Impaired amygdala astrocytic signaling worsens neuropathic pain-associated neuronal functions and behaviors.

Introduction: Pain is a clinically relevant health care issue with limited therapeutic options, creating the need for new and improved analgesic strategies. The amygdala is a limbic brain region critically involved in the regulation of emotional-affective components of pain and in pain modulation. The central nucleus of amygdala (CeA) serves major output functions and receives nociceptive information via the external lateral parabrachial nucleus (PB). While amygdala neuroplasticity has been linked causally to pain behaviors, non-neuronal pain mechanisms in this region remain to be explored. As an essential part of the neuroimmune system, astrocytes that represent about 40-50% of glia cells within the central nervous system, are required for physiological neuronal functions, but their role in the amygdala remains to be determined for pain conditions. In this study, we measured time-specific astrocyte activation in the CeA in a neuropathic pain model (spinal nerve ligation, SNL) and assessed the effects of astrocyte inhibition on amygdala neuroplasticity and pain-like behaviors in the pain condition. Methods and Results: Glial fibrillary acidic protein (GFAP, astrocytic marker) immunoreactivity and mRNA expression were increased at the chronic (4 weeks post-SNL), but not acute (1 week post-SNL), stage of neuropathic pain. In order to determine the contribution of astrocytes to amygdala pain-mechanisms, we used fluorocitric acid (FCA), a selective inhibitor of astrocyte metabolism. Whole-cell patch-clamp recordings were performed from neurons in the laterocapsular division of the CeA (CeLC) obtained from chronic neuropathic rats. Pre-incubation of brain slices with FCA (100 µM, 1 h), increased excitability through altered hyperpolarization-activated current (Ih) functions, without significantly affecting synaptic responses at the PB-CeLC synapse. Intra-CeA injection of FCA (100 µM) had facilitatory effects on mechanical withdrawal thresholds (von Frey and paw pressure tests) and emotional-affective behaviors (evoked vocalizations), but not on facial grimace score and anxiety-like behaviors (open field test), in chronic neuropathic rats. Selective inhibition of astrocytes by FCA was confirmed with immunohistochemical analyses showing decreased astrocytic GFAP, but not NeuN, signal in the CeA. Discussion: Overall, these results suggest a complex modulation of amygdala pain functions by astrocytes and provide evidence for beneficial functions of astrocytes in CeA in chronic neuropathic pain.

Ginger alleviates mechanical hypersensitivity and anxio-depressive behavior in rats with diabetic neuropathy through beneficial actions on gut microbiome composition, mitochondria, and neuroimmune cells of colon and spinal cord.

The relationship among gut microbiota, mitochondrial dysfunction/neuroinflammation, and diabetic neuropathic pain (DNP) has received increased attention. Ginger has antidiabetic and analgesic effects because of its anti-inflammatory property. We examined the effects of gingerols-enriched ginger (GEG) supplementation on pain-associated behaviors, gut microbiome composition, and mitochondrial function and neuroinflammation of colon and spinal cord in DNP rats. Thirty-three male rats were randomly divided into 3 groups: control group, DNP group (high-fat diet plus single dose of streptozotocin at 35 mg/kg body weight, and GEG group (DNP+GEG at 0.75% in the diet for 8 weeks). Von Frey and open field tests were used to assess pain sensitivity and anxio-depressive behaviors, respectively. Colon and spinal cord were collected for gene expression analysis. 16S rRNA gene sequencing was done from cecal samples and microbiome data analysis was performed using QIIME 2. GEG supplementation mitigated mechanical hypersensitivity and anxio-depressive behavior in DNP animals. GEG supplementation suppressed the dynamin-related protein 1 protein expression (colon) and gene expression (spinal cord), astrocytic marker GFAP gene expression (colon and spinal cord), and tumor necrosis factor-α gene expression (colon, P < .05; spinal cord, P = .0974) in DNP rats. GEG supplementation increased microglia/macrophage marker CD11b gene expression in colon and spinal cord of DNP rats. GEG treatment increased abundance of Acinetobacter, Azospirillum, Colidextribacter, and Fournierella but decreased abundance of Muribaculum intestinale in cecal feces of rats. This study demonstrates that GEG supplementation decreased pain, anxio-depression, and neuroimmune cells, and improved the composition of gut microbiomes and mitochondrial function in rats with diabetic neuropathy.

Associations Between Vitamin D Deficiency/Insufficiency and Depression Expose Health Disparities in Older Rural West Texans: A Project FRONTIER Study.

OBJECTIVE: To determine associations between Vitamin D (VD) levels and clinical depression through the Geriatric Depression Scale (GDS) and its questions and subdomains, stratified by demographics and Hispanic/Latino ethnicity (HLE). DESIGN, SETTING, AND PARTICIPANTS: A cohort of 299 Project FRONTIER participants aged 62.6 ± 11.7 years old, 70.9% female, and 40.5% HLE were used. Standard correlation and regression analyses were employed. MEASUREMENTS: The main outcome measures were VD (serum 25(OH)-VD) level, GDS-30 (30-item questionnaire), GDS-30 subfactors and questions, and HLE status. VD categories were defined as VD deficiency (VDD; ≤20 ng/mL), VD insufficiency (VDI; 21-29 ng/mL), VD sufficiency (30-38 ng/mL) and high VD sufficiency (>38 ng/mL). RESULTS: The majority (61.5%) of samples fell into VDD/VDI categories. A significant negative association was found between VD level and GDS-30 total score. VD level was negatively correlated with Dysphoria and Meaninglessness GDS-30 subfactors. Although GDS subfactors were similar between HLE and non-HLE groups, VD levels were significantly lower in HLE samples. Finally, HLE/non-HLE groups were differentially stratified across VD categories. Only 4% of HLEs fell into the high VD sufficient category, suggesting low VD supplementation. CONCLUSION: A significant negative association between VD level and depressive symptoms was revealed in our aging Project FRONTIER participants. HLE individuals were overrepresented in VDD/VDI samples, and VDD/VDI was associated primarily with the Dysphoria GDS subdomain. Regression analysis predicted high VD sufficiency (95.5 ng/mL) to be associated with no depressive symptoms (GDS=0). Our results underscore troubling disparities in VD-related depressive symptoms between HLE and non-HLE populations.

Alcohol withdrawal and pain: Peripheral mechanisms join central circuits.

Negative affective aspects of alcohol withdrawal and pain involve converging brain circuits. In this issue of Neuron, Son et al.1 identify a peripheral mechanism of an alcohol-withdrawal-induced headache-like condition, which is centered on mast-cell-specific receptor MrgprB2 activated by corticotropin-releasing factor (CRF) in dura mater to drive nociception.

A "double-edged" role for type-5 metabotropic glutamate receptors in pain disclosed by light-sensitive drugs.

Knowing the site of drug action is important to optimize effectiveness and address any side effects. We used light-sensitive drugs to identify the brain region-specific role of mGlu5 metabotropic glutamate receptors in the control of pain. Optical activation of systemic JF-NP-26, a caged, normally inactive, negative allosteric modulator (NAM) of mGlu5 receptors, in cingulate, prelimbic and infralimbic cortices and thalamus inhibited neuropathic pain hypersensitivity. Systemic treatment of alloswitch-1, an intrinsically active mGlu5 receptor NAM, caused analgesia, and the effect was reversed by light-induced drug inactivation in in the prelimbic and infralimbic cortices, and thalamus. This demonstrates that mGlu5 receptor blockade in the medial prefrontal cortex and thalamus is both sufficient and necessary for the analgesic activity of mGlu5 receptor antagonists. Surprisingly, when light was delivered in the basolateral amygdala, local activation of systemic JF-NP-26 reduced pain thresholds, whereas inactivation of alloswitch-1 enhanced analgesia. Electrophysiological analysis showed that alloswitch-1 increased excitatory synaptic responses in prelimbic pyramidal neurons evoked by stimulation of BLA input, and decreased feedforward inhibition of amygdala output neurons by BLA. Both effects were reversed by optical silencing and reinstated by optical reactivation of alloswitch-1. These findings demonstrate for the first time that the action of mGlu5 receptors in the pain neuraxis is not homogenous, and suggest that blockade of mGlu5 receptors in the BLA may limit the overall analgesic activity of mGlu5 receptor antagonists. This could explain the suboptimal effect of mGlu5 NAMs on pain in human studies and validate photopharmacology as an important tool to determine ideal target sites for systemic drugs.

Investigating the Relationship Between Marital Status and Ethnicity on Neurocognitive Functioning in a Rural Older Population: A Project FRONTIER Study.

OBJECTIVES: Research indicates being married is related to better physical and psychological health. Little is known regarding the relationship between marital status and neurocognitive functioning and whether it differs based on ethnicity (Hispanic vs non-Hispanic). This is the first study to examine this relationship in a sample of aging adults in rural Texas. METHODS: Data from 1,864 participants (Mage = 59.68, standard deviation [SD]age = 12.21), who were mostly Hispanic (n = 1,053), women (n = 1,295), and married (n = 1,125) from Project Facing Rural Obstacles to Healthcare Now Through Intervention, Education, & Research were analyzed. Neuropsychological testing comprised Repeatable Battery for the Assessment of Neuropsychological Status, Trails Making Test, and Clock Drawing. Participants were dichotomized, married, and unmarried. RESULTS: There was a significant interaction between Hispanic identity and marital status on overall neurocognitive functioning (F(1, 1,480) = 4.79, p < .05, ηp2 = 0.003). For non-Hispanic individuals, married individuals had higher overall neurocognitive functioning compared to unmarried individuals, whereas neurocognitive functioning for Hispanic individuals did not significantly differ between married and unmarried individuals. There were significant main effects as married individuals (M = 84.95, SD = 15.56) had greater overall neurocognitive functioning than unmarried individuals (M = 83.47, SD = 15.86; F(1, 1,480) = 14.67, p < .001, ηp2 = 0.01), Hispanic individuals (M = 78.02, SD = 14.25) had lower overall neurocognitive functioning than non-Hispanic individuals (M = 91.43, SD = 15.07; F(1, 1,480) = 284.99, p < .001, ηp2 = 0.16). DISCUSSION: Hispanics living in rural areas experience additional stressors that could lead to worse neurocognitive functioning, which is supported by the Lifespan Biopsychosocial Model of Cumulative Vulnerability and Minority Health, which postulates that race/ethnicity/socioeconomic-status-related stressors exacerbate the impact of other life stressors. Reduction of stress on rural Hispanics should be a priority as it could positively affect their neurocognitive functioning.

Opposing Effects on Descending Control of Nociception by µ and κ Opioid Receptors in the Anterior Cingulate Cortex.

BACKGROUND: The efficiency of descending pain modulation, commonly assessed with the conditioned pain modulation procedure, is diminished in patients with chronic pain. The authors hypothesized that the efficiency of pain modulation is controlled by cortical opioid circuits. METHODS: This study evaluated the effects of µ opioid receptor activation in the anterior cingulate cortex on descending control of nociception, a preclinical correlate of conditioned pain modulation, in male Sprague-Dawley rats with spinal nerve ligation-induced chronic pain or in sham-operated controls. Additionally, the study explored the consequences of respective activation or inhibition of κ opioid receptor in the anterior cingulate cortex of naive rats or animals with neuropathic pain. Descending control of nociception was measured as the hind paw withdrawal response to noxious pressure (test stimulus) in the absence or presence of capsaicin injection in the forepaw (conditioning stimulus). RESULTS: Descending control of nociception was diminished in the ipsilateral, but not contralateral, hind paw of rats with spinal nerve ligation. Bilateral administration of morphine in the anterior cingulate cortex had no effect in shams but restored diminished descending control of nociception without altering hypersensitivity in rats with neuropathic pain. Bilateral anterior cingulate cortex microinjection of κ opioid receptor antagonists, including nor-binaltorphimine and navacaprant, also re-established descending control of nociception in rats with neuropathic pain without altering hypersensitivity and with no effect in shams. Conversely, bilateral injection of a κ opioid receptor agonist, U69,593, in the anterior cingulate cortex of naive rats inhibited descending control of nociception without altering withdrawal thresholds. CONCLUSIONS: Anterior cingulate cortex κ opioid receptor activation therefore diminishes descending control of nociception both in naive animals and as an adaptive response to chronic pain, likely by enhancing net descending facilitation. Descending control of nociception can be restored by activation of µ opioid receptors in the anterior cingulate cortex, but also by κ opioid receptor antagonists, providing a nonaddictive alternative to opioid analgesics. Navacaprant is now in advanced clinical trials.

Turmeric Bioactive Compounds Alleviate Spinal Nerve Ligation-Induced Neuropathic Pain by Suppressing Glial Activation and Improving Mitochondrial Function in Spinal Cord and Amygdala.

This study examined the effects of turmeric bioactive compounds, curcumin C3 complex® (CUR) and bisdemethoxycurcumin (BDMC), on mechanical hypersensitivity and the gene expression of markers for glial activation, mitochondrial function, and oxidative stress in the spinal cord and amygdala of rats with neuropathic pain (NP). Twenty-four animals were randomly assigned to four groups: sham, spinal nerve ligation (SNL, an NP model), SNL+100 mg CUR/kg BW p.o., and SNL+50 mg BDMC/kg BW p.o. for 4 weeks. Mechanical hypersensitivity was assessed by the von Frey test (VFT) weekly. The lumbosacral section of the spinal cord and the right amygdala (central nucleus) were collected to determine the mRNA expression of genes (IBA-1, CD11b, GFAP, MFN1, DRP1, FIS1, PGC1α, PINK, Complex I, TLR4, and SOD1) utilizing qRT-PCR. Increased mechanical hypersensitivity and increased gene expression of markers for microglial activation (IBA-1 in the amygdala and CD11b in the spinal cord), astrocyte activation (GFAP in the spinal cord), mitochondrial dysfunction (PGC1α in the amygdala), and oxidative stress (TLR4 in the spinal cord and amygdala) were found in untreated SNL rats. Oral administration of CUR and BDMC significantly decreased mechanical hypersensitivity. CUR decreased CD11b and GFAP gene expression in the spinal cord. BDMC decreased IBA-1 in the spinal cord and amygdala as well as CD11b and GFAP in the spinal cord. Both CUR and BDMC reduced PGC1α gene expression in the amygdala, PINK1 gene expression in the spinal cord, and TLR4 in the spinal cord and amygdala, while they increased Complex I and SOD1 gene expression in the spinal cord. CUR and BDMC administration decreased mechanical hypersensitivity in NP by mitigating glial activation, oxidative stress, and mitochondrial dysfunction.

Tai Chi exercise reduces circulating levels of inflammatory oxylipins in postmenopausal women with knee osteoarthritis: results from a pilot study.

Background: Tai Chi (TC) controls pain through mind-body exercise and appears to alter inflammatory mediators. TC actions on lipid biomarkers associated with inflammation and brain neural networks in women with knee osteoarthritic pain were investigated. Methods: A single-center, pre- and post-TC group (baseline and 8 wk) exercise pilot study in postmenopausal women with knee osteoarthritic pain was performed. 12 eligible women participated in TC group exercise. The primary outcome was liquid chromatography tandem mass spectrometry determination of circulating endocannabinoids (eCB) and oxylipins (OxL). Secondary outcomes were correlations between eCB and OxL levels and clinical pain/limitation assessments, and brain resting-state function magnetic resonance imaging (rs-fMRI). Results: Differences in circulating quantitative levels (nM) of pro-inflammatory OxL after TC were found in women. TC exercise resulted in lower OxL PGE1 and PGE2 and higher 12-HETE, LTB4, and 12-HEPE compared to baseline. Pain assessment and eCB and OxL levels suggest crucial relationships between TC exercise, inflammatory markers, and pain. Higher plasma levels of eCB AEA, and 1, 2-AG were found in subjects with increased pain. Several eCB and OxL levels were positively correlated with left and right brain amygdala-medial prefrontal cortex functional connectivity. Conclusion: TC exercise lowers pro-inflammatory OxL in women with knee osteoarthritic pain. Correlations between subject pain, functional limitations, and brain connectivity with levels of OxL and eCB showed significance. Findings indicate potential mechanisms for OxL and eCB and their biosynthetic endogenous PUFA precursors that alter brain connectivity, neuroinflammation, and pain. Clinical Trial Registration: ClinicalTrials.gov, identifier: NCT04046003.

Pain and aging: A unique challenge in neuroinflammation and behavior.

Chronic pain is one of the most common, costly, and potentially debilitating health issues facing older adults, with attributable costs exceeding $600 billion annually. The prevalence of pain in humans increases with advancing age. Yet, the contributions of sex differences, age-related chronic inflammation, and changes in neuroplasticity to the overall experience of pain are less clear, given that opposing processes in aging interact. This review article examines and summarizes pre-clinical research and clinical data on chronic pain among older adults to identify knowledge gaps and provide the base for future research and clinical practice. We provide evidence to suggest that neurodegenerative conditions engender a loss of neural plasticity involved in pain response, whereas low-grade inflammation in aging increases CNS sensitization but decreases PNS sensitivity. Insights from preclinical studies are needed to answer mechanistic questions. However, the selection of appropriate aging models presents a challenge that has resulted in conflicting data regarding pain processing and behavioral outcomes that are difficult to translate to humans.

Hmgb1 Silencing in the Amygdala Inhibits Pain-Related Behaviors in a Rat Model of Neuropathic Pain.

Chronic pain presents a therapeutic challenge due to the highly complex interplay of sensory, emotional-affective and cognitive factors. The mechanisms of the transition from acute to chronic pain are not well understood. We hypothesized that neuroimmune mechanisms in the amygdala, a brain region involved in the emotional-affective component of pain and pain modulation, play an important role through high motility group box 1 (Hmgb1), a pro-inflammatory molecule that has been linked to neuroimmune signaling in spinal nociception. Transcriptomic analysis revealed an upregulation of Hmgb1 mRNA in the right but not left central nucleus of the amygdala (CeA) at the chronic stage of a spinal nerve ligation (SNL) rat model of neuropathic pain. Hmgb1 silencing with a stereotaxic injection of siRNA for Hmgb1 into the right CeA of adult male and female rats 1 week after (post-treatment), but not 2 weeks before (pre-treatment) SNL induction decreased mechanical hypersensitivity and emotional-affective responses, but not anxiety-like behaviors, measured 4 weeks after SNL. Immunohistochemical data suggest that neurons are a major source of Hmgb1 in the CeA. Therefore, Hmgb1 in the amygdala may contribute to the transition from acute to chronic neuropathic pain, and the inhibition of Hmgb1 at a subacute time point can mitigate neuropathic pain.

Studies on diketopiperazine and dipeptide analogs as opioid receptor ligands.

Using the structure of gliotoxin as a starting point, we have prepared two different chemotypes with selective affinity to the kappa opioid receptor (KOR). Using medicinal chemistry approaches and structure-activity relationship (SAR) studies, structural features required for the observed affinity were identified, and advanced molecules with favorable Multiparameter Optimization (MPO) and Ligand Lipophilicity (LLE) profiles were prepared. Using the Thermal Place Preference Test (TPPT), we have shown that compound2 blocks the antinociceptive effect of U50488, a known KOR agonist. Multiple reports suggest that modulation of KOR signaling is a promising therapeutic strategy in treating neuropathic pain (NP). As a proof-of-concept study, we tested compound 2 in a rat model of NP and recorded its ability to modulate sensory and emotional pain-related behaviors. Observed in vitro and in vivo results suggest that these ligands can be used to develop compounds with potential application as pain therapeutics.

Pain-related cortico-limbic plasticity and opioid signaling.

Neuroplasticity in cortico-limbic circuits has been implicated in pain persistence and pain modulation in clinical and preclinical studies. The amygdala has emerged as a key player in the emotional-affective dimension of pain and pain modulation. Reciprocal interactions with medial prefrontal cortical regions undergo changes in pain conditions. Other limbic and paralimbic regions have been implicated in pain modulation as well. The cortico-limbic system is rich in opioids and opioid receptors. Preclinical evidence for their pain modulatory effects in different regions of this highly interactive system, potentially opposing functions of different opioid receptors, and knowledge gaps will be described here. There is little information about cell type- and circuit-specific functions of opioid receptor subtypes related to pain processing and pain-related plasticity in the cortico-limbic system. The important role of anterior cingulate cortex (ACC) and amygdala in MOR-dependent analgesia is most well-established, and MOR actions in the mesolimbic system appear to be similar but remain to be determined in mPFC regions other than ACC. Evidence also suggests that KOR signaling generally serves opposing functions whereas DOR signaling in the ACC has similar, if not synergistic effects, to MOR. A unifying picture of pain-related neuronal mechanisms of opioid signaling in different elements of the cortico-limbic circuitry has yet to emerge. This article is part of the Special Issue on "Opioid-induced changes in addiction and pain circuits".

Kappa opioid receptor agonists produce sexually dimorphic and prolactin-dependent hyperalgesic priming.

ABSTRACT: Repeated stress produces hyperalgesic priming in preclinical models, but underlying mechanisms remain uncertain. As stress engages kappa opioid receptors (KORs), we hypothesized that repeated administration of KOR agonists might mimic, in part, stress-induced hyperalgesic priming. The potential contribution of circulating prolactin (PRL) and dysregulation of the expression of PRL receptor (PRLR) isoforms in sensory neurons after KOR agonist administration was also investigated. Mice received 3 daily doses of U-69593 or nalfurafine as a "first-hit" stimulus followed by assessment of periorbital tactile allodynia. Sixteen days after the first KOR agonist administration, animals received a subthreshold dose of inhalational umbellulone, a TRPA1 agonist, as the second-hit stimulus and periorbital allodynia was assessed. Cabergoline, a dopamine D2 receptor agonist, was used to inhibit circulating PRL in additional cohorts. Prolactin receptor isoforms were quantified in the V1 region of the trigeminal ganglion after repeated doses of U-69593. In both sexes, KOR agonists increased circulating PRL and produced allodynia that resolved within 14 days. Hyperalgesic priming, revealed by umbellulone-induced allodynia in animals previously treated with the KOR agonists, also occurred in both sexes. However, repeated U-69593 downregulated the PRLR long isoform in trigeminal neurons only in female mice. Umbellulone-induced allodynia was prevented by cabergoline co-treatment during priming with KOR agonists in female, but not male, mice. Hyperalgesic priming therefore occurs in both sexes after either biased or nonbiased KOR agonists. However, a PRL/PRLR-dependence is observed only in female nociceptors possibly contributing to pain in stress-related pain disorders in females.

Chronic pain recruits hypothalamic dynorphin/kappa opioid receptor signalling to promote wakefulness and vigilance.

Increased vigilance in settings of potential threats or in states of vulnerability related to pain is important for survival. Pain disrupts sleep and conversely, sleep disruption enhances pain, but the underlying mechanisms remain unknown. Chronic pain engages brain stress circuits and increases secretion of dynorphin, an endogenous ligand of the kappa opioid receptor (KOR). We therefore hypothesized that hypothalamic dynorphin/KOR signalling may be a previously unknown mechanism that is recruited in pathological conditions requiring increased vigilance. We investigated the role of KOR in wakefulness, non-rapid eye movement (NREM) sleep and rapid eye movement (REM) sleep in freely moving naïve mice and in mice with neuropathic pain induced by partial sciatic nerve ligation using EEG/EMG recordings. Systemic continuous administration of U69,593, a KOR agonist, over 5 days through an osmotic minipump decreased the amount of NREM and REM sleep and increased sleep fragmentation in naïve mice throughout the light-dark sleep cycle. We used KORcre mice to selectively express a Gi-coupled designer receptor activated by designer drugs (Gi-DREADD) in KORcre neurons of the hypothalamic paraventricular nucleus, a key node of the hypothalamic-pituitary-adrenal stress response. Sustained activation of Gi-DREADD with clozapine-N-oxide delivered in drinking water over 4 days, disrupted sleep in these mice in a similar way as systemic U69,593. Mice with chronic neuropathic pain also showed disrupted NREM and total sleep that was normalized by systemic administration of two structurally different KOR antagonists, norbinaltorphimine and NMRA-140, currently in phase II clinical development, or by CRISPR/Cas9 editing of paraventricular nucleus KOR, consistent with endogenous KOR activation disrupting sleep in chronic pain. Unexpectedly, REM sleep was diminished by either systemic KOR antagonist or by CRISPR/Cas9 editing of paraventricular nucleus KOR in sham-operated mice. Our findings reveal previously unknown physiological and pathophysiological roles of dynorphin/KOR in eliciting arousal. Physiologically, dynorphin/KOR signalling affects transitions between sleep stages that promote REM sleep. Furthermore, while KOR antagonists do not promote somnolence in the absence of pain, they normalized disrupted sleep in chronic pain, revealing a pathophysiological role of KOR signalling that is selectively recruited to promote vigilance, increasing chances of survival. Notably, while this mechanism is likely beneficial in the short-term, disruption of the homeostatic need for sleep over longer periods may become maladaptive resulting in sustained pain chronicity. A novel approach for treatment of chronic pain may thus result from normalization of chronic pain-related sleep disruption by KOR antagonism.

Recent Advances in the Modulation of Pain by the Metabotropic Glutamate Receptors.

Metabotropic glutamate receptors (mGluR or mGlu) are G-protein coupled receptors activated by the binding of glutamate, the main classical neurotransmitter of the nervous system. Eight different mGluR subtypes (mGluR1-8) have been cloned and are classified in three groups based on their molecular, pharmacological and signaling properties. mGluRs mediate several physiological functions such as neuronal excitability and synaptic plasticity, but they have also been implicated in numerous pathological conditions including pain. The availability of new and more selective allosteric modulators together with the canonical orthosteric ligands and transgenic technologies has led to significant advances in our knowledge about the role of the specific mGluR subtypes in the pathophysiological mechanisms of various diseases. Although development of successful compounds acting on mGluRs for clinical use has been scarce, the subtype-specific-pharmacological manipulation might be a compelling approach for the treatment of several disorders in humans, including pain; this review aims to summarize and update on preclinical evidence for the roles of different mGluRs in the pain system and discusses knowledge gaps regarding mGluR-related sex differences and neuroimmune signaling in 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.

Kappa Opioid Receptor Blockade in the Amygdala Mitigates Pain Like-Behaviors by Inhibiting Corticotropin Releasing Factor Neurons in a Rat Model of Functional Pain.

Functional pain syndromes (FPS) occur in the absence of identifiable tissue injury or noxious events and include conditions such as migraine, fibromyalgia, and others. Stressors are very common triggers of pain attacks in various FPS conditions. It has been recently demonstrated that kappa opioid receptors (KOR) in the central nucleus of amygdala (CeA) contribute to FPS conditions, but underlying mechanisms remain unclear. The CeA is rich in KOR and encompasses major output pathways involving extra-amygdalar projections of corticotropin releasing factor (CRF) expressing neurons. Here we tested the hypothesis that KOR blockade in the CeA in a rat model of FPS reduces pain-like and nocifensive behaviors by restoring inhibition of CeA-CRF neurons. Intra-CeA administration of a KOR antagonist (nor-BNI) decreased mechanical hypersensitivity and affective and anxiety-like behaviors in a stress-induced FPS model. In systems electrophysiology experiments in anesthetized rats, intra-CeA application of nor-BNI reduced spontaneous firing and responsiveness of CeA neurons to peripheral stimulation. In brain slice whole-cell patch-clamp recordings, nor-BNI increased feedforward inhibitory transmission evoked by optogenetic and electrical stimulation of parabrachial afferents, but had no effect on monosynaptic excitatory transmission. Nor-BNI decreased frequency, but not amplitude, of spontaneous inhibitory synaptic currents, suggesting a presynaptic action. Blocking KOR receptors in stress-induced FPS conditions may therefore represent a novel therapeutic strategy.

Sex Differences in CGRP Regulation and Function in the Amygdala in a Rat Model of Neuropathic Pain.

The amygdala has emerged as a key player in the emotional response to pain and pain modulation. The lateral and capsular regions of the central nucleus of the amygdala (CeA) represent the "nociceptive amygdala" due to their high content of neurons that process pain-related information. These CeA divisions are the targets of the spino-parabrachio-amygdaloid pain pathway, which is the predominant source of calcitonin gene-related peptide (CGRP) within the amygdala. Changes in lateral and capsular CeA neurons have previously been observed in pain models, and synaptic plasticity in these areas has been linked to pain-related behavior. CGRP has been demonstrated to play an important role in peripheral and spinal mechanisms, and in pain-related amygdala plasticity in male rats in an acute arthritis pain model. However, the role of CGRP in chronic neuropathic pain-related amygdala function and behaviors remains to be determined for both male and female rats. Here we tested the hypothesis that the CGRP1 receptor is involved in neuropathic pain-related amygdala activity, and that blockade of this receptor can inhibit neuropathic pain behaviors in both sexes. CGRP mRNA expression levels in the CeA of male rats were upregulated at the acute stage of the spinal nerve ligation (SNL) model of neuropathic pain, whereas female rats had significantly higher CGRP and CGRP receptor component expression at the chronic stage. A CGRP1 receptor antagonist (CGRP 8-37) administered into the CeA in chronic neuropathic rats reduced mechanical hypersensitivity (von Frey and paw compression tests) in both sexes but showed female-predominant effects on emotional-affective responses (ultrasonic vocalizations) and anxiety-like behaviors (open field test). CGRP 8-37 inhibited the activity of CeA output neurons assessed with calcium imaging in brain slices from chronic neuropathic pain rats. Together, these findings may suggest that CGRP1 receptors in the CeA are involved in neuropathic pain-related amygdala activity and contribute to sensory aspects in both sexes but to emotional-affective pain responses predominantly in females. The sexually dimorphic function of CGRP in the amygdala would make CGRP1 receptors a potential therapeutic target for neuropathic pain relief, particularly in females in chronic pain conditions.