RESUMEN
BACKGROUND AND PURPOSE: Selective nociceptor fibre block is achieved by introducing the cell membrane impermeant sodium channel blocker lidocaine N-ethyl bromide (QX-314) through transient receptor potential V1 (TRPV1) channels into nociceptors. We screened local anaesthetics for their capacity to activate TRP channels, and characterized the nerve block obtained by combination with QX-314. EXPERIMENTAL APPROACH: We investigated TRP channel activation in dorsal root ganglion (DRG) neurons by calcium imaging and patch-clamp recordings, and cellular QX-314 uptake by MS. To characterize nerve block, compound action potential (CAP) recordings from isolated nerves and behavioural responses were analysed. KEY RESULTS: Of the 12 compounds tested, bupivacaine was the most potent activator of ruthenium red-sensitive calcium entry in DRG neurons and activated heterologously expressed TRPA1 channels. QX-314 permeated through TRPA1 channels and accumulated intracellularly after activation of these channels. Upon sciatic injections, QX-314 markedly prolonged bupivacaine's nociceptive block and also extended (to a lesser degree) its motor block. Bupivacaine's blockade of C-, but not A-fibre, CAPs in sciatic nerves was extended by co-application of QX-314. Surprisingly, however, this action was the same in wild-type, TRPA1-knockout and TRPV1/TRPA1-double knockout mice, suggesting a TRP-channel independent entry pathway. Consistent with this, high doses of bupivacaine promoted a non-selective, cellular uptake of QX-314. CONCLUSIONS AND IMPLICATIONS: Bupivacaine, combined with QX-314, produced a long-lasting sensory nerve block. This did not require QX-314 permeation through TRPA1, although bupivacaine activated these channels. Regardless of entry pathway, the greatly extended duration of block produced by QX-314 and bupivacaine may be clinically useful.
Asunto(s)
Anestésicos Locales/farmacología , Bupivacaína/farmacología , Lidocaína/análogos & derivados , Bloqueo Nervioso , Bloqueadores de los Canales de Sodio/metabolismo , Anestésicos Locales/administración & dosificación , Animales , Conducta Animal/efectos de los fármacos , Bupivacaína/administración & dosificación , Calcio/metabolismo , Línea Celular , Traumatismos de los Pies , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Inyecciones , Lidocaína/metabolismo , Masculino , Ratones Noqueados , Técnicas de Placa-Clamp , Nervios Periféricos/efectos de los fármacos , Cultivo Primario de Células , Ratas , Ratas Sprague-Dawley , Nervio Ciático/efectos de los fármacos , Canal Catiónico TRPA1 , Canales de Potencial de Receptor Transitorio/genética , Canales de Potencial de Receptor Transitorio/metabolismoRESUMEN
OBJECTIVES: To evaluate whether augmenting neuronal protective mechanisms might slow or arrest experimental diabetic peripheral neuropathy (DPN). DPN is one of the most common neurodegenerative disorders and is rising in prevalence. How it targets sensory neurons is uncertain; the disorder is irreversible and untreatable. We explored the intrinsic protective properties of overexpressed human HSP27 on experimental DPN. HSP27 is a small pro-survival heat shock protein that also increases axonal regeneration. METHODS: Experimental diabetes was superimposed on mice overexpressing a human HSP27 transgene and its impact was evaluated on epidermal innervation, behavioral tests of sensation and electrophysiological indices of DPN. RESULTS: Mice that overexpress human HSP27 in their sensory and motor neurons and that were made diabetic for 6 months by streptozotocin treatment were protected from a range of neuropathic abnormalities, including loss of footpad thermal sensation, mechanical allodynia, loss of epidermal innervation, and slowing of sensory conduction velocity. The protection was selective for sensory neurons in comparison to motor neurons and at 6 months provided better protection in female than male mice. Markers of RAGE-NFκB activation were attenuated by the transgene. CONCLUSIONS: The findings support the idea that diabetic polyneuropathy involves a unique, sensory-centric neurodegenerative process which can be reduced by overexpressing a single gene, an important starting point for new disease-modifying therapeutic approaches.
Asunto(s)
Diabetes Mellitus Experimental/patología , Ganglios Espinales/patología , Regulación de la Expresión Génica/genética , Proteínas de Choque Térmico HSP27/metabolismo , Células Receptoras Sensoriales/metabolismo , Factores de Edad , Análisis de Varianza , Animales , Glucemia/efectos de los fármacos , Glucemia/genética , Caspasa 3/metabolismo , Diabetes Mellitus Experimental/inducido químicamente , Modelos Animales de Enfermedad , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Hemoglobina Glucada/metabolismo , Proteínas de Choque Térmico HSP27/genética , Humanos , Hiperalgesia/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fibras Nerviosas/metabolismo , Fibras Nerviosas/patología , Fibras Nerviosas/fisiología , Conducción Nerviosa/efectos de los fármacos , Conducción Nerviosa/genética , Umbral del Dolor/fisiología , Nervios Periféricos/patología , Nervios Periféricos/fisiopatología , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Piel/inervación , Piel/metabolismo , Estreptozocina/farmacología , Factores de Tiempo , Quinasa de Factor Nuclear kappa BRESUMEN
BACKGROUND AND PURPOSE: We have developed a strategy to target the permanently charged lidocaine derivative lidocaine N-ethyl bromide (QX-314) selectively into nociceptive sensory neurons through the large-pore transient receptor potential cation channel subfamily V (TRPV1) noxious heat detector channel. This involves co-administration of QX-314 and a TRPV1 agonist to produce a long-lasting local analgesia. For potential clinical use we propose using lidocaine as the TRPV1 agonist, because it activates TRPV1 at clinical doses. EXPERIMENTAL APPROACH: We conducted experiments in rats to determine optimal concentrations and ratios of lidocaine and QX-314 that produce the greatest degree and duration of pain-selective block when administered nearby the sciatic nerve: reduction in the response to noxious mechanical (pinch) and to radiant heat stimuli, with minimal disruption in motor function (grip strength). KEY RESULTS: A combination of 0.5% QX-314 and 2% lidocaine produced 1 h of non-selective sensory and motor block followed by >9 h of pain-selective block, where grip strength was unimpaired. QX-314 at this concentration had no effect by itself, while 2% lidocaine by itself produced 1 h of non-selective block. The combination of 0.5% QX-314 and 2% lidocaine was the best of the many tested, in terms of the duration and selectivity of local analgesia. CONCLUSIONS AND IMPLICATIONS: Targeting charged sodium channel blockers into specific sets of axons via activation of differentially expressed large-pore channels provides an opportunity to produce prolonged local analgesia, and represents an example of how exploiting ion channels as a drug delivery port can be used to increase the specificity and efficacy of therapeutics.
Asunto(s)
Analgésicos/farmacología , Lidocaína/análogos & derivados , Nociceptores/efectos de los fármacos , Células Receptoras Sensoriales/metabolismo , Bloqueadores de los Canales de Sodio/farmacología , Canales Catiónicos TRPV/antagonistas & inhibidores , Analgésicos/farmacocinética , Animales , Sistemas de Liberación de Medicamentos/métodos , Lidocaína/farmacocinética , Lidocaína/farmacología , Masculino , Nociceptores/metabolismo , Dolor/tratamiento farmacológico , Dolor/metabolismo , Ratas , Bloqueadores de los Canales de Sodio/farmacocinética , Canales Catiónicos TRPV/metabolismoRESUMEN
BACKGROUND: Oesophageal acid infusion induces enhanced pain hypersensitivity in non-acid exposed upper oesophagus (secondary hyperalgesia) in patients with non-cardiac chest pain, thus suggesting central sensitisation contributes to visceral pain hypersensitivity in functional gut disorders (FGD). Perceptual wind-up (increased pain perception to constant intensity sensory stimuli at frequencies>or=0.3 Hz) is used as a proxy for central sensitisation to investigate pain syndromes where pain hypersensitivity is important (for example, fibromyalgia). AIMS: Wind-up in central sensitisation induced human visceral pain hypersensitivity has not been explored. We hypothesised that if wind-up is a proxy for central sensitisation induced human visceral pain hypersensitivity, then oesophageal wind-up should be enhanced by secondary hyperalgesia. METHODS: In eight healthy volunteers (seven males; mean age 32 years), perception at pain threshold to a train of 20 electrical stimuli applied to the hand and upper oesophagus (UO) at either 0.1 Hz (control) or 2 Hz was determined before and one hour after a 30 minute lower oesophageal acid infusion. RESULTS: Wind-up occurred only with the 2 Hz train in the UO and hand (both p=0.01). Following acid infusion, pain threshold decreased (17 (4)%; p=0.01) in the UO, suggesting the presence of secondary hyperalgesia. Wind-up to the 2 Hz train increased in the UO (wind-up ratio 1.4 (0.1) to 1.6 (0.1); p=0.03) but not in the hand (wind-up ratio 1.3 (0.1) and 1.3 (0.1); p=0.3) CONCLUSION: Enhanced wind-up after secondary oesophageal hyperalgesia suggests that visceral pain hypersensitivity induced by central sensitisation results from increased central neuronal excitability. Wind-up may offer new opportunities to investigate the contribution of central neuronal changes to symptoms in FGD.
Asunto(s)
Esófago/fisiología , Aprendizaje/fisiología , Umbral del Dolor/fisiología , Adulto , Estimulación Eléctrica , Mano , Humanos , Ácido Clorhídrico/farmacología , Concentración de Iones de Hidrógeno , Masculino , Persona de Mediana Edad , Dimensión del Dolor , Distorsión de la PercepciónRESUMEN
Cyclooxygenase-2 (COX-2) after induction peripherally, and within the CNS, plays an important role in producing inflammatory pain. However, its role in neuropathic pain models is controversial. Recently a robust and persistent model of partial nerve injury pain, the spared nerve injury (SNI) model, has been developed. The aim of the present study was to examine the regulation of COX-2 in the rat SNI model and to evaluate the effectiveness of the selective COX-2 inhibitor rofecoxib in preventing neuropathic allodynia and hyperalgesia. RNase protection assays revealed only a very small and transient increase in COX-2 mRNA in the dorsal horn of the spinal cord in the SNI model with a maximum change at 24 h. Immunohistochemical analysis showed a small increase in COX-2 protein in the deep layers of the dorsal horn 10 h following SNI surgery. Rofecoxib (100 microM) did not affect spontaneous excitatory postsynaptic currents or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propanoic acid (AMPA) and N-methyl-d-aspartate (NMDA) responses in lamina II neurons from spinal cords of animals with SNI indicating no detectable action on transmitter release or postsynaptic activity. Furthermore, rofecoxib treatment (1 and 3.2 mg/kg for 5 and 3 days respectively starting on the day of surgery) failed to modify the development of allodynia and hyperalgesia in the SNI model. However, rofecoxib significantly reduced inflammatory hypersensitivity evoked by injection of complete Freund's adjuvant into one hindpaw, indicating that the doses used were pharmacologically active. The pain hypersensitivity produced by the SNI model is not COX-2-dependent.
Asunto(s)
Isoenzimas/metabolismo , Neuralgia/enzimología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Animales , Conducta Animal , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/análisis , Inhibidores de la Ciclooxigenasa/sangre , Inhibidores de la Ciclooxigenasa/farmacología , Soluciones para Diálisis/química , Modelos Animales de Enfermedad , Electrofisiología , Hiperalgesia/prevención & control , Hiperestesia/prevención & control , Inmunohistoquímica , Isoenzimas/genética , Lactonas/análisis , Lactonas/sangre , Lactonas/farmacología , Microdiálisis , Neuralgia/fisiopatología , Ensayos de Protección de Nucleasas , Concentración Osmolar , Umbral del Dolor/efectos de los fármacos , Prostaglandina-Endoperóxido Sintasas/genética , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Ribonucleasas , SulfonasRESUMEN
Despite identification of GABA(B) receptors with gb1a-gb2 composition and the alpha2delta calcium channel subunit as putative molecular targets for gabapentin (GBP), its cellular mechanism of action has remained elusive. Therefore, we have used an in vitro spinal cord slice preparation to study the effects of GBP on lamina II neurons. The frequency and amplitude of spontaneous EPSCs and IPSCs were unaffected by GBP, suggesting presynaptic neurotransmitter release is not regulated. Direct modulation of postsynaptic membrane excitability is also unlikely since the level of holding current required to maintain neurons at -70, 0 and +45 mV was unaffected by GBP. Effects on excitatory and inhibitory synaptic transmission were variable across the population. Primary afferent-evoked fast glutamatergic EPSCs were unaffected by GBP, while evoked NMDA receptor-mediated EPSCs and IPSCs were variably affected. In contrast, GBP enhanced responses to bath applied NMDA in 71% of neurons. Thus, in adult rat dorsal horn, synaptic and extrasynaptic NMDA receptors may be differentially regulated by GBP perhaps due to differences in subunit composition.
Asunto(s)
Acetatos/farmacología , Aminas , Ácidos Ciclohexanocarboxílicos , Células del Asta Posterior/efectos de los fármacos , Ácido gamma-Aminobutírico , Animales , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Gabapentina , Técnicas In Vitro , Masculino , N-Metilaspartato/farmacología , Células del Asta Posterior/fisiología , Ratas , Ratas Sprague-Dawley , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiologíaAsunto(s)
Analgesia/veterinaria , Analgésicos/uso terapéutico , Neuronas Aferentes/fisiología , Dolor/veterinaria , Analgesia/métodos , Analgésicos/efectos adversos , Animales , Neuronas Aferentes/clasificación , Dolor/fisiopatología , Manejo del Dolor , Dimensión del Dolor/veterinaria , Dolor Intratable/fisiopatología , Dolor Intratable/terapia , Dolor Intratable/veterinariaRESUMEN
Although visceral hypersensitivity is thought to be important in generating symptoms in functional gastrointestinal disorders, the neural mechanisms involved are poorly understood. We recently showed that central sensitization (hyperexcitability of spinal cord sensory neurones) may play an important role. In this study, we demonstrate that after a 30-min infusion of 0.15 M HCl acid into the healthy human distal esophagus, we see a reduction in the pain threshold to electrical stimulation of the non-acid-exposed proximal esophagus (9.6 +/- 2.4 mA) and a concurrent reduction in the latency of the N1 and P2 components of the esophageal evoked potentials (EEP) from this region (10.4 +/- 2.3 and 15.8 +/- 5.3 ms, respectively). This reduced EEP latency indicates a central increase in afferent pathway velocity and therefore suggests that hyperexcitability within the central visceral pain pathway contributes to the hypersensitivity within the proximal, non-acid-exposed esophagus (secondary hyperalgesia/allodynia). These findings provide the first electrophysiological evidence that central sensitization contributes to human visceral hypersensitivity.
Asunto(s)
Sistema Nervioso Central/fisiopatología , Esófago/fisiopatología , Adulto , Estimulación Eléctrica , Esófago/efectos de los fármacos , Potenciales Evocados/efectos de los fármacos , Femenino , Humanos , Ácido Clorhídrico/farmacología , Hiperestesia/inducido químicamente , Hiperestesia/fisiopatología , Masculino , Persona de Mediana Edad , Umbral del Dolor/efectos de los fármacos , Tiempo de Reacción/efectos de los fármacos , Sensación/efectos de los fármacos , Cloruro de Sodio/farmacologíaAsunto(s)
Analgésicos/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Neuropéptidos/genética , Canales de Sodio/genética , Analgésicos/uso terapéutico , Animales , Antiinflamatorios no Esteroideos/farmacología , Etiquetas de Secuencia Expresada , Biblioteca de Genes , Marcación de Gen , Humanos , Canal de Sodio Activado por Voltaje NAV1.8 , Narcóticos/farmacología , Neuronas Aferentes/metabolismo , Neuropéptidos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Dolor/tratamiento farmacológico , Dolor/etiología , Reacción en Cadena de la Polimerasa/métodos , Canales de Sodio/metabolismoRESUMEN
Cutaneous sensory neurons that detect noxious stimuli project to the dorsal horn of the spinal cord, while those innervating muscle stretch receptors project to the ventral horn. DRG11, a paired homeodomain transcription factor, is expressed in both the developing dorsal horn and in sensory neurons, but not in the ventral spinal cord. Mouse embryos deficient in DRG11 display abnormalities in the spatio-temporal patterning of cutaneous sensory afferent fiber projections to the dorsal, but not the ventral spinal cord, as well as defects in dorsal horn morphogenesis. These early developmental abnormalities lead, in adults, to significantly attenuated sensitivity to noxious stimuli. In contrast, locomotion and sensori-motor functions appear normal. Drg11 is thus required for the formation of spatio-temporally appropriate projections from nociceptive sensory neurons to their central targets in the dorsal horn of the spinal cord.
Asunto(s)
Proteínas de Homeodominio/metabolismo , Músculo Esquelético/inervación , Proteínas del Tejido Nervioso , Neuronas Aferentes/fisiología , Dolor/fisiopatología , Células del Asta Posterior/fisiología , Piel/inervación , Médula Espinal/fisiología , Factores de Transcripción/metabolismo , Vías Aferentes/embriología , Vías Aferentes/fisiología , Secuencia de Aminoácidos , Animales , Animales Recién Nacidos , Tipificación del Cuerpo , Sondas de ADN , Embrión de Mamíferos , Desarrollo Embrionario y Fetal , Exones , Proteínas de Homeodominio/genética , Calor , Mecanorreceptores/fisiología , Ratones , Ratones Noqueados , Datos de Secuencia Molecular , Actividad Motora , Mutagénesis Sitio-Dirigida , Nociceptores/fisiología , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Transcripción/deficiencia , Factores de Transcripción/genéticaRESUMEN
The development of neuronal excitability involves the coordinated expression of different voltage-gated ion channels. We have characterized the expression of two sensory neuron-specific tetrodotoxin-resistant sodium channel alpha subunits, Na(v)1. (SNS/PN3) and Na(v)1.9 (SNS2/NaN), in developing rat lumbar dorsal root ganglia (DRGs). Expression of both Na(v)1.8 and Na(v)1.9 increases with age, beginning at embryonic day (E) 15 and E17, respectively, and reaching adult levels by postnatal day 7. Their distribution is restricted mainly to those subpopulations of primary sensory neurons in developing and adult DRGs that give rise to unmyelinated C-fibers (neurofilament 200 negative). Na(v)1.8 is expressed in a higher proportion of neuronal profiles than Na(v)1.9 at all stages during development, as in the adult. At E17, almost all Na(v)1.8-expressing neurons also express the high-affinity NGF receptor TrkA, and only a small proportion bind to IB4, a marker for c-ret-expressing (glial-derived neurotrophic factor-responsive) neurons. Because IB4 binding neurons differentiate from TrkA neurons in the postnatal period, the proportion of Na(v)1.8 cells that bind to IB4 increases, in parallel with a decrease in the proportion of Na(v)1.8-TrkA co-expressing cells. In contrast, an equal number of Na(v)1.9 cells bind IB4 and TrkA in embryonic life. The differential expression of Na(v)1.8 and Na(v)1.9 in late embryonic development, with their distinctive kinetic properties, may contribute to the development of spontaneous and stimulus-evoked excitability in small diameter primary sensory neurons in the perinatal period and the activity-dependent changes in differentiation they produce.
Asunto(s)
Ganglios Espinales/metabolismo , Regulación del Desarrollo de la Expresión Génica , Neuronas Aferentes/metabolismo , Neuropéptidos/metabolismo , Canales de Sodio/metabolismo , Envejecimiento/metabolismo , Animales , Antígenos de Diferenciación/análisis , Antígenos de Diferenciación/biosíntesis , Northern Blotting , Ganglios Espinales/citología , Ganglios Espinales/embriología , Inmunohistoquímica , Canal de Sodio Activado por Voltaje NAV1.8 , Canal de Sodio Activado por Voltaje NAV1.9 , Neuronas Aferentes/clasificación , Neuronas Aferentes/citología , Neuronas Aferentes/efectos de los fármacos , Neuropéptidos/efectos de los fármacos , Neuropéptidos/genética , Subunidades de Proteína , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Receptor trkA/análisis , Receptor trkA/biosíntesis , Canales de Sodio/efectos de los fármacos , Canales de Sodio/genética , Tetrodotoxina/farmacologíaRESUMEN
Excitotoxicity due to excessive synaptic glutamate release is featured in many neurological conditions in which neuronal death occurs. Whether activation of primary sensory pathways can ever produce sufficient over-activity in secondary sensory neurons in the dorsal horn of the spinal cord to induce cell death, however, has not been determined. In this study, we asked whether activity in myelinated afferents (A fibers), which use glutamate as a transmitter, can induce cell death in the dorsal horn. Using stereological estimates of neuron numbers from electron microscopic sections, we found that stimulation of A-fibers in an intact sciatic nerve at 10 Hz, 20 Hz, and 50 Hz in 10-minute intervals at a stimulus strength that activates both Abeta and Adelta fibers resulted in the loss of 25% of neurons in lamina III, the major site of termination of large Abeta fibers, but not in lamina I, where Adelta fibers terminate. Furthermore, sciatic nerve lesions did not result in detectable neuron loss, but activation of A fibers in a previously sectioned sciatic nerve did cause substantial cell death not only in lamina III but also in laminae I and II. The expansion of the territory of A-fiber afferent-evoked cell death is likely to reflect the sprouting of the fibers into these laminae after peripheral nerve injury. The data show, therefore, that primary afferent A-fiber activity can cause neuronal cell death in the dorsal horn with an anatomical distribution that depends on whether intact or injured fibers are activated. Stimulation-induced cell death potentially may contribute to the development of persistent pain.
Asunto(s)
Vías Aferentes/metabolismo , Muerte Celular/fisiología , Ácido Glutámico/metabolismo , Fibras Nerviosas Mielínicas/metabolismo , Células del Asta Posterior/metabolismo , Transmisión Sináptica/fisiología , Vías Aferentes/ultraestructura , Animales , Recuento de Células , Estimulación Eléctrica/efectos adversos , Masculino , Microscopía Electrónica , Fibras Nerviosas Mielínicas/ultraestructura , Plasticidad Neuronal/fisiología , Dolor/etiología , Dolor/patología , Dolor/fisiopatología , Enfermedades del Sistema Nervioso Periférico/patología , Enfermedades del Sistema Nervioso Periférico/fisiopatología , Células del Asta Posterior/ultraestructura , Ratas , Ratas Sprague-Dawley , Nervio Ciático/patología , Nervio Ciático/fisiopatología , Nervio Ciático/cirugíaRESUMEN
Mammalian brain sodium channel (BNaC, also known as BNC/ASIC) proteins form acid-sensitive and amiloride-blockable sodium channels that are related to putative mechanosensory channels. Certain BNaC isoforms are expressed exclusively in dorsal root ganglia (DRG) and have been proposed to form the ion channels mediating tissue acidosis-induced pain. With antibody labeling, we find that the BNaC1alpha isoform is expressed by most large DRG neurons (low-threshold mechanosensors not involved in acid-induced nociception) and few small nociceptor neurons (which include high-threshold mechanoreceptors). BNaC1alpha is transported from DRG cell bodies to sensory terminals in the periphery, but not to the spinal cord, and is located specifically at specialized cutaneous mechanosensory terminals, including Meissner, Merkel, penicillate, reticular, lanceolate, and hair follicle palisades as well as some intraepidermal and free myelinated nerve endings. Accordingly, BNaC1alpha channels might participate in the transduction of touch and painful mechanical stimuli.
Asunto(s)
Ganglios Espinales/metabolismo , Canales Iónicos/metabolismo , Mecanorreceptores/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas Aferentes/metabolismo , Canales de Sodio/metabolismo , Canales Iónicos Sensibles al Ácido , Animales , Células CHO , Clonación Molecular , Cricetinae , Canales de Sodio Degenerina , Canales Epiteliales de Sodio , Ganglios Espinales/citología , Canales Iónicos/genética , Masculino , Mecanorreceptores/citología , Ratones , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/genética , Neuronas Aferentes/citología , Especificidad de Órganos , Técnicas de Placa-Clamp , Mapeo Físico de Cromosoma , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Transporte de Proteínas , Ratas , Ratas Sprague-Dawley , Piel/citología , Piel/inervación , Piel/metabolismo , Canales de Sodio/genética , TransfecciónRESUMEN
Inflammation causes the induction of cyclooxygenase-2 (Cox-2), leading to the release of prostanoids, which sensitize peripheral nociceptor terminals and produce localized pain hypersensitivity. Peripheral inflammation also generates pain hypersensitivity in neighbouring uninjured tissue (secondary hyperalgesia), because of increased neuronal excitability in the spinal cord (central sensitization), and a syndrome comprising diffuse muscle and joint pain, fever, lethargy and anorexia. Here we show that Cox-2 may be involved in these central nervous system (CNS) responses, by finding a widespread induction of Cox-2 expression in spinal cord neurons and in other regions of the CNS, elevating prostaglandin E2 (PGE2) levels in the cerebrospinal fluid. The major inducer of central Cox-2 upregulation is interleukin-1beta in the CNS, and as basal phospholipase A2 activity in the CNS does not change with peripheral inflammation, Cox-2 levels must regulate central prostanoid production. Intraspinal administration of an interleukin-converting enzyme or Cox-2 inhibitor decreases inflammation-induced central PGE2 levels and mechanical hyperalgesia. Thus, preventing central prostanoid production by inhibiting the interleukin-1beta-mediated induction of Cox-2 in neurons or by inhibiting central Cox-2 activity reduces centrally generated inflammatory pain hypersensitivity.
Asunto(s)
Sistema Nervioso Central/fisiología , Inflamación/fisiopatología , Interleucina-1/fisiología , Isoenzimas/metabolismo , Dolor , Prostaglandina-Endoperóxido Sintasas/metabolismo , Sistema Nervioso Central/enzimología , Ciclooxigenasa 2 , Dinoprostona/líquido cefalorraquídeo , Inducción Enzimática , Adyuvante de Freund , Inflamación/metabolismo , Neuronas/enzimología , Neuronas/fisiología , Médula Espinal/citología , Médula Espinal/enzimología , Médula Espinal/fisiologíaRESUMEN
Remarkable progress has been made recently in identifying a new gene family related to the capsaicin (vanilloid) receptor, VR1. Using a combination of in silico analysis of expressed sequence tag (EST) databases and conventional molecular cloning, we have isolated a novel vanilloid-like receptor, which we call VRL-2, from human kidney. The translated gene shares 46% and 43% identity with VR1 and VRL-1, respectively, and maps to chromosome 12q23-24.1, a locus associated with bipolar affective disorder. VRL-2 mRNA was most strongly expressed in the trachea, kidney, and salivary gland. An affinity-purified antibody against a peptide incorporating the COOH terminal of the receptor localized VRL-2 immunolabel in the distal tubules of the kidney, the epithelial linings of both trachea and lung airways, serous cells of submucosal glands, and mononuclear cells. Unlike VR1 and VRL-1, VRL-2 was not detected in cell bodies of dorsal root ganglia (DRG) or sensory nerve fibers. However, VRL-2 was found on sympathetic and parasympathetic nerve fibers, such as those innervating the arrector pili smooth muscle in skin, sweat glands, intestine, and blood vessels. At least four vanilloid receptor-like genes exist, the newest member, VRL-2 is found in airway and kidney epithelia and in the autonomic nervous system.
Asunto(s)
Proteínas de Transporte de Catión , Canales Iónicos , Receptores de Droga/genética , Secuencia de Aminoácidos , Animales , Northern Blotting , Línea Celular , Mapeo Cromosómico , Cromosomas Humanos Par 12/genética , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , ADN Complementario/aislamiento & purificación , Femenino , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Masculino , Datos de Secuencia Molecular , ARN/genética , ARN/metabolismo , Mapeo de Híbrido por Radiación , Ratas , Receptores de Droga/metabolismo , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Aminoácido , Canales Catiónicos TRPV , Distribución TisularRESUMEN
Pathological pain, consisting of tissue injury-induced inflammatory and nerve injury-induced neuropathic pain, is an expression of neuronal plasticity. One component of this is that the afferent input generated by injury and intense noxious stimuli triggers an increased excitability of nociceptive neurons in the spinal cord. This central sensitization is an activity-dependent functional plasticity that results from activation of different intracellular kinase cascades leading to the phosphorylation of key membrane receptors and channels, increasing synaptic efficacy. Central sensitization is both induced and maintained in a transcription-independent manner. Several different intracellular signal transduction cascades converge on MAPK (mitogen-activated protein kinase), activation of which appears to be a master switch or gate for the regulation of central sensitization. In addition to posttranslational regulation, the MAPK pathway may also regulate long-term pain hypersensitivity, via transcriptional regulation of key gene products. Pharmacological intervention targeted specifically at the signal transduction pathways in nociceptive neurons may provide, therefore, new therapeutic opportunities for pathological pain.
Asunto(s)
Enfermedades del Sistema Nervioso/fisiopatología , Plasticidad Neuronal/fisiología , Neuronas/fisiología , Nociceptores/fisiología , Dolor/fisiopatología , Transducción de Señal/fisiología , Animales , HumanosRESUMEN
Whole-cell patch-clamp and intracellular recording techniques have been used to study the action of prostaglandin E2 (PGE2) on neurons in adult rat transverse spinal cord slices. Bath-applied PGE2 (1-20 microm) induced an inward current or membrane depolarization in the majority of deep dorsal horn neurons (laminas III-VI; 83 of 139 cells), but only in a minority of lamina II neurons (6 of 53 cells). PGE2 alone never elicited spontaneous action potentials; however, it did convert subthreshold EPSPs to suprathreshold, leading to action potential generation. PGE2-induced inward currents were unaffected by perfusion with either a Ca(2+)-free/high Mg(2+) (5 mm) solution or tetrodotoxin (1 microm), indicating a direct postsynaptic action. Both 17-phenyl trinor prostaglandin E2 (an EP1 agonist) and sulprostone (an EP3 agonist) had little effect on membrane current, whereas butaprost methyl ester (an EP2 agonist) mimicked the effect of PGE2. Depolarizing responses to PGE2 were associated with a decrease in input resistance, and the amplitude of inward current was decreased as the holding potential was depolarized. PGE2-induced inward currents were reduced by substitution of extracellular Na(+) with N-methyl-d-glucamine and inhibited by flufenamic acid (50-200 microm), which is compatible with activation of a nonselective cation channel. These results suggest that PGE2, acting via an EP2-like receptor, directly depolarizes spinal neurons. Moreover, these findings imply an involvement of spinal cord-generated prostanoids in modulating sensory processing through an alteration in dorsal horn neuronal excitability.