Pain Pathophysiology

Introduction

The use of pain-relieving substances dates back to the dawn of humanity. In particular, in animistic cultures, psychotropic substances were used for shamanic practices to seek a state of ecstasy that brought them closer to the primary source of life and knowledge. This laid the foundations for a proto-medicine whose primary goal was the relief of pain caused by wounds or illness. Pain, like illness, was often considered the result of a curse from demons or hostile deities, which could only be driven away through magic and the "food of the gods" or Vedic soma ("All the gods drink soma mixed with cow's milk and pressed by the priests..." from a passage of the Rig Veda) .

Moving forward in history, as early as the 18th Egyptian Dynasty, the effects of the sleep-inducing poppy on the psyche were known, likely imported from Palestine where it was indigenous. By the 3rd century BC, sleep-inducing poppy was cultivated (in the Faiyum) to extract oil. The Greeks were aware of the effects of opium, learned from Arab physicians and traders, and introduced it to the Romans, who used and abused it in wars and celebrations. With Pliny, we have the first recorded use of opium in medicine. During the Middle Ages, the use of plants and psychotropic substances was often attributed to witchcraft—women who, having learned the secrets of nature, used them as a trade, replacing official medicine. After centuries of abuse and bans, opium returned to medical practice with Paracelsus in the early 16th century .

On the other side of the ocean, Native Americans had been using psychotropic substances and willow bark extract for centuries to relieve pain and reduce fever. It wasn't until the 18th century, however, that willow bark was introduced to European medicine when an English physician described its effects on malarial fevers to the president of the Royal Society. In 1829, Leroux isolated a bitter-tasting glucoside from willow bark, called salicin, which was identified as the active compound and demonstrated its antipyretic effect. In 1875, sodium salicylate was formed by hydrolyzing and carboxylating salicin, and it was used in antipyretic therapy. Shortly thereafter, Hoffman synthesized acetylsalicylic acid, which was marketed by Bayer in 1899 under the name Aspirin. Soon after, synthetic salicylates replaced the more expensive natural ones, and by the end of the 19th century, new drugs with similar effects were synthesized .

In 1975, R.G. Black of the University of Washington suggested the term "chronic pain syndrome" to describe patients suffering from persistent and intractable painful conditions. Around the same time, Stenbarch introduced the concept of chronic benign pain syndrome, in contrast to chronic malignant pain associated with cancerous conditions. Subsequently, other authors have discussed benign non-neoplastic chronic pain or intractable benign pain syndrome .

In the 1980s, Cervero advocated for the differentiation between visceral and somatic pain. Pain of ectodermal origin (skin and mucous membranes) is characterized by good localization and definition, while mesodermal-origin pain corresponds to the current concept of deep pain. Endodermal-origin pain, on the other hand, is represented by visceral pain: "the imprecise localization and dull nature would be greater in endodermal-origin pain than in mesodermal-origin pain" .

The temporal evolution and peculiar characteristics of pain allow for the identification of two types: acute and chronic. New classifications based on the type of stimulation received by pain receptors divide it into transient (without significant tissue or nerve damage), reversible (significant damage that heals within days or weeks), and irreversible (significant damage with persistent inflammation, tissue loss, or neuronal damage) .

Before delving into the discussion on pain, it is essential to define some commonly used terms:

Inflammatory pain: refers to pain that originates from tissue damage, followed by an inflammatory reaction.

Neurogenic pain: a pain syndrome resulting from an alteration in the mechanisms of pain transmission and modulation in the central and peripheral nervous systems. When the pain is caused by damage to peripheral nerves, it is referred to as neuropathic pain.

Allodynia: refers to a decrease in pain threshold; a stimulus that is normally not painful is perceived as such. Allodynia can be classified based on the type of stimulus (thermal, mechanical, or chemical) and the mode of stimulation (pressure, prick, touch, heat, cold, type of chemical substance).

Hyperalgesia: refers to an erroneous perception of a painful stimulus that is nonetheless exaggerated in intensity. -

Hyperpathy: is a syndrome characterized by an increased threshold for pain, where the stimulus is not perceived as painful until it reaches a certain intensity or frequency. Subsequently, the stimuli are perceived in an exaggerated manner and for a prolonged period .

Definition of Pain

The International Association for the Study of Pain (IASP) defines PAIN as "an unpleasant sensory and emotional experience associated with actual or potential tissue damage, or at least the subjective sensation of it" . For this reason, pain falls within the area of somatosensory perception.

There are many qualities that can be attributed to pain, but the fundamental one that defines it is "unpleasant," both in terms of the sensory experience and the emotional response, which is the result of personal reprocessing. The activity induced in nociceptors and nociceptive pathways by a painful stimulus is not pain; pain is always a psychological state, although we must consider that it often has an underlying physical cause .

As we have seen, pain is a subjective sensory experience that manifests at various levels of consciousness and triggers a series of behavioral patterns aimed at preserving the individual's integrity. The multidimensional perception includes strong emotionality, increased motivational drive to reduce or prevent pain, cognitive and motor strategies aimed at preventing tissue damage, and the memory of the unpleasant affective and discriminatory experience.

Between 1960 and 1965, Melzack and Wall developed this theory, bringing order to the confusion surrounding the explanation of certain pain phenomena.

This phenomenon, essential for the regulation of pain, involves the Substantia Gelatinosa (lamina II) of the spinal cord, which consists of a dense network of interconnected interneurons and the second sensory neuron (lamina V). It receives four types of fibers: Aδ and C fibers (nociceptive) from the periphery, Aβ fibers from the periphery (tactile and nociceptive with excitatory action), and the descending inhibitory system from the thalamus via the dorsolateral funiculus.

The theory is based on the following assumptions:

• The interneurons of the Substantia Gelatinosa (SG) have an inhibitory effect on lamina V of the spinal cord, from which the lateral spinothalamic tract originates, transmitting the nociceptive stimulus centrally.
• $A\delta$ fibers project onto the SG with excitatory action, promoting the inhibitory effect on lamina V through the release of opioid peptides.
• $C$ fibers project onto the SG wi$A\delta$th inhibitory effects, blocking the release of endorphins and opening the gate, allowing the pain stimulus to pass.
• $A\beta$ fibers and the descending inhibitory system release opioid peptides and block the pain stimulus.
• Lamina V gives rise to the axons of T cells, which form the lateral spinothalamic tract. This tract ascends to the ventral posterolateral (VPL) nucleus of the thalamus, where nociceptive sensitivity is processed.

Reversible Pain

Acute Inflammatory Pain

Among the various pain syndromes, acute inflammatory pain is the most representative of reversible pain. The inflammatory response is the common pathway of many events that cause tissue damage and is also one of the most frequent causes of peripheral pain. Historically, the signs of inflammation are “rubor, calor, tumor, dolor, and functio laesa.” Tissue damage induces the release of pain and inflammation mediators from damaged or necrotic cells. The pain initially perceived is due to an appropriate stimulus to low-threshold nociceptive fibers (polymodal C fibers), resulting in neuronal firing and the induction of a local vasoconstriction reflex; if this stimulus is not repeated, the firing tends to subside, as does the pain perception.

The most important mediators of acute pain are the same ones that actively participate in inflammation: histamine, serotonin, bradykinin, substance P, and eicosanoids. The local release of intracellular enzymes triggers the complement and kinin cascades, which, together with high concentrations of $K^{+}$ and ATP lost from cells, stimulate the activation of high-threshold nociceptive fibers. The identification of acid-sensitive ion channels in small-diameter nerve fibers opens up the possibility of their involvement in hyperalgesia mechanisms in inflamed and hypoperfused tissues (local acidosis) .

The products of inflammatory cascades and cellular degradation act as chemotactic factors for inflammatory cells, inducing increased blood flow, increased vascular permeability, and sensitization to painful stimuli. The activation of phospholipase A2, triggered by tissue damage or pro-inflammatory cytokines ($IL-1$,$IL-6$,$TNF-\alpha$ and $INF$), induces the release of arachidonic acid from cell membranes. This comes into contact with 5-lipoxygenase (the most important enzyme in neutrophils) and cyclooxygenase (COX), both type 1 (constitutive in vascular endothelium) and type 2 (inducible by cytokines in inflammatory cells) . This leads to the release of leukotrienes (LTB4), thromboxanes (TxA2), and prostaglandins (PGD2, PGI2, PGF2α, and PGE2), important mediators of edema, leukocyte recruitment, and pain . Prostaglandins induce hyperalgesia and sensitization by binding to metabotropic receptors present in polymodal C and Aδ fibers. The increase in intracellular $Ca^{2+}$and IP3/DAG activates kinases (Tyr kinase), leading to the phosphorylation of membrane channels and a reduction in the activation threshold for subliminal stimuli .

Acute Neurogenic Pain

Pain is defined as neurogenic when it originates in the nervous system. This type of pain can be divided into central or peripheral, depending on whether it originates in the central nervous system (cortex, thalamus, hypothalamus, or spinal cord) or in peripheral nerve trunks, respectively .

Central pain is a very common symptom in spinal cord pathologies but less so in brain conditions.

Peripheral pain manifests in two main types: truncal and dysesthetic. Both are generally present, but one tends to prevail over the other in a characteristic and different way in each pathology.

Truncal pain has familiar characteristics for the patient as it resembles pain from tissue injury and is caused by an inflammatory process involving the perineurium with the activation of visceral nociceptors (projection via nervi nervorum). This type of pain is prevalent in leprous neuropathy, acute herpetic NP, and compressive NP caused by osteophytes or disc herniation. It is generally reversible, and if treated early, it may resolve with the healing of the inflammatory process. However, if left untreated, it often results in nerve degeneration, leading to the development of dysesthetic pain and chronicization.

Irreversible Pain

Chronic Inflammatory Pain

The mechanisms of chronic inflammatory pain are the most representative of irreversible pain. The chronicity of pain depends on both the persistence of the inflammatory process and spinal hyperalgesia mechanisms. When an inflammatory stimulus is prolonged (weeks or months), regardless of its cause, it tends to induce a response different from the acute phase: the cellular infiltrate changes towards a persistent reaction, with recruitment of mononuclear cells (activated macrophages and giant cells, $T$ and $B$ lymphocytes, and plasma cells), extensive tissue destruction, and attempts at repair through connective tissue replacement (fibrosis and neoangiogenesis). Macrophages play a fundamental role in maintaining the inflammatory process through the production of numerous substances (enzymes, active oxygen metabolites, cytokines, growth factors, and eicosanoids) that, on the one hand, cause further tissue damage and, on the other, stimulate the recruitment of lymphocytes and neovascularization, creating a vicious cycle. COX-2 is found in high concentrations in activated macrophages but not in other cells of the chronic infiltrate .

The prolonged alteration of the tissue microenvironment establishes a vicious cycle involving nociception, with increased vascular permeability, infiltration of algogenic substances, and progressive disruption of the microenvironment itself. This results in increased excitability of peripheral nerve fibers.

The chronicity of spinal pain involves complex mechanisms, including spinal reflexes that trigger positive feedback between sensitized peripheral fibers, lamina I neurons, and anterior horn motor neurons; a local increase in the production of algogenic prostaglandins through COX-1 ; and alterations in the gene expression of various molecules, including neurotransmitters and receptors (especially among endorphins, nociceptin, nocistatin, and the ORL1 receptor¹, as well as increased expression of the capsaicin/VR1 receptor in the dorsal root ganglia and the posterior horns of the spinal cord) . Finally, at a higher level, hypothalamic or cortical mechanisms, such as neuronal damage, deafferentation, or altered control mechanisms over subspinal structures, can contribute to pain chronicity .

Chronic Neurogenic Pain

In spinal cord injuries, pain often coexists with sensory alterations. It can take on various forms: burning, stabbing, cramping, or constrictive. Generally, it is persistent and exacerbated by peripheral stimulation or movement. When caused by acute-onset conditions, it is often accompanied by autonomic reflexes such as piloerection, profuse sweating, and skin vasodilation; however, these associated symptoms tend to disappear when the pain becomes chronic. The most common cause of spinal pain is traumatic spinal cord injury, with a significant prevalence of thoracolumbar localization.

¹The ORL1 receptor has been identified in three species (mouse, rat, and human), showing a high degree of homology with "classical" opioid receptors. There is an ongoing debate regarding the naming of the endogenous peptide agonist for the ORL1 receptor, currently referred to as "nociceptin" or "orphanin FQ." Although this receptor is structurally classified among opioid receptors, there is no correlation in terms of pharmacological characteristics. Non-selective ligands with high affinity for μ, κ, and δ receptors show very low affinity for ORL1, which is why this receptor has been called the "orphan opioid receptor" .

Central pain can occur as a result of cerebrovascular events at the cortical, thalamic, or hypothalamic levels. The most well-known syndrome is Dejerine and Roussy’s thalamic syndrome: persistent burning pain, sometimes oppressive or heavy, can be exacerbated by tactile stimuli and have characteristics of allodynia and hyperpathy. Pain is associated with ipsilateral hypoesthesia and neurological symptoms. Similar symptomatology can be seen in pseudothalamic syndromes, where afferent or efferent connection fibers are damaged. Multiple sclerosis is another condition with peculiar painful symptoms, presenting with a range of paroxysmal pain crises, including trigeminal neuralgia, painful tonic spasms, Lhermitte’s sign (a painful shock along the spine triggered by head flexion), paroxysmal burning pain in the limbs, and pain associated with optic neuritis. In general, these pains are persistent, burning, or oppressive, and resistant to therapy .

Dysesthetic, or neuropathic pain, is initiated or caused by a primary lesion or dysfunction of the nervous system . It has unusual perceptual characteristics for the patient, never experienced before. It occurs during neuronal regeneration and prevails in major peripheral neuropathies. It can manifest as pain associated with dysesthesia, burning, and stabbing sensations, sometimes accompanied by allodynia and hyperpathy. Even when treated, it rarely resolves completely .

Neuropathic Pain

Neuropathic pain refers to pain initiated or caused by a primary lesion or dysfunction of the nervous system . It is caused by heterogeneous alterations of the Peripheral and/or Central Nervous System (traumatic, metabolic, neoplastic, inflammatory, etc.), resulting in functional abnormalities characterized by hyperactivity of the nociceptive system, central induction of neuronal plasticity phenomena, and subsequent memory formation . It can also be evoked in the absence of peripheral stimuli .

Pathophysiology of Neuropathic Pain

Here are some of the mechanisms that may underlie neuropathic pain:

Ectopic generators along the fiber**:

• Regeneration neuritis and neuroma formation;

• Site of chronic damage
• Mechanical stimuli along the fiber or in the ganglion.

Synaptic modifications

• Previously ineffective synapses become active
• Increased efficacy in residual synapses
• Formation of aberrant connections
• Hypersensitivity to transmitters and neuromodulators.

Loss of inhibition

• Deficit in peripheral tactile input from $A\beta$ fibers
• Deficit in descending inhibitory systems.

Mixed pain: Central Sensitization

• Alteration of spinal receptive fields
• Alteration of cortical perception .

Currently, the most accepted hypothesis is neuronal damage. According to this hypothesis, the lesion affecting the peripheral or central somatosensory and visceral pathways would cause the onset of abnormal impulses originating from neuromas, axonal alterations, and sensitized nociceptors . This damage increases the sensitivity of the primary afferent nociceptor (peripheral sensitization) and the second-order sensory neuron (central sensitization), and it alters spinal and supraspinal modulatory systems .

Patients with painful neuropathy develop two closely related phenomena: hyperalgesia and allodynia. These are considered characteristic of this type of pain, as they are symptoms of anatomically or functionally detectable lesions.

Peripheral Mechanisms of Hyperalgesia

Nociceptors

Generally, the nociceptors involved are unmyelinated free nerve fibers located in the skin, muscles, and viscera. The cutaneous nociceptors are numerous (200/cm²); most of them are polymodal $C$ fibers and $A\delta$ fibers, which respond to various types of stimulation, while less frequently, there are specific C fibers (thermal or mechanical stimuli). $A\delta$ fibers seem to mediate immediate sensitivity, while $C$ fibers mediate delayed perception. The receptive fields of these fibers tend to overlap, so that a punctate stimulus can activate multiple fibers. Among the afferent fibers, $C$ fibers predominate (80%) .

Sensitization of Nociceptors

The mechanisms of pain transduction are still only partially understood. Ion mobilization across membranes is responsible for the action potential, but this alone does not explain some complex processes.

The reactivity of this system can be modified by the local microenvironment in the tissues. It has been shown that some substances can be algogenic, provoking nociceptive stimuli (e.g., 5HT, bradykinin, H+ and K+ ions), while other substances can be sensitizing (e.g., prostaglandins, leukotrienes, and histamine). Substance P,($SP$) produced by both central and peripheral nerve terminals, can cause vasodilation and plasma exudation. The cellular infiltrate, caused by trauma and inflammation, releases various peptides locally, including Nerve Growth Factor ($NGF$), $SP$, and $CGRP$. All of this underlies the different responses elicited by various stimuli and lesions. Reduced threshold and latency of discharge, increased response to painful stimuli, and persistent discharges after prolonged stimuli, or even spontaneous activity, are phenomena suggesting increased system sensitivity.

Some authors have demonstrated that sensitization can propagate from mechanical allodynia and hyperalgesia mechanisms to thermal ones when receptive fields are restored through axon reflexes. After an initial tissue injury, the skin becomes much more sensitive to thermal stimuli.

A pro-inflammatory effect and nociceptor sensitization have been obtained after antidromic activation of $C$ fibers, without the involvement of the CNS, and this effect persisted even after nerve sectioning. Finally, it has been demonstrated that there are normally silent visceral and articular receptors that activate only after sensitization due to inflammatory processes. The recruitment of these receptors may be one of the causes of hyperalgesia and allodynia .

Studies on induced neuropathic pain in guinea pigs and patients with neuropathic pain have demonstrated abnormal ectopic activity near or at the sites of nerve injury. The mechanisms identified include abnormal distribution of $Na^{+}$ and $Ca^{2+}$ channels on regenerating nerve sprouts and/or neuromas; sensitization to various mediators or modulators (e.g., bradykinin, histamine, 5HT, capsaicin, etc.); increased sympathetic response, which may result in greater sensitivity to pain mediators; and production of cytokines ($IL-1$ and$TNF-\alpha$) involved in central and peripheral pain sensitization, mediated by the protein kinase cascade .

Central Mechanisms of Hyperalgesia

Primary and Secondary Hyperalgesia

The area considered primary hyperalgesia, where peripheral mechanisms predominate, is where the skin is inflamed. The surrounding area is that of secondary hyperalgesia, where central processes are more significant, characterized by a reduced threshold to mechanical and thermal stimuli. Among the mechanisms of mechanical hyperalgesia, pressure stimuli are more extensive and last longer than tactile stimuli.

It is also highly likely that these hyperalgesic phenomena result from a combination of peripheral and central mechanisms: the extension of hyperalgesia progresses alongside receptor sensitization. Recent studies have shown that secondary hyperalgesia can extend beyond the area of edema and that these processes can be dissociated .

Central Sensitization

Central sensitization manifests as a lasting reduction in the activation threshold and increased responsiveness to cutaneous stimulation by dorsal horn neurons. NMDA receptors, sensitized by inputs from high-threshold primary afferents, play an important role in its induction and maintenance .

Neuronal Sensitization

The stimulation of hyperalgesia after local injection of capsaicin can be prevented by the prior administration of local anesthetic, either partially through infiltration or totally through nerve block. When the anesthetic is administered after hyperalgesia has been induced, no effect is observed. It appears that this phenomenon is initiated by nociceptive impulses that do not originate peripherally. Using microneurography, it has been demonstrated in humans that the capsaicin receptor (VR1) is responsible for the sensitization of spinothalamic neurons. This increased sensitivity is not altered by local anesthesia .

Receptive Field Modifications

The plasticity of neuronal receptive fields is another important factor in hyperalgesia. Inflammation and thermal or electrical stimulation are responsible for a significant increase in receptive field size. The stimulation of multiple receptive fields could partially explain allodynia and hyperalgesia through the activation of multiple afferent fibers and quiescent synapses. D-CPP, a competitive antagonist, and MK-801, a non-competitive NMDA receptor antagonist, block the development and maintenance of central sensitization and hyperalgesia. Additionally, MK-801 reduces the area of the receptive fields of nociceptive neurons in the dorsal horn, confirming their actual expansion in hyperalgesia processes .

Some studies have revealed forms of neuronal plasticity with pronounced spinal localization, the existence of which could clarify the nature of hyperalgesic states and ensure better clinical treatment. Three of these plasticity mechanisms have assumed primary importance in this context.

The Wind-up Phenomenon The wind-up phenomenon consists of a frequency-dependent progressive facilitation of a neuron's response to the repetitive application of a constant-intensity stimulus . This mechanism was explained by Thompson through intracellular recordings of potentials in spinal dorsal horn neurons and motor neurons in the anterior horn: for each 0.5 Hz stimulus at the posterior root level, a burst of action potentials is induced in the anterior horn, which can last up to 60 seconds after the stimulation ends. This phenomenon is abolished by NMDA receptor antagonists. These results suggest that the wind-up phenomenon can be seen as the temporal summation of excitatory postsynaptic potentials (EPSPs) mediated by NMDA receptors.

Long-Term Potentiation (LTP) Long-term potentiation refers to the tetanic electrical stimulation of C fibers, which lasts for a few seconds and causes a significant increase in postsynaptic potentials (around 200%) during subsequent stimulations. LTP persists after the stimulation ends for a period of 8 hours. NMDA glutamate receptors and tachykinin receptors (NK1) seem to be involved, inducing the activation of PKC, PKA, and other kinases, leading to extensive phosphorylation of membrane and channel receptors. Such mechanisms have been widely reported for peripheral nerve injuries .

Long-Term Depression (LTD) Long-term depression (LTD) involves a prolonged reduction in synaptic transmission efficacy, triggered by a moderate increase in intracellular calcium and subsequent hyperpolarization. In this phenomenon, the same receptors involved in LTP are inhibited by phosphatases .

Nociceptive Reflex

Many studies have used the nociceptive reflex to analyze nervous system sensitization, which occurs due to the activation of $C$ fibers from deep tissues like muscles but not from superficial tissues like the skin. The difference lies in the various substances released by these fibers. However, some studies may have been influenced by specific lesions present in the animal models used (spinalization or decerebration), which can increase sensitivity due to the interruption of descending cortical control pathways .

Peripheral and Central Sensitization Mechanisms

Lewis and Hardy, using peripheral nerve block techniques, demonstrated that secondary hyperalgesia depends on both central and peripheral activation. In inflammatory damage, as in capsaicin injection, the effectiveness of the peripheral block lies in preventing the development of secondary hyperalgesia, suggesting that central sensitization plays a prominent role. The development of allodynia and hyperalgesia requires the sensitization of both the central and peripheral nervous systems. Peripheral stimuli are essential for system activation and the maintenance of sensitization .

The hypotheses outlined above appear valid in most cases, except for deafferentation neuropathies. The poor results from neurolesive therapies highlight alternative mechanisms that require more effective treatments .

Molecular Sensitization Mechanisms

The phenomena of wind-up and hyperexcitability underlie the hyperalgesia and allodynia that develop in neuropathic pain. These phenomena are more complex than initially described, both biophysically and neurochemically.

Electrical Parameters

Wind-up is a frequency-dependent phenomenon of $C$-fiber discharge. Below a critical frequency of 0.2-0.3 Hz, the phenomenon cannot be observed, whereas at frequencies above 20 Hz, a habituation effect called wind-down is observed. The largest response occurs at frequencies between 1 and 2 Hz. Wind-up also depends on other parameters that determine adequate stimulation of C fibers; in fact, the duration of stimulation is crucial (increasing the stimulation time can generate wind-up even at lower discharge frequencies). Therefore, wind-up is generated by the temporal and spatial summation of impulses from a larger population of $C$ fibers, recruitable by broader impulses. Another important parameter for wind-up generation is the initial excitability level of spinal neurons: in cases of spinal hyperexcitability, lower frequencies and discharge amplitudes are required .

Mechanisms of Genesis

Several types of neurons in the dorsal horn of the spinal cord participate in the genesis of wind-up. The neurons most prone to developing this phenomenon are undoubtedly class 2 neurons in the deep areas of the dorsal horns, with a response to cutaneous stimulation of 200%; those located in more superficial areas show a lower response, equal to 44% of the normal; finally, class 3 neurons, which are even more superficial, do not show wind-up at all. Conversely, the afferent $C$-fibers exhibit a functional anatomy that is specular to this, likely due to the fact that the deeper fibers present polysynaptic potentials, with lower synchrony, and a role in neuronal inhibition. The potential fields of superficial $C$-fibers in the dorsal horns are widely represented in lamina II, corresponding to the central neurons of the Substantia Gelatinosa; these are neurons with excitatory activity, inhibited by afferent stimuli (inverse neurons). This is due to the connections that $C$-fibers make in lamina II with the dendrites of neurons whose somas are located outside this lamina.

Characteristically, the responses of wind-up and nociceptive reflexes in the neurons of the dorsal horns of the spinal cord block signal progression after 16 and 8 stimuli, respectively; by contrast, the potential fields of C fibers show an increased response, up to 70-100 stimulations with a discharge frequency of 1 Hz. The reason for this difference is unclear, and some authors have hypothesized the existence of some inhibitory mechanism capable of counteracting the increase in response in the first two cases, which nevertheless does not alter the process of wind-up genesis in C fibers. This inhibitory mechanism has been described by Le Bars and colleagues as supraspinal and can be summarized in a few points:

• Neurons on the surface of the dorsal horns receive less control from descending supraspinal structures compared to those located deeper;
• There is a post-tetanic supraspinal depression of neuronal discharge;
• Wind-up can be modulated by supraspinal structures due to other converging nociceptive messages.

Alarcon and Cervero have demonstrated that viscerosomatic neurons in the spinal cord also show wind-up of their somatic afferents, functionally dissociated from the reflex circuit involving somatosensory neurons and trunk motor neurons .

Presynaptic Mechanisms

Schouenborg proposed a model involving a presynaptic mechanism for wind-up generation: increased synaptic efficacy through a phenomenon similar to post-tetanic potentiation. Many similarities exist: post-tetanic potentiation describes an increase in the magnitude of a postsynaptic response; systems that exhibit it are subject to response facilitation during a train of conditioning stimuli. It decreases exponentially over time, and the time constant is inversely proportional to the number of conditioning stimuli. It appears to result from a progressive increase in intracellular free $Ca^{2+}$in nerve terminals caused by multiple depolarizations, leading to an increased number of vesicles (quanta) released. The similarities between wind-up and post-tetanic potentiation suggest that wind-up may be presynaptic and act by increasing neurotransmitter release from primary afferents. In normal circumstances, wind-up is induced only by primary afferent $C$ fibers, which release GLUT and various peptides, including SP, in independent ways. The fact that different discharge frequencies can modulate the release of different molecules or peptides might seem inconsistent with the theory that wind-up facilitates their release, particularly when considering the low discharge frequencies that characterize the phenomenon. However, high frequencies are not necessary for adequate and consistent neuropeptide release, and it has been demonstrated that SP release at low frequencies is maximal .

Postsynaptic Mechanisms

The most important substances normally released in the dorsal horns of the spinal cord fall into two classes: opioid peptides and excitatory amino acids. The receptors also belong to two groups: ionotropic and metabotropic.

Ionotropic receptors include NMDA, AMPA, kainate, and AP4 (activating protein): opening these channels allows $Ca^{2+}$ ions to enter, which activates intracellular second messengers.

Metabotropic receptors are associated with G proteins that activate phospholipase C or adenylate cyclase; the second messenger activates an intracellular cascade that includes the production of nitric oxide (NO) and cyclic GMP.

Evidence that glutamate (GLU) receptors are involved in the transmission of nociceptive information in the dorsal horns of the spinal cord comes from numerous immunocytochemical and autoradiographic studies. It has been shown that intense noxious stimulation causes the release and significant increase of GLU concentration in the dorsal roots, leading to an increase in the excitability of spinal sensory neurons (central sensitization); this is thought to be involved in the genesis of wind-up and LTP .

Substance P ($SP$) and $NKA$, belonging to the neurokinin family, appear to be involved in the genesis of the wind-up phenomenon. There are also transcription regulators, such as the c-fos and c-jun proteins, which promote the transcription of important genes, including those for proenkephalin and prodynorphin. Some studies have shown the concomitant localization of c-fos with dynorphin and enkephalin mRNA in the superficial layers of the dorsal horns. The temporal expression of these mediators suggests that they may play a role in the long-term activation and sensitization of neurons: intrathecal administration of dynorphin has been shown to increase the size of receptive fields by 50% .

Ion Channels

These are a subtype of channel receptors, whose opening is determined not by a neurotransmitter but by membrane depolarization (activation threshold). Each subtype has its own kinetic characteristics for opening and closing, as well as different activation thresholds. They can be selective and specific for certain ions based on their charge or molecular weight, or they may not exhibit particular selectivity and allow multiple elements to pass through.

Voltage-gated $Na^{+}$ Channels

Various subtypes of voltage-gated $Na^{+}$ channels (VGSCs), including $Na_{V}1.6$,$Na_{V}1.7$ ,$Na_{V}1.8$, and$Na_{V}1.9$, are normally expressed in the peripheral nervous system. Their expression and organization vary between $A$ and $C$ fibers under normal conditions and undergo significant changes during neuronal damage. Characterized by an activation potential of -40 mV, NaV1.8 is the isoform most expressed in the C fibers of dorsal root ganglia (DRG) and is crucial for action potentials. Immunological, electrophysiological, and functional studies have demonstrated the involvement of $Na_{V}1.8$ in the genesis of experimental neuropathic pain (sciatic nerve ligation) in guinea pigs: it undergoes significant redistribution and reorganization in DRG cells, with initial upregulation in damaged fibers within the first 48 hours. Subsequently, at 7-10 days post-injury, after extensive degeneration of damaged fibers, $Na_{V}1.8$ shows high expression in residual afferent fibers. Hyperalgesia and allodynia resulting from nerve injury are eliminated by intrathecal administration of antisense oligonucleotides specific to $Na_{V}1.8$.

$Na_{V}1.9$ with an activation potential of -70 mV, is selectively expressed in nociceptive neurons, where it modulates resting membrane potential and the response to sub-threshold stimuli. It becomes inactivated by persistent stimulation. Antisense oligonucleotide studies have not shown interference with hyperalgesia or hypersensitivity mechanisms. Immunohistological studies have shown that NaV1.9 is downregulated in severed fibers but not in those with chronic lesions.

The $Na_{V}1.3$ subtype is normally expressed at very low levels in primary nociceptors but undergoes rapid upregulation in DRGs following axonotomy, ligation, or chronic peripheral nerve constriction. Its kinetic characteristics (fast opening and closing) make it the best candidate for generating spontaneous ectopic discharges, although a direct correlation between these discharges and $Na_{V}1.3$ expression has yet to be demonstrated. The distribution of $Na_{V}1.3$ in large- and small-diameter fibers in DRG neurons suggests that transcript and protein upregulation occurs after axonotomy in both A and C fibers, although ectopic discharges are predominantly found in the former. This highlights the importance of VGSCs in the genesis of neuropathic pain .

$L$-type $Ca^{2+}$ Channels

In the spinal cord, L-type $Ca^{2+}$ channels are expressed by motor neurons and sensory interneurons, where they mediate a slow inward current in response to depolarization and show no significant inactivation. The slow activation and inactivation kinetics lead to plateau potentials. As a result, two different functional states of stability arise in both sensory and motor compartments, one of which is wind-up. Both types of phenomena, similar to wind-up, are voltage-dependent and dihydropyridine-sensitive, involving L-type $Ca^{2+}$ channels. The gradual increase in depolarization in response to repeated current pulses is not due to cumulative depolarization of the membrane potential during the interstimulus interval but to depolarization-induced facilitation of L-type $Ca^{2+}$ channels: this mechanism is known as warm-up. The simplest scheme to explain the warm-up phenomenon is a voltage-dependent transition between two closed states of L-type $Ca^{2+}$ channels: one infrequent state with a high activation threshold and a more frequent state with a lower activation threshold. L-type $Ca^{2+}$ channels are regulated by metabotropic receptors: in the sensory compartment, they are positively regulated by GLUT and SP and negatively by GABAB. These modulators contribute to the dynamic regulation of excitability and the intrinsic characteristics of plateau currents .

Transmitters and Receptors

Adenosine (P1) and ATP (P2) Receptorsqui

Adenosine and ATP are ubiquitous mediators released by a variety of cells. Adenosine acts by binding to metabotropic receptors ($A_{1},A_{2A},A_{2B},A_{3}$) associated with excitatory or inhibitory G proteins, while ATP acts through both ion channel receptors (P2X) and metabotropic receptors (P2Y). Interactions between the adenosine receptor and other receptors, both metabotropic and ion channels, contribute to the fine regulation of nervous function . Adenosine and ATP exert numerous influences on peripheral and spinal pain transmission:

• In peripheral nerve terminals, stimulation of the $A_{1}$ receptor increases intracellular cAMP, resulting in analgesia, while activation of the $A_{2}$ receptor induces pain or facilitates nerve sensitization.
• At the spinal level, activation of the $A_{1}$ adenosine receptor produces analgesia for both nociceptive and neuropathic pain, at doses lower than those with motor effects. Spinal analgesia is mediated by the inhibition of interneurons through hyperpolarization (increased $K^{+}$ conductance) and presynaptic inhibition of the release of Substance P (SP) and possibly glutamate .

In the spinal cord, morphine can induce analgesia indirectly by releasing adenosine from capsaicin-sensitive afferents via PKC activation associated with μ-opioid receptors. PKC appears responsible for morphine-induced hypotension and the muscle rigidity associated with fentanyl administration . Significant interactions also seem possible with the spinal monoaminergic system: in pain perception tests, the effectiveness of adenosine, or its analogs, in inducing analgesia is dependent on the integrity of the noradrenergic system . Recent studies have highlighted the relevance of the $P2X_{3}$subtype in pain pathways: it is selectively expressed at high levels by nociceptive sensory neurons and plays a role in the presynaptic regulation of glutamate release in the first afferent neuron in DRG .

Glutamate and NMDA

Glutamate (GLU) is the most important excitatory amino acid present in the nervous system. It exerts its effects through various membrane receptors, both ionotropic (NMDA, AMPA, and kainate) and metabotropic ($mGLUR_{1-8}$), all of which are widely represented. Over the years, evidence has accumulated that NMDA receptors (NMDAR) are involved in the induction and maintenance of both central and peripheral sensitization during pain states, including visceral pain. This is due to its unique characteristics:

• NMDARs control an ion channel with high permeability to monovalent ions and calcium;
• Efficient activation requires the simultaneous binding of GLU and GLY;
• At resting membrane potential, the receptor is blocked by extracellular Mg2+, and both agonist binding and depolarization are required for channel opening.

NMDARs are composed of three subunits (NR1, NR2, NR3), with various isoforms that determine their chemical and biophysical properties. Recent studies have shown that NMDARs are present in both the CNS and peripheral nerves, contributing to pain perception. During inflammatory processes, the number of NMDA receptors on peripheral nerve fibers increases significantly, contributing to peripheral sensitization, while in neuropathic pain, once hyperalgesia has already been established, NMDARs no longer contribute to nociceptor sensitization. At the spinal level, NMDARs are involved in central sensitization, a state of hyperexcitability in the dorsal horns of the spinal cord characterized by sensory facilitation, expressed as hyperalgesia and allodynia. Increased NMDAR activity reflects an increased number of receptors and prolonged channel opening times due to transcriptional, translational, and post-translational modifications. PKC-mediated phosphorylation of NMDAR is of particular importance, reducing dependence on depolarization and Mg2+ block. PKC also regulates NMDAR function by activating the Src tyrosine kinase cascade, interacting with the cytoskeleton, and inducing up-regulation. Another protein that mediates many aspects of signal transduction is Ca2+-calmodulin-dependent protein kinase (CaMKII), which is persistently activated after NMDAR stimulation. It binds to the cytoplasmic domain of the receptor in a state that cannot be inactivated by phosphatases. Moreover, this kinase causes calmodulin entrapment, which may promote receptor down-regulation. Recent studies have shown that CaMKII is predominantly expressed in regions of the CNS involved in pain perception, such as the spinal lamina II and DRG, and undergoes up-regulation following inflammatory processes or peripheral lesions. This highlights the importance of the relationship between CaMKII and NMDARs in the development and maintenance of nociceptive hypersensitivity .

At the spinal level, NMDARs are also present presynaptically in the afferent terminals of small-caliber fibers, and their activation triggers the release of Substance P (SP). These fibers are part of a positive feedback loop for GLU, in response to sequential stimuli, facilitating and prolonging the transmission of the pain signal through the release of GLU, SP, and Calcitonin Gene-Related Peptide (CGRP). Multiple studies conducted on animal models have demonstrated the presence of NMDARs in both the peripheral and central afferents of visceral nociceptive pathways; notably, in this case, unlike in somatic perceptive pathways, the innocuous visceral stimulus is sensitive to NMDAR antagonists .

SP and NKA

International literature reports the involvement of neurokinins in the genesis and transmission of pain, both peripherally and centrally, although they do not appear to play a significant role in neuropathic pain. However, their interactions with the endogenous opioid system, the glutamatergic system, molecular transduction mechanisms, and their involvement in wind-up phenomena warrant a brief discussion. Basbaum, in a 1999 review, noted that deletion of the preprotachykinin A (PPT-A) gene, the precursor of SP and NKA, leads to altered perception of acute pain (thermal, mechanical, and chemical) but not chronic pain, suggesting that these neuromediators do not participate in the genesis of neuropathic pain . Hueda highlights peripheral and spinal molecular mechanisms closely related to the endogenous opioid system. In the periphery, chemical stimulation with BK or His induces Ca2+-mediated SP release; in the spinal cord, Orphanin FQ or kitorphin, through activation of the ORL1 receptor, trigger Gi1 activation and Ca2+ influx, resulting in SP release. SP, in turn, stimulates the activation of Gq/11, which activates PLC and the IP3 cascade, with prolonged calcium influx leading to NaV channel activation and depolarization .

In the spinal cord, SP plays a facilitating role in pain signal transmission . Other authors have shown that SP increases the activity of GLU and NMDAR in the dorsal horn neurons of the spinal cord, as well as the presence of NMDA, AMPA receptors for GLU, and NK1R for SP in the peripheral terminals of unmyelinated axons. Carlton and colleagues have demonstrated that GLU injected peripherally generates pain, SP enhances its effects, and the simultaneous administration of both has a synergistic effect, meaning it exceeds the sum of their individual effects. The authors support the hypothesis that primary neurons play a role in the genesis of wind-up phenomena and central sensitization . Studies in knockout mice for the NK1 receptor gene for SP have provided evidence supporting the involvement of this receptor in the aforementioned phenomena, concluding that NK1R is critical for central hyperexcitability observed in wind-up and central sensitization phenomena . Recent data suggest that NK1R may modulate the activity of L-type Ca2+ channels and, consequently, the plateau potentials observed in neurons of the dorsal horn of the spinal cord . These findings, demonstrated by Russo and colleagues in the turtle's spinal cord, seem to have been confirmed for some mammals; for these as well, SP and NKA increased calcium currents in the neuron, leading to plateau potentials . Despite the extensive knowledge on these channels and the role of calcium, numerous studies present conflicting evidence; the complexity of the topic highlights the need for further in-depth research on the role of these channels in mammals, particularly in humans.

• • CCK and Galanin**. It is well known that cholecystokinin (CCK) reduces the antinociceptive effects of opioids. The presence of CCK has been shown to overlap with the expression of opioids and their receptors throughout the CNS. CCK exerts its action via Gi/Go protein-coupled receptors, attenuating the action of endorphins and morphine. The level of CCK and its receptors, as well as its release, show considerable plasticity following nerve injury and inflammation, conditions associated with chronic pain. Such altered CCK release, along with receptor level changes in some cases, may be the cause of altered opioid sensitivity in various clinical pain conditions.

Neuropathic pain resulting from central or peripheral nervous system injury does not respond well to opioid treatment, likely due to increased activity in the endogenous CCKergic system. CCK receptor antagonists may, therefore, be useful as analgesics in combination with opioids for treating neuropathic pain . Galanin is a peptide involved in various functions, including pain perception, and it exerts its function through Gi/Go-coupled metabotropic receptors. It is normally expressed in small DRG neurons in rats, which also contain SP and CGRP, and it appears to be located in some neurons in lamina II. Electrophysiological and behavioral studies in rodents have shown that galanin produces complex effects on spinal pain perception, with a predominant inhibitory effect. Intrathecal administration of galanin enhances the analgesic effects of morphine, particularly when administered together with a CCK2 receptor inhibitor. Galanin reduces spinal hyperexcitability and the pain effects of SP. After peripheral nerve injury, a consistent increase in galanin expression and release can be observed in DRG, but not in dorsal horn interneurons of the spinal cord; additionally, there is no alteration in galanin receptor expression. The probable role of this peptide is tonic activation to suppress painful sensation in injured nerves, suggesting that low levels of galaninergic control may contribute to the development of neuropathic pain .

• • Opioids**. The main groups of opioid peptides, enkephalins, dynorphins, and β-endorphins, are derived from proenkephalin, prodynorphin, and proopiomelanocortin, respectively. Recently, a new group of peptides called endomorphins (-1 and -2), with atypical structure and high selectivity for the μ-opioid receptor, has been discovered. Another group of endogenous opioids is derived from pronociceptin, acting on the ORL1 receptor. Three members of the opioid receptor family were cloned in the early 1990s: first the δ-receptor in mice (DOR1), followed by the μ (MOR1) and κ (KOR1) receptors. These three receptors belong to the seven-transmembrane domain superfamily, coupled to G proteins, and share significant structural similarities .

These receptors and peptides are significantly involved in antinociception processes and are found in nociceptive pathways. Peripheral inflammation affects central structures and alters opioid activity; on one hand, it increases the activity of some receptor antagonists, while on the other hand, it increases the affinity and number of μ receptors, enhancing the analgesic potency of opioids. This is achieved by altering the expression of certain genes in the dorsal horn of the spinal cord . Numerous studies have found evidence of the inefficient inhibition exerted by the endogenous opioid system in neuropathic pain and related hyperalgesia; indeed, both this system and the descending inhibitory system may be inadequate for controlling pain at the spinal level. The spinal pain transmission system is under continuous control from the basal nuclei, particularly the periaqueductal gray matter and the locus coeruleus. Zimmermann and colleagues have shown that in animals with neuropathy, although these inhibitory systems are still functioning, they provide less than 50% of the normal inhibition ; similarly, Porreca and colleagues have demonstrated tonic facilitation in pain transmission in the dorsal horn of the spinal cord in neuropathic animals, driven by neurons located in the ventromedial medulla . Several studies have shown altered prodynorphin systems following peripheral inflammatory processes; furthermore, the biosynthesis of dynorphin is increased in various conditions associated with neuropathic pain following spinal or peripheral nerve injury. Although morphine is not able to exert its efficacy in neuropathic pain, a wide range of evidence suggests that it is not completely resistant, but only shows reduced sensitivity, and higher doses are needed to achieve the same response . Mayer and colleagues demonstrated that a nerve injury induced 8 days before the morphine test caused the dose-response curve to shift towards higher doses, by a factor of 6, meaning six times higher doses were needed to achieve the same response as in the control. Of particular interest is the fact that pretreatment with an NMDA receptor inhibitor (MK-801) prevents desensitization to morphine . Studies on the molecular processes of opioid regulation, tolerance, and dependence have shown that nitric oxide (NO) is closely linked to these mechanisms: not only do opioids influence NO release, but NO itself also participates in the processes of tolerance and dependence. In the first case, it has been shown that chronic pain activates NMDA receptors, allowing calcium entry, which activates Nitric Oxide Synthase (NOS) and, downstream, guanylate cyclase. The increase in cGMP causes thermal and mechanical hyperalgesia, and tactile allodynia. On the other hand, chronic activation of μ-opioid receptors causes PKC translocation, which phosphorylates NMDA receptors, increasing calcium levels and activating NOS; in this case, NO induces tolerance and dependence. This theory has been confirmed by studies in rodents that have demonstrated significant reductions in hyperalgesia, allodynia, and tolerance following the administration of NOS inhibitors such as Agmatine and N(G)-nitro-L-arginine methyl ester (L-NAME). Interestingly, these desensitization mechanisms do not occur with endomorphins, indicating the existence of different pathways for these molecules .

• • BDNF: neuromodulator**

During development, brain-derived neurotrophic factor (BDNF) supports the survival of a neuronal population in both the central and peripheral nervous systems. In maturity, BDNF appears to act as an important modulator of synaptic plasticity. BDNF is synthesized by primary sensory neurons (presynaptic neurons) whose expression is regulated in models of inflammatory and neuropathic pain. The high-affinity receptor for BDNF, tropomyosine receptor kinase B (TrkB), is expressed by postsynaptic neurons in the dorsal horn of the spinal cord. Stimulation of presynaptic nociceptive afferent fibers induces the release of BDNF and the consequent activation of TrkB receptors, leading to postsynaptic excitability. Electrophysiological recordings in vitro show that BDNF increases discharge potential induced by stimulation of C fibers in ventral roots. Additionally, behavioral data indicate that BDNF exerts antagonism by attenuating both the second phase of hyperalgesia induced by formalin (in animals treated with NGF) and the thermal hyperalgesia induced by carrageenan antigen: this suggests that BDNF is involved in some aspects of central sensitization under conditions of peripheral inflammation. In conclusion, BDNF meets many of the criteria needed to be defined as a neurotransmitter/neuromodulator in small-diameter nociceptive neurons .

• • Wind-up and neuropathic pain**

In recent years, numerous experimental models of neuropathic pain have been developed, and the multiple changes characterizing spinal neurons have been studied, yet very few have emphasized the wind-up phenomenon. Dorsal horn neurons in animals with experimental mononeuropathy exhibit normal wind-up to electrical stimulation of C fibers. Some have shown reduced sensitivity to wind-up after dizocilpine administration, an NMDAR inhibitor. In a group of 16 patients with neuropathic pain from spinal cord injury, repeated stimulation with a von Frey filament revealed wind-up-like pain more commonly in denervated and painful skin areas than in denervated but non-painful areas . Similarly, Price and colleagues report temporal summation with repeated von Frey filament stimulation in some patients with areas of mechanical hyperalgesia or tactile allodynia; these patients also demonstrated greater intensity of wind-up-like pain . In conclusion, while there is limited evidence of changes in the intensity or quality of wind-up in experimental animal models, it is well observed in patients with neuropathic pain, although it is present in less than 50% of these patients .

• • Nitric Oxide Synthase (NOS), Heme Oxygenase (HO), and Reactive Oxygen Species (ROS) activity**

Local inflammation resulting from peripheral nerve injury plays an important role in neuropathic pain. Levy and Zochodne demonstrated the presence of endothelial and neuronal NOS immunoreactivity near the nerve lesion within 48 hours of the injury; additionally, late reactivity for the inducible isoform of NOS (iNOS) was noted 7 and 14 days after the lesion ; these findings were confirmed by Cizkova and collaborators . Heme oxygenase is an enzyme that catalyzes the formation of biliverdin and iron and carbon monoxide monoxides through the heme structure. In humans, two isoforms have been identified: HO-1 and HO-2, the latter of which is present in both neurons and glial cells in the CNS . Both of these enzymes produce highly toxic substances (nitric oxide and carbon monoxide) but play a role as neuromediators in the CNS . These two mediators are described in the literature for their influence on opioid dependence and tolerance phenomena, as well as hyperalgesia. Similar results have been reported using various techniques: Liang and colleagues first used NOS and HO-2 inhibitors and then molecular biology techniques to demonstrate that these enzymes independently modulate the molecular changes that occur during chronic opioid exposure, tolerance, and the resulting behavioral alterations. Activation of the NOS system by chronic morphine stimulation limits the analgesic capacity of the opioid; additionally, NOS knock-out rodents exhibit reduced hyperalgesia. Similarly, HO-2 knock-out rodents show reduced mechanical allodynia after withdrawal from chronic morphine therapy; while wild-type rodents (not genetically altered) exhibit two- to three-fold increased expression of NOS, NMDAR, and prodynorphin. Morphine administration increases cGMP levels in spinal neurons, and cGMP analogs cause hyperalgesia. Administration of NOS and HO-2 inhibitors significantly reduces cGMP production induced by morphine and the resulting hyperalgesia . These data collectively demonstrate that NOS and HO alter opioid action and open new therapeutic strategies. Khalil and Khodr studied the effects of reactive oxygen and nitrogen species on nerve lesion healing in rodents by measuring xanthine oxidase (XO) and lipoperoxidase (LPO) activity: XO was more active in the young population (+400% compared to control), while LPO was higher in the older population (+300% compared to control). In the younger population, healing was more frequent and occurred after the fifth week, whereas in the older population, healing occurred less frequently after the ninth or tenth week, with persistent symptoms. Early or late administration of the antioxidant tirilazad mesylate (20 mg/kg) reduced LPO levels with contrasting effects, depending on the timing of administration: it either prolonged or reduced thermal hyperalgesia, respectively. These results led the authors to conclude that reactive oxygen and nitrogen species may be responsible for delayed healing in older individuals but are still necessary for healing itself: early administration of antioxidants may negatively affect nerve lesion repair .

Neuronal Apoptosis in Neuropathies

Apoptosis is defined as programmed cell death, a phenomenon in which a cell, whether damaged or not, undergoes a series of events, either spontaneously or induced, that culminate in the disintegration of DNA and the compaction of cytological material into elements that can be easily phagocytosed by neighboring cells. Naturally, for it to be termed "programmed," there must be genetic elements regulating the process; among these are the proto-oncogenes jun, fos, bcl-2, and bax. Dysfunction of these genes and their products is involved in the pathogenesis of neoplasms, particularly in cell immortalization (Fig.1). The induction of c-jun by NGF seems to have dual properties, both in axonal regeneration and in programmed cell death. Studies on the bcl-2 oncogene have shown that its intense expression protects neurons from cell death following axotomy, while the Bax protein promotes apoptosis. In reality, these proteins interact in regulating these processes, so the expression ratio between Bax and Bcl-2 determines whether apoptosis progresses. The c-Jun protein is a regulator of the expression of Bax and Bcl-2 and is itself controlled by jun kinase (JNK): phosphorylation of c-Jun facilitates apoptosis, whereas the non-phosphorylated form promotes neuronal regeneration . Some data suggest that, in chronic pain, specific genes involved in apoptosis are active, contributing to critical changes in cell survival and the establishment of chronic pain states . Following axonal injury, some neurons in the dorsal root ganglia undergo apoptosis, resulting in deafferentation of postsynaptic spinal neurons. These in turn degenerate due to the lack of tonic inhibitory stimulation of apoptosis normally exerted by the presynaptic neuron. The preventive administration of MK-801, a competitive NMDAR antagonist, prevents cell death due to axotomy in nearly all cases . Furthermore, Whiteside and Munglani have demonstrated that, following chronic nerve ligation injury, hyperalgesia develops in parallel with neuronal apoptosis. The administration of MK-801 prevents the former and significantly reduces the latter; from this, the authors suggest that apoptosis may contribute to the development and maintenance of hyperalgesia .

Bibliography & references