28Stomatologija, Baltic Dental and Maxillofacial Journal, 2010, Vol. 12, No. 1
Functional activity of rabbit salivary glands in reduced
and restored regional arterial blood supply conditions
Kaspars Stamers, Andrejs Skagers, Kalvis Pastars, Nonna Tomisheva, Maija Ratniece
Background. Although the vascular pathology of carotid arteries is widespread, the function of
salivary glands in reduced arterial flow conditions is not much investigated clinically and in experi-
ments. At the same time blood supply is a keystone to normal functioning of every organ and
especially of salivary secretion. The aim of this study was to estimate functional activity of salivary
glands in reduced and restored blood supply conditions in experiment by sialoscintigraphy which is
an approved method for functional assessment of salivary glands.
Methods. The ligature of a. carotis communis dextra was performed on 20 Californian rabbits.
After 28 days sialoscintigraphy with Tc99 pertechnetate and revascularization through resection of
the occluded part of a. carotis communis and reconstruction with venous autograft was performed.
One month later sialoscintigraphy was done.
Results. The functional activity of rabbit salivary glands after the ligature of a. carotis commu-
nis is strongly depressed. The revascularized glands accumulated isotope slowly, but the level of
accumulation was higher than on the control side.
Conclusion. The ligature and reconstruction of a common carotid artery on rabbits confirm the
important role of the arterial blood supply in functional activity of salivary glands and may be an
appropriate experimental model for investigation of ischemic disease of salivary glands.
Key words: sialoscintigraphy, carotid artery ligature.
Stomatologija, Baltic Dental and Maxillofacial Journal, 12: 28-32, 2010
The occlusive diseases of carotid arteries are wide-
spread and often accompanied by disturbances of ce-
rebral circulation, like stroke and other vascular dis-
eases which represent a significant public health prob-
lem. Stroke accounted for about one of every 16 deaths
in the USA in 2004 and up to 25% are related to steno-
sis of the carotid arteries treated by carotid endarter-
ectomy procedures on 98,000 inpatients in 2004 (1).
Regional hypoxia and ischemia cannot be without
consequences also for tissue and organs of oro-maxillo-
facial region having blood supply from external carotid
artery (2). Vascular pathology largely accounts for dif-
ferent diseases of salivary gland. In Sjogren’s syndrome
five stages for morphogenesis of arteritis are proposed:
endothelial swelling, thrombosis, fibrinoid degeneration,
necrotizing panarteritis and endarteritis obliterans which
are closely connected with glandular tissue pathology
(3). The experimental model of oro-maxillo-facial is-
chemic disease may be useful for evaluation of angio-
genic pathology of different organs in this region and
perhaps also for development of new treatment meth-
ods. Ductal obstruction is more often used in investiga-
tions on experimental pathology of salivary glands (4)
and has vascular and secretory responses (5).
Functional activity of major salivary glands may
be tested using scintigraphy with 99mTc – technetium
pertechnetate (Tc-99) which has been used in clinical
conditions during the three last decades for functional
assessment in cases of Sjogren’s syndrome (6,7,8,9,10),
xerostomy after radiotherapy of thyroid and oro-facial
cancer (11,12,13,14), sialolithiasis (15), and gastro-
esophagel reflux disease (16). The use of Tc-99 scintig-
raphy is proposed for checking radioprotection of sali-
1Department of Oral and Maxillofacial Surgery, Riga Stradins Uni-
versity, Riga, Latvia
2Department of Reconstructive Microsurgery, University Hospital
"Gailezers", Riga, Latvia
3Department of Radionuclide Diagnostics, Stradins University Hos-
pital, Riga, Latvia
Kaspars Stamers1 – MD, D.D.S.
Andrejs Skagers1 – MD, Dr. hab. med, professor, Head of De-
partment of Oral and Maxillofacial Surgery
Kalvis Pastars2 – MD
Nonna Tomisheva2 – MD
Maija Ratniece3 – MD
Address correspondence to Kaspars Stamers, Department of Oral
and Maxillofacial Surgery, Dzirciema str. 20, Riga LV 1007, Latvia.
E-mail address: firstname.lastname@example.org
Stomatologija, Baltic Dental and Maxillofacial Journal, 2010, Vol. 12, No. 129
Fig. 1. Scintigramms of rabbit's salivary glands with Tc99:
Right side – 28 days after ligature of common carotid ar-
tery; left side – control; stimulation at 15th minute
vary glands (17). There is not yet a clear relationship
between Tc-99 scintigraphic data and saliva secretion
but there is a significant correlation between healthy
and xerostomic patients (18), age and gender differences
are also estimated (19). The quantitative analysis of time
active scintigraphy curve usually includes the uptake
rate and the washout fraction parameters enable objec-
tive testing of salivary gland function (20). The uptake
parameters are only sensitive to differences exceeding
25% of gland mass and are not useful to detect the kind
of disease causing xerostomia (10).
Our previous report (21) showed depressed functional
activity of salivary glands after ligature of a. carotis com-
munis on rabbits. The aim of this study was to estimate func-
tional activity of salivary glands in reduced blood supply
conditions in experiment by sialoscintigraphy.
MATERIALS AND METHODS
The experiments were authorized by the Animal
Ethics Committee of the Latvian Food and Veterinary
SCIENTIFIC ARTICLESK. Stamers et al.
Service. Twenty male Californian rabbits weighing ap-
proximately 3 kg were used. Under intravenous general
anesthesia with Diazepam 2 mg/kg and 5% Ketamin
hydrochloride 15 mg/kg and local infiltration with 0.5%
Lidocain solution 5 ml, the ligature of a. carotis com-
munis dextra was performed with 4-0 silk and the wound
was closed. After 28 days under general anesthesia 6
MBq/kg Tc99 pertechnetate was injected intravenously
and sialoscintigraphy using Siemens E.CAM equipment
was performed on 12 rabbits (Fig. 1).
After 15 minutes salivary excretion was stimu-
lated with five drops of lemon juice. Following the
scintigraphy, 12 rabbits were sacrificed with over dos-
age of anesthesia and salivary glands were taken out
for histological and immunohistochemical evaluation.
After general and local anesthesia, re-
vascularisation was attempted on 8 rabbits: in two cases
with extensive thrombosis resection of the occluded
part of common carotid artery, reconstruction was per-
formed with autovenous graft from v. jugularis interna
using Monosof 8-0 sutures and 6X magnification; two
Fig. 2. Scintigramms of rabbit's salivary glands with Tc99:
Right side – 28 days after reconstruction of common ca-
rotid artery; left side – control; stimulation at 15th minute
Table 2. The Bolton anterior ratio in different malocclusionTable 1. Maximal level of counts per second in accumula-
tion phase (counts/sec) on side of a. carotis communis liga-
ture (test) and control sides
30Stomatologija, Baltic Dental and Maxillofacial Journal, 2010, Vol. 12, No. 1
K. Stamers et al.SCIENTIFIC ARTICLES
Table 3. Response of both side salivary glands after stimula-
tion with lemon juice (counts/sec).
Fig. 3. Maximal counts per second in accumulation phase (counts/sec) in test (ligature of a.
carotis communis dxt) and control sides
of those rabbits died from suppurative complications
in early postoperative period. On 6 rabbits thrombosis
of common carotid artery was not extensive and after
resection the end-to-end anastomosis was possible;
these rabbits survived and 28 days later sialoscinti-
graphy (Fig. 2) was performed on them, then the rab-
bits were sacrificed under general anesthesia.
After intravenous injection of Tc99 in salivary
glands on the side of a. carotis communis ligature, small
accumulation of radioactive isotope (mean 0.14 counts/
sec, SD 0.04) was observed; while on the healthy side
the glands accumulated isotope (mean 0.48 counts/sec
SD 0.12) 2–3 times more (Table 1 and Fig 3). Compar-
ing mean values with t test the result was P<0.01
The mean difference in Tc99 accumulation be-
tween the test and the control side was 0.34 counts per
sec with SD 0.12.
The duration of isotope accumulation was differ-
ent for the test and the control side. On the control side
the maximum of accumu-
lation was reached in 5
minutes on average (SD
was 1.16), but on the test
side it was reached only
in 14 minutes on average
(SD was 4.18); in two
cases the graphic repre-
sentation was constant
with lemon juice, the re-
sponse in salivary glands
of both sides showed fast
excretion, the mean be-
ing –0.42 counts per sec
(SD was 0.17) on the
control side, and small secretion +0.15 counts per sec-
ond (SD was 0.07) on the ligature side (Table 3).
After intravenous injection of Tc99, accumulation
of isotope in salivary glands on the side of a. carotis
communis ligature was on average 0.14 counts/min.,
SD 0.04. While salivary glands on the healthy side
accumulated Tc 99 on average 0.48 counts/min., SD
0.12. The maximum of Tc99 accumulation on the
healthy side was reached on average in 5 minutes, but
on the test side – on average in 14 minutes.
On 6 rabbits 28 days after revascularization the
sialoscintigraphy showed changes in functions as com-
pared to the glands only with ligatures (Fig 4). During
the accumulation phase, the control side showed the
maximal accumulation in the 6th minute (SD 0.75),
but the reconstructed (test) side reached the maximum
mean accumulation in the 13th minute (SD 1.21)
(Table 4). Statistical significance for revascularized
values was P<0.001.
Comparing the differences in accumulation be-
tween both sides, they were quite similar, even the test
side reached a higher accumulation than the control
side (Table 5).
The mean difference in accumulation between the
test and the control side was 0.12 counts/per sec with
SD 0.13. The revascularized glands can slowly accu-
mulate isotope, but the accumulation remains spastic
until the end of examination.
There is a close relationship between function of
autonomous neural system and blood supply in sali-
vary secretion. Salivary secretion is an active two-stage
process. Primary secrete produced by acini has ionic
concentration similar to blood plasma (22,23). Con-
tinuous stimulation of the parasympathetic ramus com-
Stomatologija, Baltic Dental and Maxillofacial Journal, 2010, Vol. 12, No. 131
SCIENTIFIC ARTICLES K. Stamers et al.
Fig. 4. Maximal counts per second in accumulation phase (counts/sec)
in test (revascularization of a. carotis communis dxt) and control sides
municans of mandibular ganglion increased salivary
gland arterial inflow and salivary secretion at low and
moderate levels of parasympathetic stimulation not
affected by reduction or cessation in arterial inflow,
whereas the response to high level parasympathetic
stimulation was significantly alleviated if blood flow
to the gland was maintained at a level less than that of
the resting arterial inflow (12). Clamping of carotid
artery in rats reduced interstitial fluid pressure and glan-
dular blood flow (–56.5±8.4% and –53.1±6.4 respec-
tively, whereas clamping of jugular vein decreased
glandular blood flow (21.6±14.3%) and increased in-
terstitial fluid pressure (141.2±27.4%) (24).
Different methods in experiments are used to cre-
ate carotid artery stenosis and occlusion. Tsuruta used
rotation of balloon catheter/guidewire system inserted
retrogradely from the external carotid artery into the
common carotid artery and got homogenous stenoses
due to intimal hyperplasia (25). Reduction of regional
blood flow by intracarotid infusion of vasoconstrictor
peptide endothelin-1 (ET-1) which acts directly on vas-
cular smooth muscle cells diminished blood flow of sub-
mandibular salivary glands of anesthetized sheep by
48±4% and the flow of saliva to parasympathetic
chorda – lingual stimulation by 50±1% due to diminu-
tion in the output of Na+, K+, and protein in saliva. Flow
of saliva consistently amounted to 10% of the blood
flow before, during and after the infusion of endothelin-
1 (26). Another group reported on reduction of
blood flow through submandibular gland also
in sheep by 56±5% and salivary secretion by
44±6% after intracarotid infusion of ET-1 (27).
In anesthetized cat’s intracarotid infusion of ET-
1 reduced blood flow through submandibular
gland by 64±7% (28). Withdrawal of blood also
was used as a method to diminish blood flow
in salivary glands (29). Submandibular secre-
tor response to stimulation of parasympathetic
chorda – lingual nerve in anaesthetized cats
before, during and after withdrawal of about
50% blood resulted in significant reduction in
submandibular blood flow and the secretion of
both saliva and protein during stimulation. Un-
der each set of conditions the flow of saliva was
linearly related to the blood flow through the gland.
Experimental pathology of salivary glands more
often was created through ligation of main salivary
ducts. In some of these experiments the vascular re-
sponse of salivary was also examined. On cat’s sub-
maxillary glands the increase in blood flow following
a 10-sec stimulation of the chorda-lingual nerve was
reduced to about 75% of control values (5).
The role of microcirculation is very important in
all fields of surgery, especially in free flaps (30). Also
the submandibular glands transferred for treatment of
ceratoconjungtivitis sica show tracer uptake and ex-
cretion in scintigraphic findings. Consequently, the
glandular tissue is able to function in compromised
In our experiment we used ligature of common
carotid artery that can create a condition close to the
acute occlusive diseases and ligature of common ca-
rotid artery clinically and in human pathology. No neu-
rological or general symptoms, or behavioral changes
in rabbits were observed.
There was a large difference between the
sialoscintigraphy data from the side with ligature of
common carotid artery and the opposite control side
regarding the uptake rate of Tc99 and the washout frac-
tion as well. The reconstruction of closed common
carotid artery 4 weeks after ligature resulted in increase
of functional activity in salivary glands with slower
yet higher accumulation of isotope.
It can be concluded that the functional
activity of rabbit salivary glands tested by
scintigraphy with Tc99 four weeks after liga-
ture of common carotid artery is strongly de-
pressed. Small activity shows that four weeks
after decrease of arterial blood supply on the
test side some salivary gland tissue is still
functioning. Four weeks after restoration of
blood flow in common carotid artery sali-
Table 5. Maximal level of
counts per second in accu-
mulation phase (counts/sec)
in test and control sides after
Table 4. Time of maximal accumulation of
Tc99 (min) after revascularization
32 Stomatologija, Baltic Dental and Maxillofacial Journal, 2010, Vol. 12, No. 1
Received: 13 10 2009
Accepted for publishing: 26 03 2010
K. Stamers et al. SCIENTIFIC ARTICLES
1. American Heart Association Statistics Committee and Stroke
Statistics Subcommittee. Heart diseases and stroke statistics –
2007 update. Circulation 2007;115:69-171.
2. Elverdin JC, Chiarenza AP, Frid AB, Giglio MJ. Effects of
chronic hypoxia on the secretory response of rat salivary
gland. Arch Oral Biol 1995;40:459-62.
3. Takahashi H, Tezuka F, Fujita S, Okabe H. Vascular changes
in major and lingual minor salivary glands in primary
Sjogren’s syndrome. Anal Cell Pathol 1995;9:243–56.
4. Carpenter GH, Osailan SM, Correia P, Paterson KP, Proctor
GB. Rat salivary gland ligation causes reversible secretory
hypofunction. Acta Physiol(Oxf) 2007;189:241-9.
5. Darke AC, Smaje LH. The effect of chronic duct ligation on
the vascular and secretory responses of the cat’s submaxil-
lary gland. J Physiol 1973;228:361-76.
6. Saito T, Fukuda H, Horikawa M, Ohmori K, Shindoh M,
Amemiya A. Salivary gland scintigraphy with 99mTc –
pertechnetate in Sjogren’s syndrome: relationship to clini-
copathologic features of salivary and lacrimal glands. J Oral
Pathol Med 1997;26:46–50.
7. Hermann GA, Vivino FB, Goin JE. Scintigraphic features of
chronic sialadenitis and Sjogren’s syndrome: a comparison.
Nucl Med Commun 1999;20:1123–32.
8. Umehara I, Yamada I, Murata Y, Takahashi Y, Okada N,
Shibuya H. Quantitative evaluation of salivary gland scin-
tigraphy in Sjogren’s syndrome. J Nucl Med 1999;40:64–9.
9. Aung W, Murata Y, Ishida R, Takahashi Y, Okada N, Shibuya
H. Study of quantitative oral radioactivity in salivary gland
scintigraphy and determination of the clinical stage of
Sjogren’s syndrome. J Nucl Med 2001;42:38–43.
10. Booker J, Howarth D, Taylor L, Voutnis D, Sutherland D.
Appropriate utilization of semi-quantitative analysis in sali-
vary scintigraphy. Nucl Med Commun 2004;25:1203–10.
11. Valdes Olmos RA, Keus RB, Takes RP, van Tinteren H, Baris
G, Hilgers FJ, Hoefnagel CA, Balm AJ. Scintigraphic as-
sessment of salivary function and excretion response in ra-
diation – induced injury of the major salivary glands. Can-
12. Liem IH, Olmos RA, Balm AJ. Evidence for early and per-
sistent impairment of salivary gland excretion after irradia-
tion of head and neck tumours. Eur J Nucl Med
13. Kosuda S, Satoh M, Yamamoto F, Uematsu M, Kusano S.
Assessment of salivary gland dysfunction following
chemoradiotherapy using quantitative salivary gland scin-
tigraphy. Int J Radiat Biol Phys 1999;45:379–84.
14. Raza H, Khan AU, Hameed A, Khan A. Quantitative evalu-
ation of salivary gland dysfunction after radioiodine therapy
using salivary gland scintigraphy. Nucl Med Commun
15. Yoshimura Y, Morishita T, Sugihara T: Salivary gland func-
tion after sialolithiasis: scintigraphic examination of subman-
dibular glands with 99mTc-pertechnetate. J Oral Maxillofac
16. Urita Y, Domon K, Yanagisawa T, Ishihara S, Hoshina M,
Akimoto T,Kato H, Hara N, Honda Y, Naigai Y, Nakanishi
K,Shimda N,Takano M, Watanabe T,Sugimoto M, Miki K.
Salivary gland scintigraphy in gastro-esophageal reflux dis-
ease. Inflammopharmacology 2007;15:141-5.
17. Mateos JJ, Setoain X, Ferre J. Salivary scintigraphy for as-
sessing the protective effect of pilocarpine in head and neck
irradiated tumors. Nucl Med Commun 2001;22:651–6.
18. Kohn WG, Ship JA, Atkinson JC, Patton LL, Fox PC. Sali-
vary gland 99mTcscintigraphy: a grading scale and correla-
tion with major salivary gland flow rates. J Oral Pathol Med
19. Firat F, Cermik TF, Sarikaya A, Berkarda S. Effects of gen-
der and age on the quantitative parameters of [99mTc]
pertechnetate salivary gland scintigraphy in normal subjects.
Nucl Med Commun 2006;27:447–53.
20. Loutfi I, Nair MK, Ebrahim AK. Salivary gland scintigra-
phy: the use of semiquantative analysis for uptake and clear-
ance. J Nucl Med Technol 2003;31:81–5.
21. Stamers K, Ratniece M, Skagers A, Pastars K. Scintigraphy
of salivary glands after ligation of common carotid artery in
experiment. Acta Chir Latviensis 2006;6:46-9.
22. Lung MA. Autonomic nervous control of venous pressure
and secretion in the submandibular glands of anesthetized
dogs. Am J Physiol Gastrointest Liver Physiol 1998;275:331–
23. Lung MA. Variations in blood flow on mandibular glandular
secretion to autonomic nervous stimulation in anaesthetized
dogs. J Physiol 1990;431:479–93.
24. Berggreen E, Wiig H, Heyeraas KJ. Blood flow and intersti-
tial fluid pressure in the rat submandibular gland during
changes in perfusion. J Dent Res 2003;82:899-902.
25. Tsuruta W, Yamamoto T, Suzuki k, Yoshida F, Matsumura A.
Simple new method for making a rat carotid artery post-
angioplasty stenosis model. Neurol Med Chir (Tokyo)
26. Thakor AS, Brown CN, Edwards AW. Effects of prolonged
reduction of blood flow on submandibular secretory func-
tion in anesthetized sheep. J Appl Physiol 2003;95:751-7.
27. Harrison AP, Cunningham ME, Edwards AV. Effects of
endothelin on submandibular salivary responses to parasym-
pathetic stimulation in anaesthetized sheep. Auton Neurosci
28. Rourke K, Edwards AV. Submandibular secretory and vascu-
lar responses to stimulation of the parasympathetic inervation
in anesthetized cats. J Appl Physiol 2000;89:1964–70.
29. Hanna SJ, Brelen ME, Edwards AV. Effects of reducing sub-
mandibular blood flow on secretory responses to parasym-
pathetic stimulation in anaesthetized cats. Exp Physiol
30. Hölze F, Loefflbein DJ, Nolte D, Wolff KD. Free flap moni-
toring using simultaneous non-invasive laser Doppler
flowmetry and tissue spectrometry. J Craniomaxillofac Surg
31. Jacobsen HC, Hakim SG, Lauer I, Dendorfer A, Wedel T,
Sieg P. Long-term results of atologous submandibular gland
transfer for the surgical treatment of sever keratoconjunc-
tivitis sicca. J Craniomaxillofac Surg 2008;36:227-233.
vary glands on the experimental side accumulation of
isotope is slow, but the level of accumulation is higher
than on the control side.
The ligature and reconstruction of a common ca-
rotid artery on rabbits confirm the important role of
the arterial blood supply in functional activity of sali-
vary glands and may be an appropriate experimental
model for investigation of ischemic disease of sali-
This study was supported by European Social Fund.