New saliva secretion model based on the expression of Na+-K+ pump and K+ channels in the apical membrane of parotid acinar cells.

The plasma membrane of parotid acinar cells is functionally divided into apical and basolateral regions. According to the current model, fluid secretion is driven by transepithelial ion gradient, which facilitates water movement by osmosis into the acinar lumen from the interstitium. The osmotic gradient is created by the apical Cl- efflux and the subsequent paracellular Na+ transport. In this model, the Na+-K+ pump is located exclusively in the basolateral membrane and has essential role in salivary secretion, since the driving force for Cl- transport via basolateral Na+-K+-2Cl- cotransport is generated by the Na+-K+ pump. In addition, the continuous electrochemical gradient for Cl- flow during acinar cell stimulation is maintained by the basolateral K+ efflux. However, using a combination of single-cell electrophysiology and Ca2+-imaging, we demonstrate that photolysis of Ca2+ close to the apical membrane of parotid acinar cells triggered significant K+ current, indicating that a substantial amount of K+ is secreted into the lumen during stimulation. Nevertheless, the K+ content of the primary saliva is relatively low, suggesting that K+ might be reabsorbed through the apical membrane. Therefore, we investigated the localization of Na+-K+ pumps in acinar cells. We show that the pumps appear evenly distributed throughout the whole plasma membrane, including the apical pole of the cell. Based on these results, a new mathematical model of salivary fluid secretion is presented, where the pump reabsorbs K+ from and secretes Na+ to the lumen, which can partially supplement the paracellular Na+ pathway.

A Study on Duration of Effect of Transcutaneous Electrical Nerve Stimulation Therapy on Whole Saliva Flow.

BACKGROUND: Saliva is a complex fluid, whose important role is to maintain the well being of oral cavity. Salivary gland hypofunction or hyposalivation is the condition of having reduced saliva production which leads to the subjective complaint of oral dryness termed xerostomia.(7) Management of xerostomia includes palliative therapy using topical agents or systemic therapy. Electrostimulation to produce saliva was studied in the past and showed moderate promise but never became part of mainstream therapy. Hence, this study was undertaken to evaluate the effect of transcutaneous electrical nerve stimulation (TENS) on whole salivary flow rate in healthy adults and to evaluate how long this effect of TENS lasts on salivary flow. MATERIALS AND METHODS: One hundred healthy adult subjects were divided into five age groups with each group containing 20 subjects equally divided into males and females in each group. Unstimulated saliva was collected using a graduated test tube fitted with funnel and quantity was measured. Transcutaneous electrical nerve stimulation unit was activated and stimulated saliva was collected. Saliva was again collected 30 minutes and 24 hours post stimulation. RESULTS: The mean unstimulated whole saliva flow rate for all subjects (n = 100) was 2.60 ml/5 min. During stimulation, it increased to 3.60 ± 0.39 ml/5 min. There was 38.46% increase in salivary flow. Ninety six out of 100 responded positively to TENS therapy. Salivary flow remained increased 30 minutes and 24 hours post stimulation with the values being 3.23 ± 0.41 ml/5 min and 2.69 ± 0.39 ml/5 min respectively. Repeated measures One way analysis of variance (ANOVA) test showed that the difference between these values were statistically significant. CONCLUSION: Transcutaneous electrical nerve stimulation therapy was effective for stimulation of whole saliva in normal, healthy subjects and its effect retained till 30 minutes and a little up to 24 hours. Transcutaneous electrical nerve stimulation may work best synergistically with other sialagogues and can be used for the management of xerostomia.

The effects of electrostimulation on parotid saliva flow: a pilot study.

OBJECTIVE: Saliva is a critical fluid necessary for oral health. Medications, radiation therapy, and systemic conditions can decrease salivary function and increase a patient's risk for caries and other oral infections. Palliative management of xerostomia includes wetting agents such as ice chips and saliva substitutes. Systemic agents stimulate salivary flow but often have unfavorable side effects. All have met with limited success. The purpose of this study is to assess the effectiveness of transcutaneous electric nerve stimulation (TENS) as a means of stimulating salivary function in healthy adult subjects. STUDY DESIGN: Twenty-two healthy, adult subjects with no history of salivary gland disorder enrolled in the protocol. The TENS electrode pads were placed externally on the skin overlying the parotid glands. Unstimulated saliva was collected for 5 minutes via the Carlson-Crittenden cup into preweighed vials using standardized collection techniques. The TENS unit was then activated and stimulated saliva collected for an additional 5 minutes. RESULTS: Fifteen of 22 subjects demonstrated increased parotid salivary flow when stimulated via the TENS unit. Five experienced no increase and 2 experienced a decrease. The mean unstimulated salivary flow rate was 0.02418 mL/min (SD 0.03432) and mean stimulated salivary flow rate was 0.04946 mL/min (SD 0.04328). Statistical analysis of flow rates utilizing the paired t test demonstrated the difference to be statistically significant, P < .001. In 7 subjects with 0 baseline flow, 5 continued to have no flow. CONCLUSIONS: The TENS unit was effective in increasing parotid gland salivary flow in two-thirds of healthy adult subjects. A further study in a cohort of patients with salivary gland disorders is warranted.

Rab3D is not required for exocrine exocytosis but for maintenance of normally sized secretory granules.

Rab3D, a member of the Rab3 subfamily of the Rab/ypt GTPases, is expressed on zymogen granules in the pancreas as well as on secretory vesicles in mast cells and in the parotid gland. To shed light on the function of Rab3D, we have generated Rab3D-deficient mice. These mice are viable and have no obvious phenotypic changes. Secretion of mast cells is normal as revealed by capacitance patch clamping. Furthermore, enzyme content and overall morphology are unchanged in pancreatic and parotid acinar cells of knockout mice. Both the exocrine pancreas and the parotid gland show normal release kinetics in response to secretagogue stimulation, suggesting that Rab3D is not involved in exocytosis. However, the size of secretory granules in both the exocrine pancreas and the parotid gland is significantly increased, with the volume being doubled. We conclude that Rab3D exerts its function during granule maturation, possibly by preventing homotypic fusion of secretory granules.

The influence of sensory stimulation (acupuncture) on the release of neuropeptides in the saliva of healthy subjects.

In recent studies we have shown that xerostomia (dry mouth) can be treated successfully with sensory stimulation (acupuncture). The increase of saliva secretion lasted often for at least one year. Some neuropeptides have been found to influence the secretion of saliva. The aim of this study was to investigate the mechanisms behind the effect of acupuncture on salivary secretion by measuring the release of neuropeptides in saliva under the influence of sensory stimulation. VIP-like immunoreactivity (VIP-LI), NPY-LI, SP-LI, CGRP-LI and NKA-LI were analysed in the saliva of eight healthy subjects. Manual acupuncture and acupuncture with low-frequency electrical stimulation (2 Hz) were used. The saliva was collected during 20 minutes before the start of acupuncture stimulation, then during 20 minutes while the needles were in situ and then for another 20 minutes after the needles were removed. Four different saliva sampling techniques were used: whole resting saliva, whole saliva stimulated by paraffin-chewing, whole saliva stimulated by citric acid (1%), and parotid saliva, also stimulated with citric acid (1%). The results showed significant increases in the release of CGRP, NPY and VIP both during and after acupuncture stimulation, especially in connection with electro-acupuncture. SP showed only few increases, mainly in connection with electro-acupuncture, whereas NKA generally was unaffected by the acupuncture stimulation. The sensory stimulation-induced increase in the release of CGRP, NPY and VIP in the saliva could be an indication of their role in the improvement of salivary flow rates in xerostomic patients who had been treated with acupuncture.

Further evidence for a hypothalamus-parotid gland endocrine axis in the rat.

The existence of a hypothalamus-parotid gland endocrine axis that stimulates intradentinal dye penetration (IDDP) in rat teeth was suggested in earlier studies and IDDP-stimulating factors were isolated or purified from porcine parotid glands and hypothalamic tissues, respectively. In the present study, infusion of carbamyl-DL-aspartic acid (CAA) into rats was used to demonstrate the role of the endogenous hormones of the hypothalamus-parotid gland endocrine axis in stimulating IDDP, as observed by fluorescence microscopy of longitudinal sections of molar teeth. Intra-arterial infusion of CAA into intact rats stimulated IDDP in a dose-related fashion (between 49-390 nmol/100 g body weight); however, infusion of 390 nmol into parotidectomized rats was ineffective. Infusion of plasma from CAA-treated rats was equally effective in stimulating IDDP in intact and in parotidectomized animals. In contrast, plasma obtained from parotidectomized, CAA-treated rats stimulated IDDP in intact recipient animals but not in parotidectomized ones. Moreover, plasma from adult rats treated with CAA after an electrolytic lesion of the hypothalamus, and infused back into young intact rats, was ineffective in stimulating IDDP. These results indicate that: (1) CAA requires the functional integrity of the parotid gland to express its IDDP-stimulating activity, (2) a hormonal factor is secreted by the parotids in response to CAA stimulation and is directly responsible for IDDP stimulation, (3) release of the endocrine parotid IDDP-stimulating factor after infusion of CAA involves a second endocrine factor that appears to originate from the hypothalamus.(ABSTRACT TRUNCATED AT 250 WORDS)

Biological and chemical evidence for the existence of a porcine hypothalamic parotid hormone-releasing factor.

Methodology has been developed to achieve partial purification of a parotid hormone-releasing peptide from porcine hypothalamus-thalamus tissue using an in vivo parotid hormone stimulation test in pigs and a dentinal fluid transport stimulation test in rats. The purification steps include: acetone-water extraction of the tissue at pH 5.2, ultrafiltration through Amicon PM10 membrane, size exclusion chromatography on Bio-Gel P2, and open-column reversed-phase chromatography on Lichroprep RP8. A 100-fold increase in specific activity was attained. Intravenous infusion of porcine hypothalamus-thalamus extract stimulates a dentinal fluid transport mechanism in teeth of intact anesthetized rats, and the release of plasma immunoreactive parotid hormone in conscious catheterized pigs. Parotidectomy in both species suppresses the response, suggesting that the expression of parotid hormone-releasing activity requires the integrity of a putative hypothalamus-parotid gland endocrine axis.

Vitamin D and parotid gland function in the rat.

1. We previously reported that parotid gland secretion is decreased in rats deprived of vitamin D (Glijer, Peterfy & Tenenhouse, 1985). In the present study we examine whether this effect is a direct result of the absence of vitamin D or due to the secondary systemic effects of vitamin D deficiency. 2. Offspring of rats maintained on a calcium-supplemented (1.2%), vitamin-D-deficient diet were weaned onto the same diet and examined after 8 weeks. Using this method it was possible to maintain serum calcium and parathyroid hormone concentrations within normal limits. Serum 25-hydroxyvitamin D (25(OH)D3) was not detectable, but 1,25-dihydroxyvitamin D (1,25(OH)2D3) concentrations were normal. 3. Pilocarpine-stimulated flow of parotid saliva was reduced 57% in vitamin-D-deprived animals, but amylase secretion was unchanged. Treatment with vitamin D3 returned flow rates to normal. 4. The concentration of calcium in parotid saliva was normal in vitamin-D-deprived rats, although total parotid calcium output was reduced 57%. 5. Pilocarpine-stimulated salivary flow from submandibular gland, a tissue which does not possess 1,25(OH)2D3 receptors, was normal in vitamin-D-deprived rats. 6. Heart rate and arterial blood pressure changes in response to I.V. pilocarpine administration were identical in normal and vitamin-D-deficient rats. 7. Auriculotemporal nerve-stimulated flow of parotid saliva was also reduced by 50% and administration of vitamin D3 to these rats corrected this abnormality. 8. It is concluded that fluid and electrolyte secretion from parotid gland is directly dependent on vitamin D; abnormal parotid gland function seen in vitamin-D-deficient rats is not due to secondary hypocalcaemia or hyperparathyroidism, nor can it be explained by haemodynamic changes evoked during systemic administration of pilocarpine. We further conclude that the metabolite of vitamin D responsible for this effect is not 1,25(OH)2D3.

A comparison of the haemodynamic effects of dietary sodium restriction and acute sodium depletion in the conscious sheep.

The use of a low Na, low K sorghum grain diet supplemented with intraruminal electrolyte infusions has enabled dietary manipulation of sodium status to be studied in the sheep. Dietary sodium restriction reduced urinary sodium excretion within 24 h with maximal retention after 3 days. There were no other substantial metabolic or haemodynamic changes. A more severe form of sodium deficiency produced by parotid salivary drainage resulted after only 2 days in a sodium deficit 3-4 times that seen with 14 days of sodium restriction. Extracellular fluid volume and cardiac output decreased. Blood pressure was unchanged but there was an increase in peripheral resistance and plasma renin concentration.

Adrenocortical function in hypophysectomized dogs with parotid gland transplants in direct contact with the basal hypothalamus.

Adrenocortical activity was studied in three groups of dogs: control-normal, hypophysectomized (hypox), and dogs with a piece of parotid gland grafted into the sella turcica immediately after removal of the whole hypophysis (hypox + graft). Cortisol plasma level (Fk) was estimated by the competitive protein binding method. The mean base-line Fk for 21 normal dogs was 0.8 +/- 0.1 microgram/dl; for 12 hypox dogs 0.03 +/- 0.01 microgram/dl, and for hypox + graft dogs 0.5 +/- 0.2 microgram/dl. After mild or severe stresses the normal and hypox + graft dogs showed an increment in Fk; hypox dogs showed no change. Adrenal glands of hypox dogs revealed striking diminution in fasciculata and reticularis layers, whereas hypox + graft dogs approached normal. Light microscopy studies of the parotid gland graft showed signs of cellular differentiation. Groups of proliferating cells forming follicular structures were present mainly in the part of the parotid tissue closely associated to third ventricle presenting different staining affinities.

A study of parotid salivation in the horse.

1. Saliva flowed from the horse's parotid duct only during mastication.2. The surface-active local anaesthetic administered by mouth inhibited salivary secretion.3. Salivary secretion was stimulated by pilocarpine and inhibited by atropine.4. The volume and composition of saliva secreted in 24 hr from one parotid duct was determined.5. The concentration of sodium, potassium, calcium, chloride and bicarbonate depended upon the rate of flow. The highest concentrations of these electrolytes were observed during periods of high flow rates.6. Horse parotid saliva contained a high concentration of calcium.7. In the absence of a dietary supplement of sodium bicarbonate, the sodium concentration of the saliva fell after about 21 days. There was an associated increase in the potassium concentration. The addition of a sodium supplement restored the sodium concentration of the saliva within 24 hr.