doi:10.1111/iej.
13400
REVIEW
Dental nerves: a neglected mediator of pulpitis
C. Zhan, M. Huang, X. Yang & J. Hou
Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou, China
Abstract
Zhan C, Huang M, Yang X, Hou J. Dental nerves: a
neglected mediator of pulpitis. International Endodontic
Journal, 54, 85–99, 2021.
As one of the most densely innervated tissues, the den-
tal pulp contains abundant nerve fibres, including sen-
sory, sympathetic and parasympathetic nerve fibres.
Studies in animal models and human patients with
pulpitis have revealed distinct alterations in protein
expression and histological appearance in all types of
dental nerve fibres. Various molecules secreted by neu-
rons, such as classical neurotransmitters, neuropep-
tides and amino acids, not only contribute to the
induction, sensitization and maintenance of tooth pain,
but also regulate non-neuronal cells, including
Introduction
Dental pulp is a peripheral end-organ without collat-
eral nerves, and its dense innervation has been the
subject of extensive research throughout the last cen-
tury. The major function of dental nerves is thought
to be the transmission of afferent signals to the cen-
tral nervous system (CNS), which ultimately triggers
various responses. More specifically, the pathway for
signal transmission in the dental pulp starts with the
detection of stimuli by primary afferent fibres. The
first-order neurons in trigeminal ganglion project cen-
tral terminals to the neurons that are located within
the brainstem; these signals are then sent to the tha-
lamus, and the somatosensory cortex finally receives
the thalamic inputs (Barnett et al. 1995). Almost all
stimuli applied to the dental pulp result in the
Correspondence: Jin Hou, Department of Stomatology, Nanfang Hospital, Southern Medical University, Guangzhou Da Dao Bei
1838, 510515, Guangzhou, China (fax: 0086-20-62787679; e-mail: houjin@smu.edu.cn).
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd
fibroblasts, odontoblasts, immune cells and vascular
endothelial cells. Dental nerves are particularly impor-
tant for the microcirculatory and immune responses in
pulpitis via their release of a variety of functional sub-
stances. Further, nerve fibres are found to be involved
in dental soft and hard tissue repair. Thus, understand-
ing how dental nerves participate in pulpitis could
have important clinical ramifications for endodontic
treatment. In this review, the roles of dental nerves in
regulating pulpal inflammatory processes are high-
lighted and their implications for future research on
this topic are discussed.
Keywords: dental nerve, immune system, inflam-
mation, peripheral nerve, pulpitis.
Received 14 June 2020; accepted 26 August 2020
sensation of pain (Hossain et al. 2019), which as one
of the most commonly reported symptoms, can vary
in intensity. Whilst several previous studies have
focused on exploring the cellular and molecular
mechanisms underlying dental pain, the role of the
dental nerves as a mediator of inflammation has been
neglected. Over the past two decades, the traditional
concept that the dental nerve merely serves a sensory
function has been challenged by numerous studies,
which have provided evidence for its complex regula-
tory roles, especially in pulpitis (Haug & Heyeraas
2006, Caviedes-Bucheli et al. 2008a,b).
Pulpitis is one of the most common oral diseases
and is often associated with severe dental pain. In
pulpitis, dental nerve fibres are not only responsible
for transmitting pain impulses (Hossain et al. 2019)
but have also been implicated in regulating
International Endodontic Journal, 54, 85–99, 2021 85
 Dental nerves in pulpitis Zhan et al.
inflammatory responses. Several studies have shown
that at least two different branches of the peripheral
nervous system are stimulated during pulpal inflam-
mation: the sensory nerve fibres, which act mainly in
a pro-inflammatory manner, and the sympathetic
nerve fibres, which may have anti-inflammatory
effects. These regulatory functions are accomplished
through the actions of various molecules secreted by
nerve endings, such as classical neurotransmitters,
neuropeptides and excitatory amino acids. Under
physiological conditions, there exists a dynamic equi-
librium state that can be disrupted by a variety of
pathological processes, such as the production and
release of bacterial virulence factors as well as inflam-
matory factors, thermal stimuli, electrical stimuli and
so on. Under these conditions, nerve fibres begin to
release functional molecules to regulate the vascular
system and immune system in addition to transmis-
sion of pain sensation. Based on these recent findings,
an in-depth understanding of this neglected role of
dental nerves will provide novel insights for the devel-
opment of strategies to explore their therapeutic
potential in the prevention of pulpal inflammation.
The present review attempts to provide a summary
of the current studies on the regulatory functions of
dental nerves in pulpitis, with emphasis on the regu-
lation of vascular function, immune response and tis-
sue repair activities.
Innervation of normal pulp
The dental pulp is a peripheral end-organ without
collateral nerves. The majority of the axons enter the
pulp cavity through the apical foramen. Based on
observations under electron microscopy, unmyeli-
nated axons (C-fibres) account for about 80–90% of
nerve fibres entering teeth (Nair 1995). Thinly myeli-
nated fibres (Ad-fibres) and a very small amount of
thickly myelinated fibres (Ab-fibres) constitute the
remaining portion of axons (Nair 1995), and myelin
of myelinated fibres is formed by Schwann cells in the
peripheral nervous system. Previous studies have
shown that many unmyelinated axons express the
neurofilament heavy, a marker for myelinated axons,
within the dental pulp, suggesting that these axons
arise from parent axons that are myelinated at proxi-
mal sites (Henry et al. 2012). Similar to most of the
peripheral nerves, dental nerves are mixed nerves that
contain sensory (afferent) fibres and sympathetic
fibres originating from the trigeminal ganglion and
cervical sympathetic ganglion, respectively (Qian &
86 International Endodontic Journal, 54, 85–99, 2021
Naftel 1994, Ibuki et al. 1996). A portion of sympa-
thetic nerves leaves the superior cervical ganglion
and runs into dental pulp along with the superior or
inferior alveolar artery. The other part runs towards
the trigeminal ganglion first and then enters the pulp
with sensory nerve fibres (Wakisaka 1990). An over-
whelming majority of unmyelinated axons are sen-
sory fibres, and very few unmyelinated axons are
sympathetic fibres. However, controversy persists
regarding the parasympathetic innervation of the
human dental pulp (Caviedes-Bucheli et al. 2008a,b).
Both sensory and sympathetic axons accompany
blood vessels and even encircle them. Whilst the
axons emit a few branches in the radicular portion of
the pulp, they arborize branch extensively in the
coronal pulp to form the subodontoblastic plexus of
Raschkow (Hossain et al. 2019). Of note, only the
sensory axons enter dentinal tubules and penetrate
the predentin/dentine (Ibuki et al. 1996), whereas
sympathetic axons end in close proximity to odonto-
blast cell bodies (Shimeno et al. 2008). At present, a
clear interaction between dental nerves and odonto-
blasts is not yet established. However, given their
close anatomical relationship, there may exist bidirec-
tional cross-talks between dental nerves and odonto-
blasts. This hypothesis is also supported by the fact
that odontoblasts express membrane-bound receptors
for functional molecules from dental nerves (Kido
et al. 2005). In addition, the release of ATP by odon-
toblast cells in response to noxious stimulation and
the substantial expression of P2X receptors in dental
nerves suggest that ATP might also act as a sig-
nalling molecule between odontoblast cells and neu-
rons. Such transmission of signals supports the
odontoblastic receptor theory of dentine sensitivity
(Sol
e-Magdalena et al. 2018).
Neural changes in pulpitis
Normal tooth development, maturation and ageing
result in neural changes (Sarram et al. 1997, Mahdee
et al. 2019), but the most typical and well-studied
changes are observed in pulpitis. Dental pulp fibrob-
lasts that are localized just beneath the carious lesion
can secrete nerve growth factors (NGF) and thereby
promote neurite outgrowth towards the injured site
(Chmilewsky et al. 2016, 2018). Nerve terminals
begin to sprout new processes within the pulp horn
region in response to carious lesions. Even intrapulpal
abscesses, when developed, are found to be sur-
rounded by nerve sprouts (Rodd & Boissonade 2001).
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd

In the apical pulp of human teeth with irreversible
pulpitis, myelinated nerves display moderate-to-severe
degenerative changes. These include granular degen-
eration, peeling degeneration, myelin sheath disinte-
gration and vacuolization (Mendoza et al. 1987,
Pisßkin et al. 1993). In addition, some axons of myeli-
nated and unmyelinated fibres appear shrunken and
display increases in the number and size of mitochon-
dria (Mendoza et al. 1987, Pisßkin et al. 1993).
Together, sprouting and degeneration of nerve fibres
are primary events that occur in pulpitis, and they
can either increase or decrease the innervation den-
sity of the dental pulp, dynamically regulating by
NGF (Byers 1994, Minnone et al. 2017). NGF, as a
homodimeric peptide, can promote neuronal differen-
tiation of human dental pulp stem cells via ERK and
AKT signalling pathway (Zhang et al. 2017, Darabi
et al. 2019). The up-regulation of pulpal NGF in pul-
pitis has been shown to induce increased innervation
density of nerve fibres, which will subside once heal-
ing occurs, but continue in states of persistent inflam-
mation (Byers 1994). However, at the most severe
levels of injury, tissue damages caused by the inflam-
matory response would overwhelm the advantage
taken from an increased expression of NGF, which
results in decreased innervation density within the
dental pulp.
The expression levels of various ion channels also
change during pulpitis. Many previous results have
shown that unmyelinated and demyelinated sensory
neurons exhibit altered expression of voltage-gated
sodium channel (Nav) in pulpitis, and these changes
are implicated in the generation of pain states (Luo
et al. 2008, 2010). The Nav1.6 isoform, a major form
located at nodes of Ranvier, maintains its expression
in pulpitis, and this stable expression is accompanied
by increased expression of other isoforms, such as
Nav1.7, Nav1.8 and Nav1.9 (Renton et al. 2005, Luo
et al. 2008, 2010, Suwanchai et al. 2012). Further,
the numbers of these channels are also significantly
elevated in the trigeminal ganglion (Wells et al.
2007a,b). On the contrary, the expression of Kv1.4, a
potassium channel subunit contributing to the sup-
pression of action potential generation, is decreased in
neurons of painful human pulp (Wells et al. 2007).
All of these changes participate in the generation and
maintenance of hyperalgesia in pulpitis.
In addition to the above two ion channels, abnor-
mal cellular location and altered expression of tran-
sient receptor potential (TRP) channels also occur in
pulpitis. TRP channels are a large family of
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd
Zhan et al. Dental nerves in pulpitis
nonselective cation channels involved in sensory sys-
tems that detect and respond to endogenous and envi-
ronmental stimuli (temperature, mechanical forces
and chemical stimuli) and the transmission of pain
signals in pulpitis (Caterina & Julius 2001). In pulpi-
tis, inflammatory mediators, such as lipopolysaccha-
ride (LPS), can induce post-translational modifications
to TRP channel proteins, thereby increasing their
expression in nerve fibres and finally leading to a
hyperalgesic state (Morgan et al. 2005, Hossain et al.
2019).
Intriguingly, apart from local neural changes in
inflamed pulps, distant neural cells in the ipsilateral
trigeminal ganglion and even the brainstem also dis-
play distinct pathological changes following pulp
injury (Tønder 1983, Matsuura et al. 2013, Lee et al.
2017). Indeed, nerve fibres lack protein synthesis
machinery, and the axonal transport (from cell body
to axon) supplies materials consumed in the axon
(Okada et al. 1995). Whilst the cell body receives sig-
nals from axons that results in changes in protein
expression level in the cell body, the level of neural
molecules in the nerve terminal change accordingly
through anterograde axonal transport. Overall, the
dental nerve fibres within the pulp are cytoplasmic
extensions of neurons whose cell bodies are located in
the ganglion; it is not these fibres but neurons them-
selves that receive stimuli and regulate the inflamma-
tory response in the pulp tissue.
Neurosecretion in pulpitis
Classical neurotransmitters and neuropeptides, acting
as functional molecules, appear to play important
roles in mediating pulpal inflammation. In normal
pulps, expression of substance P (SP), neurokinin A
(NKA), calcitonin gene-related peptide (CGRP), neu-
ropeptide Y (NPY), vasoactive intestinal polypeptide
(VIP) and secretoneurin (SN) can be detected (Awaw-
deh et al. 2002, Caviedes-Bucheli et al. 2004, 2006,
Steiner et al. 2018). However, it should be noted that
the expression level of neuropeptides is low in clini-
cally normal pulps (Caviedes-Bucheli et al. 2004).
These short-chain peptides (approximately 4–40
amino acids long) are mainly produced in the neuron
cell body in the ganglion and are transported via axo-
nal flow to the nerve endings in the dental pulp
(Holmgren & Jensen 2001). Intriguingly, non-neural
cells including dental pulp fibroblasts can also express
neuropeptides (Killough et al. 2009). Once pulpitis
has occurred, the inflammatory environment leads to
International Endodontic Journal, 54, 85–99, 2021 87
 Dental nerves in pulpitis Zhan et al.
the production of greater amounts of neuropeptides in
pulps, except for VIP and SN levels, which remain
practically unaltered (Awawdeh et al. 2002, Bowles
et al. 2003a,b, Caviedes-Bucheli et al. 2004, 2006,
Steiner et al. 2018).
Given that neural functional molecules should be
secreted to the outside of cells prior to binding with
their receptors located on the cell surface membrane,
it is crucial to understand the secretion mechanisms
involved. Classical neurotransmitters are packaged in
small clear vesicles (SCVs), which can be supplied via
endocytic pathways. In contrast, secretory proteins
are produced in the cell body and condensed in large
dense core vesicles (LDCVs), which initially form at
the trans-Golgi network and then are transported to
the vicinity of the plasma membrane where they wait
to be secreted (Xu & Xu 2008). Although different
molecular players are involved in SCV and LDCV
release (Xu & Xu 2008), the exocytosis of both shares
conserved features and depends on a localized
increase in intracellular Ca2+ concentration (S€ udhof
2012). Taking neuropeptides as an example, whilst
the rate of basal release of peptides is low (Iversen
et al. 1980), enhanced activation of both voltage- and
ligand-gated Ca2+ channels can trigger the release of
neuropeptide (Wang et al. 2017). Thus, it seems that
thermal, chemical and electric stimuli of sufficient
intensity to dental nerves would lead to an increased
secretion of functional molecules. However, the
release of neuropeptides is highly variable between
different individuals even under the same stimulating
conditions (Burns et al. 2016). It is worth noting that
this increased release might not be the only cause of
high extracellular levels of neuropeptides in pulpitis
(Bowles et al. 2003a,b). It is also possible that this
phenomenon could be the result of increased synthe-
sis of neuropeptides in the ganglion or decreased
levels of peptidases.
There is overwhelming evidence that various resi-
dent cell types within pulp, such as fibroblasts,
odontoblasts, inflammatory cells and the endothelial
cells of capillaries and post-capillary venules, can
express membrane-bound receptors for sensory and
sympathetic neuropeptides, allowing these cells to
respond to neuronal stimuli (Fristad et al. 1999,
2003, Kido et al. 2005, El Karim et al. 2008, Kil-
lough et al. 2010). Notably, in the presence of
inflammation, the expression of these receptors
increases significantly (Caviedes-Bucheli et al. 2005,
Rethnam et al. 2010), which indicates an increased
response of neuropeptides.
88 International Endodontic Journal, 54, 85–99, 2021
Neuromodulation in pulpitis
During pulpitis, the density of innervation of the den-
tal pulp is altered and the balance amongst sensory,
sympathetic and parasympathetic nerves is disrupted
at the site of inflammation. These changes corre-
spondingly affect the inflammatory response in the
pulp tissue and facilitate tissue repair (Fig. 1). Byers
& Taylor (1993) found that the pulpal survival rate
was lower in denervated rat mandibular molars than
in contralateral innervated teeth. More specifically,
the denervated teeth exhibit more rapid progression
of pulpitis as well as more extensive pulp necrosis
than the innervated teeth, although denervation pro-
vides rapid and effective pain relief in experimental
animals. Therefore, a better understanding of the
mediator role of dental nerves will yield insights for
the development of novel control strategies for pulpal
diseases.
Regulation of blood vessels in pulpitis
The microcirculation is essential for the preservation
of pulp functions: first by providing optimal nutrition
to the tissue as well as removing metabolites and
waste products, and secondly, for maintaining suffi-
cient blood pressure within the vascular system of the
pulp that is in harmony with the interstitial fluid
pressure. The pulpal vasculature constantly changes
under varying conditions such as growth, maturation
and especially inflammation throughout life. It has
been shown that increased blood flow and vascular
permeability occur in the vascular system in the first
stage of inflammation, followed by fluid exudation
and leucocyte migration in the venular network (Tøn-
der 1983, Takahashi 1990), thereby maintaining an
adequate supply of oxygen and nutrients to the com-
promised tissue.
Within human pulps, about 20% of pulpal blood
vessels have definite intersections with nerve fibres
(Rodd & Boissonade 2003), strongly indicating a
functional relationship exists between the microvascu-
lature and dental nerves. A rat experiment demon-
strated that electrical stimulation of the incisor or
molar promptly increased pulpal blood flow, whereas
denervation of the inferior alveolar nerve diminishes
blood flow upon stimulation (Kerezoudis et al. 1995a,
b, Fristad et al. 1997). In addition, intra-arterial
administration of SP and CGRP antagonists also abol-
ish pulp vasodilation following nerve stimulation
(Berggreen & Heyeraas 2000), whereas intravenous
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd
 Zhan et al. Dental nerves in pulpitis

Figure 1 Imbalance of dental nerves in pulpitis. Pulpal sensory, sympathetic and parasympathetic nerve fibres play important roles
in regulating the vasculature and immune response in pulpitis via communication with relevant cells through neuron-secreted fac-
tors. Vasodilator function of neuropeptides and glutamate overwhelm the vasoconstrictor function of sympathetic nerves in pulpi-
tis. Meanwhile, to some extent, acetylcholine and vasoactive intestinal polypeptide secreted by parasympathetic nerve fibres also
lead to vasodilation and increased vascular permeability. Substance P (SP) promotes an immune reaction, resulting in a pro-inflam-
matory response, whilst neuronal signalling molecules from sympathetic nerve fibres have an immune-suppressive function. Calci-
tonin gene-related peptide (CGRP) may either promote migration of immune cells or be involved in immunosuppression. Various
molecules, including SP, neurokinin A, CGRP and sonic hedgehog (Shh), released from sensory nerves participate in tissue repair
activity. In contrast, catecholamine decreases tertiary dentine formation.

injection of SP can enhance vasodilation in dener-
vated pulps (Olgart et al. 1993). A laboratory study
reported that SP could exert its vasodilation effect
partly through production of a key endogenous
vasodilator, nitric oxide (NO), from endothelial cells
(Karabucak et al. 2005). However, NO synthase inhi-
bitor potentiates SP-induced vasodilatation in vivo.
This unexpected result may be due to a negative feed-
back mechanism, which needs to be verified by fur-
ther experiments (Hsu et al. 2003).
Local application of SP or CGRP can increase dental
pulp vascular permeability, as confirmed by the Evans
Blue dye method, and CGRP is more active than SP
(Maltos et al. 2004). However, another study reported
that SP, but not CGRP, is involved in neurogenic
plasma extravasation (Kerezoudis et al. 1994). Consid-
ering the variable gene expression patterns of nerve
cells in different contexts, the discrepancy between
the above studies may be attributed to different
experimental schemes (exposure of pulp vs. electrical
stimulation).
Neuropeptides from sensory nerves are not the only
neuromodulators that can induce vascular changes.
Glutamate, a type of excitatory amino acid, can also
elicit vasodilation. This excitatory amino acid can be
released from neurons expressing the glutamate trans-
porter in the pulp (Zerari-Mailly et al. 2012) and act
through metabotropic glutamate receptors (GluRs),
but not ionotropic GluRs, to regulate pulpal blood
flow (Hofman et al. 2002). However, the specific types
of nerve fibres expressing this transporter and the
localization of these receptors remain uninvestigated.
Although GluRs have been found in odontoblasts
(Nishiyama et al. 2016), it is unclear whether other
cell types in the pulp, especially endothelial cells,
express GluRs.
The role of parasympathetic fibres in the control of
pupal vasodilation has been a controversial issue. A

© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd International Endodontic Journal, 54, 85–99, 2021 89
 Dental nerves in pulpitis Zhan et al.
laboratory study of isolated pig pulpal arterioles
revealed that acetylcholine can induce endothelium-
dependent vasodilation via atropine-sensitive mus-
carinic receptors (Yu et al. 2001). By contrast, in an
in vivo study, the parasympathetic vasodilator func-
tion was not observed (Sasano et al. 1995). It seems
that the influence of cholinergic modulation on pulpal
microcirculation is weak. Additionally, there was no
significant increase in VIP expression during the
inflammatory phenomenon (Caviedes-Bucheli et al.
2006). This finding confirms a weak influence of
parasympathetic fibres in pulpitis.
Post-ganglionic sympathetic fibres and trigeminal
sensory fibres have opposing effects on pulpal blood
flow. There is evidence that the effects of sympathetic
nerve activation on blood flow are mediated primarily
through classical neurotransmitter catecholamines
and NPY. Like adrenoceptor, NPY receptor is also
located on the walls of blood vessels in the human
pulp tissue (El Karim et al. 2008). In addition, it has
been reported that the majority of NPY-immunoreac-
tive fibres encircling pulpal blood vessels are identical
to adrenergic ones (Uddman et al. 1984, El Karim
et al. 2006). An electrical stimulation of sympathetic
nerve causes vasoconstriction and reduction in pulpal
blood flow (Kim et al. 1996). Alpha-adrenoceptor
antagonists, phenoxybenzamine or phentolamine, and
NPY antagonist, D-myo-inositol-1,2,6-trisphosphate,
can attenuate the reduction in pulpal blood flow
caused by electrical stimulation (Edwall et al. 1985,
Kim et al. 1996). Furthermore, stimulation of sympa-
thetic nerves inhibits sensory nerve-induced plasma
extravasation (Kerezoudis et al. 1993). The vasocon-
strictor function of norepinephrine is realized in both
direct (vasoconstriction) and indirect (inhibiting
vasodilatation function of sensory nerves) ways. The
latter is achieved through inhibiting the release of
vasodilator from trigeminal sensory fibres. For exam-
ple, the application of a-adrenoceptor or b2-adreno-
ceptor agonists in the dental pulp significantly blocks
exocytosis of CGRP from pulpal sensory neurons. This
effect can be blocked by the a-adrenoceptor antago-
nists (phentolamine or phenoxybenzamine) and b2-
adrenoceptor antagonist (ICI 118551; Bowles et al.
2003a,b, Hargreaves et al. 2003a,b). These studies
suggest that sympathetic nerves can modulate the
function of sensory nerves in the regulation of blood
vessels in pulpitis through the inhibition of exocytosis
from sensory nerves. In contrast, to date, no study
has identified functional molecules from sensory
nerves that can inhibit the secretion of
90 International Endodontic Journal, 54, 85–99, 2021
catecholamines and NPY from autonomic neurons
innervating dental pulp.
In pulpal inflammation, the state of microcircula-
tion, in turn, affects nerve activity. Given that the
pulp is encased in rigid dentinal walls, increases in
pulpal blood flow and vascular permeability will tend
to increase intrapulpal pressure in the interstitial fluid
surrounding dental nerves and cause an excitation
effect on nerves in this low compliance environment
(Tønder 1983). It is likely that when the tissue pres-
sure exceeds a critical pressure threshold, the periph-
eral nerve is rapidly damaged, followed by a
progressive loss of neural function (Gelberman et al.
1983).
Regulation of immune reaction in pulpitis
In the para-odontoblastic region of the normal dental
pulp, class II antigen-expressing (OX6+) dendritic cells
(DCs) have much close positional association with
nerve fibres (Okiji et al. 1997). When untreated caries
progress into dentine, DCs and nerve fibres gather
together promptly along the pulp–dentine border cor-
responding to the area of carious dentine, instead of
noncarious regions of the same teeth (Sakurai et al.
1999). Of note, pattern recognition receptors, Toll-like
receptor (TLR)-4 and CD14, have been identified on
myelinated nerve fibres in human dental pulps
(Wadachi & Hargreaves 2006). This means that den-
tal nerves, similar to DCs, may represent a substantial
immune surveillance component, and the positional
similarity of dental nerve endings to DCs may par-
tially result from functional resemblance between
them, all of which can potentiate neuro-immune
paracrine interactions during the initial stage of the
carious pulpitis. Besides, activation of TLR-4 through
LPS from microorganisms causes sensitization of other
receptors, such as TRP vanilloid 1 (TRPV1), in
trigeminal sensory neurons (Diogenes et al. 2011, Fer-
raz et al. 2011). This response reduces the threshold
of neuronal activation, leading to a hyperalgesic
state.
As described before, neuropeptides take part in reg-
ulating the local microcirculation in pulpitis.
Increases in blood flow and vascular permeability
may serve as prerequisites for promoting and guiding
immune cells to local tissues. Also, it has been shown
that the rat denervated molar pulps exhibit consider-
ably lower recruitment of immunocompetent cells in
the inflammatory pulpal tissue subjacent to artificial
dentinal cavities than the innervated teeth (Fristad
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd

et al. 1995a,b). In addition, numerous receptors for
these neuronal signalling molecules exist on immune
cells, such as monocytes/macrophages (Ho et al.
1997), indicating that dental nerves are directly
involved in the recruitment of immune cells. More-
over, cell experiments confirmed the ability of SP and
CGRP to chemoattract human monocytes and T lym-
phocytes, respectively (Ruff et al. 1985, Foster et al.
1992). Taken together, these findings indicate that
neuropeptides promote chemotaxis of immune cells
into dental pulps by increasing vascular permeability
and directly acting on immune cells. Notably, the
number of immunocompetent cells is almost unaf-
fected in the denervated pulp in the absence of injury
stimulus (Fristad et al. 1995a,b). Also, considering the
fact that the production and secretion of neuropep-
tides are significantly low in healthy pulps (Caviedes-
Bucheli et al. 2006), it could be speculated that dental
nerves have little effects on the number of immune
cells in healthy dental pulps. It could be speculated
that nerve activity is not the main factor maintaining
the number of immune cells in healthy dental pulps,
because the production and secretion of the peptides
is not sufficient to affect immune cells under normal
conditions. Periapical lesions often occur as a conse-
quence of pulpal inflammation. Thus, functional inter-
actions between the immune and nervous systems
may also play roles in the progression of apical peri-
odontitis (Austah et al. 2016). For instance, increased
infiltration of OX6+ cells as well as a close association
between OX6+ cells and nerve fibres was detected
around the apex or in the periphery of lesions during
active lesion expansion and stabilization (Yang et al.
2007).
In addition to their capability of attracting
immunocompetent cells towards the inflammatory
site, neuropeptides also play an important role in reg-
ulating immunomodulation by affecting immune cell
proliferation and function. SP is a major immune-ac-
tive substance, due to the fact that it can stimulate
the release of interleukin (IL)-1, IL-6, IL-12 and
tumour necrosis factor (TNF)-a from monocytes/
macrophages (Lotz et al. 1988, Kincy-Cain & Bost
1997). It has been shown that SP activates the mast-
cell-specific receptor Mas-related G-protein-coupled
receptor MRGPRX2, instead of a canonical receptor,
resulting in the release of TNF-a, IL-8, CCL2, CCL3,
etc. (Green et al. 2019). These pro-inflammatory
cytokines are key mediators involved in the inflamma-
tory response. Additionally, a large number of labora-
tory experiments have proven that SP can stimulate
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd
Zhan et al. Dental nerves in pulpitis
the production of pro-inflammatory cytokines in
human dental pulp fibroblasts. So far, IL-1b, IL-6, IL-
8 and TNF-a have been observed at the physiological
level of SP stimulation (Bowles et al. 2003a,b, Park
et al. 2004, Yamaguchi et al. 2004). However, con-
troversy still exists regarding the role of CGRP in
mediating immune responses, even though more
studies have suggested that CGRP has an immuno-
suppression function (Holzmann 2013). In pulpitis,
CGRP signalling can control the anti-inflammatory
macrophage phenotype (M2) to regulate the immune
response (Caviedes-Bucheli et al. 2008a,b). In addi-
tion, pulpal Schwann cells also help induce M2 phe-
notypic polarization of macrophages, and abundant
colocalization of Schwann cells and M2 macrophages
can be detected under the lesion in active and deep
caries with no episodes of spontaneous pain, but faint
sensitivity to air (Yoshiba et al. 2020).
Under physiological conditions, it is apparent that
NPY fibres encircle blood vessels (El Karim et al.
2006), whereas, in pathological conditions, NPY has
no visible association with blood vessels (El Karim
et al. 2006). Additionally, NPY receptors are located
in inflammatory cells (El Karim et al. 2008). These
findings provide clues that sympathetic nerves could
have a modulatory role, in addition to inducing vaso-
constriction, during pulpitis. For example, activation
of sympathetic nerves promotes recruitment of CD43+
polymorphonuclear cells (Csillag et al. 2004). More-
over, sympathectomy decreases the occurrence of
abscess formation 4 days after cavity preparation
(Haug et al. 2001) but has no influence on the degree
of the pulpal inflammation when the inflammation is
extensive (Haug & Heyeraas 2003).
Paradoxically, unlike the inferior alveolar nerve axo-
tomy (complete loss of sensory nerve fibres and partial
loss of sympathetic nerve fibres) mentioned above,
sympathectomy (only loss of sympathetic nerve fibres)
has an impact on pulpal immunity under noninflam-
matory conditions. Previous research showed that
sympathectomy can stimulate migration and prolifera-
tion of immunoglobulin-producing cells, such as
plasma cells (Haug & Heyeraas 2005). Two possible
factors may explain this discrepancy: first, the differ-
ence in observation time (18 days vs. 6 days). In the
denervation experiments (Fristad et al. 1995a,b),
6 days of observation may be too short to allow the
immune system to respond to denervation in nonin-
flammatory pulp tissue. Secondly, sympathectomy
relieves repression of sensory nerves and activates sen-
sory nerves to release SP and CGRP. This possibility is
International Endodontic Journal, 54, 85–99, 2021 91
 Dental nerves in pulpitis Zhan et al.
supported by the fact that knockout of NPY receptors
or application of adrenergic antagonists increases the
release of SP and CGRP (Bowles et al. 2003a,b, Harg-
reaves et al. 2003, Shi et al. 2006).
Products, such as b-endorphin, somatostatin and
IL-1, from immune cells can interact with receptors
on dental afferent nerves, resulting in an increase in
peripheral antinociception or nociception (Mudie &
Holland 2006, Rutz et al. 2007, Bletsa et al. 2009).
For example, immune cells express more corti-
cotropin-releasing factor (CRF) receptors to bind CRFs,
and this progression promotes immune cells them-
selves to release more b-endorphin, which can inter-
act with opioid receptors to decrease the sensitivity of
dental nerve endings in pulpitis (Rutz et al. 2007).
Up-regulation of IL-1 during pulpitis, accompanied
with an increased amount of IL-1 receptor type I on
sensory CGRP-immunoreactive and sympathetic NPY-
immunoreactive nerve fibres, leads a nociceptor sensi-
tization through the synthesis and release of prosta-
glandins and sympathetic amines (Bletsa et al. 2009).
Furthermore, IL-1 increases the neuronal content of
SP, and IL-1a is significantly more efficient than IL-
1b at up-regulating SP expression (Skoff et al. 2009).
Thus, inflammatory mediators form a positive feed-
back loop in promoting the pulpal inflammation pro-
cess.
The role of nerves in apical periodontitis
In the later stage of pulpitis, inflammation or necrosis
extends beyond the apical foramen and into the peri-
apical tissue, ultimately resulting in acute/chronic
apical periodontitis. It has been found that alterations
in the number of macrophages and dendritic cells in
periapical lesions are particularly well synchronized
with neural changes; and once the density of dental
sensory nerves reaches a peak, periapical tissue repair
starts to take place. These phenomena suggest that
dental nerves participate actively in the immune reac-
tion and tissue repair in apical periodontitis (Toriya
et al. 1997, Yang et al. 2007).
Infection-stimulated bone destruction at the periapi-
cal area is characteristic of apical periodontitis, whose
process is accompanied by the sprouting of dental
nerves (Kimberly & Byers 1988, Haug & Heyeraas
2003). Intriguingly, the role of sympathetic nerves in
bone remodelling appears to be distinct from its role
in the formation of tertiary dentine. Sympathetic
nerves inhibit the production of the bone-resorbing
pro-inflammatory cytokine IL-1a that can increase
92 International Endodontic Journal, 54, 85–99, 2021
the number of osteoclasts (Bletsa et al. 2004). Fur-
thermore, VIP from parasympathetic fibres might also
participate in the development and healing of periapi-
cal lesions (Azuero-Holguin et al. 2003). What is
worth mentioning, in particular, is that both osteo-
blasts and osteoclasts have been shown to express
VIP receptors. Importantly, VIP can exert a transient
inhibitory effect on osteoclast activity, which results
in a delayed stimulation of osteoclastic pit forma-
tion in bone slices; whereas, in the presence of osteo-
blast stromal cells, it increases the number of
resorption pits significantly (Lundberg et al. 2000).
Effects on healing in pulpitis
In injured pulpal tissues, the inflammation process is
always accompanied by tissue repair that involves the
activation of fibroblasts, odontoblasts and undifferenti-
ated mesenchymal cells. Importantly, this repair pro-
cess has been suggested to be under neuronal control.
Indeed, neuronal protein receptors are found on hard
tissue-forming cells, including odontoblasts, amelo-
blasts and cementoblasts of rat teeth (Fristad et al.
2003). Moreover, changes in pulpal innervation may
be partly responsible for degenerative alterations in
rodent teeth. Several studies have reported that dener-
vated incisors exhibit reduced microhardness and turn
chalky about 1 week after operation and fracture
1 month after denervation (Krage et al. 2005, Zhao
et al. 2014, Hayano et al. 2018, Liu et al. 2020).
Within the dental pulp, fibroblasts are the most
common cells responsible for the production and
maintenance of the pulp extracellular matrix (Jean-
neau et al. 2017). Once these cells are less active, or
even dead, the repair of the pulp tissue may be slowed
during pulpitis. It has been reported that SP, NKA
and CGRP can act as competence factors involved in
pulpal fibroblast proliferation in the process of tissue
repair (Bongenhielm et al. 1995, Trantor et al. 1995).
On the one hand, these neuropeptides may have a
negligible impact on glycosaminoglycan synthesis
(Trantor et al. 1995). On the other hand, they can
regulate the angiogenic functions of fibroblasts by
altering the expression of angiogenic growth factors
(El Karim et al. 2009). Of note, this mechanism may
operate in conjunction with other mechanisms such
as direct modulation of endothelial cells or the attrac-
tion of the immune cells with angiogenic potential to
the site of injury (Caviedes-Bucheli et al. 2017).
Amongst these neuropeptides, CGRP may act as a
paracrine factor, promoting proliferation of
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd

endothelial cells by stimulating cyclic AMP formation
(Haegerstrand et al. 1990).
Tertiary dentine formation, as another form of heal-
ing in pulpitis, can protect the pulp from injury. Not
only the formation of reparative dentine in pulpitis but
also the formation of dentine bridges following pulpo-
tomy is synchronized with the morphological reaction
of CGRP-immunoreactive nerve fibres (Taylor & Byers
1990). It has been reported that innervation maintains
the protein-synthetic repair processes of odontoblasts
(Chiego et al. 1987), and chemical or surgical denerva-
tion will cause delayed formation of reparative dentine
formation (Jacobsen & Heyeraas 1996). Moreover,
stimulation of human pulp-derived cells, containing
odontoblast-like cells, with CGRP results in increased
expression of bone morphogenetic protein-2 (BMP-2)
mRNA, potentially promoting dentine formation (Cal-
land et al. 1997). Treating pulp with CGRP has been
shown to enhance the formation of osteodentine and
tertiary dentine following pulpotomy (Kline & Yu
2009). In addition to neuropeptides, other molecules
released from dental nerves are also involved in den-
tine formation. For example, inhibition of sonic hedge-
hog (Shh) secretion from dental nerves can reduce
dentine formation in vivo (Zhao et al. 2014). It
therefore appears that various molecules secreted from
the nerve facilitate odontoblast differentiation and
function. Previous studies have implicated serotonin
(5-hydroxytryptamine, 5-HT) as an important neuro-
transmitter in both the peripheral and CNS (Ren et al.
2018, Li et al. 2020). Of note, serotonin has been
reported to be involved in dental tissue repair (Baudry
et al. 2019), although dental pulpal nerves do not
serve as the main source of serotonin (Kerezoudis et al.
1995a,b). Moreover, dental pulp stem cells can express
5-HT receptors that bind to the 5-HT released from
themselves and pulp injury-activated platelets, which
is critical for stem cell-mediated dental tissue repair
(Baudry et al. 2015), whereas deletion and inhibition
of 5-HT receptor provoke a significant reduction in
dentine formation (Dimitrova-Nakov et al. 2014,
Baudry et al. 2015).
In the case of severe pulpitis, reparative dentine for-
mation was found more frequently in sympathec-
tomized rats compared with control rats (Haug &
Heyeraas 2003). Although it remains unexplainable
that the ipsilateral sides of sympathectomy form more
reparative dentine than the other side, sympathetic
nerves can still be considered to have inhibited repara-
tive dentine formation to some extent (Haug & Heyer-
aas 2003). Also, it has been shown that
© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd
Zhan et al. Dental nerves in pulpitis
norepinephrine decreases tertiary dentine formation
via b2-adrenoceptors existing in the odontoblastic layer
(Gu et al. 2015).
Clinical implications
There remain considerable challenges associated with
the success rate of vital pulp treatments (VPT). However,
existing evidence suggests that dental nerves, apart from
the transmission of pain impulses, are also involved in
the modulation of various aspects of pulpal inflamma-
tion. It is therefore expected to improve treatment out-
comes and prognosis of VPT through manipulating
dental nerve activity, especially in the initial stage of pul-
pal injury. This may help dampen local inflammatory
responses of pulp tissues directly beneath the sites of
injury (caries, trauma, etc.), thereby contributing to the
preservation of its vitality and integrity.
At present, there are two potential ways to manipu-
late dental nerve activity. The first one relies on the
use of lasers. Treatment with either an Er:YAG or Nd:
YAG laser has been reported to change neuropeptide
expression in the pulp (Takamori 2000, Yoo et al.
2014), and this alteration induces marked fibroblast
proliferation and the formation of reparative dentine
(Takamori 2000). More importantly, benefiting from
the rapid development of a drug delivery system
(Thongkukiatkun et al. 2015, Ji et al. 2018), coordi-
nated interventions against the complex pathophysiol-
ogy of pulpitis through the dental nerve can be a novel
therapeutic approach for pulpitis. Although progress
has been made in understanding the mediator role of
dental nerves in pulpitis, more work remains to be
done. For example, many regulatory factors mentioned
above are also pain-promoting molecules, especially SP
and CGRP. Thus, this raises questions regarding the
situations in which the healing-promoting effects of
these molecules could be maximized and the stimula-
tion-related adverse effects could be minimized, as well
as whether inhibitors of specific channels can effec-
tively alleviate pain symptoms without influencing the
healing-promoting effects of neuropeptides as well as
other molecules in pulpitis. Answering these questions
will extend current knowledge in this field and make it
possible to deliver optimal molecule agonists and
antagonists to pulpal tissues at the right time.
Conclusions
The developments described in this review demon-
strate a complex and pivotal role of dental nerves in
International Endodontic Journal, 54, 85–99, 2021 93
 Dental nerves in pulpitis Zhan et al.
mediating pulpal inflammatory responses, which goes
far beyond the traditional concept related to pain sig-
nal transmission. Significantly, nerve fibres reaching
the para-odontoblastic region underneath areas of
carious dentine are now regarded as essential for reg-
ulating both immune cell recruitment and dentine
regeneration. These advances represent a start to fill-
ing the current gap in knowledge concerning the
functional interplay between the nervous system and
dental pulp, thus opening the way for precise investi-
gations of the neurobiology of the dental pulp as well
as the mechanisms underlying nerve-mediated regula-
tion of pulpal inflammation and regeneration. This
may open new therapeutic perspectives in endodon-
tics, and future development of novel strategies target-
ing nerve cells should be eagerly anticipated.
Acknowledgements
This work was supported by the Natural Science Founda-
tion of China (Grant Nos. 81500870, 81971902) and
Student’s Platform for Innovation and Entrepreneurship
Training Program (Grant No. c1051222).
Conflict of interest
The authors have stated explicitly that there are no
conflicts of interest in connection with this article.
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