Chronic pain has three unique symptoms, painful response to innocuous stimuli (allodynia) that may manifest as burning or itching sensation, increased sensitivity to noxious stimuli (hyperalgesia) and the pain in the absence of stimuli (spontaneous activity)62. These behaviours come about as a result of a complex series of interconnecting events that cause sensitisation of the nervous system21. In most cases, these changes are driven by the compensatory immune response that is triggered by neuronal injury. After healing, inflammation usually dissipates and the sensory circuitry involved to returns to normal, however in chronic pain the peripheral and central nervous system become permanently sensitised19.
Sensitisation is caused by changes in ion channel expression and activity. Of particular interest is the up-regulation of Nav1 channel family. Several attempts have been made to pin- point the major Nav1 subtype involved in hyperexcitability. Through the use of channel blockers and knock-down (KD) mouse models, Nav1.3, Nav1.7 and Nav1.8, have all been implicated in neuron hypersensitivity; KD models involve a reduction in the expression of a vital genes 19,63. Another cation channel, that has been attributed to sensitisation, is the HCN (hyperpolarisation-activated cyclic nucleotide-gated) channel. HCN channels are non-specific cation channels that are activated close to the threshold and help achieve depolarisation required for action potentials. These are known to be up-regulated during chronic pain and HCN antagonists are able to reduce spontaneous activity 64. Down-regulation of K+ channels
and up-regulation of Ca2+, also play central roles in membrane hyperexcitability. In the case of temperature dependant channels (e.g. TRPV1-4), the activation threshold can be lowered so significantly that nociceptors are triggered by normal body temperature, thus giving the illusion of pain without a stimulus3.
Altered expression and function of ion channels is the end-point of transcriptional events that are driven by restorative inflammation22. Pro-inflammatory mediators (e.g. ATP, bradykinin, PGE2, etc.) trigger a series of calcium-dependant intracellular events, usually
involving protein kinases, that lead to the activation of transcription factors65.
These events are complex and not well understood however a number key transcription factors have been identified, including SOX-11 (Sry-related HMG box), AP-1 (activator protein 1) and ATF3 (Activating transcription factor 3)19. Inflammatory mediated up-regulation is not
just limited to activation of transcriptional pathways. Inflammatory proteins can aid the
Figure 1.7. Structures of prostaglandin E2 (PGE2) (top left), adenosine triphosphate (ATP) (top right)
trafficking and integration of ion channels, as well as direct or indirect agonism. For example, TRPV1 opening is mediated by inhibitory PIP2 (phosphatidylinositol 4, 5-bisphosphate).
Bradykinin acting on bradykinin receptor (B2) receptors triggers the breakdown of PIP2 by PLC
(phospholipase C) into DAG (diacyl glycerol) and IP3 (inositol 1, 4, 5-trisphosphate). IP3
increases intracellular Ca2+, while DAG activates protein kinase C (PKC) which can down- regulate potassium channels (Figure 1.8)66. Bradykinin, may also trigger the conversion of arachidonic acid into 12-HPETE (hydroperoxyeicosatetraenoic acid). 12 HPETE mimics the effect of vanilloids on TRPV channels67; 68. While, NF-κB acts on PAR2, triggering a kinase cascade, that involves ERK1/2 (extracellular-signal-regulated kinases) and JNK (c-Jun N- terminal kinases). This ultimately leads to activation of the previously mentioned PIP2-PLC
pathway33.
Phosphatidylinositol 4, 5-bisphosphate (PIP2)
Diacyl glycerol (DAG)
Inositol 1, 4, 5-trisphosphate (IP3)
PLC
Figure 1.8. Breakdown of PIP2into IP3 and DAG via the enzyme PLC: This pathway is triggered by
bradykinin acting on B2 receptors. PIP2 regulates the opening of TRPV1, while IP3 increases intracellular Ca2+ and DAG down-regulate potassium channels66.
Another prominent inflammatory mediator in sensitisation is brain-derived neurotrophic factor (BDNF). This neurotrophin is released from affected neurons and immune cells during inflammatory pain and is heavily involved in the sensitisation of the nervous system65. BDNF increases glutamate activity via phosphorylation of NMDA subunits, NR1; more activate subunits means more functional receptors. Blocking BDNF production has been shown to reduce both hyperalgesia and allodynia69.
Spontaneous activity is likely to be caused by the activation thresholds of nociceptive neurons lowering to that of normal body temperature70. Allodynia is the perception of pain following an innocuous stimulus (i.e. light touch). Malfunction of non-nociceptive neurons usually occurs at the site of injury (e.g. sensitisation) or in the dorsal horn (e.g. disinhibition), where sensory neurons from the PNS and CNS synapse. In most cases this type of pain occurs following mechanical stimuli (e.g. pressure, contraction, distension and sound), so allodynia is often referred to as mechanical allodynia71.
Normally, neurons specialising mechanoreception are large, fast conducting myelinated Aβ-fibres3. Intracellular kinase cascades, such as the MAPK/ERK (mitogen-activated protein kinase/ extracellular signal-regulated kinases) pathway that is normally seen in noxious signalling has be revealed to occur in Aβ-fibres after neuronal injury72. Pro-inflammatory proteins (e.g. BDNF) are known to upregulate mechanosensory ion channels, PIEZO and TRPC. PIEZO2 (Piezo-type mechanosensitive ion channel component 2) is the pore forming subunit of cation channels found in sensory neurons and has been implicated in the development of allodynia73; 74. It is probable that increased sensitivity of both primary nociceptive and sensory fibres may influence projection neurons across the dorsal horn, i.e. activation of peripheral
mechanosensory neurons creates nociceptive inputs in the CNS. In addition, the activation threshold of, mechanically insensitive, TRPV channels may become so low that they respond to mechanical stimulus73. As well as neuronal sensitisation, it was a suggested that growth factors, released during inflammation, act on uninjured Aβ-fibres causing them to sprout new axons that grow into laminae II75. Recently it has been shown that these ‘sprouts’ may be an undiscovered sub-class of nociceptive neurons that become far more pronounced following nerve injury, the precise nature of these neurones remains a mystery76.
Hyperexcitability of the nervous system is just one of the two major contributors to dysfunctional pain. The other is a malfunction of natural pain regulation between the PNS and CNS. Loss of inhibitory activity in the dorsal horn of the spinal cord is different from the descending pathways that involve 5-HT, NA and opioids77. Inhibition of the ascending pain pathway is heavily involved in regulation of signals from the PNS to the CNS and the interactions between nociceptive and non-nociceptive neurons.