Studying Independent Kcna6 Knock-out Mice Reveals Toxicity of Exogenous LacZ to Central Nociceptor Terminals and Differential Effects of Kv1.6 on Acute and Neuropathic Pain Sensation.

The potassium channel Kv1.6 has recently been implicated as a major modulatory channel subunit expressed in primary nociceptors. Furthermore, its expression at juxtaparanodes of myelinated primary afferents is induced following traumatic nerve injury as part of an endogenous mechanism to reduce hyperexcitability and pain-related hypersensitivity. In this study, we compared two mouse models of constitutive Kv1.6 knock-out (KO) achieved by different methods: traditional gene trap via homologous recombination and CRISPR-mediated excision. Both Kv1.6 KO mouse lines exhibited an unexpected reduction in sensitivity to noxious heat stimuli, to differing extents: the Kv1.6 mice produced via gene trap had a far more significant hyposensitivity. These mice (Kcna6lacZ ) expressed the bacterial reporter enzyme LacZ in place of Kv1.6 as a result of the gene trap mechanism, and we found that their central primary afferent presynaptic terminals developed a striking neurodegenerative phenotype involving accumulation of lipid species, development of "meganeurites," and impaired transmission to dorsal horn wide dynamic range neurons. The anatomic defects were absent in CRISPR-mediated Kv1.6 KO mice (Kcna6-/-) but were present in a third mouse model expressing exogenous LacZ in nociceptors under the control of a Nav1.8-promoted Cre recombinase. LacZ reporter enzymes are thus intrinsically neurotoxic to sensory neurons and may induce pathologic defects in transgenic mice, which has confounding implications for the interpretation of gene KOs using lacZ Nonetheless, in Kcna6-/- mice not affected by LacZ, we demonstrated a significant role for Kv1.6 regulating acute noxious thermal sensitivity, and both mechanical and thermal pain-related hypersensitivity after nerve injury.SIGNIFICANCE STATEMENT In recent decades, the expansion of technologies to experimentally manipulate the rodent genome has contributed significantly to the field of neuroscience. While introduction of enzymatic or fluorescent reporter proteins to label neuronal populations is now commonplace, often potential toxicity effects are not fully considered. We show a role of Kv1.6 in acute and neuropathic pain states through analysis of two mouse models lacking Kv1.6 potassium channels: one with additional expression of LacZ and one without. We show that LacZ reporter enzymes induce unintended defects in sensory neurons, with an impact on behavioral data outcomes. To summarize we highlight the importance of Kv1.6 in recovery of normal sensory function following nerve injury, and careful interpretation of data from LacZ reporter models.

A study of cortical and brainstem mechanisms of diffuse noxious inhibitory controls in anaesthetised normal and neuropathic rats.

Diffuse noxious inhibitory controls (DNIC) are a mechanism of endogenous descending pain modulation and are deficient in a large proportion of chronic pain patients. However, the pathways involved remain only partially determined with several cortical and brainstem structures implicated. This study examined the role of the dorsal reticular nucleus (DRt) and infralimbic (ILC) region of the medial prefrontal cortex in DNIC. In vivo electrophysiology was performed to record from dorsal horn lamina V/VI wide dynamic range neurones with left hind paw receptive fields in anaesthetised sham-operated and L5/L6 spinal nerve-ligated (SNL) rats. Evoked neuronal responses were quantified in the presence and absence of a conditioning stimulus (left ear clamp). In sham rats, DNIC were reproducibly recruited by a heterotopically applied conditioning stimulus, an effect that was absent in neuropathic rats. Intra-DRt naloxone had no effect on spinal neuronal responses to dynamic brush, punctate mechanical, evaporative cooling and heat stimuli in sham and SNL rats. In addition, intra-DRt naloxone blocked DNIC in sham rats, but had no effect in SNL rats. Intra-ILC lidocaine had no effect on spinal neuronal responses to dynamic brush, punctate mechanical, evaporative cooling and heat stimuli in sham and SNL rats. However, differential effects were observed in relation to the expression of DNIC; intra-ILC lidocaine blocked activation of DNIC in sham rats but restored DNIC in SNL rats. These data suggest that the ILC is not directly involved in mediating DNIC but can modulate its activation and that DRt involvement in DNIC requires opioidergic signalling.

Supraspinal Opioid Circuits Differentially Modulate Spinal Neuronal Responses in Neuropathic Rats.

BACKGROUND: The anterior cingulate cortex and central nucleus of the amygdala connect widely with brainstem nuclei involved in descending modulation, including the rostral ventromedial medulla. Endogenous opioids in these circuits participate in pain modulation. The hypothesis was that a differential opioidergic role for the brain nuclei listed in regulation of spinal neuronal responses because separable effects on pain behaviors in awake animals were previously observed. METHODS: This study utilized in vivo electrophysiology to determine the effects of morphine microinjection into the anterior cingulate cortex, right or left central nucleus of the amygdala, or the rostral ventromedial medulla on spinal wide dynamic range neuronal responses in isoflurane-anesthetized, male Sprague-Dawley rats. Ongoing activity in the ventrobasal thalamus was also measured. In total, 33 spinal nerve ligated and 26 control age- and weight-matched control rats were used. RESULTS: Brainstem morphine reduced neuronal firing to 60-g von Frey stimulation in control rats (to 65 ± 12% of control response (means ± 95% CI), P < 0.001) with a greater inhibition in neuropathic rats (to 53 ± 17% of control response, P < 0.001). Contrasting anterior cingulate cortex morphine had only marginal modulatory effects on spinal neuronal responses with limited variance in effect between control and neuropathic rats. The inhibitory effects of morphine in the central nucleus of the amygdala were dependent on pain state and laterality; only right-side morphine reduced neuronal firing to 60-g stimulation in neuropathic rats (to 65 ± 14% of control response, P = 0.001). In addition, in neuropathic rats elevated ongoing neuronal activity in the ventral posterolateral thalamus was not inhibited by anterior cingulate cortex morphine, in contrast to evoked responses. CONCLUSIONS: Cumulatively the data support opioid modulation of evoked responses predominately through a lateralized output from the right amygdala, as well as from the brainstem that is enhanced in injured conditions. Minimal modulation of dorsal horn responses was observed after anterior cingulate cortex opioid administration regardless of injury state.

Osteochondroma of the Zygomatic Arch: A Case Report and Review of the Literature.

An osteochondroma, when reported in the maxillofacial region, is a benign neoplasm that involves the skull base, maxillary sinus, zygomatic arch, or mandible. Most commonly, the osteochondroma occurs in the coronoid process and the condyle. One rare subtype of osteochondroma reported in the literature, termed Jacob disease, arises from the coronoid process and interferes with the zygomatic arch. This report describes a unique case of an isolated osteochondroma arising from the zygomatic arch and interfering with the coronoid process, which was treated through surgical excision by an intraoral approach. The literature search indicated that this is 1 among only 9 other reported cases since 1964 in which an osteochondroma arose primarily from the zygomatic arch.

Reported community-level indoor residual spray coverage from two-stage cluster surveys in sub-Saharan Africa.

BACKGROUND: Malaria is an important cause of morbidity and mortality in malaria-endemic areas. Indoor residual spray is an effective intervention to control malaria, but high community-level coverage is needed to maximize its impact. METHODS AND RESULTS: Using thirty-four two-stage cluster surveys (e.g., demographic and health surveys) and lot quality assurance sampling, indoor residual spray was estimated at the community level (i.e. enumeration-area) across sub-Saharan Africa since 2010. For communities receiving indoor residual spray a logistic regression predicted whether community-level coverage exceeded 50% or not. Household-level coverage was equitable both in terms of wealth and urban/rural, with poorer and rural houses more likely to be sprayed than richer and urban houses. Coverage of indoor residual spray at the community level is poor across the continent, with 54% of communities receiving the intervention not reaching 50% coverage. Having >50% coverage at the community-level was not associated with increasing the number of houses sprayed in the country. CONCLUSIONS: Implementation and monitoring of indoor residual coverage at small geographical scales need to improve greatly to receive maximum benefit of the intervention.

Neuronal hyperexcitability in the ventral posterior thalamus of neuropathic rats: modality selective effects of pregabalin.

Neuropathic pain represents a substantial clinical challenge; understanding the underlying neural mechanisms and back-translation of therapeutics could aid targeting of treatments more effectively. The ventral posterior thalamus (VP) is the major termination site for the spinothalamic tract and relays nociceptive activity to the somatosensory cortex; however, under neuropathic conditions, it is unclear how hyperexcitability of spinal neurons converges onto thalamic relays. This study aimed to identify neural substrates of hypersensitivity and the influence of pregabalin on central processing. In vivo electrophysiology was performed to record from VP wide dynamic range (WDR) and nociceptive-specific (NS) neurons in anesthetized spinal nerve-ligated (SNL), sham-operated, and naive rats. In neuropathic rats, WDR neurons had elevated evoked responses to low- and high-intensity punctate mechanical stimuli, dynamic brushing, and innocuous and noxious cooling, but less so to heat stimulation, of the receptive field. NS neurons in SNL rats also displayed increased responses to noxious punctate mechanical stimulation, dynamic brushing, noxious cooling, and noxious heat. Additionally, WDR, but not NS, neurons in SNL rats exhibited substantially higher rates of spontaneous firing, which may correlate with ongoing pain. The ratio of WDR-to-NS neurons was comparable between SNL and naive/sham groups, suggesting relatively few NS neurons gain sensitivity to low-intensity stimuli leading to a "WDR phenotype." After neuropathy was induced, the proportion of cold-sensitive WDR and NS neurons increased, supporting the suggestion that changes in frequency-dependent firing and population coding underlie cold hypersensitivity. In SNL rats, pregabalin inhibited mechanical and heat responses but not cold-evoked or elevated spontaneous activity.

Ionic mechanisms of spinal neuronal cold hypersensitivity in ciguatera.

Cold hypersensitivity is evident in a range of neuropathies and can evoke sensations of paradoxical burning cold pain. Ciguatoxin poisoning is known to induce a pain syndrome caused by consumption of contaminated tropical fish that can persist for months and include pruritus and cold allodynia; at present no suitable treatment is available. This study examined, for the first time, the neural substrates and molecular components of Pacific ciguatoxin-2-induced cold hypersensitivity. Electrophysiological recordings of dorsal horn lamina V/VI wide dynamic range neurones were made in non-sentient rats. Subcutaneous injection of 10 nm ciguatoxin-2 into the receptive field increased neuronal responses to innocuous and noxious cooling. In addition, neuronal responses to low-threshold but not noxious punctate mechanical stimuli were also elevated. The resultant cold hypersensitivity was not reversed by 6-({2-[2-fluoro-6-(trifluoromethyl)phenoxy]-2-methylpropyl}carbamoyl)pyridine-3-carboxylic acid, an antagonist of transient receptor potential melastatin 8 (TRPM8). Both mechanical and cold hypersensitivity were completely prevented by co-injection with the Nav 1.8 antagonist A803467, whereas the transient receptor potential ankyrin 1 (TRPA1) antagonist A967079 only prevented hypersensitivity to innocuous cooling and partially prevented hypersensitivity to noxious cooling. In naive rats, neither innocuous nor noxious cold-evoked neuronal responses were inhibited by antagonists of Nav 1.8, TRPA1 or TRPM8 alone. Ciguatoxins may confer cold sensitivity to a subpopulation of cold-insensitive Nav 1.8/TRPA1-positive primary afferents, which could underlie the cold allodynia reported in ciguatera. These data expand the understanding of central spinal cold sensitivity under normal conditions and the role of these ion channels in this translational rat model of ciguatoxin-induced hypersensitivity.

α2δ-1 gene deletion affects somatosensory neuron function and delays mechanical hypersensitivity in response to peripheral nerve damage.

The α2δ-1 subunit of voltage-gated calcium channels is upregulated after sensory nerve injury and is also the therapeutic target of gabapentinoid drugs. It is therefore likely to play a key role in the development of neuropathic pain. In this study, we have examined mice in which α2δ-1 gene expression is disrupted, to determine whether α2δ-1 is involved in various modalities of nociception, and for the development of behavioral hypersensitivity after partial sciatic nerve ligation (PSNL). We find that naive α2δ-1(-/-) mice show a marked behavioral deficit in mechanical and cold sensitivity, but no change in thermal nociception threshold. The lower mechanical sensitivity is mirrored by a reduced in vivo electrophysiological response of dorsal horn wide dynamic range neurons. The CaV2.2 level is reduced in brain and spinal cord synaptosomes from α2δ-1(-/-) mice, and α2δ-1(-/-) DRG neurons exhibit lower calcium channel current density. Furthermore, a significantly smaller number of DRG neurons respond to the TRPM8 agonist menthol. After PSNL, α2δ-1(-/-) mice show delayed mechanical hypersensitivity, which only develops at 11 d after surgery, whereas in wild-type littermates it is maximal at the earliest time point measured (3 d). There is no compensatory upregulation of α2δ-2 or α2δ-3 after PSNL in α2δ-1(-/-) mice, and other transcripts, including neuropeptide Y and activating transcription factor-3, are upregulated normally. Furthermore, the ability of pregabalin to alleviate mechanical hypersensitivity is lost in PSNL α2δ-1(-/-) mice. Thus, α2δ-1 is essential for rapid development of mechanical hypersensitivity in a nerve injury model of neuropathic pain.

Novel TRPM8 antagonist attenuates cold hypersensitivity after peripheral nerve injury in rats.

Abnormal cold sensitivity is a common feature of a range of neuropathies. In the murine somatosensory system, multiple aspects of cold sensitivity are dependent on TRPM8, both short term and in response to peripheral nerve injury. The specialized nature of cold-sensitive afferents and the restricted expression of TRPM8 render it an attractive target for the treatment of cold hypersensitivity. This current study examines the effect of a novel TRPM8 antagonist (M8-An) in naive and spinal nerve-ligated rats through behavioral and in vivo electrophysiological approaches. In vitro, M8-An inhibited icilin-evoked Ca(2+) currents in HEK293 cells stably expressing human TRPM8 with an IC(50) of 10.9 nM. In vivo, systemic M8-An transiently decreased core body temperature. Deep dorsal horn recordings were made in vivo from neurons innervating the hind paw. M8-An inhibited neuronal responses to innocuous and noxious cooling of the receptive field in spinal nerve-ligated rats but not in naive rats. No effect on neuronal responses to mechanical and heat stimulation was observed. In addition, M8-An also attenuated behavioral responses to cold but not mechanical stimulation after nerve ligation without affecting the uninjured contralateral response. The data presented here support a contribution of TRPM8 to the pathophysiology of cold hypersensitivity in this model and highlight the potential of the pharmacological block of TRPM8 in alleviating the associated symptoms.

A back-translational study of descending interactions with the induction of hyperalgesia by high-frequency electrical stimulation in rat and human.

ABSTRACT: In humans and animals, high-frequency electrocutaneous stimulation (HFS) induces an "early long-term potentiation-like" sensitisation, where synaptic plasticity is underpinned by an ill-defined interaction between peripheral input and central modulatory processes. The relative contributions of these processes to the initial pain or nociceptive response likely differ from those that underpin development of the heightened response. To investigate the impact of HFS-induced hyperalgesia on pain and nociception in perception and neural terms, respectively, and to explore the impact of descending inhibitory pathway activation on the development of HFS-induced hyperalgesia, we performed parallel studies utilising identical stimuli to apply HFS concurrent to (1) a conditioned pain modulation paradigm during psychophysical testing in healthy humans or (2) a diffuse noxious inhibitory controls paradigm during in vivo electrophysiological recording of spinal neurones in healthy anaesthetised rats. High-frequency electrocutaneous stimulation alone induced enhanced perceptual responses to pinprick stimuli in cutaneous areas secondary to the area of electrical stimulation in humans and increased the excitability of spinal neurones which exhibited stimulus intensity-dependent coded responses to pinprick stimulation in a manner that tracked with human psychophysics, supporting their translational validity. Application of a distant noxious conditioning stimulus during HFS did not alter perceived primary or secondary hyperalgesia in humans or the development of primary or secondary neuronal hyperexcitability in rats compared with HFS alone, suggesting that, upon HFS-response initiation in a healthy nervous system, excitatory signalling escapes inhibitory control. Therefore, in this model, dampening facilitatory mechanisms rather than augmenting top-down inhibitions could prevent pain development.

The circuit basis for chronic pain and its comorbidities.

PURPOSE OF REVIEW: Chronic pain is poorly treated with many developing disabling comorbidities such as anxiety, depression and insomnia. Considerable evidence supports the idea that pain and anxiodepressive disorders share a common neurobiology and can mutually reinforce, which has significant long-term implications as the development of comorbidities leads to poorer treatment outcomes for both pain and mood disorders. This article will review recent advances in the understanding of the circuit basis for comorbidities in chronic pain. RECENT FINDINGS: A growing number of studies have aimed to determine the mechanisms underlying chronic pain and comorbid mood disorders by using modern viral tracing tools for precise circuit manipulation with optogenetics and chemogenetics. These have revealed critical ascending and descending circuits, which advance the understanding of the interconnected pathways that modulate the sensory dimension of pain and the long-term emotional consequences of chronic pain. SUMMARY: Comorbid pain and mood disorders can produce circuit-specific maladaptive plasticity; however, several translational issues require addressing to maximise future therapeutic potential. These include the validity of preclinical models, the translatability of endpoints and expanding analysis to the molecular and system levels.

Nerve injury increases native Ca V 2.2 trafficking in dorsal root ganglion mechanoreceptors.

ABSTRACT: Neuronal N-type (Ca V 2.2) voltage-gated calcium channels are essential for neurotransmission from primary afferent terminals in the dorsal horn. In this study, we have used a knockin mouse containing Ca V 2.2 with an inserted extracellular hemagglutinin tag (Ca V 2.2_HA), to visualise the pattern of expression of endogenous Ca V 2.2 in dorsal root ganglion (DRG) neurons and their primary afferents in the dorsal horn. We examined the effect of partial sciatic nerve ligation (PSNL) and found an increase in Ca V 2.2_HA only in large and medium dorsal root ganglion neurons and also in deep dorsal horn synaptic terminals. Furthermore, there is a parallel increase in coexpression with GFRα1, present in a population of low threshold mechanoreceptors, both in large DRG neurons and in their terminals. The increased expression of Ca V 2.2_HA in these DRG neurons and their terminals is dependent on the presence of the auxiliary subunit α 2 δ-1, which is required for channel trafficking to the cell surface and to synaptic terminals, and it likely contributes to enhanced synaptic transmission at these synapses following PSNL. By contrast, the increase in GFRα1 is not altered in α 2 δ-1-knockout mice. We also found that following PSNL, there is patchy loss of glomerular synapses immunoreactive for Ca V 2.2_HA and CGRP or IB4, restricted to the superficial layers of the dorsal horn. This reduction is not dependent on α 2 δ-1 and likely reflects partial deafferentation of C-nociceptor presynaptic terminals. Therefore, in this pain model, we can distinguish 2 different events affecting specific DRG terminals, with opposite consequences for Ca V 2.2_HA expression and function in the dorsal horn.

Sagittal synostosis: does choice of intervention and its timing affect the long-term aesthetic and neurodevelopmental outcome? A single-institution study of 167 children.

OBJECTIVE: Sagittal craniosynostosis (SC) is the most commonly encountered form of craniosynostosis. Despite its relative frequency, there remains significant heterogeneity in both operative management and follow-up between centers and a relative paucity of long-term outcome data in the literature. At the authors' institution, families of children presenting with SC are offered the following options: 1) conservative management with ophthalmic surveillance, 2) minimally invasive surgery at < 6 months of age (spring-assisted cranioplasty [SAC]) or 3) calvarial vault remodeling at any age (CVR). The authors reviewed outcomes for all children presenting with SC during a 5-year period, regardless of the treatment received. METHODS: Consecutive children born between January 1, 2008, and December 31, 2012, presenting with SC were identified, and detailed chart reviews were undertaken. Demographic, surgical, perioperative, head shape, scar, and neurodevelopmental (behavioral, education, speech, and language) data were analyzed. The cohort was divided by type of surgery (none, SAC, or CVR) and by age at surgery (early, defined as ≤ 6 months; or late, defined as > 6 months) for comparison purposes. RESULTS: A total of 167 children were identified, 129 boys and 38 girls, with a median age at presentation of 5.0 (range 0.4-135) months. Three families opted for conservative management. Of the 164 children who underwent surgery, 83 underwent SAC, 76 underwent CVR, and 5 underwent a "hybrid" procedure (CVR with springs). At a median age of 7.0 (range 0.5-12.3) years, there was no significant difference in concerns regarding head shape, scar, or neurodevelopmental outcomes between the early and late intervention groups over all procedures performed, or between the early or late SAC and CVR cohorts. There were more head shape concerns in the SAC group than in the CVR group overall (25.7% vs 11.8%, respectively; p = 0.026), although most of these concerns were minor and did not require revision. CONCLUSIONS: In this cohort, regardless of operative intervention and timing of intervention, infants achieved similar neurodevelopmental outcomes. Minimally invasive surgery (SAC) appears to result in less complete correction of head shape than CVR, but this may be balanced by advantages in reduced operative time, hospitalization, and blood loss. SAC was equal to CVR in neuropsychological outcomes.

Neuropharmacological basis for multimodal analgesia in chronic pain.

Managing chronic pain remains a major unmet clinical challenge. Patients can be treated with a range of interventions, but pharmacotherapy is the most common. These include opioids, antidepressants, calcium channel modulators, sodium channel blockers, and nonsteroidal anti-inflammatory drugs. Many of these drugs target a particular mechanism; however, chronic pain in many diseases is multifactorial and induces plasticity throughout the sensory neuroaxis. Furthermore, comorbidities such as depression, anxiety, and sleep disturbances worsen quality of life. Given the complexity of mechanisms and symptoms in patients, it is unsurprising that many fail to achieve adequate pain relief from a single agent. The efforts to develop novel drug classes with better efficacy have not always proved successful; a multimodal or combination approach to analgesia is an important strategy in pain control. Many patients frequently take more than one medication, but high-quality evidence to support various combinations is often sparse. Ideally, combining drugs would produce synergistic action to maximize analgesia and reduce side effects, although sub-additive and additive analgesia is still advantageous if additive side-effects can be avoided. In this review, we discuss pain mechanisms, drug actions, and the rationale for mechanism-led treatment selection.Abbreviations: COX - cyclooxygenase, CGRP - calcitonin gene-related peptide, CPM - conditioned pain modulation, NGF - nerve growth factor, NNT - number needed to treat, NMDA - N-methyl-d-aspartate, NSAID - nonsteroidal anti-inflammatory drugs, TCA - tricyclic antidepressant, SNRI - serotonin-noradrenaline reuptake inhibitor, QST - quantitative sensory testing.

Physiological CTG categorization in types of hypoxia compared with MRI and neurodevelopmental outcome in infants with HIE.

BACKGROUND: Commonly used methods of CTG classification do not reliably predict neonatal hypoxic-ischemic encephalopathy (HIE). OBJECTIVE: To examine whether a relationship exists between the types of hypoxia as identified on the cardiotocograph using novel physiology-based CTG classification and patterns of injury on neonatal cerebral MRI and later neurodevelopmental outcomes. STUDY DESIGN: A retrospective study of term-born infants admitted to four neonatal units with HIE as part of a brain injury biomarkers study between January 2014 and December 2015. Intrapartum CTG traces were analyzed by two obstetricians trained in physiological CTG classification, blind to neonatal outcomes. Neonatal cerebral MR images were assessed independently by a neuroradiologist and an expert neonatologist. CTG traces were classified into types of hypoxia and allocated to groups; (1) chronic hypoxia or antepartum injury; (2) gradually evolving or subacute hypoxia; and (3) acute hypoxia. RESULTS: Of 106 infants recruited to the study, records were available for 58 cases. Of these, CTGs were available for 37. All 37 had abnormal CTGs. Twenty-four infants, all of whom had received therapeutic hypothermia had cerebral MRI. Fourteen of the 24 (58%) infants had abnormal MRI. In group 1 (chronic hypoxia/antenatal injury), total brain injury was most predominant (4/6 infants). Group 2 (gradually evolving/subacute hypoxia) was associated with peripheral brain injury (5/5 infants). Group 3 (acute hypoxia) was associated with basal-ganglia thalamic injury pattern (3/3 infants). Later neurodevelopmental outcomes were available for 35 cases. Infants suspected to have a pre-labor injury on CTG (group 1) had a higher proportion of adverse neurodevelopmental outcomes (4/10, 40%) compared to groups 2 and 3 (4/25, 16%). CONCLUSION: Using this novel physiology-based CTG classification, we demonstrate an association between types of hypoxia observed on the CTG and MRI patterns of hypoxic brain injury. Infants with CTG trace suggestive of chronic hypoxia or other antenatal injuries were overrepresented in this cohort and were also more likely to have a poor neurodevelopmental outcome.

Sodium-calcium exchanger-3 regulates pain "wind-up": From human psychophysics to spinal mechanisms.

Repeated application of noxious stimuli leads to a progressively increased pain perception; this temporal summation is enhanced in and predictive of clinical pain disorders. Its electrophysiological correlate is "wind-up," in which dorsal horn spinal neurons increase their response to repeated nociceptor stimulation. To understand the genetic basis of temporal summation, we undertook a GWAS of wind-up in healthy human volunteers and found significant association with SLC8A3 encoding sodium-calcium exchanger type 3 (NCX3). NCX3 was expressed in mouse dorsal horn neurons, and mice lacking NCX3 showed normal, acute pain but hypersensitivity to the second phase of the formalin test and chronic constriction injury. Dorsal horn neurons lacking NCX3 showed increased intracellular calcium following repetitive stimulation, slowed calcium clearance, and increased wind-up. Moreover, virally mediated enhanced spinal expression of NCX3 reduced central sensitization. Our study highlights Ca2+ efflux as a pathway underlying temporal summation and persistent pain, which may be amenable to therapeutic targeting.

FTY720 ameliorates MOG-induced experimental autoimmune encephalomyelitis by suppressing both cellular and humoral immune responses.

FTY720, an oral sphingosine 1-phosphate (S1P) receptor modulator, has shown efficacy in phase II trials in patients with relapsing-remitting multiple sclerosis (MS). Although this molecule is thought to immunosuppress by inhibiting lymphocyte egress from the lymph nodes, the full spectrum of FTY720's actions has not yet been uncovered. In this study, we investigated the effects of FTY720 treatment on disease severity and histopathology of MOG-induced experimental autoimmune encephalomyelitis (EAE) in the dark agouti (DA) rat, a model that closely mimics several features of MS. The effects of FTY720 on T-cell subsets, anti-MOG antibody production, and mRNA expression of a number of cytokines and other genes were also examined. Commencement of treatment before disease onset prevented the appearance of clinical disease. Therapeutic treatment after established disease reduced clinical scores and substantially attenuated inflammation, demyelination, and axon loss. EAE suppression was associated with a reduction in all measured T-cell subsets in blood and spleen and a significant decrease in serum IgG(2a) levels. However, in the lymph nodes, all T-cell subsets except for naïve T cells and recent thymic emigrants remained unaffected. In addition, FTY720 treatment led to a significant inhibition in interferon-gamma, inducible nitric oxide synthase, and glial cell line-derived neurotrophic factor mRNA expression in the MOG-EAE spinal cord. In conclusion, our findings indicate that FTY720-mediated S1P receptor modulation ameliorates chronic relapsing MOG-EAE by suppressing both cellular and humoral immune responses.

Translational issues in precision medicine in neuropathic pain.

Neuropathic pain remains poorly treated, with most new drugs falling through the translational gap. The traditional model of bench-to-bedside research has relied on identifying new mechanisms/targets in animal models and then developing clinical applications. Several have advocated bridging the translational gap by beginning with clinical observations and back-translating to animal models for further investigation of mechanisms. There is good evidence that phenotyping of patients through quantitative sensory testing can lead to improved treatment selection and hence improved patient outcomes. This practice has been widely adopted in clinical investigations, but its application in preclinical research is not mainstream. In this review, we retrospectively examine our historical rodent data sets with the aim of reconsidering drug effects on sensory neuronal endpoints, their alignment with clinical observations, and how these might guide future clinical studies.

La douleur neuropathique reste mal traitée, alors que la plupart des nouveaux médicaments ne réussissent pas à franchir le fossé translationnel. Le modèle traditionnel de la recherche, du laboratoire au chevet du patient, repose sur l'identification de nouveaux mécanismes ou cibles dans des modèles animaux, suivie du développement d'applications cliniques. Certains préconisent de combler le fossé de la recherche translationnelle en commençant par des observations cliniques et en les transposant ensuite sur des modèles animaux afin d'approfondir l'étude des mécanismes. Il est bien établi que le phénotypage des patients par des tests sensoriels quantitatifs peut conduire à une meilleure sélection des traitements et, par conséquent, à de meilleurs résultats pour les patients. Ces pratiques ont été largement adoptées dans les enquêtes cliniques, mais leur application dans la recherche préclinique n'est pas généralisée. Dans cette revue, nous examinons rétrospectivement nos ensembles de données historiques sur les rongeurs dans le but de reconsidérer les effets des médicaments sur les paramètres neuronaux sensoriels, leur alignement avec les observations cliniques et la manière dont celles-ci pourraient orienter les études cliniques futures.

GPR160 de-orphanization reveals critical roles in neuropathic pain in rodents.

Treating neuropathic pain is challenging and novel non-opioid-based medicines are needed. Using unbiased receptomics, transcriptomic analyses, immunofluorescence, and in situ hybridization, we found that the expression of the orphan GPCR Gpr160 and GPR160 increased in the rodent dorsal horn of the spinal cord following traumatic nerve injury. Genetic and immunopharmacological approaches demonstrated that GPR160 inhibition in the spinal cord prevented and reversed neuropathic pain in male and female rodents without altering normal pain response. GPR160 inhibition in the spinal cord attenuated sensory processing in the thalamus, a key relay in the sensory discriminative pathways of pain. We also identified cocaine- and amphetamine-regulated transcript peptide (CARTp) as a GPR160 ligand. Inhibiting endogenous CARTp signaling in spinal cord attenuated neuropathic pain, whereas exogenous intrathecal CARTp evoked painful hypersensitivity through GPR160-dependent ERK and cAMP response element-binding protein (CREB). Our findings de-orphanize GPR160, identify it as a determinant of neuropathic pain and potential therapeutic target, and provide insights into its signaling pathways. CARTp is involved in many diseases including depression and reward and addiction; de-orphanization of GPR160 is a major step forward understanding the role of CARTp signaling in health and disease.

Neuropathy following spinal nerve injury shares features with the irritable nociceptor phenotype: A back-translational study of oxcarbazepine.

BACKGROUND: The term 'irritable nociceptor' was coined to describe neuropathic patients characterized by evoked hypersensitivity and preservation of primary afferent fibres. Oxcarbazepine is largely ineffectual in an overall patient population, but has clear efficacy in a subgroup with the irritable nociceptor profile. We examine whether neuropathy in rats induced by spinal nerve injury shares overlapping pharmacological sensitivity with the irritable nociceptor phenotype using drugs that target sodium channels. METHODS: In vivo electrophysiology was performed in anaesthetized spinal nerve ligated (SNL) and sham-operated rats to record from wide dynamic range (WDR) neurones in the ventral posterolateral thalamus (VPL) and dorsal horn. RESULTS: In neuropathic rats, spontaneous activity in the VPL was substantially attenuated by spinal lidocaine, an effect that was absent in sham rats. The former measure was in part dependent on ongoing peripheral activity as intraplantar lidocaine also reduced aberrant spontaneous thalamic firing. Systemic oxcarbazepine had no effect on wind-up of dorsal horn neurones in sham and SNL rats. However, in SNL rats, oxcarbazepine markedly inhibited punctate mechanical-, dynamic brush- and cold-evoked neuronal responses in the VPL and dorsal horn, with minimal effects on heat-evoked responses. In addition, oxcarbazepine inhibited spontaneous activity in the VPL. Intraplantar injection of the active metabolite licarbazepine replicated the effects of systemic oxcarbazepine, supporting a peripheral locus of action. CONCLUSIONS: We provide evidence that ongoing activity in primary afferent fibres drives spontaneous thalamic firing after spinal nerve injury and that oxcarbazepine through a peripheral mechanism exhibits modality-selective inhibitory effects on sensory neuronal processing. SIGNIFICANCE: The inhibitory effects of lidocaine and oxcarbazepine in this rat model of neuropathy resemble the clinical observations in the irritable nociceptor patient subgroup and support a mechanism-based rationale for bench-to-bedside translation when screening novel drugs.