Participants completed a cross-sectional online survey assessing relevant constructs. We conducted bivariate correlations and network analyses to test for relationships between key constructs. Nearly 85% of participants reported severe levels of FORP. We found that people with greater existential concerns and more unhelpful metacognitive beliefs but not stronger interpretation bias reported greater FORP. However, only existential concerns contributed independent variance in FORP. We also found that individuals with heightened FORP reported greater physical pain and poorer overall well-being. These results suggest that FORP is common and important. Furthermore, metacognitive beliefs and existential concerns were identified as important factors. Testing whether these factors are potentially modifiable intervention targets through longitudinal and intervention studies could ultimately lead to reductions in the impact of endometriosis-associated pain.

Pelvic pain is a common and often debilitating experience with limited treatment options. Improving understanding of the psychosocial mechanisms involved in pelvic pain experiences will enhance the efficacy of pain management interventions. Interpretation bias is a promising psychosocial factor in leading models of chronic pain, yet its relevance for pelvic pain remains poorly understood. The current study sought to assess disorder specific interpretation bias in people with endometriosis and other pelvic pain-related conditions. To do so, we developed a new interpretation bias assessment tool based on existing literature and used this to investigate interpretation bias and relationships with pain, menstrual symptoms, and psychological distress across pelvic pain-related conditions. The sample comprised 342 individuals including those with endometriosis, adenomyosis, chronic pelvic pain, polycystic ovarian syndrome, and those without pelvic pain-related conditions. Participants completed an online survey including demographics, new and modified interpretation bias scenarios, pain-related outcomes, and psychological distress. After a rigorous item selection process, 27 ambiguous pelvic pain-related scenarios were retained, forming the Pelvic Pain Bias Assessment (PPBA). Using the PPBA, we replicated our previous finding of elevated interpretation bias among individuals with endometriosis relative to controls and found interpretation bias was also elevated among individuals with other pelvic pain-related conditions relative to controls. Stronger interpretation bias was associated with worse pelvic pain and menstrual symptoms among individuals with and without pelvic pain-related conditions. This study adds to the growing evidence that interpretation bias is associated with a range of pain conditions and poorer pain-related outcomes in pelvic pain.

Complex regional pain syndrome (CRPS) involves disproportionate chronic pain alongside sensory, autonomic, motor, and trophic disturbances. The pathophysiology of CRPS remains unclear, but evidence indicates that cutaneous neuroimmune dysregulation contributes to symptom onset and maintenance. We conducted a high-parameter imaging mass cytometry and targeted immunofluorescence study of skin biopsies from 18 CRPS patients and 18 healthy controls using a panel of heavy metal isotope-conjugated antibodies to assess nerve fibre, immune cell, and vascular markers in situ. Quantitative data analysis revealed a reduction in proliferating keratinocytes, increased human leukocyte antigen-DR+ (HLA-DR+) Langerhans cell (LC) abundance, and increased HLA-DR+ LC physiological interactions with intraepidermal nerve fibres in patients with CRPS. Langerhans cells also showed sexual dimorphism, being higher in CRPS-affected skin in women compared with men. HLA-DR+CD206- dermal dendritic cells (DC) and CD68+CXCR3+ pro-inflammatory macrophages were increased in CRPS-affected skin. Nerve fibre and blood vessel densities were unchanged. However, intraepidermal nerve fibre density decreased in proportion to temperature asymmetry between disease-affected and contralateral unaffected limbs, being lowest in patients with a cool CRPS-affected limb. These findings highlight significant decreases in epidermal keratinocyte proliferation, increases in LC abundance and nerve fibre interactions, especially the HLA-DR+ subset, and increases in dermal DC cells in CRPS-affected skin, supporting a role for neuroimmune dysfunction and autoimmunity in CRPS pathophysiology.

Parent emotional and behavioral responses play a critical role in shaping youth pain-related outcomes, yet little research has examined these processes day-to-day overtime. This study investigated the contemporaneous and temporal associations between child pain intensity, pain-related distress, and child functional impairment and parent pain-related distress, protective behaviors, and pain acceptance in a sample of youth with chronic pain and their parents. Daily assessments were collected from 68 youth-parent dyads participating in a study investigating a multimodal intervention for chronic pain. Longitudinal Actor-Partner Interdependence Models were used to examine within-day and next-day associations for child and parent variables. Within the same day, higher parent pain-related distress was associated with greater child pain-related distress (β = 0.10, P = 0.001), as well as increased parent protective behaviors (β = 0.44, P < 0.001). Increased parent pain acceptance was associated with decreased child functional impairment (β = 0.15, P <0.001) and protective behaviors (β = -0.09, P = 0.03). Next-day analyses revealed that higher parent pain acceptance predictors lower child pain-related distress and parent protective behaviors. Child pain reductions predicted lower functional impairment and fewer parent protective behaviors the next day. Findings highlight the importance of targeting parent psychological processes, particularly distress and pain acceptance, in pediatric chronic pain interventions. Addressing parent distress and fostering pain acceptance may lead to improvements in both child and parent outcomes, reinforcing the value of dyadic, acceptance-based approaches in pediatric pain treatment.

Assessment of pain in chronic pancreatitis (CP) has largely focused on intensity and pattern, unable to address its complexity. To evaluate pain in a multidimensional fashion, we aimed to identify phenotypes based on the Comprehensive Pain Assessment Tool Short Form (COMPAT-SF) questionnaire and examine their associations with clinical factors. A cross-sectional study including 248 patients with painful CP from Asia, Europe, and the United States was performed. A cluster analysis including the 5 pain dimensions from the COMPAT-SF questionnaire (severity, fluctuation, provocative factors, spreading pain, and qualitative descriptors) identified pain phenotypes. The phenotypes were compared to demographic and clinical data, including patient-reported outcomes and quantitative sensory testing. Three phenotypes were identified in the cluster analysis: a low-burden phenotype, cluster 1 (n = 151); a high-intensity, constant pain phenotype, cluster 2 (n = 75); and a widespread pain, multidimensional phenotype, cluster 3 (n = 22). Quality of life and sleep scores were worse in cluster 3 than in the other phenotypes (all P < 0.001). The degree of anxiety, depression, and catastrophizing was also worse in cluster 3 (all P < 0.001). Cluster 3 showed increased hyperalgesia on sensory testing with a lower sum of pressure pain detection thresholds than cluster 1 (P = 0.008) and higher temporal summation than cluster 2 (P = 0.023). The COMPAT-SF questionnaire thereby identified 3 clinically relevant phenotypes in CP. Widespread, multidimensional pain correlated with increased hyperalgesia, higher psychological distress, and worse overall well-being. Phenotyping based on the COMPAT-SF questionnaire may prove helpful in guiding treatment plans and more accurately allocating patients in clinical trials.

This study investigated the effects of experimentally induced acute pain on body image and sensory testing. A total of 91 pain-free participants were randomly assigned to 1 of 3 groups: injection (pain), sham injection, or control. Measures were taken 3 times in each group: at baseline, during the testing phase, and once the pain had subsided. The study included both sensory testing and body image measures. Participants in the injection group showed significant increases in body image distortion during the testing phase (P < 0.001, η2 = 0.19), alongside larger area of pain distribution (P < 0.001, η2 = 0.40) and greater size distortion of the lower back (P < 0.001, η2 = 0.14) compared with the other groups. A drawing task further revealed that the painful body part was perceived as larger (P = 0.01, η2 = 0.14), particularly on the right side. Body esteem indicated a subtle but significant decrease in body satisfaction during pain (P = 0.05, η2 = 0.06). Sensory testing measures remained unchanged, suggesting intact sensory processing. Correlation analyses showed positive associations between Polish version of the Fremantle Back Awareness Questionnaire scores and perceived pain area, pain intensity, pain catastrophizing, and general fear. These findings highlight the selective influence of acute pain on body image, suggesting that perceptual and emotional factors-driven by higher-order mechanisms play a central role in pain-related body distortions rather than changes in basic sensory processing.

Impairments in lumbar sensory perception, including reduced tactile acuity, occur in patients with nonspecific low back pain (LBP). Tactile acuity is linked to primary somatosensory cortex (S1) activity and structure, but neural markers of lumbar-specific tactile acuity tests remain unvalidated. This cross-sectional study investigated associations between lumbar two-point discrimination (TPD) and estimation (TPE) with functional and structural properties of S1, as well as S1-thalamic connectivity. Resting-state functional MRI and diffusion-weighted MRI assessed S1-thalamic functional connectivity (FC) and structural connectivity, as well as regional homogeneity (ReHo) and mean diffusivity (MD) of S1 grey matter in 78 LBP patients and 39 pain-free controls. Participants with LBP were subdivided into 2 groups: 1 with pain (LBP+, n = 39) and 1 without pain (LBP-, n = 39) on the day of assessment. Higher TPD (ie, worse tactile acuity) was associated with higher contralateral S1-thalamic FC (β = 19.97 mm, 95% CI = 8.47-31.46 mm) and lower contralateral S1-MD (β = -76.98 mm, 95% CI = -142.83 to -11.13 mm). Higher TPE was associated with higher S1-ReHo (β = 19.67 mm, 95% CI = 0.35-39 mm). Two-point discrimination and two-point estimation were positively correlated (r = 0.25, P < 0.001). No between-group differences were found for the MRI variables or TPE, but the LBP+ group showed higher TPD thresholds than pain-free controls (MDiff. = 6.05 mm, Padj. = 0.023). Our findings question the validity of TPE as a measure of tactile acuity. Both neural markers of TPD may not explain tactile acuity impairments in LBP but instead reflect a baseline indicator of tactile performance capability, suggesting poor validity as an LBP-specific marker of neuroplasticity.

The endocannabinoid (eCB) system comprises cannabinoid receptors; endogenous ligands, including anandamide (AEA), 2-arachidonoylglycerol (2-AG); and related N-acylethanolamines (NAEs), N-palmitoylethanolamide (PEA), and N-oleoylethanolamide (OEA); and metabolizing enzymes (eg, fatty acid amide hydrolase [FAAH]). The eCB system modulates nociceptive circuits in rodents. In humans, the FAAH C385A polymorphism is associated with reduced pain sensitivity, suggesting eCB tone contributes to individual pain differences, but this has yet to be tested. Here, we determined whether the eCB system is associated with somatosensory and pain sensitivity measured with quantitative sensory testing (QST) in 91 healthy participants (39 males, 52 females). We tested 3 hypotheses: (1) FAAH C385A polymorphism, cannabis use, and sex affect serum eCB/NAE concentrations; (2) FAAH C385A carriers show altered pain sensitivity vs noncarriers; and (3) baseline serum eCB/NAE concentrations are associated with QST measures. eCB/NAE concentrations were not statistically different based on sex (P > 0.05), FAAH genotype (P > 0.05), or cannabis use (P > 0.05). To address collinearity of AEA, OEA, and PEA, we performed a principal components analysis, which identified a single component of FAAH substrates. Linear regressions found that FAAH genotype did not affect QST measures and that baseline 2-AG and FAAH substrate concentrations were not associated with QST measures, except pressure pain thresholds (PPT; P = 0.003), which were associated with AEA and OEA. Baseline eCB/NAE levels and FAAH genotype are not associated with the outcome measures of standard QST tests that rely on point estimates in healthy adult humans; nonetheless, circulating FAAH substrate levels were associated with PPT.

Neuropathic chronic low-back pain (neuCLBP) is associated with worse clinical outcomes compared with non-neuropathic or axial CLBP (non-neuCLBP) and has limited effective nonsurgical treatment options, reflecting poor understanding of its underlying pathophysiology. In this study, we compared neuCLBP and non-neuCLBP patients using standardized clinical phenotyping of the neuropathic component alongside multimodal brain functional magnetic resonance imaging (fMRI). We hypothesized that, consistent with the definition of neuropathic pain as pain arising from injury to the somatosensory nervous system, neuCLBP patients would exhibit reduced thalamic volume and/or altered thalamic shape, reduced primary somatosensory cortex (S1) thickness, and altered resting-state functional connectivity of these structures compared with non-neuCLBP patients and pain-free healthy controls. Consistent with previous literature, we observed that neuCLBP patients (n = 28) presented with more severe clinical symptoms than non-neuCLBP patients (n = 28). Structurally, neuCLBP patients exhibited extensive differences in thalamic shape but no significant differences in thalamic volume or S1 gray matter thickness. By contrast, by examining resting-state thalamic connectivity gradient maps, we found that non-neuCLBP patients exhibited the most pronounced alterations in these gradients. This study is the first to combine multimodal fMRI with rigorous, standardized phenotyping to investigate neuCLBP. While our results may be influenced by greater symptom severity in the neuCLBP patients, they indicate that these patients may display distinct central plasticity patterns. The findings also highlight the importance of distinguishing between these clinical phenotypes to reduce heterogeneity in future studies.

Sodium channel variants are associated with small fiber neuropathy. While in vitro recordings allow detailed assessment of sodium channel function, their contribution to peripheral small nerve fiber excitability remains unstudied. The perception threshold tracking (PTT) method allows indirect assessment of small fiber function by transcutaneous electrical stimulation and psychophysics. Here, we developed a Sodium channel Excitability Nociceptor Test that allows for the identification of alterations in selected subtypes of sodium channels in small fibers of awake humans. We hypothesize that due to the unique dynamics of sodium channels, it is possible to derive a nerve excitability test to uniquely identify alterations in the Nav1.7 channel. Using a multicompartmental nerve fiber model, a set of 5 electrical pulse shapes was developed. These pulse shapes consist of rectangular and ramp test pulses preceded by either a hyperpolarizing or depolarizing prepulse. To validate the predictive power of our in silico simulations, we used an automated whole-cell patch clamp on Nav1.7 expressed in HEK293T cells and a PTT experiment in healthy participants. The computational model predicted that an alteration of Nav1.7 can be classified with an accuracy of 92% (n = 30) when the standard deviation of the perception threshold is 11%, which was measured by the PTT experiment. The peak Nav1.7 current elicited in vitro corresponded well with the prediction of the in silico model. The study thus provides an excitability test to predict the involvement of Nav1.7, and potentially also Nav1.8 and Nav1.9, in the generation of neuropathic pain in small fiber neuropathy.

A substantial body of evidence points to menstrual pain as a risk factor for chronic pain, yet measures of central sensitization of pain have failed to account for this transition. New research suggests that multisensory sensitivity (MSS) may better characterize those at risk for chronic pain. This study aimed to determine the contribution of MSS to menstrual pain and widespread pain trajectories over 1 year in adolescents with and without primary dysmenorrhea and no other chronic pain conditions at baseline. One hundred forty-one regularly menstruating participants (ages 13-19 years) completed the Sensory Hypersensitivity Scale (SHS) at baseline, and assessments of menstrual pain and widespread pain were completed monthly for 11 months to determine menstrual pain and widespread pain trajectories. There were no differences in reported initiation of hormonal or pain medications or rates of persistent pain between those in the "increase" vs "no increase" in menstrual pain groups. The results indicated main effects of SHS scores on menstrual pain and SHS scores on widespreaed pain. However, for participants with an increase in menstrual pain, there was a significant interaction between SHS scores and widespread pain over time for those with average (β = 0.11, SE = 0.05, P = 0.03) and high (β = 0.19, SE = 0.07, P = 0.01), but not low, SHS scores. These results suggest that MSS plays an important role in understanding the trajectories of both menstrual pain and widespread pain.

There is limited and conflicting evidence about associations between prescription opioid discontinuation and mortality. Two nested case-control studies explored associations between prescription opioid discontinuation and death due to suicide or unintentional overdose. The studies were undertaken within a population-based cohort study linking dispensing data to health and mortality data sets for 371,048 adult residents of New South Wales, Australia, who were dispensed opioids for ≥6 months between July 1, 2003, and December 31, 2018. Cases were individuals with a suicide death or fatal unintentional overdose, matched using risk-set sampling to 10 controls by age, sex, and time. Opioid discontinuation, vs ongoing use, was measured using time-varying exposure periods quantified from dispensing records. Conditional logistic regression was undertaken, with models adjusted for sociodemographics, comorbidities including mental health conditions and substance use disorders, and psychotropic dispensings. Overall, 523 people died by suicide (median age 50 years [interquartile range 39-66], 368 male [70.4%]) and were matched to 5230 controls. Compared with people with ongoing opioid use, discontinuation was not associated with increased odds of experiencing a suicide death (adjusted odds ratio 0.88, 95% confidence interval 0.72-1.07). In addition, 671 people experienced a fatal unintentional overdose (median age 42 years [interquartile range 35-50], 395 male [58.9%]) and were matched to 6710 controls. Opioid discontinuation was associated with reduced odds of experiencing a fatal unintentional overdose (adjusted odds ratio 0.45, 95% confidence interval 0.37-0.54), relative to ongoing use, with the magnitude of this effect greatest when people were unexposed to opioids for >90 days. These findings provide evidence that opioid discontinuation is not necessarily associated with adverse mortality outcomes.

The high failure rate in translating novel analgesics into the clinic has highlighted the need for more translatable biomarkers of analgesic target engagement. The N13 component of spinal somatosensory evoked potential (SEP) has been proposed as a biomarker of spinal nociceptive processing in humans, but it is not known whether this can be back translated into rodents. Tapentadol, lacosamide, and pregabalin were used as pharmacological probes to assess the sensitivity of spinal SEPs to drug action. In anaesthetised, naïve rats (n = 44), a multielectrode silicon probe was inserted into the L4 spinal cord to record SEPs from the dorsal horn after electrical stimulation of the sciatic nerve. At baseline, the N1 component (rodent equivalent of the human N13) had an amplitude of 1.33 ± 0.07 mV at a latency of 4.6 ± 0.2 milliseconds after low-intensity stimulation (average 0.32 mA), with an intensity-dependent amplitude increase into the noxious range (0.4-3.2 mA). The N1 amplitude was significantly reduced by 10 mg/kg tapentadol (40.2 ± 12.5% vs vehicle 96.2 ± 8.0%) and 30 mg/kg lacosamide (46.3 ± 20.9% lacosamide vs vehicle 115 ± 5.9%) at 60 minutes after intraperitoneal administration. Tapentadol also reduced the N1 amplitude in the noxious range. Lacosamide increased the stimulus current required to evoke the half maximal N1 response (EC50), without reducing the maximum N1 amplitude in the noxious range. Pregabalin (at any dose up to 30 mg/kg) did not modulate the N1 amplitude. These results show that the spinal N1 is differentially modulated in a way that reflects distinct mechanisms of analgesic action consistent with it being a translatable biomarker of target engagement.

Mindfulness meditation, a slow breathing practice that is predicated on cultivating nonreactive awareness, also reduces pain. The existing literature has been mixed in demonstrating whether mindfulness meditation is more effective than slow breathing alone (ie, sham-mindfulness meditation) at reducing pain. Furthermore, smaller sample sizes in prior work have hindered a clear understanding of how respiration rate and state anxiety contribute to analgesia during meditation. This study pooled data from 5 randomized controlled trials using paralleling interventions and methodologies in 245 healthy, pain-free, meditation-naïve individuals. Noxious heat (49°C; right calf) was used to test whether changes in respiration and state anxiety mediate the effects of (1) mindfulness meditation (n = 113), (2) a slow-breathing sham-mindfulness meditation technique (n = 73), and (3) a book-listening control (n = 60) on pain intensity ratings ("0" no pain; "10" worst pain imaginable). Three separate 3 (group) × 2 (pre- vs postmanipulation) repeated-measures analysis of variances assessed whether mindfulness and sham-mindfulness meditation significantly reduced pain, anxiety, and respiration, respectively. Mediation analyses examined whether changes in state anxiety and respiration, respectively, mediated analgesia during meditation. Mindfulness meditation produced significantly greater analgesia when compared with sham-mindfulness meditation and controls. Slower respiration rate partially mediated analgesia for mindfulness and sham-mindfulness meditation as compared with controls. Both meditation techniques significantly lowered anxiety, but these improvements did not mediate pain relief. The present findings demonstrate that mindfulness meditation is more effective than slow-breathing (sham-mindfulness) meditation at reducing pain and highlight the role of slower breathing in the direct modulation of acutely evoked pain by meditation.

Small nerve fibre pathology has been observed in several chronic pain conditions not typically classified as neuropathic (e.g., fibromyalgia). This systematic review compared morphometric data of small nerve fibres in people with non-neuropathic chronic pain conditions, defined using ICD-11 classifications, with pain-free controls. MEDLINE, EMBASE, Cochrane, Web of Science, and CINAHL were searched. Two independent reviewers performed screening, risk of bias assessment, and data extraction. Outcomes of interest were intraepidermal nerve fibre density (IENFD) and metrics derived from corneal confocal microscopy (CCM), compared by meta-analysis. Quality of evidence was assessed using Grades of Recommendation, Assessment, Development, and Evaluation. Fifty-two studies were included, examining fibromyalgia (N = 28), complex regional pain syndrome I (N = 5), migraine (N = 4), and other conditions (N = 15). Intraepidermal nerve fibre density was assessed in 41 studies and CCM in 14 studies. Risk of bias was low in 24 studies and moderate in 28. Meta-analysis revealed lower IENFD (fibres/mm2) at the distal leg (MD: -3.06 [95% CI: -3.80 to -2.31]; I2 = 74%; 16 studies; 1123 participants) and proximal thigh (MD: -3.77 [-5.10 to -2.43]; I2 = 86%; 12 studies; 896 participants) and reduced corneal nerve fibre density (MD: -7.14 [-13.56 to -0.72]; I2 = 98%; 8 studies; 768 participants) in participants with non-neuropathic chronic pain compared with controls. Innervation measures differed in participants with chronic pain vs controls for other IENFD and CCM metrics. Quality of evidence ranged from high to low. Associations between clinical features (e.g., pain, disability) and IENFD/CCM data were less commonly examined, mostly indicating no relationships. There is morphological evidence of small nerve fibre pathology in non-neuropathic chronic pain conditions; however, the clinical significance of these findings is unclear.

Nav1.7 has emerged as a promising target for developing novel analgesics that avoid central side effects, yet clinical trials involving systemically administered Nav1.7 inhibitors have thus far yielded disappointing outcomes. In this study, we explored whether delivering PF-05089771-a highly selective Nav1.7 inhibitor previously tested in clinical settings-directly into the intrathecal space could elicit potent analgesic effects. We assessed pain responses using a range of behavioral assays, including the Hargreaves, hot plate, von Frey hair, and Randall-Selitto tests, while evaluating motor coordination and gastrointestinal transit through the rotarod test and a charcoal meal gavage method, respectively. Intrathecal administration of PF-05089771 produced rapid (within 15 minutes) and long-lasting (>4 hours) analgesia across multiple pain models, including nociceptive, inflammatory, neuropathic, and morphine-tolerant pain, as well as both acute and chronic itch. However, no significant effect was observed in a visceral pain model. The analgesic effects were abolished by naloxone pretreatment, implicating endogenous opioid signaling in the mechanism of action. Importantly, repeated intrathecal injections did not lead to analgesic tolerance, and no adverse effects on motor function or gastrointestinal motility were detected. These results indicate that changing the delivery route of Nav1.7 inhibitors may overcome limitations seen with systemic administration and highlight the potential of intrathecal PF-05089771 as a powerful and well-tolerated analgesic.

Pain states fluctuate over time and across situations. Similarly, there is variation in risk and protective factors and how they impact on these pain-related transitions. We are interested in whether such variations are more than random, and whether they can be accounted for by observed variables. The availability of large longitudinal datasets, such as UK Biobank ( https://www.ukbiobank.ac.uk/ ), offers a unique opportunity to study these variations at scale. However, such datasets bring a high risk of bias (eg, confounding) and danger of over-interpretation. It is therefore important to be transparent about our causal thinking. Directed acyclic graphs (DAGs) are graphical representations of the hypothesized causal relationships between variables. They are used to identify the smallest set of variables that need to be adjusted to remove confounding bias in estimating the causal effect of an exposure on an outcome. However, use of DAGs in pain research is not common, despite their potential to guide study design and data analysis. In this article, we present a workflow for building a DAG using domain knowledge from 3 different sources: researchers (theory-based), people with lived experience (person-based), and the literature (evidence-based). We created a DAG for the putative effect of executive function on the maintenance of chronic high-impact pain. The resulting DAG provides a valuable framework for guiding future research on the role of executive functioning in pain, and it underscores the broader potential of using DAGs to improve causal inference in pain research.

Neuropathic pain is a chronic, intractable condition characterized by a complex pathogenesis. Hydrogen-rich water (HRW) has been reported to have possible analgesic properties, but the underlying mechanisms remain unclear. To investigate the effects of HRW, we utilized an L4 spinal nerve ligation (SNL) model in both male and female mice. Pain behaviors were quantitatively assessed using paw withdrawal threshold, paw withdrawal latency, and acetone response tests. Fecal microbiota composition was analyzed through 16S rRNA sequencing, and serum bile acids were assessed by liquid chromatography-mass spectrometry. The expression of bile acid membrane receptor Takeda G-protein-coupled receptor 5 (TGR5) and autophagy markers were evaluated in spinal cord, dorsal root ganglia, and colon through biochemistry experiments. Autophagic structures were examined using transmission electron microscopy. We further validated the roles of TGR5 and autophagy in neuropathic pain through experiments using rapamycin treatment and TGR5 knockout mouse models. The effect of HRW on intestinal barrier function was also tested. We found that HRW effectively ameliorated behavioral deficits associated with SNL-induced neuropathic pain. In addition, HRW facilitated the repair of the damaged gut barrier, restored the disrupted intestinal microbiota, and modulated bile acid metabolism in SNL mice. Interventions with hyodeoxycholic acid and deoxycholic acid produced analgesic effects similar to those of HRW. Takeda G-protein-coupled receptor 5 knockdown reversed the therapeutic effects of HRW and rapamycin. Mechanistically, HRW alleviates neuropathic pain by modulating bile acid metabolites linked to intestinal microbiota and directly or indirectly activating TGR5 in the spinal cord, dorsal root ganglia, and colon, thereby promoting autophagy.