Behavioural
responses following tooth injury in
rats
Chudler EH, Byers MR
Department of
Anesthesiology, University of Washington,
Seattle
Introduction
Dental pain is estimated to affect 12-14% of
the US population annually. Despite its high
incidence, dental pain relies on century-old
methods for its diagnosis. Although clinical
treatment of dental pain has improved with new
understanding of psychological, sensory and
pharmacological mechanisms of trigeminal pain,
there are still significant areas for
improvement.
Several experimental models of orofacial
nerve injury have been developed in recent
years. Such methods have provided a wealth of
behavioural, neurophysiological, neuroanatomical
and neurochemical data concerning peripheral and
central nervous system changes subsequent to
injuries of the trigeminal system. Although
acute dental pain models have been developed in
rats and ferrets, few investigations concerning
the behavioural consequences of tooth injuries
in animals have been conducted.
Most studies investigating the behavioural
consequences of dental pain in animals have
relied on single indices of pain often in
response to electrical, mechanical or chemical
stimulation of the tooth. For example, the
jaw-opening reflex evoked by electrical
stimulation of the tooth has been used
extensively to investigate pain in rats. More
recently, Chattipakorn et al. used bacterial
lipopolysacchande to inflame the ferret tooth
pulp to evaluate pain behaviour based on the
number of tongue protrusions and Chidiac et al.
devised a pain scale based on abnormal grooming
and movements following intrapulpal capsaicin or
formalin applied to the rat incisor. Tooth
extraction has also been proposed to model
dental pain, but questions have been raised
about its usefulness.
The present study sought to determine the
shortterm behavioural effects of tooth injuries
with crystals of Fluorogold (FG) implanted into
molar teeth of adult rats. We were interested in
developing a set of quantitative measures of
behavioural changes that could be used with a
variety of pain models and retrograde neural
labelling techniques.
Discussion
Changes in spontaneous behaviour and body
weight following injuries to the infraorbitat
nerve, sciatic nerve and experimental arthritis
have been used as indices of pain in rats.
Alterations in steep patterns have also been
observed within the first few days after chronic
constriction of the sciatic nerve. Tooth
injuries, as produced in the present study,
result in behaviouraL changes similar to those
observed in these other pain models: reduced
body weight gain, increased time spent freezing
and decreased time spent exploring as welt as
inducing yawning behaviour in some of the
injured rats. Comparable behavioural
observations, including changes in movement,
steep and food intake, have been used to
evaluate human pain behaviour. Our data showing
greater behavioural changes when three to four
teeth are involved compared to the group with
two injured teeth are consistent with our
previous observation that larger injuries for
rat molar teeth caused a slower return to normal
weight gain and more extensive ganglionic
satellite cell can central c-Fos responses.
Experiments can therefore be designed to study
dental injuries and their effects on trigeminal
neuronal cytochemistry and function without
behavioural changes (two teeth) or with
behavioural changes (three or more teeth).
The behavioural tests presented here can now
be used to establish the behavioural
consequences of the various dental neuronal
retrograde labelling procedures. This would be
especially important for investigations that
label dental neurons for subsequent patch clamp
in vitro analyses, in order to know whether
those neurons come from animals with significant
pain behaviour. Previous studies have often
assumed that rats return to normal behaviour
when they resume feeding after surgery. Our
data, however, show that body weight, yawn and
motor behaviour can show delayed recovery.
FG labelled many dental neurons in the
corresponding trigeminal division of the
ipsilateral ganglion, and we estimate that more
than 500 dental neurons were routinely labelled
in right trigeminal ganglia in the three tooth
labelling group. We were surprised to find that
less than 20% of the FG neurons co-localized
CGRP, since others have strong evidence that FG
is an excellent tracer for following dental
neuronal changes after tooth injury. It is
likely that our implantation of FG crystals
directly onto pulp exceeded toxicity limits for
some of the neurons. Furthermore, we were
analysing the neurons at three days after
implant, while Pan et al. placed 200 µl of
5% FG into shallow cavities along the buccal
surface of the first maxillary molar, sealing it
in place, and then waited three to five weeks
before analysing the cytochemistry of neurons.
Such dentinal labelling procedure and another FG
technique that shows penetration of FG from oral
sulcus into intact rat molars did not cause
significant weight loss, showing that exposure
of rats to FG does not by itself cause reduced
feeding.
Although we observed differences in body
weight, exploration and freezing in
tooth-injured and sham-operated rats two days
after injury, these behavioural changes were
statistically significant only at the three-day
observation period. It is possible that the
absence of significant differences between
groups at the two-day period was due to residual
anaesthetic effects. However, other movement
assays (rearing, grooming, jaw motions) were
normal at two to three days after these FG
implants. Unlike changes in body weight,
exploration and freezing, statistically
significant alterations in jaw movements,
rearing and facial and body grooming were not
observed within the time frame of the present
study. Using the infraorbital nerve constriction
model, Vos et al. reported that rats displayed
increased facial grooming within three days
after surgery, but Benoliel et al did not
observe such changes until two weeks after a
similar surgery. It is possible that tooth
injuries do not result in abnormal sensory
phenomena (i.e., paresthesia) during the
observation period of the current
experiments.
A unique observation was the increased
number of yawns seen after tooth
injuries. Although the significance of
yawning following tooth injuries is
unclear, it may be related to stress.
Yawning can be elicited in horses by
abdominal stress and in rats by foot shock
stress. Yawning is also a symptom of some
migraine headaches in humans. It is unlikely
that the yawns after tooth injuries were
caused by damage to the jaw or temporomandibular
joint or to stress of the novel testing
environment because sham-operated rats did not
yawn. Moreover, because rats do not yawn
following infraorbital nerve injury, it is
likely that yawning is made in direct
response to sensory information from the tooth.
The use of a rat strain that displays a high
frequency of yawning may help resolve
this question because such animals might display
a higher number of yawns following tooth
injuries and reduce the variability of this
behavioural marker. Additional experiments to
observe rats at other times of the day and for
longer observation periods are also warranted.
It is possible that yawning is an
important behavioural consequence of tooth
injury that has been overlooked in previous
investigations.
Tooth injuries with implants of the tracer
Di-I, similar to the injury procedure in the
present study, increase the number of trigeminal
ganglion neurons encircled by glial fibrillary
acidic protein (GFAP) three days after surgery.
Such alteration in satellite cell cytochemistry
may not indicate altered electrophysiology, as
no changes in trigeminal neuronal function were
detected in the maxillary division where many
uninjured neurons exhibited prolonged increase
of GFAP expression, after injuries to the
mandibular neurons that project into the
inferior alveolar nerve. However, significant
increases in the expression of P/Q-type (Ca2.1)
calcium channels three days after tooth pulp
exposure have also been observed 20 after pulp
exposure injury.
The long-term behavioural consequences of
tooth injury were not determined in this study.
Moreover, motor behaviours were measured by the
time spent in locomoting, rearing and freezing.
A more detailed analysis of fine motor movement
may reveaL additional changes. Benoliel et al.
reported reduced bite force and altered bite
patterns within a few days after application of
a pro-inflammatory agent to the infraorbital
nerve of the rat. A recent report suggests that
bite force in trained rats is also a valid
measure for craniofacial muscle inflammatory
pain and it would be interesting to test such an
assay for behavioural responses to tooth injury.
Nevertheless, our results demonstrate that
changes in normal behaviour patterns can be used
to assess pain in rats.
Equine Vet J.1990; 22; 4; 241-243
Naloxone-induced abdominal distress in the
horse.
Kamerling SG, Hamra JG, Bagwell CA.
Department of Veterinary
Physiology, Pharmacology and Toxicology, School
of Veterinary Medicine, Baton Rouge,
Louisiana
Endogenous opioid peptides have been
implicated in the regulation of pain perception,
behaviour, gastrointestinal activity and other
physiological responses. However, the functional
role of these peptides in the horse has yet to
be elucidated. The opioid antagonist, naloxone,
is often administered to infer endogenous opioid
effects. In the present study, naloxone (0.75
mg/kg bodyweight) was administered to eight
Thoroughbred racehorses and a number of
behavioural and autonomic responses were
measured. Naloxone produced rapid onset
diarrhoea, restlessness, abdominal checking,
tachycardia, tachypnoea, paradoxical
yawning and diaphoresis. These responses
described an acute abdominal distress syndrome
similar to spasmodic colic. Results from this
study suggest that, in the horse, endogenous
opioids: 1) influence behaviour, 2) modify
intestinal activity and sensation, and 3) if
perturbed, may be involved in pathophysiology of
colic.
