Wilson SP’s research while affiliated with University of South Carolina and other places

Antidepressants that inhibit the recapture of noradrenaline have variable effects in chronic pain which may be related to the complex role of noradrenaline in pain modulation. Whereas at the spinal cord noradrenaline blocks nociceptive transmission, both antinociception and pronociception were reported after noradrenaline release in the brain. To study the role of noradrenaline in pain modulatory areas of the brain, we elected the dorsal reticular nucleus (DRt), a key pain facilitatory area located at the medulla oblogata. Three studies were performed. First, we show that the infusion in the DRt of nomifensine, which increases local extracellular levels of noradrenaline as shown by in vivo microdialysis, also enhances pain behavioral responses during both phases of the formalin test, a classic inflammatory pain model. Then, we demonstrate that the formalin test triggers the release of noradrenaline in the DRt in a biphasic pattern that matches the two phases of the test. Finally, we show that reducing noradrenaline release into the DRt, using an HSV-1 vector which decreases the expression of tyrosine hydroxylase in noradrenergic DRt-projecting neurons, attenuates pain behavioral responses in both phases of the formalin test. The increased noradrenaline levels induced by the infusion of nomifensine at the DRt, along with the hyperalgesic effects of noradrenaline released at the DRt upon noxious stimulation, indicates that noradrenaline may enhance pain facilitation from the brain. It is important to evaluate if antidepressants that inhibit noradrenaline recapture enhance pain facilitation from the brain herein attenuating their analgesic effects.

N-methyl-D-aspartate receptors (NMDAR) have a role in cardiovascular control at the nucleus tractus solitarii (NTS), eliciting increases or decreases in blood pressure (BP), depending on the area injected with the agonists. In spite of the association between cardiovascular control and pain modulation, the effects of manipulating NMDAR in pain responses have never been evaluated. In this study, we decreased the expression of NMDAR in the NTS using gene transfer to target receptor subunits and evaluate long-term effects. Seven days after the injection of lentiviral vectors containing the NR1a subunit cDNA of NMDAR, in antisense orientation, into the intermediate NTS of Wistar rats, BP was measured, and the formalin test of nociception was performed. The antisense vector induced a decrease of NR1 expression in the NTS and elicited BP rises and hypoalgesia. Antisense vectors inhibited formalin-evoked c-Fos expression in the spinal cord, indicating decreased nociceptive activity of spinal neurons. Using a time-course approach, we verified that the onset of both the increases in BP and the hypoalgesia was at 4 days after vector injection into the NTS. The injection of NMDA into the NTS reversed the effects of antisense vectors in pain behavioral responses and spinal neuronal activation and decreased BP and heart rate. The present study shows that the NR1 subunit of the NMDAR at the NTS is critical in the regulation of tonic cardiovascular and nociceptive control and shows an involvement of the nucleus in the modulation of sustained pain.

Aims Noradrenaline is involved in pain transmission but its role in pain facilitation is poorly understood. We sought to study noradrenaline effects at a supraspinal pain facilitatory area, the dorsal reticular nucleus (DRt). We determined: i) the effect of inflammatory (formalin model) and neuropathic pain (spared nerve injury model; SNI) in the release of noradrenaline at the DRt by in vivo microdialysis , and ii) the effects of noradrenaline at the DRt through local injection of α-adrenoreceptor antagonists or decrease of noradrenaline release using a HSV-1 vector (THa) which decreases tyrosine hydroxlase expression. Methods In vivo microdialysis was performed on naïve male Wistar rats, upon injection of formalin or saline into the hind paw and on SNI or sham rats upon application of hyperalgesic (pin prick) or allodynic (acetone) stimuli into the hind paw. Prazosin and atipamezole, α1 and α2 antagonists respectively, and THa were injected at the DRt and their effects were assessed by evaluating the number of paw jerks induced by formalin or the withdrawal responses induced by pin prick or acetone. Results Noradrenaline released at the DRt increased after formalin injection and hyperalgesic and allodynic stimulation of SNI rats. Prazosin induced antihyperalgesia and antiallodynia in both pain models indicating that noradrenaline enhances pain facilitation through α1-adrenoreceptors. Atipamezole induced opposite effects, likely mediated by presynaptic adrenoreceptors. HSV-1 mediated decrease of noradrenaline significantly attenuated behavioral responses in both pain models. Conclusions Our results indicate that increased noradrenaline release at the DRt contributes to descending pain facilitation. Acknowlegments TDC/SAU-NSC/110954/2009.

Targeting supraspinal pain control centers by gene transfer is known to induce sustained analgesia. In this study, we evaluated the effects of injecting a Herpes Simplex Virus type 1 vector which expresses enkephalin (HSV-ENK vector) in the lateralmost part of the caudal ventrolateral medulla (VLMlat), a pain control center that exerts mainly descending inhibitory effects on pain modulation. Overexpression of enkephalin at the VLMlat reduced the number of flinches during the early and delayed phases of the formalin test and decreased c-fos expression in the spinal cord. These antinociceptive effects were detected at 2 and 10days after injection of HSV-ENK in the VLMlat and were completely reversed by local administration of naloxone. Virally driven-enkephalin was expressed from transduced neurons located in the VLMlat and, at lower extent, in the rostral ventromedial medulla. Our results show that HSV-mediated expression of enkephalin in the VLMlat induced antinociceptive effects, likely due to an enhancement of the opioidergic input to the VLMlat which accounted for descending inhibition of the nociceptive transmission at the spinal cord. This study also demonstrates the value of HSV-1 derived vectors to manipulate, in a sustained and directed manner, pain modulatory pathways in the brain, which is important in the study of supraspinal pain control circuits.

Descending modulation of nociceptive transmission depends on the release of noradrenaline at the spinal cord. The role of noradrenaline in the control of nociceptive transmission at the supraspinal pain control system remains understudied. As chronic pain is associated with enhanced descending facilitation of nociceptive transmission, we sought to determine the role of noradrenaline in pain facilitation from the brain during neuropathic pain. We determined the action of the noradrenergic input to the dorsal reticular nucleus (DRt), a unique pain facilitatory area, using the spared nerve injury model. Injections of the α1-adrenoreceptor agonist phenylephrine into the DRt induced hyperalgesia and allodynia, indicating that α1-adrenoreceptors enhance the facilitatory action of the nucleus. This led us to reduce noradrenaline release at the DRt using a viral vector derived from the Herpes Simplex virus type 1 (HSV-1) which carried the tyrosine hydroxylase (TH) transgene in antisense orientation. The reduction of noradrenaline release, confirmed by microdialysis experiments, induced a long-lasting attenuation of pain responses, which was reverted by the local administration of phenylephrine. The present study indicates that the noradrenergic modulation of a pronociceptive area at the supraspinal pain control system accounts for pain facilitation, through the activation of α1-adrenoreceptors. The study also shows that sustained effects on chronic pain can be achieved by decreasing the release of noradrenaline in a pain facilitatory centre of the brain using gene transfer.

Gene therapy for pain control has mainly focused on the application of viral vectors at the peripheral nervous system. However, targeting pain inhibitory supraspinal control centres, such as the caudal ventrolateral medulla (VLM), could be a more effective strategy. We tested the analgesic effects of a recombinant herpes simplex virus (HSV-1) driving the expression of the human pre-proenkephalin (PPE) cDNA, into the VLM, in the formalin model of inflammatory pain. Male Wistar rats were stereotaxically injected into the VLM with the HSV-1 vector carrying PPE or lacZ, under control of the human cytomegalovirus promoter. Two days after injection, the animals received a subcutaneous injection of 50 ul of formalin at 5% into the hindpaw and the number of flinches was recorded during 5-min periods for 60 min. The animals were transcardially perfused following 2 hours after formalin injection, the brains and the L4 spinal segment were removed and serially cut. The spinal sections were immunocytochemically processed to evaluate the activation of the proto-oncogene c-fos. Brain sections from animals injected with the HSV-1 vector carrying lacZ were processed with an antibody against -galactosidase to establish the patterns of neuronal transduction. The HSV-1 vector carrying PPE significantly reduced the number of flinches both in the early and delayed phases of the formalin test and significantly reduced the numbers of superficial dorsal horn cells expressing c-fos. Numerous -galactosidase labelled neurons were observed at the injection site with only a few neurons found in some VLM brain afferents, which indicates that the antinociceptive effects of the PPE vector is mediated by the VLM. The reduction of c-fos expression in the dorsal horn along with the behavioural effects suggests that the PPE vector enhances descending inhibition from the VLM. The results reinforce the value of gene transfer directed to supraspinal pain control areas in the treatment of pain.

Herpes Simplex Virus type 1 (HSV-1) vectors are known to inhibit nociceptive transmission at the spinal cord after peripheral applications. Similar approaches may also be useful when applied at the supraspinal pain control system as the system includes pronociceptive (facilitatory) components. We performed a study aimed to analyse the migration of HSV-1 along with the inhibition of pronociception from the medullary dorsal reticular nucleus (DRt), a major facilitatory component of the supraspinal pain control system. To study the migration, a HSV-1 vector expressing lacZ under control of the human cytomegalovirus (hCMV) promoter was injected in the DRt and the expression of beta-galactosidase (beta-gal) was detected at 2, 4, 7, 10 and 14 days. Numerous beta-gal-immunoreactive neurons were observed at the injection site until day 4, and at some of the brain areas projecting to the DRt until day 7. To block the pronociceptive effects of the DRt, a HSV-1 vector expressing the preproenkephalin transgene, under the control of hCMV promoter, was injected into the DRt. Behavioural evaluation was performed at the time-points referred above, using the paw withdrawal latency test to evaluate thermal nociceptive responses. Anti-hyperalgesic effects persisted during 4 days, decreasing after that time-point. The present study demonstrates that selective migration of HSV-1 should be considered in gene therapy strategies based on HSV-1 injections into the brain. The study also shows that it is possible to decrease pain facilitation from the brain using opioidergic inhibition of pronociceptive supraspinal areas.