Neuropathic pain: neurochemical aspects

Abstracts

JUSTIFICATIVA E OBJETIVOS: A dor neuropática é causada por lesão ou inflamação do sistema nervoso. É síndrome complexa, com mecanismos biológicos pouco esclarecidos, envolvendo teorias inflamatórias e imunes. O objetivo desta revisão foi descrever os principais fatores biológicos relacionados com a dor neuropática, associando de forma lógica as hipóteses sugeridas pela literatura. CONTEÚDO: Foram descritos os principais neuromediadores, canais iônicos e células, incluindo as do sistema imune envolvidos na excitabilidade neuronal, assim como enfatizada possível seqüência de ativação ou interação desses agentes na alteração neuroplástica decorrente da agressão ao nervo. CONCLUSÕES: Do estudo, foi possível concluir que os avanços no conhecimento da fisiopatologia da dor neuropática podem determinar novos alvos para abordagem farmacológica dessa síndrome.

DOR; FISIOLOGIA

REVIEW ARTICLE

Neuropathic pain - neurochemical aspects^*

Dolor neuropático - aspectos neuroquímicos

Durval Campos Kraychete, TSA^I; Judymara Lauzi Gozzani, TSA^II; Angiolina Campos Kraychete^III

^IProfessor Adjunto de Anestesiologia - UFBA; Coordenador do Ambulatório de Dor - UFBA

^IIEditor-Chefe da Revista Brasileira de Anestesiologia; Coordenador do Serviço de Dor da Faculdade de Ciências Médicas da Santa Casa de São Paulo, SP

^IIIMédica Estagiária do Ambulatório de Dor - UFBA

Correspondence to

SUMMARY

BACKGROUND AND OBJECTIVES: Neuropathic pain is caused by damage or inflammation of the nervous system. It is a complex syndrome and its biological mechanisms, involving inflammatory and immunologic theories, are not clear. The objective of this review was to describe the main biologic factors associated with neuropathic pain, making a logical association between hypotheses suggested in the literature.

CONTENTS: The main neuromediators, ion channels, and cells, including cells in the nervous system involved in neuronal excitation are described, and the possible activation sequence or interaction among those agents in the neoplastic change secondary to nerve damage are emphasized.

CONCLUSIONS: It was possible to conclude that the advances on the knowledge of the pathophysiology of neuropathic pain can determine new pharmacologic approaches for this syndrome.

Key Words: PAIN: neuropathic; PHYSIOLOGY: neurotransmitters.

RESUMEN

JUSTIFICATIVA Y OBJETIVOS: El dolor Neuropático lo causa la lesión o inflamación del sistema nervioso. Es un síndrome complejo, con mecanismos biológicos poco aclarados, que envuelve teorías inflamatorias e inmunes. El objetivo de esta revisión fue describir los principales factores biológicos relacionados con el dolor Neuropático, asociando de forma lógica a las hipótesis sugeridas por la literatura.

CONTENIDO: Fueron descritos los principales neuromediadores, canales iónicos y células, incluyendo las del sistema inmune involucrados en la excitabilidad neuronal, como también la posible secuencia de activación o interacción de esos agentes en la alteración neuroplástica proveniente e la agresión al nervio.

CONCLUSIONES: De ese estudio, se pudo concluir que los avances en el conocimiento de la fisiopatología del dolor Neuropático, pueden determinar nuevos objetivos para el abordaje farmacológico de ese síndrome.

INTRODUCTION

Neuropathic pain is defined as pain secondary to nerve damage or dysfunction and, more widely, as a consequence of damage or disease of the somesthetic system ¹. It is a complex syndrome whose biological mechanisms are not clear, involving inflammatory and immunologic theories.

EXPERIMENTAL MODELS FOR THE STUDY OF NEUROPATHIC PAIN

Most experimental models described in the literature involve mice and were developed from traumatic, metabolic, or toxic peripheral lesions:

1. Spinal nerve ligature (SNL) - one or more spinal nerves innervating the paw are ligated and cut.

2. Partial sciatic ligature (PSL) - part of the sciatic nerve is ligated.

3. Chronic Constrictive Injury (CCI) - includes the placement of four ligatures on the sciatic nerve tightened by a chrome suture.

4. Spared nerve injury (SNI) - fibular and posterior tibial nerves are severed and the sural nerve is spared. This technique causes long-term behavioral changes.

Other methods include the intraperitoneal injection of streptozocin to mimic diabetic neuropathy, or paclitaxel and vincristine for chemotherapy induced neuropathy. Models of central pain use contusion (trauma using the force of impact of tissue dislocation), or ischemic lesions from slow compression by clamping or balloon insufflation. Cytotoxic methods use injections of glutamate analogues (cainate) or substances that cause lesion in specific areas of the gray matter. The techniques described are aimed at causing mechanical and thermal hyperalgesia ^2,3.

SECONDARY HYPERALGesIA AND CENTRAL SENSITIZATION

Secondary hyperalgesia is due to damage of neural and non-neural tissues adjacent to the primary lesion, and it is associated with central sensitization. Thus, patients with neuropathic pain can feel mechanical allodynia on the skin related to nerve transmission through Aβ fibers. This occurs because when the noxious stimulus from Aδ fibers reaches the dorsal horn of the spinal cord (lamina I), it can activate wide dynamic range neurons (lamina V) and increase the synaptic efficacy of Aβ fibers. Therefore, the loss of tactile function in patients with neuropathic pain can lead to an end of the allodynia ⁴.

Role of Primary Afferents

Since receptors that are expressed predominantly in small-diameter nerve fibers, such as cannabinoid and neutrotrophic receptors reverse thermal and mechanical hyperalgesia after SNL when they are stimulated, this reinforces the hypothesis that neuropathic pain is related to trauma of the primary afferent ^5,6.

Hypothesis of the Damaged Primary Afferent

In the lesion of the primary afferent, it has been widely documented that there is spontaneous and ectopic activity of the traumatic neuroma to thermal, chemical, and mechanical stimulus ⁷. Damage of fibers distal to the dorsal root ganglion causes local valerian degeneration associated with inflammatory phenomena and activation of macrophages, facilitating abnormal electric discharges from areas not affected by the lesion. An increase in the expression of TRVP1 receptors (transient receptor potential vanilloid-1), brain derived neurotrophic factors (BDNF), and excitatory neurotransmitters, such as the calcitonin gene-related peptide (CGRP) is seen. Since adjacent intact C fibers belong to the same damaged nerve or fasciculus, they develop an increased sensitivity to catecholamines and in the expression of Nav 1.8 sodium channel receptors. This could explain why L₄ rhizotomy can alleviate the mechanical hyperalgesia secondary to the L₅ lesion in mice ^2,3. The injection of a local anesthetic directly in the L₅ dorsal root ganglion in the model of SNL in mice reversed the changes secondary to neuropathic pain ⁸. This occurred despite the fact that the L₅ lesion causes spontaneous neuronal activity preferentially in Aδ fibers, which suggests that the spontaneous activity of the Aδ fiber initiates the central sensitization and changes the phenotypic expression of the primary afferent (C fibers). A new expression of neuropeptides normally related with C fibers can occur, as well as an increase in the spontaneous activity of Aβ fibers ^2,3.

Hypothesis of the Intact Primary Afferent

After damage of a peripheral nerve, spontaneous neural activity develops in primary afferents that share the same innervation with the severed nerves. The incidence of spontaneous neuronal activity is high, around 50%. However, the damage of a spinal nerve in rodents leads in intact nerves to an increased sensitivity to adrenaline and tumor necrosis factor (TNFα) ⁹. There is an increase in the response to heat and cold secondary to the increased expression of proteins for transitory receptor potential vanilloid (TRVP1) and cold-sensitive channels (TRPA1) in the dorsal horn of the spinal cord ¹⁰. There is also an increased expression of messenger ribonucleic acid (mRNA) for the calcitonin gene-related peptide (CGRP), brain-derived neurotrophic factor (BDNF), and purinergic receptors (P2X3) ^2,3.

Pain Mediated by the Sympathetic Nervous System

The influence of the sympathetic nervous system on neuropathic pain is very relevant. Complex regional syndrome usually manifests itself with severe pain in one extremity. Patients present with edema, hyperalgesia, or changes in motor function, which might improve with a sympathetic block or with α-adrenergic receptors antagonists, indicating a sympathetic component. In animal models of neuropathic pain secondary to SNL at the level of L₆, more than 60% of the intact nociceptors show spontaneous activity and more than 50% respond to α-adrenergic agonists ^2,3.

Sodium Channels

Nav 1.3, 1.7, 1.8, and 1.9 sodium channels can be found in the dorsal root ganglion and are involved in the generation of action potentials and conduction of nociceptors. Nav 1.3 and 1.7 are sensitive to tetrodotoxin and the other two are not. There is an increase in the expression of Nav 1.3 in the dorsal root ganglion of damaged axons. This channel has kinetic properties that facilitate repetitive discharges. The β2 subunit regulates the opening of the channel and mice that do not express this subunit do not develop mechanical hyperalgesia after nerve damage. Therefore, the high density of voltage-dependent sodium channels in the damaged nerve can cause neuronal depolarization to last longer than the refractory period of adjacent sodium channels, allowing the antidromic propagation of the action potential (from proximal to distal) in the nerves. The frequency of fast pulses in large-caliber fibers can lead to central sensitization because they can stimulate adjacent nociceptive fibers. Mutations in Nav 1.7 sodium channels can reduce the excitability of the sympathetic nervous system and cause hyperexcitability in small-caliber fibers. This would explain the edema, erythema, and pain of erythromelalgia ¹¹.

Role of Central Sensitization

Central sensitization involves homo- and heterosynaptic mechanisms. In homosynaptic sensitization, the test stimulus and conditioning are associated with the same afferent. This can be exemplified by the wind up phenomenon, when the continuous low-frequency stimulation of afferent C fibers causes an increase in the response of specific cells in the dorsal horn of the spinal cord. In heterosynaptic sensitization, the test stimulus and conditioning are related in a different way. In this case, nociceptive stimuli increase the synaptic efficacy of mechanoreceptors connected with Aβ fibers. Thus, neuropathic pain is accompanied by homo- and heterosynaptic sensitization, which was demonstrated in models of L₅ lesion using the SNL technique. L₅ myelinated fibers (Aβ and Aδ) develop spontaneous activity. The impulse from those fibers can lead to homosynaptic sensitization in the spinothalamic tract, explaining the persistent chronic pain. On the other hand, projection to the adjacent segment at the L₄ level can lead to heterosynaptic sensitization. Therefore, when applying mechanical stimulus to Aβ fibers in the skin corresponding to the L₄ path, it can lead to sensitization of afferent fibers in the cells related with L₄ in the dorsal horn of the spinal cord. Thus intact L₄ afferents show mechanical and thermal hyperalgesia ^2,3.

Central sensitization occurs by an increase in synaptic efficacy or due to excessive release of excitatory neurotransmitters.

Pre-Synaptic Changes

Release of glutamate is inhibited by the activation of GABAergic (type B), adenosine, and opioid (µ) ¹² receptors. The reduction or failure of the function of those receptors can lead to neuronal hyperexcitability in the damaged nerve. Calcium channels a2d subunits in the dorsal root ganglion and in the spinal cord can also be increased ¹³, which cause the release of excitatory neurotransmitters. The phenotypic change of Ad fibers in neuropathic pain would similarly cause the pre-synaptic release of substance P facilitating sensitization of the dorsal horn of the spinal cord.

Post-Synaptic Changes

The release of substance P and CGRP, besides other excitatory neurotransmitters (aspartate and glutamate) in the synaptic cleft activates NMDA (N-methyl-D-aspartate) and AMPA (amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptors leading to an increase of calcium influx into the cell, formation of the calcium-calmodulin complex, activation of the enzyme calcium-calmodulin kinase II, and the neuronal pathways of nitric oxide synthase and production of nitric oxide. This promotes a specific action of protein kinases (A and C), mitogen-activated protein kinases (ERK ½, JNK p38, MAPK), and the transcriptional factors CREB, ATF-2 (activating transcription factor-2) that when phosphorylated bind to c-fos and c-jun (immediate-early genes) promoters, resulting in the synthesis of nuclear gene products that undergo dimerization to form the AP-1 complex and several other genes that facilitate neuronal excitability and changes in the neuroplasticity of the dorsal horn ¹⁴. It is possible that the damaged nerve shows increased expression of mRNA for AMPA and NMDA receptors in the dorsal horn of the spinal cord. Gene transcription seems to be connected more to the activation of NMDA and neurokinin (NK1) receptors, besides the pre- and post-synaptic action of nitric oxide ¹⁴.

On the other hand, a reduction in the expression of inhibitory receptors after nerve damage and neuronal sensitization would probably be secondary to facilitating mechanisms. Long-term depression is opposed to the long-term potentiation that occurs in GABA type inhibitor cells in the dorsal horn of the spinal cord and would lead to neuronal sensitization. The expression of potassium and chloride channels can be increased and if opened they could inhibit GABAergic neurons causing consequently nerve excitation. In experimental models of neuropathic pain GABAergic neurons can also undergo apoptosis or glycine-containing neurons can be lost resulting in facilitation of the nerve conduction ^2,3.

CELLS AND NEUROPATHIC PAIN

Mast Cells

Mast cells are present in peripheral nerves and can be activated after the increase of adenosine or bradykinin in the site of damage ¹⁵. This causes the release of granules containing histamine, proteases (tryptases and PAR-2 receptors), cytokines, and neurotrophic factors (NGF) capable of: direct excitation of nociceptors and dorsal root ganglion cells; facilitate the action of SP and CGRP; and provoke spontaneous burning pain ¹⁶. Synthesis, transcription, translation, and secretion of prostaglandins, cytokines, and chemokines ¹⁷ at the site of nerve damage besides recruitment of leukocytes including neutrophils and macrophages that affect directly the neurovegetative nervous system can also occur ¹⁸.

Neutrophils

Neutrophils adhere to the vascular endothelium and migrate to the site of inflammation in the damaged nerve releasing lipoxigenases and cytokines that affect pain receptors directly. Thus, depletion of circulating neutrophils before experimental nerve lesion attenuates the resulting hyperalgesia ¹⁹. Neutrophils release chemokines and defensins that are chemotactic for macrophages and lymphocytes. On the other hand, macrophages that reside in the central and peripheral nervous systems phagocyte degenerated or dead neurons and Schwann cells ²⁰. Macrophages release prostaglandins, cytokines (interleukin-6 IL-6, TNFα, and interleukin 1β IL-1β), and superoxide radicals -implicated on neuropathic pain, and the depletion of macrophages on mice with damaged nerves reduces hyperalgesia ²¹.

Lymphocytes

Lymphocytes responsible for cell-mediated immunity (T cells) can be found on the damaged site in models of neuropathic pain, expressing themselves in the periphery and in the central nervous system. Thus, mice that lack T cells develop less mechanical allodynia and thermal hyperalgesia when subjected to damage of the sciatic nerve ²².

T cells can be divided in CD4⁺ (helper) and CD8⁺ (cytotoxic), which provoke specific TH1 and TH2 reactions, respectively, according to the cytokines secreted. The TH1 response releases interferon-gamma and IL-2, and it is involved in cell-mediated inflammatory responses, while the TH2 response (IL-4, IL-5, IL-6, IL-9, and IL-10) is involved in the allergic response with the production of antibodies and inhibition of the synthesis of pro-inflammatory cytokines ²³. Those responses could possibly have opposing effects in the evolution of neuropathic pain, since the transference of CD4⁺ lymphocytes increases the response to painful stimuli in mice, while the transference of CD8⁺ cells reduces hyperalgesia ²².

Glial Cells

Glial and Schwann cells interact with neurons, promoting maintenance of the homeostasis, regulating the concentration of neurotransmitters and ions, and the extracellular pH. In neuropathic pain, microglial cells seem to play a fundamental role in the beginning of the lesion and astrocytes in its maintenance ²⁴. Microglia are activated by several neuromediators, such as ATP, bradykinin, substance P, fractalkine, and Toll-like receptor 4 (TLR₄) ^25,26. ATP activates P2X₄ receptors in the central nervous system; fractalkine is a chemokine expressed on the surface of spinal neurons that activates the microglial CX3CR1 receptor, and TLR₄ receptors recognize molecules of different structures released during the nerve damage. However, it is not known how neuropathic pain develops after microglial activation. It is possible that glial cells release several excitatory neurotransmitters, such as prostaglandins, nitric oxide, cytokines, and chemokines, activating the sensitive afferent directly. Activation can also occur in the contralateral side of the body by propagation of calcium waves between neural junctions, facilitating the release of excitatory neuromediators ²⁷.

On the other hand, Schwann cells interact with T cells expressing MHC class II molecules ²⁸. Schwann cells secrete cytokines (IL-6, IL-1, TNFα), neurotrophic factors (NGF), prostaglandin E2, and ATP ²⁹. They also express ion channels, and glutamate and cytokine receptors ³⁰. Thus, it is possible that they contribute to the genesis of neuropathic pain.

NEUROMEDIATORS

Bradykinin

Bradykinin and kalidine are formed in the blood and tissues. Bradykinin exerts its actions on B1 and B2 receptors, causes sensitization of peripheral nociceptors (desinhibiting vanilloid receptors - TRVP1), potentiates glutaminergic synaptic transmission in the spinal cord ³¹, stimulates the release of cytokines by macrophages, secretes neutrophil and monocyte chemotactic factors, and facilitates the release of histamine by mast cells. In mice, damage of the sciatic nerve increases the expression of B2 and B1 receptors in the dorsal root ganglion, with predominance of B1 receptors in the soma of myelinated axons. This suggests that B2 receptors are involved in central sensitization ³¹.

ATP and Adenosine

ATP is a classical neurotransmitter, but it is also released by damaged non-neuronal cells and tissues. It exerts its actions in purinergic receptors (P1 or P2). P2 receptors can be divided in P2X and P2Y, which are coupled to protein G and ion channels, respectively ³².

In experimental models of neuropathic pain P2X₃ receptors are reduced (after axotomy or partial nerve ligature) or increased (chronic constrictive lesion); however, even in the face of reduction those receptors show increased sensitivity. Blockade of P2X₃ receptors attenuates thermal and mechanical allodynia in mice ³³. The expression of P2X₄ receptors is also increased in microglia after nerve damage, and its pharmacological blockade reverses allodynia ³⁴. P2X₇ receptors can be found in T cells and macrophages. Mice that do not express this receptor do not develop neuropathic pain ³⁵. On the other side, P2Y₁ receptors show a 70% increase after damage of the sciatic nerve in mice ³⁶.

Serotonin

Serotonin is a neurotransmitter synthesized and released by central nervous system (dorsal root ganglion, cytoplasm, and Schwann cells) neurons. In the periphery, serotonin is released by platelets and induces hyperalgesia by direct action in the primary afferent via the 5HT_1A receptors ³⁷. In partial nerve lesions in mice, serotonin contributes with mechanical hyperalgesia through 5HT_2A and 5HT₃ receptors ³⁸.

Eicosanoids

Arachidonic acid metabolites include prostaglandins, thromboxanes, and leukotrienes. Prostaglandins PGE₂ and PGI₂ exert their action on protein G-coupled receptors (EP14 and IP, respectively) and induce hyperalgesia in peripheral ³⁹ and central nervous system ⁴⁰ nociceptors. The expression of cyclooxygenase (COX-2) in the damaged area, in the spinal cord (COX2 and COX1), and in the thalamus is increased in models of neuropathic pain, associated with the increase in the number of macrophages and in the production of PGE₂ by mast cells ^41,42. This phenomenon is related with the actions of IL1β, TNFα, nerve growth factor (NGF), MCP-1 (monocyte chemoattractant protein-1), and reactive oxygen species (ROS). Thus, PGE₂ can depolarize directly cells with wide dynamic range in the dorsal horn of the spinal cord, activate tetrodotoxin-resistant sodium channels and voltage-dependent calcium channels, inhibit potassium channels, and increase the release of glutamate, substance P, or CGRP, or block glycine inhibition on specific nociceptive neurons. Treatment of cultures of dorsal horn neurons with PGE₂ increases the expression of substance P NK1 receptors, and in models of nerve lesions it induces greater expression of basic fibroblast growth factor (bFGF), nerve growth factor, nitric oxide synthase (NOS), Nav 1.7 and 1.8 sodium channels, TRVP1 receptors (transitory receptor potential vanilloid-1), and metalloproteinases (MMPs) involved in axonal degeneration, loss of myelin sheath, recruitment of leukocytes and macrophages to the site of damage, and disruption of the hematoencephalic barrier. Those factors contribute for the maintenance of persistent chronic pain⁴³. Despite the reversal of thermal and mechanical hyperalgesia after the subcutaneous injection of anti-inflammatories ³⁸ in laboratory animals, in humans anti-inflammatories are ineffective in clinical practice for the treatment of neuropathic pain.

On the other side, leukotriene B4 (LTB4) produces hyperalgesia by releasing neuromediators from neutrophils ⁴⁴. Besides, neurotrophic factors (NGF) also produce hyperalgesia by inducing the release of LTB₄ from mast cells, and increase the recruitment of neutrophils ⁴⁵.

Cytokines

Cytokines are small molecules that mediate the interactions among cells over distances. IL1β, IL-6, and TNFα are pro-inflammatory, induce the sequential production of each one, and are synergistic ⁴⁶. The exogenous administration of those substances can also induce pain and hyperalgesia ⁴⁷. Binding of IL1β to the IL1-RI receptor triggers a series of intracellular events activating transcriptional factors, inducing the expression of COX-2, nitric oxide synthase, IL1β, IL-6, and TNFα. Thus, IL1β affects nociceptors direct and indirectly ⁴⁸, in nerve damage its production is increased in peripheral nerves ⁴⁹, and in mice the administration of antibodies for the IL-1 receptor alleviates neuropathic pain.

Interleukin-6 is synthesized by mast cells, monocytes, lymphocytes, neurons, and glia. Lesions of the sciatic nerve promote the local increase in IL-6 expression, as well as in the dorsal horn of the spinal cord ⁵⁰. In mice, the injection of IL-6 in the lateral ventricles produces thermal hyperalgesia ⁵¹. Despite controversial studies, in mice that do not produce IL-6 the development of mechanical allodynia after peripheral nerve lesion is delayed ⁵².

After acting on specific receptors, TNFα activates transcriptional factors (p36 MAPK and NFkβ) and releases COX-2-dependent factors. The intraplantar injection of TNFα in rodents induces mechanical hyperalgesia ⁵³ and after topical and intraneural administration it induces thermal hyperalgesia and mechanical allodynia ⁵⁴. Lesion of the primary afferent leads to an increase in TNFα in the dorsal horn of the spinal cord, locus ceruleus, and hippocampus ⁵⁵.

Neurotrophins

Neurotrophins are dimeric proteins synthesized and released by several immune cells (mast cells and lymphocytes) and they are essential for the normal development of the nervous system in vertebrates ⁵⁶. This family includes nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neutrophin 3 and neutrophin 4/5. Glial cell-line derived neurotrophic factor (GDNF) is another protein with non-dimeric neurotrophic properties.

Neurotrophins act on specific receptors - tyrosine kinases (TrK) A, for NGF, B, for BDNF, and NT-4/5 and C, for NT-3, and their expression is greater in the embryo ⁵⁷.

Nerve damage induces changes in the expression of growth factors. This change affects the tissue without innervation, Schwann cells, dorsal horn of the spinal cord, and dorsal root ganglion. However, after nerve damage the synthesis of those substances as well as of their receptors increases ⁵⁸. In the experimental L₅ SNL damage neurotrophic factor (GNF) is increased in the L₄-innervated territory with the retrograde transport of neurotrophins to the dorsal root ganglion modifying the expression of brain derived neurotrophic factors (BDNF). Therefore, an influence of neurotrophic factors in damaged and intact neurons is seen. Neurotrophic factors (NGF) increase the expression of channels related with transduction of the mechanical stimulus (TRPV4). Thus, cold hyperalgesia can be secondary to the abnormal expression of TRPA1 and TRPM8 channels. Nerve growth factor besides causing direct sensitization of peripheral nociceptors affects immune and sympathetic cells ⁵⁹, while BDNF facilitates the excitation of dorsal horn neurons ⁶⁰. In experimental models, expression of those substances increases in the area related with the damaged nerve secondary to constriction as well as the corresponding dorsal root ganglion ⁶¹. The endoneural injection of neurotrpphic factor causes budding of synapses and signs of thermic hyperalgesia, while the perineural administration of this substance determines the forthcoming of mechanical hyperalgesia.

Growth factors play a fundamental role in the development and maintenance of small neurons and the sympathetic nervous system. After nerve damage, α-adrenergic receptors are increased in the soma in intact axons, with the consequent sensitivity of those nerves to circulating adrenaline. In animal models, the blockade of damaged fibers by local anesthetics reduces significantly their spontaneous activity and nerve budding in the sympathetic nervous system up to five weeks after the lesion, with the blockade of NGF production by inhibiting tyrosine kinase.

Nitric Oxide and Superoxide Radicals

Superoxide radicals are released by mast cells and glia (astrocytes and microglia). On the other hand, nitric oxide has constitutive endothelial and neuronal forms, while the induced form is expressed in cells of the immunologic system. Nitric oxide causes hyperalgesia after injection in the skin and joints ¹⁶² and it is implicated in central sensitization ⁴⁸, increasing the effects of PG₂ in neuropathic pain models ⁶³.

Lysophosphatidic acid

Lysophosphatidic acid (LPA) is derived from the lipid metabolism, it is released after tissue damage and might be involved in neuropathic pain. Platelets are the greatest producers of LPA and the plasma is its major source. Lysophosphatidic acid receptors (LPA_{1 to 4}) are distributed in the central and peripheral nervous system. They are also coupled to protein Ga_12/13 and when activated, they activate GTPase and RHoA and a series of second messengers, including Rho kinase or ROCK. The intrathecal injection of LPA in animals causes mechanical allodynia and thermal hyperalgesia. There is a drastic reduction of myelin-associated proteins (myelin basic and peripheral protein, MPB and PMP 22, respectively) and an increase in the expression of phosphokinase C and calcium channels (Ca_vα₂δ1) with nerve degeneration, synaptic budding, increased neuronal excitability ⁶⁴.

NEUROPATHIC PAIN AND NERVE DEGENERATION

Calcium influx is one of the first events after nerve damage. Calcium initiates tissue cicatrization grouping vesicles in the axolema around damaged distal and proximal nerve endings. The increase in intracellular calcium also activates calcium-dependent protein kinases and other proteases (UPS) that facilitate the release of cytokines and destruction of microtubules and microfilaments, which are important in the axonal transport of substances and in the mechanism of neuropathic pain. This leads to an accumulation of SP, CGRP, nitric oxide, neurotrophic factors, and sodium channels that, together, amplify neuronal excitability ⁶⁵.

Although peripheral lesions of C fibers lead to a reduction of SP, neurotrophins (BDNF), receptors (VR1 and P2X₃), and type N high-voltage calcium channels in the dorsal horn of the spinal cord, there is ascending regulation of type III TTX-sensitive (TTXs) channels and translocation of TTX-resistant (TTXr) sodium channels from the soma to the neuroma, increasing nerve excitability ⁶⁶. The expression of Nav 1.3 TTXs channels, whose kinetic characteristic is to be easily activated and inactivated, can increase by two to 30-fold in models of dorsal root ganglion axotomy, ligature of the spinal nerve, chronic constrictive damage, diffuse nerve damage, diabetic neuropathy, and post-herpetic neuropathy ¹¹.

When the nerve fiber is intact (inflammation), excitatory neuromediators increase in the dorsal horn of the spinal cord, as well as the expression of sodium channels (TTXr), which facilitates neuronal hyperexcitability and hinders the response to treatment with local anesthetics ⁶⁶.

In both situations described, especially in lesions of C fibers, there can also be an increase in SP and BDNF in Aβ fibers (low threshold mechanoreceptors), as well as budding of those fibers at the site of afferent connections of C fibers (lamina II), widening the receptive field of the neuron and leading to the interpretation of innocuous peripheral mechanical stimuli as aggressive ⁶⁷. This explains, for example, the mechanical allodynia that is seen in post-herpetic neuralgia ⁶⁸. Sprouting of the sympathetic noradrenergic dorsal root ganglion axon around large-caliber neurons (Aδ fibers) would be another possibility, suggesting the hypothesis of activation of sensitive afferent fibers after sympathetic stimulation ⁶⁹. Besides, some disproportion between excitatory and pain suppression pathways can be present, with reduction in the inhibitory activity of glycine, GABA, and opioids ⁶⁶.

CENTRAL ASPECTS

In models of neuropathic pain in animals, the inhibitory effect of descending fibers originating in the periaqueductal gray matter and locus ceruleus can show a 50% reduction from the baseline. There is reduced efficacy of the opioid system in the spinal cord, with reduction of β-endorphins in the brain and spinal cord, with the consequent reduction of the analgesic effects of morphine, both by systemic and spinal administration. This suggests that opioid receptors or other factors that are necessary for the activation of the spinal opioid system are compromised after nerve damaged ^2,3. On the other hand, programmed cell death can occur in dorsal horn neurons that express c-jun. This involves studies of the Bcl-2 and Bax gene families that inhibit and favor cell death, respectively. In mice with sectioned nerve and in which the expression of c-jun is increased, the Bcl-2/Bax relationship is reduced, indicating a tendency for apoptosis and deafferentation of post-synaptic spinal neurons ¹⁴.

The destruction of the posterior cord or the gracilis nucleus ipsilateral to the lesion can abolish mechanical allodynia in mice. On the other hand, allodynia can also be inhibited by the selective destruction of descending pathways of the brainstem and dorsolateral cord, indicating a role of ascending and descending pathways in changes secondary to neuropathic pain. Thermal allodynia can be abolished by the injection of lidocaine or cholecystokinin (CCK) receptor antagonists in the ventromedial area of the medulla (raphe magnus nucleus). Descending facilitation involving the median raphe nucleus in the medulla (NRM) and CCK can occur in nerve damage. Therefore, severing descendent fibers can reduce considerably mechanical or thermal allodynia in animals. Medulla-spinal impulses are also capable to activate post-synaptic sympathetic neurons that contribute for the maintenance of neuropathic pain ^2,3,14.

As for the neurophysiological aspects of neuropathic pain in SNL, the spontaneous neuronal activity of somatosensorial regions of the thalamus (ventral posteromedial nucleus, VPM, and ventral posterior lateral nucleus, VPL) is increased and it can be inhibited by substances that block the expression of Nav 1.3 sodium channels. On the other hand, functional imaging of the thalamus contralateral to the neuropathic pain shows reduced metabolism. In the cerebral cortex, the behavior of 10% of the neurons is similar to the thalamic neurons, especially those located in the periphery of the deafferented zone of the primary somesthetic cortex. The electroencephalogram shows an increase in theta frequency (4 to 5 Hz) and the metabolism or the cortical output of the insular, posterior parietal, pre-frontal, and cingulate areas is elevated. The opposite is seen in the medial (Brodmann area 24), and anterior and perigenual (Brodmann areas 32 and 35) cingulate cortices. Some researchers demonstrated that the increased perigenual output after motor cortical stimulation of the Gasser ganglion reduces pain severity. This strengthens the hypothesis of the relationship between perigenual changes and failure of the descending inhibitory control. A reduction of opioid receptors in the periaqueductal gray matter, in medial thalamus, pre-frontal cortex, and ínsula due to internalization is also seen. This reduction is bilateral and symmetrical and preponderant in the ipsilateral side in the case of a central lesion ².

CONCLUSION

Neuropathic pain is still a challenge for clinical and experimental researchers. Its mechanisms, which are complex and not completely understood result occasionally in dynamics with contradictory results. Understanding the neurobiology of neuropathic pain is a step towards the improvement in the treatment of this syndrome. This understanding may result in the development of drugs aimed at specific targets with effective responses.

REFERENCES

• 01. Dworkin B and the Members of the Classification Subcommittee of the Special Interest Group of the International Association for the Study of Pain. Disponível em http://www.neupsig.org/
• 02. Garcia-Larrea L, Magnin M - Physiopathologie de la douleur neuropathique: revue des modèles expérimentaux et des mécanismes proposés. Presse Med, 2008;37:315-340.
• 03. Campbell JN, Meyer RA - Mechanisms of neuropathic pain. Neuron, 2006;52:77-92.
• 04. Meyer RA, Ringkamp M, Campbell JN et al. - Peripheral Mechanisms of Cutaneous Nociception, em: McMahon SB, Koltzenburg M - Wall and Melzack's - Textbook of Pain. London, Elsevier, 2006;3-34.
• 05. Ibrahim MM, Deng H, Zvonok A et al. - Activation of CB2 cannabinoid receptors by AM1241 inhibits experimental neuropathic pain: pain inhibition by receptors not present in the CNS. Proc Natl Acad Sci USA, 2003;100:10529-10533.
• 06. Gardell LR, Wang R, Ehrenfels C et al. - Multiple actions of systemic artemin in experimental neuropathy. Nat Med, 2003;9: 1383-1389.
• 07. Devor M - Response of Nerves to Injury in Relation to Neuropathic Pain, em: McMahon SB, Koltzenburg M - Wall and Melzack's - Textbook of Pain. London, Elsevier, 2006;905-927.
• 08. Sukhotinsky I, Ben Dor E, Raber P et al. - Key role of the dorsal root ganglion in neuropathic tactile hypersensibility. Eur J Pain, 2004;8:135-143.
• 09. Schäfers M, Lee DH, Brors D et al. - Increased sensitivity of injured and adjacent uninjured rat primary sensory neurons to exogenous tumor necrosis factor-alpha after spinal nerve ligation. J Neurosci, 2003;23:3028-3038.
• 10. Katsura H, Obata K, Mizushima T et al. - Antisense knock down of TRPA1, but not TRPM8, alleviates cold hyperalgesia after spinal nerve ligation in rats. Exp Neurol, 2006;200:112-123.
• 11. Rogers M, Tang L, Madge DJ et al. - The role of sodium channels in neuropathic pain. Semin Cell Dev Biol, 2006;17:571-81.
• 12. KohnoT, Ji RR, Ito N et al. - Peripheral axonal injury results in reduced mu opioid receptor pre- and post-synaptic action in the spinal cord. Pain, 2005;117:77-87.
• 13. Li CY, Song YH, Higuera ES et al. - Spinal dorsal horn calcium channel alpha2delta-1 subunit upregulation contributes to peripheral nerve injury-induced tactile allodynia. J Neurosci, 2004; 24:8494-8499.
• 14. Zimmermann M - Pathobiology of neuropathic pain. Eur J Pharmacol, 2001;429:23-37.
• 15. McLean PG, Ahluwalia A, Perretti M - Association between kinin B (1) receptor expression and leukocyte trafficking across mouse mesenteric postcapillary venules. J Exp Med, 2000;192: 367- 380.
• 16. Baron R, Schwarz K, Kleinert A et al. - Histamine-induced itch converts into pain in neuropathic hiperalgesia. Neuroreport, 2001;12:3475-78.
• 17. Mekori YA, Metcalfe DD - Mast cells in innate immunity. Immunol Rev, 2000;173:131-140.
• 18. Woolf CJ - Dissecting out mechanisms responsible for peripheral neuropathic pain: implications for diagnosis and therapy. J Life Sci. 2004;74:2605- 2610.
• 19. Zuo Y, Perkins NM, Tracey DJ et al. - Inflammation and hyperalgesia induced by nerve injury in the rat: a key role of mast cells. Pain, 2003;105:467-479.
• 20. Brück W - The role of macrophages in Wallerian degeneration. Brain Pathol, 1997;7:741-752.
• 21. Liu T, Van Rooijen N, Tracey DJ - Depletion of macrophages reduces axonal degeneration and hyperalgesia following nerve injury. Pain, 2000;86:25-32.
• 22. Moalem G, Xu K, Yu L T - lymphocytes play a role in neuropathic pain following peripheral nerve injury in rats. Neuroscience, 2004;134:1399 -1411.
• 23. Mosmann TR, Sad S - The expanding universe of T-cell subsets: Th1, Th2 and more. Immunol Today, 1996;17:138-146.
• 24. Raghavendra V, Tanga F, DeLeo JA - Inhibition of microglial activation attenuates the development but not existing hipersensitivity in a rat model neuropathy. J Pharmacol Exp Ther, 2003;306:624-630.
• 25. Tanga FY, Nutile-McMenemy N, DeLeo JA - The CNS role of Toll-like receptor 4 in innate neuroimmunity and painful neuropathy. Proc Natl Acad Sci USA, 2005;102:5856-5861.
• 26. Watkins LR, Milligan ED, Maier SF - Glial activation: a driving force for pathological pain. Trends Neurosci, 2001;24:450-455.
• 27. Spataro LE, Sloane EM, Milligan ED et al. - Spinal gap junctions potential involvement in pain facilitation. Clin J Pain, 2004;5:392-405.
• 28. Bergsteinsdottir K, Kingston A, Jessen KR - Rat Schwann cells can be induced to express major histocompatibility complex class II molecules in vivo. J Neurocytol, 1992;21:382-390.
• 29. Liu GJ, Werry EL, Bennett MR - Secretion of ATP from Schwann cells in response to uridine triphosphate. Eur J Neurosci, 2005; 21:151-160.
• 30. Skundric DS, Bealmear B, Lisak RP - Induced upregulation of IL-1, IL-1RA and IL-1R type I gene expression by Schwann cells. J Neuroimmunol, 1997;74:9-18.
• 31. Wang H, Kohno T, Amaya F et al. - Bradykinin produces pain hupersensitivity by potentiating spinal cord glutamatergic synaptic transmission. J Neurosci, 2005;25:7986 -7992.
• 32. Burnstock G, Knight GE - Cellular distribution and functions of P2 receptor subtypes in different systems. Int Rev Cytol, 2004; 240:281-304.
• 33. Javis MF, Bugard EC, McGaraughty S et al. - A-317491, a novel potent and selective non-nucleotide P2X3 and P2X2/3 receptors, reduces chronic inflammatory and neuropathic pain in rats. Proc Natl Acad Sci USA, 2002;99:17179-17184.
• 34. Tsuda M, Shigemoto-Mogami Y, Koizumi S et al. - P2X4 receptors induced in spinal microglia gate tactile alodynia after nerve injury. Nature, 2003;424:778-783.
• 35. Chessell IP, Hatcher JP, Bountra C et al. - Disruption of the P2X7 purinoreceptor gene4 abolishes chronic inflammatoty and neuropathic pain. Pain, 2005;114:386-96.
• 36. Xiao HS, Huang QH, Zhang FX et al. - Identification of gene expression profile of dorsal root ganglion in the rat peripheral axotomy model of neuropathic pain. Proc Natl Acad Sci USA, 2002;99:8360-8365.
• 37. Hong Y, Abbott FV - Behavioural effects of intraplantar injection of inflammatory mediators in the rat. Neuroscience, 1994;63: 827-836.
• 38. Moalem G, Grafe P, Tracey DJ - Chemical mediators enhance the excitability of unmyelinated sensory axons in normal and injured peripheral nerve of the rat. Neuroscience, 2005;134:1399-1411.
• 39. Taiwo YO, Levine JD - Prostaglandin effects after elimination of indirect hyperalgic mechanisms in the skin of the rat. Brain Res, 1989;492:397-399.
• 40. Malberg AB, Yaksh TL - Hyperalgesia mediated by spinal glutamate or substance P receptor blocked by spinal cyclooxygenase inhibition. Science, 1992;257:1276-1279.
• 41. Ma W, Einsenach JC - Morphological and pharmacological evidence of the role of peripheral prostaglandins in the pathogenesis of neuropathic pain. Eur J Neurosci, 2002;15:1037-1047.
• 42. Zhao Z, Chen SR, Eisenach JC et al. - Spinal cyclooxygenase-2 is involved in development of allodynia after nerve injury in rats. Neuroscience, 2000;97:743-748.
• 43. Ma W, Quirion R - Does COX2-dependent PGE2 play a role in neuropathic pain? Neurosci Lett, 2008;437:165-169.
• 44. Levine JD, Lau W, Kwiat G et al. - Leukotriene B4 produces hyperalgesia that is dependent on polymorphonuclear leukocytes. Science, 1984;225:743-745.
• 45. Bennett G, al-Rashed S, Hoult JR et al. - Nerve growth factor induced hyperalgesia in the rat hind paw is dependent on circulating neutrophils. Pain, 1998;77:315-322.
• 46. Watikins LR, Hansen MK, Nguyen KT et al. - Dynamic regulation of the pro-inflammatory cytokines, interleukin-1beta: molecular biology for non molecular biologists. Life Sci, 1999;65:449-481.
• 47. Sommer C, Kress M - Recent findings on how proinflammatory cytokines cause pain: Peripheral mechanisms in inflammatory and neuropathic hyperalgesia. Neurosci Lett, 2004;361:184-187.
• 48. Sung CS, Wen ZH, Chang WK et al. - Intrathecal Interleukin-1beta administration induces thermal hiperalgesia by activating inducible nitric oxide synthase expression in the rat spinal cord. Brain Res, 2004;1015:145-153.
• 49. Gillen C, Jander S, Stroll G - Sequential expression of mRNA for proinflammatory cytokines and interleukin-10 in the rat peripheral nervous system: comparison between immune-mediated demylination and Wallerian degeneration. J Neurosci, 1998;51: 489-496.
• 50. DeLeo JA, Yezierski RP - The role of neuroinflammation and neuroimmune activation in persistent pain. Pain, 2001;90:1-6.
• 51. Oka T, Oka K, Hosoi M et al. - Intracerebroventricular injection of interleukin-6 induces thermal hyperalgesia in rats. Brain Res, 1995;692:123-128.
• 52. Murphy PG, Ramer MS, Borthwick L et al. - Endogenous interleukin-6 contributes to hipersensitivity to cutaneous stimuli and changes in europeptides associated with chronic nerve constriction in mice. Eur J Neurosci, 1999;11:2243-2253.
• 53. Cunha FQ, Poole S , Lorenzetti BB et al. - The pivotal role of tumor necrosis factor alpha in the development of inflammatory hyperalgesia. Br J Pharmacol, 1992;107:660-664.
• 54. Zelenka M, Shäfers M, Sommer C - Intraneural injection of interleukin-1beta and tumor necrois factor-alpha into rat sciatic nerve at physiological doses induces signs of neuropathic pain. Pain, 2005;116:257-263.
• 55. Ignatowski TA, Covey WC, Knight PR et al. - Brain-derived TNFalpha mediates neuropathic pain. Brain Res, 1999;841:70-77.
• 56. Moalem G, Gdalyahu A, Shani Y et al. - Production of neurotrophins by activated T cells: implications for neuroprotective autoimmunity. J Autoimmun, 2000;15:331-345.
• 57. Nykjaer A, Willnow TE, Petersen CM - P75(NTR).... Live or let die. Curr Opin Neurobiol, 2005;15:49-57.
• 58. Funakoshi H, Frisen J, Barbany G et al. - Differential expression of mRNAs for neurotrophins and their receptors after axotomy of the sciatic nerve. J Cell Biol, 1993;123:455-465.
• 59. Woolf CJ, Safieh-Garabedian B, Ma QP et al. - Nerve growth factor contributes to the generation of inflammatory sensory hypersensitivity. Neuroscience, 1994;62:327-331.
• 60. Pezet S, Malcangio M, McMahon SB - BDNF: a neuromodulator in nociceptive pathways? Brain Res Rev, 2002;40:240-249.
• 61. Miletic G, Miletic V. - Increases in the concentration of brain derived neurotrophic factor in the lumbar spinal dorsal horn are associated with pain behavior following chronic constriction injury in rats. Neurosci Lett, 2002;319:137-140.
• 62. Aley KO, McCarter G, Levine JD - Nitric oxide signaling in pain and nociceptor sensitization in the rat. J Neurosci, 1998;18: 7008-7014.
• 63. Levy D, Zochodne DW - No pain: potential roles of nitric oxide in neuropathic pain. Pain Pract, 2004;4:11-18.
• 64. Ueda H- Molecular mechanisms of neuropathic pain phenotypic switch and initiation mechanisms. Clin Pharmacol Ther, 2006;109:57-77.
• 65. Üçeler N, Sommer C - Wallerian degeneration and neuropathic pain. Drug Discov Today Dis Mech, 2006:3:351-356.
• 66. McMahon SB - Neuropathic Pain Mechanisms, em: Giamberardino MA - Pain 2002 - an updated review: Refresher course syllabuss. IASP Press: Seattle, 2002;155-161.
• 67. Mannion RJ, Woolf CJ - Pain mechanisms and management: a central perspective. Clin J Pain, 2000;16:S144-156.
• 68. Rowbotham MC, Fields HL - The relationship of pain, allodynia and thermal sensation in post-herpetic neuralgia. Brain, 1996; 119:347-354.
• 69. Choi B, Rowbotham MC - Effect of adrenergic receptor activation on post-herpetic neuralgia pain and sensory disturbances. Pain, 1997;69:55-63.

*

Recebido da Faculdade de Medicina da Universidade Federal da Bahia (UFBA), Salvador, BA

Publication Dates

• Publication in this collection
  12 Sept 2008
• Date of issue
  Oct 2008

History

• Received
  16 Feb 2008
• Accepted
  23 June 2008