Spontaneous Ongoing Pain Research Articles

Peripheral mu-opioid receptor activation by dermorphin [D-Arg2, Lys4] (1–4) amide alleviates behavioral and neurobiological aberrations in rat model of chemotherapy-induced neuropathic pain

Paclitaxel, a frequently utilized chemotherapeutic agent, often gives rise to severe and distressing sensory neuropathy in patients undergoing chemotherapy. Unfortunately, current therapeutics for chemotherapy-induced neuropathic pain (CINP) demonstrate limited effectiveness and are burdened with the potential for central side effects such as sedation, respiratory depression, cognitive impairment, and addiction, posing substantial clinical challenges. In light of these limitations, present study is designed to investigate the therapeutic potential of Dermorphin [D-Arg2, Lys4] (1–4) amide (DALDA), a preferential peripherally acting mu-opioid receptor agonist, in rat model of CINP. The primary objective was to assess the analgesic properties of DALDA and elucidate the underlying mechanisms governing its therapeutic activity. Our findings revealed that DALDA treatment significantly ameliorated paclitaxel-induced evoked and spontaneous ongoing pain in rats without causing drug addiction and other central side effects. Molecular analyses further unveiled that paclitaxel administration resulted in increased expression of TRP channels, NR2B, voltage-gated sodium channels (VGSCs) and neuroinflammatory markers in both the dorsal root ganglion (DRG) and the spinal cord (L4-L5 region) of rats. DALDA treatment significantly downregulated ion channels (TRPs, VGSCs) and NR2B expressions, concomitant with the inhibition of microglial activation, resulting in the suppression of oxido-nitrosative stress and neuroinflammatory cascade. Findings from the current study suggests that peripheral mu-opioid receptors may offer a potential target for the treatment of patients suffering from CINP, offering new avenues for improved pain relief while minimizing central side effects.

Β-Lactam Antibiotic Ceftriaxone As a Potential Therapeutic Intervention for Chronic Pain in Sickle Cell Disease

Sickle cell disease (SCD) features the presence of complex physiologic changes beyond a simple vasculopathy. Among its most debilitating neurological complications, chronic pain remains a challenging condition with limited effective treatments. Patients with SCD suffer tremendous pain every day. β-Lactam antibiotics have been shown to offer neuroprotection by promoting glutamate transporter GLT1 expression in the nervous system. This study aimed to investigate the effect of ceftriaxone, a prototype beta-lactam antibiotic, on chronic pain in SCD. In a humanized mouse model of SCD, spontaneous ongoing pain as well as evoked hypersensitivity to mechanical and thermal stimuli were present in mice carrying human sickle hemoglobin, but not non-sickle control littermates. Notably, prominent activation of astrocytes was observed in the spinal cord dorsal horn region in the mice with SCD. As compared with non-sickle littermates, the spinal expression of GLT1 was dramatically decreased in sickle cell mice. Repeated intraperitoneal administration of ceftriaxone effectively attenuated spontaneous pain, mechanical allodynia, and heat hyperalgesia in mice with SCD. We found the pain reversal effect by ceftriaxone (200 mg/kg, i.p., for 7 days) lasted for at least 4 weeks. Correlating with the behavioral manifestations, ceftriaxone significantly deactivated astrocytes and elevated the level of GLT1 in the spinal cord in the mice with SCD. These findings suggest that ceftriaxone alleviates both non-evoke ongoing pain and evoked pain in SCD by up-regulating spinal GLT1 expression, which highlights the possibility of a new clinical strategy to treat chronic pain in SCD. This study sheds light on the role of astrocyte activation in SCD and lays the foundation for the development of a potential new intervention for chronic pain in sickle cell disease.

Bergenin ameliorates chemotherapy-induced neuropathic pain in rats by modulating TRPA1/TRPV1/NR2B signalling

Chemotherapy-induced neuropathic pain (CINP) is one of the most prominent and incapacitating complication associated with chemotherapeutic regimens. The exact mechanisms underlying CINP are not fully understood yet, which hampers the development of effective therapeutics. The current study has been designed to investigate the effect of bergenin on CINP and dissect the underlying cellular and molecular mechanisms. Behavioural responsiveness assays were conducted in rats before and after CINP induction and at different time points post-bergenin treatment. We also measured alterations in tight junction proteins, pro-inflammatory cytokines, microglia activity, transient receptor potential (TRP) channels (TRPV1, TRPA1 and TRPM8) and N-methyl-D-aspartate receptor subtype 2 (NR2B) in dorsal root ganglion (DRG) and spinal tissues of neuropathic rats. Bergenin treatment leads to a significant and dose-dependent reduction in evoked and spontaneous ongoing pain without causing central side effects in neuropathic rats. Furthermore, treatment with bergenin and gabapentin did not affect the baseline pain threshold in healthy, non-chemotherapy-treated rats, as evaluated through tail-flick and tail-clip assays. Chemotherapy administration leads to a significant activation of TRP channels, concurrent with microglial activation, disruption of spinal cord tight junction proteins, and subsequent infiltration of pro-inflammatory cytokines, as well as NR2B activation. Notably, bergenin treatment effectively reversed all of these alterations, with the exception of TRPM8, in both the DRG and spinal cord of neuropathic rats. Findings from the present study suggests that bergenin mitigates neuropathic pain by modulating the TRPA1/TRPV1/NR2B signalling and presents a promising therapeutic avenue for the treatment of chemotherapy-induced neuropathic pain.

Gut microbiota dysbiosis alters chronic pain behaviors in a humanized transgenic mouse model of sickle cell disease.

Pain is the most common symptom experienced by patients with sickle cell disease (SCD) throughout their lives and is the main cause of hospitalization. Despite the progress that has been made towards understanding the disease pathophysiology, major gaps remain in the knowledge of SCD pain, the transition to chronic pain, and effective pain management. Recent evidence has demonstrated a vital role of gut microbiota in pathophysiological features of SCD. However, the role of gut microbiota in SCD pain is yet to be explored. We sought to evaluate the compositional differences in the gut microbiota of transgenic mice with SCD and nonsickle control mice and investigate the role of gut microbiota in SCD pain by using antibiotic-mediated gut microbiota depletion and fecal material transplantation (FMT). The antibiotic-mediated gut microbiota depletion did not affect evoked pain but significantly attenuated ongoing spontaneous pain in mice with SCD. Fecal material transplantation from mice with SCD to wild-type mice resulted in tactile allodynia (0.95 ± 0.17 g vs 0.08 ± 0.02 g, von Frey test, P < 0.001), heat hyperalgesia (15.10 ± 0.79 seconds vs 8.68 ± 1.17 seconds, radiant heat, P < 0.01), cold allodynia (2.75 ± 0.26 seconds vs 1.68 ± 0.08 seconds, dry ice test, P < 0.01), and anxiety-like behaviors (Elevated Plus Maze Test, Open Field Test). On the contrary, reshaping gut microbiota of mice with SCD with FMT from WT mice resulted in reduced tactile allodynia (0.05 ± 0.01 g vs 0.25 ± 0.03 g, P < 0.001), heat hyperalgesia (5.89 ± 0.67 seconds vs 12.25 ± 0.76 seconds, P < 0.001), and anxiety-like behaviors. These findings provide insights into the relationship between gut microbiota dysbiosis and pain in SCD, highlighting the importance of gut microbial communities that may serve as potential targets for novel pain interventions.

Inhibition of T-Type Calcium Channels With TTA-P2 Reduces Chronic Neuropathic Pain Following Spinal Cord Injury in Rats

Spinal cord injury (SCI)-induced neuropathic pain (SCI-NP) develops in up to 60 to 70% of people affected by traumatic SCI, leading to a major decline in quality of life and increased risk for depression, anxiety, and addiction. Gabapentin and pregabalin, together with antidepressant drugs, are commonly prescribed to treat SCI-NP, but their efficacy is unsatisfactory. The limited efficacy of current pharmacological treatments for SCI-NP likely reflects our limited knowledge of the underlying mechanism(s) responsible for driving the maintenance of SCI-NP. The leading hypothesis in the field supports a major role for spontaneously active injured nociceptors in driving the maintenance of SCI-NP. Recent data from our laboratory provided additional support for this hypothesis and identified the T-type calcium channels as key players in driving the spontaneous activity of SCI-nociceptors, thus providing a rational pharmacological target to treat SCI-NP. To test whether T-type calcium channels contribute to the maintenance of SCI-NP, male and female SCI and sham rats were treated with TTA-P2 (a blocker of T-type calcium channels) to determine its effects on mechanical hypersensitivity (as measured with the von Frey filaments) and spontaneous ongoing pain (as measured with the conditioned place preference paradigm), and compared them to the effects of gabapentin, a blocker of high voltage–activated calcium channels. We found that both TTA-P2 and gabapentin reduced mechanical hypersensitivity in male and females SCI rats, but surprisingly only TTA-P2 reduced spontaneous ongoing pain in male SCI rats. PerspectivesSCI-induced neuropathic pain, and in particular the spontaneous ongoing pain component, is notoriously very difficult to treat. Our data provide evidence that inhibition of T-type calcium channels reduces spontaneous ongoing pain in SCI rats, supporting a clinically relevant role for T-type channels in the maintenance of SCI-induced neuropathic pain.

Reshaping the gut microbiota affects pain and anxiety-like behaviors in a transgenic model of sickle cell disease

Sickle cell disease (SCD) patients experience recurrent episodes of excruciating acute pain and persistent chronic ongoing pain which negatively impacts their quality of life and is the main cause for hospitalizations. Although major progress has been made in understanding the complex SCD pathophysiology, marked deficiencies in the knowledge regarding SCD pain, the factors underlying the transition from acute to chronic pain, pain comorbidities and effective pain management have been realized. Several studies have reported a crucial role of gut microbiota in the pathophysiological features of SCD; however, its role in pain has not been explored. Our study aimed at evaluating the compositional differences in the gut microbiota of SCD transgenic mice and non-sickle control mice and differences in the gut-derived metabolites. We further investigated the role of gut microbiota in SCD pain by utilizing antibiotic-mediated gut microbiota depletion and fecal microbiota transplantation (FMT). We report significant differences in the alpha and beta diversity between the two cohorts. The antibiotic mediated gut microbiota depletion did not affect evoked pain but significantly attenuated ongoing spontaneous pain in SCD mice. FMT from sickle to non-sickle mice resulted in reduced pain threshold for von Frey (0.95±0.17g vs 0.08±0.02g, p&lt;0.001), reduced withdrawal latency for heat hyperalgesia and cold allodynia (15.10±0.79s vs 8.68±1.17s, p&lt;0.01and 2.75±0.26s vs 1.68±0.08s, p&lt;0.01) along with an increase in anxiety-like behaviors tested using elevated-plus maze (EPM) and open field test (OFT). The transfer of gut microbiota from SCD mice to non-sickle control mice was sufficient to evoke anxiety-like behaviors in the recipient mice as evidenced by decreased time spent in the open arm of the EPM and decreased time spent in the central compartment of the open field (76.18±13.18s vs 36.15±7.34s, p&lt;0.05 for EPM and 126.4±35.3s vs 53.98±19.33s for OFT). Additionally, reshaping the gut microbiota of SCD mice with FMT resulted in reduced mechanical allodynia (0.05±0.01g vs 0.25±0.03g, p&lt;0.001) and heat hyperalgesia (5.89±0.67s vs 12.25±0.76s, p&lt;0.001) and reduced anxiety-like behaviors as evidenced by increased amount of time spent in open arms of the elevated plus maze (20.2±6.3s vs 42.1±15.03s) and increased amount of time spent in the central chamber of the open field (44.03±14.49s vs 104.4±[WJ1] 32.79s). These findings provide novel insights into the relationship between gut microbiota and pain in SCD, highlighting the different gut microbial communities which may serve as potential targets for pain management. Funding: This study was supported in part by a grant from the National Heart, Lung, and Blood Institute (NHLBI) R35 HL140021 (Z.W.). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NHLBI or NIH. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Development of a spontaneous pain indicator based on brain cellular calcium using deep learning

Chronic pain remains an intractable condition in millions of patients worldwide. Spontaneous ongoing pain is a major clinical problem of chronic pain and is extremely challenging to diagnose and treat compared to stimulus-evoked pain. Although extensive efforts have been made in preclinical studies, there still exists a mismatch in pain type between the animal model and humans (i.e., evoked vs. spontaneous), which obstructs the translation of knowledge from preclinical animal models into objective diagnosis and effective new treatments. Here, we developed a deep learning algorithm, designated AI-bRNN (Average training, Individual test-bidirectional Recurrent Neural Network), to detect spontaneous pain information from brain cellular Ca2+ activity recorded by two-photon microscopy imaging in awake, head-fixed mice. AI-bRNN robustly determines the intensity and time points of spontaneous pain even in chronic pain models and evaluates the efficacy of analgesics in real time. Furthermore, AI-bRNN can be applied to various cell types (neurons and glia), brain areas (cerebral cortex and cerebellum) and forms of somatosensory input (itch and pain), proving its versatile performance. These results suggest that our approach offers a clinically relevant, quantitative, real-time preclinical evaluation platform for pain medicine, thereby accelerating the development of new methods for diagnosing and treating human patients with chronic pain.

Gingerol-Enriched Ginger Supplementation Mitigates Neuropathic Pain via Mitigating Intestinal Permeability and Neuroinflammation: Gut-Brain Connection.

Objectives: Emerging evidence suggests an important role of the gut-brain axis in the development of neuropathic pain (NP). We investigated the effects of gingerol-enriched ginger (GEG) on pain behaviors, as well as mRNA expressions of inflammation via tight junction proteins in GI tissues (colon) and brain tissues (amygdala, both left and right) in animals with NP. Methods: Seventeen male rats were randomly divided into three groups: Sham, spinal nerve ligation (SNL, pain model), and SNL+0.375% GEG (wt/wt in diet) for 4 weeks. Mechanosensitivity was assessed by von Frey filament tests and hindpaw compression tests. Emotional responsiveness was measured from evoked audible and ultrasonic vocalizations. Ongoing spontaneous pain was measured in rodent grimace tests. Intestinal permeability was assessed by the lactulose/D-mannitol ratio in urine. The mRNA expression levels of neuroinflammation (NF-κB, TNF-α) in the colon and amygdala (right and left) were determined by qRT-PCR. Data was analyzed statistically. Results: Compared to the sham group, the SNL group had significantly greater mechanosensitivity (von Frey and compression tests), emotional responsiveness (audible and ultrasonic vocalizations to innocuous and noxious mechanical stimuli), and spontaneous pain (rodent grimace tests). GEG supplementation significantly reduced mechanosensitivity, emotional responses, and spontaneous pain measures in SNL rats. GEG supplementation also tended to decrease SNL-induced intestinal permeability markers. The SNL group had increased mRNA expression of NF-κB and TNF-α in the right amygdala and colon; GEG supplementation mitigated these changes in SNL-treated rats. Conclusion: This study suggests GEG supplementation palliated a variety of pain spectrum behaviors in a preclinical NP animal model. GEG also decreased SNL-induced intestinal permeability and neuroinflammation, which may explain the behavioral effects of GEG.

Cold allodynia is correlated to paroxysmal and evoked mechanical pain in complex regional pain syndrome (CRPS).

Mechanisms of complex regional pain syndrome (CRPS) are still debated. Identifying subgroups of patients have been attempted in the hope of linking clinical findings to possible mechanisms. The aim of the present study was to investigate whether subgroups of CRPS (based on quantitative sensory testing (QST)-results) differed with respect to different characteristics of pain like spontaneous ongoing or paroxysmal pain and mechanical dynamic allodynia. 61 CRPS-patients (type 1 and 2) were examined clinically and with QST, in affected and contralateral extremity, with assessment of thresholds for warmth, cold and heat-and cold pain. 43 patients (20 men, 23 men) were diagnosed with CRPS 1 (70.5%) and 18 patients (8 women and 10 men) with CRPS 2 (29.5%). Three subgroups were defined based on thermal thresholds; A (thermal allodynia 22.9%), B (thermal hyposensitivity 37.3%), C (thermal allodynia and hyposensitivity 39.3%). Paroxysmal pain was more prevalent in patients with thermal allodynia (merging group A+C, 25/38-65.8%) compared to patients without thermal allodynia (group B, 5/23-21.7%) (p-value=0.00085). We suggest that cold allodynia is based on hyper-excitability of very superficial skin nociceptors. The correlation between paroxysmal pain, allodynia to light touch and cold allodynia suggests that activity in those peripheral nociceptors can drive both, paroxysmal pain and spinal sensitization leading to stroke evoked allodynia. Mechanistically, the physical cold stimulus can unmask disease-related hyperexcitability by closure of temperature-sensitive potassium channels or induction of resurgent currents. Small fiber degeneration alone may not be the crucial mechanism in CRPS, nor explain pain.

Inhibition of pan-Aurora kinase attenuates evoked and ongoing pain in nerve injured rats via regulating KIF17-NR2B mediated signaling

Kinesins (KIF’s) are the motor proteins which are recently reported to be involved in the trafficking of nociceptors leading to chronic pain. Aurora kinases are known to be involved in the regulation of KIF proteins which are associated with the activation of N-methyl-D-aspartate (NMDA) receptors. Here, we investigated the effect of tozasertib, a pan-Aurora kinase inhibitor, on nerve injury-induced evoked and chronic ongoing pain in rats and the involvement of kinesin family member 17 (KIF17) and NMDA receptor subtype 2B (NR2B) crosstalk in the same. Rats with chronic constriction injury showed a significantly decreased pain threshold in a battery of pain behavioural assays. We found that tozasertib [10, 20, and 40 mg/kg intraperitoneally (i.p.)] treatment showed a significant and dose-dependent inhibition of both evoked and chronic ongoing pain in rats with nerve injury. Tozasertib (40 mg/kg i.p.) and gabapentin (30 mg/kg i.p.) treatment significantly inhibits spontaneous ongoing pain in nerve injured rats but did not produce any place preference behaviour in healthy naïve rats pointing towards their non-addictive analgesic potential. Moreover, tozasertib (10, 20, and 40 mg/kg i.p.) and gabapentin (30 mg/kg i.p.) treatment did not altered the normal pain threshold in healthy naïve rats and didn’t produce central nervous system associated side effects as well. Western blotting and reverse transcription polymerase chain reaction studies suggested enhanced expressions of NR2B and KIF-17 along with increased nuclear factor kappa β (NFkβ), tumour necrosis factor-α (TNF-α), interleukin 1β (IL-1β), and interleukin 6 (IL-6) levels in dorsal root ganglion (DRG) and spinal cord of nerve injured rats which was significantly attenuated on treatment with different does of Tozasertib. Findings from the current study suggests that inhibition of pan-Aurora kinase decreased KIF-17 mediated NR2B activation which further leads to significant inhibition of evoked and chronic ongoing pain in nerve-injured rats.

Protein kinase Cδ as a neuronal mechanism for headache in a chronic intermittent nitroglycerin model of migraine in mice.

Migraine is one of the most common neurological disorders characterized by recurrent attacks of typically throbbing and unilateral headaches, affecting up to 20% of the population worldwide. Despite the high prevalence and severity of this primary headache disorder, it remains to be a challenge to fully understand and treat migraine headaches. By characterizing and validating a mouse migraine model, this study aimed to investigate the functional contribution of protein kinase C (PKC) isoforms in migraine. In this study, we identified the presence of migraine-like ongoing pain in mice after chronic intermittent treatment with nitroglycerin (NTG). The peptide antagonist of calcitonin gene-related peptide α-CGRP (8-37), but not topiramate nor sumatriptan, effectively blocked ongoing pain and elicited pain relief-induced conditioned place preference in NTG-treated mice. Prominent activation of PKCδ was observed in chronic NTG-treated mice. Functional inhibition of PKCδ significantly attenuated ongoing spontaneous pain in chronic NTG-treated mice. Furthermore, we recapitulated the NTG-triggered migraine behavior in wild-type mice, but not in PKCδ-null mice. In response to repeated administration of NTG, ongoing spontaneous pain was not developed in mice lacking the specific PKC isoform. This study identified the presence of ongoing pain in mice treated with NTG, a known human migraine trigger that closely resembles the common manifestation of spontaneous migraine attacks in humans. These findings demonstrated a critical regulatory role of PKCδ in migraine pathophysiology, which may offer new pharmacological targets for antimigraine treatment.

Dorsal root ganglion stimulation of injured sensory neurons in rats rapidly eliminates their spontaneous activity and relieves spontaneous pain.

Dorsal root ganglion field stimulation (GFS) relieves evoked and spontaneous neuropathic pain by use-dependent blockade of impulse trains through the sensory neuron T-junction, which becomes complete within less than 1 minute for C-type units, also with partial blockade of Aδ units. We used this tool in the spinal nerve ligation (SNL) rat model to selectively block sensory neuron spontaneous activity (SA) of axotomized neurons at the fifth lumbar (L5) level vs blockade of units at the L4 level that remain uninjured but exposed to inflammation. In vivo dorsal root single-unit recordings after SNL showed increased SA in L5 units but not L4 units. Ganglion field stimulation blocked this SA. Ganglion field stimulation delivered at the L5 dorsal root ganglion blocked mechanical hyperalgesia behavior, mechanical allodynia, and ongoing spontaneous pain indicated by conditioned place preference, whereas GFS at L4 blocked evoked pain behavior but not spontaneous pain. In vivo single-unit recordings of spinal cord dorsal horn (DH) wide-dynamic-range neurons showed elevated SA after SNL, which was reduced by GFS at the L5 level but not by GFS at the L4 level. In addition, L5 GFS, but not L4 GFS, increased mechanical threshold of DH units during cutaneous mechanical stimulation, while L5 GFS exceeded L4 GFS in reducing evoked firing rates. Our results indicate that SA in injured neurons supports increased firing of DH wide-dynamic-range neurons, contributing to hyperalgesia, allodynia, and ongoing pain. Ganglion field stimulation analgesic effects after nerve injury are at least partly attributable to blocking propagation of this SA.

The bivalent ligand, MMG22, reduces neuropathic pain after nerve injury without the side effects of traditional opioids.

Functional interactions between the mu opioid receptor (MOR) and the metabotropic glutamate receptor 5 (mGluR5) in pain and analgesia have been well established. MMG22 is a bivalent ligand containing MOR agonist (oxymorphamine) and mGluR5 antagonist (MPEP) pharmacophores tethered by a 22-atom linker. MMG22 has been shown to produce potent analgesia in several models of chronic inflammatory and neuropathic pain (NP). This study assessed the efficacy of systemic administration of MMG22 at reducing pain behavior in the spared nerve injury (SNI) model of NP in mice, as well as its side-effect profile and abuse potential. MMG22 reduced mechanical hyperalgesia and spontaneous ongoing pain after SNI, with greater potency early (10 days) as compared to late (30 days) after injury. Systemic administration of MMG22 did not induce place preference in naive animals, suggesting absence of abuse liability when compared to traditional opioids. MMG22 also lacked the central locomotor, respiratory, and anxiolytic side effects of its monomeric pharmacophores. Evaluation of mRNA expression showed the transcripts for both receptors were colocalized in cells in the dorsal horn of the lumbar spinal cord and dorsal root ganglia. Thus, MMG22 reduces hyperalgesia after injury in the SNI model of NP without the typical centrally mediated side effects associated with traditional opioids.

Activation of the Intrinsic Pain Inhibitory Circuit from the Midcingulate Cg2 to Zona Incerta Alleviates Neuropathic Pain.

Neuropathic pain is one of the most common and notorious neurological diseases. The changes in cerebral structures after nerve injury and the corresponding contributions to neuropathic pain are not well understood. Here we found that the majority of glutamatergic neurons in the area 2 of midcingulate cortex (MCC Cg2Glu) were inhibited by painful stimulation in male mice. Optogenetic manipulation revealed that these neurons were tonically involved in the inhibitory modulation of multimodal nociception. We further identified the projections to GABAergic neurons in the zona incerta (ZIGABA) mediated the pain inhibitory role. However, MCC Cg2Glu became hypoactive after nerve injury. Although a brief activation of the MCC Cg2Glu to ZIGABA circuit was able to relieve the aversiveness associated with spontaneous ongoing pain, consecutive activation of the circuit was required to alleviate neuropathic allodynia. In contrast, glutamatergic neurons in the area 1 of MCC played opposite roles in pain modulation. They became hyperactive after nerve injury and only consecutive inhibition of their activity relieved allodynia. These results demonstrate that MCC Cg2Glu constitute a component of intrinsic pain inhibitory circuitry and their hypoactivity underlies neuropathic pain. We propose that selective and persistent activation of the MCC Cg2Glu to ZIGABA circuit may serve as a potential therapeutic strategy for this disease.SIGNIFICANCE STATEMENT Glutamatergic neurons in the area 2 of midcingulate cortex (MCC Cg2Glu) are tonically involved in the intrinsic pain inhibition via projecting to GABAergic neurons in the zona incerta. They are hypoactive after nerve injury. Selective activation of the circuit compensates the reduction of its analgesic strength and relieves neuropathic pain. Therefore, MCC Cg2Glu and the related analgesic circuit may serve as therapeutic targets for neuropathic pain. In contrast, MCC Cg1Glu have an opposite role in pain modulation and become hyperactive after nerve injury. The present study provides novel evidence for the concept that neuropathic pain is associated with the dysfunction of endogenous pain modulatory system and new perspective on the treatment of neuropathic pain.

Gabapentin alleviates chronic spontaneous pain and acute hypoxia-related pain in a mouse model of sickle cell disease.

Pain is the main complication of sickle cell disease (SCD). Individuals with SCD experience acute pain episodes and chronic daily pain, both of which are managed with opioids. Opioids have deleterious side effects and use-associated stigma that make them less than ideal for SCD pain management. After recognizing the neuropathic qualities of SCD pain, clinically-approved therapies for neuropathic pain, including gabapentin, now present unique non-opioid based therapies for SCD pain management. These experiments explored the efficacy of gabapentin in relieving evoked and spontaneous chronic pain, and hypoxia/reoxygenation (H/R)-induced acute pain in mouse models of SCD. When administered following H/R, a single dose of gabapentin alleviated mechanical hypersensitivity in SCD mice by decreasing peripheral fibre activity. Gabapentin treatment also alleviated spontaneous ongoing pain in SCD mice. Longitudinal daily administration of gabapentin failed to alleviate H/R-induced pain or chronic evoked mechanical, cold or deep tissue hypersensitivity in SCD mice. Consistent with this observation, voltage-gated calcium channel (VGCC) α2 δ1 subunit expression was similar in sciatic nerve, dorsal root ganglia and lumbar spinal cord tissue from SCD and control mice. Based on these data, gabapentin may be an effective opioid alternative for the treatment of chronic spontaneous and acute H/R pain in SCD.

Chronic pain after blast-induced traumatic brain injury in awake rats

Explosive blast-induced traumatic brain injury (blast-TBI) in military personnel is a leading cause of injury and persistent neurological abnormalities, including chronic pain. We previously demonstrated that chronic pain after spinal cord injury results from central sensitization in the posterior thalamus (PO). The presence of persistent headaches and back pain in veterans with blast-TBI suggests a similar involvement of thalamic sensitization. Here, we tested the hypothesis that pain after blast-TBI is associated with abnormal increases in activity of neurons in PO thalamus. We developed a novel model with two unique features: (1) blast-TBI was performed in awake, un-anesthetized rats, to simulate the human experience and to eliminate confounds of anesthesia and surgery inherent in other models; (2) only the cranium, rather than the entire body, was exposed to a collimated blast wave, with the blast wave striking the posterior cranium in the region of the occipital crest and foramen magnum. Three weeks after blast-TBI, rats developed persistent, ongoing spontaneous pain. Contrary to our hypothesis, we found no significant differences in the activity of PO neurons, or of neurons in the spinal trigeminal nucleus. There were also no significant changes in gliosis in either of these structures. This novel model will allow future studies on the pathophysiology of chronic pain after blast-TBI.

Protein kinase C‐beta isoform as a mechanism promoting chronic pain in sickle cell disease

Pain is a hallmark symptom in sickle cell disease (SCD). Besides acute painful vaso‐occlusive crises, SCD is also accompanied by intractable chronic pain. This persistent, and often unrelieved, pain starts early in childhood and continues throughout life. The neurobiological mechanisms of chronic pain in SCD remain unclear, which limits effective pain management and the quality of life in patients with SCD. Taking advantage of a humanized mouse model of SCD, this study aimed to investigate protein kinase C (PKC)‐beta isoform mechanism for chronic pain in SCD. We characterized pain phenotypes in the Townes' mouse model (TOW) of SCD that exclusively expresses human alleles encoding normal α‐ and sickle β‐globin. TOW mice exhibited ongoing spontaneous pain as well as hypersensitivity to evoked pain stimuli compared with littermate control mice. We then examined spinal PKCbeta‐mediated signaling and behavioral consequences. Prominent activation of PKCbeta was identified in the superficial laminae of the spinal cord in TOW sickle mice. Functional inhibition and genetic silencing of the PKCbeta attenuated sickle cell pain phenotypes, abolishing PKCbeta activation, spontaneous pain, mechanical allodynia, and heat hyperalgesia in the sickle cell mice. Taken together, these data suggest PKCbeta may be one of the mechanisms for the generation and maintenance of ongoing and evoked pain in SCD. These findings offer new insights into sickle cell pain mechanisms, which may offer new pharmacological interventions.Support or Funding InformationR35 HL140031 and K99 HL133590This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Protein Phosphorylation Mechanisms Underlying Chronic Pain in Sickle Cell Disease

Pain is one of the most dreadful symptoms in sickle cell disease (SCD) and is often refractory to currently available analgesics. Besides acute painful vaso-occlusive crises, SCD is also accompanied by intractable chronic pain. This persistent, and often unrelieved, pain starts early in childhood and continues throughout life. The neurobiological mechanisms of chronic pain in SCD remain unclear, which markedly limits effective pain management and the quality of life in patients with SCD. Taking advantage of two humanized mouse models of SCD, this study aimed to investigate protein phosphorylation mechanisms for chronic pain in SCD.We characterized pain in two transgenic mice models of SCD that exclusively express human alleles encoding normal α- and sickle β-globin. Berkeley SCD mice (BERK mice) and Townes' SCD mice (TOW mice) exhibited ongoing spontaneous pain behavior and increased sensitivity to evoked pain stimuli compared with littermate control mice expressing normal human hemoglobins. To investigate the underlying protein phosphorylation mechanisms of chronic pain in SCD, we examined PKC isoform mediated nociceptive signaling. Prominent activation of multiple PKC isoforms were observed in the superficial laminae of the spinal cord dorsal horn in BERK and TOW mice. Functional inhibition and silencing of specific PKC isoforms attenuated spontaneous pain, mechanical allodynia, and heat hyperalgesia in both transgenic SCD mice. Furthermore, employing hematopoietic stem cell transplantation approach, we were able to generate a sickle cell anemia model in PKC-null mice, allowing us to specifically target neuronal PKC in SCD. Neither spontaneous pain nor evoked pain was detected in the mice lacking specific PKC isoform despite the full establishment of SCD phenotypes.In summary, this study is the first to identify the presence of ongoing spontaneous pain in preclinical sickle cell models, which closely mimic the most prevalent manifestation in patients with SCD. Moreover, we found that spinal PKC is a critical mechanism for the generation and maintenance of ongoing and evoked pain in SCD. These findings offer insights into sickle cell pain mechanisms, which may become a potential target for pharmacological interventions. DisclosuresNo relevant conflicts of interest to declare.

Spinal Contusion Injury Induces Long‐Lasting Changes in Home Cage Activity and Respiration which Correlate with Spontaneous and Evoked Indices of Neuropathic Pain

Spinal cord injury (SCI) often results in chronic neuropathy and dysregulation of motor and respiratory behavior. Quantifying these changes in rodents is difficult, often invasive, and can alter the target measure. Despite its importance, few studies have undertaken the attempt [Urban et al 2011; 10.1016/j.pain.2010.12.003]. The objective of this work is to correlate measures of pain after SCI to changes in physiological events of naturally‐behaving animals using non‐invasive home‐cage monitoring.All animal work conformed to Emory University Department of Animal Resources and FASEB guidelines. Animal home cages were instrumented with non‐contact electric potential integrated circuit sensors (Plessey Inc.) able to non‐invasively detect animal activity and respiratory variables from outside the home cage [Noble et al 2017; doi.org/10.1016/j.jneumeth.2016.12.007]. After three weeks of acclimation and baseline recordings, seven female adult c57/bl6 mice were recorded for 15 hours overnight for six weeks subsequent to a thoracic T10 spinal cord contusion injury (SCI; 50 kDyne impact). Hindpaw mechanical sensitivity was assessed with von Frey hairs using Chaplan's up‐down protocol. At 6 weeks post‐injury, SCI animals (n=7) and age‐matched naïve c57/bl6 mice (n=3) were euthanized, and in vitro electrophysiologic recordings of spontaneous afferent population activity were obtained from isolated lumbar dorsal root ganglia (DRG). Spinal cords and DRG were immersion fixed for subsequent histological assessment. Spontaneous activity was calculated as average firing rates over 5 trials (5 sec per trial). Data were analyzed using 1‐way ANOVAs and Tukey's post‐hoc test for significance. Linear regressions were calculated to assess correlation between data.Relative to baseline, SCI animals exhibited heightened mechanical sensitivity consistent with evoked allodynia at all time points except week 1 (p&lt;0.001, Fig 1A). SCI animals were less active after SCI (p&lt;0.034, Fig 1B). While there was no change in resting respiratory rate (RR, 3.6±1.4 Hz), there was an increase in RR variability at weeks 5 and 6 relative to baseline (p&lt;0.045, Fig 1C). The response for mechanical sensitivity and change in active time significantly correlated (R2=0.11, p=0.005) with a biphasic shape, exhibiting an acute response right after surgery then recovering to baseline levels before declining into a chronic response. Spontaneous DRG firing rates were higher in SCI animals than naïve animals (p=0.045, Fig 1D). With the removal of one outlying animal, spontaneous DRG activity strongly correlated with respiration rate variability (R2=0.87, p=0.0002) and with near significance to mechanical sensitivity (R2=0.42, p=0.055).In summary, contusion SCI leads to increased RR variability, which correlated to increased afferent spontaneous activity. SCI also reduced motor activity, which correlated to mechanical allodynia; These correlations may be causally linked. We hypothesize that ongoing spontaneous pain (i.e. increased spontaneous nociceptor activity) increases RR variability, and ongoing hindpaw mechanical allodynia reduces motor activity. While further studies are planned to more definitively link pain to alterations in physiological events in naturally‐behaving animals, it is clear that non‐invasive home‐cage monitoring can detect important consequences of SCI.Support or Funding InformationThis work was supported by the Craig H. Neilsen Foundation and NIH/NIGMS Institutional Research and Academic Career Development Award 5k12‐GM000680‐17This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Neuropathic Pain: Epidemiology, Classification, Mechanisms, and Therapy

Neuropathic pain develops because of lesions or disease affecting the somatosensory nervous system either in the periphery or centrally. Examples of neuropathic pain include painful polyneuropathy, post herpetic neuralgia, trigeminal neuralgia, and post-stroke pain. Clinically, neuropathic pain is characterized by spontaneous ongoing or shooting pain and evoked amplified pain responses after noxious or non-noxious stimuli. Methods such as questionnaires for screening and assessment focus on the presence and quality of neuropathic pain. Basic research is enabling the identification of different pathophysiological mechanisms, and clinical assessment of symptoms and signs can help to determine which mechanisms are involved in specific neuropathic pain disorders. Management of neuropathic pain requires an interdisciplinary approach, centered on pharmacological treatment. A better understanding of neuropathic pain and, in particular, of the translation of pathophysiological mechanisms into sensory signs will lead to a more effective and specific mechanism-based treatment approach.