J Oral Pathol Med (2009) 38: 29–33

doi: 10.1111/j.1600-0714.2008.00721.x ª 2009 John Wiley & Sons A/S Æ All rights reserved

www.blackwellmunksgaard.com/jopm

Calcitonin gene-related peptide levels in saliva of patients

with burning mouth syndrome
Jasna Zidverc-Trajkovic1, Dragan Stanimirovic2, Radmila Obrenovic1, János Tajti3, László Vécsei3,
János Gardi4, József Németh5, Milija Mijajlovic1, Nadezda Sternic1, Ljiljana Jankovic2
1
Institute of Neurology, Clinical Center of Serbia, Belgrade, Serbia; 2Clinic for Periodontology and Oral medicine, Faculty of
Stomatology, University of Belgrade, Belgrade, Serbia; 3Department of Neurology, Albert Szent-Gyorgyi Medical University,
Szeged, Hungary; 41st Department of Internal Medicine, Albert Szent-Gyorgyi Medical University, Szeged, Hungary;
5
Department of Pharmacology and Pharmacotherapy, University of Debrecen, Debrecen, Hungary

Burning mouth syndrome (BMS) is an intraoral burning
sensation for which no medical or dental cause can be
found. Recent studies suggest that primary neuropathic
dysfunction might be involved in the pathogenesis of
BMS. Calcitonin gene-related peptide (CGRP) plays an
important role in the development of pain and serves as a
biological marker of trigeminovascular activation. The
aim of this study was to determine the levels of CGRP in
the saliva of BMS patients and estimate the trigemino-
vascular activation in BMS. CGRP levels were measured,
by RIA method in 78 BMS patients and 16 healthy sub-
jects. The levels of CGRP were non-significantly
decreased in BMS patients in comparison to healthy
subjects. These results suggest that trigeminal nerve
degeneration may be the underlying cause of BMS.
J Oral Pathol Med (2009) 38: 29–33
Keywords: burning mouth syndrome; calcitonin gene-related
peptide; saliva
Introduction
According to the International Classification of Head-
ache Disorders, burning mouth syndrome (BMS) is an
intraoral burning sensation for which no medical or
dental cause can be found (1). BMS, coded 13.18.5, is
classified as a separate group with other cranial neural-
gias and central causes of facial pain. Diagnostic criteria
include: pain in the mouth that is present daily and
persisting for most of the day, normal appearance of
intraoral mucosa and exclusion of local and systemic
diseases (1). Epidemiological studies have estimated
BMS to be prevalent in 1–15% of the general population
Correspondence: Jasna Zidverc-Trajkovic, Institute of Neurology,
Clinical Center of Serbia, 11 000 Belgrade, Serbia. Tel: +381112614122,
ext.106, Fax: +381112684577, E-mail: jzidverc@gmail.com
Accepted for publication September 9, 2008
and it is a disorder which is seven times more common
disorder in females than males (2).
Despite the large number of clinical and epidemio-
logical studies, pathogenesis and aetiology of BMS
remains unclear (3). Recent clinical, electrophysiological
(4, 5) and histological (6) studies suggest that primary
neuropathic dysfunction might be involved in the
pathogenesis of BMS. On the other hand, a central
mechanism with the involvement of dopamine receptors
in the basal ganglia was suggested to play a role in the
pathogenesis of the disease (7).
Calcitonin gene-related peptide (CGRP) plays an
important role in the development of pain and hyper-
algesia. CGRP is a potent vasodilator that is contained
in and released from sensory nerve endings during
migraine attacks (8). CGRP modulates nociceptive
transmission in the trigeminovascular system (9). In
the jugular venous blood plasma concentrations of
CGRP, but not of other neuropeptides, were elevated
during the headache phase of migraine (10). In migraine
patients the baseline CGRP levels were considerably
higher and the changes in plasma CGRP levels during
migraine attacks significantly correlated with the head-
ache intensity (11). The cerebral vasculature is prefer-
entially innervated by CGRP-containing sensory nerves
(12). Therefore, it is now generally accepted that CGRP
is a biological marker of trigeminovascular activation
(13–15).
The aim of this study was to determine the levels of
CGRP in saliva of our group of patients with BMS and
in that way to try to estimate the trigeminovascular
activation in this painful disorder.
Patients and methods
Subjects
Between April 2004 and April 2005 the examined group
was formed from patients who was referred and
examined to the Clinic for Periodontology and Oral
medicine, Faculty of Stomatology, University of Bel-
grade, Serbia for unremitting sensory disturbances in the
 CGRP in saliva of patients with BMS
Zidverc-Trajkovic et al.
30
mouth for at least 6 months. Seventy-eight patients were
diagnosed as primary BMS, according to the Interna-
tional Headache Classification criteria. The control
group consisted of 16 young healthy medical staff and
served to standardize normal CGRP saliva levels. All
patients underwent a general medical examination, a
detailed blood test examination, including glucose
tolerance testing and B12 vitamin levels. Selected cases
were screened for connective tissue disorders and
thyroid gland function. Oral infections were ruled out
by microbiological culture. A thorough neurolo-
gical examination was performed in all patients and
additional diagnostic procedures (cranial computed
tomography or brain magnetic resonance and electro-
myography) in patients with abnormal results of neu-
rological examination.
Patients with oral mucosal diseases, bacterial and
fungal oral infections, diabetes, anaemia, connective
tissue disorders and diseases of thyroid gland were
excluded from the study. All patients and control
subjects gave their written informed consent to the
study that was approved by the Ethical Committee of
Faculty of Stomatology, University of Belgrade.
The following information was recorded for every
BMS patient: age, gender, duration of the disorder
expressed in months, presence of taste disturbances
and⁄or subjective feeling of dry mouth, the extent of
burning sensation classified in three grades (I – involving
only tongue, II – extending from tongue to other mouth
organs and III – spreading in the whole oral cavity), as
well as the speed of salivary flow (SF; normal value
1.5 ml ⁄ 15 min).
Saliva collection
The participants were asked to retrain from eating,
drinking and smoking for at least 1 h prior to saliva
collection. Each individual participating in the study
was tested in the morning, approximately at the same
time, considering the differences in saliva production
during the day. The saliva was collected for 15 min as
unstimulated salivary flow. The saliva samples were
collected in small test tubes, stored in ice bath and
centrifuged at 1600 g on 4C for 10 min. The super-
natants were frozen at )80C immediately after the end
of the session. This part of the study was carried out at
the Clinic for Periodontology and Oral medicine,
Faculty of Stomatology, University of Belgrade and
Institute of Neurology, Clinical Center of Serbia in
Belgrade.
Measurement of CGRP in saliva extract
Tyr-a-CGRP (23–27) (Bachem, Bubendorf, Switzer-
land), acetic acid (Reanal, Budapest, Hungary), aceto-
nitrile were used. 125I-Tyr-a-CGRP (23–37) was
prepared by the method described earlier (16). CGRP
antiserum was provided by T. Gorcs, Semmelweis
Medical University, Budapest. The saliva sample0073
was centrifuged, diluted with 4% acetic acid
(pH = 4.0), and then loaded into Sep-Pak C18 mini
columns. The peptide was eluted with 60% acetonitrile
in 0.5% acetic acid (pH = 4.0), lyophilized, and stored
J Oral Pathol Med
at )70C. Concentration of CGRP in saliva extracts was
determined by means of radioimmunoassay as described
previously (17). Sensitivity of the assay was 0.2 fmol ⁄
tube. Measurements of CGRP levels were performed at
the University of Szeged, Hungary.
Statistical analyses
The data are presented as arithmetic mean ± SD or as
percentages. The Kolmogorov–Smirnov test was applied
to assess the normality of the studied continuous data.
Independent samples t-test was used to compare data
between the two groups and one-way ANOVA to
compare data between three groups. The level of
significance for the analysis was set at 5% (P < 0.05).
Results
Our sample consisted of 78 patients diagnosed with
primary BMS. Of the 78 subjects included in this study,
51 (65.4%) were females. Mean age of the studied BMS
patients was 64.6 ± 10.9 years (range 25–81), 58
(74.36%) patients were older than 60 years. Forty-six
(59%) patients suffered from BMS disorder longer than
1 year (range 6–180 month). Subjective feeling of dry
mouth was reported by 76.5% of the patients and taste
disturbances by 35.5% of the patients. The distribution
of BMS complaints which were confined to the tongue
had 23 patients (30.1%), while 21 patients (27.4%)
complained of burning sensation extending from the
tongue to the hard palate, lips or alveolar ridges.
Burning sensation in the whole oral cavity was reported
by 33 patients (42.5%). Mean values of salivary flow in
BMS patients were 4.40 ± 2.40 ml ⁄ 15 min and only 7
(9%) patients had salivary flow equal or below
1.5 ml ⁄ 15 min. The values of SF in BMS patients were
significantly higher (t = 2.114, d.f. = 92, P = 0.037)
than in healthy subjects (3.06 ± 1.81 ml ⁄ 15 min).
Twenty patients had abnormal neurological findings.
Lacunar infarctions with or without leucoaraiosis were
detected in five (6.4%) patients, polyneuropathy was
detected in 10 (12.8%) patients and extrapyramidal
disease in five (6.4%) patients. In the extrapyramidal
disorders group, two patients had initial form of
Parkinson’s disease and three of them had essential
tremor.
Comparison of CGRP values between the examined
group of patients and the control group showed lower
values in patients (2.93 ± 1.43 nmol ⁄ l) than in controls
(3.45 ± 0.81 nmol ⁄ l) without significant difference
(t = )1.396, d.f. = 92, P = 0.166). CGRP values were
compared between BMS patients according to different
demographic features (Table 1). CGRP levels were
significantly higher in BMS patients with longer disease
duration. No significant difference was found in CGRP
levels of BMS patients according to gender and age
(Table 1).
Calcitonin gene-related peptide values were compared
between BMS patients with different BMS features
(Table 2). No significant difference was found in CGRP
levels of BMS patients according to presence of
subjective feeling of dry mouth, taste disturbances,
 CGRP in saliva of patients with BMS
Zidverc-Trajkovic et al.

31
Table 1 Calcitonin gene-related peptide (CGRP) levels in burning majority of patients were seniors. The demographical
mouth syndrome patients compared according to different demo- features detected in our patients’ group are in accor-
graphical features
dance with results of other authors who estimated that
Demographical feature CGRP levels (nmol ⁄ l) Statistics BMS is most frequently reported by post-menopausal
women (18). Patients with BMS frequently have other
Gender: females vs. males 2.73 ± 1.48 vs. 3.32 ± 1.28 P = 0.083 complaints and amongst these symptoms, xerostomia
Age: older than 3.01 ± 1.42 vs. 2.70 ± 1.47 P = 0.412
60 years vs. and loss or altered sense of taste are most frequently
yonger than 60 years mentioned (18). A majority of our BMS patients
Duration of the disorder: 3.22 ± 1.58 vs. 2.52 ± 1.08 P = 0.034 complained of subjective feeling of dry mouth and more
longer than 1 year vs. than one-third of patients reported abnormalities in
shorter than 1 year
taste perception. However, the decreased salivary flow
was detected in <10% of examined patients. The rate of
salivary flow was even higher in BMS group in
comparison with healthy controls. Current researches
Table 2 CGRP leves in BMS patients compared according to
have indicated that altered compositions of saliva such
different BMS features as a lower expression of low-molecular weight proteins
in individuals with BMS may contribute to changes in
BMS feature CGRP levels (nmol ⁄ l) Statistics
the lubricating function of saliva and in the perception
properties of the oral mucosa (19). Elevated salivary
Subjective feeling 2.97 ± 1.49 vs. 2.69 ± 1.72 P = 0.759 viscosity may result in a thin and discontinuous salivary
of dry mouth: reported film covering the oral mucosa that triggers the sensation
vs. not reported
Taste disturbances:reported 2.95 ± 1.90 vs. 2.88 ± 1.32 P = 0.932 of a dry mouth and causes lingual receptors such as
vs. not reported vanilloid receptors to be more exposed to stimuli (20).
Distribution of BMS 2.57 ± 1.39 vs. 2.81 ± 1.49 P = 0.345 The vanilloid receptors are localized in oral mucosa of
complaints:tongue vs. vs. 3.15 ± 1.48 the tongue, which is the area most frequently reported
extension to the other
mouth
by BMS patients. All our patients had burning sensation
organs vs.whole oral cavity of the tongue, and two-thirds of them complained that
Salivary flow: normal 3.00 ± 1.45 vs. 2.25 ± 1.11 P = 0.188 these sensations extended to the other mouth organs or
vs. decreased the whole oral cavity.
The main result of this study is that the levels of
BMS, burning mouth syndrome; CGRP, calcitonin gene-related pep-
tide. CGRP are not elevated in saliva of patients with BMS.
CGRP plays an important role in the development of
pain and hyperalgesia, particularly at the level of
trigeminal caudal nuclei (21). It has also been hypoth-
Table 3 CGRP leves in BMS patients compared according to esized that CGRP is implicated in the pathogenesis of
different neurological disorders migraine attacks (10, 12, 22). In the last decade, clinical,
psychophysical, and, more recently, electrophysiological
Neurological disease CGRP levels (nmol ⁄ l) Statistics studies suggested that primary neuropathic dysfunction
Cerebrovascular disorder: 4.32 ± 0.73 vs. 2.84 ± 1.42 P = 0.024 might be involved in the pathogenesis of BMS. A
present vs. absent neuropathic basis for BMS has been supported by
Polyneuropathy: 2.62 ± 1.43 vs. 2.98 ± 1.44 P = 0.469 observations that it is frequently accompanied by
present vs. absent changes in taste, altered pain or sensory perceptions.
Extrapyramidal disease: 3.32 ± 0.81 vs. 2.91 ± 1.46 P = 0.539
present vs. absent
Earlier studies had shown chemosensory and heat
tolerance impairment (23). Later, these findings were
BMS, burning mouth syndrome; CGRP, calcitonin gene-related pep- strengthened by the evidence of altered thermal and
tide. nociception thresholds in the tongue (4, 5) that have
been reported in BMS patients, as well as abnormalities
in the blink reflexes responses mediated by small-
distribution of complaints or values of SF (Table 2). diameter nerve fibres (4). Superficial biopsy of the
CGPR values were compared between BMS patients tongue demonstrated diffuse degeneration of epithelial
with different neurological disorders (Table 3). BMS and sub-papillary nerve fibres in the anterior two-thirds
patients with cerebrovascular disorders had higher levels of the tongue as a sign of a small-fibre sensory
of CGRP than BMS patients without these disorders. trigeminal nerve neuropathy (6). Decreased levels of
No significant difference was found in CGRP levels of CGRP measured in our BMS patients could be addi-
BMS patients according to presence of polyneuropathy tional proof of the trigeminal nerve degeneration in this
or extrapyramidal disorder (Table 3). disorder.
Age, gender, distribution of burning sensations,
salivary flow, as well as subjective feelings of dry mouth
Discussion and taste disturbances did not influence the CGRP levels
The examined group of BMS patients consisted of 78 of our BMS patients. Polyneuropathy was the most
subjects. Nearly two-thirds of them were women and common neurological disorder detected in 13% of our

J Oral Pathol Med

 CGRP in saliva of patients with BMS
Zidverc-Trajkovic et al.
32
patients. The causal relationships between BMS and
diabetes that have been proposed include poor glycemic
control and diabetic neuropathy (24). BMS was more
prevalent in diabetic patients with distal symmetrical
polyneuropathy (25). We excluded patients with diabe-
tes from our study, so the polyneuropathy of our
patients was not diabetic. However, we could not
exclude that the same cause, other than diabetes, was
responsible for polyneuropathy and trigeminal neurop-
athy in our BMS patients.
According to the results of our study, the patients with
longer disease duration had higher values of CGPR levels
in comparison to patients with shorter disease duration.
This result pointed to the role of centrally mediated
mechanisms of pain control in BMS. CGRP is one of the
neurotransmitters found in the nerve fibres of both the
sensory and autonomic nervous system that is involved in
salivary secretion. Hypoactivity of the nigrostriatal
dopaminergic system has been documented in BMS
patients (7, 26). Only five patients in our BMS group
had extrapyramidal disorder. However, the CGRP levels
were not different in this group of patients in comparison
with BMS patients without extrapyramidal disorder. By
using functional magnetic resonance imaging brain hyp-
oactivity similar to those of patients with other chronic
neuropathic pain conditions was displayed in BMS
patients (27). The CGRP levels were higher in five
patients with cerebral lacunar infarctions. Cerebral lac-
unes are secondary to thrombotic occlusion of the small
penetrating arteries and are located mainly in deep white
matter of cerebral hemispheres periventriculary (28). It
could be speculated that the lesions of anatomical
structures that are involved in the transmission or
modulation of nociceptive signals, such as the periaqu-
eductal grey matter, may lead to an impairment of
antinociceptive activity and subsequently result in a
permanent pain feeling, like in chronic migraine (29).
Decreased levels of CGRP measured in this study support
the premise that trigeminal nerve degeneration may be the
underlying cause of BMS and further studies are neces-
sary to determine the origin of neuropathic process.
References
1. Headache Classification subcommittee of the International
Headache society. The international classification of head-
ache disorders, 2nd edn. Cephalalgia 2004; 24 (Suppl. 1):
1–160.
2. Lipton JA, Ship JA, Larach-Robinson D. Estimated
prevalence and distribution of reported orofacial pain in
the United States. J Am Dent Assoc 1993; 124: 115–21.
3. Zakrzewska JM. The burning mouth syndrome remains an
enigma. Pain 1995; 62: 253–7.
4. Forssell H, Jaaskelainen S, Tenovuo O, Hinkka S. Sensory
dysfunction in burning mouth syndrome. Pain 2002; 99:
41–7.
5. Gao S, Wang Y, Wang Z. Assessment of trigeminal
somatosensory evoked potentials in burning mouth syn-
drome. Chin J Dent Res 2000; 3: 40–6.
6. Lauria G, Majorana A, Borgna M, et al. Trigeminal
small-fiber sensory neuropathy causes burning mouth
syndrome. Pain 2005; 115: 332–7.
J Oral Pathol Med
7. Hagelberg N, Forssell H, Rinne JO, et al. Striatal dopa-
mine D1 and D2 receptors in burning mouth syndrome.
Pain 2003; 101: 149–54.
8. Uddman R, Edvinsson L, Ekblad E, Hakanson R, Sundler
F. Calcitonin gene-related peptide (CGRP): perivascular
distribution and vasodilatatory effects. Regul Pept 1986;
15: 1–23.
9. Brain SD. Calcitonin gene-related peptide (CGRP) anta-
gonists: blockers of neuronal transmission in migraine. Br
J Pharmacol 2004; 142: 1053–4.
10. Goadsby PJ, Edvinsson L, Ekman R. Vasoactive peptide
release in the extracerebral circulation of human during
migraine headache. Ann Neurol 1990; 28: 183–97.
11. Juhasz G, Zsombok T, Modos EA, et al. NO-induced
migraine attack: strong increase in plasma calcitonin gene-
related peptide (CGRP) concentration and negative cor-
relation with platelet serotonin release. Pain 2003; 106:
461–70.
12. Edvinsson L. Blockade of CGRP receptors in the intra-
cranial vasculature: a new target in the treatment of
headache. Cephalalgia 2004; 24: 611–22.
13. Edvinsson L, Uddman R. Neurobiology in primary
headaches. Brain Res Rev 2005; 48: 438–56.
14. Edvinsson L. Novel migraine therapy with calcitonin gene-
regulated peptide receptorantagonists. Expert Opin Ther
Targets 2007; 11: 1179–88.
15. Link AS, Kuris A, Edvinsson L. Treatment of migraine
attacks based on the interaction with the trigemino-
cerebrovascular system. J Headache Pain 2008; 9: 5–12.
16. Nemeth J, Oroszi G, Jakab B, et al. 125I-labeling and
purification of peptide hormones and bovine serum
albumin. J Radioanal Nucl Chem 2002; 251: 129–33.
17. Nemeth J, Gorcs T, Helyes Z, et al. Development of a new
sensitive CGRP radioimmunoassay for neuropharmaco-
logical research. Neurobiology 1998; 6: 473–5.
18. Scala A, Checchi L, Montevecchi M, Marini I, Giamber-
ardino MA. Update on burning mouth syndrome: over-
view and patient management. Crit Rev Oral Biol Med
2003; 14: 275–91.
19. De Moura SA, De Sousa JM, Lima DF, Negreiros AN,
Silva F de V, Da Costa LJ. Burning mouth syn-
drome (BMS): sialometric and sialochemical analysis
and salivary protein profile. Gerodontology 2007; 24:
173–6.
20. Yilmaz Z, Renton T, Yiangou Y, et al. Burning mouth
syndrome as a trigeminal small fibre neuropathy: increased
heat and capsaicin receptor TRPV1 in nerve fibres
correlates with pain score. J Clin Neurosci 2007; 14: 864–
71.
21. Greco R, Tassorelli C, Sandrini G, Di Bella P, Buscone S,
Nappi G. Role of calcitonin gene-related peptide and
substance P in different models of pain. Cephalalgia 2008;
28: 114–26.
22. Fusayasu E, Kowa H, Takeshima T, Nakaso K, Naka-
shima K. Increased plasma substance P and CGRP levels,
and high ACE activity in migraineurs during headache-
free periods. Pain 2007; 128: 209–14.
23. Grushka M, Sessle BJ. Burning mouth syndrome. Dent
Clin North Am 1991; 35: 171–84.
24. Carrington J, Getter L, Brown RS. Diabetic neuropathy
masquerading as glossodynia. J Am Dent Assoc 2001; 132:
1549–51.
25. Moore PA, Guggenheimer J, Orchard T. Burning mouth
syndrome and peripheral neuropathy in patients with type
1 diabetes mellitus. J Diabetes Complications 2007; 21:
397–402.
 CGRP in saliva of patients with BMS
Zidverc-Trajkovic et al.

26. Jaaskelainen SK, Rinne JO, Forssell H, et al. Role of the
dopaminergic system in chronic pain – a fluorodopa-PET
study. Pain 2001; 90: 257–60.
27. Albuquerque R, De Leeuw R, Carlson C, Okeson J, Miller
C, Andersen A. Cerebral activation during thermal stim-
ulation of patients who have burning mouth disorder: an
fMRI study. Pain 2006; 122: 223–34.
28. Fisher CM. Lacunar infarcts – an overview. Cerebrovasc
Dis 1991; 1: 311–20.
33
29. Welch KM. Concepts of migraine headache pathogenesis:
insights into mechanisms of chronicity and new drug
targets. Neurol Sci 2003; 24: 149–53.
Acknowledgements
We would like to thank Bojana and Nathan Abahari for the English
corrections.

J Oral Pathol Med