JOURNAL OF CELLULAR PHYSIOLOGY 209:812 (2006)

REVIEW ARTICLES

Neurobiology of Pain
M.C. PACE, L. MAZZARIELLO, M.B. PASSAVANTI, P. SANSONE, M. BARBARISI, AND C. AURILIO*
Department of Anaesthesiological, Surgical and Emergency Sciences,
Second University of Naples, Naples, Italy
The neurobiology of pain had a notable interest in research focused on the study of neuronal plasticity development, nociceptors,
molecular identity, signaling mechanism, ionic channels involved in the generation, modulation and propagation of action
potential in all type of excitable cells. All the findings open the possibility for developing new therapeutic treatment. Nociceptive/
inflammatory pain and neuropathic pain represent two different kinds of persistent chronic pain. We have reviewed the different
mechanism suggested for the maintenance of pain, like descending nociceptive mechanism and their changes after tissue damage,
including suppression and facilitation of defence behavior during pain. The role of these changes in inducing NMDA and AMPA
receptors gene expression, after prolonged inflammation is emphasized by several authors. Furthermore, a relation between a
persistent pain and amygdale has been shown. Molecular biology is the new frontier in the study of neurobiology of pain. Since the
entire genome has been studied, we will able to find new genes involved in specific condition such as pain, because an altered gene
expression can regulate neuronal activity after inflammation or tissue damage. J. Cell. Physiol. 209: 812, 2006.
2006 Wiley-Liss, Inc.

Research on pain is focused on neurobiology studies

concerning neuronal plasticity development, nociceptors molecular identity, signaling mechanisms, ionic
channels involved in the generation, modulation and
propagation of action potentials in all type of excitable
cells. All the findings open the possibility for developing
new therapeutic treatment.
The interest of researchers for receptors, neurotransmitters, second messengers, transcription factors
involved in neuronal processing, in spinal cord and in
cortical areas, increased dramatically. There are evidences clarifying the origin of chronic pain.
Now it is well known the existence of two different
kind of persistent chronic pain: nociceptive/inflammatory pain and neuropathic pain. The first, inflammation
associated, is caused by tissue damage (Fig. 1A,B). The
first lesion and the inflammatory process cause Ad and C
fibers alteration. These fibers are responsible of sensitization, recruitment of nociceptors normally silent and
ionic channels and membrane receptors activation.
Neurogenic pain syndromes arise as consequence of
central and peripheral nerve damage.
During inflammation and neuropathies there are
phenotypic changes of the dorsal root ganglions (DRGs)
with an increase in excitability, immune system signal
alteration in the CNS, endocrine modifications. It
has been demonstrated that after damage nociceptors
become hyperexcitable.
IMMUNE CELL INVOLVMENT IN
NEUROPATHIC PAIN

The activation of the immune system has a main role

in both peripheral and central abnormal sensory
processing. Zuo et al. (2003) indicated that mast cells
were activated in a model of partial sciatic nerve injury.
Perkins and Tracey (2000) studied and showed an
invasion of endoneural neutrophils into the damaged
nervea process that peaked 24 h after injury. Neuropathic pain symptoms did not develop after depleting
circulating neutrophilis at the time of nerve injury but
established symptoms did not reverse (Perkins and
Tracey, 2000). Therefore, neutrophils could have an
important role in the early stage if neuropathic could
have an important role in the early stage of neuropathic
pain development.
2006 WILEY-LISS, INC.

Several lines of evidence indicate that macrophages

and the development of allodynia or hyperalgesia
(Heumann, 1987; Myers et al., 1996; Sommer and
Schafers, 1998; Cui et al., 2000; Liu et al., 2000). A
temporal correlation between the invasion of bloodborn, macrophages and the development of allodynia or
hyperalgesia was shown. Furthermore, a lack of
thermal hyperalgesia in a neuropathic model in the
WDl mouse, which shows delayed recruitment for nonresident macrophages, has been reported. Rutkowski
et al. (2002) failed to relieve mechanical allodynia after
clodronate administration.
Along with or after macrophage recruitment, T cells
are infiltrated into damage nerves, but their involvement in neuropathic pain has been poorly studied.
Few studies have focused on the infiltration of
immune cells into the spinal cord after peripheral nerve
injury, in particular hematogenous leukocytes and
resident microglia (Sweitzer et al., 2002).
Fluorocitrate treatment, which blocks astrocyte
and microglia metabolism, inhibits neuropathic pain,
whereas minocycline, specific microglial inhibitor,
blocks the development of neuropathic pain states but
does not reduce pain that is already established
(Raghavendra et al., 2003).
The spinal implantation of microglia, activated
in vitro, simulated signs of neuropathic pain (mechanical allodynia). Microglial activation, as new studies
reveal (Zhuang et al., 2005), seems to be the first step in
the activation of immune responses in CNS. In fact,
microglia might be responsible for the initiation of
neuropathic pain states, and astrocytes may be involved
in their maintenance (Tanga et al., 2005; Zhuang
et al., 2005).

*Correspondence to: C. Aurilio, Department of Anaesthesiological, Surgical and Emergency Sciences, Second University of
Naples, Via Petrarca, 151-80151 Naples, Italy.
E-mail: caterina.aurilio@unina2.it
Received 8 March 2006; Accepted 14 April 2006
DOI: 10.1002/jcp.20693

NEUROBIOLOGY

Fig. 1. A: Skin lesion, noxious stimuli and the beginning of pain transmission. B: Skin lesion and nerve
damage. C: NGF and NGF receptor. D: Spontaneus abnormal activity and neuronal firing.

ROLE OF CITOKINES AND GROWTH FACTOR

Not only immune cells but also citochines play a role in

the perception and transmission of pain, for example,
TNFa IL-1, and IL-6 (De Jongh, 2003). The importance
of growth factors and of cytokines in the development
and maintenance of pain, as a result of different kind of
tissue damage, is still on debating. Mendell L. was the
first to identify the relation between NGF and pain. In
his laboratory he studied the mechanism of hyperalgesia
after NGF stimulus, probably dependent by mastocyte
degranulation and by the increasing discharge in
capsacin-inducted isolated sensory neurons (Fig. 1C).
The high-affinity NGF receptors (tyrosine kinase receptor ATrkA) are expressed by about 50% of nociceptors
and their activation leads to phosphorylation and
sensitization of TRPV1 receptors, which might account
for NGF-induced heat hyperalgesia. The intracellular
signaling pathways remain controversial (Bonnington
and McNaughton, 2003) NGF modulates nociceptor
gene expression (such as TRPV1, BDNF, substance P)
after retrograde transport of NGF-TrkA to nucleus,
which might underline increases in long-term nociceptor sensitivity.
McMahon focused his work on neurotopic factors
which during inflammation could play a role of neurotransmitters/neuromodulators in the dorsal horn neurons of the spinal cord, where they are released by small
caliber afferent roots, increasing the excitability of
dorsal horn neurons. The whole sensory neurons,
coming along with damaged fibers, in neuropathic
models show different gene expression. These axons,
close to progressive degenerated nerves, in some
amyelinic afferent fibers, recruit an elevated number
of macrophages, which are cause of an increase of
Journal of Cellular Physiology DOI 10.1002/jcp

neuroactive molecules such as growth factors and

chemokines. In this way intact nerves along with
degenerated afferent fibers, could receive abnormal
molecular signals from target and nerves. After nerve
damage some afferent neurons fire spontaneously.
Na CHANNEL AND GENE EXPRESSION
AFTER PERIPHERAL NERVE INJURY

Until few years ago electrophysiological data showed

that peripheral damage was the origin of abnormal
activity (Fig. 1D). Only recently it has been established
that only some fibers become origin of abnormal activity.
Primary sensory neurons could be divided in two
functionally different groups: nociceptors with fibers
C, amyelinic with low conduction, and mechanoreceptors with large myelinic fibers and rapid conduction Ad.
We can easily imagine how an abnormal or ectopic
activity derives from nociceptors, which could determinate the onset of pain in many neuropathies. We have to
consider the Ad activity, which can be the cause of pain
in presence of central sensitization. Fibers C activity
starts central sensitization and following this event the
Ad activity plays an important role inducing allodynia in
human neuropathy. Usually after lesion of spinal nerve
L5 spontaneous activity arises exclusively from myelinic
fibers, in particular during the first and the second week
after damage, when neuropathic pain stabilizes (Boucher et al., 2000). Many data show that intact nerves as
well as L4 nerve, after L5 lesion, have many plastic
changes including the presence of spontaneous activity.
Therefore, myelinic fibers show changes similar to the
damaged afferent nerves. Many Ad afferents begin to
produce high frequency impulses or trains of potentials
action against the spinal cord (Michaelis and Liu, 2000).

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PACE ET AL.

Fig. 2. A: Painful Pathway B: Na Channel TTX-Resistent. C: Two Human C-fos:C-Jun:DNA

Complexes. D: NMDA Channel.

Besides electrophysiological changes already described,

models of experimental neuropathy show changes of
primary sensory neuron gene expression. Peripheral
axons of damaged sensory fibers reveal a progressive
degeneration.
Consequently, damaged neuron cells lose their binding with target and show a different gene expression.
Some damaged fibers A with a phenotypic change
express molecules normally associated to nociceptors
(substance P, BDNF).
Altered gene expression of afferent intact nerves could
be explained by an increasing of NGF available
(Fokuoka et al., 2000). Moreover, substance P and
VR1, abundant in intact fibers C, are regulated by
NGF (Fig. 2A).
A different expression of ionic channels could contribute to explain the mechanism of spontaneous
neuronal firing. Waxman et al. (1999) demonstrated
that different type of damage causes alteration in
sodium channel expression: axonal lesion is associated
with a downregulation of sodium channel TTX-resistant
expression, whereas inflammatory damage is associated
with upregulation. Porreca et al. (1999) indicated that
sodium channels TTX-resistant, called SNS/PN3, trigger and maintain hyperalgesia and allodynia nerve
damage induced. Gold et al. demonstrated the importance of sodium channels TTX-resistant during inflammation: the discharge is modulated by mediators as
prostaglandin E2, 5-HT, and adenosin, responsible of
peripheral sensitization (Fig. 2B).
Several neuron specific sodium channels have been
cloned and sequenced using molecular techniques. The
latest revealed new nociceptive mechanisms, involving
molecules receptors and a neuronal network in the
Journal of Cellular Physiology DOI 10.1002/jcp

spinal cord and in the brain, developed after peripheral

nerve injury or nerve damage.
Almost 10 years ago, studying memory in Aplysia,
Kandel (1986) found that oncogenes, first identified in
the virus, were present also inside stimulated neurons.
One of the genes, identical to the oncogene v-fos, is
involved in neuron final changes after temporary
stimuli. This gene is constitutive and is called c-fos
(Fig. 2C). Hunt et al. (1987) observed that nociceptive
stimuli induce the expression of c-fos in the DRG and
that c-fos and other early genes, as c-jun, bind to
regulator factors in the nucleus of many activated cells.
Curran and Franza (1988) described the binding of fos
and jun proteins to a regulatory region of AP-1, as third
messenger in many cells already activated by second
messenger as c-AMP or ions. Ji and Rupp (1997)
emphasized that c-AMP responsive binding element
(CREB) was involved in the transcription of many genes,
in spinal neuron long-term potentiation and in c-fos
induction after formalin test. Draisci and Iadarola
(1989) demonstrated that c-fos mRNA is present in rat
dorsal horn neurons few minutes after the beginning of a
peripheral inflammation. During peripheral chronic
inflammation, the continuous activation of fibers C
result in gene transcription alteration in the DRG and in
the posterior horn neurons. Following peripheral lesion,
changes in the neuron excitability and in mRNA levels
in sensory neurons are substrate for chronic pain.
CENTRAL EVENTS ASSOCIATED WITH
PERIPHERAL NEUROPATHY

The prolonged activity of fibers C, even if with

moderate frequency, is able to induce a synaptic

NEUROBIOLOGY

11

Fig. 3. A: Sagittal Section showing the guide cannula rout leaved into cerebellum and reaching the Vc.
B: Sagittal section shoeing the guide cannula rout reaching the trigeinal caudalis nucleus. C: SOD
activity in right cortex. D: SOD activity in left cortex. E. Up: coronal section stained with the
SOD histochemical assay. Squares show the areas sampled to compare ROD in the Vc. Down: measured
ROD from both sides of the Vc.

conduction increase in dorsal root neurons. This central

sensitization has been demonstrated in many experimental models of inflammatory pain. Few seconds of
fibers C activity result in minutes of post-synaptic
depolarization. The latest is caused by the activation of
NMDA, glutamate, tachikinin NK-1, substance P, and
neurochinin A receptors.
The importance of glutamate, substance P, neurokinin, in central sensitization has been demonstrated
for their ability to prevent depolarization using antagonist of NMDA receptors in particular not competitive
antagonists of NMDA receptors (Fig. 2D), which
reduce nociceptive behavior formalin induced. Competitive antagonists (AP-5 for NMDA receptor) as well as
not competitive (MK-801 for phencyclidin FCP) reduce
the persistent activity of dorsal roots in association with
peripheral formalin injection (Berrino et al., 2003).
In our study (Viggiano et al., 2004) we observed after
formaldehyde injection in rat lip skin, an altered
releasing of GABA in the spinal nucleus of trigeminus
(Fig. 3A); moreover, NO synthesis mediates the
increased releasing of amino acids in the same nucleus.
Recently, it has been demonstrated that free radicals
could be involved in signal transduction of pain as
cellular mediators in a sub-toxicity condition (Smythies,
1999).
The strong evidence that nitric oxide is an important
mediator of hyperalgesia is in the CNS, but evidence for
a peripheral action is less clear. Nitric oxide is induced in
tissues during inflammation, probably through both
inducible and neuronal nitric oxide synthase (iNOS and
Journal of Cellular Physiology DOI 10.1002/jcp

nNOS). Nitric oxide donors can induce pain in humans

and NOS inhibitors can reduce inflammatory hyperalgesia in PGE2 dependent manner (Omote, 2001;
Thomsen and Olesen, 2001; McMahon et al., 2005).
For these reasons we decided to evaluate, in experimental model of pain, formalin injection induced,
superoxide dismutase (SOD) activity in section of
brain sensory nuclei using immunohystochimic technique to quantify SOD. We found a difference, statistic
significant, in SOD activity between the two side of the
encephalic trunk, in trigeminus sensory nuclei, confirming SOD as modulator, fully involved in pain
transmission (Viggiano et al., 2005; Fig. 3BE).
International scientific literature emphasized the
prevalence of benign pain chronic syndromes, more
intense, frequent and prolonged in women compared
to men. For this reason we decided to investigate
the different aspects of visceral pain, in standard
condition, in rat models, male and female, with induced
ureter stones. The rats showed behavior indicating
visceral pain (ureteral crisis) and muscle hyperalgesia
directly correlated to organ pain potential. Such a
model reproduced the same condition found in clinical
practice.
Moreover, the sexual dysmorphism present in painful
stimuli appeared us to be an important factor in order to
evaluate the role of sexual hormones in the modulation
of pain.
A brief peripheral nociceptive stimulus induces c-fos
expression in central nervous system within 30 min,
with spikes after 1 or 2 h. Then c-fos expression

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PACE ET AL.

disappears within 8 h. Therefore, c-fos seems to be a

neuronal activation index, which could localize sensory
and motoneurons involved.
CONCLUSIONS

We have reviewed the different mechanisms suggested for the maintenance of pain. Dubner (2004)
studied descending nociceptive mechanisms and their
changes after tissue damage, including suppression and
facilitation of defense behavior during pain. Many
researchers emphasize the role of these changes in
inducing NMDA and AMPA receptors gene expression,
after prolonged inflammation.
Neugebauer et al. (2004) demonstrated a relation
between a persistent pain and amygdale. The amygdale
is the center of negative affective status such as
anxiety, depression, fear. Capsular lateral division of
amygdale central nucleus has recently been defined as
nociceptive-amygdale, since it has been demonstrated,
through neuroimaging techniques, neuroplastic biochemical pharmacological, and electrophysiological
changes during persistent pain.
Molecular biology is important to clarify how altered
gene expression can regulate neuronal activity after
inflammation or tissue damage. Since the entire genome
has been studied, we will be able to find new genes
involved in specific condition such as pain, hoping that
in the next future it will be possible control pain through
gene transfer.
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