Secretion from minor salivary glands following ablation of the major salivary glands in rats

Since minor salivary glands are tiny and dispersed, ductal cannulation cannot be used when studying their function. The present study was devised to develop a method of measuring minor salivary gland function by excision of the major glands. Female rats (230–280 g) were anaesthetized with sodium pentobarbital. Ablation of the submandibular, sublingual and parotid glands was performed through a sagittal neck incision. Sham-operated rats served as controls. Groups of sialadenectomized animals were investigated immediately and after 1 week, 2 weeks and 3 months. To study secretory function, the mouth was rinsed with 250 μl water in every 5 min and protein and amylase concentrations were measured. After an initial 50 min of basal secretion pilocarpine (1 mg/kg, i.p.) was given. Bilateral ablation of both submandibular, sublingual and parotid glands led to a moderate loss of body weight and a considerable increase in water intake. No other obvious abnormality was observed for periods up to 90 days following surgery. We deduce that the minor glands secrete approximately 14% of protein and 1% of amylase in whole saliva. Secretion is maintained even after 90 days following removal of the major glands. Surgical removal of the major salivary glands allows the secretory function of the minor glands in rats to be studied in vivo.     

Differences in the regulatory mechanism of amylase release by rat parotid and submandibular glands

It is not known whether the mechanisms involved in amylase release in submandibular and parotid glands are similar. Here, the participation of different signalling pathways in amylase release by the parotid and submandibular glands of the male rat was compared by studying the secretory response after beta-adrenergic stimulation. The beta-adrenergic agonist isoproterenol induced an increase of cAMP in both salivary glands, but while in the parotid it triggered amylase release, in the submandibular it was unable to increase amylase secretion. Parotid amylase release was dependent on adenylate cyclase activation, as SQ-22536 inhibited the secretory effect. In contrast, submandibular amylase secretion did not depend on the intracellular concentration of cAMP, as SQ-22536 did not modify its secretory response. Moreover, other activators of adenylate cyclase, such as forskolin and prostaglandin E2, also failed to modify amylase release by the submandibular gland. Neither ionophores nor calcium-blocking agents, as well as calcium-calmodulin and nitric oxide synthase inhibitors, were effective in modifying basal amylase release by the submandibular gland. However, the disruption of microfilaments with cytochalasin B, but not the disruption of microtubules with colchicine, prevented amylase release in that gland. It is concluded that amylase exocytosis in the submandibular gland is a constitutive non-regulated phenomenon, as it is independent of extracellular or intracellular signals. It depends only on the integrity of the microfilaments, probably used by the vesicles to travel from the Golgi apparatus to the plasma membrane.     

Changes in amylase activity in submandibular salivary glands of adult male mice following castration.

The right and left submandibular salivary glands of 25 adult, castrated male Swiss Webster mice, aged 20–30 weeks, were removed for determination of amylase activity and for comparison with the corresponding glands from 28 control mice. The amylase activities in the glands of the intact and castrated animals did not differ significantly until the animals were 28–30 weeks old; then activity was greater in the castrated animal. In the control animals, there appeared to be an age-dependent rate of amylase synthesis, which was possibly disturbed by castration. It is suggested that castration disclosed the contributing, or compensating, influence of one or more regulatory factors, other than testosterone, pertaining to the synthesis of amylase.     

A biochemical analysis of parotid and submandibular salivary gland function with age after simultaneous stimulation with pilocarpine and isoproterenol in female NIA Fischer 344 rats.

This analysis of physiological, biochemical and molecular changes related to aging was made in 3-, 12- and 24-month-old rats. The salivary gland weight/body weight ratio and the structural membrane proteins did not change with age for either gland, but a significant age-related decline in DNA synthesis for both glands was detected, unrelated to the hormonal responsiveness at the level of the plasma membrane. There was a marked increase in the concentration of soluble proteins in adolescent parotid gland and, for the two older age groups, in submandibular gland. The saliva flow rate was different when expressed as volume per time, as volume per time and g glandular wet weight, and/or kg body weight. The concentration of secreted proteins was not affected by age in either gland. The total amount of proteins secreted over 30 min revealed no age-related perturbation for the parotid gland, but showed a significant age-related increase in submandibular saliva. Sodium dodecyl sulphate-polyacrylamide gel analysis revealed changes in the protein bands between 39 and 50 kDa in the Coomassie blue-stained gels from 12-month-old animals. Amylase showed an initial increase (12 months), followed by a marked decline in its activity in parotid saliva. The glandular supernatant had low residual cellular amylase activity after stimulation. Therefore, secretory impairment with age after pilocarpine-isoproterenol stimulation was excluded. Analysis of total RNA showed a pronounced decrease of amylase mRNA in the parotid gland between 12 and 24 months of age. No amylase mRNA was expressed in any of the submandibular samples. For epidermal growth factor, total saliva showed a decrease with age. It seemed that the submandibular gland followed the same picture with age as the parotid gland, with a specific decline in the biosynthesis of single secretory proteins.     

Altered glycogen metabolism in the submandibular and parotid salivary glands of rats with streptozotocin-induced diabetes

Experimental animal models of diabetes induced either by alloxan or streptozotocin have been used to study aspects of the pathophysiology of this disease. The purpose of this study was to examine the metabolism of glycogen in the submandibular and parotid salivary glands of diabetic rats. Diabetes was induced by an intraperitoneal injection of streptozotocin. Eight weeks after the induction of diabetes, the animals were sacrificed and the submandibular and parotid salivary glands were removed. The glands were analyzed for glycogen concentration, and activities of glycogen synthase and phosphorylase. Although the diabetic rats consumed more food than controls, they had a lower body weight eight weeks after diabetes induction. Glycogen concentration in the submandibular and parotid glands increased by about 27% and 130%, respectively. Glycogen phosphorylase a in the submandibular gland of diabetic rats showed a reduction of between 75% and 68% compared with controls. In parotid glands, phosphorylase a was reduced by between 84% and 79% compared with controls. The increase in the activity of glycogen synthase a (active) varied from 64% to 130% for the submandibular glands and from 75% to 110% for the parotid compared with controls. These results suggest that the diabetic state influences glycogen metabolism in the submandibular and parotid salivary glands of rats.     

Developmental changes in the volumes, protein, and some electrolyte concentrations of male and female rat submandibular saliva secreted in response to methoxamine and pilocarpine

Saliva secreted in response to methoxamine and pilocarpine was collected from the cannulated ducts of the submandibular glands of male and female rats at weekly intervals from two to 10 weeks of age. It was analyzed for volume and for concentrations of protein, potassium, calcium, and inorganic phosphate. Following the collection of saliva, the submandibular glands were removed and weighted. The wet weights of the glands increased substantially up to seven weeks of age and then reached almost plateau values in both sexes. The salivary volumes secreted in response to both agents in both sexes were positively correlated with the gland weights, except that after five to six weeks of age there was no correlation between gland weight and methoxamine-stimulated salivary volume. The concentrations of protein, potassium, and inorganic phosphate were inversely related to the flow rates only at relatively low rates of flow. The concentration of calcium was positively correlated with the protein concentration and was independent of the nature of the stimulus and of sex differences during postnatal development.     

Postnatal changes in calcium and amylase of rat salivary glands, including calcium changes with senescence

Postnatal changes occur in glandular Ca concentration of rat parotid and submandibular glands. At 4 days of age, Ca concentration was low in both glands (only one-third to one-half that of adults) and increased gradually with age. The pattern of change was generally similar for male and female rats, but in submandibular gland, adult levels of 9-10 m-equiv./kg were reached by weaning, whereas for parotid gland, a gradual increase in Ca concentration occurred with adult levels of 9-10 m-equiv./kg reached by 7 weeks of age. The pattern of change was the same whether Ca concentration was expressed per kg wet or dry weight albeit water content changed with age. The changes in Ca concentration of parotid paralleled the age-associated increases in amylase activity of parotid gland. Amylase activity of submandibular gland was much less than that of parotid and similarly low at all ages examined, and did not parallel the age-associated increases in Ca concentration. The regulatory role of the sympathetic innervation on glandular Ca concentration was examined by effecting surgical denervation of parotid and submandibular glands at 8 days of age, and then determining Ca concentration of the denervated glands at 32 days. A three-fold increase in Ca concentration, similar to that following acute sympathectomy in adults, occurred in submandibular gland but no change was seen in parotid. An unexpectedly high concentration of Ca was also found in submandibular (but not parotid) gland of old rats.     

Epidermal growth factor in the submandibular salivary glands of congenitally athymic mice.

The submandibular salivary glands of a group of congenitally athymic (“nude”) mice were assayed for their epidermal growth factor (EGF) content and their histology was examined by light microscopy. The ability of the submandibular salivary glands from athymic mice to respond to an androgenic agent was assessed. The histology of the submandibular salivary glands resembled that of normal mice. Athymic mice had concentrations of EGF in their submandibular salivary glands which were similar to those reported previously for normal mice As in normal mice, male athymic mice had more prominent granular convoluted tubules than female mice, but, as in normal mice, testosterone treatment of female athymic mice resulted in an increase in both the EGF content and in the number of granules within the cells of the granular tubules of the duct system of the submandibular salivary gland.These results provide no evidence for a relationship between the level of submandibular salivary gland EGF and the immunological deficiency of nude mice, and show that the abnormalities in development of nude mice do not extend to their submandibular salivary glands.     

Transplantation of a reverse facial artery pedicled submandibular gland in a dog

This study was performed to evaluate the possibility of using reverse facial artery pedicled submandibular gland transfer for treating keratoconjunctivitis sicca. A reverse facial artery pedicled submandibular gland, including the duct, from a beagle dog was transplanted into the temporal region in the same animal. 99mTc pertechnetate scintigraphy was used to monitor graft viability, and the grafted glands were examined histologically 12 weeks after transplantation. Postoperative 99mTc pertechnetate scintigraphy demonstrated viable salivary gland tissue in the transplanted region. Histologically, some of the acinar cells in the graft had atrophied. The transplantation of a reverse facial artery pedicled submandibular gland was successful and may be a simple, reliable treatment for patients with keratoconjunctivitis sicca.     

Effect of feeding different levels of corn oil on the composition of submandibular salivary gland fatty acids in rats

Male, weanling rats were fed synthetic diets containing 0, 3, 10 and 20 per cent corn oil. Rats were killed after 3 weeks, their plasma and submandibular salivary glands were extracted for lipids, and the fatty acid composition was determined. There were significant changes in the fatty acid composition of both plasma and submandibular salivary glands as a result of feeding different levels of corn oil in the diet. The proportions of monoenoic fatty acids (16:1, 18:1) decreased whereas those of linoleic acid increased as the level of corn oil was increased from 0 to 20 per cent. Arachidonic acid level was the lowest in the fat-free group as compared with the other groups. Plasma and submandibular salivary gland lipids of rats fed 0 per cent corn oil in their diet contained another fatty acid which appears to be eicosadicnoic fatty acid. The changes in submandibular salivary gland fatty acids were correlated with the changes in plasma fatty acids. Fractions of neutral lipids and phospholipids which were isolated from the submandibular salivary glands showed changes in their fatty acid composition similar to those observed in the total lipid extract.     

Epidermal growth factor concentration in hyperplastic and hypertrophic submandibular salivary glands of mice.

Submandibular salivary glands of mice contain high concentrations of epidermal growth factor (EGF). The EGF content of mouse submandibular salivary glands undergoing hyperplastic and hypertrophic changes was measured and compared to that in the glands of control mice. Salivary gland hyperplasia was induced by giving mice a single injection of isoprenaline and hypertrophy was produced either by repeated, daily injections of isoprenaline, repeated amputation of a lower incisor tooth or by removing the right submandibular salivary gland and thus producing a compensatory hypertrophy of the left submandibular gland. The EGF content of the hyperplastic submandibular salivary glands was not different from that of the control glands. While the EGF content of the hypertrophied glands resulting from either repeated isoprenaline injections or partial sialoadenectomy did not differ from that of the control glands, the concentration of EGF was significantly lower. This reduced concentration is probably a reflection of acinar hypertrophy with a resultant smaller proportionate contribution of the granular tubules to the mass of the gland. Incisor-amputation-induced hypertrophy did not result in a reduced concentration of EGF in the submandibular salivary glands, but the reason for the different response is unknown.The findings provide no evidence for the involvement of EGF in the induced changes of submandibular salivary gland hyperplasia or hypertrophy resulting from either isoprenaline treatment or partial sialoadenectomy. The reason for the higher concentration of EGF in hypertrophied submandibular salivary glands resulting from incisor amputation compared to that measured in hypertrophied glands resulting from the other stimuli used remains unresolved.     

Mechanisms of increase in amylase activity in rat submandibular and sublingual glands after administration of pilocarpine

The increase in parotid rather than pancreatic-type amylase activity in the submandibular and sublingual glands of rats caused by administration of pilocarpine was abolished or diminished when pilocarpine was injected into rats which had been parotidectomized, sympathectomized by superior cervical ganglionectomy or pretreated with reserpine. These results suggest that the increases in amylase activity in the submandibular and sublingual glands by pilocarpine are not due to increase in enzyme synthesis, but to uptake of enzyme released into the blood in large quantities from the parotid gland and that the release from the parotid gland by pilocarpine is primarily mediated by sympathetic nerves.     

下颌下腺涎石病伴发黏液表皮样癌1例报告

黏液表皮样癌是最常见的唾液腺恶性肿瘤之一,最常发生于腮腺（约占2/3）,其次为腭部、小唾液腺（特别是磨牙后腺）及下颌下腺,而发生于由涎石病导致的慢性炎性下颌下腺者极为罕见。本文报告1例发生于慢性炎性下颌下腺中的黏液表皮样癌病例,并对其诊断和治疗方法进行讨论。     

The effect of alloxan diabetes and insulin in vivo on peroxidase activity in the rat submandibular gland.

Protein content and peroxidase activity were determined in submandibular salivary gland homogenates from control and alloxan-diabetic rats. Fourteen days after alloxan administration, peroxidase activity and protein in the glands of diabetic rats were lower than in control rats. Stimulation of salivary secretion with pilocarpine produced a decrease in the difference between the peroxidase activities in the glands from control and diabetic animals. Insulin treatment of alloxan-diabetic animals produced an increase in submandibular gland peroxidase activity to control levels within 3 hours.     

Activity, distribution and regulation of phosphofructokinase in salivary gland of rats with streptozotocin-induced diabetes

Although the influence of diabetes on salivary glands is well studied, it still presents conflicting results. In this work, the regulation of the phosphofructokinase-1 enzyme (PFK-1) was studied utilizing the salivary glands of rats. Diabetes was induced by a single intraperitoneal injection of streptozotocin (60 mg/Kg of body weight) in rats (180-200 g). The animals were killed 30 days after the induction of diabetes and the submandibular and parotid salivary glands were used. Hyperglycemia was evaluated by blood sugar determination. The distribution of PFK-1 between the soluble and cytoskeleton fractions, the phosphate content of PFK-1, the content of fructose-2,6-bisphosphate and the activity of the PFK-2 enzyme were determined. The calculated relative glandular weight showed a higher value for the parotid gland in comparison with the control, but not for the submandibular gland. The activity of PFK-1 expressed per gland showed no variation between diabetic and control animals. However, considering the specific activity, the soluble enzyme presented a value 50% higher than that of the control and the cytoskeleton bound form increased by 84% compared to the control. For the parotid gland, no difference in the specific activity between diabetic and control animals was observed. On the other hand, the activity per gland of the soluble enzyme increased in the diabetic animals. The phosphate content of PFK-1 increased in the submandibular and parotid glands of diabetic rats. Both the content of fructose-2,6-bisphosphate and the active form of PFK-2 were reduced in the diabetic glands. In conclusion, the increase in the activity of PFK-1 observed in the salivary glands of rats with streptozotocin-induced diabetes does not seem to be due to its modulator fructose-2,6-bisphosphate.     

Physostigmine-induced salivary secretion in the rat

The purpose of this study was to see if physostigmine, a reversible cholinesterase inhibitor, affects the secretion and composition of saliva of the major salivary glands of the rat. Low doses of physostigmine did not elicit secretion. At higher doses there was significant flow from the parotid and submandibular glands within 5 min; however, no sublingual secretion was observed. The submandibular flow rate was highest for the first 5 min, then declined rapidly. The parotid flow rate initially was one-fifth of the maximum submandibular rate and then gradually decreased. The concentrations of Ca, Na and K of physostigmine-induced parotid saliva, and the Na of submandibular saliva, were similar to those with carbachol stimulation. The Ca and K concentrations of submandibular saliva were significantly higher than with carbachol or parasympathetic stimulation, and resembled those of alpha-adrenergic stimulation. The protein concentrations of physostigmine-evoked saliva from both glands were similar. The amylase activity of physostigmine-evoked parotid saliva was much higher than that of carbachol or parasympathetic stimulation. Physostigmine-evoked secretion was completely blocked by atropine, a cholinergic antagonist, and by reserpine, partially blocked by phentolamine, an alpha-adrenergic antagonist and not affected by surgical sympathectomy. Morphologically, physostigmine resulted in a moderate decrease in the number of acinar, but not ductal, secretory granules of both the parotid and submandibular glands, while the sublingual gland was unaffected. Numerous patches of parotid acini also developed vacuoles or vesicles. These results suggest that physostigmine-induced salivary secretion is mediated primarily by direct effects on cholinergic and alpha-adrenergic receptors.     

Glycogen content and activities of enzymes involved in the carbohydrate metabolism of the salivary glands of rats during postnatal development

Carbohydrate metabolism was examined in the developing rat salivary glands by analysing enzymatic activity and glycogen content in the postnatal parotid and submandibular glands. The following enzymes of the carbohydrate metabolism, hexokinase (HK), phosphofructokinase-1 (PFK-1), pyruvate kinase (PK), glucose-6-phosphate dehydrogenase (G6PD), and lactate dehydrogenase (LDH) as well as the content of glycogen were determined in the salivary glands of rats aged 2, 7, 14, 21, 30 and 60 days. The specific activity of HK increased from days 2 to 21 and then it decreased up to 60 days old. The values found for the submandibular glands were from 2.5 to 4.9 times higher than those found for the parotid gland, except for rats aged 60 days. PFK-1 showed a different pattern of variation between the glands. In the submandibular gland there was a statistically significant increase in PFK-1 specific activity from 2 to 30 days of age and then, in the 60 days old group a return to level of the rats aged 2 days. In parotid gland, the specific activity of PFK-1 decreased between 2 and 7 days of age, from 7 to 14 days the specific activity increased markedly and from 14 to 60 days old it gradually decreased. The specific activity of PK followed the same pattern of variation in the submandibular and parotid glands, showing no great variation. The specific activity of LDH decreased from 2 to 60 days old in the submandibular glands. In the parotid glands the mean values for this enzyme were higher for the 2 days old group, and then decreased to remained more or less constant. The potential capacity of the pentose phosphate pathway was greater than that of glycolysis at early ages. The glycogen content showed similar variation in both glands. It was initially high and then decreased. In conclusion, our results on the activities of enzymes involved in carbohydrate metabolism in submandibular and parotid glands may be relevant to the initiation of saliva secretion in these animals.     

Comparison of phospholipid N-methylation activity in rat submandibular salivary gland and liver

Successive phospholipid N-methylation from phosphatidylcholine to phosphatidylethanolamine in submandibular gland and liver microsomes proceeded without the addition of exogenous phospholipid substrate. Methylation activity in the submandibular microsomes showed different susceptibilities to various detergents than the liver enzyme and also partially required Mg2+. However, the three methylation steps could not be distinguished by their Mg2+ requirements. Ca2+ had no effect on the activity. The methylation activity in submandibular gland was much lower than in liver. Chronic administration of isoproterenol, which causes an increase of phosphatidylcholine in membrane phospholipids of salivary glands, decreased methylation activity in the submandibular gland. Thus the increase in phosphatidylcholine in isoproterenol-treated rat salivary glands may not be derived from the phospholipid methylation pathway, but may be due to stimulation of other routes of phosphatidylcholine metabolism.     

一次性18Gy放射对大鼠颌下腺损伤的实验研究

目的:观察一次性18 Gy放射对大鼠颌下腺组织学形态和唾液流率的改变。方法:40只大鼠随机分为实验组和对照组,每组20只。实验组一次性18 Gy局部照射大鼠颌下腺区域,对照组只麻醉不放射。8周后处死所有大鼠,处死前插管法提取大鼠颌下腺唾液,称取其质量,计算唾液流率,比较两组唾液流率的改变。处死后取颌下腺组织,经4%多聚甲醛固定,切片,HE染色,镜下观察颌下腺的组织学形态。结果:放射后实验组进食进水量下降、活动减少。实验组饮水频率高于对照组。对照组的唾液流率为(18.64±8.23)μL/min,实验组的唾液流率是对照组的57.42%,为(10.70±2.22)μL/min。HE染色显示,放射后实验组颌下腺细胞变性,间质血管充血,腺泡细胞内的空泡数量明显增多。结论:放射后大鼠颌下腺唾液流率明显降低,放射后8周,组织学形态出现显著变化。放疗对大鼠颌下腺组织学形态和唾液分泌功能的远期影响,尚需进一步观察和研究。