Salivary Glands of the Rat in a Hot Environment

Elmér , M. and P. Ohlin . Salivary glands of the rat in a hot environment. Acta physiol. scand. 1970. 79. 129–132. A marked hypertrophy of the normally innervated submaxillary gland was observed in rats subjected to heat (34°C) for 3 weeks. The increase in weight of parasympathetically de-xiervated glands was much less while that of sympathetically denervated glands was still pronounced. The size of both synipathetically and parasympathetically denervated glands was not affected in a hot environment. It seems likely that the hypertrophy of the submaxillary gland is to a large extent secondary to an increased secretory activity elicited by the para-sympathetic secretory fibres. A corresponding increase in weight of the sublingual gland was not seen.     

The problem of the effect of parasympathetic denervation on the protein metabolism of the submaxillary salivary glands of the cat

Summary The author studied the influence of unilateral and bilateral section of the chorda tympani on the protein metabolism and the secretory function of the submaxillary salivary glands in cats. The reflex and the paralytic secretion in the denervated glands was absent. The nitrogen content of mucin in the tissues of these glands increased.Changes in protein metabolism and in the secretory activity of the glands were more marked with bilateral denervation. Denervation did not stop secretion and synthesis of protein. Administration of pilocarpine caused typical changes in the salivary secretion and in the content of nitrogenous substances in the saliva and in the tissue of the salivary glands: 1) thelatent period decreased; 2) the maximum quantity of the secreted saliva was changed; 3) after denervation the total amount of saliva secreted in 2 hours decreased (the content of the nitrogenous substances, however, was much higher than before denervation; 4) after pilocarpine the nitrogen content of mucin in the tissues of the denervated glands decreased and the content of protein which was not precipitated with mucin and which is typical of an actively secreting gland increased.Thus, the parasympathetic denervation alters protein metabolism and the secretory function of the submaxillary salivary glands in cats.(Presented by Active Member AMN SSSR V. N. Chernigovskii) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 50, No. 10, pp. 67–73, October, 1960     

Effects of Sympathetic Nerve Stimulation in the Presence of a Slow Parasympathetic Secretion in the Parotid and Submaxillary Glands of the Rabbit

Sympathetic secretory responses from the parotid and the submaxillary glands of the rabbit were studied; sympathetic excitation occurred either alone or during a slow parasympathetic background secretion, imitating the resting secretion normally present. The fairly rapid sympathetically evoked flow of saliva from the parotid gland was, in the background secretion, further increased and obtained at low frequencies, which at sympathetic stimulation alone were subthreshold. The effects, which could be repeated with sympathomimetic drugs in a pilocarpine induced secretion, were abolished by αand βadrenoceptor blockade in combination. The submaxillary gland gave a very scanty secretion on sympathetic excitation, the effects being mediated via β-adrenoceptors. The responses were not increased in a background secretion but the main effect of sympathetic activation was to retard the salivary flow. The retardation was attributed to vasoconstriction accompanying sympathetic excitation. Eliminating the vasoconstrictor responses, while retaining the sympathetic secretory effects, did not reveal any increase of the sympathetic salivary responses in the submaxillary gland.     

The role of salivary glands in the removal of excessive blood plasma norepinephrine

Standard high-performance liquid chromatography was used to study the contents of epinephrine and norepinephrine in rat submaxillary salivary glands, oral cavity mucosa, and in saliva during background and evoked secretion as well as in plasma after parenteral injection of norepinephrine solution. The pilocarpine-induced stimulation of secretory function of the salivary glands increases epinephrine content in the oral cavity mucosa, while in the salivary gland its concentration decreases, although the norepinephrine content remains at the control level. The concentrations of these amines do not vary in saliva, but their release with saliva increases. Presumably, the salivary glands accumulate and release catecholamines during the secretion cycle. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 125, No. 1, pp. 15–17, January, 1998     

Nitric oxide-dependent in vitro secretion of amylase from innervated or chronically denervated parotid glands of the rat in response to isoprenaline and vasoactive intestinal peptide

The basal in vitro release of amylase was similar from rat parotid lobules of innervated and chronically denervated glands and was unaffected by the inhibitors used in this study. The secretion of amylase induced by isoprenaline or vasoactive intestinal peptide (VIP) was reduced by one-third to one-half from the lobules of the innervated glands and even more so from the lobules of the denervated glands by ODQ, an inhibitor of soluble guanyl cyclase which is activated by nitric oxide (NO) and catalyses the cGMP production. The use of N (omega)-propyl-L-arginine (N-PLA) revealed that the evoked secretion of amylase in the denervated glands depended on the activity of neuronal type NO synthase to synthesize NO. Since the denervated gland is virtually devoid of NO synthase-containing nerve fibres, the neuronal type NO synthase was most probably of a non-neuronal source. NO-dependent amylase secretion was agonist related, since amylase secretion evoked by bethanechol and neuropeptide Y was not reduced by ODQ or N-PLA. Hence, under physiological conditions, activation of beta-adrenoceptors (sympathetic activity) and VIP receptors (parasympathetic activity) is likely to cause secretion of parotid amylase partly through a NO/cGMP-dependent intracellular pathway involving the activity of neuronal type NO synthase, possibly of acinar origin.     

The action of physalaemin on electrolyte excretion by the mandibular and sublingual salivary glands of the rat

The effect of physalaemin, an undecapeptide belonging to a family known collectively as the tachykinins, on water and electrolyte excretion of the mandibular and sublingual salivary glands of the rat has been investigated and compared to that of acetylcholine. Drugs were administered intravenously or by close-arterial infusion. Physalaemin is a powerful stimulant of fluid secretion by both glands although less potent than acetylcholine. The Na and K excretion patterns in physalaemin-evoked saliva resembled, but were by no means identical to those evoked by acetylcholine and other parasympathomimetic drugs: saliva evoked by physalaemin was considerably poorer in Na and K at all secretory rates. The HCO₃ excretion curves, on the other hand, seemed to be identical to those evoked by parasympathomimetic drugs. From an analysis of the Na and K excretion patterns, it can be concluded, both for the mandibular and the sublingual glands, that physalaemin stimulates Na reabsorption and K secretion across the gland duct epithelium, whereas acetylcholine has the opposite effect. These findings agree nicely with what has previously been demonstrated in vitro in the isolated perfused main excretory duct of the rat mandibular gland.     

Exocrine and endocrine release of kallikrein after reflex-induced salivary secretion

Exocrine and endocrine release of rat submandibular gland kallikrein has been shown to be low after parasympathetic and beta-adrenergic stimulation but greatly increased after alpha-adrenergic stimulation. In the present study, release of glandular kallikrein was investigated under conditions known to give a reflex-induced salivary gland response. Heat stress induced a rich flow of saliva originating in the submandibular glands. Salivary kallikrein secretory rate was higher than after parasympathetic stimulation but lower than after sympathetic stimulation (P less than 0.005). Only heat stress increased circulating glandular kallikrein (12.7 +/- 0.8 ng ml-1 before heat exposure and 53.3 +/- 14.1 ng ml-1 40 min afterwards, P less than 0.005). There were no indications that the endocrine release of kallikrein was due to non-specific leakage. Atropine abolished heat-induced salivation and endocrine kallikrein secretion, possibly through interference with central pathways (P less than 0.05). However, phentolamine did not, which may indicate as an yet unidentified mediator of endogenous kallikrein release. The salivary gland response to acid and ether was comparable to that observed after parasympathetic nerve stimulation and was abolished by atropine (P less than 0.005). Stimuli known to influence other salivary gland ductal cells, such as aggression and starvation followed by drinking, also did not increase the plasma concentration of glandular kallikrein. The fact that various conditions which induce salivation did not increase circulating glandular kallikrein, coupled with the fact that kallikrein concentration was the highest in animals that died from heat stress, may suggest that the increase in circulating glandular kallikrein seen after heat stress may be pathological and could contribute to the development of heat shock.     

The enteric nervous system participates in the secretory response to the heat stable enterotoxins of Escherichia coli in rats and cats

Intestinal secretion was evoked in periarterially denervated jejunal segments of anesthetized rats and cats by exposing the intestines to the heat stable (ST) toxins from a strain of Escherichia coli producing both STa and STb toxins. The secretion was significantly inhibited and to about the same relative extent by the addition of each one of the three following drugs: hexamethonium (i.v., rats), lidocaine (applied on the serosal surface, rats) and tetrodotoxin (intra-arterial, cats). Atropine inhibited fluid secretion in some experiments. It is proposed that a nervous mechanism is mediating part of the secretory response to Escherichia coli heat stable toxins, since three different drugs, which influence nervous activity in different ways, significantly diminished the secretory response. A model for the secretory nervous reflex(es) within the enteric nervous system is proposed; Escherichia coli heat stable toxins activate a "receptor cell" in the epithelium, which then stimulates surrounding dendritic nerve endings via the release of unknown substance(s). A nicotinic receptor is involved but further characteristics of the nervous reflex(es) remain to be elucidated.     

Measurement of heat production in dog submandibular gland

In dog submandibular glands, we measured changes in metabolic heat production during a secretory period which was induced by electrical stimulation of the chorda tympani. When measured with fine thermocouples, the temperature of both the gland itself and venous blood from the gland increased by 0.1-0.7 degrees C at the maximum. The blood flow was about 0.3 g.min-1.g-1 wet weight of gland at a resting period and increased to 1-3 g.min-1.g-1 at the maximum of secretory response. The resting heat production in vitro, measured by a microcalorimeter, was 3.3 mW.g-1 at 27 degrees C and 7.8 mW-g-1 at 37 degrees C. The specific heat of the blood and the salivary gland, measured by a microcalorimeter, were 3.97 and 3.91 J.g-1, degrees C-1, respectively. The change in conductive heat loss, measured by a thermopile under stimulated conditions, ranged from 4 to 6 mW.g-1 at the maximum. Metabolic heat production during secretion in vivo (37 degrees C) was estimated from temperature changes in the glandular tissue and blood, blood flow, specific heat and resting heat production. Values obtained were in the range from 15 to 80 mW.g-1 at the maximum. Heat production increased rapidly within the first 20 sec to reach a maximum and then declined with time, but slow heat production continued. On the basis of the present results together with those from former studies on ionic transport in the salivary gland, we discussed the relation of heat production to transport of substances during secretory processes.     

Compensatory Hypertrophy of the Rat's Submaxillary Gland

The secretory activity of the submaxillary gland in the conscious rat is increased in the acute experiment when the secretion from one or more of the other salivary glands is prevented from reaching the oral cavity. In chronic experiments the weight of the submaxillary gland is augmented after duct ligation of other salivary glands. It seems likely that increased secretory activity as observed in the acute experiments, is the cause of this glandular hypertrophy.     

Pituitary adenylate cyclase activating peptide (PACAP) in salivary glands of the rat: origin,and secretory and vascular effects

Pituitary adenylate cyclase activating peptide (PACAP)-38. injected Lv. to the anaesthetized rat. evoked secretion of saliva from the three major salivary glands. the submandibular glands responding with the greatest and the sublingual glands with the smallest volumes. The parotid saliva was rich in amylase and protein. In vitro. pieces of parotid and submandibular gland tissues released K⁺ and protein in response to PACAP-38. with atropine and adrenoceptor antagonists present. The blood flow in the submandibular gland increased in response to PACAP-38. despite a marked fall in mean aortic blood pressure. PACAP is a vasoactive intestinal peptide (VIP)-like neuropeptide. A comparison between the two peptides showed PACAP-38 to be more effective than VIP with respect to vascular responses and less or equi-effective with VIP with respect to the secretory responses. thus suggesting the involvement of PACAP type I and type II receptors. respectively PACAP-38 and -27 were present in the parotid gland as judged by radioimmunoassay. the concentration of the former being about twice that of the latter. Parasympathetic denervation. by cutting the auricula-temporal nerve. reduced the total parotid gland contents of PACAP-38 and -27 by 23 and 44%. respectively (compared with a previously demonstrated 95% reduction of VIP). Sympathetic de nervation. section of the facial nerve or treatment with the sensory neurotoxin capsaicin did not affect the content of PACAP. The difference in efficacy between PACAP and VIP in the vascular and secretory responses as well as the difference in localization suggest that the two peptides play different physiological roles in the salivary glands.     

Fractionated irradiation and early changes in salivary glands. Different effects on potassium efflux, exocytotic amylase release and gland morphology.

Irradiation is a potent treatment modality of head and neck cancer. However, the irradiation is usually associated with an influence on salivary glands with ensuing dryness and discomfort for the patients. In the present study we used different in vitro secretory models and morphologic characterization of rat parotid gland. Radiation was given to one gland on a 5-day schedule with 6 MV photons (total dose 20, 30, 35, 40, 45 Gy). The contralateral gland served as control, and the analysis of glands were performed 10 days after the last irradiation treatment. The noradrenaline stimulated electrolyte secretion (86rubidium tracer for potassium) was decreased in relation to the irradiation dose and in comparison to contralateral control glands. Noradrenaline stimulated exocytotic amylase release was not affected by irradiation and, there were no signs of obvious quantitative morphologic alterations after irradiation compared with controls. The results suggest that there are differences in the sensitivity to radiation for the two different secretory processes in salivary glands, and, thus, the structures regulating electrolyte and fluid secretion seem to be more vulnerable to irradiation than the process of exocytosis. The results, however, do not allow discrimination between temporary cellular impairment and irreversible damage leading to cell death.     

Rat salivary gland ligation causes reversible secretory hypofunction

Aim: To determine the influence of inflammation on salivary secretion. Secretion by salivary glands involves interactions between nerves, blood vessels and salivary cells. The present study investigated the effects of inflammation on rat submandibular gland function following acute ductal obstruction. Methods: Under recovery anaesthesia a metal clip was placed on the main duct of the submandibular gland. After 24 h salivary secretion was evoked by nerve and methacholine stimulation. For recovery experiments the clip was removed after 24 h and the animal left to recover for 3 days when salivary function was again assessed. Results: By 24 h of obstruction an inflammatory infiltrate had developed within the obstructed gland and stimulated salivary flows were just 20% of the normal secretion, whilst protein secretion and ion reabsorption were also severely impaired. If ductal obstruction was removed after 24 h the salivary function returned to normal after 3 days of recovery. In vitro analysis of cells from 24‐h ligated glands revealed normal changes in intracellular calcium (the main secondary messenger involved in fluid secretion) in response to methacholine stimulation. Protein secretion from isolated cells indicated some changes in particular to methacholine‐induced protein secretion although a significant protein secretion was still seen in response to isoprenaline – the main stimulus for protein secretion. Conclusion: This report demonstrates reversible salivary inhibition associated with an inflammatory infiltrate within the salivary gland.     

Beta-adrenergic receptors and salivary gland secretion during aging.

Beta-adrenergic signal transduction is primarily responsible for the control of the protein secretions by salivary cells. To examine the relationship between beta-adrenergic signal transduction and beta-adrenergic agonist-stimulated salivary secretion, we simultaneously assessed beta-adrenergic receptor number and pilocarpine-isoproterenol-stimulated salivary flow and secreted proteins in parotid and submandibular glands from 3-, 12- and 24-month-old female NNIA F-344 rats. There were no age-related changes in the density of beta-adrenergic receptors in the parotid gland or in the submandibular gland. In the parotid gland there was a significant increase in saliva flow rate in the oldest age group and no changes in the amount of total proteins secreted over 30 min. However, when normalized to gland weight, flow rate was unchanged and the amount of total secreted proteins decreased with age. In the submandibular gland there were age-related increases in both absolute volume and total secreted protein, but when normalized to gland weight there were no longer changes with age. Changes in flow rate were paralleled by reciprocal changes in protein secretory function such that changes in the salivary protein concentrations for the most part were unchanged with age for both the parotid and the submandibular gland. These parameters were compared to our previous data on adenylate cyclase activity, and collectively, these data suggest that in the submandibular gland salivary secretory function does not correlate with changes in beta-adrenergic receptor density or isoproterenol-stimulated adenylate cyclase activity.     

The involvement of the enteric nervous system in the intestinal secretion evoked by cyclic adenosine 3‘,5’-monophosphate

Intestinal fluid secretion was evoked in vivo in rats and cats by introducing dibutyrylcyclic adenosine 3,5-monophosphate (db-cAMP) or theophylline, a phosphodiesterase inhibitor, in the intestinal lumen. The intestines were denervated periarterially. It was demonstrated that three compounds of varying chemical structure and with different modes of action on nerves (tetrodotoxin, lidocaine, hexamethonium) decreased the secretory response 60–70%. It is concluded that the secretion induced by increasing the intracellular cAMP concentrations is in part evoked via the enteric nervous system     

Some factors influencing stimulation-induced release of potassium from the cat submandibular gland to fluid perfused through the gland

1. The release of K from the cat submandibular gland to the extracellular fluid (ECF) after stimulation with acetylcholine (ACh) and the subsequent uptake of K from the ECF was studied in glands perfused artificially with Locke solutions.2. The first injection of ACh after shift of the perfusion fluid from control to K-free Locke solution evoked a normal loss of K and a normal secretion of saliva. The second injection only evoked a small release of K and a reduced secretion.3. Perfusion with dinitrophenol (DNP) (10(-4)M) containing solutions, Na-free Li Locke solutions and chloride-free nitrate Locke solutions inhibited salivary secretion and the uptake of K. The first injection of ACh after shift of the perfusion fluid from control to test solution gave a normal K loss, but thereafter the ACh-induced K-loss declined.4. Perfusion with g-strophanthin (10(-5)-10(-4)M) always inhibited K uptake whereas K release was not affected primarily. The sensitivity of the secretory mechanism of different glands to strophanthin varied considerably.5. Perfusion with tetraethylammonium Locke solution inhibited secretion, K uptake and release of K.6. It is suggested that the release of K from salivary glands to the ECF after stimulation with ACh can be explained by diffusion as a consequence of an enhanced permeability of the cell membranes to K. Concomitantly with the release of K, Na is taken up. It is suggested that the subsequent uptake of K and extrusion of Na is due to active transport processes probably involving a Na-K activated ATP-ase.     

Immunohistochemical localization of Clara cell secretory proteins (CC10-CC26) and Annexin-1 protein in rat major salivary glands

The oral cavity is continuously bathed by saliva secreted by the major and minor salivary glands. Saliva is the first biological medium to confront external materials that are taken into the body as part of food or drink or inhaled volatile substances, and it contributes to the first line of oral defence. In humans, it has been shown that sputum and a variety of biological fluids contain Clara cell secretory proteins (CC10–CC26). Various studies of the respiratory apparatus have suggested their protective effect against inflammatory response and oxidative stress. Recently, CC10 deficiency has been related to the protein Annexin‐1 (ANXA1), which has immunomodulatory and anti‐inflammatory properties. Considering the defensive role of both Clara cell secretory proteins and ANXA1 in the respiratory apparatus, and the importance of salivary gland secretion in the first line of oral defence, we decided to evaluate the expression of CC10, CC26 and ANXA1 proteins in rat major salivary glands using immunohistochemistry. CC10 expression was found only in the ductal component of the sublingual gland. Parotid and submandibular glands consistently lacked CC10 immunoreactivity. In the parotid gland, both acinar and ductal cells were always CC26‐negative, whereas in the submandibular gland, immunostaining was localized in the ductal component and in the periodic acid Schiff (PAS)‐positive area. In the sublingual gland, ductal cells were always positive. Acinar cells were not immunostained at all. ANXA1 was expressed in ductal cells in all three major glands. In parotid and sublingual glands, acinar cells were negative. In submandibular glands, immunostaining was present in the mucous PAS‐positive portion, whereas serous acinar cells were consistently negative. The existence of some CC10‐CC26–ANXA1‐positive cells in rat salivary glandular tissue is an interesting preliminary finding which could support the hypothesis, suggested for airway tissue, that these proteins have a defensive and protective role. Protein expression heterogeneity in the different portions of the glands could be an important clue in further investigations of their role.     

The involvement of the enteric nervous system in the intestinal secretion evoked by cyclic adenosine 3'5'-monophosphate

Intestinal fluid secretion was evoked in vivo in rats and cats by introducing dibutyrylcyclic adenosine 3,5-monophosphate (db-cAMP) or theophylline, a phosphodiesterase inhibitor, in the intestinal lumen. The intestines were denervated periarterially. It was demonstrated that three compounds of varying chemical structure and with different modes of action on nerves (tetrodotoxin, lidocaine, hexamethonium) decreased the secretory response 60-70%. It is concluded that the secretion induced by increasing the intracellular cAMP concentrations is in part evoked via the enteric nervous system.     

Secretory effects of 5-hydroxytryptamine following neonatal sympathectomy in rat parotid gland.

Unilateral sympathetic denervation of rat parotid glands was performed within 4 h after birth. Nine weeks later the glands were used for in-vitro studies of amylase secretion, and 86Rb+ was used as a marker for potassium efflux. The non-denervated contralateral glands served as controls. The tissue concentrations of 5-hydroxytryptamine and its metabolite 5-hydroxyindole acetic acid were also measured. 5-Hydroxytryptamine caused a significant dose-dependent increase in amylase secretion, which was inhibited by methysergide. There was no difference between controls and denervated glands. 5-Hydroxytryptamine was without effect on potassium efflux from either denervated or control glands. The sympathectomy caused increased levels of 5-hydroxytryptamine and 5-hydroxyindole acetic acid as compared with contralateral controls. The results suggest that 5-hydroxytryptamine influences the two main secretory processes in rat parotid gland differently. A significant amylase discharge was seen following 5-hydroxytryptamine stimulation, whereas no effect was seen on 86Rb+ efflux. Although it is also proposed that there are no 5-hydroxytryptamine-associated nerves in the superior cervical ganglion innervating parotid tissue, it seems that there is a complex connection between the sympathetic pathway and the serotoninergic system.     

Defective signalling in salivary glands precedes the autoimmune response in the non-obese diabetic mouse model of sialadenitis

The spontaneous non‐obese diabetic (NOD) mouse model of Sjögren's syndrome provides a valuable tool to study the onset and progression of both the autoimmune response and secretory dysfunction. Our purpose was to analyse the temporal decline of salivary secretion in NOD mice in relation to the autoimmune response and alterations in various signalling pathways involved in saliva secretion within each salivary gland. A progressive loss of nitric oxide synthase activity in submandibular and parotid glands started at 12 weeks of age and paralleled the decline in salivary secretion. This defect was associated with a lower response to vasoactive intestinal peptide in salivary flow rate, cAMP and nitric oxide/cGMP production. No signs of mononuclear infiltrates or local cytokine production were detectable in salivary glands in the time period studied (10–16 weeks of age). Our data support a disease model for sialadenitis in NOD mice in which the early stages are characterized by defective neurotransmitter‐mediated signalling in major salivary glands that precedes the autoimmune response.