Microglial STING activation alleviates nerve injury-induced neuropathic pain in male but not female mice.

Microglia, resident immune cells in the central nervous system, play a role in neuroinflammation and the development of neuropathic pain. We found that the stimulator of interferon genes (STING) is predominantly expressed in spinal microglia and upregulated after peripheral nerve injury. However, mechanical allodynia, as a marker of neuropathic pain following peripheral nerve injury, did not require microglial STING expression. In contrast, STING activation by specific agonists (ADU-S100, 35 nmol) significantly alleviated neuropathic pain in male mice, but not female mice. STING activation in female mice leads to increase in proinflammatory cytokines that may counteract the analgesic effect of ADU-S100. Microglial STING expression and type I interferon-ß (IFN-ß) signaling were required for the analgesic effects of STING agonists in male mice. Mechanistically, downstream activation of TANK-binding kinase 1 (TBK1) and the production of IFN-ß, may partly account for the analgesic effect observed. These findings suggest that STING activation in spinal microglia could be a potential therapeutic intervention for neuropathic pain, particularly in males.

STING recognition of viral dsDNA by nociceptors mediates pain in mice.

Pain is often one of the initial indicators of a viral infection, yet our understanding of how viruses induce pain is limited. Immune cells typically recognize viral nucleic acids, which activate viral receptors and signaling, leading to immunity. Interestingly, these viral receptors and signals are also present in nociceptors and are associated with pain. Here, we investigate the response of nociceptors to nucleic acids during viral infections, specifically focusing on the role of the viral signal, Stimulator of Interferon Genes (STING). Our research shows that cytosolic double-stranded DNA (dsDNA) from viruses, like herpes simplex virus 1 (HSV-1), triggers pain responses through STING expression in nociceptors. In addition, STING agonists alone can elicit pain responses. Notably, these responses involve the direct activation of STING in nociceptors through TRPV1. We also provided a proof-of-concept showing that STING and TRPV1 significantly contribute to the mechanical hypersensitivity induced by HSV-1 infection. These findings suggest that STING could be a potential therapeutic target for relieving pain during viral infections.

The Anti-Inflammatory Agent Bindarit Attenuates the Impairment of Neural Development through Suppression of Microglial Activation in a Neonatal Hydrocephalus Mouse Model.

Neonatal hydrocephalus presents with various degrees of neuroinflammation and long-term neurologic deficits in surgically treated patients, provoking a need for additional medical treatment. We previously reported elevated neuroinflammation and severe periventricular white matter damage in the progressive hydrocephalus (prh) mutant which contains a point mutation in the Ccdc39 gene, causing loss of cilia-mediated unidirectional CSF flow. In this study, we identified cortical neuropil maturation defects such as impaired excitatory synapse maturation and loss of homeostatic microglia, and swimming locomotor defects in early postnatal prh mutant mice. Strikingly, systemic application of the anti-inflammatory small molecule bindarit significantly supports healthy postnatal cerebral cortical development in the prh mutant. While bindarit only mildly reduced the ventricular volume, it significantly improved the edematous appearance and myelination of the corpus callosum. Moreover, the treatment attenuated thinning in cortical Layers II-IV, excitatory synapse formation, and interneuron morphogenesis, by supporting the ramified-shaped homeostatic microglia from excessive cell death. Also, the therapeutic effect led to the alleviation of a spastic locomotor phenotype of the mutant. We found that microglia, but not peripheral monocytes, contribute to amoeboid-shaped activated myeloid cells in prh mutants' corpus callosum and the proinflammatory cytokines expression. Bindarit blocks nuclear factor (NF)-kB activation and its downstream proinflammatory cytokines, including monocyte chemoattractant protein-1, in the prh mutant. Collectively, we revealed that amelioration of neuroinflammation is crucial for white matter and neuronal maturation in neonatal hydrocephalus. Future studies of bindarit treatment combined with CSF diversion surgery may provide long-term benefits supporting neuronal development in neonatal hydrocephalus.SIGNIFICANCE STATEMENT In neonatal hydrocephalus, little is known about the signaling cascades of neuroinflammation or the impact of such inflammatory insults on neural cell development within the perinatal cerebral cortex. Here, we report that proinflammatory activation of myeloid cells, the majority of which are derived from microglia, impairs periventricular myelination and cortical neuronal maturation using the mouse prh genetic model of neonatal hydrocephalus. Administration of bindarit, an anti-inflammatory small molecule that blocks nuclear factor (NF)-kB activation, restored the cortical thinning and synaptic maturation defects in the prh mutant brain through suppression of microglial activation. These data indicate the potential therapeutic use of anti-inflammatory reagents targeting neuroinflammation in the treatment of neonatal hydrocephalus.

Single-cell analysis of dorsal root ganglia reveals metalloproteinase signaling in satellite glial cells and pain.

Satellite glial cells (SGCs) are among the most abundant non-neuronal cells in dorsal root ganglia (DRGs) and closely envelop sensory neurons that detect painful stimuli. However, little is still known about their homeostatic activities and their contribution to pain. Using single-cell RNA sequencing (scRNA-seq), we were able to obtain a unique transcriptional profile for SGCs. We found enriched expression of the tissue inhibitor metalloproteinase 3 (TIMP3) and other metalloproteinases in SGCs. Small interfering RNA and neutralizing antibody experiments revealed that TIMP3 modulates somatosensory stimuli. TIMP3 expression decreased after paclitaxel treatment, and its rescue by delivery of a recombinant TIMP3 protein reversed and prevented paclitaxel-induced pain. We also established that paclitaxel directly impacts metalloproteinase signaling in cultured SGCs, which may be used to identify potential new treatments for pain. Therefore, our results reveal a metalloproteinase signaling pathway in SGCs for proper processing of somatosensory stimuli and potential discovery of novel pain treatments.

Targeting Nociceptive Neurons and Transient Receptor Potential Channels for the Treatment of Migraine.

Migraine is a neurovascular disorder that affects approximately 12% of the global population. While its exact causes are still being studied, researchers believe that nociceptive neurons in the trigeminal ganglia play a key role in the pain signals of migraine. These nociceptive neurons innervate the intracranial meninges and convey pain signals from the meninges to the thalamus. Targeting nociceptive neurons is considered promising due to their accessibility and distinct molecular profile, which includes the expression of several transient receptor potential (TRP) channels. These channels have been linked to various pain conditions, including migraine. This review discusses the role and mechanisms of nociceptive neurons in migraine, the challenges of current anti-migraine drugs, and the evidence for well-studied and emerging TRP channels, particularly TRPC4, as novel targets for migraine prevention and treatment.

Venom Peptide Toxins Targeting the Outer Pore Region of Transient Receptor Potential Vanilloid 1 in Pain: Implications for Analgesic Drug Development.

The transient receptor potential vanilloid 1 (TRPV1) ion channel plays an important role in the peripheral nociceptive pathway. TRPV1 is a polymodal receptor that can be activated by multiple types of ligands and painful stimuli, such as noxious heat and protons, and contributes to various acute and chronic pain conditions. Therefore, TRPV1 is emerging as a novel therapeutic target for the treatment of various pain conditions. Notably, various peptides isolated from venomous animals potently and selectively control the activation and inhibition of TRPV1 by binding to its outer pore region. This review will focus on the mechanisms by which venom-derived peptides interact with this portion of TRPV1 to control receptor functions and how these mechanisms can drive the development of new types of analgesics.

Key role of CCR2-expressing macrophages in a mouse model of low back pain and radiculopathy.

Chronic low back pain is a common condition, with high societal costs and often ineffectual treatments. Communication between macrophages/monocytes (MØ) and sensory neurons has been implicated in various preclinical pain models. However, few studies have examined specific MØ subsets, although distinct subtypes may play opposing roles. This study used a model of low back pain/radiculopathy involving direct local inflammation of the dorsal root ganglia (DRG). Reporter mice were employed that had distinct fluorescent labels for two key MØ subsets: CCR2-expressing (infiltrating pro-inflammatory) MØ, and CX3CR1-expressing (resident) macrophages. We observed that local DRG inflammation induced pain behaviors in mice, including guarding behavior and mechanical hypersensitivity, similar to the previously described rat model. The increase in MØ in the inflamed DRG was dominated by increases in CCR2+ MØ, which persisted for at least 14 days. The primary endogenous ligand for CCR2, CCL2, was upregulated in inflamed DRG. Three different experimental manipulations that reduced the CCR2+ MØ influx also reduced pain behaviors: global CCR2 knockout; systemic injection of INCB3344 (specific CCR2 blocker); and intravenous injection of liposomal clodronate. The latter two treatments when applied around the time of DRG inflammation reduced CCR2+ but not CX3CR1+ MØ in the DRG. Together these experiments suggest a key role for the CCR2/CCL2 system in establishing the pain state in this model of inflammatory low back pain and radiculopathy. Intravenous clodronate given after pain was established had the opposite effect on pain behaviors, suggesting the role of macrophages or their susceptibility to clodronate may change with time.

The inhibition of Kir2.1 potassium channels depolarizes spinal microglial cells, reduces their proliferation, and attenuates neuropathic pain.

Spinal microglia change their phenotype and proliferate after nerve injury, contributing to neuropathic pain. For the first time, we have characterized the electrophysiological properties of microglia and the potential role of microglial potassium channels in the spared nerve injury (SNI) model of neuropathic pain. We observed a strong increase of inward currents restricted at 2 days after injury associated with hyperpolarization of the resting membrane potential (RMP) in microglial cells compared to later time-points and naive animals. We identified pharmacologically and genetically the current as being mediated by Kir2.1 ion channels whose expression at the cell membrane is increased 2 days after SNI. The inhibition of Kir2.1 with ML133 and siRNA reversed the RMP hyperpolarization and strongly reduced the currents of microglial cells 2 days after SNI. These electrophysiological changes occurred coincidentally to the peak of microglial proliferation following nerve injury. In vitro, ML133 drastically reduced the proliferation of BV2 microglial cell line after both 2 and 4 days in culture. In vivo, the intrathecal injection of ML133 significantly attenuated the proliferation of microglia and neuropathic pain behaviors after nerve injury. In summary, our data implicate Kir2.1-mediated microglial proliferation as an important therapeutic target in neuropathic pain.

Local Sympathectomy Promotes Anti-inflammatory Responses and Relief of Paclitaxel-induced Mechanical and Cold Allodynia in Mice.

BACKGROUND: Patients undergoing cancer treatment often experience chemotherapy-induced neuropathic pain at their extremities, for which there is no U.S. Food and Drug Administration-approved drug. The authors hypothesized that local sympathetic blockade, which is used in the clinic to treat various pain conditions, can also be effective to treat chemotherapy-induced neuropathic pain. METHODS: A local sympathectomy (i.e., cutting the ipsilateral gray rami entering the spinal nerves near the L3 and L4 dorsal root ganglia) was performed in mice receiving intraperitoneal injections every other day of the chemotherapeutic drug paclitaxel. Sympathectomy effects were then assessed in chemotherapy-induced pain-like behaviors (i.e., mechanical and cold allodynia) and neuroimmune and electrophysiologic responses. RESULTS: Local microsympathectomy produced a fast recovery from mechanical allodynia (mean ± SD: sympathectomy vs. sham at day 5, 1.07 ± 0.34 g vs. 0.51 ± 0.17g, n = 5, P = 0.030 in male mice, and 1.08 ± 0.28 g vs. 0.62 ± 0.16 g, n = 5, P = 0.036 in female mice) and prevented the development of cold allodynia in both male and female mice after paclitaxel. Mechanistically, microsympathectomy induced transcriptional increases in dorsal root ganglia of macrophage markers and anti-inflammatory cytokines, such as the transforming growth factor-ß. Accordingly, depletion of monocytes/macrophages and blockade of transforming growth factor-ß signaling reversed the relief of mechanical allodynia by microsympathectomy. In particular, exogenous transforming growth factor-ß was sufficient to relieve mechanical allodynia after paclitaxel (transforming growth factor-ß 100 ng/site vs. vehicle at 3 h, 1.21 ± 0.34g vs. 0.53 ± 0.14 g, n = 5, P = 0.001 in male mice), and transforming growth factor-ß signaling regulated neuronal activity in dorsal root ganglia. CONCLUSIONS: Local sympathetic nerves control the progression of immune responses in dorsal root ganglia and pain-like behaviors in mice after paclitaxel, raising the possibility that clinical strategies already in use for local sympathetic blockade may also offer an effective treatment for patients experiencing chemotherapy-induced neuropathic pain.

Paclitaxel-activated astrocytes produce mechanical allodynia in mice by releasing tumor necrosis factor-α and stromal-derived cell factor 1.

BACKGROUND: Paclitaxel is a widely used and potent chemotherapeutic agent for the treatment of cancer. However, patients receiving paclitaxel often develop an acute pain syndrome for which there are few treatment options. Astrocytes play an important role in the pathogenesis of pain in multiple preclinical models, as well as in paclitaxel-treated rodents. However, it is still unclear what the exact contribution of astrocytes may be in paclitaxel-associated acute pain syndrome (P-APS). METHODS: P-APS was modeled by a single systemic or intrathecal injection of paclitaxel and astrocyte contribution tested by immunohistochemical, pharmacological, and behavioral approaches. Cell cultures were also prepared to assess whether paclitaxel treatment directly activates astrocytes and whether intrathecal injection of paclitaxel-treated astrocytes produces pain that is reminiscent of P-APS. RESULTS: Systemic injection of paclitaxel resulted in increased expression of glial fibrillary acidic protein (a common marker of astrocytic activation), as well as both systemic or intrathecal injection of paclitaxel induced pain hypersensitivity indicated by the development of mechanical allodynia, which was significantly reversed by the astrocytic inhibitor L-α-AA. Cultured astrocytes were activated by paclitaxel with significant increases in protein levels for tumor necrosis factor-α (TNF-α) and stromal-derived cell factor 1 (SDF-1). Importantly, intrathecal injection of paclitaxel-activated astrocytes produced mechanical allodynia that was reversed by TNF-α and SDF-1 neutralizing antibodies. CONCLUSION: Our results suggest for the first time that paclitaxel can directly activate astrocytes, which are sufficient to produce acute pain by releasing TNF-α and SDF-1. Targeting astrocytes and these cytokines may offer new treatments for P-APS.

5-Hydroxymethylcytosine (5hmC) and Ten-eleven translocation 1-3 (TET1-3) proteins in the dorsal root ganglia of mouse: Expression and dynamic regulation in neuropathic pain.

Epigenetic mechanisms are increasingly implicated in chronic pain pathology. In this study, we demonstrate that the novel epigenetic mark 5-hydroxymethylcytosine (5hmC) is present in dorsal root ganglia (DRG) neurons and glia, and its levels increase following nerve injury. Furthermore, we show that the 5hmC-generating Ten-eleven translocation 1-3 (TET1-3) proteins are expressed in a cell-type specific manner in the DRG, with Tet3 displaying differential upregulation after injury, suggesting a potential role in neuropathic pain.

Unconventional Role of Caspase-6 in Spinal Microglia Activation and Chronic Pain.

Chronic pain affects ~20% of the worldwide population. The clinical management of chronic pain is mostly palliative and results in limited success. Current treatments mostly target the symptoms or neuronal signaling of chronic pain. It has been increasingly recognized that glial cells, such as microglia, and inflammatory signaling play a major role in the pathogenesis of chronic pain. Caspases (CASPs) are a family of protease enzymes involved in apoptosis and inflammation. They are pivotal components in a variety of neurological diseases. However, little is known about the role of CASPs in microglial modulation as to chronic pain. In particular, our recent studies have shown that CASP6 regulates chronic pain via microglial inflammatory signaling. Inhibition of microglia and CASP signaling might provide a new strategy for the prevention and treatment of chronic pain.

Neuropathic pain is constitutively suppressed in early life by anti-inflammatory neuroimmune regulation.

Peripheral nerve injury can trigger neuropathic pain in adults but not in infants; indeed, for unknown reasons, neuropathic pain is rare before adolescence. We show here that the absence of neuropathic pain response in infant male rats and mice following nerve injury is due to an active, constitutive immune suppression of dorsal horn pain activity. In contrast to adult nerve injury, which triggers a proinflammatory immune response in the spinal dorsal horn, infant nerve injury triggers an anti-inflammatory immune response, characterized by significant increases in IL-4 and IL-10. This immediate anti-inflammatory response can also be evoked by direct C-fiber nerve stimulation in infant, but not adult, mice. Blockade of the anti-inflammatory activity with intrathecal anti-IL10 unmasks neuropathic pain behavior in infant nerve injured mice, showing that pain hypersensitivity in young mice is actively suppressed by a dominant anti-inflammatory neuroimmune response. As infant nerve injured mice reach adolescence (postnatal day 25-30), the dorsal horn immune profile switches from an anti-inflammatory to a proinflammatory response characterized by significant increases in TNF and BDNF, and this is accompanied by a late onset neuropathic pain behavior and increased dorsal horn cell sensitivity to cutaneous mechanical and cold stimuli. These findings show that neuropathic pain following early life nerve injury is not absent but suppressed by neuroimmune activity and that "latent" pain can still emerge at adolescence, when the neuroimmune profile changes. The data may explain why neuropathic pain is rare in young children and also why it can emerge, for no observable reason, in adolescent patients.

Lipoxin A4 inhibits microglial activation and reduces neuroinflammation and neuropathic pain after spinal cord hemisection.

BACKGROUND: Spinal cord injury (SCI) is a severe neurological disorder with many disabling consequences, including persistent neuropathic pain, which develops in about 40 % of SCI patients and is induced and sustained by excessive and uncontrolled spinal neuroinflammation. Here, we have evaluated the effects of lipoxin A4 (LXA4), a member of a unique class of endogenous lipid mediators with both anti-inflammatory and analgesic properties, on spinal neuroinflammation and chronic pain in an experimental model of SCI. METHODS: Spinal hemisection at T10 was carried out in adult male CD1 mice and Wistar rats. To test if LXA4 can reduce neuroinflammation and neuropathic pain, each animal received two intrathecal injections of LXA4 (300 pmol) or vehicle at 4 and 24 h after SCI. Sensitivity to mechanical stimulation of the hind paws was evaluated using von Frey monofilaments, and neuroinflammation was tested by measuring the mRNA and/or protein expression levels of glial markers and cytokines in the spinal cord samples after SCI. Also, microglia cultures prepared from murine cortical tissue were used to assess the direct effects of LXA4 on microglial activation and release of pro-inflammatory TNF-α. RESULTS: LXA4 treatment caused significant reductions in the intensity of mechanical pain hypersensitivity and spinal expression levels of microglial markers and pro-inflammatory cytokines induced by SCI, when compared to rodents receiving control vehicle injections. Notably, the increased expressions of the microglial marker IBA-1 and of the pro-inflammatory cytokine TNF-α were the most affected by the LXA4 treatment. Furthermore, cortical microglial cultures expressed ALX/FPR2 receptors for LXA4 and displayed potentially anti-inflammatory responses upon challenge with LXA4. CONCLUSIONS: Collectively, our results suggest that LXA4 can effectively modulate microglial activation and TNF-α release through ALX/FPR2 receptors, ultimately reducing neuropathic pain in rodents after spinal cord hemisection. The dual anti-inflammatory and analgesic properties of LXA4, allied to its endogenous nature and safety profile, may render this lipid mediator as new therapeutic approach for treating various neuroinflammatory disorders and chronic pain with only limited side effects.

Spinal inhibition of p38 MAP kinase reduces inflammatory and neuropathic pain in male but not female mice: Sex-dependent microglial signaling in the spinal cord.

Previous studies have shown that activation of p38 mitogen-activating kinase (MAPK) in spinal microglia participates in the generation of inflammatory and neuropathic pain in various rodent models. However, these studies focused on male mice to avoid confounding effects of the estrous cycle of females. Recent studies have shown that some spinal pro-inflammatory signaling such as Toll-like receptor 4-mediated signaling contributes to pain hypersensitivity only in male mice. In this study we investigated the distinct role of spinal p38 in inflammatory and neuropathic pain using a highly selective p38 inhibitor skepinone. Intrathecal injection of skepinone prevented formalin induced inflammatory pain in male but not female mice. Furthermore, intrathecal skepinone reduced chronic constriction injury (CCI) induced neuropathic pain (mechanical allodynia) in male mice on CCI-day 7 but not CCI-day 21. This male-dependent inhibition of neuropathic pain also occurred in rats following intrathecal skepinone. Nerve injury induced spinal p38 activation (phosphorylation) in CX3CR1-GFP(+) microglia on CCI-day 7, and this activation was more prominent in male mice. In contrast, CCI induced comparable microgliosis and expression of the microglial markers CX3CR1 and IBA-1 in both sexes. Notably, intraperitoneal or local perineural administration of skepinone inhibited CCI-induced mechanical allodynia in both sexes of mice. Finally, skepinone only reduced the frequency of spontaneous excitatory postsynaptic currents (sEPSCs) in lamina IIo neurons of spinal cord slices of males 7days post CCI. Therefore, the sex-specific p38 activation and signaling is confined to the spinal cord in inflammatory and neuropathic pain conditions.

Connexin-43 induces chemokine release from spinal cord astrocytes to maintain late-phase neuropathic pain in mice.

Accumulating evidence suggests that spinal cord astrocytes play an important role in neuropathic pain sensitization by releasing astrocytic mediators (e.g. cytokines, chemokines and growth factors). However, it remains unclear how astrocytes control the release of astrocytic mediators and sustain late-phase neuropathic pain. Astrocytic connexin-43 (now known as GJ1) has been implicated in gap junction and hemichannel communication of cytosolic contents through the glial syncytia and to the extracellular space, respectively. Connexin-43 also plays an essential role in facilitating the development of neuropathic pain, yet the mechanism for this contribution remains unknown. In this study, we investigated whether nerve injury could upregulate connexin-43 to sustain late-phase neuropathic pain by releasing chemokine from spinal astrocytes. Chronic constriction injury elicited a persistent upregulation of connexin-43 in spinal astrocytes for >3 weeks. Spinal (intrathecal) injection of carbenoxolone (a non-selective hemichannel blocker) and selective connexin-43 blockers (connexin-43 mimetic peptides (43)Gap26 and (37,43)Gap27), as well as astroglial toxin but not microglial inhibitors, given 3 weeks after nerve injury, effectively reduced mechanical allodynia, a cardinal feature of late-phase neuropathic pain. In cultured astrocytes, TNF-α elicited marked release of the chemokine CXCL1, and the release was blocked by carbenoxolone, Gap26/Gap27, and connexin-43 small interfering RNA. TNF-α also increased connexin-43 expression and hemichannel activity, but not gap junction communication in astrocyte cultures prepared from cortices and spinal cords. Spinal injection of TNF-α-activated astrocytes was sufficient to induce persistent mechanical allodynia, and this allodynia was suppressed by CXCL1 neutralization, CXCL1 receptor (CXCR2) antagonist, and pretreatment of astrocytes with connexin-43 small interfering RNA. Furthermore, nerve injury persistently increased excitatory synaptic transmission (spontaneous excitatory postsynaptic currents) in spinal lamina IIo nociceptive synapses in the late phase, and this increase was suppressed by carbenoxolone and Gap27, and recapitulated by CXCL1. Together, our findings demonstrate a novel mechanism of astrocytic connexin-43 to enhance spinal cord synaptic transmission and maintain neuropathic pain in the late-phase via releasing chemokines.

Systemic Progesterone Administration in Early Life Alters the Hyperalgesic Responses to Surgery in the Adult: A Study on Female Rats.

BACKGROUND: There has recently been a substantial increase in the survival of prematurely born neonates and an increase of in utero surgeries. Noxious stimulation in the newborn alters the pain response to injury in adult life. Progesterone, an effective antihyperalgesic agent in the adult, is at high concentration in the pregnant mother. Therefore, we investigated the effects of early-life progesterone on postsurgical outcomes in adult rats. METHODS: Female rat pups were administered progesterone or vehicle during the first 7 days postpartum (P1-P7). A second control group had no injections. Half of each of these groups received an incision of the hindpaw at P3 and the other half did not. At P60, all groups of these now adult rats received a second paw incision. Tactile sensitivity and thermal sensitivity were measured weekly at P14-P42 (period I), at P60 (just before the second incision), and every 2 days of P61-P70 (period II). At P67, rats were fixed by systemic paraformaldehyde perfusion and their spinal cords taken for staining and immunocytochemical analysis of activated p-p38 mitogen-activated protein kinase. RESULTS: Rats with surgery at P3 had greater tactile and thermal hyperalgesia in period I than the nonoperated rats, a difference abolished by progesterone treatment. P3 incision also resulted in long-lasting tactile and thermal hyperalgesia after the P60 incision (period II), both of which were markedly smaller in degree and faster to resolve in rats receiving early progesterone. Even in rats that were not operated on in period I, neonatal progesterone lessened the tactile hyperalgesia in period II. More spinal cells showed p-p38 staining in vehicle-treated rats as a result of the early-life incision but not in those treated with progesterone. CONCLUSIONS: These findings suggest that endogenously high progesterone in utero may have a similarly protective action and that the development of nociceptive circuitry can be strongly influenced by neonatal progesterone.

Neuroprotectin/protectin D1 protects against neuropathic pain in mice after nerve trauma.

Prevalence of neuropathic pain is high after major surgery. However, effective treatment for preventing neuropathic pain is lacking. Here we report that perisurgical treatment of neuroprotectin D1/protectin D1 (NPD1/PD1), derived from docosahexaenoic acid, prevents nerve injury-induced mechanical allodynia and ongoing pain in mice. Intrathecal post-treatment of NPD1/PD1 also effectively reduces established neuropathic pain and produces no apparent signs of analgesic tolerance. Mechanistically, NPD1/PD1 treatment blocks nerve injury-induced long-term potentiation, glial reaction, and inflammatory responses, and reverses synaptic plasticity in the spinal cord. Thus, NPD1/PD1 and related mimetics might serve as a new class of analgesics for preventing and treating neuropathic pain.

MicroRNA let-7b enhances spinal cord nociceptive synaptic transmission and induces acute and persistent pain through neuronal and microglial signaling.

ABSTRACT: Secreted microRNAs (miRNAs) have been detected in various body fluids including the cerebrospinal fluid, yet their direct role in regulating synaptic transmission remains uncertain. We found that intrathecal injection of low dose of let-7b (1 µg) induced short-term (<24 hours) mechanical allodynia and heat hyperalgesia, a response that is compromised in Tlr7-/- or Trpa1-/- mice. Ex vivo and in vivo calcium imaging in GCaMP6-report mice revealed increased calcium signal in spinal cord afferent terminals and doral root ganglion/dorsal root ganglia neurons following spinal perfusion and intraplantar injection of let-7b. Patch-clamp recordings also demonstrated enhanced excitatory synaptic transmission (miniature excitatory postsynaptic currents [EPSCs]) in spinal nociceptive neurons following let-7b perfusion or optogenetic activation of axonal terminals. The elevation in spinal calcium signaling and EPSCs was dependent on the presence of toll-like receptor-7 (TLR7) and transient receptor potential ion channel subtype A1 (TRPA1). In addition, endogenous let-7b is enriched in spinal cord synaptosome, and peripheral inflammation increased let-7b in doral root ganglion/dorsal root ganglia neurons, spinal cord tissue, and the cerebrospinal fluid. Notably, let-7b antagomir inhibited inflammatory pain and inflammation-induced synaptic plasticity (EPSC increase), suggesting an endogenous role of let-7b in regulating pain and synaptic transmission. Furthermore, intrathecal injection of let-7b, at a higher dose (10 µg), induced persistent mechanical allodynia for >2 weeks, which was abolished in Tlr7-/- mice. The high dose of let-7b also induced microgliosis in the spinal cord. Of interest, intrathecal minocycline only inhibited let-7b-induced mechanical allodynia in male but not female mice. Our findings indicate that the secreted microRNA let-7b has the capacity to provoke pain through both neuronal and glial signaling, thereby establishing miRNA as an emerging neuromodulator.