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1.
Nat Commun ; 14(1): 366, 2023 Jan 23.
Artigo em Inglês | MEDLINE | ID: mdl-36690629

RESUMO

Sensory neurons of the dorsal root ganglion (DRG) are critical for maintaining tissue homeostasis by sensing and initiating responses to stimuli. While most preclinical studies of DRGs are conducted in rodents, much less is known about the mechanisms of sensory perception in primates. We generated a transcriptome atlas of mouse, guinea pig, cynomolgus monkey, and human DRGs by implementing a common laboratory workflow and multiple data-integration approaches to generate high-resolution cross-species mappings of sensory neuron subtypes. Using our atlas, we identified conserved core modules highlighting subtype-specific biological processes related to inflammatory response. We also identified divergent expression of key genes involved in DRG function, suggesting species-specific adaptations specifically in nociceptors that likely point to divergent function of nociceptors. Among these, we validated that TAFA4, a member of the druggable genome, was expressed in distinct populations of DRG neurons across species, highlighting species-specific programs that are critical for therapeutic development.


Assuntos
Gânglios Espinais , Transcriptoma , Camundongos , Humanos , Animais , Cobaias , Gânglios Espinais/metabolismo , Macaca fascicularis , Nociceptores/metabolismo , Células Receptoras Sensoriais/metabolismo , Sensação , Citocinas/metabolismo
2.
Molecules ; 28(2)2023 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-36677929

RESUMO

Arthroplasty is an orthopedic surgical procedure that replaces a dysfunctional joint by an orthopedic prosthesis, thereby restoring joint function. Upon the use of the joint prosthesis, a wearing process begins, which releases components such as titanium dioxide (TiO2) that trigger an immune response in the periprosthetic tissue, leading to arthritis, arthroplasty failure, and the need for revision. Flavonoids belong to a class of natural polyphenolic compounds that possess antioxidant and anti-inflammatory activities. Hesperidin methyl chalcone's (HMC) analgesic, anti-inflammatory, and antioxidant effects have been investigated in some models, but its activity against the arthritis caused by prosthesis-wearing molecules, such as TiO2, has not been investigated. Mice were treated with HMC (100 mg/kg, intraperitoneally (i.p.)) 24 h after intra-articular injection of 3 mg/joint of TiO2, which was used to induce chronic arthritis. HMC inhibited mechanical hyperalgesia, thermal hyperalgesia, joint edema, leukocyte recruitment, and oxidative stress in the knee joint (alterations in gp91phox, GSH, superoxide anion, and lipid peroxidation) and in recruited leukocytes (total reactive oxygen species and GSH); reduced patellar proteoglycan degradation; and decreased pro-inflammatory cytokine production. HMC also reduced the activation of nociceptor-sensory TRPV1+ and TRPA1+ neurons. These effects occurred without renal, hepatic, or gastric damage. Thus, HMC reduces arthritis triggered by TiO2, a component released upon wearing of prosthesis.


Assuntos
Artrite , Chalconas , Hesperidina , Camundongos , Animais , Nociceptores/metabolismo , Chalconas/uso terapêutico , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Artrite/tratamento farmacológico , Estresse Oxidativo , Antioxidantes/farmacologia , Anti-Inflamatórios/farmacologia , Hiperalgesia/tratamento farmacológico , Citocinas/metabolismo
3.
Mol Pain ; 19: 17448069221148351, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36526437

RESUMO

Sensory neuron hyperexcitability is a critical driver of pathological pain and can result from axon damage, inflammation, or neuronal stress. G-protein coupled receptor signaling can induce pain amplification by modulating the activation of Trp-family ionotropic receptors and voltage-gated ion channels. Here, we sought to use calcium imaging to identify novel inhibitors of the intracellular pathways that mediate sensory neuron sensitization and lead to hyperexcitability. We identified a novel stimulus cocktail, consisting of the SSTR2 agonist L-054,264 and the S1PR3 agonist CYM5541, that elicits calcium responses in mouse primary sensory neurons in vitro as well as pain and thermal hypersensitivity in mice in vivo. We screened a library of 906 bioactive compounds and identified 24 hits that reduced calcium flux elicited by L-054,264/CYM5541. Among these hits, silymarin, a natural product derived from milk thistle, strongly reduced activation by the stimulation cocktail, as well as by a distinct inflammatory cocktail containing bradykinin and prostaglandin E2. Silymarin had no effect on sensory neuron excitability at baseline, but reduced calcium flux via Orai channels and downstream mediators of phospholipase C signaling. In vivo, silymarin pretreatment blocked development of adjuvant-mediated thermal hypersensitivity, indicating potential use as an anti-inflammatory analgesic.


Assuntos
Nociceptores , Silimarina , Camundongos , Animais , Nociceptores/metabolismo , Cálcio/metabolismo , Silimarina/metabolismo , Silimarina/farmacologia , Dor/metabolismo , Células Receptoras Sensoriais/metabolismo , Anti-Inflamatórios não Esteroides/farmacologia , Gânglios Espinais/metabolismo
4.
Sci Rep ; 12(1): 20995, 2022 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-36470947

RESUMO

Multiple Sclerosis (MS) is an autoimmune disease with notable sex differences. Women are not only more likely to develop MS but are also more likely than men to experience neuropathic pain in the disease. It has been postulated that neuropathic pain in MS can originate in the peripheral nervous system at the level of the dorsal root ganglia (DRG), which houses primary pain sensing neurons (nociceptors). These nociceptors become hyperexcitable in response to inflammation, leading to peripheral sensitization and eventually central sensitization, which maintains pain long-term. The mouse model experimental autoimmune encephalomyelitis (EAE) is a good model for human MS as it replicates classic MS symptoms including pain. Using EAE mice as well as naïve primary mouse DRG neurons cultured in vitro, we sought to characterize sex differences, specifically in peripheral sensory neurons. We found sex differences in the inflammatory profile of the EAE DRG, and in the TNFα downstream signaling pathways activated intracellularly in cultured nociceptors. We also found increased cell death with TNFα treatment. Given that TNFα signaling has been shown to initiate intrinsic apoptosis through mitochondrial disruption, this led us to investigate sex differences in the mitochondria's response to TNFα. Our results demonstrate that male sensory neurons are more sensitive to mitochondrial stress, making them prone to neuronal injury. In contrast, female sensory neurons appear to be more resistant to mitochondrial stress and exhibit an inflammatory and regenerative phenotype that may underlie greater nociceptor hyperexcitability and pain. Understanding these sex differences at the level of the primary sensory neuron is an important first step in our eventual goal of developing sex-specific treatments to halt pain development in the periphery before central sensitization is established.


Assuntos
Encefalomielite Autoimune Experimental , Esclerose Múltipla , Neuralgia , Feminino , Camundongos , Masculino , Humanos , Animais , Gânglios Espinais/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Caracteres Sexuais , Nociceptores/metabolismo , Neuralgia/etiologia , Neuralgia/metabolismo , Esclerose Múltipla/metabolismo
5.
Proc Natl Acad Sci U S A ; 119(45): e2210053119, 2022 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-36322763

RESUMO

Choreographic dendritic arborization takes place within a defined time frame, but the timing mechanism is currently not known. Here, we report that the precisely timed lin-4-lin-14 regulatory circuit triggers an initial dendritic growth activity, whereas the precisely timed lin-28-let-7-lin-41 regulatory circuit signals a subsequent developmental decline in dendritic growth ability, hence restricting dendritic arborization within a set time frame. Loss-of-function mutations in the lin-4 microRNA gene cause limited dendritic outgrowth, whereas loss-of-function mutations in its direct target, the lin-14 transcription factor gene, cause precocious and excessive outgrowth. In contrast, loss-of-function mutations in the let-7 microRNA gene prevent a developmental decline in dendritic growth ability, whereas loss-of-function mutations in its direct target, the lin-41 tripartite motif protein gene, cause further decline. lin-4 and let-7 regulatory circuits are expressed in the right place at the right time to set start and end times for dendritic arborization. Replacing the lin-4 upstream cis-regulatory sequence at the lin-4 locus with a late-onset let-7 upstream cis-regulatory sequence delays dendrite arborization, whereas replacing the let-7 upstream cis-regulatory sequence at the let-7 locus with an early-onset lin-4 upstream cis-regulatory sequence causes a precocious decline in dendritic growth ability. Our results indicate that the lin-4-lin-14 and the lin-28-let-7-lin-41 regulatory circuits control the timing of dendrite arborization through antagonistic regulation of the DMA-1 receptor level on dendrites. The LIN-14 transcription factor likely directly represses dma-1 gene expression through a transcriptional means, whereas the LIN-41 tripartite motif protein likely indirectly promotes dma-1 gene expression through a posttranscriptional means.


Assuntos
Proteínas de Caenorhabditis elegans , MicroRNAs , Animais , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Nociceptores/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteínas com Motivo Tripartido/genética , Plasticidade Neuronal , Proteínas Repressoras/metabolismo , Proteínas de Membrana/metabolismo
6.
Elife ; 112022 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-36200757

RESUMO

Mechanical nociception is an evolutionarily conserved sensory process required for the survival of living organisms. Previous studies have revealed much about the neural circuits and sensory molecules in mechanical nociception, but the cellular mechanisms adopted by nociceptors in force detection remain elusive. To address this issue, we study the mechanosensation of a fly larval nociceptor (class IV da neurons, c4da) using a customized mechanical device. We find that c4da are sensitive to mN-scale forces and make uniform responses to the forces applied at different dendritic regions. Moreover, c4da showed a greater sensitivity to localized forces, consistent with them being able to detect the poking of sharp objects, such as wasp ovipositor. Further analysis reveals that high morphological complexity, mechanosensitivity to lateral tension and possibly also active signal propagation in dendrites contribute to the sensory features of c4da. In particular, we discover that Piezo and Ppk1/Ppk26, two key mechanosensory molecules, make differential but additive contributions to the mechanosensitivity of c4da. In all, our results provide updates into understanding how c4da process mechanical signals at the cellular level and reveal the contributions of key molecules.


Being able to sense harm is essential for survival. Animals have to be able to tell the difference between a gentle touch and a dangerous pressure. They do this using nerve cells called mechanical nociceptors which switch on when the body feels a potentially painful pressure, such as a sharp object poking the skin. Once activated, the nerves send outputs to other parts of the central nervous system which coordinate the motions needed to escape the source of the pain. One popular model to understand harm-sensing is the larvae of fruit flies which automatically roll back and forth when they sense the pointy sting of a wasp. This process is initiated by sensory nerve cells called class IV dendritic arborization neurons (or c4da for short) which sit under the fly's skin. However, it is still not fully understood how these mechanical nociceptors detect the poking forces of the wasp's tail. To investigate, Liu, Wu et al. built a device that could poke sections of fly larvae under a microscope so they could see how different types of pressure affected the activity and shape of c4da cells. This revealed that c4da nerves were most sensitive to sharp objects that illicit a more localized force, which may explain why these cells are so good at responding to wasp attacks. Further analysis showed that this sensitivity was due to the high number of branches, or dendrites, protruding from the body of c4da nerves. Liu, Wu et al. discovered that the dendrites were coated in a touch-sensitive protein that can sense and amplify both squashing and pulling, resulting in a signal that activates c4da nerves to send outputs to other parts of the central nervous system. This mechanism increases the likelihood that a c4da cell will detect a mechanical pressure even if it is far away from the body of the nerve. These findings shed light on how sensory cells like c4da are optimized to carry out specific roles. This could be important for understanding other nerve systems which sense mechanical pressure, such as those involved in touch or auditory processes. However, further work is needed to see whether the molecules and mechanism identified by Liu, Wu et al. are also present in humans.


Assuntos
Proteínas de Drosophila , Drosophila , Animais , Drosophila/metabolismo , Nociceptores/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Larva/fisiologia , Canais Iônicos , Canais Epiteliais de Sódio
7.
Cell ; 185(22): 4190-4205.e25, 2022 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-36243004

RESUMO

Neuroepithelial crosstalk is critical for gut physiology. However, the mechanisms by which sensory neurons communicate with epithelial cells to mediate gut barrier protection at homeostasis and during inflammation are not well understood. Here, we find that Nav1.8+CGRP+ nociceptor neurons are juxtaposed with and signal to intestinal goblet cells to drive mucus secretion and gut protection. Nociceptor ablation led to decreased mucus thickness and dysbiosis, while chemogenetic nociceptor activation or capsaicin treatment induced mucus growth. Mouse and human goblet cells expressed Ramp1, receptor for the neuropeptide CGRP. Nociceptors signal via the CGRP-Ramp1 pathway to induce rapid goblet cell emptying and mucus secretion. Notably, commensal microbes activated nociceptors to control homeostatic CGRP release. In the absence of nociceptors or epithelial Ramp1, mice showed increased epithelial stress and susceptibility to colitis. Conversely, CGRP administration protected nociceptor-ablated mice against colitis. Our findings demonstrate a neuron-goblet cell axis that orchestrates gut mucosal barrier protection.


Assuntos
Colite , Células Caliciformes , Camundongos , Humanos , Animais , Células Caliciformes/metabolismo , Nociceptores/metabolismo , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Colite/metabolismo , Muco/metabolismo , Proteína 1 Modificadora da Atividade de Receptores/metabolismo
8.
Eur J Pain ; 26(10): 2238-2256, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36097791

RESUMO

BACKGROUND: Kv4 channels are key components controlling neuronal excitability at membrane potentials below action potential thresholds. It remains elusive whether Kv4.1 participates in pain regulation. METHODS: We raised a Kv4.1 antibody to map Kv4.1+ neurons in the superficial dorsal horn of the spinal cord and dorsal root ganglion (DRG) of rats. Behavioural, biochemical and immunohistochemical methods were used to examine whether the activity of Kv4.1+ neurons or Kv4.1 expression level is altered after peripheral nerve injury. RESULTS: In lamina I of the spinal cord, Kv4.1 immunoreactivity (IR) was detected in neurokinin-1 receptor positive (NK1R)+ projection neurons (the secondary nociceptive neurons) and NK1R+ excitatory interneurons. Kv4.1, KChIP2 and DPP10 were co-expressed in these neurons. Peripheral nerve injury evoked by lumbar spinal nerve ligation (SNL) immediately induced phosphorylated extracellular regulated protein kinase (pERK, an indicator of enhanced neuronal activity) in lamina I Kv4.1+ neurons and lamina II Kv4.2/Kv4.3+ neurons of the spinal cord. Furthermore, Kv4.1 appeared in 59.9% of DRG neurons with variable sizes. Kv4.1 mRNA and protein levels in DRG neurons were gradually decreased after SNL. Following intrathecal injection of Kv4.1 antisense oligodeoxynucleotide (ASO) into naive rats, Kv4.1 protein level was reduced in the DRG, and mechanical but not thermal hypersensitivity was induced. CONCLUSIONS: Kv4.1 appears in the secondary nociceptive neurons, and peripheral nerve injury increases the activity of these neurons. Kv4.1 expression in DRG neurons (including half of the nociceptors) is gradually reduced after peripheral nerve injury, and knockdown of Kv4.1 in DRG neurons induces pain. Thus, Kv4.1 participates in pain regulation. SIGNIFICANCE: Based on the expression of Kv4.1 and Kv4.3 in the nociceptors, Kv4.1 in the secondary nociceptive neurons, Kv4.1 in spinal lamina I excitatory interneurons that regulate the activity of the secondary nociceptive neurons, as well as Kv4.2 and Kv4.3 in spinal lamina II excitatory interneurons that also regulate the activity of the secondary nociceptive neurons, developing Kv4 activators or genetic manipulation to increase Kv4 channel activity in these pain-related Kv4+ neurons will be useful in future pain therapeutics.


Assuntos
Nociceptores , Traumatismos dos Nervos Periféricos , Canais de Potássio Shal , Animais , Nociceptores/metabolismo , Oligodesoxirribonucleotídeos/metabolismo , Dor/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Proteínas Quinases/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores da Neurocinina-1 , Canais de Potássio Shal/metabolismo , Corno Dorsal da Medula Espinal/metabolismo
9.
Neuron ; 110(22): 3727-3742.e8, 2022 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-36087585

RESUMO

Mechanical nociception is essential for animal survival. However, the forces involved in nociceptor activation and the underlying mechanotransduction mechanisms remain elusive. Here, we address these problems by investigating nocifensive behavior in Drosophila larvae. We show that strong poking stimulates nociceptors with a mixture of forces including shear stress and stretch. Unexpectedly, nociceptors are selectively activated by shear stress, but not stretch. Both the shear stress responses of nociceptors and nocifensive behavior require transient receptor potential A1 (TrpA1), which is specifically expressed in nociceptors. We further demonstrate that expression of mammalian or Drosophila TrpA1 in heterologous cells confers responses to shear stress but not stretch. Finally, shear stress activates TrpA1 in a membrane-delimited manner, through modulation of membrane fluidity. Together, our study reveals TrpA1 as an evolutionarily conserved mechanosensitive channel specifically activated by shear stress and suggests a critical role of shear stress in activating nociceptors to drive mechanical nociception.


Assuntos
Nociceptores , Canais de Potencial de Receptor Transitório , Animais , Nociceptores/metabolismo , Drosophila/metabolismo , Nociceptividade/fisiologia , Mecanotransdução Celular , Canal de Cátion TRPA1/metabolismo , Canais de Potencial de Receptor Transitório/metabolismo , Mamíferos/metabolismo
10.
Cells ; 11(15)2022 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-35954234

RESUMO

Hypersensitivity to mechanical stimuli is a cardinal symptom of neuropathic and inflammatory pain. A reduction in spinal inhibition is generally considered a causal factor in the development of mechanical hypersensitivity after injury. However, the extent to which presynaptic inhibition contributes to altered spinal inhibition is less well established. Here, we used conditional deletion of GABAA in NaV1.8-positive sensory neurons (Scn10aCre;Gabrb3fl/fl) to manipulate selectively presynaptic GABAergic inhibition. Behavioral testing showed that the development of inflammatory punctate allodynia was mitigated in mice lacking pre-synaptic GABAA. Dorsal horn cellular circuits were visualized in single slices using stimulus-tractable dual-labelling of c-fos mRNA for punctate and the cognate c-Fos protein for dynamic mechanical stimulation. This revealed a substantial reduction in the number of cells activated by punctate stimulation in mice lacking presynaptic GABAA and an approximate 50% overlap of the punctate with the dynamic circuit, the relative percentage of which did not change following inflammation. The reduction in dorsal horn cells activated by punctate stimuli was equally prevalent in parvalbumin- and calretinin-positive cells and across all laminae I-V, indicating a generalized reduction in spinal input. In peripheral DRG neurons, inflammation following complete Freund's adjuvant (CFA) led to an increase in axonal excitability responses to GABA, suggesting that presynaptic GABA effects in NaV1.8+ afferents switch from inhibition to excitation after CFA. In the days after inflammation, presynaptic GABAA in NaV1.8+ nociceptors constitutes an "open gate" pathway allowing mechanoreceptors responding to punctate mechanical stimulation access to nociceptive dorsal horn circuits.


Assuntos
Hiperalgesia , Nociceptores , Animais , Adjuvante de Freund , Hiperalgesia/metabolismo , Inflamação/metabolismo , Camundongos , Nociceptores/metabolismo , Ácido gama-Aminobutírico
11.
Cells ; 11(15)2022 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-35954288

RESUMO

Migraines constitute a common neurological and headache disorder affecting around 15% of the world's population. In addition to other mechanisms, neurogenic neuroinflammation has been proposed to play a part in migraine chronification, which includes peripheral and central sensitization. There is therefore considerable evidence suggesting that inflammation in the intracranial meninges could be a key element in addition to calcitonin gene-related peptide (CGRP), leading to sensitization of trigeminal meningeal nociceptors in migraines. There are several studies that have utilized this approach, with a strong focus on using inflammatory animal models. Data from these studies show that the inflammatory process involves sensitization of trigeminovascular afferent nerve terminals. Further, by applying a wide range of different pharmacological interventions, insight has been gained on the pathways involved. Importantly, we discuss how animal models should be used with care and that it is important to evaluate outcomes in the light of migraine pathology.


Assuntos
Peptídeo Relacionado com Gene de Calcitonina , Transtornos de Enxaqueca , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Inflamação , Meninges , Nociceptores/metabolismo
12.
Pharmacol Res ; 183: 106392, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35940396

RESUMO

This study aimed to investigate the anti-neuropathic pain activity and its underlying molecular mechanism of Ajugarin-I (Aju-I) in a rat model of diabetic neuropathic pain. The rats were given a single injection of 60 mg/kg of streptozotocin (STZ) intraperitoneally (i.p.) to induce diabetic neuropathic pain. After two weeks, rats were given Aju-I (1 and 5 mg/kg/day) i.p. for four consecutive weeks. The results demonstrated that in diabetic rats, treatment with Aju-I decreased STZ-induced hyperglycemia. It reduced the pain hypersensitivity (mechanical, thermal, and cold nociception) caused by STZ. It effectively restored STZ-associated pathological changes in the pancreas. In the sciatic nerve and spinal cord, it attenuated STZ-associated histopathological alterations and DNA damage. It suppressed oxidative stress by increasing the expression of nuclear factor-erythroid factor 2-related factor 2 (Nrf2), thioredoxin (Trx), and heme oxygenase (HO-1), but decreasing the immunoreactivity of Kelch-like ECH-associated protein 1 (Keap1). Additionally, TRPV1 (transient receptor potential vanilloid 1) and TRPM8 (transient receptor potential melastatin 8) expression levels were considerably reduced by Aju-I treatment. It enhanced antioxidant levels and suppressed inflammatory cytokines production. Taken together, this research suggests that Aju-I treatment reduces pain behaviors in the STZ model of diabetic neuropathy via modulating Nrf2/Keap-1/HO-1 signaling and TRPV1/TRPM8 nociceptors.


Assuntos
Diabetes Mellitus Experimental , Neuropatias Diabéticas , Neuralgia , Canais de Cátion TRPM , Animais , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/tratamento farmacológico , Neuropatias Diabéticas/induzido quimicamente , Neuropatias Diabéticas/tratamento farmacológico , Neuropatias Diabéticas/metabolismo , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Neuralgia/tratamento farmacológico , Neuralgia/metabolismo , Nociceptores/metabolismo , Ratos , Estreptozocina/efeitos adversos , Canais de Cátion TRPM/metabolismo , Canais de Cátion TRPV/metabolismo
13.
Proc Natl Acad Sci U S A ; 119(33): e2118501119, 2022 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-35943985

RESUMO

Pain and itch are distinct sensations arousing evasion and compulsive desire for scratching, respectively. It's unclear whether they could invoke different neural networks in the brain. Here, we use the type 1 herpes simplex virus H129 strain to trace the neural networks derived from two types of dorsal root ganglia (DRG) neurons: one kind of polymodal nociceptors containing galanin (Gal) and one type of pruriceptors expressing neurotensin (Nts). The DRG microinjection and immunosuppression were performed in transgenic mice to achieve a successful tracing from specific types of DRG neurons to the primary sensory cortex. About one-third of nuclei in the brain were labeled. More than half of them were differentially labeled in two networks. For the ascending pathways, the spinothalamic tract was absent in the network derived from Nts-expressing pruriceptors, and the two networks shared the spinobulbar projections but occupied different subnuclei. As to the motor systems, more neurons in the primary motor cortex and red nucleus of the somatic motor system participated in the Gal-containing nociceptor-derived network, while more neurons in the nucleus of the solitary tract (NST) and the dorsal motor nucleus of vagus nerve (DMX) of the emotional motor system was found in the Nts-expressing pruriceptor-derived network. Functional validation of differentially labeled nuclei by c-Fos test and chemogenetic inhibition suggested the red nucleus in facilitating the response to noxious heat and the NST/DMX in regulating the histamine-induced scratching. Thus, we reveal the organization of neural networks in a DRG neuron type-dependent manner for processing pain and itch.


Assuntos
Galanina , Gânglios Espinais , Rede Nervosa , Neurotensina , Nociceptores , Dor , Prurido , Animais , Galanina/metabolismo , Gânglios Espinais/ultraestrutura , Herpesvirus Humano 1 , Camundongos , Camundongos Transgênicos , Rede Nervosa/ultraestrutura , Neurotensina/metabolismo , Nociceptores/metabolismo , Dor/fisiopatologia , Prurido/fisiopatologia , Núcleo Solitário/ultraestrutura
14.
J Neurosci Res ; 100(10): 1951-1966, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-35839339

RESUMO

Spastic cerebral palsy (CP) is a movement disorder marked by hypertonia and hyperreflexia; the most prevalent comorbidity is pain. Since spinal nociceptive afferents contribute to both the sensation of painful stimuli as well as reflex circuits involved in movement, we investigated the relationship between prenatal hypoxia-ischemia (HI) injury which can cause CP, and possible changes in spinal nociceptive circuitry. To do this, we examined nociceptive afferents and mechanical and thermal sensitivity of New Zealand White rabbit kits after prenatal HI or a sham surgical procedure. As described previously, a range of motor deficits similar to spastic CP was observed in kits born naturally after HI (40 min at ~70%-80% gestation). We found that HI caused an expansion of peptidergic afferents (marked by expression of calcitonin gene-related peptide) in both the superficial and deep dorsal horn at postnatal day (P)5. Non-peptidergic nociceptive afferent arborization (labeled by isolectin B4) was unaltered in HI kits, but overlap of the two populations (peptidergic and non-peptidergic nociceptors) was increased by HI. Density of glial fibrillary acidic protein was unchanged within spinal cord white matter regions important in nociceptive transmission at P5. We found that mechanical and thermal nociception was enhanced in HI kits even in the absence of motor deficits. These findings suggest that prenatal HI injury impacts spinal sensory pathways in addition to the more well-established disruptions to descending motor circuits. In conclusion, changes to spinal nociceptive circuitry could disrupt spinal reflexes and contribute to pain experienced by individuals with CP.


Assuntos
Paralisia Cerebral , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Paralisia Cerebral/complicações , Feminino , Nociceptividade , Nociceptores/metabolismo , Dor , Gravidez , Coelhos , Medula Espinal/metabolismo
15.
Neuropharmacology ; 216: 109189, 2022 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-35820471

RESUMO

Maresin-2 (MaR2) is a specialized pro-resolution lipid mediator (SPM) that reduces neutrophil recruitment in zymosan peritonitis. Here, we investigated the analgesic effect of MaR2 and its mechanisms in different mouse models of pain. For that, we used the lipopolysaccharide (LPS)-induced mechanical hyperalgesia (electronic version of the von Frey filaments), thermal hyperalgesia (hot plate test) and weight distribution (static weight bearing), as well as the spontaneous pain models induced by capsaicin (TRPV1 agonist) or AITC (TRPA1 agonist). Immune cell recruitment was determined by immunofluorescence and flow cytometry while changes in the pro-inflammatory mediator landscape were determined using a proteome profiler kit and ELISA after LPS injection. MaR2 treatment was also performed in cultured DRG neurons stimulated with capsaicin or AITC in the presence or absence of LPS. The effect of MaR2 on TRVP1- and TRPA1-dependent CGRP release by cultured DRG neurons was determined by EIA. MaR2 inhibited LPS-induced inflammatory pain and changes in the cytokine landscape as per cytokine array assay. MaR2 also inhibited TRPV1 and TRPA1 activation as observed by a reduction in calcium influx in cultured DRG neurons, and the number of flinches and time spent licking the paw induced by capsaicin or AITC. In corroboration, MaR2 reduced capsaicin- and AITC-induced CGRP release by cultured DRG neurons and immune cell recruitment to the paw skin close the CGRP+ fibers. In conclusion, we show that MaR2 is an analgesic SPM that acts by targeting leukocyte recruitment, nociceptor TRPV1 and TRPA1 activation, and CGRP release in mice.


Assuntos
Capsaicina , Canais de Potencial de Receptor Transitório , Analgésicos/farmacologia , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Capsaicina/farmacologia , Citocinas/farmacologia , Ácidos Docosa-Hexaenoicos , Hiperalgesia , Lipopolissacarídeos/farmacologia , Camundongos , Monócitos/metabolismo , Neurônios , Neutrófilos , Nociceptores/metabolismo , Dor , Canal de Cátion TRPA1 , Canais de Cátion TRPV
16.
J Vis Exp ; (184)2022 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-35848838

RESUMO

Somatosensory neurons have evolved to detect noxious stimuli and activate defensive reflexes. By sharing means of communication, nociceptor neurons also tune host defenses by controlling the activity of the immune system. The communication between these systems is mostly adaptive, helping to protect homeostasis, it can also lead to, or promote, the onset of chronic diseases. Both systems co-evolved to allow for such local interaction, as found in primary and secondary lymphoid tissues and mucosa. Recent studies have demonstrated that nociceptors directly detect and respond to foreign antigens, immune cell-derived cytokines, and microbes. Nociceptor activation not only results in pain hypersensitivity and itching, but lowers the nociceptor firing threshold, leading to the local release of neuropeptides. The peptides that are produced by, and released from, the peripheral terminals of nociceptors can block the chemotaxis and polarization of lymphocytes, controlling the localization, duration, and type of inflammation. Recent evidence shows that sensory neurons interact with innate immune cells via cell-cell contact, for example, engaging group 2D (NKG2D) receptors on natural killer (NK) cells. Given that NK cells express the cognate receptors for various nociceptor-produced mediators, it is conceivable that nociceptors use neuropeptides to control the activity of NK cells. Here, we devise a co-culture method to study nociceptor neuron-NK cell interactions in a dish. Using this approach, we found that lumbar nociceptor neurons decrease NK cell cytokine expression. Overall, such a reductionist method could be useful to study how tumor-innervating neurons control the anticancer function of NK cells and how NK cells control the elimination of injured neurons.


Assuntos
Neuropeptídeos , Nociceptores , Citocinas/metabolismo , Humanos , Células Matadoras Naturais , Neuropeptídeos/metabolismo , Nociceptores/metabolismo , Dor , Células Receptoras Sensoriais/metabolismo
17.
Exp Neurol ; 357: 114190, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-35907583

RESUMO

TREK2 is a member of the 2-pore domain family of K+ channels (K2P) preferentially expressed by unmyelinated, slow-conducting and non-peptidergic isolectin B4-binding (IB4+) primary sensory neurons of the dorsal root ganglia (DRG). IB4+ neurons depend on the glial-derived neurotrophic factor (GDNF) family of ligands (GFL's) to maintain their phenotype. In our previous work, we demonstrated that 7 days after spinal nerve axotomy (SNA) of the L5 DRG, TREK2 moves away from the cell membrane resulting in a more depolarised resting membrane potential (Em). Given that axotomy deprives DRG neurons from peripherally-derived GFL's, we hypothesized that they might control the expression of TREK2. Using a combination of immunohistochemistry, immunocytochemistry, western blotting, in vivo pharmacological manipulation and behavioral tests we examined the ability of the GFL's (GDNF, neurturin and artemin) and their selective receptors (GFRα1, GFRα2 and GFRα3) to regulate the expression and function of TREK2 in the DRG. We found that TREK2 correlated strongly with the three receptors normally and ipsilaterally for all GFR's after SNA. GDNF, but not NGF, neurturin or artemin up-regulated the expression of TREK2 in cultured DRG neurons. In vivo continuous, subcutaneous administration of GDNF restored the subcellular distribution of TREK2 ipsilaterally and reversed mechanical and cold allodynia 7 days after SNA. This is the first demonstration that GDNF controls the expression of a K2P channel in nociceptors. As TREK2 controls the Em of C-nociceptors affecting their excitability, our finding has therapeutic potential in the treatment of chronic pain.


Assuntos
Fator Neurotrófico Derivado de Linhagem de Célula Glial , Neuralgia , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Animais , Axotomia , Gânglios Espinais/metabolismo , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Neuralgia/metabolismo , Neurturina , Nociceptores/metabolismo , Ratos
18.
J Physiol ; 600(16): 3819-3836, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35775903

RESUMO

Visceral pain is a leading cause of morbidity in gastrointestinal diseases, which is exacerbated by the gut-related side-effects of many analgesics. New treatments are needed and further understanding of the mediators and mechanisms underpinning visceral nociception in disease states is required to facilitate this. The pro-inflammatory cytokine TNFα is linked to pain in both patients with inflammatory bowel disease and irritable bowel syndrome, and has been shown to sensitize colonic sensory neurons. Somatic, TNFα-triggered thermal and mechanical hypersensitivity is mediated by TRPV1 signalling and p38 MAPK activity respectively, downstream of TNFR1 receptor activation. We therefore hypothesized that TNFR1-evoked p38 MAPK activity may also be responsible for TNFα sensitization of colonic afferent responses to the TRPV1 agonist capsaicin, and noxious distension of the bowel. Using Ca2+ imaging of dorsal root ganglion sensory neurons, we observed TNFα-mediated increases in intracellular [Ca2+ ] and sensitization of capsaicin responses. The sensitizing effects of TNFα were dependent on TNFR1 expression and attenuated by p38 MAPK inhibition. Consistent with these findings, ex vivo colonic afferent fibre recordings demonstrated an enhanced response to noxious ramp distention of the bowel and bath application of capsaicin following TNFα pre-treatment. Responses were reversed by p38 MAPK inhibition and absent in tissue from TNFR1 knockout mice. Our findings demonstrate a contribution of TNFR1, p38 MAPK and TRPV1 to TNFα-induced sensitization of colonic afferents, highlighting the potential utility of these drug targets for the treatment of visceral pain in gastrointestinal disease. KEY POINTS: The pro-inflammatory cytokine TNFα is elevated in gastrointestinal disease and sensitizes colonic afferents via modulation of TRPA1 and NaV 1.8 activity. We further develop this understanding by demonstrating a role for p38 MAPK and TRPV1 in TNFα-mediated colonic afferent sensitization. Specifically, we show that: TNFα sensitizes sensory neurons and colonic afferents to the TRPV1 agonist capsaicin. TNFα-mediated sensitization of sensory neurons and colonic nociceptors is dependent on TNFR1 expression. TNFα sensitization of sensory neurons and colonic afferents to capsaicin and noxious ramp distension is abolished by inhibition of p38 MAPK. Collectively these data support the utility of targeting TNFα, TNFR1 and their downstream signalling via p38 MAPK for the treatment of visceral pain in gastrointestinal disease.


Assuntos
Nociceptores , Dor Visceral , Animais , Capsaicina/farmacologia , Gânglios Espinais/metabolismo , Camundongos , Nociceptores/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/farmacologia , Canais de Cátion TRPV/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Dor Visceral/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
19.
J Gen Physiol ; 154(8)2022 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-35819364

RESUMO

PIEZO2 channels mediate rapidly adapting mechanically activated currents in peripheral sensory neurons of the dorsal root ganglia (DRG), and they are indispensable for light touch and proprioception. Relatively little is known about what other proteins regulate PIEZO2 activity in a cellular context. TMEM120A (TACAN) was proposed to act as a high threshold mechanically activated ion channel in nociceptive DRG neurons. Here, we find that Tmem120a coexpression decreased the amplitudes of mechanically activated PIEZO2 currents and increased their threshold of activation. TMEM120A did not inhibit mechanically activated PIEZO1 and TREK1 channels and TMEM120A alone did not result in the appearance of mechanically activated currents above background. Tmem120a and Piezo2 expression in mouse DRG neurons overlapped, and siRNA-mediated knockdown of Tmem120a increased the amplitudes of rapidly adapting mechanically activated currents and decreased their thresholds to mechanical activation. Our data identify TMEM120A as a negative modulator of PIEZO2 channel activity, and do not support TMEM120A being a mechanically activated ion channel.


Assuntos
Canais Iônicos/metabolismo , Mecanotransdução Celular , Animais , Gânglios Espinais/metabolismo , Mecanotransdução Celular/fisiologia , Camundongos , Nociceptores/metabolismo , Células Receptoras Sensoriais/fisiologia
20.
Pflugers Arch ; 474(9): 1003-1019, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-35867188

RESUMO

We studied the efficacy of a near-infrared laser (1475 nm) to activate rat dorsal root ganglion (DRG) neurons with short punctate radiant heat pulses (55 µm diameter) and investigated temporal and spatial summation properties for the transduction process for noxious heat at a subcellular level. Strength-duration curves (10-80 ms range) indicated a minimum power of 30.2mW for the induction of laser-induced calcium transients and a chronaxia of 13.9 ms. However, threshold energy increased with increasing stimulus duration suggesting substantial radial cooling of the laser spot. Increasing stimulus duration demonstrated suprathreshold intensity coding of calcium transients with less than linear gains (Stevens exponents 0.29/35mW, 0.38/60mW, 0.46/70mW). The competitive TRPV1 antagonist capsazepine blocked responses to short near-threshold stimuli and significantly reduced responses to longer duration suprathreshold heat. Heating 1/3 of the soma of a neuron was sufficient to induce calcium transients significantly above baseline (p < 0.05), but maximum amplitude was only achieved by centering the laser over the entire neuron. Heat-induced calcium increase was highest in heated cell parts but rapidly reached unstimulated areas reminiscent of spreading depolarization and opening of voltage-gated calcium channels. Full intracellular equilibrium took about 3 s, consistent with a diffusion process. In summary, we investigated transduction mechanisms for noxious laser heat pulses in native sensory neurons at milliseconds temporal and subcellular spatial resolution and characterized strength duration properties, intensity coding, and spatial summation within single neurons. Thermal excitation of parts of a nociceptor spread via both membrane depolarization and intracellular calcium diffusion.


Assuntos
Temperatura Alta , Nociceptores , Animais , Cálcio/metabolismo , Células Cultivadas , Gânglios Espinais/metabolismo , Lasers , Nociceptores/metabolismo , Ratos , Ratos Sprague-Dawley
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