dynorphins and Neuralgia

Chronic pain is one of the most ubiquitous diseases in the world, but treatment is difficult with conventional methods, due to undesirable side effects of treatments and unknown mechanisms of pathological pain states. The endogenous peptide, dynorphin A has long been established as a target for the treatment of pain. Interestingly, this unique peptide has both inhibitory (opioid in nature) and excitatory activities (nonopioid) in the CNS. Both of these effects have been found to play a role in pain and much work has been done to develop therapeutics to enhance the inhibitory effects. Here we will review the dynorphin A compounds that have been designed for the modulation of pain and will discuss where the field stands today.

Topics: Analgesics, Non-Narcotic; Chronic Disease; Dynorphins; Humans; Neuralgia; Neurotransmitter Agents; Receptors, Bradykinin; Receptors, N-Methyl-D-Aspartate; Receptors, Opioid, kappa; Structure-Activity Relationship

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, whi

Topics: Animals; Chronic Pain; Dynorphins; Mice; Narcotic Antagonists; Neuralgia; Receptors, Opioid, kappa; Wakefulness

Panax notoginseng (Chinese ginseng, Sanqi), one of the major ginseng species, has been traditionally used to alleviate different types of chronic pain. The raw P. notoginseng powder is commonly available in China as a non-prescription drug to treat various aliments including arthritic pain. However, strong scientific evidence is needed to illustrate its pain antihypersensitive effects, effective ingredients and mechanism of action. The oral P. notoginseng powder dose-dependently alleviated formalin-induced tonic hyperalgesia, and its total ginsenosides remarkably inhibited neuropathic pain hypersensitivity. Ginsenoside Rb1, the most abundant ginsenoside of P. notoginseng, dose-dependently produced neuropathic pain antihypersensitivity. Conversely, ginsenosides Rg1, Re and notoginseng R1, the other major saponins from P. notoginseng, failed to inhibit formalin-induced tonic pain or mechanical allodynia in neuropathic pain. Ginsenoside Rb1 metabolites ginsenosides Rg3, Compound-K and protopanaxadiol also had similar antineuropathic pain efficacy to ginsenoside Rb1. Additionally, intrathecal ginsenoside Rb1 specifically stimulated dynorphin A expression which was colocalized with microglia but not neurons or astrocytes in the spinal dorsal horn and primary cultured cells. Pretreatment with microglial metabolic inhibitor minocycline, dynorphin A antiserum and specific κ-opioid receptor antagonist GNTI completely blocked Rb1-induced mechanical antiallodynia in neuropathic pain. Furthermore, the specific glucocorticoid receptor (GR) antagonist Dex-21-mesylate (but not GPR30 estrogen receptor antagonist G15) also entirely attenuated ginsenoside Rb1-related antineuropathic pain effects. All these results, for the first time, show that P. notoginseng alleviates neuropathic pain and ginsenoside Rb1 is its principal effective ingredient. Furthermore, ginsenoside Rb1 inhibits neuropathic pain by stimulation of spinal microglial dynorphin A expression following GR activation.

Topics: Dynorphins; Ginsenosides; Hyperalgesia; Microglia; Neuralgia; Panax notoginseng; Powders

Pain is a worldwide public health problem and its treatment is still a challenge since clinically available drugs do not completely reverse chronic painful states or induce undesirable effects. Crotalphine is a 14 amino acids synthetic peptide that induces a potent and long-lasting analgesic effect on acute and chronic pain models, peripherally mediated by the endogenous release of dynorphin A and the desensitization of the transient receptor potential ankyrin 1 (TRPA1) receptor. However, the effects of crotalphine on the central nervous system (CNS) and the signaling pathway have not been investigated. Thus, the central effect of crotalphine was evaluated on the partial sciatic nerve ligation (PSNL)-induced chronic neuropathic pain model. Crotalphine (100 µg/kg, p.o.)-induced analgesia on the 14th day after surgery lasting up to 24 h after administration. This effect was prevented by intrathecal administration of CB1 (AM251) or CB2 (AM630) cannabinoid receptor antagonists. Besides that, crotalphine-induced analgesia was reversed by CTOP, nor-BNI, and naltrindole, antagonists of

Topics: Amino Acids; Analgesia; Analgesics; Analgesics, Opioid; Ankyrins; beta-Endorphin; Cannabinoid Receptor Antagonists; Cannabinoids; Dynorphins; Enkephalin, Methionine; Humans; Interleukin-6; Lipopolysaccharides; Microglia; Minocycline; Neuralgia; Peptides; Phenotype; Receptors, Opioid; Spinal Cord

New remedies are required for the treatment of neuropathic pain due to insufficient efficacy of available therapies. This study provides a novel approach to develop painkillers for chronic pain treatment.. The rat formalin pain test and spinal nerve ligation model of neuropathic pain were used to evaluate antinociception of protopanaxadiol. Primary cell cultures, immunofluorescence staining, and gene and protein expression were also performed for mechanism studies.. Gavage protopanaxadiol remarkably produces pain antihypersensitive effects in neuropathic pain, bone cancer pain and inflammatory pain, with efficacy comparable with gabapentin. Long-term PPD administration does not induce antihypersensitive tolerance, but prevents and reverses the development and expression of morphine analgesic tolerance. Oral protopanaxadiol specifically stimulates spinal expression of dynorphin A in microglia but not in astrocytes or neurons. Protopanaxadiol gavage-related pain antihypersensitivity is abolished by the intrathecal pretreatment with the microglial metabolic inhibitor minocycline, dynorphin antiserum or specific κ-opioid receptor antagonist GNTI. Intrathecal pretreatment with glucocorticoid receptor)antagonists RU486 and dexamethasone-21-mesylate, but not GPR-30 antagonist G15 or mineralocorticoid receptor antagonist eplerenone, completely attenuates protopanaxadiol-induced spinal dynorphin A expression and pain antihypersensitivity in neuropathic pain. Treatment with protopanaxadiol, the glucocorticoid receptor agonist dexamethasone and membrane-impermeable glucocorticoid receptor agonist dexamethasone-BSA in cultured microglia induces remarkable dynorphin A expression, which is totally blocked by pretreatment with dexamthasone-21-mesylate.. All the results, for the first time, indicate that protopanaxadiol produces pain antihypersensitivity in neuropathic pain probably through spinal microglial dynorphin A expression after glucocorticoid receptor activation and hypothesize that microglial membrane glucocorticoid receptor/dynorphin A pathway is a potential target to discover and develop novel painkillers in chronic pain.

Topics: Animals; Dynorphins; Glucocorticoids; Hyperalgesia; Microglia; Neuralgia; Rats; Rats, Wistar; Receptors, Glucocorticoid; Sapogenins; Spinal Cord

Isotalatizidine is a representative C. A chronic constrictive injury (CCI)-induced model of neuropathic pain was established in mice, and the limb withdrawal was evaluated by the Von Frey filament test following isotalatizidine or placebo administration. The signaling pathways in primary or immortalized microglia cells treated with isotalatizidine were analyzed by Western blotting and immunofluorescence.. Intrathecal injection of isotalatizidine attenuated the CCI-induced mechanical allodynia in a dose-dependent manner. At the molecular level, isotalatizidine selectively increased the phosphorylation of p38 and ERK1/2, in addition to activating the transcription factor CREB and increasing dynorphin A production in cultured primary microglia. However, the downstream effects of isotalatizidine were abrogated by the selective ERK1/2 inhibitor U0126-EtOH or CREB inhibitor of KG-501, but not by the p38 inhibitor SB203580. The results also were confirmed in in vivo experiments.. Taken together, isotalatizidine specifically activates the ERK1/2 pathway and subsequently CREB, which triggers dynorphin A release in the microglia, eventually leading to its anti-nociceptive action.

Topics: Aconitine; Analgesics; Animals; Chronic Pain; Cyclic AMP Response Element-Binding Protein; Dynorphins; MAP Kinase Signaling System; Mice; Microglia; Neuralgia; Signal Transduction

Dynorphin A is increased in neuropathic pain models. Activation of α7 n acetylcholine receptor (nAchR) reduces inflammation and pain. Whether activation of α7 nAchR affects dynorphin A release is unknown. The experiments evaluated the proinflammatory effect of dynorphin A in the spinal nerve ligation-induced neuropathic pain models and the effect of α7 nAchR activation on the dynorphin A content. α7 nAchR agonist, PHA-543613 and its antagonist, methyllycaconitine citrate were used and dynorphin A content was measured after spinal nerve ligation and in microglia cultures to test the analgesic mechanisms of α7 nAchR activation. The results showed that dynorphin A content peaked 3 to 7 days after nerve injury, and dynorphin A anti-serum intrathecal injection decreased IL-β and TNF-α content a week after nerve injury. Activation of α7 nAchR by PHA-543613 alleviated neuropathic pain behaviors and decreased dynorphin A concentration in the ipsilateral spinal cords. Also, PHA-543613 decreased dynorphin A release from the microglia cultures to LPS stimulation by activation of α7 nAchR. Our results suggest that dynorphin A contribute to the development and maintenance of neuropathic pain and that decreasing dynorphin A content by activation of α7 AchR of microglia is a potential therapeutic target for treating neuropathic pain.

Topics: Aconitine; alpha7 Nicotinic Acetylcholine Receptor; Animals; Bridged Bicyclo Compounds, Heterocyclic; Dynorphins; Injections, Spinal; Male; Microglia; Neuralgia; Nicotinic Agonists; Nicotinic Antagonists; Quinuclidines; Rats; Rats, Sprague-Dawley; Spinal Cord; Spinal Nerves

Aconiti brachypodi Radix (Xue-shang-yi-zhi-hao) has been prescribed to manage chronic pain, arthritis, and traumatic injuries. Bullatine A, a C20-diterpenoid alkaloid, is one of its principle effective compounds. This study aimed to investigate the anti-hypersensitivity of bullatine A in a variety of rat pain models and explore its mechanisms of action.. Rat neuropathic pain, inflammatory pain, diabetic neuropathic pain, and bone cancer pain models were used. Dynorphin A and pro-inflammatory cytokines were measured in the spinal cord and cultured primary microglia. Double immunofluorescence staining of dynorphin A and glial and neuronal cellular markers was also measured in the spinal cord.. Subcutaneous and intrathecal injection of bullatine A dose-dependently attenuated spinal nerve ligation-, complete Freud's adjuvant-, diabetes-, and bone cancer-induced mechanical allodynia and thermal hyperalgesia, with the efficacies of 45-70 % inhibition, and half-effective doses of 0.9-1.9 mg/kg for subcutaneous injection. However, bullatine A was not effective in blocking acute nociceptive response in the normal condition. Bullatine A specifically stimulated dynorphin A expression in microglia in the spinal cord in vivo and cultured primary microglia in vitro; the stimulatory effects were completely inhibited by the microglial inhibitor minocycline. In contrast, bullatine A did not have an inhibitory effect on peripheral nerve injury- or lipopolysaccharide-induced pro-inflammatory cytokine expression. The spinal anti-allodynic effects of bullatine A were entirely blocked by intrathecal injection of minocycline, the specific dynorphin A antiserum, and the selective k-opioid receptor antagonist.. We, for the first time, demonstrate that bullatine A specifically attenuates pain hypersensitivity, regardless of the pain models employed. The results also suggest that stimulation of spinal microglial dynorphin A expression mediates bullatine A anti-nociception in pain hypersensitivity conditions.

Topics: Alkaloids; Analgesics; Animals; Animals, Newborn; Cells, Cultured; Disease Models, Animal; Diterpenes; Dose-Response Relationship, Drug; Dynorphins; Female; Gene Expression; Hyperalgesia; Injections, Subcutaneous; Male; Neuralgia; Rats; Rats, Sprague-Dawley; Rats, Wistar; Spinal Cord

Neuropathic pain often results from peripheral nerve damage, which can involve immune response. Local leukocyte-derived opioid peptides or exogenous opioid agonists inhibit neuropathy-induced mechanical hypersensitivity in animal models. Since neuropathic pain can also be augmented by heat, in this study we investigated the role of opioids in the modulation of neuropathy-evoked heat hypersensitivity. We used a chronic constriction injury of the sciatic nerve in wild-type and opioid peptide-knockout mice, and tested opioid effects in heat and mechanical hypersensitivity using Hargreaves and von Frey tests, respectively. We found that although perineural exogenous opioid agonists, including peptidergic ligands, were effective, the endogenous opioid peptides β-endorphin, Met-enkephalin and dynorphin A did not alleviate heat hypersensitivity. Specifically, corticotropin-releasing factor, an agent triggering opioid peptide secretion from leukocytes, applied perineurally did not attenuate heat hypersensitivity in wild-type mice. Exogenous opioids, also shown to release opioid peptides via activation of leukocyte opioid receptors, were equally analgesic in wild-type and opioid peptide-knockout mice, indicating that endogenous opioids do not contribute to exogenous opioid analgesia in heat hypersensitivity. Furthermore, exogenously applied opioid peptides were ineffective as well. Conversely, opioid peptides relieved mechanical hypersensitivity. Thus, both opioid type and sensory modality may determine the outcome of neuropathic pain treatment.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesics, Opioid; Animals; beta-Endorphin; Corticotropin-Releasing Hormone; Dynorphins; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Methionine; Hot Temperature; Male; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Opioid Peptides; Peripheral Nervous System; Receptors, Opioid

During the inflammation which occurs following nerve damage, macrophages are recruited to the site of injury. Phenotypic diversity is a hallmark of the macrophage lineage and includes pro-inflammatory M1 and anti-inflammatory M2 populations. Our aim in this study was to investigate the ability of polarized M0, M1, and M2 macrophages to secrete opioid peptides and to examine their relative contribution to the modulation of neuropathic pain.. Mouse bone marrow-derived cells were cultured as unstimulated M0 macrophages or were stimulated into an M1 phenotype using lipopolysaccharide and interferon-γ or into an M2 phenotype using interleukin-4. The macrophage phenotypes were verified using flow cytometry for surface marker analysis and cytokine bead array for cytokine profile assessment. Opioid peptide levels were measured by radioimmunoassay and enzyme immunoassay. As a model of neuropathic pain, a chronic constriction injury (CCI) of the sciatic nerve was employed. Polarized M0, M1, and M2 macrophages (5 × 10. Compared to M0 and M1 cells, M2 macrophages contained and released higher amounts of opioid peptides, including Met-enkephalin, dynorphin A (1-17), and β-endorphin. M2 cells transferred perineurally at the nerve injury site reduced mechanical, but not heat hypersensitivity following the second injection. The analgesic effect was reversed by the perineurally applied opioid receptor antagonist naloxone methiodide. M2 cells did not affect sensitivity following sham surgery. Neither M0 nor M1 cells altered mechanical and heat sensitivity in CCI or sham-operated animals. Tracing the fluorescently labeled M0, M1, and M2 cells ex vivo showed that they remained in the nerve and preserved their phenotype.. Perineural transplantation of M2 macrophages resulted in opioid-mediated amelioration of neuropathy-induced mechanical hypersensitivity, while M1 macrophages did not exacerbate pain. Therefore, rather than focusing on macrophage-induced pain generation, promoting opioid-mediated M2 actions may be more relevant for pain control.

Topics: Acyltransferases; Adoptive Transfer; Animals; beta-Endorphin; Cell Polarity; Cytokines; Disease Models, Animal; Dynorphins; Flow Cytometry; Histocompatibility Antigens Class II; Hyperalgesia; Lipopolysaccharides; Macrophages; Male; Mice; Mice, Inbred C57BL; Neuralgia; Opioid Peptides; Pain Threshold; Physical Stimulation

Infraorbital nerve constriction (CION) causes hypersensitivity to facial mechanical, heat and cold stimulation in rats and mice and is a reliable model to study trigeminal neuropathic pain. In this model there is evidence that mechanisms operated by kinin B1 and B2 receptors contribute to heat hyperalgesia in both rats and mice. Herein we further explored this issue and assessed the role of kinin receptors in mechanical hyperalgesia after CION. Swiss and C57Bl/6 mice that underwent CION or sham surgery or dynorphin A (1-17) administration were repeatedly submitted to application of either heat stimuli to the snout or mechanical stimuli to the forehead. Treatment of the animals on the fifth day after CION surgery with DALBK (B1 receptor antagonist) or HOE-140 (B2 receptor antagonist), both at 0.01-1μmol/kg (i.p.), effectively reduced CION-induced mechanical hyperalgesia. Knockout mice for kinin B1, B2 or B1/B2 receptors did not develop heat or mechanical hyperalgesia in response to CION. Subarachnoid dynorphin A (1-17) delivery (15nmol/5μL) also resulted in orofacial heat hyperalgesia, which was attenuated by post-treatment with DALBK (1 and 3μmol/kg, i.p.), but was not affected by HOE-140. Additionally, treatment with an anti-dynorphin A antiserum (200μg/5μL, s.a.) reduced CION-induced heat hyperalgesia for up to 2h. These results suggest that both kinin B1 and B2 receptors are relevant in orofacial sensory nociceptive changes induced by CION. Furthermore, they also indicate that dynorphin A could stimulate kinin receptors and this effect seems to contribute to the maintenance of trigeminal neuropathic pain.

Topics: Animals; Bradykinin; Bradykinin B1 Receptor Antagonists; Bradykinin B2 Receptor Antagonists; Disease Models, Animal; Dynorphins; Facial Pain; Hot Temperature; Hyperalgesia; Male; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Neurotransmitter Agents; Pain Measurement; Receptors, Bradykinin; Touch

Interleukin-17 (IL-17) is involved in a wide range of inflammatory disorders and in recruitment of inflammatory cells to injury sites. A recent study of IL-17 knock-out mice revealed that IL-17 contributes to neuroinflammation and neuropathic pain after peripheral nerve injury. Surprisingly, little is known of micro-environment modulation by IL-17 in injured sites and in pathologically related neuroinflammation and chronic neuropathic pain. Therefore, we investigated nociceptive sensitization, immune cell infiltration, myeloperoxidase (MPO) activity, and expression of multiple cytokines and opioid peptides in damaged nerves of wild-type (IL-17(+/+)) and IL-17 knock-out (IL-17(-/-)) mice after partial sciatic nerve ligation. Our results demonstrated that the IL-17(-/-) mice had less behavioral hypersensitivity after partial sciatic nerve ligation, and inflammatory cell infiltration and pro-inflammatory cytokine (tumor necrosis factor-α, IL-6, and interferon-γ) levels in damaged nerves were significantly decreased, with the levels of anti-inflammatory cytokines IL-10 and IL-13, and expressions of enkephalin, β-endorphin, and dynorphin were also decreased compared to those in wild-type control mice. In conclusion, we provided evidence that IL-17 modulates the micro-environment at the level of the peripheral injured nerve site and regulates progression of behavioral hypersensitivity in a murine chronic neuropathic pain model. The attenuated behavioral hypersensitivity in IL-17(-/-) mice could be a result of decreased inflammatory cell infiltration to the injured site, resulting in modulation of the pro- and anti-inflammatory cytokine milieu within the injured nerve. Therefore, IL-17 may be a critical component for neuropathic pain pathogenesis and a novel target for therapeutic intervention for this and other chronic pain states.

Topics: Animals; Behavior, Animal; beta-Endorphin; Central Nervous System Sensitization; Cytokines; Disease Models, Animal; Dynorphins; Enkephalins; Hyperalgesia; Inflammation; Interleukin-10; Interleukin-13; Interleukin-17; Interleukin-1beta; Interleukin-2; Interleukin-6; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Neutrophils; Nociception; Peripheral Nerve Injuries; Peroxidase; Sciatic Nerve; T-Lymphocytes; Tumor Necrosis Factor-alpha

Dynorphin A is an endogenous opioid peptide derived from the precursor prodynorphin. The proteolytic fragment dynorphin A (1-17) exhibits inhibitory effects via opioid receptors. Paradoxically, the activity of the dynorphin system increases with chronic pain and neuropathy is associated with the up-regulation of dynorphin biosynthesis. Dynorphin A (1-17) is cleaved in vivo to produce a non-opioid fragment, dynorphin A (2-17). Previously, a mechanism by which the non-opioid fragment promotes pain through agonist action at bradykinin receptors was revealed. Bradykinin receptor expression is up-regulated after nerve injury and both a truncated version of non-opioid fragment dynorphin A (2-17), referred to as 'Ligand 10', and novel bradykinin receptor antagonist 'Ligand 14', are known to bind to the bradykinin receptor. Here we show that Ligand 10 facilitates the response of wide dynamic range (WDR) neurons to innocuous and noxious mechanical stimuli in neuropathic, but not naïve, animals, while Ligand 14 exhibits inhibitory effects in neuropathic animals only. Furthermore, we reveal an inhibitory effect of Ligand 14 in naïve animals by pre-dosing with either Ligand 10 or a 5-HT3 receptor agonist to reflect activation of descending excitatory drives. Thus remarkably, by mimicking pro-excitatory pharmacological changes that occur after nerve injury in a naïve animal, we induce a state whereby the inhibitory actions of Ligand 14 are now effective. Ultimately our data support an increasing number of studies that suggest that blocking spinal bradykinin receptors may have a therapeutic potential in chronic pain states, here, in particular, in neuropathic pain.

Topics: Analgesics, Non-Narcotic; Animals; Bradykinin B2 Receptor Antagonists; Disease Models, Animal; Dynorphins; Male; Neuralgia; Neuronal Plasticity; Neurons; Nociception; Peptide Fragments; Physical Stimulation; Rats, Sprague-Dawley; Serotonin 5-HT3 Receptor Agonists; Spinal Cord; Spinal Nerves

Despite many advances, our understanding of the involvement of prodynorphin systems in the development of neuropathic pain is not fully understood. Recent studies suggest an important role of neuro-glial interactions in the dynorphin effects associated with neuropathic pain conditions. Our studies show that minocycline reduced prodynorphin mRNA levels that were previously elevated in the spinal and/or dorsal root ganglia (DRG) following sciatic nerve injury. The repeated intrathecal administration of minocycline enhanced the analgesic effects of low-dose dynorphin (0.15 nmol) and U50,488H (25-100 nmol) and prevented the development of flaccid paralysis following high-dose dynorphin administration (15 nmol), suggesting a neuroprotective effect. Minocycline reverts the expression of IL-1β and IL-6 mRNA within the spinal cord and IL-1β mRNA in DRG, which was elevated following intrathecal administration of dynorphin (15 nmol). These results suggest an important role of these proinflammatory cytokines in the development of the neurotoxic effects of dynorphin. Similar to minocycline, a selective inhibitor of MMP-9 (MMP-9 levels are reduced by minocycline) exerts an analgesic effect in behavioral studies, and its administration prevents the occurrence of flaccid paralysis caused by high-dose dynorphin administration (15 nmol). In conclusion, our results underline the importance of neuro-glial interactions as evidenced by the involvement of IL-1β and IL-6 and the minocycline effect in dynorphin-induced toxicity, which suggests that drugs that alter the prodynorphin system could be used to better control neuropathic pain.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesics, Non-Narcotic; Animals; Disease Models, Animal; Dynorphins; Ganglia, Spinal; Injections, Spinal; Interleukin-1beta; Interleukin-6; Male; Matrix Metalloproteinase 9; Minocycline; Neuralgia; Neuroprotective Agents; Paraplegia; Rats, Wistar; RNA, Messenger; Sciatic Neuropathy; Spinal Cord

Although morphine and other mu-opioid agonists are the main analgesics for severe pain, these compounds have potential for abuse and/or addiction. This has complicated the use of mu-agonists in the treatment of chronic pain. However, clinical studies show that when mu-agonist analgesics are appropriately used to control pain, actual abuse or addiction does not usually occur, although some risk factors that increase vulnerability need to be considered, including genetic variation. We review recent findings on molecular adaptations in sustained pain models, and propose how these adaptations (including sustained release of the endogenous mu-agonist beta-endorphin) can result in decreased abuse potential of mu-agonists in chronic pain states. We also review data on particular gene polymorphisms (e.g. in the mu-receptor gene) that could also influence the relative abuse potential of mu-agonists in clinical pain populations.

Topics: Analgesics, Opioid; Chronic Disease; Down-Regulation; Dynorphins; Endorphins; Extracellular Signal-Regulated MAP Kinases; Humans; Neuralgia; Opioid-Related Disorders; Pain; Protein Kinase C; Receptors, Dopamine; Receptors, Opioid, mu; Reward; Up-Regulation; Ventral Tegmental Area

Dynorphin A is an endogenous opioid peptide that produces non-opioid receptor-mediated neural excitation. Here we demonstrate that dynorphin induces calcium influx via voltage-sensitive calcium channels in sensory neurons by activating bradykinin receptors. This action of dynorphin at bradykinin receptors is distinct from the primary signaling pathway activated by bradykinin and underlies the hyperalgesia produced by pharmacological administration of dynorphin by the spinal route in rats and mice. Blockade of spinal B1 or B2 receptor also reverses persistent neuropathic pain but only when there is sustained elevation of endogenous spinal dynorphin, which is required for maintenance of neuropathic pain. These data reveal a mechanism for endogenous dynorphin to promote pain through its agonist action at bradykinin receptors and suggest new avenues for therapeutic intervention.

Topics: Animals; Calcium; Calcium Channels; Dynorphins; Male; Mice; Mice, Knockout; Nerve Degeneration; Neuralgia; Neurons, Afferent; Rats; Rats, Sprague-Dawley; Receptors, Bradykinin; Signal Transduction; Single-Blind Method; Spinal Nerves

It is generally accepted that neuropathic pain is resistant to amelioration by morphine in clinical studies and insensitivity to intrathecal (i.t.) administered morphine in experimental models of neuropathic pain has been demonstrated. This study is to determine if endogenous dynorphin, substance P or cholecystokinin is involved in the lack of anti-allodynia of morphine in a partial sciatic nerve ligation (PSL) model of CD-1 mice. Mice exhibited tactile allodynia in the ipsilateral hind paw 1 day after PSL, and reached its maximal allodynic effect at 2 days and remained allodynic for 7 days. Morphine (3.0 nmol) given i.t. did not alter the tactile allodynic threshold in ipsilateral paw of mice pretreated i.t. with normal rabbit serum 2 days after PSL. However, the same dose of morphine (3.0 nmol) given i.t. reduced markedly allodynia in mice pretreated for 2h with antiserum against dynorphin A(1-17) (200 microg); the morphine-produced anti-allodynia developed slowly, reached its peak effect at 30 min and returned to an allodynic state in 60 min. Similarly, i.t. injection of morphine reduced the allodynia in PSL mice pretreated with antiserum against substance P (10 microg) or cholecystokinin (200 microg) for 2h. Intrathecal pretreatment with antiserum against dynorphin A(1-17), substance P or cholecystokinin for 2h injected alone did not affect the baseline mechanical tactile threshold in ipsilateral paw 2 days after PSL. The results indicate that endogenous dynorphin A(1-17), substance P and cholecystokinin are involved in PSL-induced neuropathic allodynia to attenuate the anti-allodynic effect of morphine.

Topics: Analgesics, Opioid; Animals; Antibodies; Cholecystokinin; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Resistance; Drug Synergism; Dynorphins; Immune Sera; Injections, Spinal; Ligation; Male; Mice; Morphine; Neuralgia; Peripheral Nervous System Diseases; Sciatic Nerve; Sciatic Neuropathy; Substance P

Release of endogenous dynorphin opioids within the spinal cord after partial sciatic nerve ligation (pSNL) is known to contribute to the neuropathic pain processes. Using a phosphoselective antibody [kappa opioid receptor (KOR-P)] able to detect the serine 369 phosphorylated form of the KOR, we determined possible sites of dynorphin action within the spinal cord after pSNL. KOR-P immunoreactivity (IR) was markedly increased in the L4-L5 spinal dorsal horn of wild-type C57BL/6 mice (7-21 d) after lesion, but not in mice pretreated with the KOR antagonist nor-binaltorphimine (norBNI). In addition, knock-out mice lacking prodynorphin, KOR, or G-protein receptor kinase 3 (GRK3) did not show significant increases in KOR-P IR after pSNL. KOR-P IR was colocalized in both GABAergic neurons and GFAP-positive astrocytes in both ipsilateral and contralateral spinal dorsal horn. Consistent with sustained opioid release, KOR knock-out mice developed significantly increased tactile allodynia and thermal hyperalgesia in both the early (first week) and late (third week) interval after lesion. Similarly, mice pretreated with norBNI showed enhanced hyperalgesia and allodynia during the 3 weeks after pSNL. Because sustained activation of opioid receptors might induce tolerance, we measured the antinociceptive effect of the kappa agonist U50,488 using radiant heat applied to the ipsilateral hindpaw, and we found that agonist potency was significantly decreased 7 d after pSNL. In contrast, neither prodynorphin nor GRK3 knock-out mice showed U50,488 tolerance after pSNL. These findings suggest that pSNL induced a sustained release of endogenous prodynorphin-derived opioid peptides that activated an anti-nociceptive KOR system in mouse spinal cord. Thus, endogenous dynorphin had both pronociceptive and antinociceptive actions after nerve injury and induced GRK3-mediated opioid tolerance.

Topics: Animals; Astrocytes; Disease Models, Animal; Disease Progression; Drug Tolerance; Dynorphins; Enkephalins; G-Protein-Coupled Receptor Kinase 3; Hyperalgesia; Lumbosacral Region; Mice; Mice, Inbred C57BL; Mice, Knockout; Narcotic Antagonists; Narcotics; Neuralgia; Neurons; Protein Precursors; Protein Serine-Threonine Kinases; Receptors, Opioid; Receptors, Opioid, kappa; Sciatic Neuropathy; Spinal Cord

Cytokines such as IL-1beta, IL-6 and tumor necrosis factor-alpha (TNF-alpha) have been shown to contribute directly to central and peripheral neuropathic pain. Recently, exogenous interleukin-10 (IL-10) was shown to impede development of dynorphin-induced allodynia presumably by inhibiting IL-1beta. We therefore wanted to determine whether endogenous IL-10 had a role in pain perception. By measuring the latency of the paw licking response, we show in IL-10 knockout mice and in normal mice treated with anti-IL-10 that latency times are increased, suggesting that endogenous IL-10 increases nociception. This does not appear to be directly correlated with IL-10's regulation of DREAM, a transcriptional regulator of prodynorphin synthesis.

Topics: Animals; Antibodies; Calcium-Binding Proteins; Cell Line; Central Nervous System; Dynorphins; Interleukin-10; Kv Channel-Interacting Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Neuralgia; Nociceptors; Pain Measurement; Peripheral Nervous System Diseases; Reaction Time; Repressor Proteins

Nerve injury-induced afferent discharge is thought to elicit spinal sensitization and consequent abnormal pain. Experimental neuropathic pain, however, also depends on central changes, including descending facilitation arising from the rostral ventromedial medulla (RVM) and upregulation of spinal dynorphin. A possible intersection of these influences at the spinal level was explored by measuring evoked, excitatory transmitter release in tissues taken from nerve-injured animals with or without previous manipulation of descending modulatory systems. Spinal nerve ligation (SNL) produced expected tactile and thermal hyperesthesias. Capsaicin-evoked calcitonin gene-related peptide (CGRP) release was markedly enhanced in lumbar spinal tissue from SNL rats when compared with sham-operated controls. Enhanced, evoked CGRP release from SNL rats was blocked by anti-dynorphin A(1-13) antiserum; this treatment did not alter evoked release in tissues from sham-operated rats. Dorsolateral funiculus lesion (DLF) or destruction of RVM neurons expressing mu-opioid receptors with dermorphin-saporin, blocked tactile and thermal hypersensitivity, as well as SNL-induced upregulation of spinal dynorphin. Spinal tissues from these DLF-lesioned or dermorphin-saporin-treated SNL rats did not exhibit enhanced capsaicin-evoked CGRP-IR release. These data demonstrate exaggerated release of excitatory transmitter from primary afferents after injury to peripheral nerves, supporting the likely importance of increased afferent input as a driving force of neuropathic pain. The data also show that modulatory influences of descending facilitation are required for enhanced evoked transmitter release after nerve injury. Thus, convergence of descending modulation, spinal plasticity, and afferent drive in the nerve-injured state reveals a mechanism by which some aspects of nerve injury-induced hyperesthesias may occur.

Topics: Afferent Pathways; Analgesics, Opioid; Animals; Calcitonin Gene-Related Peptide; Capsaicin; Disease Models, Animal; Dynorphins; Enkephalins; Hyperesthesia; Ligation; Lumbosacral Region; Male; Medulla Oblongata; Microinjections; N-Glycosyl Hydrolases; Nerve Compression Syndromes; Neuralgia; Neuronal Plasticity; Neurotransmitter Agents; Oligopeptides; Opioid Peptides; Pain Measurement; Protein Precursors; Rats; Rats, Sprague-Dawley; Receptors, Opioid, mu; Recombinant Fusion Proteins; Ribosome Inactivating Proteins, Type 1; Saporins; Sensory Thresholds; Spinal Cord; Spinal Nerves

Whereas tissue injury increases spinal dynorphin expression, the functional relevance of this upregulation to persistent pain is unknown. Here, mice lacking the prodynorphin gene were studied for sensitivity to non-noxious and noxious stimuli, before and after induction of experimental neuropathic pain. Prodynorphin knock-out (KO) mice had normal responses to acute non-noxious stimuli and a mild increased sensitivity to some noxious stimuli. After spinal nerve ligation (SNL), both wild-type (WT) and KO mice demonstrated decreased thresholds to innocuous mechanical and to noxious thermal stimuli, indicating that dynorphin is not required for initiation of neuropathic pain. However, whereas neuropathic pain was sustained in WT mice, KO mice showed a return to baselines by post-SNL day 10. In WT mice, SNL upregulated lumbar dynorphin content on day 10, but not day 2, after injury. Intrathecal dynorphin antiserum reversed neuropathic pain in WT mice at post-SNL day 10 (when dynorphin was upregulated) but not on post-SNL day 2; intrathecal MK-801 reversed SNL-pain at both times. Opioid (mu, delta, and kappa) receptor density and G-protein activation were not different between WT and KO mice and were unchanged by SNL injury. The observations suggest (1) an early, dynorphin-independent phase of neuropathic pain and a later dynorphin-dependent stage, (2) that upregulated spinal dynorphin is pronociceptive and required for the maintenance of persistent neuropathic pain, and (3) that processes required for the initiation and the maintenance of the neuropathic pain state are distinct. Identification of mechanisms that maintain neuropathic pain appears important for strategies to treat neuropathic pain.

Topics: Animals; Chronic Disease; Disease Models, Animal; Dizocilpine Maleate; Dynorphins; Excitatory Amino Acid Antagonists; Hyperesthesia; Immune Sera; Injections, Spinal; Ligation; Lumbosacral Region; Male; Mice; Mice, Knockout; Neuralgia; Pain Measurement; Pain Threshold; Physical Stimulation; Reaction Time; Receptors, Opioid; Spinal Cord; Spinal Nerves

The aim of this investigation was to determine whether murine models of inflammatory, neuropathic and cancer pain are each characterized by a unique set of neurochemical changes in the spinal cord and sensory neurons. All models were generated in C3H/HeJ mice and hyperalgesia and allodynia behaviorally characterized. A variety of neurochemical markers that have been implicated in the generation and maintenance of chronic pain were then examined in spinal cord and primary afferent neurons.Three days after injection of complete Freund's adjuvant into the hindpaw (a model of persistent inflammatory pain) increases in substance P, calcitonin gene-related peptide, protein kinase C gamma, and substance P receptor were observed in the spinal cord. Following sciatic nerve transection or L5 spinal nerve ligation (a model of persistent neuropathic pain) significant decreases in substance P and calcitonin gene-related peptide and increases in galanin and neuropeptide Y were observed in both primary afferent neurons and the spinal cord. In contrast, in a model of cancer pain induced by injection of osteolytic sarcoma cells into the femur, there were no detectable changes in any of these markers in either primary afferent neurons or the spinal cord. However, in this cancer-pain model, changes including massive astrocyte hypertrophy without neuronal loss, increase in the neuronal expression of c-Fos, and increase in the number of dynorphin-immunoreactive neurons were observed in the spinal cord, ipsilateral to the limb with cancer. These results indicate that a unique set of neurochemical changes occur with inflammatory, neuropathic and cancer pain in C3H/HeJ mice and further suggest that cancer induces a unique persistent pain state. Determining whether these neurochemical changes are involved in the generation and maintenance of each type of persistent pain may provide insight into the mechanisms that underlie each of these pain states.

Topics: Animals; Astrocytes; Axotomy; Behavior, Animal; Disease Models, Animal; Dynorphins; Fluorescent Antibody Technique; Freund's Adjuvant; Ganglia, Spinal; Male; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Neuralgia; Neuritis; Neurons, Afferent; Osteolysis; Pain; Palpation; Physical Stimulation; Proto-Oncogene Proteins c-fos; Receptors, Neurokinin-1; Sarcoma, Experimental; Sciatic Nerve; Spinal Cord; Spinal Nerves; Tumor Cells, Cultured

The co-administration of morphine at spinal (i.th.) and supraspinal (i.c.v.) sites to the same rat produces antinociceptive synergy, a phenomenon which may underlie the clinical analgesic utility of this drug. In animals with peripheral nerve injury, however, the antinociceptive potency and efficacy of i.th. morphine is significantly decreased. Here, the possible loss of spinal/supraspinal morphine antinociceptive synergy and relationship to elevation of spinal dynorphin content was studied. Ligation of lumbar spinal nerves resulted in elevated dynorphin in the ipsilateral lumbar and sacral spinal cord. In sham-operated rats supraspinal/spinal co-administration of morphine produced synergistic antinociception which was unaffected by i.th. MK-801 or dynorphin A((1-17)) antiserum. In nerve-injured rats, i.th. morphine was inactive against tactile allodynia and showed diminished in potency against acute nociception without supraspinal/spinal antinociceptive synergy. Antiserum to dynorphin A((1-17)) or the non-competitive NMDA antagonist MK-801 increased the antinociceptive potency of i.th. morphine, restored supraspinal/spinal morphine antinociceptive synergy and elicited a dose-related i.th. morphine antiallodynic action. These agents did not demonstrate antinociceptive or antiallodynic activity alone and did not alter morphine actions in sham-operated animals. The loss of spinal/supraspinal antinociceptive synergy and lack of antiallodynic activity of spinal morphine appear to be due to the elevation across multiple spinal segments of dynorphin following nerve injury. Pathological actions of elevated dynorphin may directly or indirectly modulate the NMDA receptor, result in a loss of supraspinal/spinal morphine synergy and may thus account for the decreased clinical analgesic efficacy of morphine in peripheral neuropathies.

Topics: Analgesics, Opioid; Animals; Dizocilpine Maleate; Drug Synergism; Dynorphins; Excitatory Amino Acid Antagonists; Immune Sera; Injections, Intraventricular; Injections, Spinal; Male; Morphine; Neuralgia; Pain Measurement; Peripheral Nerve Injuries; Rats; Rats, Sprague-Dawley; Touch

Neuropathic pains arising from peripheral nerve injury can result in increased sensitivity to both noxious and non-noxious stimuli and are accompanied by a number of neuroplastic alterations at the level of the spinal cord including upregulation of neurotransmitters including dynorphin, cholecystokinin and neuropeptide Y. Additionally, such pain states appear to be associated with activation of excitatory amino acid receptors including the N-methyl-D-aspartate (NMDA) receptor. Neuropathic pains have often been classified as 'opioid resistant' in both clinical and laboratory settings. As it is known that dynorphin produces 'non-opioid' effects through interaction with NMDA receptors and this peptide is upregulated after peripheral nerve injury, the present studies were undertaken to determine the possible importance of this substance in the neuropathic state. Nerve injury was produced in rats by tight ligation of the L5 and L6 spinal roots of the sciatic nerve. Catheters were inserted for the intrathecal (i.t.) delivery of drug to the lumbar spinal cord. Tactile allodynia was determined by measuring responses to probing the plantar surface of the affected limb with von Frey filaments, and acute nociception was determined in the 55 degrees C hot-water tail-flick test in nerve-ligated and sham-operated subjects. Intrathecal administration of MK-801 or antisera to dynorphin A (1-13) did not alter the tactile allodynia associated with nerve-ligation injury or the baseline tail-flick latency in either sham-operated or nerve-injured animals. As previously reported, i.t. morphine did not alter tactile allodynia and showed reduced potency and efficacy to block the tail-flick reflex in nerve-injured animals. Co-administration, however, of i.t. morphine with MK-801, or i.t. antisera to dynorphin A (1-13) given prior to morphine elicited both a full antiallodynic response and a complete block of the tail-flick reflex in nerve-injured animals. These results suggest that tonic activation of NMDA receptors, following peripheral nerve injury, is involved with the attenuation of the effectiveness of spinal morphine in a model of neuropathic pain. Additionally, this tonic NMDA activity may be mediated, in part, by increased levels of endogenous dynorphin associated with peripheral nerve injury.

Topics: Analgesics, Opioid; Animals; Constriction; Dizocilpine Maleate; Dynorphins; Excitatory Amino Acid Antagonists; Immune Sera; Injections, Spinal; Male; Morphine; Neuralgia; Pain Measurement; Peptide Fragments; Peripheral Nervous System Diseases; Rats; Rats, Sprague-Dawley; Touch