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Abstract Background Individuals who have experienced mild traumatic brain injuries (mTBIs) suffer from several comorbidities, including chronic pain. Despite extensive studies investigating the underlying mechanisms of mTBI-associated chronic pain, the role of inflammation in long-term pain after mTBIs is not fully elucidated. Given the shifting dynamics of inflammation, it is important to understand the spatial-longitudinal changes in inflammatory processes following mTBIs and their effects on TBI-related pain. Methods We utilized a recently developed transgenic caspase-1 luciferase reporter mouse model to monitor caspase-1 activation through a thinned skull window in the in vivo setting following three closed-head mTBI events. Organotypic coronal brain slice cultures and acutely dissociated dorsal root ganglion (DRG) cells provided tissue-relevant context of inflammation signal. Mechanical allodynia was assessed by mechanical withdrawal threshold to von Frey and thermal hyperalgesia withdrawal latency to radiant heat. Mouse grimace scale (MGS) was used to detect spontaneous or non-evoked pain. In some experiments, mice were prophylactically treated with MCC950, a potent small molecule inhibitor of NLRP3 inflammasome assembly to inhibit injury-induced inflammatory signaling. Bioluminescence spatiotemporal dynamics were quantified in the head and hind paws, and caspase-1 activation was confirmed by immunoblot. Immunofluorescence staining was used to monitor the progression of astrogliosis and microglial activation in ex vivo brain tissue following repetitive closed-head mTBIs. Results Mice with repetitive closed-head mTBIs exhibited significant increases of the bioluminescence signals within the brain and paws in vivo for at least one week after each injury. Consistently, immunoblotting and immunofluorescence experiments confirmed that mTBIs led to caspase-1 activation, astrogliosis, and microgliosis. Persistent changes in MGS and hind paw withdrawal thresholds, indicative of pain states, were observed post-injury in the same mTBI animals in vivo. We also observed enhanced inflammatory responses in ex vivo brain slice preparations and DRG for at least 3 days following mTBIs. In vivo treatment with MCC950 significantly reduced caspase-1 activation-associated bioluminescent signals in vivo and decreased stimulus-evoked and non-stimulus evoked nociception. Conclusions Our findings suggest that the inflammatory states in the brain and peripheral nervous system following repeated mTBIs are coincidental with the development of nociceptive sensitization, and that these events can be significantly reduced by inhibition of NLRP3 inflammasome activation.

Background: Individuals who have experienced mild traumatic brain injuries (mTBIs) suffer from several comorbidities, including chronic pain. Despite extensive studies investigating the underlying mechanisms of mTBI-associated chronic pain, the role of inflammation in long-term pain after mTBIs is not fully elucidated. Given the shifting dynamics of inflammation, it is important to understand the spatial-longitudinal changes in inflammatory processes following mTBIs and their effects on TBI-related pain. Methods: We utilized a recently developed transgenic caspase-1 luciferase reporter mouse model to monitor caspase-1 activation through a thinned skull window in the in vivo setting following three closed-head mTBI events. Organotypic coronal brain slice cultures and acutely dissociated dorsal root ganglion (DRG) cells provided tissue-relevant context of inflammation signal. Mechanical allodynia was assessed by mechanical withdrawal threshold to von Frey and thermal hyperalgesia withdrawal latency to radiant heat. Mouse grimace scale (MGS) was used to detect spontaneous or non-evoked pain. In some experiments, mice were prophylactically treated with MCC950, a potent small molecule inhibitor of NLRP3 inflammasome assembly to inhibit injury-induced inflammatory signaling. Bioluminescence spatiotemporal dynamics were quantified in the head and hind paws, and caspase-1 activation was confirmed by immunoblot. Immunofluorescence staining was used to monitor the progression of astrogliosis and microglial activation in ex vivo brain tissue following repetitive closed-head mTBIs. Results: Mice with repetitive closed-head mTBIs exhibited significant increases of the bioluminescence signals within the brain and paws in vivo for at least one week after each injury. Consistently, immunoblotting and immunofluorescence experiments confirmed that mTBIs led to caspase-1 activation, astrogliosis, and microgliosis. Persistent changes in MGS and hind paw withdrawal thresholds, indicative of pain states, were observed post-injury in the same mTBI animals in vivo. We also observed enhanced inflammatory responses in ex vivo brain slice preparations and DRG for at least 3 days following mTBIs. In vivo treatment with MCC950 significantly reduced caspase-1 activation-associated bioluminescent signals in vivo and decreased stimulus-evoked and non-stimulus evoked nociception. Conclusions: Our findings suggest that the inflammatory states in the brain and peripheral nervous system following repeated mTBIs are coincidental with the development of nociceptive sensitization, and that these events can be significantly reduced by inhibition of NLRP3 inflammasome activation. [ABSTRACT FROM AUTHOR]

It has been unexplained why chronic pain does not invariably accompany chronic pain-prone disorders. This question-driven, hypothesis-based article suggests that the reason may be varying occurrence of concomitant peripheral compressive proximal neural lesion (cPNL), e.g., radiculopathy and entrapment plexopathies. Transition of acute to chronic pain may involve development or aggravation of cPNL. Nociceptive hypersensitivity induced and/or maintained by cPNL may be responsible for all types of general chronic pain as well as for pain in isolated tissue conditions that are usually painless, e.g., neuroma, scar, and Dupuytren's fibromatosis. Compressive PNL induces focal neuroinflammation, which can maintain dorsal root ganglion neuron (DRGn) hyperexcitability (i.e., peripheral sensitization) and thus fuel central sensitization (i.e., hyperexcitability of central nociceptive pathways) and a vicious cycle of chronic pain. DRGn hyperexcitability and cPNL may reciprocally maintain each other, because cPNL can result from reflexive myospasm-induced myofascial tension, muscle weakness, and consequent muscle imbalance- and/or pain-provoked compensatory overuse. Because of pain and motor fiber damage, cPNL can worsen the causative musculoskeletal dysfunction, which further accounts for the reciprocity between the latter two factors. Sensitization increases nerve vulnerability and thus catalyzes this cycle. Because of these mechanisms and relatively greater number of neurons involved, cPNL is more likely to maintain DRGn hyperexcitability in comparison to distal neural and non-neural lesions. Compressive PNL is associated with restricted neural mobility. Intermittent (dynamic) nature of cPNL may be essential in chronic pain, because healed (i.e., fibrotic) lesions are physiologically silent and, consequently, cannot provide nociceptive input. Not all patients may be equally susceptible to develop cPNL, because occurrence of cPNL may vary as vary patients' predisposition to musculoskeletal impairment. Sensitization is accompanied by pressure pain threshold decrease and consequent mechanical allodynia and hyperalgesia, which can cause unusual local pain via natural pressure exerted by space occupying lesions or by their examination. Worsening of local pain is similarly explainable. Neuroma pain may be due to cPNL-induced axonal mechanical sensitivity and hypersensitivity of the nociceptive nervi nervorum of the nerve trunk and its stump. Intermittence and symptomatic complexity of cPNL may be the cause of frequent misdiagnosis of chronic pain.

Individual sensory neurons can be tuned to many stimuli, each driving unique, stimulus-relevant behaviors, and the ability of multimodal nociceptor neurons to discriminate between potentially harmful and innocuous stimuli is broadly important for organismal survival. Moreover, disruptions in the capacity to differentiate between noxious and innocuous stimuli can result in neuropathic pain. Drosophila larval class III (CIII) neurons are peripheral noxious cold nociceptors and innocuous touch mechanosensors; high levels of activation drive cold-evoked contraction (CT) behavior, while low levels of activation result in a suite of touch-associated behaviors. However, it is unknown what molecular factors underlie CIII multimodality. Here, we show that the TMEM16/anoctamins subdued and white walker (wwk; CG15270) are required for cold-evoked CT, but not for touch-associated behavior, indicating a conserved role for anoctamins in nociception. We also evidence that CIII neurons make use of atypical depolarizing chloride currents to encode cold, and that overexpression of ncc69—a fly homologue of NKCC1—results in phenotypes consistent with neuropathic sensitization, including behavioral sensitization and neuronal hyperexcitability, making Drosophila CIII neurons a candidate system for future studies of the basic mechanisms underlying neuropathic pain.

Injury occurring in the neonatal period in mammals is known to induce plasticity in pain pathways that may lead to pain dysfunction in later life. Whether these effects are unique to the mammalian nervous system is not well understood. Here, we investigate whether similar effects of early-life injury are found in a large-brained comparative model, the cephalopod Euprymna scolopes. We show that the peripheral nervous system of E. scolopes undergoes profound and permanent plasticity after injury of peripheral tissue in the early post-hatching period, but not after the same injury given in the later juvenile period. Additionally, both innate defensive behaviour and learning are impaired by injury in early life. We suggest that these similar patterns of nervous system and behavioural remodelling that occur in squid and in mammals indicate an adaptive value for long-lasting plasticity arising from early-life injury, and suggest that injuries inflicted in very early life may signal to the nervous system that the environment is highly dangerous. Thus, neonatal pain plasticity may be a conserved pattern whose purpose is to set the developing nervous system's baseline responsiveness to threat. This article is part of the Theo Murphy meeting issue 'Evolution of mechanisms and behaviour important for pain'. [ABSTRACT FROM AUTHOR]

Important insights into the selection pressures and core molecular modules contributing to the evolution of pain-related processes have come from studies of nociceptive systems in several molluscan and arthropod species. These phyla, and the chordates that include humans, last shared a common ancestor approximately 550 million years ago. Since then, animals in these phyla have continued to be subject to traumatic injury, often from predators, which has led to similar adaptive behaviors (e.g., withdrawal, escape, recuperative behavior) and physiological responses to injury in each group. Comparisons across these taxa provide clues about the contributions of convergent evolution and of conservation of ancient adaptive mechanisms to general nociceptive and pain-related functions. Primary nociceptors have been investigated extensively in a few molluscan and arthropod species, with studies of long-lasting nociceptive sensitization in the gastropod, Aplysia, and the insect, Drosophila, being especially fruitful. In Aplysia, nociceptive sensitization has been investigated as a model for aversive memory and for hyperalgesia. Neuromodulator-induced, activity-dependent, and axotomy-induced plasticity mechanisms have been defined in synapses, cell bodies, and axons of Aplysia primary nociceptors. Studies of nociceptive sensitization in Drosophila larvae have revealed numerous molecular contributors in primary nociceptors and interacting cells. Interestingly, molecular contributors examined thus far in Aplysia and Drosophila are largely different, but both sets overlap extensively with those in mammalian pain-related pathways. In contrast to results from Aplysia and Drosophila, nociceptive sensitization examined in moth larvae (Manduca) disclosed central hyperactivity but no obvious peripheral sensitization of nociceptive responses. Squid (Doryteuthis) show injury-induced sensitization manifested as behavioral hypersensitivity to tactile and especially visual stimuli, and as hypersensitivity and spontaneous activity in nociceptor terminals. Temporary blockade of nociceptor activity during injury subsequently increased mortality when injured squid were exposed to fish predators, providing the first demonstration in any animal of the adaptiveness of nociceptive sensitization. Immediate responses to noxious stimulation and nociceptive sensitization have also been examined behaviorally and physiologically in a snail (Helix), octopus (Adopus), crayfish (Astacus), hermit crab (Pagurus), and shore crab (Hemigrapsus). Molluscs and arthropods have systems that suppress nociceptive responses, but whether opioid systems play antinociceptive roles in these phyla is uncertain.

Diabetes-associated nociceptive hypersensitivity affects diabetic patients with hard-to-treat chronic pain. Because multiple tissues are affected by systemic alterations in insulin signaling, the functional locus of insulin signaling in diabetes-associated hypersensitivity remains obscure. Here, we used Drosophila nociception/nociceptive sensitization assays to investigate the role of Insulin receptor (Insulin-like receptor, InR) in nociceptive hypersensitivity. InR mutant larvae exhibited mostly normal baseline thermal nociception (absence of injury) and normal acute thermal hypersensitivity following UV-induced injury. However, their acute thermal hypersensitivity persists and fails to return to baseline, unlike in controls. Remarkably, injury-induced persistent hypersensitivity is also observed in larvae that exhibit either type 1 or type 2 diabetes. Cell type-specific genetic analysis indicates that InR function is required in multidendritic sensory neurons including nociceptive class IV neurons. In these same nociceptive sensory neurons, only modest changes in dendritic morphology were observed in the InRRNAi-expressing and diabetic larvae. At the cellular level, InR-deficient nociceptive sensory neurons show elevated calcium responses after injury. Sensory neuron-specific expression of InR rescues the persistent thermal hypersensitivity of InR mutants and constitutive activation of InR in sensory neurons ameliorates the hypersensitivity observed with a type 2-like diabetic state. Our results suggest that a sensory neuron-specific function of InR regulates the persistence of injury-associated hypersensitivity. It is likely that this new system will be an informative genetically tractable model of diabetes-associated hypersensitivity.

Important insights into the selection pressures and core molecular modules contributing to the evolution of pain-related processes have come from studies of nociceptive systems in several molluscan and arthropod species. These phyla, and the chordates that include humans, last shared a common ancestor approximately 550 million years ago. Since then, animals in these phyla have continued to be subject to traumatic injury, often from predators, which has led to similar adaptive behaviors (e.g., withdrawal, escape, recuperative behavior) and physiological responses to injury in each group. Comparisons across these taxa provide clues about the contributions of convergent evolution and of conservation of ancient adaptive mechanisms to general nociceptive and pain-related functions. Primary nociceptors have been investigated extensively in a few molluscan and arthropod species, with studies of long-lasting nociceptive sensitization in the gastropod, Aplysia, and the insect, Drosophila, being especially fruitful. In Aplysia, nociceptive sensitization has been investigated as a model for aversive memory and for hyperalgesia. Neuromodulator-induced, activity-dependent, and axotomy-induced plasticity mechanisms have been defined in synapses, cell bodies, and axons of Aplysia primary nociceptors. Studies of nociceptive sensitization in Drosophila larvae have revealed numerous molecular contributors in primary nociceptors and interacting cells. Interestingly, molecular contributors examined thus far in Aplysia and Drosophila are largely different, but both sets overlap extensively with those in mammalian pain-related pathways. In contrast to results from Aplysia and Drosophila, nociceptive sensitization examined in moth larvae (Manduca) disclosed central hyperactivity but no obvious peripheral sensitization of nociceptive responses. Squid (Doryteuthis) show injury-induced sensitization manifested as behavioral hypersensitivity to tactile and especially visual stimuli, and as hypersensitivity and spontaneous activity in nociceptor terminals. Temporary blockade of nociceptor activity during injury subsequently increased mortality when injured squid were exposed to fish predators, providing the first demonstration in any animal of the adaptiveness of nociceptive sensitization. Immediate responses to noxious stimulation and nociceptive sensitization have also been examined behaviorally and physiologically in a snail (Helix), octopus (Adopus), crayfish (Astacus), hermit crab (Pagurus), and shore crab (Hemigrapsus). Molluscs and arthropods have systems that suppress nociceptive responses, but whether opioid systems play antinociceptive roles in these phyla is uncertain. [ABSTRACT FROM AUTHOR]

Serotonin is a widely studied modulator of neural plasticity. Here we investigate the effect of fluoxetine, a selective serotonin reuptake inhibitor, on short-term, peripheral nociceptive plasticity in the neurologically complex invertebrate, octopus. After crush injury to isolated mantle (body wall) tissue, application of 10 nM fluoxetine increased spontaneous firing in crushed preparations, but had a minimal effect on mechanosensory sensitization. Effects largely did not persist after washout. We suggest that transiently elevated, endogenous serotonin may help promote initiation of longer-term plasticity of nociceptive afferents and drive immediate and spontaneous behaviors aimed at protecting wounds and escaping dangerous situations. [ABSTRACT FROM AUTHOR]

Pain signaling in vertebrates is modulated by neuropeptides like Substance P (SP). To determine whether such modulation is conserved and potentially uncover novel interactions between nociceptive signaling pathways we examined SP/Tachykinin signaling in a Drosophila model of tissue damage-induced nociceptive hypersensitivity. Tissue-specific knockdowns and genetic mutant analyses revealed that both Tachykinin and Tachykinin-like receptor (DTKR99D) are required for damage-induced thermal nociceptive sensitization. Electrophysiological recording showed that DTKR99D is required in nociceptive sensory neurons for temperature-dependent increases in firing frequency upon tissue damage. DTKR overexpression caused both behavioral and electrophysiological thermal nociceptive hypersensitivity. Hedgehog, another key regulator of nociceptive sensitization, was produced by nociceptive sensory neurons following tissue damage. Surprisingly, genetic epistasis analysis revealed that DTKR function was upstream of Hedgehog-dependent sensitization in nociceptive sensory neurons. Our results highlight a conserved role for Tachykinin signaling in regulating nociception and the power of Drosophila for genetic dissection of nociception.

Animals with detectable injuries are at escalated threat of predation. The anti-predation tactic of schooling reduces individual predation risk overall, but it is not known how schooling behavior affects injured animals, or whether risks are reduced equally for injured animals versus other school members. In this laboratory study we examined the effects of minor fin injury on schooling decisions made by squid. Schooling behavior of groups of squid, in which one member was injured, was monitored over 24 h. Injured squid were more likely to be members of a school shortly after injury (0.5–2 h), but there were no differences compared with sham-injured squid at longer time points (6–24 h). Overall, the presence of an injured conspecific increased the probability that a school would form, irrespective of whether the injured squid was a member of the school. When groups containing one injured squid were exposed to a predator cue, injured squid were more likely to join the school, but their position depended on whether the threat was a proximate visual cue or olfactory cue. We found no evidence that injured squid oriented themselves to conceal their injury from salient threats. Overall we conclude that nociceptive sensitization after injury changes grouping behaviors in ways that are likely to be adaptive. [ABSTRACT FROM AUTHOR]

It has been unexplained why chronic pain does not invariably accompany chronic pain-prone disorders. This question-driven, hypothesis-based article suggests that the reason may be varying occurrence of concomitant peripheral compressive proximal neural lesion (cPNL), e.g., radiculopathy and entrapment plexopathies. Transition of acute to chronic pain may involve development or aggravation of cPNL. Nociceptive hypersensitivity induced and/or maintained by cPNL may be responsible for all types of general chronic pain as well as for pain in isolated tissue conditions that are usually painless, e.g., neuroma, scar, and Dupuytren's fibromatosis. Compressive PNL induces focal neuroinflammation, which can maintain dorsal root ganglion neuron (DRGn) hyperexcitability (i.e., peripheral sensitization) and thus fuel central sensitization (i.e., hyperexcitability of central nociceptive pathways) and a vicious cycle of chronic pain. DRGn hyperexcitability and cPNL may reciprocally maintain each other, because cPNL can result from reflexive myospasm-induced myofascial tension, muscle weakness, and consequent muscle imbalance- and/or pain-provoked compensatory overuse. Because of pain and motor fiber damage, cPNL can worsen the causative musculoskeletal dysfunction, which further accounts for the reciprocity between the latter two factors. Sensitization increases nerve vulnerability and thus catalyzes this cycle. Because of these mechanisms and relatively greater number of neurons involved, cPNL is more likely to maintain DRGn hyperexcitability in comparison to distal neural and non-neural lesions. Compressive PNL is associated with restricted neural mobility. Intermittent (dynamic) nature of cPNL may be essential in chronic pain, because healed (i.e., fibrotic) lesions are physiologically silent and, consequently, cannot provide nociceptive input. Not all patients may be equally susceptible to develop cPNL, because occurrence of cPNL may vary as vary patients' predisposition to musculoskeletal impairment. Sensitization is accompanied by pressure pain threshold decrease and consequent mechanical allodynia and hyperalgesia, which can cause unusual local pain via natural pressure exerted by space occupying lesions or by their examination. Worsening of local pain is similarly explainable. Neuroma pain may be due to cPNL-induced axonal mechanical sensitivity and hypersensitivity of the nociceptive nervi nervorum of the nerve trunk and its stump. Intermittence and symptomatic complexity of cPNL may be the cause of frequent misdiagnosis of chronic pain., Competing Interests: The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Macionis.)

Individual sensory neurons can be tuned to many stimuli, each driving unique, stimulus-relevant behaviors, and the ability of multimodal nociceptor neurons to discriminate between potentially harmful and innocuous stimuli is broadly important for organismal survival. Moreover, disruptions in the capacity to differentiate between noxious and innocuous stimuli can result in neuropathic pain. Drosophila larval class III (CIII) neurons are peripheral noxious cold nociceptors and innocuous touch mechanosensors; high levels of activation drive cold-evoked contraction (CT) behavior, while low levels of activation result in a suite of touch-associated behaviors. However, it is unknown what molecular factors underlie CIII multimodality. Here, we show that the TMEM16/anoctamins subdued and white walker ( wwk; CG15270 ) are required for cold-evoked CT, but not for touch-associated behavior, indicating a conserved role for anoctamins in nociception. We also evidence that CIII neurons make use of atypical depolarizing chloride currents to encode cold, and that overexpression of ncc69 -a fly homologue of NKCC1 -results in phenotypes consistent with neuropathic sensitization, including behavioral sensitization and neuronal hyperexcitability, making Drosophila CIII neurons a candidate system for future studies of the basic mechanisms underlying neuropathic pain., Competing Interests: NH, AS, AP, SB, JL, MB, TG, GC, DC No competing interests declared, (© 2023, Himmel et al.)

Abstract: In recent years, evidence that sensitization of primary afferent nociceptors is an important event associated with chronic pain has been accumulating. The present study aimed to evaluate the participation of the prostaglandin and sympathetic components in the long-lasting sensitization of nociceptors induced by acute inflammation in mice. The intraplantar administration of carrageenan (100μg) enhanced the nociceptive response to a small dose of PGE2 (9ng/paw) or dopamine (3μg/paw) up to 30days later. This long-lasting sensitization is dependent on dopaminergic and prostanoid systems, since the pre-treatment with chlorpromazine (3μg/paw) or indomethacin (100μg/paw), but not local (6μg/paw) or systemic (6mg/kg) treatment with morphine, prevented its development. In agreement with this idea, the previous intraplantar administration of hyperalgesic doses of PGE2 or dopamine also induced long-lasting sensitization, which was fully prevented by pretreatment with EP4 and D1 antagonists, respectively. In summary, the present work described in mice a long-lasting sensitization of nociceptors, initiated by an acute inflammatory stimulation and dependent on dopaminergic and prostanoid systems. The present data represent new insights on the mechanisms of peripheral sensitization that could contribute to establish the basis of new therapeutic strategies for acute and chronic inflammatory pain. [Copyright &y& Elsevier]

Molluscan preparations have yielded seminal discoveries in neuroscience, but the experimental advantages of this group have not, until now, been complemented by adequate molecular or genomic information for comparisons to genetically defined model organisms in other phyla. The recent sequencing of the transcriptome and genome of Aplysia californica, however, will enable extensive comparative studies at the molecular level. Among other benefits, this will bring the power of individually identifiable and manipulable neurons to bear upon questions of cellular function for evolutionarily conserved genes associated with clinically important neural dysfunction. Because of the slower rate of gene evolution in this molluscan lineage, more homologs of genes associated with human disease are present in Aplysia than in leading model organisms from Arthropoda (Drosophila) or Nematoda (Caenorhabditis elegans). Research has hardly begun in molluscs on the cellular functions of gene products that in humans are associated with neurological diseases. On the other hand, much is known about molecular and cellular mechanisms of long-term neuronal plasticity. Persistent nociceptive sensitization of nociceptors in Aplysia displays many functional similarities to alterations in mammalian nociceptors associated with the clinical problem of chronic pain. Moreover, in Aplysia and mammals the same cell signaling pathways trigger persistent enhancement of excitability and synaptic transmission following noxious stimulation, and these highly conserved pathways are also used to induce memory traces in neural circuits of diverse species. This functional and molecular overlap in distantly related lineages and neuronal types supports the proposal that fundamental plasticity mechanisms important for memory, chronic pain, and other lasting alterations evolved from adaptive responses to peripheral injury in the earliest neurons. Molluscan preparations should become increasingly useful for comparative studies across phyla that can provide insight into cellular functions of clinically important genes. Copyright © 2009 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Experiments on snails showed that extracellular application of dibutyryl-cAMP (db-cAMP) or intracellular application of cAMP for 30 min evoked increases in excitability and synaptic facilitation in responses to sensory stimulation of defensive behavior command neurons LP11 and RP11. Extitability increased 45–60 min after the start of addition of db-cAMP or cAMP and remained elevated until the end of the experiment (3–4h). Synaptic facilitation started 50–60 min after the onset of extracellular application of db-cAMP and remained detectable in the responses of neurons to tactile stimulation of the head for 1 h and to application of dilute quinine solution for 2–4 h. Application of db-cAMP produced no changes in responses to tactile stimulation of the foot or mantle ridge. Intracellular injection of cAMP induced facilitation of neuron responses only to weak quinine solutions. The responses of neurons to tactile stimulation of the head, foot, and mantle ridge did not change after injections of cAMP. It is suggested that cAMP is involved in the mechanisms controlling the excitability of neurons LP11 and RP11. In addition, cAMP is selectively involved in the postsynaptic mechanism inducing the transient stage of long-term facilitation of synaptic “inputs”, which mediates excitation evoked by chemical stimuli. This set of effects of cAMP is similar to effects arising during the development of nociceptive sensitization and in response to serotonin. [ABSTRACT FROM AUTHOR]

• Understanding of chemosensory nociception and neuro-immune interactions. • Breathing patterns in rats exposed to upper, bronchial, and pulmonary irritants. • Alternative approach for estimating PODs from respiratory function data. • Hazard characterization and assessment for irritants causing nociceptor sensitization. Contemporary approaches to human health risk assessment for respiratory tract irritants are variable and controversial. This manuscript provides an in-depth analysis and assessment of the applicability of the classical respiratory depression 50 % (RD 50) assay with focus on the Log-linear extrapolation of the non-sensory irritant threshold (RD 0 or RD 10) relative to the contemporary Point of Departure (POD) U.S.-EPA benchmark approach. Three prototypic volatile chemically reactive irritants are used to exemplify the pros and cons of this alternative approach. These irritants differ in physicochemical properties affecting water-solubility and lipophilicity. Depending on these variables, a vapor may preferentially be retained in the extrathoracic region (ET), the tracheobronchial region (TB), and the pulmonary region (PU); although a smooth transition between these regions occurs at increasingly high concentrations. Each region has its specific nociceptors sensing irritants and regional-specific response to injury. The alternative approach using rats identified the chemical-specific critical region of respiratory tract injury. Statistically derived PODs on ET-TB related sensory irritation provide important information for ET-TB irritants but not for PU irritants. The POD of ET-TB irritants from acute and repeated studies decreased substantially. In summary, statistically derived PODs improve the risk assessment of respiratory tract irritants; however, those from repeated exposures should be given preference to those from acute exposures. [ABSTRACT FROM AUTHOR]

Abstract Background Kisspeptin is a neuropeptide known for its role in the hypothalamic regulation of the reproductive axis. Following the recent description of kisspeptin and its 7-TM receptor, GPR54, in the dorsal root ganglia and dorsal horns of the spinal cord, we examined the role of kisspeptin in the regulation of pain sensitivity in mice. Results Immunofluorescent staining in the mouse skin showed the presence of GPR54 receptors in PGP9.5-positive sensory fibers. Intraplantar injection of kisspeptin (1 or 3 nmol/5 μl) induced a small nocifensive response in naive mice, and lowered thermal pain threshold in the hot plate test. Both intraplantar and intrathecal (0.5 or 1 nmol/3 μl) injection of kisspeptin caused hyperalgesia in the first and second phases of the formalin test, whereas the GPR54 antagonist, p234 (0.1 or 1 nmol), caused a robust analgesia. Intraplantar injection of kisspeptin combined with formalin enhanced TRPV1 phosphorylation at Ser800 at the injection site, and increased ERK1/2 phosphorylation in the ipsilateral dorsal horn as compared to naive mice and mice treated with formalin alone. Conclusion These data demonstrate for the first time that kisspeptin regulates pain sensitivity in rodents and suggest that peripheral GPR54 receptors could be targeted by novel drugs in the treatment of inflammatory pain.

Introduction: Central sensitization (CS) was first defined in animal studies to be increased nociceptive responsiveness due to sensitization of neurons in the central nervous system, usually the result of prolonged nociceptive input or a disease state. Recently, the concept of CS has been adopted in clinical assessments of chronic pain, but its diagnosis in humans has expanded to include the enhancement of a wide range of nociceptive, sensory, and emotional responses. Many poorly understood pain disorders are referred to as "central sensitivity syndrome," a term associated with a broad range of hypervigilant sensory and emotional responses. Diagnosis often involves a review of medical records and an assessment of behaviour, emotional disposition, and overall sensitivity of a patient. Obviously, these assessments are unable to directly capture the responsiveness of nociceptive neurons. The purpose of this review is to ascertain whether self-report questionnaires associated with central sensitization and the diagnosis of central sensitivity syndrome are associated with enhanced nociceptive responses or whether they more validly measure sensitivity in a broader sense (ie, including emotional responses)., Methods: Following the PRISMA guidelines, a detailed search of studies that involve the Central Sensitization Inventory or Pain Sensitivity Questionnaire correlated with either nociceptive sensory tests (quantitative sensory testing) or emotional hypervigilance (anxiety, depression, stress, etc) will be conducted on MEDLINE, PsychINFO, and Web of Science., Perspective: The review is expected to synthesize correlations between sensitivity questionnaires and nociceptive or emotional sensitivity to determine whether these questionnaires reflect a broadened understanding of the term "central sensitization.", Competing Interests: The authors have no conflict of interest to declare.Sponsorships or competing interests that may be relevant to content are disclosed at the end of this article., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of The International Association for the Study of Pain.)

N-Acetylcysteine, one of the most prescribed antioxidant drugs, enhances pain threshold in rodents and humans by activating mGlu2 metabotropic glutamate receptors. Here, we assessed the analgesic activity of N-acetylcysteine in the streptozotocin model of painful diabetic neuropathy and examined the effect of N-acetylcysteine on proteins that are involved in mechanisms of nociceptive sensitization. Mice with blood glucose levels ≥250 mg/dl in response to a single intraperitoneal (i.p.) injection of streptozotocin (200 mg/kg) were used for the assessment of mechanical pain thresholds. Systemic treatment with N-acetylcysteine (100 mg/kg, i.p., either single injection or daily injections for seven days) caused analgesia in diabetic mice. N-acetylcysteine-induced analgesia was abrogated by the Sx c − inhibitors, sulfasalazine (8 mg/kg, i.p.), erastin (30 mg/kg, i.p.), and sorafenib (10 mg/kg, i.p.), or by the mGlu2/3 receptor antagonist, LY341495 (1 mg/kg, i.p.). Repeated administrations of N-acetylcysteine in diabetic mice reduced ERK1/2 phosphorylation in the dorsal region of the lumbar spinal cord. The analgesic activity of N-acetylcysteine was occluded by the MEK inhibitor, PD0325901 (25 mg/kg, i.p.), the TRPV1 channel blocker, capsazepine (40 mg/kg, i.p.), or by a cocktail of NMDA and mGlu5 metabotropic glutamate receptor antagonists (memantine, 25 mg/kg, plus MTEP, 5 mg/kg, both i.p.). These findings offer the first demonstration that N-acetylcysteine relieves pain associated with diabetic neuropathy and holds promise for the use of N-acetylcysteine as an add-on drug in diabetic patients. [ABSTRACT FROM AUTHOR]

Diabetes-associated nociceptive hypersensitivity affects diabetic patients with hard-to-treat chronic pain. Because multiple tissues are affected by systemic alterations in insulin signaling, the functional locus of insulin signaling in diabetes-associated hypersensitivity remains obscure. Here, we used Drosophila nociception/nociceptive sensitization assays to investigate the role of Insulin receptor (Insulin-like receptor, InR) in nociceptive hypersensitivity. InR mutant larvae exhibited mostly normal baseline thermal nociception (absence of injury) and normal acute thermal hypersensitivity following UV-induced injury. However, their acute thermal hypersensitivity persists and fails to return to baseline, unlike in controls. Remarkably, injury-induced persistent hypersensitivity is also observed in larvae that exhibit either type 1 or type 2 diabetes. Cell type-specific genetic analysis indicates that InR function is required in multidendritic sensory neurons including nociceptive class IV neurons. In these same nociceptive sensory neurons, only modest changes in dendritic morphology were observed in the InR RNAi -expressing and diabetic larvae. At the cellular level, InR -deficient nociceptive sensory neurons show elevated calcium responses after injury. Sensory neuron-specific expression of InR rescues the persistent thermal hypersensitivity of InR mutants and constitutive activation of InR in sensory neurons ameliorates the hypersensitivity observed with a type 2-like diabetic state. Our results suggest that a sensory neuron-specific function of InR regulates the persistence of injury-associated hypersensitivity. It is likely that this new system will be an informative genetically tractable model of diabetes-associated hypersensitivity., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2018. Published by The Company of Biologists Ltd.)

Pain signaling in vertebrates is modulated by neuropeptides like Substance P (SP). To determine whether such modulation is conserved and potentially uncover novel interactions between nociceptive signaling pathways we examined SP/Tachykinin signaling in a Drosophila model of tissue damage-induced nociceptive hypersensitivity. Tissue-specific knockdowns and genetic mutant analyses revealed that both Tachykinin and Tachykinin-like receptor (DTKR99D) are required for damage-induced thermal nociceptive sensitization. Electrophysiological recording showed that DTKR99D is required in nociceptive sensory neurons for temperature-dependent increases in firing frequency upon tissue damage. DTKR overexpression caused both behavioral and electrophysiological thermal nociceptive hypersensitivity. Hedgehog, another key regulator of nociceptive sensitization, was produced by nociceptive sensory neurons following tissue damage. Surprisingly, genetic epistasis analysis revealed that DTKR function was upstream of Hedgehog-dependent sensitization in nociceptive sensory neurons. Our results highlight a conserved role for Tachykinin signaling in regulating nociception and the power of Drosophila for genetic dissection of nociception.