RESUMEN
The periaqueductal gray (PAG) is a critical brain region involved in opioid analgesia and provides efferents to descending pathways that modulate nociception. In addition, the PAG contains ascending pathways to regions involved in the regulation of reward, including the substantia nigra (SN) and the ventral tegmental area (VTA). SN and VTA contain dopaminergic neurons that are critical for the maintenance of positive reinforcement. Interestingly, the PAG is also reported to contain a population of dopaminergic neurons. In this study, the distribution of catecholaminergic neurons within the ventrolateral (vl) PAG was examined using immunocytochemical methods. In addition, the catecholaminergic PAG neurons were examined to determine whether these neurons are integrated into ascending (VTA, SN) and descending rostral ventral medulla (RVM) efferent pathways from this region. The immunocytochemical analysis determined that catecholaminergic neurons in the PAG are both dopaminergic and noradrenergic and these neurons have a distinct rostrocaudal distribution within the ventrolateral column of PAG. Dopaminergic neurons were concentrated rostrally and were significantly smaller than noradrenergic neurons. Combined immunocytochemistry and tract tracing methods revealed that catecholaminergic neurons are distinct from, but closely associated with, both ascending and descending efferent projection neurons. Finally, by electron microscopy, catecholaminergic neurons showed close dendritic appositions with other neurons in PAG, suggesting a possible nonsynaptic mechanism for regulation of PAG output by these neurons. In conclusion, our data indicate that there are two populations of catecholaminergic neurons in the vlPAG that form dendritic associations with both ascending and descending efferents suggesting a possible nonsynaptic modulation of vlPAG neurons.
Asunto(s)
Catecolaminas/metabolismo , Vías Eferentes/metabolismo , Neuronas/metabolismo , Sustancia Gris Periacueductal/metabolismo , Animales , Vías Eferentes/ultraestructura , Masculino , Bulbo Raquídeo/metabolismo , Bulbo Raquídeo/ultraestructura , Neuronas/ultraestructura , Sustancia Gris Periacueductal/ultraestructura , Ratas , Ratas Sprague-Dawley , Área Tegmental Ventral/metabolismo , Área Tegmental Ventral/ultraestructuraRESUMEN
UNLABELLED: Individuals with chronic craniofacial pain experience symptoms that are consistent with central sensitization. In fact, central sensitization may constitute the major disease process in these conditions, particularly if the original injury has healed or the condition is idiopathic. To understand central sensitization we have developed a conjugate of substance P and cholera toxin (SP-CTA). SP-CTA is selectively taken up by cells that express neurokinin receptors. Twenty-four hours following intracisternal administration of SP-CTA, wild-type rats and mice demonstrated signs of persistent background nociception, but when tested for facial cold sensitivity, they did not differ from controls. However, treating the SP-CTA-injected animals with naloxone exposed cold hypersensitivity in the face. Mu-opioid receptor knockout mice treated with SP-CTA demonstrated hypersensitivity without naloxone treatment. These findings suggest that central sensitization leads to activation of an endogenous opioid system. The data also demonstrate that the intracisternal administration of SP-CTA in rodents is a useful model for studying central sensitization as a disease process without having to induce a peripheral injury. PERSPECTIVE: Central sensitization is a concern in many craniofacial pain conditions. In this project, we utilize a conjugate of substance P and the catalytic subunit of cholera toxin to induce central sensitization in the nucleus caudalis of rodents. The data indicate that the injected animals become hypersensitive in the face.
Asunto(s)
Toxina del Cólera/farmacología , Sustancia P/farmacología , Núcleo Caudal del Trigémino/efectos de los fármacos , Animales , Conducta Animal/efectos de los fármacos , Western Blotting , Toxina del Cólera/uso terapéutico , Cisterna Magna , Condicionamiento Operante/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Dolor Facial/tratamiento farmacológico , Femenino , Inmunohistoquímica , Inyecciones , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Dimensión del Dolor , Ratas , Ratas sin Pelo , Ratas Sprague-Dawley , Receptores Opioides mu/genética , Receptores Opioides mu/fisiología , Sustancia P/uso terapéuticoRESUMEN
BACKGROUND: Visceral hypersensitivity is a clinical observation made when diagnosing patients with functional bowel disorders. The cause of visceral hypersensitivity is unknown but is thought to be attributed to inflammation. Previously we demonstrated that a unique set of enteric neurons, colospinal afferent neurons (CANs), co-localize with the NR1 and NR2D subunits of the NMDA receptor as well as with the PAR2 receptor. The aim of this study was to determine if NMDA and PAR2 receptors expressed on CANs contribute to visceral hypersensitivity following inflammation. Recently, work has suggested that dorsal root ganglion (DRG) neurons expressing the transient receptor potential vanilloid-1 (TRPV1) receptor mediate inflammation induced visceral hypersensitivity. Therefore, in order to study CAN involvement in visceral hypersensitivity, DRG neurons expressing the TRPV1 receptor were lesioned with resiniferatoxin (RTX) prior to inflammation and behavioural testing. RESULTS: CANs do not express the TRPV1 receptor; therefore, they survive following RTX injection. RTX treatment resulted in a significant decrease in TRPV1 expressing neurons in the colon and immunohistochemical analysis revealed no change in peptide or receptor expression in CANs following RTX lesioning as compared to control data. Behavioral studies determined that both inflamed non-RTX and RTX animals showed a decrease in balloon pressure threshold as compared to controls. Immunohistochemical analysis demonstrated that the NR1 cassettes, N1 and C1, of the NMDA receptor on CANs were up-regulated following inflammation. Furthermore, inflammation resulted in the activation of the PAR2 receptors expressed on CANs. CONCLUSION: Our data show that inflammation causes an up-regulation of the NMDA receptor and the activation of the PAR2 receptor expressed on CANs. These changes are associated with a decrease in balloon pressure in response to colorectal distension in non-RTX and RTX lesioned animals. Therefore, these data suggest that CANs contribute to visceral hypersensitivity during inflammation.
Asunto(s)
Colon/inervación , Hipersensibilidad/metabolismo , Inflamación/complicaciones , Neuronas Aferentes/metabolismo , Receptor PAR-2/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Vísceras/patología , Animales , Conducta Animal/efectos de los fármacos , Colon/efectos de los fármacos , Colon/patología , Diterpenos/farmacología , Ganglios Espinales/efectos de los fármacos , Ganglios Espinales/metabolismo , Hipersensibilidad/etiología , Hipersensibilidad/patología , Inflamación/patología , Canal de Sodio Activado por Voltaje NAV1.9 , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Neuronas Aferentes/efectos de los fármacos , Neuronas Aferentes/patología , Neuropéptidos/metabolismo , Especificidad de Órganos/efectos de los fármacos , Subunidades de Proteína/metabolismo , Ratas , Ratas Sprague-Dawley , Canales de Sodio/metabolismo , Canales Catiónicos TRPV/metabolismo , Tetrodotoxina/farmacología , Ácido Trinitrobencenosulfónico , Vísceras/efectos de los fármacos , Vísceras/metabolismoRESUMEN
Women are more sensitive to most noxious visceral stimuli, both in terms of intensity and frequency. The spinoparabrachial (spino-PBn) pathway is an essential neural circuit for the central relay of viscerosensitive information, but studies characterizing the anatomical and physiological characteristics of this pathway have only been conducted in males. Sex differences in the anatomical and/or physiological organization of the spino-PBn may contribute to the sexually dimorphic incidence rate for visceral pain syndromes. Retrograde labeling and colorectal distention (CRD) induced Fos expression was used to delineate the spino-PBn circuit in male and cycling female Sprague-Dawley rats. The ability of morphine to suppress CRD evoked responses was also examined. Neurons retrogradely labeled from the PBn were localized primarily within the superficial dorsal horn and sacral parasympathetic nucleus of the L5-S1 spinal cord. While no sex differences were noted in either the distribution of spino-PBn neurons or in CRD-induced Fos expression, significantly greater Fos expression was noted specifically in spino-PBn neurons in males compared to females. Morphine selectively attenuated Fos expression in spino-PBn neurons in males, but not females. Subsequent anatomical studies showed significantly reduced mu opioid receptor protein levels and radioligand binding within the PBn of females in comparison to males. Together, these data indicate that there are profound sex differences in how a noxious visceral stimulus and opiates engage the spino-PBn pathway, which may account for the observed clinical differences in visceral pain sensitivity and morphine antinociception.
Asunto(s)
Colon/inervación , Bulbo Raquídeo/patología , Dolor/patología , Caracteres Sexuales , Médula Espinal/patología , Analgésicos Opioides/farmacología , Animales , Autorradiografía , Mapeo Encefálico , Encefalina Ala(2)-MeFe(4)-Gli(5)/metabolismo , Femenino , Masculino , Bulbo Raquídeo/metabolismo , Morfina/farmacología , Vías Nerviosas , Dolor/etiología , Dimensión del Dolor , Ratas , Ratas Sprague-Dawley , Receptores Opioides mu/metabolismo , Médula Espinal/efectos de los fármacos , Estilbamidinas/metabolismo , Estrés Mecánico , Tritio/metabolismoRESUMEN
BACKGROUND: Several investigators have coupled toxins to neuropeptides for the purpose of lesioning specific neurons in the central nervous system. By producing deficits in function these toxin conjugates have yielded valuable information about the role of these cells. In an effort to specifically stimulate cells rather than kill them we have conjugated the neuropeptide substance P to the catalytic subunit of cholera toxin (SP-CTA). This conjugate should be taken up selectively by neurokinin receptor expressing neurons resulting in enhanced adenylate cyclase activity and neuronal firing. RESULTS: The conjugate SP-CTA stimulates adenylate cyclase in cultured cells that are transfected with either the NK1 or NK2 receptor, but not the NK3 receptor. We further demonstrate that intrathecal injection of SP-CTA in rats induces the phosphorylation of the transcription factor cyclic AMP response element binding protein (CREB) and also enhances the expression of the immediate early gene c-Fos. Behaviorally, low doses of SP-CTA (1 microg) injected intrathecally produce thermal hyperalgesia. At higher doses (10 microg) peripheral sensitivity is suppressed suggesting that descending inhibitory pathways may be activated by the SP-CTA induced sensitization of spinal cord neurons. CONCLUSION: The finding that stimulation of adenylate cyclase in neurokinin receptor expressing neurons in the spinal cord produces thermal hyperalgesia is consistent with the known actions of these neurons. These data demonstrate that cholera toxin can be targeted to specific cell types by coupling the catalytic subunit to a peptide agonist for a g-protein coupled receptor. Furthermore, these results demonstrate that SP-CTA can be used as a tool to study sensitization of central neurons in vivo in the absence of an injury.