Modulation Of Spinal Nociceptive Transmission Research Articles

Inhibition of itch by neurokinin 1 receptor (Tacr1) -expressing ON cells in the rostral ventromedial medulla in mice.

The rostral ventromedial medulla (RVM) is important in descending modulation of spinal nociceptive transmission, but it is unclear if the RVM also modulates spinal pruriceptive transmission. RVM ON cells are activated by noxious algesic and pruritic stimuli and are pronociceptive. Many RVM-spinal projection neurons express the neurokinin-1 receptor (Tacr1), and ON-cells are excited by local administration of substance P (SP). We hypothesized that Tacr1-expressing RVM ON cells exert an inhibitory effect on itch opposite to their pronociceptive action. Intramedullary microinjection of SP significantly potentiated RVM ON cells and reduced pruritogen-evoked scratching while producing mild mechanical sensitization. Chemogenetic activation of RVM Tacr1-expressing RVM neurons also reduced acute pruritogen-evoked scratching. Optotagging experiments confirmed RVM Tacr1-expressing neurons to be ON cells. We conclude that Tacr1-expressing ON cells in RVM play a significant role in the modulation of pruriceptive transmission.

Modulation of Spinal Nociceptive Transmission by Sub-Sensory Threshold Spinal Cord Stimulation in Rats After Nerve Injury

High-frequency spinal cord stimulation (SCS) administered below the sensory threshold (subparesthetic) can inhibit pain, but the mechanisms remain obscure. We examined how different SCS paradigms applied at intensities below the threshold of Aβ-fiber activation (sub-sensory threshold) affect spinal nociceptive transmission in rats after an L5 spinal nerve ligation (SNL). Electrophysiology was used to record local field potential (LFP) at L4 spinal cord before, during, and 0-60 min after SCS in SNL rats. LFP was evoked by high-intensity paired-pulse test stimulation (5 mA, 0.2 msec, 400 msec interval) at the sciatic nerve. Epidural SCS was delivered through a miniature electrode placed at T13-L1 and L2-L3 spinal levels. Four patterns of SCS (200 Hz, 1 msec; 500 Hz, 0.5 msec; 1200 Hz; 0.2 msec; 10,000 Hz, 0.024 msec, 30 min, bipolar) were tested at 90% Aβ-threshold as a subthreshold intensity. As a positive control, traditional SCS (50 Hz, 0.2 msec) was tested at 100% Aβ-plateau as a suprathreshold intensity. Traditional suprathreshold SCS at T13-L1 level significantly reduced LFP to C-fiber inputs (C-LFP). Subthreshold SCS of 200 and 500 Hz, but not 1200 or 10,000 Hz, also reduced C-LFP, albeit to a lesser extent than did traditional SCS (n = 7-10/group). When SCS was applied at the L2-L3 level, only traditional SCS and subthreshold SCS of 200 Hz inhibited C-LFP (n = 8-10/group). Traditional suprathreshold SCS acutely inhibits spinal nociceptive transmission. Low-frequency subthreshold SCS with a long pulse width (200 Hz, 1 msec), but not higher-frequency SCS, also attenuates C-LFP.

In Silico Screening Identified Novel Small-molecule Antagonists of PAC1 Receptor.

Pituitary adenylate cyclase-activating polypeptide (PACAP) and its receptors are present in the spinal dorsal horn and dorsal root ganglia, suggesting an important role of PACAP signaling systems in the modulation of spinal nociceptive transmission. Previously, we found that intrathecal injection of PACAP or maxadilan, a selective PACAP type I (PAC1) receptor agonist, induced transient aversive responses followed by a long-lasting mechanical allodynia in mice, suggesting that PACAP-PAC1 receptor systems are involved in chronic pain and that selective PAC1 antagonists may become a new class of analgesics. Although several PAC1 antagonists, such as PACAP 6-38, have been reported, all of them are peptide compounds. In the present study, we identified new small-molecule antagonists of the PAC1 receptor using in silico screening and in vitro/vivo pharmacological assays. The identified small-molecule compounds, named PA-8 and PA-9, dose dependently inhibited the phosphorylation of CREB induced by PACAP in PAC1-, but not VPAC1- or VPAC2-receptor-expressing CHO cells. PA-8 and PA-9 also dose dependently inhibited PACAP-induced cAMP elevation with an IC50 of 2.0 and 5.6 nM, respectively. In vivo pharmacological assays showed that intrathecal injection of these compounds blocked the induction of PACAP-induced aversive responses and mechanical allodynia in mice. In contrast, the compounds when administered alone exerted neither agonistic nor algesic actions in the in vitro/vivo assays. The compounds identified in the present study are new and the first small-molecule antagonists of the PAC1 receptor; they may become seed compounds for developing novel analgesics.

Venlafaxine and oxycodone effects on human spinal and supraspinal pain processing: a randomized cross-over trial.

Severe pain is often treated with opioids. Antidepressants that inhibit serotonin and norepinephrine reuptake (SNRI) have also shown a pain relieving effect, but for both SNRI and opioids, the specific mode of action in humans remains vague. This study investigated how oxycodone and venlafaxine affect spinal and supraspinal pain processing. Twenty volunteers were included in this randomized cross-over study comparing 5-day treatment with venlafaxine, oxycodone and placebo. As a proxy of the spinal pain transmission, the nociceptive withdrawal reflex (NWR) to electrical stimulation on the sole of the foot was recorded at the tibialis anterior muscle before and after 5days of treatment. For the supraspinal activity, 61-channel electroencephalogram evoked potentials (EPs) to the electrical stimulations were simultaneously recorded. Areas under curve (AUCs) of the EMG signals were analyzed. Latencies and AUCs were computed for the major EP peaks and brain source analysis was done. The NWR was decreased in venlafaxine arm (P=0.02), but the EP parameters did not change. Oxycodone increased the AUC of the EP response (P=0.04). Oxycodone also shifted the cingulate activity anteriorly in the mid-cingulate-operculum network (P<0.01), and the cingulate activity was increased while the operculum activity was decreased (P=0.02). Venlafaxine exerts its effects on the modulation of spinal nociceptive transmission, which may reflect changes in balance between descending inhibition and descending facilitation. Oxycodone, on the other hand, exerts its effects at the cortical level. This study sheds light on how opioids and SNRI drugs modify the human central nervous system and where their effects dominate.

Spinal astrocytic activation contributes to both induction and maintenance of pituitary adenylate cyclase-activating polypeptide type 1 receptor-induced long-lasting mechanical allodynia in mice.

BackgroundPituitary adenylate cyclase-activating polypeptide (PACAP) and its receptors are present in the spinal dorsal horn and dorsal root ganglia, suggesting an important role of PACAP–PACAP receptors signaling system in the modulation of spinal nociceptive transmission. We have previously reported that a single intrathecal injection of PACAP or a PACAP specific (PAC1) receptor selective agonist, maxadilan, in mice induced dose-dependent aversive behaviors, which lasted more than 30 min, and suggested that the maintenance of the nociceptive behaviors was associated with the spinal astrocytic activation.ResultsWe found that a single intrathecal administration of PACAP or maxadilan also produced long-lasting hind paw mechanical allodynia, which persisted at least 84 days without affecting thermal nociceptive threshold. In contrast, intrathecal application of vasoactive intestinal polypeptide did not change mechanical threshold, and substance P, calcitonin gene-related peptide, or N-methyl-D-aspartate induced only transient mechanical allodynia, which disappeared within 21 days. Western blot and immunohistochemical analyses with an astrocytic marker, glial fibrillary acidic protein, revealed that the spinal PAC1 receptor stimulation caused sustained astrocytic activation, which also lasted more than 84 days. Intrathecal co-administration of L-α-aminoadipate, an astroglial toxin, with PACAP or maxadilan almost completely prevented the induction of the mechanical allodynia. Furthermore, intrathecal treatment of L-α-aminoadipate at 84 days after the PAC1 stimulation transiently reversed the mechanical allodynia accompanied by the reduction of glial fibrillary acidic protein expression level.ConclusionOur data suggest that spinal astrocytic activation triggered by the PAC1 receptor stimulation contributes to both induction and maintenance of the long-term mechanical allodynia.

Presynaptic modulation of spinal nociceptive transmission by glial cell line-derived neurotrophic factor (GDNF).

The role of glial cell line-derived neurotrophic factor (GDNF) in nociceptive pathways is still controversial, as both pronociceptive and antinociceptive actions have been reported. To elucidate this role in the mouse, we performed combined structural and functional studies in vivo and in acute spinal cord slices where C-fiber activation was mimicked by capsaicin challenge. Nociceptors and their terminals in superficial dorsal horn (SDH; laminae I-II) constitute two separate subpopulations: the peptidergic CGRP/somatostatin+ cells expressing GDNF and the nonpeptidergic IB4+ neurons expressing the GFRα1-RET GDNF receptor complex. Ultrastructurally the dorsal part of inner lamina II (LIIid) harbors a mix of glomeruli that either display GDNF/somatostatin (GIb)-IR or GFRα1/IB4 labeling (GIa). LIIid thus represents the preferential site for ligand-receptor interactions. Functionally, endogenous GDNF released from peptidergic CGRP/somatostatin+ nociceptors upon capsaicin stimulation exert a tonic inhibitory control on the glutamate excitatory drive of SDH neurons as measured after ERK1/2 phosphorylation assay. Real-time Ca(2+) imaging and patch-clamp experiments with bath-applied GDNF (100 nM) confirm the presynaptic inhibition of SDH neurons after stimulation of capsaicin-sensitive, nociceptive primary afferent fibers. Accordingly, the reduction of the capsaicin-evoked [Ca(2+)]i rise and of the frequency of mEPSCs in SDH neurons is specifically abolished after enzymatic ablation of GFRα1. Therefore, GDNF released from peptidergic CGRP/somatostatin+ nociceptors acutely depresses neuronal transmission in SDH signaling to nonpeptidergic IB4+ nociceptors at glomeruli in LIIid. These observations are of potential pharmacological interest as they highlight a novel modality of cross talk between nociceptors that may be relevant for discrimination of pain modalities.

Diabetic neuropathy affects serotoninergic and Noradrenergic neurons at brainstem centers devoted to pain control

Event Abstract Back to Event Diabetic neuropathy affects serotoninergic and Noradrenergic neurons at brainstem centers devoted to pain control Carla Morgado1*, Terra P. Pereira1, Fani L. Neto1 and Isaura Tavares1 1 IBMC-Instituto de Biologia Celular e Molecular, University of Porto, Institute of Histology and Embryology, Faculty of Medicine of Porto, Portugal Painful diabetic neuropathy is a common complication of diabetes which strongly affects the life quality of diabetic patients. Spinal cord and brain impairments were recently reported in humans and animals during painful diabetic neuropathy. Spinal neuronal hyperactivity observed in streptozotocin-diabetic rats (STZ rat) is probably a contributing factor for pain, but its causes remain unclear. It may result from increased peripheral input or local neurons hyperactivity but may also reflect impaired inhibitory descending modulation of spinal nociceptive transmission, which is mainly mediated by serotonin and noradrenaline. Here, we quantified the serotoninergic and noradrenergic neurons in the brainstem centres involved in descending pain control in STZ rats, by immunodetection of the synthesizing enzymes tryotophan hydroxilase (TpH) and tyrosine hydroxilase (TH), respectively.Male Wistar rats received an intraperitoneal injection of streptozotocin (STZ rats; 60 mg/kg) or saline (controls) and were sacrificed, under chloral hydrate anaesthesia, at 10 weeks post-injection (n= 5 rats per group). Upon vascular perfusion, brain sections were immunoreacted against TpH or TH, using the ABC method. Neurons immunoreactive neurons for TpH (TpH-IR) were counted in dorsal raphe nucleus (DRN), median raphe nucleus (MRN), caudal linear raphe (CLR), nucleus of the tractus solitarius (NTS), rostroventromedial medulla (RVM) and caudal ventrolateral medulla (CVLM). Neurons immunoreactive for TH (TH-IR) were counted in A1, A2, A5 and A7 noradrenergic cell groups. The numbers of TpH-IR neurons were significantly lower in STZ than in control rats at DRN (STZ: 29,6±3,82 vs control: 52,7±8,08; p<0,05), RVM (STZ: 4,9±1,71 vs control: 15,8±4,17; p<0,05) and CVLM (STZ: 1,8±0,28 vs control: 8,6±1,99; p<0,05). The A1 (STZ: 13,5±2,34 vs control: 19,2±0,50; p<0,05), A2 (STZ: 13,8±2,62 vs control: 35,6±3,40; p<0,01) and A5 (STZ: 9,8±0,73 vs control: 13,9±1,34; p<0,05) noragrenergic cell groups presented significant low numbers of TH-IR neurons in STZ than in control rats.These findings point to a deficient inhibitory descending modulation of spinal neurotransmission in STZ rats, which may contribute to spinal cord hyperactivity and concur to spontaneous pain during diabetes.Support by FCT project – PTDC/SAU/64643/2006. Conference: 11th Meeting of the Portuguese Society for Neuroscience, Braga, Portugal, 4 Jun - 6 Jun, 2009. Presentation Type: Oral Presentation Topic: Symposium 5 – Pain Citation: Morgado C, Pereira TP, Neto FL and Tavares I (2009). Diabetic neuropathy affects serotoninergic and Noradrenergic neurons at brainstem centers devoted to pain control. Front. Neurosci. Conference Abstract: 11th Meeting of the Portuguese Society for Neuroscience. doi: 10.3389/conf.neuro.01.2009.11.027 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 05 Aug 2009; Published Online: 05 Aug 2009. * Correspondence: Carla Morgado, IBMC-Instituto de Biologia Celular e Molecular, University of Porto, Institute of Histology and Embryology, Faculty of Medicine of Porto, Porto, Portugal, cmorgado@med.up.pt Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Carla Morgado Terra P Pereira Fani L Neto Isaura Tavares Google Carla Morgado Terra P Pereira Fani L Neto Isaura Tavares Google Scholar Carla Morgado Terra P Pereira Fani L Neto Isaura Tavares PubMed Carla Morgado Terra P Pereira Fani L Neto Isaura Tavares Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Neuropathic pain is maintained by brainstem neurons co-expressing opioid and cholecystokinin receptors

Descending input from the rostral ventromedial medulla (RVM) provides positive and negative modulation of spinal nociceptive transmission and has been proposed to be critical for maintaining neuropathic pain. This study tests the hypothesis that neuropathic pain requires the activity of a subset of RVM neurons that are distinguished by co-expression of mu opioid receptor (MOR) and cholecystokinin type 2 receptor (CCK2). Using male Sprague-Dawley rats, we demonstrate that discrete RVM neurons express MOR and CCK2; over 80% of these cells co-express both receptors. Agonist-directed cell lesion in the RVM with the cytotoxin, saporin, using either CCK-saporin to target CCK receptor expressing cells, or dermorphin-saporin to target MOR expressing cells, resulted in concomitant loss of CCK2 and MOR expressing cells, did not alter the basal sensory thresholds but abolished the hyperalgesia induced by microinjection of CCK into the RVM. The findings suggest that these CCK2-MOR co-expressing RVM neurons facilitate pain and can be directly activated by CCK input to the RVM. Furthermore, lesion of these RVM neurons did not affect the initial development of neuropathic pain in the hind paw upon injury to the sciatic nerve, but the abnormal pain states were short lived such that by about day 9 the sensory thresholds had reverted to pre-injury baselines despite the existing neuropathy. These data support our hypothesis and identify CCK2-MOR co-expressing neurons in the RVM as potential therapeutic targets for neuropathic pain.

Orexin affects dorsal root ganglion neurons: a mechanism for regulating the spinal nociceptive processing.

Orexins (orexin A and B) are initially known to be a hypothalamic peptide critical for feeding and normal wakefulness. In addition, emerging evidence from behavioral tests suggests that orexins are also involved in the regulation of nociceptive processing, suggesting a novel potential therapeutic approach for pain treatment. Both spinal and supraspinal mechanisms appear to contribute to the role of orexin in nociception. In the spinal cord, dorsal root ganglion (DRG) neurons are primary afferent neurons that transmit peripheral stimuli to the pain-processing areas. Morphological results show that both orexin A and orexin-1 receptor are distributed in DRG neurons. Moreover, by using whole-cell patch-clamp recordings and calcium imaging measurements we found that orexin A induced excitability and intracellular calcium concentration elevation in the isolated rat DRG neurons, which was mainly dependent on the activation of spinal orexin-1 receptor. Based on these findings, we propose a hypothesis that the direct effect of orexin A on DRG neurons would represent a possible mechanism for the orexinergic modulation of spinal nociceptive transmission.

Intrathecal sensory neuron-specific receptor agonists bovine adrenal medulla 8-22 and (Tyr 6)-γ2-msh-6-12 inhibit formalin-evoked nociception and neuronal Fos-like immunoreactivity in the spinal cord of the rat

The finding that sensory neuron-specific G-protein-coupled receptor mRNA is solely expressed in small primary sensory neurons suggests involvement of the receptor in nociceptive modulation. The present study was designed to assess effects of intrathecal administration of bovine adrenal medulla 8-22 and (Tyr 6)-γ2-MSH-6-12, selective sensory neuron-specific receptor agonists, on nocifensive behaviors and expression of spinal c-Fos-like immunoreactivity evoked by intraplantar injection of 2.5% formalin in rats. The agonists were administered 10 min before (pretreatment) and/or after (post-treatment) injection of formalin. Pretreatment with bovine adrenal medulla 8-22 dose-dependently (3, 10 and 30 nmol) decreased time lifting and licking the paw mainly in the second phase. Intrathecal bovine adrenal medulla 8-22 (30 nmol) remarkably suppressed nocifensive behaviors in the first and second phases and the expression of formalin-evoked c-Fos-like immunoreactivity in laminae I–II and V–VI of the spinal dorsal horn at L4-5. Moreover, naloxone (20 μg, intrathecal) failed to antagonize the inhibitory effects of bovine adrenal medulla 8-22. Post-treatment with bovine adrenal medulla 8-22 also exerted inhibition on the second phase behaviors in a dose-dependent manner with a similar efficacy observed in pretreatment groups. Furthermore, post-treatment with (Tyr 6)-γ2-MSH-6-12 (0.5, 1.5 and 5 nmol) also suppressed formalin-evoked nocifensive behaviors in the second phase and c-Fos-like immunoreactivity in the spinal dorsal horn similar with bovine adrenal medulla 8-22. Our results suggest that sensory neuron-specific receptor may play an important role in modulation of spinal nociceptive transmission. This is the first to demonstrate that activation of sensory neuron-specific receptor produces analgesia in the persistent pain model.

Anterior cingulate cortex contributes to the descending facilitatory modulation of pain via dorsal reticular nucleus

Supraspinal centres biphasically modulate spinal nociceptive transmission, including descending inhibition and facilitation. Recent studies have revealed that descending facilitatory modulation is a key mechanism underlying induction and maintenance of neuropathic and inflammatory pain. The anterior cingulate cortex (ACC) is not only involved in the transmission of pain sensation but also plays a role in processing pain-related emotion. The ACC also widely connects with relevant regions of the descending modulation system. Here we used electrophysiological and behavioural techniques to study the possible pathways behind the modulation of spinal nociceptive transmission from the ACC. C-fibre-evoked field potentials in the spinal dorsal horn were produced by electrical stimulation of the sciatic nerve at an intensity high enough to excite C fibres, and paw withdrawal latencies (PWLs) to noxious heating were recorded. The results showed that high-frequency tetanic electrical stimulation of the ACC both unilaterally enhanced the C-fibre-evoked field potentials in the spinal dorsal horn and bilaterally shortened PWLs, indicating a facilitation of spinal nociception. A similar effect was observed after microinjection of N-methyl-d-aspartic acid (NMDA; 10 nm, 1 microL) or homocysteic acid (HCA; 0.1 m, 1 microL) into the ACC. When the dorsal reticular nucleus (DRt) was electrolytically lesioned, ACC-induced facilitation of spinal nociception was blocked. These results imply that: (i) activation of the ACC may facilitate spinal nociception; (ii) NMDA receptors in the ACC may be involved in descending facilitation; and (iii) the DRt plays a crucial role in mediating ACC-induced facilitation of spinal nociception.

Spinal inhibitory effects of cardiopulmonary afferent inputs in monkeys: neuronal processing in high cervical segments.

Noxious stimulation of spinal afferents inhibits primate spinothalamic tract (STT) neurons in segments distant from the region of afferent entry. Inhibitory effects of cardiopulmonary sympathetic afferent (CPSA) stimulation remain after C(1) transection but disappear with spinal transection between C(3) and C(7). We hypothesized that spinal inhibitory effects produced by CPSA stimulation are processed by neurons in C(1)-C(3) segments. One purpose of this study in anesthetized monkeys was to determine whether chemical activation of high cervical neurons reduced sacral STT cell responses to colorectal distension (CRD) and urinary bladder distension (UBD). First, effects and interactions of pelvic and cardiopulmonary visceral afferent inputs were determined in 10 monkeys on extracellular activity of sacral STT neurons recorded in deep dorsal horn. CRD and UBD increased activity in 95 and 91% of sacral STT neurons, respectively. CPSA and cardiopulmonary vagal stimulation decreased activity in 84 and 56% of STT neurons, respectively. CPSA stimulation decreased CRD-evoked activity in six of eight sacral STT neurons and decreased UBD-evoked activity in five of eight STT neurons tested. Excitatory amino acid application at C2 segment decreased CRD-evoked responses in 7 of 10 sacral STT neurons and decreased UBD-evoked responses in 9 of 12 STT neurons. The second purpose of this study was to examine responses of C(1)-C(3) descending propriospinal neurons to stimulation of cardiopulmonary afferent fibers. If C(1)-C(3) neurons process CPSA input to suppress STT transmission, then CPSA stimulation should excite C(1)-C(3) neurons with descending projections. Effects of thoracic vagus nerve stimulation also were examined. Vagal stimulation inhibits STT neurons in segments below C(3) but excites C(1)-C(3) STT neurons; we theorized that vagal inhibition of sensory transmission might relay in high cervical segments and, therefore, excite C(1)-C(3) descending propriospinal neurons. Extracellular discharge rate was recorded for C(1)-C(3) neurons antidromically activated from thoracic or lumbar spinal cord in 24 monkeys. CPSA stimulation increased activity of 16 of 45 neurons and inhibited one cell. Thoracic vagus stimulation increased activity of 20 of 43 neurons and inhibited one cell; stimulation of abdominal vagus fibers did not affect activity of six of six cells that were excited by thoracic vagal input. Mechanical stimulation of somatic fields excited 30 of 41 neurons tested. All neurons activated by visceral input received convergent somatic input from noxious pinch of somatic receptive fields that generally included the neck and upper body; 11 C(1)-C(3) propriospinal neurons did not respond to any afferent input examined. Results of these studies were consistent with the idea that modulation of spinal nociceptive transmission might involve neuronal connections in high cervical segments.

Effects of 5-HT 2 and 5-HT 3 receptors on the modulation of nociceptive transmission in rat spinal cord according to the formalin test

We used the formalin test to clarify the 5-hydroxytryptamine (5-HT) receptor subtypes involved in the modulation of spinal nociceptive transmission in rats. Intrathecal administration of a 5-HT 1A receptor agonist, 8-hydroxy-2-(di- n-propylamino)-tetraline (8-OH-DPAT; 1, 10, and 30 μg), or a 5-HT 1B receptor agonist, 1, 4-dihydro-3-(1, 2, 3, 6-tetrahydro-4-pyridinyl)-5H-pyrrol (3, 2- b) pyridin-5-one (CP 93129; 1 and 10 μg), produced no significant change in the number of flinches. A 5-HT 2 receptor agonist, (±)-2, 5-dimethoxy-4-iodoamphetamine (DOI ; 10, 30, and 100 μg), and a 5-HT 3 receptor agonist, 2-methyl-5-HT (100 and 300 μg), produced dose-dependent decreases in the number of flinches in phases 1 (1 to 6 min) and 2 (10 to 61 min) of the test. The antinociceptive effects of DOI and 2-methyl-5-HT were antagonized by intrathecal pretreatment with a 5-HT 2 receptor antagonist, ketanserin, and a 5-HT 3 receptor antagonist, 3-tropanyl-3, 5-dichlorobenzoate (MDL-72222), respectively. These results suggest that 5-HT 2 and 5-HT 3 receptors in the spinal cord mediate antinociception to chemical stimuli.

Effects of spinally administered P2X receptor agonists and antagonists on the responses of dorsal horn neurones recorded in normal, carrageenan-inflamed and neuropathic rats.

1. The function and role of P2X receptors in the spinal transmission of nociception was investigated using the selective P2X receptor agonists, alpha,beta-methylene ATP (alpha,beta-me ATP) and beta, gamma-methylene-L-ATP (beta,gamma-me-L-ATP) and the P2X receptor antagonists pyridoxal-phosphate-6-azophenyl-2',4'-disulphonate (PPADS) and suramin. 2. Intrathecal administration of 5 and 50 microg of beta,gamma-me-L-ATP produced a significant facilitation of the C-fibre evoked response and a tendency towards increased excitability of the post-discharge, but not Abeta-fibre evoked response of dorsal horn neurones recorded in normal animals. Administration of similar doses of alpha,beta-me ATP did not produce an overall change in the response of the neuronal population. 3. Peripheral administration of 20 microg of these agonists into the paw of the rat evoked firing in the dorsal horn neurones. 4. Intrathecal administration of the antagonists, suramin (50 and 500 microg) and PPADS (5, 50 and 500 microg), to normal animals and to animals with a model of neuropathy induced by spinal nerve ligation did not alter the evoked neuronal responses. In contrast, intrathecal administration of 500 microg of suramin to animals 3 h after the induction of carrageenan inflammation produced a significant inhibition of the C-fibre evoked response of the neurones. Similar inhibitions were also seen following high doses of intrathecal PPADS, although this did not reach significance. 5. These results suggest that spinal P2X receptors may play a role in the modulation of spinal nociceptive transmission following the development of inflammation, but that these receptors play at most a minor role in spinal nociceptive processing in normal and neuropathic animals.

Activation of spinal N-methyl-D-aspartate receptors stimulates a nitric oxide/cyclic guanosine 3,5-monophosphate/glutamate release cascade in nociceptive signaling.

Increasing evidence has suggested the possibility that the activation of N-methyl-D-aspartate (NMDA) receptors modulates spinal nociceptive transmission via a nitric oxide (NO)/cyclic guanosine 3',5'-monophosphate (cGMP) pathway. However, the existence and the role of an NO/cGMP pathway in the modulation of spinal nociceptive transmission has been unclear. The authors hypothesized that the activation of NMDA receptors stimulates an NO/cGMP pathway, and this pathway evokes glutamate release within the spinal cord, modulating spinal nociceptive transmission. The authors have examined the effects of an NO synthase inhibitor and a soluble guanylate cyclase inhibitor on the concentrations of NO metabolites (NO2-/NO3-) and glutamate in the cerebrospinal fluid after intrathecal perfusion of NMDA, concomitantly observing pain-related behavior (scratching, biting, and vocalization) in unanesthetized, free-moving rats using an intrathecal microdialysis method. The contents of cGMP in the dorsal horn were also measured using enzyme immunoassay method. Intrathecal perfusion of NMDA produced pain-related behavior and increased glutamate and NO2-/NO3-concentrations in a dose-dependent manner. A competitive NMDA receptor antagonist, D,L-2-amino-5-phosphonovaleric acid, completely blocked the NMDA-induced responses. An NO synthase inhibitor, N(G)-monomethyl-L-arginine acetate, at a dose that completely blocked the increase in NO2-/NO3-, inhibited both the NMDA-induced pain-related behavior and the increase in glutamate concentration. In addition, a soluble guanylate cyclase inhibitor, 1H-[1,2,4]oxadiazole[4,3-a]quinoxaline-1-one, also inhibited significantly NMDA-induced pain-related behavior and the increase in glutamate concentration. NMDA induced an increase in cGMP in the dorsal half of the spinal cord, which was blocked by N(G)-monomethyl-L-arginine acetate. The results of this study support the hypothesis that the activation of NMDA receptors modulated pain-related behavior via an NO/cGMP/glutamate release cascade within the spinal cord.

Effects of moderate and intensive training on the sensitivity of 5-HT 1b receptors in rat brain

We used the formalin test to clarify the 5-hydroxytryptamine (5-HT) receptor subtypes involved in the modulation of spinal nociceptive transmission in rats. Intrathecal administration of a 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)-tetraline (8-OH-DPAT; 1, 10, and 30 μg), or a 5-HT1B receptor agonist, 1, 4-dihydro-3-(1, 2, 3, 6-tetrahydro-4-pyridinyl)-5H-pyrrol (3, 2-b) pyridin-5-one (CP 93129; 1 and 10 μg), produced no significant change in the number of flinches. A 5-HT2 receptor agonist, (±)-2, 5-dimethoxy-4-iodoamphetamine (DOI ; 10, 30, and 100 μg), and a 5-HT3 receptor agonist, 2-methyl-5-HT (100 and 300 μg), produced dose-dependent decreases in the number of flinches in phases 1 (1 to 6 min) and 2 (10 to 61 min) of the test. The antinociceptive effects of DOI and 2-methyl-5-HT were antagonized by intrathecal pretreatment with a 5-HT2 receptor antagonist, ketanserin, and a 5-HT3 receptor antagonist, 3-tropanyl-3, 5-dichlorobenzoate (MDL-72222), respectively. These results suggest that 5-HT2 and 5-HT3 receptors in the spinal cord mediate antinociception to chemical stimuli.

Characterization of biphasic modulation of spinal nociceptive transmission by neurotensin in the rat rostral ventromedial medulla.

Modulation of spinal nociceptive transmission by neurotensin microinjected in the rostral ventromedial medulla (RVM) was examined in anesthetized, paralyzed rats. Forty-three spinal dorsal horn neurons in the L3-L5 spinal segments responding to mechanical and noxious thermal stimulation (50 degrees C) of the plantar surface of the ipsilateral hind foot were studied. Spinal units were classified as either wide dynamic range or nociceptive specific and were located in spinal laminae I-V. Microinjection of neurotensin (0.03 pmol/0.2 microl) into the RVM produced a significant facilitation (135% of control) of spinal unit responses to noxious thermal stimulation (50 degrees C) that lasted approximately 12 min. In contrast, injection of greater doses of neurotensin (300 or 3,000 pmol) produced an inhibition of spinal unit responses to noxious heat (51.7 and 10.6% of control, respectively) that had a longer duration (60-120 min). The effects of neurotensin on wide-dynamic-range and nociceptive-specific neuron responses to noxious heat were qualitatively and quantitatively similar. Spinal unit responses to graded heating of the skin (42-50 degrees C) were completely inhibited after microinjection of 3,000 pmol of neurotensin into the RVM. Injection of a lesser dose of neurotensin (300 pmol), however, resulted in a partial inhibition of spinal unit responses and significantly reduced the slope of the stimulus-response function to graded heating of the skin. Transection of either the ipsilateral or contralateral dorsolateral funiculus (DLF) significantly reduced the inhibition of spinal nociceptive transmission produced by neurotensin (3,000 pmol) in the RVM, whereas bilateral transection of the DLFs completely blocked the effect. In contrast, bilateral transection of the DLFs had no effect on facilitation of spinal nociception by neurotensin (0.03 pmol) in the RVM. The inhibition of spinal nociceptive transmission by neurotensin (3,000 pmol) in the RVM was completely blocked by injection of the nonpeptide neurotensin receptor antagonist SR48692 (30 fmol) into the RVM 10 min before neurotensin. To confirm a specific block of neurotensin-receptor-mediated effects by the antagonist, a subsequent injection of L-glutamate into the RVM was performed. L-Glutamate (100 nmol) was found to inhibit the nociceptive responses of those spinal units whose responses were no longer inhibited by neurotensin. In contrast, injection of SR48692 (30 fmol) into the RVM failed to block the facilitation of spinal unit responses to noxious heat produced by a subsequent injection of neurotensin (0.03 pmol) into the same site. The present series of experiments demonstrate a specific role for neurotensin in the RVM in the modulation of spinal nociceptive transmission, because the peptide was found to both facilitate and inhibit spinal neuron responses to noxious thermal stimulation. Additionally, the facilitatory and inhibitory effects of neurotensin appear to occur via interaction with multiple neurotensin receptors in the RVM that activate independent systems that descend in the ventrolateral funiculi and DLFs, respectively. The results from these experiments are consistent with prior studies demonstrating that the RVM both facilitates and inhibits spinal nociceptive transmission, and they complement previous work showing that neurotensin in the RVM modulates spinal nociceptive behavioral responses.

Biphasic modulation of spinal nociceptive transmission from the medullary raphe nuclei in the rat.

The modulatory effects of electrical and chemical (glutamate) stimulation in the rostral ventromedial medulla (RVM) on spinal nociceptive transmission and a spinal nociceptive reflex were studied in rats. Electrical stimulation at a total 86 sites in the RVM in the medial raphe nuclei (n = 54) and adjacent gigantocellular areas (n = 32) produced biphasic (facilitatory and inhibitory, n = 43) or only inhibitory (n = 43) modulation of the tail-flick (TF) reflex. At these 43 biphasic sites in the RVM, facilitation of the TF reflex was produced at low intensities of stimulation (5-25 microA) and inhibition was produced at greater intensities of stimulation (50-200 microA). At 43 sites in the RVM, electrical stimulation only produced intensity-dependent inhibition of the TF reflex. Activation of cell bodies in the RVM by glutamate microinjection reproduced the biphasic modulatory effects of electrical stimulation. At biphasic sites previously characterized by electrical stimulation, glutamate at a low concentration (5 nmol) produced facilitation of the TF reflex; a greater concentration (50 nmol) only inhibited the TF reflex. In electrophysiological experiments, electrical stimulation at 62 sites in the RVM produced biphasic (n = 26), only inhibitory (n = 26), or only facilitatory (n = 10) modulation of responses of lumbar spinal dorsal horn neurons to noxious cutaneous thermal (50 degrees C) or mechanical (75.9 g) stimulation. Facilitatory effects were produced at lesser intensities of stimulation and inhibitory effects were produced at greater intensities of stimulation. The apparent latencies to stimulation-produced facilitation and inhibition, determined with the use of a cumulative sum method and bin-by-bin analysis of spinal neuron responses to noxious thermal stimulation of the skin, were 231 and 90 ms, respectively. The spinal pathways conveying descending facilitatory and inhibitory influences were found to be different. Descending facilitatory influences on the TF reflex were conveyed in ventral/ventrolateral funiculi, whereas inhibitory influences were conveyed in dorsolateral funiculi. The results indicate that descending inhibitory and facilitatory influences can be simultaneously engaged throughout the RVM, including nucleus raphe magnus, and that such influences are conveyed in different spinal funiculi.

Role of GABA receptor subtypes in inhibition of primate spinothalamic tract neurons: difference between spinal and periaqueductal gray inhibition.

1. gamma-Aminobutyric acid (GABA) is thought to inhibit both pre- and postsynaptically the transfer of nociceptive signals from primary afferent fibers to spinal dorsal horn sensory cells, including spinothalamic tract (STT) neurons. The inhibition can be mediated by both GABAA and GABAB receptors. We now attempt to characterize the synaptic inhibition of STT cells by spinal GABAA and GABAB receptors in anesthetized monkeys and to analyze the roles of these two receptor subtypes in the inhibition of STT cellular activity produced by stimulation in the periaqueductal gray (PAG). 2. Iontophoretic release of GABA or muscimol (a selective GABAA receptor agonist) onto STT cells elicited a profound and dose-related inhibition of the responses of all cells tested to noxious cutaneous stimuli. Only four cells (16.7%) were found to be inhibited when baclofen (a selective GABAB receptor agonist) was applied iontophoretically. However, a strong and dose-dependent inhibition of the responses to cutaneous mechanical and thermal stimuli was obtained in all cells examined when baclofen was administered into the dorsal horn through a microdialysis fiber. The inhibitory effects were mainly on nociceptive inputs. 3. The inhibition of cellular activity by GABAA and GABAB agonists could be selectively antagonized by specific antagonists applied through a microdialysis fiber. 4. The excitatory responses evoked by pulsed release of glutamic acid (GLUT) were also inhibited in a dose-related manner by iontophoretic application of GABA and muscimol, but not by baclofen. A high dose of baclofen administered by microdialysis resulted in only a small decrease in GLUT-evoked excitatory responses. 5. Infusion of GABAA and GABAB antagonists into the dorsal horn by microdialysis caused an increase in both background activity and responses to cutaneous stimuli, suggesting that there is a tonic GABAergic inhibition of STT cells. 6. The inhibition of responses to mechanical and thermal stimulation of the cutaneous excitatory receptive field resulting from stimulation in PAG was significantly antagonized in most of the STT cells tested when the GABAA antagonist bicuculline was infused into the spinal dorsal horn through a microdialysis fiber. In contrast, the inhibition produced by PAG stimulation in most of the cells examined was not significantly antagonized by the GABAB antagonists phaclofen or 3-amino-propyl(diethoxymethyl)phophinic acid (CGP35348) administered into the spinal dorsal horn by microdialysis. 7. Our results support the contention that GABAergic mechanisms in the spinal dorsal horn normally exert a tonic modulation of nociceptive inputs through both GABAA and GABAB receptors. The evidence provided here indicates that GABAA receptors located on primate STT neurons contribute to a postsynaptic inhibitory effect on the transmission of peripheral nociceptive inputs. A possible presynaptic GABAA action was not investigated. Our finding of a GABAB-receptor-mediated inhibition is consistent with the view that both pre- and postsynaptic GABAB receptors are involved in inhibitory modulation of spinal nociceptive transmission. Finally, it is suggested from this study that primate spinal GABAA, but not GABAB receptors, are involved in mediating the descending inhibition induced by PAG stimulation.

Effects of isoflurane concentration on the activity of pontomedullary raphe and medial reticular neurons in the rat

Neurons in the pontomedullary raphe magnus (RM) and adjacent nucleus reticularis paragigantocellularis pars α (NRPGα) are thought to participate in the modulation of spinal nociceptive transmission. In order to determine whether these cells also contribute to the suppression of nocifensive reflexes produced by general anesthetics, the spontaneous activity of RM/NRPGα cells was recorded in rats anesthetized with isoflurane (IF) at several steady state concentrations, corresponding to depths which are below, equal to, or above the threshold for blocking the motor response to noxious stimuli (minimum alveolar concentration, MAC). Neurons were classified by their spontaneous activity patterns and their responses to noxious stimulation as OFF, ON, REGULAR or NEUTRAL cells. After cell classification, unit activity, arterial blood pressure, heart rate, and EEG activity were simultaneously recorded, in the absence of somatic stimulation, for 1 h at each of two or three concentrations of IF. The concentrations tested were low (1.05–1.25%), medium (1.30–1.45%) and high (1.70–1.90%). ON, OFF and some NEUTRAL cells exhibited alternating periods of inactivity and activity when recorded during periods of low and medium anesthetic concentrations. At high steady state anesthetic concentrations, the mean discharge of most OFF, ON and NEUTRAL cells decreased by greater than 25% from their mean discharge rate at the low concentration. REGULAR cells maintained a uniform firing rate at all steady state anesthetic concentrations studied. Since high concentrations of IF do not activate OFF cells, the putative inhibitory output neuron of the RM/NRPGα, it is unlikely that the activity of RM/NRPGα neurons contributes to the suppression of nocifensive movement by the general anesthetic, IF.