CLASSIFICATION OF NERVES
The dental pulp has an abundant supply of both sensory and autonomic nerves (Figure 24). This sophisticated arrangement of neural network plays an important role, although not fully understood, in managing the microenvironment of the dental pulp. The majority of these nerves are sensory. The trigem- inal ganglion supplies sensory innervation to the pulp via the maxillary and mandibular nerves. Sympathetic nerves are less numerous with its source from the superior cervical ganglion. Parasympathetic fibers have been suggested to exist in the pulp, although the source of this innervation is still disputed.
Anatomical and electophysiological studies have classified nerves according to their diameter and con- duction velocity (Table 1). The pulp contains two types of sensory nerve fibers: A fibers that are myeli- nated and C fibers that are unmyelinated. Nerve fibers enter teeth via the apical foramen, and arborize cor- onally to form the plexus of Rashkow in the subo- dontoblastic region. Some axons leave the plexus of Raschow, lose the myelin sheath to become unmyeli- nated, and pass between odontoblasts to reach inner dentin.191Nerve fibers have been found in more than 50% of tubules in inner cuspal dentin.190–194Coronal dentin is more densely innervated than radicular den- tin. Axons do not traverse beyond 200 mm into cor- onal and radicular dentin but usually end less than 200 mm from the pulp.191A fibers include both A-b and A-d with A-d amounting to approximately 90% of the A fibers.195
Classical electrophysiological studies have shown the afferent function of sensory nerves in pain sensa- tions. Some overlapping of functions have been
shown between pulpal A and C fibers.195–199The A-d fibers in the dental pulp have been involved in trans- mitting fast pain usually perceived as sharp, piercing pain, while C fibers transmit slow pain described as dull, aching pain.200 Sympathetic nerves in general have been shown to have a vasoconstrictor effect, while parasympathetic nerves have a vasodilator effect.
NEUROPEPTIDES
In addition to their vasoactive functions, stimulated sensory and autonomic nerves release biologically active peptides, known as neuropeptides, that influence neural activity and functioning.201–203 Neuropeptides are synthesized on ribosomes in the neuron cell body, processed through the endoplasmic reticulum and Golgi complex, transported in vesicles to the nerve terminals (Figure 25), and released from the peripheral terminals of mainly A-d and C fibers.204
Neuropeptides can be classified according to the nerves from which it is derived. For example, sensory nerve-derived neuropeptides include CGRP, SP, and neurokinin A (NKA), while NPY is typically a sympathetic nerve-derived neuropeptide. Vasoactive intestinal peptide (VIP) has been shown to be in parasympathetic nerves. The introduction of immu- nohistochemical labeling of neuropeptides has con- tributed greatly to our understanding of the neural network of the dental pulp.
DISTRIBUTION
CGRP-containing or CGRP-immunoreactive (CGRP- IR) nerve fibers are the most abundant in the dental pulp (Figure 26). It is three to four times more
Table 1 Classification of Nerves in the Pulp
Type of Fiber Function Diameter (mm) Conduction Velocity (m/s) Aa Motor, proprioception 12–22 70–120 Ab Pressure, touch 5–12 30–70 Ag Motor 3–6 15–30 Ad Pain, temperature, touch 2–5 12–30 B Preganglionic autonomic <3 3–15 C Pain 0.4–1.2 0.5–2 Sympathetic Postganglionic sympathetic 0.3–1.3 0.7–2.3
Figure 24 Schematic diagram showing the course of sensory and sympathetic nerves to the dental pulp. Sensory nerves (yellow) derive from the trigeminal ganglion while sympathetic nerves (green) are from the superior cervical ganglion. Postganglionic sympathetic nerves travel with the internal carotid nerve, reach the trigeminal ganglion, and supply teeth and supporting structures via the maxillary and the inferior alveolar nerve (yellow).
abundant than SP.205CGRP-IR fibers enter the pulp via the apical foramen in bundles either surrounding the blood vessels or individually and ramify into a network of fine fibers in the coronal pulp.205–207 Coronally, individual fibers penetrate the odontoblast layer and predentin to terminate in the inner 100 mm of the dental tubules. Radicular pulp fibers run parallel to the long axis of the tooth with little arborization and little penetration of the odontoblast layer.
SP is commonly found in C fibers with similar dis- tribution to CGRP.205,208,209 NKA fibers have similar distribution to SP-IR nerve fibers.210 CGRP, SP, and NKA have been shown to coexist in the same nerve fibers in the dental pulp. NPY is usually found in sympathetic nerves and its distribution varies from the sensory neu- ropeptides. It enters the pulp mainly surrounding the blood vessels and terminates in the floor or the mid- coronal part of the pulp chamber. It is more numerous in the radicular than in the coronal pulp.205,211–214
VIP-immunoreactive (VIP-IR) fibers in the dental pulp are associated with blood vessels215 or occur as free and interlacing nerves in the central pulp and the subodontoblastic plexus.215,216 Nerves containing neuro-peptides are also found in close contact with A
B
Figure 25 Calcitonin gene-related peptide-immunoreactive (CGRP-IR) neurons in the trigeminal ganglion. Low-magnification A, view of darkly stained neurons that contain CGRP. Higher magnification B, view of neurons with axons transporting CGRP to the peripheral nerve terminals. Courtesy of SR Haug and KJ Heyeraas.
Figure 26 Immunohistochemical staining of calcitonin gene-related peptide-immunoreactive (CGRP-IR) nerve fibers in the rat dental pulp. A, Low-magnification view of darkly stained nerve fibers entering the apical foramen and traversing the long axis of the tooth as thick bundles. Coronally, nerve fibers branch extensively. B, CGRP-IR fibers penetrate the cuspal odontoblast layer and enter dentin. C, In the mid-coronal region, thick bundles of fibers are seen surrounding the blood vessels and in root pulp. D, Thick band of fibers surrounding the blood vessels are in the root pulp. Courtesy of SR Haug and KJ Heyeraas.
immunocompetent cells suggesting important neu- roimmune interactions (Figure 27).
FUNCTION
Neuropeptides are released by various stimulations. What happens when a sensory neuropeptide is released? A classical demonstration of neuropeptide activity is the flare component of the ‘‘triple response’’ in the human skin: wheal (edema formation), local reddening
(increased blood flow), and flare.217 The flare is a con- sequence of the sensory nerve-mediated axon reflex and involves antidromic stimulation of sensory nerves to release neuropeptides such as SP and CGRP. These neuropeptides can be coreleased when an antidro- mic impulse depolarizes the peripheral end of the nerve. CGRP is a strong vasodilator. SP is also a vasodilator and in addition it increases vascular permeability and stimu- lates inflammatory and immune cells (Figure 28). NPY has many physiological effects including vasoconstriction,
A B
Figure 27 Calcitonin gene-related peptide-immunoreactive (CGRP-IR) nerve fibers in close proximity to immunocompetent cells in the dental pulp. Courtesy of SR Haug and KJ Heyeraas.
Figure 28 Schematic illustration summarizing some of the effects of neuropeptides (calcitonin gene-related protein, CGRP; substance P, SP) released from sensory nerves. Sensory nerves conduct afferent sensory information (e.g., pain sensations/dentin sensitivity) to the central nervous system and have an efferent function such as vasodilation and increased permeability of blood vessels. It affects various aspects of immune function such as activation and chemotaxis of immune cells, and release of cytokine and growth factors from various cellular structures in the dental pulp. Sensory nerves have also been shown to be involved in the formation of dentin.
immune regulation, and pain perception. In the dental pulp, CGRP and SP increases the blood flow while NPY decreases it.
Perhaps one of the most important function, although, little understood is how nerves affect the immune system, a term known as neuroimmunomodu- lation. SP and NKA are considered to have a proinflam-
matory role, while NPY and VIP are said to have an anti-inflammatory role on the immune system.218–221 Classic work by Byers et al.222–225has shown that CGRP and SP-IR nerve fibers undergo sprouting during inflam- mation in the dental pulp. These fibers also change in quantity and architecture when inflamed (Figure 29). Recent studies have shown that NPY-IR nerve
A B
C D
Figure 29 Calcitonin gene-related protein-immunoreactive (CGRP-IR) nerve fibers in the dental pulp. A, Low-magnification view of a rat molar tooth with pulp exposure (top left corner) for 20 days. Extensive formation of reparative dentin din the pulp chamber. B, On the right coronal pulp wall region, spouting of CGRP-IR fibers is observed. C, In the root pulp, sprouting of CGRP-IR nerve fibers is observed. D, Architecture of nerve fibers is quite different from that of a normal dental pulp from similar region. Courtesy of SR Haug and KJ Heyeraas.
fibers also sprout in long-standing inflammatory lesions (Figure 30).212 Quantification studies have shown that SP, CGRP, NKA, NPY, and VIP are increased in pulps diagnosed clinically as irreversi- ble pulpitis (Figure 31).218,226
RECEPTORS
Neuropeptides cannot cross the cell membrane and therefore their action is dependent on the existence of
specific receptors (Table 2). Without the existence of a particular receptor, release of neurotransmitters or neuropeptides will have no effect. Therefore identify- ing the location and distribution of these receptors is important for understanding the functionality of a neuropeptide in a given location. NK1 receptors, specific for SP, are present in the blood vessels in the root pulp and smaller vessels along the odontoblast layer.227,228 Fibroblasts express NK1 receptors during mechanical orthodontic stress.212
A B
C D
Figure 30 Neuropeptide Y-immunoreactive (NPY-IR) fibers in the inflamed dental pulp. A, Pulp exposure injury (top left) on a rat molar for 6 days. B, Higher magnification of A showing ‘normal’ location of NPY-IR fibers in the central pulp and on the floor of the pulp chamber. C, Pulp exposure injury (top right) on a rat molar tooth for 20 days. There is extensive reparative dentin formation and increased number of NPY-IR nerve fibers penetrating the region of the reparative dentin. D, Higher magnification. (A and B) Courtesy of SR Haug and KJ Heyeraas (C and D).212
Antagonists to these receptors are now being widely studied as therapeutic agents against inflammatory diseases or diseases involving overexpression of neuropeptides.
NEUROPHYSIOLOGY
Experimental animal model studies are widely used to investigate the role played by nerves in the dental pulp. Removing sensory nerves either by nerve transection or chemical removal has shown that sensory nerves are important for secondary dentin formation.229Releasing neuropeptides in the dental pulp by electrical stimula- tion results in transendothelial migration of inflamma- tory cells. This was absent when sensory nerves were previously removed from the dental pulp.230 Recently studies have demonstrated that sympathetic nerves are involved in the modulation of the immune system (Figure 32). Sympathetic nerves inhibit the production of proinflammatory cytokines, while stimulating the
production of anti-inflammatory cytokines.220 Teeth lacking sympathetic nerves also tend to undergo necrosis more frequently.211
Sympathetic nerves have been shown to be impor- tant in the recruitment of granulocytes during elec- trical stimulation of teeth and experimental tooth movement.231,232 Sympathetic imbalance results in the recruitment of plasma cells in an intact tooth suggesting an immune dysfunction.233 Sympathetic nerves have also been shown to have an inhibitory effect on osteoclast resorption activity and on the cytokine IL-1a.145,212,232 NPY that is coreleased in the sympathetic nerve terminal has recently been reported to modulate immune function in a wide range of inflammatory conditions. Therefore, the effects one sees in the dental pulp and surrounding structures could be due to the neuropeptide released from the sympathetic nerve terminal since an upregulation of NPY has been seen in the dental pulp.
Table 2 Neuropeptides and Their Origins Including Some of Their Actions and Specific Receptors
Neuropeptides Released From Actions Receptors
Substance P Sensory Vasodilation, increases vascular permeability NK1 Neurokinin A Sensory Increases vascular permeability NK2
Calcitonin gene-related peptide Sensory Vasodilation CGRP1 and CGRP2 Neuropeptide Y Sympathetic Vasoconstriction, pain modulation, immune function NPY Y1-6 Vasoactive intestinal peptide Parasympathetic? Vasodilation, immune function VIP 1 and 2
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