Somatic pain--pathogenesis and prevention.
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Central Nervous System↗
Biomedical subjects
C J Woolf
Publications and source records attributed to C J Woolf.
Contribution of interleukin-1 beta to the inflammation-induced increase in nerve growth factor levels and inflammatory hyperalgesia.
1. Peripheral inflammation is associated with the local production of neuroactive inflammatory cytokines and growth factors. These may contribute to inflammatory pain and hyperalgesia by directly or indirectly altering the function or chemical phenotype of responsive primary sensory neurones. 2. To investigate this, inflammation was produced by the intraplantar injection of complete Freund's adjuvant (CFA) in adult rats. This resulted in a significant elevation in interleukin-1 beta (IL-1 beta) and nerve growth factor (NGF) levels in the inflamed tissue and of the peptides, substance P and calcitonin gene-related peptide (CGRP) in the L4 dorsal root ganglion 48 h post CFA injection. 3. The effects of a steroidal (dexamethasone) and a non-steroidal (indomethacin) anti-inflammatory drug on the levels of NGF and IL-1 beta in inflamed tissue were investigated and compared with alterations in behavioural hyperalgesia and neuropeptide expression in sensory neurones. 4. Systemic dexamethasone (120 micrograms kg-1 per day starting the day before the CFA injection) had no effect on the inflammatory hyperalgesia. When the dose was administered 3 times daily, a reduction in mechanical and to a lesser extent thermal sensitivity occurred. Indomethacin at 2 mg kg-1 daily (i.p.) had no effect on the hyperalgesia and a dose of 4 mg kg-1 daily was required to reduce significantly mechanical and thermal hypersensitivity. 5. The increase in NGF produced by the CFA inflammation was prevented by both dexamethasone and indomethacin, but only at the higher dose levels. Dexamethasone at the lower and higher dose regimes diminished the upregulation of IL-1 beta whereas indomethacin had an effect only at the higher dose. 6. The increase in SP and CGRP levels produced by the CFA inflammation was prevented by dexamethasone and indomethacin at the lower and higher dose regimes. 7. Intraplantar injections of IL-1 beta (0.01, 0.1 and 1 ng) produced a brief (6 h) thermal hyperalgesia and an elevation in cutaneous NGF levels which was prevented by pretreatment with human recombinant IL-1 receptor antagonist (IL-1 ra) (0.625 microgram, i.v.). The thermal hyperalgesia but not the NGF elevation produced by intraplantar IL-1 beta (1 ng) was prevented by administration of a polyclonal neutralizing anti-NGF serum. 8. IL-1 ra significantly reduced the mechanical hyperalgesia produced by CFA for 6 h after administration as well as the CFA-induced elevation in NGF levels. Anti-NGF pretreatment substantially reduced CFA-induced mechanical and thermal hyperalgesia without reducing the elevation in IL-1 beta. 9. Intraplantar NGF (0.02, 0.2 and 2 microg) injections produced a short lasting thermal and mechanical hyperalgesia but did not change IL-1beta levels in the hindpaw skin.10. Our results demonstrate that IL-1beta contributes to the upregulation of NGF during inflammation and that NGF has a major role in the production of inflammatory pain hypersensitivity.
Animals↗
Neurite outgrowth and GAP-43 mRNA expression in cultured adult rat dorsal root ganglion neurons: effects of NGF or prior peripheral axotomy.
Adult dorsal root ganglion (DRG) cells are capable of neurite outgrowth in vivo and in vitro after axotomy. We have investigated, in cultured adult rat DRG cells, the relative influence of nerve growth factor (NGF) or a prior peripheral nerve lesion on the capacity of these neurons to produce neurites. Since there is evidence suggesting that the growth-associated protein GAP-43 may play a crucial role in axon elongation during development and regeneration, we have also compared the effect of these treatments on GAP-43 mRNA expression. NGF increased the early neurite outgrowth in a subpopulation of DRG cells. This effect was substantially less, however, than that resulting from preaxotomy, which initiated an early and profuse neurite outgrowth in almost all cells. No difference in the expression of GAP-43 mRNA was found between neurons grown in the presence or absence of NGF over 1 week of culture, in spite of the increased growth produced by NGF. In contrast, cultures of neurons that had been preaxotomized showed substantial increases in GAP-43 mRNA and NGF had, as expected, a significant effect on substance P mRNA levels. Two forms of growth may be present in adult DRG neurons: an NGF-independent, peripheral nerve injury-provoked growth associated with substantial GAP-43 upregulation, and an NGF-dependent growth that may underlie branching or sprouting of NGF-sensitive neurons, but which is not associated with increased levels of GAP-43 mRNA.
Animals↗
Nerve growth factor levels in developing rat skin: upregulation following skin wounding.
Levels of nerve growth factor (NGF) in rat hindpaw skin, measured with a sensitive two-site enzyme-linked immunosorbent assay, show two peaks during normal development. The first (57 +/- 5 pg mg-1) occurs at embryonic days (E) 18-20 and coincides with the arrival of axon terminals into the hindpaw skin. The second, larger peak (132 +/- 10 pg mg-1), occurs later, around postnatal day (P) 21 and may be involved in maintenance of neuronal phenotype. Levels outside the two peaks stay relatively constant throughout development (30 pg mg-1). Skin wounding at birth produces a marked increase in NGF levels (149 +/- 25 pg mg-1) which declines after 4 days. This large increase is not observed if wounding is performed at older ages and may underlie the sensory hyperinnervation that accompanies neonatal wounds.
Aging↗
NK1 and NK2 receptors contribute to C-fibre evoked slow potentials in the spinal cord.
Small diameter primary afferents produce slow synaptic potentials in spinal neurones. These can be recorded as prolonged ventral root potentials (VRPs) in the isolated neonatal rat spinal cord preparation. The VRP elicited by stimulating C-fibres consists of two phases: an early phase comprising a monosynaptic and a short polysynaptic response identical to that elicited by low threshold A beta fibres and a late phase which has an initial component (0.1-1 s) which is sensitive to N-methyl-D-aspartic acid receptor antagonism and a very long lasting second component (1-20 s) which is resistant to these antagonists. We now demonstrate that the slowest component of the VRP is significantly reduced by both NK1 and NK2 tachykinin receptor antagonists and as a consequence, tachykinins have a particular contribution to the cumulative depolarization produced by low frequency (1 Hz) C-fibre stimulation.
Amino Acid Sequence↗
Cobalt accumulation in neurons expressing ionotropic excitatory amino acid receptors in young rat spinal cord: morphology and distribution.
Excitatory amino acids (EAA) acting on N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and kainate receptors play an important role in synaptic transmission in the spinal cord. Quantitative autoradiography and physiological experiments suggest that NMDA receptors are localized mainly in lamina II while kainate and AMPA receptors are found on both dorsal and ventral horn neurons. However the cell types expressing EAA receptors and their laminar distribution is not known. We have used a cobalt uptake method to study the morphology and distribution of spinal cord neurons expressing AMPA, kainate, or NMDA excitatory amino acid receptors in the lumbar enlargement of the rat spinal cord. The technique involved superfusion of hemisected spinal cords of 14 day-old rat pups in vitro with excitatory amino acid receptor ligands in the presence of CoCl2. Cobalt has been shown to enter cells through ligand-gated ion channels in place of Ca2+. Cells which accumulated cobalt ions following activation by ionotropic excitatory amino acid receptors were visualized histochemically. The cobalt uptake generated receptor-specific labeling of cells, as the NMDA receptor antagonist D-(-)-2-amino-(5)-phosphonovaleric acid (D-AP-5) (20 microM) blocked the NMDA, but not kainate-induced cobalt uptake. The kainate-induced cobalt labeling was reduced by the non-selective excitatory amino acid receptor antagonist kynurenic acid (4 mM). Passive opening of the voltage-gated Ca(2+)-channels by KCl (50 mM) did not result in cobalt uptake, indicating that cobalt enters the cells through ligand-gated Ca(2+)-channels. AMPA (500 microM), kainate (500 microM), or NMDA (500 microM) each induced cobalt uptake with characteristic patterns and distributions of neuronal staining. Overall, kainate induced cobalt uptake in the greatest number of neuronal staining. Overall, kainate induced cobalt uptake in the greatest number of neuronal perikarya while NMDA-induced uptake was the lowest. AMPA and kainate, but not NMDA superfusion, resulted in cobalt labeling of glial cells. Our results show that the cobalt uptake technique is a useful way to study the morphology and distribution of cells expressing receptors with ligand-gated Ca2+ channels.
Animals↗
The downregulation of GAP-43 is not responsible for the failure of regeneration in freeze-killed nerve grafts in the rat.
Freeze-killed nerve grafts in rats are able to support limited axonal regeneration from severed peripheral nerves, but by 6 weeks postoperation, axonal elongation through the grafts ceases. To find out whether this limited regeneration may be related to GAP-43 expression, 4-cm freeze-killed nerve grafts were attached to the proximal stumps of severed tibial nerves in adult inbred Fischer rats. For comparison, tibial nerve crush, to allow functional regeneration, or section and ligation, which allows only abortive axonal sprouting, were also performed. After survival for 3 or 6 weeks, the lumbar spinal cord and L4 dorsal root ganglia were stained for GAP-43 mRNA. Freeze-killed grafts of 3-8 weeks duration were processed for GAP-43 immunocytochemistry. Three weeks after all three operations, comparable numbers of axotomized spinal motorneurons and primary sensory DRG neurons reexpressed high levels of GAP-43 mRNA. Six weeks after tibial nerve crush, the number of tibial motorneurons and DRG cells expressing GAP-43 mRNA returned to control levels but after section and ligation or freeze-killed nerve grafting many positively stained cells were still visible. GAP-43 immunoreactivity was detectable using immunocytochemistry in many unmyelinated axons which had regenerated into the freeze-killed grafts at all times. Both axonal profiles in contact with Schwann cells and those which lacked such contact were GAP-43 positive. These results suggest that the cessation of axonal regeneration into freeze-killed tibial nerve grafts is not the result of a down-regulation of GAP-43. Furthermore, the presence of high levels of GAP-43 alone is not sufficient to ensure prolonged axonal regeneration.
Animals↗
Nerve growth factor contributes to the generation of inflammatory sensory hypersensitivity.
Experimental inflammation produced by an intraplantar injection of complete Freund's adjuvant results in local sensory hypersensitivity and up-regulates the neuropeptides substance P and calcitonin gene related peptide in the primary sensory neurons innervating the inflamed tissue. The inflammation also elevates nerve growth factor levels in the skin. Systemic administration of anti-NGF neutralizing antibodies prevent the behavioral sensitivity, the up-regulation of neuropeptides and the inflammation-induced expression of the immediate early gene c-fos in dorsal horn neurons, without modifying swelling and erythema. Elevation of the neurotrophin NGF in the periphery is a major contributor, therefore, of inflammatory pain.
Animals↗
The pathophysiology of chronic pain--increased sensitivity to low threshold A beta-fibre inputs.
Chronic pain is characterized by abnormal sensitivity, which is due to the generation of pain in response to the activation of the low-threshold mechanoreceptive A beta fibres that normally generate innocuous sensations. Three different processes in the spinal cord can account for this dramatic alteration in sensory processing in the somatosensory system: increased excitability, decreased inhibition and structural reorganization. All have been shown to occur and each may contribute separately or together to the wide range of chronic inflammatory and neuropathic pain disorders. The unravelling of the cellular mechanisms involved both offers the potential for developing novel therapeutic strategies, which reduce functional synaptic plasticity and prevent central atrophic and regenerative responses in injured neurones, and illustrates the capacity of the adult nervous system for maladaptive modification.
Animals↗
The contribution of GABAA and glycine receptors to central sensitization: disinhibition and touch-evoked allodynia in the spinal cord.
1. Pain hypersensitivity is characterized by an increase in the response to noxious stimuli (hyperalgesia) and a reduction in threshold such that innocuous stimuli begin to elicit pain (allodynia). These sensitivity changes can be produced by an increase in excitability of dorsal horn neurons; the phenomenon of central sensitization. We have now examined whether a reduction in local segmental inhibitory mechanisms produces similar changes. The model system used for studying touch-evoked allodynia has been the recruitment of a low-threshold mechanoreceptor input to the nociceptive flexion withdrawal reflex in the decerebrate-spinal rat. 2. Hamstring flexor alpha motoneurons are characterized by high-threshold cutaneous receptive fields. Mechanical stimuli (pinch or firm pressure) evoke a brisk firing response in these cells, whereas low-intensity stimuli (light touch or brush) produce little or no effect, as expected for the output neurons of the nociceptive flexion withdrawal reflex. 3. Primary afferent C fiber conditioning inputs have previously been shown to produce prolonged increases in the excitability of the flexion reflex, as measured by the augmentation of the response to high-intensity peripheral stimuli. We have now examined whether these conditioning inputs and segmental disinhibition modify the responsiveness of the reflex to low-threshold inputs. 4. Brief (20 s), low-frequency (1 Hz), C fiber conditioning stimuli to the sural nerve increased the response of the hamstring flexor motor neurons to low-intensity cutaneous touch stimuli, reduced the cutaneous mechanical threshold, and increased the response to A beta inputs from the sural nerve. 5. Intrathecal injections of subconvulsant doses of the glycine receptor antagonist, strychnine (7 nmol) or the gamma-aminobutyric acid-A (GABAA) receptor antagonist, bicuculline (8 nmol) produced similar but longer lasting changes. The GABAB antagonist P-(3-aminopropyl)-P-diethoxymethyl-phosphonic acid (CGP 35348) had no significant effects. 6. The nociceptive flexion withdrawal reflex is under the control, therefore, of segmental inhibitory mechanisms mediated by glycine and GABAA receptors. Removal of this inhibition enables the reflex to be activated by low-intensity cutaneous stimuli. Given the similarities between the stimulus-response profiles of the nociceptive flexion reflex and the production of pain in man, these findings indicate that a decrease in the efficacy of spinal inhibitory circuits may contribute to the touch-evoked allodynia that occurs in pain hypersensitivity states, where A beta inputs begin to produce pain.
Afferent Pathways↗
A new strategy for the treatment of inflammatory pain. Prevention or elimination of central sensitization.
The optimal treatment of pain requires an understanding of the mechanisms involved. Pain is a sensory end-point that can be generated by a number of dissimilar processes. Consequently, the concept of treating pain as a unitary symptom is obsolete. The mechanisms responsible for specific types of pain need to be understood, and particular treatments should be aimed selectively at the various subtypes of pain. A major breakthrough in our understanding of pain has come from the appreciation that clinical pain is qualitatively quite different from physiological or nociceptive pain and is characterised by the appearance of abnormal hypersensitivity. Clinical pain is more than a reflection of sustained peripheral input and it is, to a large extent, the expression of changes produced in the CNS, including the phenomenon of central sensitization. We need to treat both the disease/injury process in the periphery and the changes it induces or triggers in the CNS. Prevention of central sensitization will substantially eliminate the hyperalgesia and allodynia that patients find so distressing, and it offers new possibilities for the development of novel analgesics or antihypersensitivity drugs.
Analgesics↗
GAP-43 expression in primary sensory neurons following central axotomy.
Primary sensory neurons are capable of successful regenerative growth in response to peripheral nerve but not dorsal root injury. The present study is concerned with the differential expression of the mRNA for GAP-43, a growth-associated protein, in these sensory neurons, in response to injury of their central or peripheral axonal branches. Peripheral axotomy resulted in an elevation in message detectable within 24 hr, using Northern blot and in situ hybridization, which was maintained for 30 d, whereas dorsal root section produced no change except a transient and small increase if the axotomy was immediately adjacent to the dorsal root ganglia (DRG). Dorsal root section had no effect on GAP-43 mRNA levels in the dorsal horn or in neighboring intact DRG. It also failed to alter the laminar boundaries of the GAP-43 central terminal labeling produced by peripheral nerve section, even though vacant synaptic sites were produced in unstained laminae by this procedure. This indicates that the location of GAP-43 immunolabeling in the central terminals of primed sensory cells may not depend only on the location of vacant synaptic sites. We conclude that distinct control mechanisms regulate the response of DRG neurons to peripheral nerve and dorsal root injury, and these may be related both to the glial environment and the particular target influences exerted on the central and peripheral branches of the primary sensory neuron. Central denervation alone is insufficient to upregulate GAP-43 levels, and this may explain the relative absence of collateral sprouting after the production of central vacant synaptic sites. The failure of dorsal root section to increase GAP-43 expression may contribute to the poor regenerative response initiated by such lesions.
Animals↗
Preoperative morphine pre-empts postoperative pain.
Postoperative analgesia is usually inadequate, perhaps because conventional approaches to pain relief do not take account of underlying mechanisms. Pre-emptive analgesia may prevent nociceptive inputs generated during surgery from sensitising central neurons and, therefore, may reduce postoperative pain. In a randomised, double-blind study, we compared the effect of parenteral morphine when given before or after total abdominal hysterectomy in 60 patients. 10 mg of morphine were given intramuscularly 1 hour before operation (im pre), intravenously at induction of anaesthesia (iv pre), or intravenously at closure of the peritoneum (iv post). Response was assessed by morphine consumption from patient-controlled analgesia machines which was found to be significantly reduced in the iv pre group for 24 hours after operation compared with the iv post group. Pain sensitivity around the wound was reduced in both preoperative treatment groups compared with the iv post group. We conclude that pre-emptive analgesia with intravenous morphine, by preventing the establishment of central sensitisation during surgery, reduces postoperative pain, analgesic requirements, and secondary hyperalgesia.
Adult↗
Morphological and membrane properties of young rat lumbar and thoracic dorsal root ganglion cells with unmyelinated axons.
Membrane and morphological properties of thoracic (Th9-13) and lumbar (L2-5) dorsal root ganglion cells have been investigated in an in vitro dorsal root ganglion (DRG) preparation from 14-day-old rats using intracellular recordings and the intracellular injection of Neurobiotin. The passive and active membrane properties of 47 DRG cells with conduction velocities (CV) less than 0.81 m/s were studied, which were considered to possess unmyelinated axons. The action potentials elicited by the stimulation of peripheral nerves or the dorsal roots were characteristic of C-cells, with long duration, inflexion on the falling phase and long lasting after hyperpolarization. Input resistance of the C-cells varied between 16 and 158 M omega and were significantly higher in thoracic than in the lumbar ganglia. Cells in the more cranial levels also tended to be smaller than those in the caudal levels with a mean cross sectional area of 301 +/- 32.5 microns2. Twenty-five percent of the cells from both regions showed an inward rectification. The distribution of CVs, input resistances and cross sectional areas were non-normal. While a weak correlation was found between the conduction velocity and input resistance of the cells, no correlation was present between the size of the perikarya and conduction velocity or the input resistance. These results show that by the 14th day of postnatal development membrane and morphological parameters approach those of adult rats. They also suggest that in cells with unmyelinated fibres, the size of the perikaryon does not predict the thickness of the axon, and that this cell population is heterogeneous.
Action Potentials↗
Chronic peripheral nerve section results in a rearrangement of the central axonal arborizations of axotomized A beta primary afferent neurons in the rat spinal cord.
In order to investigate the reorganization of the neuropil of the dorsal horn following peripheral nerve injury, the central terminal arborizations of 35 A beta primary afferent neurons, chronically injured by a cut and ligation of the sural nerve 6-12 weeks previously, were studied by the intra-axonal injection of horseradish peroxidase. Their morphology was compared to 13 intact sural nerve hair follicle afferents. Following axotomy, three kinds of morphological abnormalities were observed in the collateral arbors of the 26 afferents that were hair follicle-like. Atrophy with thin stem axons and reduced terminal branch patterns with few boutons was seen in 5 afferents. Sprouting of bouton-containing terminals into lamina I and IIo was found in 8 afferents. Finally, abnormal arborization patterns in the deeper laminae were observed in 29% of the collateral arbors. Changes included the loss in some arbors of a flame-shaped appearance, which is characteristic of hair follicle afferents, atypical branching patterns and ventrally directed axons producing wider and deeper arbors, compared to normal. Axotomy also caused a disruption of the normal somatotopic organization of sural nerve A beta afferents. This disruption manifested as a variability in the normally mediolaterally restricted terminal sheet, with a consequent loss of the strict somatotopic register in the rostrocaudal direction. Damage to the peripheral axon of A beta primary afferents induces a structural reorganization of their central terminals in the dorsal horn of the spinal cord, which may modify sensory input to the central nervous system.
Animals↗
Morphology and somatotopy of the central arborizations of rapidly adapting glabrous skin afferents in the rat lumbar spinal cord.
The central arborizations in the dorsal horn of the spinal cord of 23 rapidly adapting (RA) A-beta primary afferent neurons innervating different regions of the glabrous skin of the hindpaw were studied by the intra-axonal injection of horseradish peroxidase in adult rats. A total of 284 arbors of the complex, simple, and blind-ending variety were recovered. The arbors of RA afferents innervating the toes, paw pads, and non-pad hindpaw differed from each other in branch pattern and dimensions. The simple and complex arbors, which are both bouton-containing, were distributed mainly in laminae III-V, although some complex arbors projected dorsally into lamina IIi. The hindpaw glabrous skin afferent terminals were located in the lumbar enlargement from caudal L3 to rostral L6. A crude somatotopic organization was observed such that toes 1-5 were represented successively in more caudal positions from mid-L4 to caudal L5. The paw pads were organized in a rostrocaudal sequence moving from the paw pads proximal to toe 1 across the foot to the paw pads proximal to toe 5, from caudal L3 to mid-L5. Non-pad hindpaw afferents were located in caudal L5. Overlap between toe, paw pad and non-pad afferent central fields was present, however, and the central terminals of afferents with non-adjacent peripheral receptive fields were shown to occupy the same region of the dorsal horn.
Adaptation, Physiological↗
The pathophysiology of peripheral neuropathic pain--abnormal peripheral input and abnormal central processing.
A review is given on the pathogenesis of peripheral neuropathic pain. Central neuropathic pain resulting from damage of the spinal cord or brain is not covered. The following conclusions are proposed. At the time of peripheral injury, an abnormal injury discharge may be sufficient to produce long term changes in the excitability of the spinal cord and/or an excitotoxic death of dorsal horn neurons. These acute changes might set the scene for the maintenance of sensory disorders both as a result of an ongoing ectopic input, which might persistently induce a state of central sensitization and for a structural reorganization of the synaptic connections of the dorsal horn. The implications of these findings is that it may be possible to prevent some of the long term consequences of nerve damage, that the treatment at the time of injury may need to be quite different for that required later, and finally that treatment directed only at the periphery may be insufficient to eliminate the sensory disturbance of chronic neuropathic pain.
Cell Death↗
Partial denervation of the medial gastrocnemius muscle results in growth-associated protein-43 immunoreactivity in sprouting axons and Schwann cells.
Regeneration in the mammalian peripheral nervous system following nerve injury is associated with the upregulation of a developmentally regulated phosphoprotein, growth-associated protein-43 (GAP-43), in the injured neurons. We have examined whether uninjured adult neurons also express GAP-43 when they sprout. The model system investigated has been the sprouting induced in the terminal axons of intact motor neurons by a partial muscle denervation. Partial denervation of the medial gastrocnemius muscle in adult rats was produced by resecting the terminal nerve supply to the anterolateral quadrant of the muscle. Three zones could be identified in the motor endplate region of the muscle after such a denervation using protein gene product (PGP) 9.5, calcitonin gene-related peptide and silver staining as axonal markers and S-100 to identify Schwann cells: a normally innervated zone, a totally denervated zone and a border or intermediate zone between the two which contained axons at the endplates with nodal and terminal sprouts. The endplates in the normally innervated zone were GAP-43 negative. In the denervated zone, Schwann cells were GAP-43 positive and had a distinctive appearance with a lack of any normal endplate organization. Endplates in the intermediate zone were GAP-43 immunoreactive. In approximately half, the GAP-43 immunoreactivity was axonal-like, identical to PGP 9.5 in an adjacent section; in the remainder it was Schwann cell-like, identical to S-100 staining. Partial muscle denervation results, therefore, in the appearance of GAP-43 both in axons and Schwann cells in the endplates bordering the denervated zone. The presence of GAP-43 in these cells may contribute to their capacity to sprout.
Acetylcholinesterase↗