Distant plasticity in the trigeminal nerve,

ganglion, and central endings

Fig 7-8 Different patterns 01 nerve libers
detected by immunocytochemistry lor CGRP in
rat molars. (a) Pattern in normal molars (bar =
0.2 mm). (Modilied from Kimberly and Byers56
with permission.) (b) Pattern 4 days after a large
abscess (asterisk) is induced near a cervical
dentinal cavity (bar = 0.2 mm). (Modilied Irom
Taylor and Byers'" with permission.) (e) Pattern
after 3 weeks 01 healing and reparative dentin
(RD) lormation at an injury site that was similar
to that in (b) (bar = 0.2 rnrn). (Modilied Irom
Taylor and Byers'" with permission.)
Fig 7-9 (a) Rat molar denervated several
days belore a small pulpal exposure. By 6 days
later, the pulpal damage and necrosis are severe
(arrowheads and bar). (Reprinted lrom Byers and
Taylor59 with permission.) (b) Innervated contralateral
tooth with many sprouting nerve libers.
Compared with the tooth in (a), it has only a
small loss 01 pulp (arrowheads and bar) alter a
small pulpal exposure. The sprouting nerve libers
show immunoreactivity for CGRP.(Reprinted lrom
Byers and Taylor59 with permission.) (e) Rat molar
root with nerves and vital pulp retained. At 14
days alter a pulpal exposure, there is already
a periapical lesion with CGRP-immunoreactive
sprouting libers (bar = 0.5 mm). (Reprinted lrom
Khayat et al60 with permission.) (d) Normal periapical
region 01 a rat molar immunoreacted lor
CGRP. The region shows normal, sparse innervation
01 the periodontal ligament. The arrow
shows the adjoining periapical nerve (bar = 0.1
mm). (Reprinted lrom Kimberly and Byers56 with
permission.) (e) Periapical changes and sprouting
nerve libers appear 3 to 5 weeks lollowing
establishment 01 irreversible pulpitis subsequent
to pulpal exposure lesions. Compared to that
observed in id), the nerve liber immunoreactivity
1O't CGRP was also enhanced in the adjoining
periapical nerve (arrow) (bar = 0.1 rnrn), (Reprinted
lrom Kimberly and Byers56 with permission.)
ganglion4,45 (see Fig 7-7), at their sensory endings in
the brainstem, and in the neurons within the central
nervous system. Many of the responses at the ganglion
are similar to those shown for spinal nerves
responding to tissue inflammation, including altered
expression of neurotrophin receptors, neuropepti
des, and voltage-gated ion channels by the neurons
and increased expression of injury proteins by
the satellite cells. Those changes can have profound
The discussion so far has focused on dental sensory
reactions in the terminal branches within the tooth
or nearby tissues. These neurons also have extensive
changes in their alveolar branches'" (see Fig 7-9), at
their cell bodies and satellite cells in the trigeminal
140
Structural and Cytochemical Responses to Tooth Injury and Infection
Fig 7-10 Conlocal micrograph to demonstrate the overall nerve innervation
pattern within the coronal region 01 a normal molar dental pulp. Nerve libers
are identilied with both neurolilament 200-kD and GAP-43 immunoreactivities
(green), while nuclei are identilied with TO-PRO immunoreactivity (blue). Axon
bundles are located within the midcoronal region (black ertow), which leads to
the many axons within the subodontoblastic plexus (arrowhead). Some 01 the
libers within the subodontoblastic plexus enter and traverse the odontoblastic
layer (white arrow) , The nuclei 01 the odontoblasts are more numerous and
larger than are the nuclei 01 other cellular proliles elsewhere in the pulp.
effects on central pain pathways. For example, tooth
injuries can cause persistent expression of the c-Fos
transcription factor by central neurons, which may
indicate altered central pain pathway functionsó2,63
Atypical chronic dental pain and referred pain both
involve long-term shifts in central processing of
peripheral inputs. Chapters 8 and 9 provide further
discussion of tooth pain and the extraordinary functional
and cytochemical plasticity of peripheral and
central neurons responding to the input of orofacial
sensory neurons.
Delayed neural reactions
Both the sensory and the sympathetic fibers can
have important reactions that are not launched until
days or weeks after tooth injury. For example, the
alveolar nerves that carry dental axons can greatly
change their neuropeptide content by several weeks
after a pulpal exposure in rats= (see Figs 7-9d and
7-ge). The sympathetic innervation initially was not
found to sprout during the early stages of neuropulpal
reactions to pulpal exposure, but, by several
weeks later, it too has focal responses directed
toward the lesion.? The late sympathetic reactions
have a major effect on immune cell invasion of the
injured pulp and may even alter the quality of tooth
pain. Thus, while the initial sensory sprouting reactions
are important, subsequent reactions in those
fibers, in the sympathetic neurons, and at central
neural pathways must also be appreciated for their
roles in tooth pain.64-66
Human teeth
The results of studies performed in animals have
provided important information regarding the neuroanatomical
responses in the diseased or damaged
dental pulp. Certainly the advantage of these
studies is that responses can be evaluated at different
time points following a standardized insult.
Another distinct advantage is the ability to evaluate
the broad effect of these injuries within the entire
trigeminal neuroaxis. Even given these advantages,
some limitations exist in animal studies, and most
notable is the relationship of these neuroanatomical
responses to pain and especially pain in humans.
In this regard, knowledge gained in animal studies
must be applied to the study of the human dental
pulp, where pain levels and response to stimuli can
be documented prior to extraction.
The human dentalpulp is richly innervated-a
common source of pain-and so its use is wellsuited
for such studies. Also, the routine extraction
of both normal third molars and diseased teeth
provides an abundant supply of specimens for study.
Together, the results from human and basic animal
studies can further the understanding of possible
correlations between neuroanatomical responses
and pain mechanisms in an attempt to more fully
understand pulpal pain and its important relationship
to the practice of endodontics. In general, the
innervation of human dental pulp (Fig 7-10) is similar
to that seen in experimental animals, and these
similarities strengthen the use of animals as a model
for understanding response to injury in the human
dental pulp.
141
Dentallnnervation and Its Responses to Tooth Injury
10
9
~8
'.",; 7

6 C>.o

'" 3
- C>. 5o 4

i':! 2 <t
1
O
None Moderate Gross
9 Degree of caries in pulp horn
10

9 e: 8
'.",; 7
C>. 6
\C!>). 5
15 4
'" 3
i':! 2 <t
1
O
None Moderate Gross
h Degree of caries in subodontoblastic plexus
10
9
~8
'.",; 7
C>. 6
\C!>). 5
15 4
'" 3
i':! 2 <t
1
O
None Moderate Gross
Degree of caries in midcoronal pulp
Fig 7-11 Differences in the distribution and morphology 01 protein gene product 9.5 (PGP 9.5)-immunoreactive nerve libers located in intact
(a and e) and carious (b,
d, e, and I) human teeth. PGP 9.5-immunoreactive libers in the pulp horn 01 an intact primary tooth are lewer (a) and thinner (e) than are
observed in a carious primary
tooth (b and d). The PGP 9.5-immunoreactive libers surround intrapulpal abscesses (AB) in the pulp horn 01 a carious permanent tooth (e),
whereas PGP 9.5-immunoreactive

libers show a Iragmentation and reduction in density within the pulp horn 01 a grossly carious primary toothm (bar = 13.5 urn in e, d, and e;
bar = 27.0 prn in a,
b, and f¡. Graphs show the mean percentage area 01 PGP 9.5 staining in the pulp horn (g), the subodontoblastic plexus (h), and the
midcoronal pulp (i) regions 01 primary
(open bars) and permanent (filled bars) teeth, according to the degree 01 caries present within the specimens. (Reprinted Irom Rodd and
Boissonade" with permission.)

Primary and erupting teeth

Although the innervation of permanent teeth has
been extensively studied." detailed descriptions of
the innervation of the primary tooth are more recent.
The coronal regions in both are more densely innervated
than radicular areas, but, while in permanent
teeth the pulp horns exhibit dense innervation, in
primary teeth the cervical third of the coronal region
is more densely innervated.v-" Dentinal tubules are
innervated in both." but primary teeth show lower
levels of overall innervation'" (Fig 7-11) and of some
neuropeptides, including CGRP, substance P (SP),
and vasoactive intestinal peptide (VIP),lo In addition,
results from animal studies show that innerva-
142
tion density can vary depending on tooth maturity
because newly erupted teeth show fewer nerves
than older teeth.l1.72
The aforementioned data are consistent with the
clinical impression that primary teeth are less sensitive
than permanent teeth?O The special anesthesia
for children's dentistry may also depend on immature
central pain perception mechanisms in infants
and children.l3.74 Specific studies of sensitivity during
tooth eruption and root maturation show sharp
increases in sensitivity once the root apices close."
and similar findings have been reported in animal
studies." That shift in sensitivity when root apices
close may depend on specific maturation of the
peptidergic system in rat molars at that time."
Structural and Cytochemical Responses to Tooth Injury and Infection
Peptidergic dental innervation
Human pulpal nerves show high levels of neuropeptides."
and their presence and changes in distribution
within carious teeth are among the most
extensively studied areas of human pulp neurobiology.
Their prominent expression in pulpal axons also
allows a convenient method for the visualization of
pulpal axons when the anatomical response to injury
is examined.
Neuropeptides that are located within human
pulpal axons include the tachykinins, neurokinin
A (NKA)78and SP,79CGRP,80VIP,81neuropeptide Y
(NPY),82cholecystokinin and somatostatin.?? galanin,
83 and methionine- and leucine-enkephalin."
The importance of neuropeptides, and especially
CGRP and SP,is well documented in the process of
neurogenic inflammation, where the release of these
peptides from sensory axons contributes to blood
flow control, inflammation, and tissue repair.14.84
Neurogenic inflammation also increases the excitability
of nociceptors and thus is important as a
peripheral pain rnechanisrn.'?
In addition to broad distributions within both
myelinated and unmyelinated sensory axons
throughout the pulp, some neuropeptides are intimately
associated with arterioles; these include
CGRP,SP,VIP,and the sympathetically derived NPy85
In contrast to the vasodilation produced by CGRP,
SP,NKA, and VIP, NPY produces a potent vasoconstriction
of pulpal blood flow,86 so its release may
counteract the eHect of the vasodilatory neuropeptides.
The release of sympathetically derived
NPY results in an anti-inflammatory effect that may
involve an inhibition of SP release." and therefore
the sympathetic nervous system is involved in the
modulation of pulpal inflarnrnation.? Because VIP is
derived from the parasympathetic nervous system
elsewhere." its presence suggests a possible parasympathetic
innervation of the pulp, although this is
a point of controversy.
In contrast to SP and NKA, which have proinflammatory
effects, VIP, like NPY, is considered an
anti-inflammatory neuropeptide beca use it inhibits
proinflammatory cytokines while upregulating the
anti-inflammatory interleukin 10 (IL-10) cytokine."?
The vasoactive effects of these neuropeptides are
critical to the control of intrapulpal pressure, and,
although involved in healing responses, the vasodilatory
effect of some result in increased intrapulpal
pressures that may lead to ischemia and pulp tissue
degeneration.
An important contribution of neuropeptides to
axonal response to injury is their effect on cytokine
expression. The release of neuropeptides results in
the recruitment of inflammatory cells and the release
of cytokines from these cells that can dramatically
increase the excitability of nociceptors.?? Neuropeptides,
and especially CGRP and SP,promote the
production of cytokines such as tumor necrosis factor
a (TNF-a), interleukin 113(IL-1I3),and interleukin
6 (IL-6) from inflammatory cells'" and from fibroblasts
derived from the human dental pulp." Inflammatory
cells that release SP include neutrophils, plasma
cells, and macrophages located within acute and
chronic periradicular granulomas.93 These findings
further support the emerging important role for
neuroimmune interactions in both inflammatory and
neuropathic pain conditions.?'
An intensive area of research over the last decade
has focused on descriptions of neuropeptide expressions
in normal dental pulp and changes seen in
the diseased and sometimes painful pulp. Many of
these studies have evaluated SP and CGRP.Results
show that tissue levels of SP are increased in teeth
with irreversible pulpitis and a history of spontaneous
pain when compared to normal controls." and
an increased SPexpression in axons that is correlated
with caries progression is significantly greater in painfui
teeth with large caries lesions when compared to
both normal controls and asymptomatic teeth with
caries lesions?" (Fig 7-12). A later study found that
increased axonal expressions of CGRP,SP,VIP, and
NPY are correlated with caries progression in both
primary and permanen! dentitions, but the study did
not evaluate whether these expressions were correlated
with pain."
Other studies have evaluated neuropeptide
expression in painful teeth and found similar increases
in SP,CGRP, NKA, and NPy97and of the receptors
for CGRP98and SP99within human dental pulp.
This upregulation of SP and CGRP receptors occurs
as a gradient because expressions are highest in
pulp tissues from teeth with irreversible pulpitis and
the presence of moderate to severe spontaneous
pain; intermediate expressions are found in pulp
tissues from teeth where inflammation is induced,
and the lowest expressions are identified within
tissues from normal control teeth. In contrast, the
expression of VIP was found to be stable among the
143
Dentallnnervation and Its Responses to Tooth Injury
three qroups.?' This last finding contrasts with the
increased VIP expression mentioned earlier70 and
with another study that found greater VIP expression
in dental tissues from teeth with moderate caries
than in normal teeth and teeth with gross caries,
although there was no indication of presence or
absence of pain among the diseased specirnens.'?'
The dental pulp expression of NPY has also been
found to vary depending on the extent of caries
present: Specimens with mild to moderate caries
express more than specimens with advanced caries,
while normal specimens contain the least." However,
the presence or absence of pain was not indicated
in the study. The NPY expression was also evaluated
with immunocytochemistry; some NPY-positive
axons show co-Iocalization with tyrosine hydroxylase
within sympathetic fibers, while others show colocalization
with sensory fibers that contained SP,
suggesting that this co-Iocalization pattern may
lead to decreased neurogenic inflammation. Other
144
_ ..•6.0 DAsyrnptomauc (n = 11) ;50 * DPain!ul (n = 10)
tn

.~ 40 * Q~* .

1Ij30
'.",;

a. 20 r <.!) a. 'O 10 I r
'~" « O Pulp harn Subadantablastic Midpulp
nerve plexus
9 Pulp region
Fig 7-12 Double-exposure photomicrographs
showing SP immunoreactivity (green) within neural
tissues identilied with PGP 9.5 labeling (red) within
intact (a, e, and e) and carious (b, d, and f) teeth.
Areas with SP immunoreactivity within PGP 9.5-
identilied nerve libers appear yellow. Few PGP 9.5
libers show SP immunoreactivity within the pulp
horn (a), the subodontoblastic region (e), and the
midcoronal region (e) 01 intact teeth, while the
numbers 01 such fibers are increased within the
pulp horn (b), the subodontoblastic region (d), and
the midcoronal region (f) 01 teeth with caries. (g)
Mean percentage area 01 PGP 9.5 labeling that
contained SP immunoreactivity within different
pulpal regions 01 both asymptomatic (apen bars)
and painlul (shaded bars) specimens with gross
caries. SP expression within PGP 9.5-identilied
nerve area was signilicantly greater (asterisk) in the
painlul specimens than in asymptomatic specimens.
(Reprinted Ira m Rodd and Boissonade'" with
permission.)
studies have documented increased expression of
SP following deep cavity preparations'?' and within
inflamed periradicular tissues.?:' increased SP and
IL-1 i3 from crevicular fJuid during orthodontic tooth
rnovement.P" and greater levels of SP and NKA (but
not CGRP) in the crevicular fluid of teeth with pulpal
pain compared to that of healthy teeth.103
Together, the results of these studies document
the increased expression of neuropeptides in teeth
with caries and furthermore identify the involvement
of neuropeptide-containing axons to the increased
innervation density seen within human primary and
permanent dental pulp with caries." An important
finding in the study by Rodd and Boissonade'? was
that increased innervation density does not corre late
with reported pain experience, although their earlier
report did see higher SP axonal expression in painful
teeth with gross caries than in grossly carious teeth
without pain. Certainly the results of these human
studies and the animal studies discussed earlier idenMembrane
Receptors and Ion Channels
tify increased neuropeptide expression as a neuroanatomical
response to injury. Even so, questions
remain concerning their role in pulpal pain mechanisms,
given the braad effects of neuropeptides on
blood flow, inflammation, and tissue healing.
Membrane Receptors and Ion
Channels
Other neuraanatomical responses to injury of the
pulpodentin complex include changes in receptors
and ion channels that control the excitability of
pulpal nociceptors by influencing the development
of generator and action potentials. Important ones
include G-protein-coupled receptors (GPCRs),transient
receptor potential ion channels (TRPs),voltagegated
ion channels, trk receptors, purinoceptors,
and others involved in neuroimmune responses,
such as IL-1, TNF-a, toll-like receptor 4, and CD14. A
role for some in human pulpal pain mechanisms has
been suggested by their presence in dental pulp.'?'
These receptors and channels allow the peripheral
terminals of nociceptors to detect and to respond
to noxious signals in their environment, and changes
in some of these have been seen within axons in the
carious and painful dental pulp. A more detailed
discussion of the rale of these in peripheral odontogenic
pain mechanisms is available elsewherew
The effects of most neuropeptides are mediated
by receptor binding, and many of these are to due
to GPCRs and subsequent activation of specific G
proteins (Ga¡/o' Gaq, and Ga,J and distinct, associated
signaling pathways.'?' For example, somatostatin
and NPY are linked to the Ga¡/o pathway, and activation
of this pathway generally leads to inhibition of
nerve activity, in part by decreasing cyclic adenosine
monophosphate levels. Therefore, an increased
expression of these neuropeptides in nociceptors
would result in decreased nerve activity and an analgesic
effect. Opioids also activate Go. GPCRs, and
this effect is thought to contribute to the analgesic
effect of peripherally administered opioids.
In contrast, the Gas GPCRs increase cyclic adenosine
monophosphate levels and lead to excitation.
The effects of prostaglandins and CGRP are
linked to this pathway, and local increases in CGRP
(mentioned earlier) and the prostaglandin E2 receptor
in periradicular exudates'?' and dental pUlp107
may contribute to the development of odontogenic
pain. The GPCRs coupled to the Gaq signaling
pathway include the SP neuropeptide, bradykinin,
pratease-activated receptors, and endothelin and
leukotriene receptors and leads to activation of
phospholipase C and protein kinase C and a stimulating
effect on nociceptors that includes a sensitization
of the TRPV1 receptor. Increased expression of
bradykinin'P" and CGRP (discussed earlier) in carious
teeth may lead to increased pain levels through this
mechanism.
The TRPs have a critical role in the transduction
of sensory stimuli, including pain and temperature,
so studies that evaluate their pulpal expressions are
important. TRPV1 represents the capsaicin receptor
and is the most intensely studied. TRPV1 activity
is gated by temperature (2 43°C),109active at .
lower temperatures in the presence of inflammatory
mediators."" and critical for the development of
inflammatory hyperalqesla.!" Given this importance
in inflammatory pain, it represents a phenotypic
marker for nociceptive neurons. Pulpal axons contain
TRPV1,112and this expression is greater in both
asymptomatic and symptomatic carious teeth than
it is in normal teeth.!" thus implicating TRPV1 in
both pulpal inflammation and pain. Indeed, the use
of capsaicin to activate TRPV1within human dental
pulp biopsies results in the activation and release
of CGRP from peripheral nociceptors; use of this
methodology appears as a pramising tool to further
evaluate the rale of TRPV1and novel pharmacologic
compounds on human nociceptor sensitivity.!"
Other putative thermoresponsive TRPs include
TRPA1115and TRPM8,116which are implicated in cold
transduction. Althouqh this cold-sensing ability suggests
a possible involvement in the exaggerated
and prolonged pain response that is often provoked
in teeth with pulpitis following the application of a
cold stimulus, TRPM8 may not be involved because
there is less axonal expression of TRPM8 in coldsensitive
and painful teeth than there is in normal
teeth.!" Given the importance of TRP channels to
inflammatory pain mechanisms and the prominent
inflammatory response in the painful dental pulp,
further studies that evaluate the role of TRPs in the
painful human pulp are warranted.
The activation of voltage-gated ion channels is
essential to the formation and propagation of action
potentials and involves calcium, potassium, and
sodium channels. Much recent interest has been
focused on sodium channels because changes in
145
Dentallnnervation and Its Responses to Tooth Injury
Fig 7-13 Conlocal micrograph
01 sodium channel (NaCh) (red)
and Caspr (green) immunoreactivities
within pulpal axons
01 a normal (a) and painlul (b)
molar tooth pulp specimen. The
NaCh antibody used in these
preparations identilies all NaCh
subtypes. Caspr is a paranodal
protein used to identily nodes 01
Ranvier in myelinated libers. The
Caspr staining within the normal
specimen is prominent in the paranodal region 01 myelinated axons (arrowheads), while NaCh staining is located at a high density at the
nodes 01 Ranvier (arrow)
and more unilormly along axons that lack Caspr and that are most likely unmyelinated. The pattern 01 NaCh and Caspr immunoreactivities
changes within axons in the
painlul specimen; the changes include an increase in the size (arrows) and density 01 NaCh accumulations, including some that show
changes in Caspr relationships.
their expression and activation may contribute to
increased neuronal excitability seen in inflammatory
and neuropathic pain conditions.!" The Nav 1.7, -1.8,
and -1.9 sodium channel subtypes are specifically
expressed within the peripheral nervous system and
thus most likely involved in pulpal pain mechanisms.
The overall expression of sodium channels has
been evaluated within the pulp of painful teeth
with large caries lesions and normal teeth with
an antibody that identifies all subtypes; the study
found that sodium channel expression varies among
these different spe cirnens.!" A common finding
in the painful specimens is an augmentation and
remodeling of sodium channels within axons (Fig
7-13). Other studies have found that nerve fiber
expression of the Nav 1.7120 and Nav 1.8121 isoforms is
greater in teeth with pain than it is in normal teeth.
The increase of Nav 1.7 within painful teeth varies
146
Fig 7-14 Conlocal micrographs 01 NaCh (red),
Caspr (qteeti), and myelin basic protein (MBP) (blue)
immunoreactivities within pulpal axons 01 a normal (a)
and painlul (b lo d) dental pulp specimens. Myelinated
libers within the normal dental pulp (a) show prominent
surface staining lor MBP (arrowheads) and NaCh
accumulations at Caspr-identilied typical nodal sites
(arrows). In contrast, painlul specimens (b lo d) show
generalized and local areas 01 decreased MBP staining
(arrowheads) and prominent NaCh accumulations at sites
that lack Caspr (thl« arrows) and at other sites that show
alterations in Caspr relationships (Ihick arrows). These
lindings identily demyelination and the remodeling 01
NaChs at demyelinated sites as common events within
the painlul human dental pulp (bar = 1O prn in a and b;
bar = 5 urn in e, and d). (Reprinted Irom Henry et al"?
with permission.)
depending on location, and the most significant
increase occurs within axons located adjacent to
areas with many inflarnlnatory cells.
Some of these isoforms, such as Nav 1.7 and
Nav 1.8,122 are also are located at nodes of Ranvier,
where changes in their expression in disease states
may contribute to spontaneous activity of myelinated
fibers and the development of sharp, shooting
pain that is characteristic of toothache. Indeed,
a common finding in studies that have examined
sodium channel expression in the painful human
dental pulp is the augmentation of sodium channels
at both intact and remodeling nodal sites that show
a dramatic loss of myelin (Fig 7-14); this finding suggests
the reorganization of ion channels at demyelinated
sites as a pulpal pain mechanism.119,120 Given
the lack of a correlation between pulpal nerve fiber
density and pain levels." pain in teeth may involve
Neurophysio!ogy of Pulpa! Nociceptors and Dentina! Sensitivity
the quality of changes within individual fibers (such
asthe remodeling of ion channels at localized sites),
as influenced by a gradient of inflammation present
within the pulp of teeth with caries lesions.V' rather
than the overall density of nerve fibers.

I Dental Nerve Degeneration

Although much is known concerning the response of
pulpal axons to physical and bacterial insults, other
important questions remain. One of the least understood
is the relationship ofaxon degeneration to
pulpal pain states. Degeneration of pulpal axons in
response to injury is a common finding when painful
pulp tissues are evaluated (see Fig 7-11f). Although
degenerating axons are observed in areas of pulpal
necrosis, they are also commonly found to be intermixed
with intact fibers within painful specimens.
The factors that influence the progression of
the degenerating response are unknown but may
relate to neuroimmune interactions that are prevalent
in the inflamed dental pulp. The presence of
degenerated fibers intermixed with intact ones also
suggests a differential response to injury among various
fiber types, with important implications for pain
mechanisms. Although degeneration ofaxons may
influence pain mechanisms, a more likely process
involves the remodeling ofaxons that occurs before
or in the absence of degeneration. This change in
structure in response to inflammatory influences
most likely involves a remodeling of ion channels
and receptors that could affect the sensitivity and
activity of nociceptors.
This response is further complicated by the gradient
of inflammatory changes that exist within the
diseased dental pUlp123and the effect of this inflammatory
gradient on different regions of the same
axon, with important implications for the development
of intense spontaneous pain that may accompany
the pain of toothache. These changes at individual
sites suggest that pulpal pain mechanisms
may relate not only to broad global changes but
also to the effect of the lesion on isolated fibers. The
painful human dental pulp presents a model system
in which future studies can relate changes at localized
sites to pain states.
Neurophysiology of Pulpal
Nociceptors and Dentinal
Sensitivity
Distinct groups of pulpal afferent nerve fibers can
be classified, as described earlier. The classification
is based on both the morphology and conduction
velocities of the afferents. A number of
recent studies indicate that these neuronal classes
are functionally different and that their activation
may mediate different types of prepain and pain
sensations.!" Generally, these studies indicate that
firing of pulpal afferents in human teeth induces
mostly, if not entirely, painful sensations12S.126 and
that temporal summation (increase of the electrical
stimulation frequency) of low-intensity electrical
stimulation changes the nonpainful (prepain) sensation
to a painful one.!" However, the type of pain
may vary according to the type of stimulus applied,
the type of fibers activated, or the condition of the
pulp. Most studies suggest that rnechanosensitivity"
or therrnosensitivity'A"? of pulpal nerve fibers does
not induce mechanical or thermal perceptions in
people.' although there is recent evidence for intradental
vibration detection by hurnans+" and dental
A¡3-dependent brain activity.!"
Tissue injury and inflammation can sensitize and
activate pulpal afferents. In previous experimental
studies on animals, pulpal inflammation has been
associated with reduced thresholds to external stimulation
and spontaneous discharges of pulpal nerve
fibers.124,132,1T3h3ese changes are probably due to
synthesis or release of' a number of different mediators,
which have been shown to activate pulpal
nerves and sensitize them to external stimuli24,132-134
(see chapter 8).
Application of a cold stimulus to hypersensitive
dentin in human subjects induces pain that, in many
cases, can reach a very high intensity.133,13M5oreover,
patients experiencing acute pulpitis often report
moderate to severe pain.!" However, this is not
invariable: Pulpitis may proceed to a total pulpal
necrosis with only minor symptoms or without any
symptoms at al1.24,13C6onsidering the exceptionally
rich nociceptive innervation of the pulp, such
asymptomatic cases ("silent pulpitis") are puzzling.
However, recent studies indicate that pulpal nociceptor
activation may be abolished by local inhibi-
147
Dentallnnervation and Its Responses to Tooth Injury
tory mediators (eg, local opioids, cannabinoids, or
somatostatin)14,24,1o3r7 by loss of functional terminals
of these fibers (eg, via apoptosis or secondary to
liquefaction necrosis). In addition to these peripheral
factors, other central neural mechanisms may have a
significant impact in the development of dental pain
conditions14,24,65,138(s,1e3e9chapter 8).
Collectively, these studies indicate that there is
a poor correlation between clinical pain symptoms
and the histopathologic status of the pUlp.136,14T0his
is not surprising considering that hyperalgesia is a
perceptual event mediated by peripheral and central
pain mechanisms at the molecular level; these
mechanisms are not necessarily discernible with
microscopes evaluating biopsies of human dental
pulp. In the following sections, the function of the
pulpal neurons in healthy teeth and their responses
to tissue injury and inflammation are described. The
role of different pulpal nerve fiber groups in the
mediation of pulpal and dentinal pain under normal
and pathologic conditions are discussed in the next
two chapters (see chapters 8 and 9).
Sensory functions of pulpal nerves under
normal conditions
A major part of current knowledge regarding the
function of dental nerves is based on electrophysiologic
recordings performed on animals (eg, cats,
dogs, and monkeys). In such experiments, single
intradental nerve fibers are identified and their
responses to various stimuli recorded (Figs 7-15 and
7-16). These electrophysiologic responses to various
external stimuli have been compared to the perceptual
responses induced by the same stimuli applied
to human teeth. Such comparisons have shed light on
how different pulpal nerve fiber groups contribute to
different pain responses under normal and pathologic
conditions. The morphologic similarity of the intradental
innervation of animals and humans serves as a
good basis for such comparisons.
The classification of the pulpal primary afferents
as A and C fibers is based on their conduction
velocities measured in single-nerve fiber recording
experiments142-145(see Figs 7-15 and 7-16). These
two classes correspond to the myelinated and unmyelinated
fibers found in morphologic studies.3,38,146
According to the results of electrophysiologic
recordings, the A and C fibers are functionally differ-
148
ent.1,13,37,127,128,I1n33a,d1d4it7ion, the A-fiber group is
not uniform beca use some slow-conducting (small)
A fibers seem to be sensitive to capsaicin, whereas
most of the faster-conducting fibers respond to
hydrodynamic stimulation but are not activated by
capsaicin.13,37,133
The results of electrophysiologic studies also
indicate that C fibers do not respond to dentinal
hydrodynamic stimulation. Instead, the sensitivity
of dentin is entirely based on the function of intradental
A fibers.135,141,1C4o5mparison of the sensory
responses from stimulated human teeth to the
electrophysiologic responses from animal studies
reveals functional differences between these two
fiber groups in response to tissue injury.l,13,124,133,145,148
As already mentioned, pain and prepain are the
only sensations that can be evoked by intradental
nerve stimulation in human subjects, although there
is recent evidence for intradental vibration perception.
P? The quality of the pain can vary depending
on the type of stimuli applied and can range from
sharp, stabbing pain to dull, aching, throbbing pain
sensations.1,125,134,143,1T4h8e,1v4a9riation is caused by
activation of different nerve fiber types and differences
in the nerve firing patterns (temporal summation)
evoked by various stimuli.127,128,135,149
The application of low-intensity electric stimulation
of human teeth can produce a nonpainful sensation.
125-127It has been proposed that intradental
low-threshold and fast-conducting A~-type afferents
mediate such prepain sensations.l=!" A~ fibers do
have low electric thresholds; however, the thresholds
of A~ and Al) fibers overlap considerably (Fig
7-17). and, accordingly, both fiber groups may be
involved in the rnediation of prepain sensations.!" It
is also important to note that painful sensations can
be induced by increasing the stimulation frequency
at prepain intensities."? a procedure that produces
temporal summation of the nerve activity at the level
of the trigeminal nuclei. Collectively, these findings
suggest that prepain and pain sensations are mediated
by the same afferent fibers.
On the basis of the single-fiber recordings (see
Fig 7-17), it can also be concluded that activation of
onlya small number of pulpal afferents is needed to
evoke prepain or pain sensations.F" This is clinically
important because it suggests that pulp testing may
produce a false-positive response, even in teeth with
extensive pulpal necrosis, as long as at least some
pulpal axons are still responsive. This could explain
Neurophysiology of Pulpal Nociceptors and Dentinal Sensitivity
R
Fig 7-15 Setup lor electrophysiologic recording
01 single intradental nerve libers. The inlerior
alveolar nerve is exposed, and the nerve lilaments
are dissected Irom the nerve trunk. Single libers
innervating the canine or incisor teeth are recorded
using metal wire electrodes (R). The nerve libers are
identilied using electrical stimulation applied to the
tooth crown. (Reprinted Irom Narhi and Hirvonen 141
with permission.)
Fig 7-16 Nerve recording Irom a nerve filament
containing one A and one C liber. The action
potential 01 the A liber shows alter a latency 01
only about 2 milliseconds (ms) after the electrical
stimulus artilact on the lelt (arrow). The C-liber
action potential on the right is delayed by about
30 ms because 01 slow conduction along the axon.
The conduction velocity 01 the recorded liber can be
calculated by dividing the conduction distance (the
length 01 the nerve liber) by the conduction delay.
the clinical observation of a positive pulpal response
in a tooth with a periradicular radiolucency (see also
chapter 17).
It has been suggested that non-noxious mecha nical
(tactile) stimulation of or pressure applied to the
intact tooth crown activates pulpal A¡3 fibers.s,131.150
On the basis of such findings, those fibers were
regarded as a discrete functional group that would
be involved with the regulation of masticatory functions,
the sensation of food texture between the
teeth, and the control of occlusal forces. However,
A¡3 and Aa fibers show similar responses to various
external stimuli and to inflammatory mediators,
124.133,142a-n1d45the results suggest that the fibers
may belong to the same functional group.
Taken together, the results of human and animal
experiments indicate that a hydrodynamic
mechanism mediates intradental nerve activation
in response to several different stimuli1.2.136.147.151-154
(see chapters 8 and 9) as well as release of neuropeptides.
134,155The responding fibers consist of the
Aa and A¡3 classes of neurons (Fig 7-18). Considering
the tissue distortion and injury in the dentin-pulp
150 -(-
-A8-
0.2 0.51 2 5 10 3050 100
Conduction velocity (mis)
Fig 7-17 Electrical thresholds 01 intradental
nerve libers 01 the cat canine tooth plotted against
their conduction velocities. Responses lor C. AS, and
A¡3 libers are shown. The A¡3 and last AS groups
both have very low thresholds compared to the
slower AS libers and C-fiber groups. (Modilied lrom
Nárhi et al124 with permission.)
border related to their actívation.l'" the responding
receptors can be classified as high-threshold mechanoreceptors
or mechanical nociceptors.
The pulpal C fibers are polymodal because they
respond to several different modes of stimulation
and have high thresholds for activation.124,128They
are activated only if stimuli reach their terminal endings
inside the pulp. In. an intact tooth, given the
insulating enamel and dentinal layers, rather intense
thermal stimuli are needed for their activation. The
insensitivity of pulpal C fibers to dentinal (hydrodynamic)
stimulation 124.14is2 consistent with the location
of their endings and receptive fields deep in the
pUlp3,ll,142-145
Pulpal C fibers also respond to histamine and
bradykinin applied to the exposed pUlp13.124(Fig
7-19), which indicates that this fiber group also may
be activated in connection with pulpal inflammatory
reactions. Thus, the dull pain induced by pulpitis
may be evoked by C-fiber activation. C fibers also
respond to capsaicin, which is a selective irritant of
small nociceptive- and neuropeptide-containing
afferents.124.156
149
Dentallnnervation and Its Responses to Tooth Injury
Fig 7-19 Responses of a single intradental C fiber (small
action potential) in the exposed pulp of a cat canine tooth
to bradykinin application (BK) and alter washing with
physiologic saline (NaCI). The A fiber (Iarge action potential)
in the same nerve filament only shows firing 01 a single
action potential at the time 01 the bradykinin application,
probably beca use of a mechanical effect. (Reprinted lrom
Narhi'3 with permission.)
Fig 7-18 Responses of a single intradental A fiber to
probing (a); an air blast (b); application 01 hypertonic,
4.9-moI/L calcium chloride to dentin (e); and drilling 01
dentin (d) over a period of 1.5 seconds. The approximate
timing 01 the stimulus application is indicated by the
horizontal fines in (a), (b), and (d) and by the arrow in (e).
(Reprinted from Nárhi et al'4S with permission.)
Fig 7-20 Responses of intradental nerve libers to intense
heating 01 an intact cat canine tooth. The timing 01 the
stimulus application is indicated by the horizontal line. The
A liber (Iarge action potential) in the lilament gives an
immediate res pon se at the beginning 01 stimulation. In
contrast, activation 01 the C fiber (small action potential) is
much delayed. (Reprinted lrom Narhill with permission.)
The application of intense heating or cooling
to human teeth produces a sharp pain sensation
with a short latency, typically within a few seconds.
If the stimulation is continued, a dull, radiating pain
response followsY28 Correspondingly, biphasic
responses to thermal stimuli are observed in cat
teeth (Fig 7-20). The first response is an immediate
or short-Iatency firing of intradental A fibers, followed
by a delayed C-fiber activation.124,142-1T45he
initial A-fiber responses are supposedly induced by
the dentinal fluid flow resulting from the rapid temperature
changesB6,148,157 The delayed C-fiber acti-
150
vation is probably induced by a direct effect of heat
and cold on the nerve endings in the pUlp.124,128,142
The results of these thermal-stimulation studies
strongly indicate that intradental A and C fibers may
mediate different perceptual qualities of dental pain,
ie, sharp and dull, respectively. In addition, certain
other stimuli, such as air drying of exposed dentin
and application of bradykinin to the exposed pulp,
which are known to activate pulpal A or C fibers
selectively, are also able to induce either sharp or
dull pain, respectively, in human experiments.125,157
Neurophysio!ogy of Pulpa! Nociceptors and Dentina! Sensitivity
Fig 7-21 Activation mechanisms 01 intradental nerve libers.
A libers in the dentin-pulp border area respond to stimulusinduced
Iluid Ilow in the dentinal tubules and consequent
delormation 01 the peripheral pulp tissues containing the nerve
endings (hydrodynamic mechanism). For C-liber activation, the
applied stimuli must reach the nerve endings, which are mostly
located deeper in the pulp. C libers also respond to certain
inllammatory mediators.
Dentinal
fluid flow
A-fiber
activation

;~lllill
Hydrodynamic
<

stimuli~ <
<
Neurophysiologic mechanisms of
dentinal sensitivity
Numerous published studies indicate that the nociceptors
in the dentin-pulp border area are activated
by hydrodynamic fluid flow in response to dentinal
stimulation (the hydrodynamic mechanism).136,15T1he
fluid flow in turn stimulates the nerve endings in the
dentin-pulp border area and causes their activation
(Fig 7-21). Movement of dentinal fluid can also be
induced in unexposed dentin, but in such cases the
capillary forces are not activated and the effect of
the stimulus is much weaker.
The results supporting the hydrodynamic mechanism
of pulpal nerve activation are based both on
in vivo studies on human subjects and experimental
animals and in vitro experiments performed on
extracted teeth. The results of the human experiments
uniformly confirm that patency of the dentinal
tubules is a prerequisite for the sensitivity of
exposed dentin.2,135,158The relationship between
the dentinal tubular condition and dentinal sensitivity
was further confirmed in experiments showing a
significant positive correlation between the degree
of the dentinal sensitivity and the density of open
dentinal tubules counted in exposed cervical dentinal
surfaces in a scanning electron microscopic
replica study on human teeth.l'" In vitro measurements
have also shown that opening or blocking of
the tubules determines the hydraulic conductance of
dentin159and, accordingly, the fluid flow in the dentinal
tubules (see chapter 4).
Several electrophysiologic studies performed on
cats and dogs have shown that acid etching of drilled
• Noxious thermal
stimulation
·Inflammatory ----- C-fiber
mediators activation
dentin significantly increases the responsiveness of
intradental nerves to air blasts, probing, and hyperosmotic
solutionsB.135.142-145,158T,1h6e0i.n1c6r1eased sensitivity
is strongly related to the patency of the dentinal
tubules.F' The sensitizing effect of acid etching can
be abolished almost completely by blocking the
tubules (eg, with oxalates or resin composites). Similar
studies conducted in human teeth indicate that acid
etching increases dentinal sensitivityB5,136
According to the hydrodynamic theory, rapid
dentinal fluid flow serves as the final stimulus activating
intradental nociceptors for many different
types of stimuli. In support of this hypothesis, single
intradental A fibers respond to a number of different
hydrodynamic stimuli, including dentinal probing, air
blasts, and hyperosmotic solutions13,141-14(s5ee Fig
7-19). Studies conducted in vitro demonstrate that
all of these stimuli induce fluid flow in the dentinal
tubules.137.160.I1t6is1the osmotic strength of solutions
and not their chemical composition that elicits pain
responses in human teeth, nerve responses in experimental
animals, and fluid flow responses in dentinal
tubules,26,124.16a2lthough some chemical solutions
may make exceptions.l'" Also, in cold stimulation of
human teeth with open or blocked dentinal tubules,
the intensity of the induced pain does not seem to
be related to the induced fluid flow, but some other
mechanisms of the nerve activation have been suggested
to playa role.l64Much current work is examining
pulp cell responses to dentinal stimulation that
may modulate or contribute in some way to the neurophysiologic
reactions to hydrodynamic force.16,22
Electrophysiologic recordings performed on
cat canine teeth indicate that a direct relationship
exists between dentinal fluid flow and intradental
151
Dentallnnervation and Its Responses to Tooth Injury
nerve activation, and a similar relationship between
induced pain and fluid flow recently has been shown
to exist in human teeth.!= Accordingly, in most cases
nerve activation seems to occur as a response to
the fluid flow, but with certain stimuli (eg, cold and
mechanosensitivity) some other mechanisms may be
activeY28,129When dentin is exposed, inflammation
may develop, leading to sensitization of the intradental
nerves.124Such changes may result in poor
responses to treatment of hypersensitive dentin
and may be significant in teeth with open dentinal
tubules that have been exposed for a long time.
Sensory functions of pulpal nerves under
conditions of pulpal inflammation
As discussed in chapter 8, the two major mechanisms
of pulpal pain are related to dentinal sensitivity
and pulpal inflammation. Injury to the pulp can
alter both of these pain mechanisms. Intense hydrodynamic
stimulation may induce tissue injury in the
dentin-pulp border area, including disruption of the
odontoblastic layer and aspiration of the cells into
the dentinal tubules47,136,152,1T6h6e nerve endings
may also be injured.i'-!"
The inflammation-induced elaboration of growth
factors can lead to subsequent morphologic and
phenotypic changes in the nociceptive nerve endings,
including sprouting and increased neuropeptide
expression=P: these changes may contribute
to long-term functional changes in the pulpal afferents4,133For
example, the local changes in the density
of the innervation in the dentin and pulp might
result in changes in the regional sensitivity of the
affected tooth. However, current knowledge about
the possible functional correlates of the morphologic
changes is limited.
The effect of various inflammatory mediators on
pulpal nerve function has been studied in cat and
dog teeth. These mediators activate intradental
nociceptors and/or sensitize them to subsequent
stimuli (ie, they reduce the threshold for firing)B,124
For example, serotonin activates A fibers and sensitizes
them to external stimulation (eg, hydrodynamic
stimuli).167Intense, repeated heating sensitizes intradental
nerves in cat canine teeth, and prostaglandins
seem to mediate this response.l" As stated earlier,
pulpal C-fiber responses are activated by histamine
and bradykinin, which may be significant for the
development of pain in pulpitis."
152
According to single-fiber recordings, the fasterconducting
pulpal afferents primarily respond to
hydrodynamic stimulation of dentin, although certain
small-diameter myelinated afferents may also
be activated.Fv':" Hydrodynamic stimulation also
affects the pulpal blood flow, indicating that the
nerve fibers activated by such stimulation are able to
induce neurogenic vascular effects.14.168
Pulpal A fibers comprise functionally distinct
classes of sensory neurons. Although most of the
intradental A fibers are activated by hydrodynamic
stimulation, there exists a rather high number of relatively
slow-conducting pulpal Aa fibers that are not
sensitive to hydrodynamic stimulation of the coronal
dentin of healthy teeth37,124,133T,1h3is5class of "silent"
A fibers can be activated only by intense heat or cold
that reaches the pulp proper, and their mechanical
receptive fields are located deep in the pUlp.169
However, the sensitivity of these silent Aa fibers is
enhanced in pulpal inflammation, when they significantly
increase their responsivenessto dentinal stimulation.
l" and they also respond to capsaicin."
Studies to date suggest that there is a functional
significance to the sprouting of sensory terminals
that occurs during inflammation. For exampie,
experiments on dog teeth indicate that nerve
sprouting may be reflected in the size of the receptive
fields of pulpal afferents responsive to hydrodynamic
stimulation of dentin." In healthy teeth, gentle
probing of the exposed dentinal surface revealed
small receptive fields that were most often composed
of a single small spot in the exposed dentin.
133,16In9contrast, gentle probing of exposed dentin
in inflamed teeth revealed a dramatic change,
with emergence of wide receptive fields, sometimes
covering the whole dentinal surface at the crown tip
in inflamed incisors.
This increase in the size of the receptive field
could be caused by sprouting as well as activation
of normally silent terminals of branched axons. An
increase in the size of receptive fields would result in
an increased overlap of receptive fields and, accordingly,
would enhance spatial summation of peripheral
nerve activity, increasing pain intensity in response
to dentinal stirnulation."?
Inflammation may also increase the regional sensitivity
of dentin in various parts of the tooth. In normal
dog teeth, the nerve fibers innervating the cervical
dentin are far less responsive to hydrodynamic
dentinal stimulation than are those innervating dentin
in the crown tipo However, in inflamed teeth the
Conclusion
sensitivity of cervical dentin can increase to the same
level as that of the crown tip.169,170
Although most inflammatory mediators activate
or sensitize peripheral neurons, some mediators
released in pulp after injury, including endogenous
opioids and somatostatin, appear to be inhibitory.
In experiments performed on inflamed dog teeth,
the local application of a somatostatin antagonist
increased firing of intradental nerves, suggesting
that the release of endogenous somatostatin reduces
firing during injury.133 In other preliminary experiments,
administration of the opioid antagonist naloxone
produced a similar effect.133 In addition, local
application of morphine in deep cavities completely
abolished the pulpal nerve responses to mustard oil,
a substance that induces inflammation and activates
nociceptive afferents.
These results suggest that in pulpal inflammation
both somatostatin and endogenous opioids effectively
reduce or abolish intradental nerve activity,
despite the presence of other inflammatory mediators
that have a stimulatory effect. These data suggest
that one possible mechanism for the frequently
reported lack of clinical symptoms in teeth with
pulpal inflammation may be based in part on the
release of local inhibitory mediators in the inflamed
tissue.

I Conclusion
Knowledge gained over the past two decades has
greatly increased the current understanding about
peripheral mechanisms of tooth sensitivity and pain,
including dentinal innervation and its sensitivity,
neurophysiology of pulpal nociceptors, sensory neuropeptides
that affect pulp cells, vasoregulation
by dental sensory and sympathetic fibers, sympathetic
interactions with immune cells, responses
of dental nerves to injury, the role of local factors
such as growth factors or inflammatory mediators
in modulating neural function, and the relationship
of these different features to clinical dental pain.
However, there is still more to be learned about the
types of nerve fibers in teeth, their functional shifts
in response to inflammation and injury, and neuropulpal
and neuro-odontoblastic interactions.
Recent advances with studies of human teeth in
vitro provide additional possibilities for odontoblast
and neural functions, along with possible interactions
of those cells in relation to tooth pain.171,172
Animal studies of odontoblast ion channels further
complicate the story by revealing at least eight different
odontoblast phenotypes, most of which do
not overlap with sensory innervation terrninations.!"
and those findings are consistent with other tissues
in which local cells utilize "neura!" genes for their
own tissue responsibilities as well as for modulation
of neural functions.174,175 Many evolving paracrine
communications between odontoblasts and
their neighboring cells continue to be identified,
including purinergic neural detection of adenosine
triphosphate release from pulp cells.'?> In addition,
recently evolving technology has enabled better dissection
and identification of cell types and functions
within the odontoblast layer of mature human teeth,
with fascinating suggestions about dental mecha noreceptor
mechanisms."?
The pace of discovery in this field suggests that
new clinical insights will be developed soon concerning
the peripheral mechanisms of dental pain
and anesthesia, diagnostic aspects of dental pain,
and the treatment of hypersensitive teeth. Some
important unresolved questions concern the different
mechanisms responsible for the transformation
of a mild toothache to asevere one that forces the
patient to seek immediate therapy. These differences
may relate to important influences provided
by the immune response on axons and the glia that
invest them and the remodeling of ion channels and
receptors within individual axons that allow them to
detect noxious signals and that control axonal excitability
and activity.
The pain attributes of individual toothaches vary,
and this variation involves the axon response to
injury and the activation of a different mix of pain
mechanisms that interact with one another to form a
unique fingerprint associated with each pain experience.
Additional study of the human dental pulp
should provide important insights into the axon
response to injury and how these relate to both
healing and pain. The puzzle of human pulpal pain
remains, but, given that endodontic therapy is highly
successful in the treatment of pulpitis, part of the
answer most likely resides in the pulp, not only
beca use infection has been arrested and removed
but also because dental pain mechanisms include
neuropulpal interactions that are only beginning to
be understood.
153