Fluid and protein secretion from ferret submandibular and parotid glands in response to sympathetic nerve stimulation or administration of sympathomimetics.

Electrical stimulation of the sympathetic innervation evoked secretion of submandibular and parotid saliva. By changing the mode of stimulation from a continuous to an intermittent one the fluid response increased and glandular blood flow improved. The volumes from the submandibular glands were larger than those from the parotid glands and further, the protein concentration of submandibular saliva was higher than that of parotid saliva. Adrenaline, isoprenaline and phenylephrine evoked larger fluid responses from submandibular than from parotid glands. However, the fluid response was small compared to the parasympathetic one. Substance P-evoked saliva was used as carrier for protein released by sympathetic nerve stimulation or administration of adrenaline and isoprenaline. In vitro tissues of submandibular and parotid glands responded to adrenaline with a dose-dependent release of protein. Taken together, the analytical pharmacology performed in vivo and in vitro, and including the antagonists phentolamine, dihydroergotamine, propranolol and metoprolol, showed that in submandibular glands, α(α₁)adrenoceptors were predominantly involved in fluid secretion and β(β₁)-adrenoceptors predominantly involved in protein secretion. In parotid glands, fluid secretion seemed solely to depend on α(α₁)-adrenoceptors, while β(β₁)-adrenoceptors seemed almost solely involved in protein secretion.     

Neurokinin A in the parotid and submandibular glands of the rat: immunohistochemical localization and effect on protein and peroxidase secretion.

An indirect immunofluorescence technique was used to study the distribution of neurokinin A immunoreactive (NKA-IR) nerve fibres in submandibular and parotid glands of the rat. The functional role of neurokinin A on protein and peroxidase secretion in these glands was evaluated by using in vitro methods. In the parotid gland neurokinin A immunoreactive fibres were mainly distributed around the secretory acini, but some were also in evidence around the stromal blood vessels and ducts. The number of the neurokinin A immunoreactive nerve fibres was lower in the submandibular gland than in the parotid gland. They were mainly distributed around the secretory acini and stromal blood vessels and ducts. In vitro, neurokinin A significantly stimulated the release of total amount of released proteins and peroxidase from parotid gland fragments, while in the submandibular gland only the release of peroxidase was increased. By using SDS polyacrylamide gel electrophoresis (SDS-PAGE) specific changes were found in the release of proteins after neurokinin A stimulation. The results of the present study demonstrate that neurokinin A immunoreactive nerve fibres are present in the rat parotid and submandibular glands. Their localization around the secretory elements of the glands and the effect of neurokinin A in vitro experiments indicates that neurokinin A might have a significant role in the regulation of salivary secretion.     

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.     

Masticatory-salivary reflexes mobilize noradrenergic,non-cholinergic secretory mechanisms in parotid glands of conscious rats

All rats were maintained on liquid diet, prepared from their ordinary hard pelleted diet, for a week, then fasted 32–33 h and subsequently either fed liquid or pelleted diet for 60–90 min or serving as non-fed controls. Both the fed and the non-fed animals had received atropine and α- and β-adrenoceptor antagonists 15 min before the test. The rats given the liquefied chow consumed about twice as much as those given the hard chow. In the parotid glands of the rats given hard chow the number of acinar granules and the total amylase activity were reduced, by 50 and 70%, respectively, as compared with the glands of control rats. In the parotid glands of the rats given liquefied chow the number of acinar granules and the total activity of amylase remained unchanged. The results suggest that non-adrenergic, non-cholinergic secretory mechanisms of the rat parotid gland participate in masticatory-salivary reflexes.     

Effect of ultrasound on major salivary glands. Dynamic of morphological changes in rat salivary glands

Morphology of rat submandibular salivary glands was examined before and after 10 sessions of ultrasonic treatment focused onto the gonial angle of the mandibular bone. The employed ultrasound protocol induced adaptive reactions and induced no degenerative and inflammatory processes. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 144, No. 11, pp. 586–589, November, 2007     

The effects of antidepressants and pilocarpine on rat parotid glands: an immunohistochemical study

OBJECTIVES:

To evaluate the effects of antidepressants and pilocarpine on the quantity of myoepithelial cells and on the proliferation index of the epithelial cells of rat parotid glands.

INTRODUCTION:

Hyposalivation, xerostomia, and alterations in saliva composition are important clinical side effects related to the use of antidepressants.

METHODS:

Ninety male Wistar rats were allocated to nine groups. The control groups received saline for 30 (group C30) or 60 days (group C60) or pilocarpine for 60 days (group Pilo). The experimental groups were administered fluoxetine (group F30) or venlafaxine for 30 days (group V30); fluoxetine (group FS60) or venlafaxine (group VS60) with saline for 60 days; or fluoxetine (group FP60) or venlafaxine (group VP60) with pilocarpine for 60 days. Parotid gland specimens were processed, and the immunohistochemical expression of calponin and proliferating cell nuclear anti-antigen on the myoepithelial and parenchymal cells, respectively, was evaluated. Analysis of variance (ANOVA), Tukey HSD and Games-Howell tests were applied to detect differences among groups (p<0.05).

RESULTS:

Compared with the controls, chronic exposure to antidepressants was associated with an increase in the number of positively stained cells for calponin. In addition, venlafaxine administration for 30 days was associated with an increase in the number of positively stained cells for proliferating cell nuclear anti-antigen. Fluoxetine and pilocarpine (group FP60) induced a significant decrease in the number of positively stained cells for calponin compared with all other groups.

CONCLUSIONS:

The number of positively stained cells for calponin increased after chronic administration of antidepressants. The proliferation index of the epithelial cells of rat parotid glands was not altered by the use of antidepressants for 60 days.     

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.     

Supersensitivity to substance P and physalaemin in rat salivary glands after denervation or decentralization

Substance P, a putative neurotransmitter in mammals, and physalaemin, present in the skin of an amphibian, are both undecapeptides and belong to the family of tachykinins. The secretory effect of these tachykinins on parotid and submaxillary glands of the rat was examined. Dose-response curves showed that in the unoperated glands maximal secretory responses were obtained to an intravenous dose of 5–10μg/kg of the tachykinins, that the amount of saliva secreted from the submaxillary gland was twice that from the parotid gland, and that physalaemin was more potent than substance P. Parasympathetic denervation of the parotid gland and decentralization of the submaxillary gland caused a marked sensitization to the tachykinins, as judged by lowered threshold doses for secretion and increased secretory responses to a series of submaximal doses 3 weeks postoperatively. Sensitization was less marked after sympathetic denervation and decentralization; in the parotid gland decentralization caused, in fact, no sensitization while in the submaxillary gland the degree of sensitization was about the same after the two types of operation. The tachykinins acted directly on the gland cells and the effect was not exerted via cholinergic, α-adrenergic or β-adrenergic receptors. The pattern of sensitization to the tachykinins, found in the present study, after the different types of operation is similar to that previously found to cholinergic and α-adrenergic agonists and different from that to a β-adrenergic agonist. Studies by others have shown that in the rat parotid gland peptidergic receptors share a common intracellular pathway with cholinergic and α-adrenergic receptors, whereas β-adrenergic receptors use another pathway. In the present study it is suggested that this intracellular arrangement is of importance for the development of supersensitivity.     

Cimetidine-induced androgenic failure causes cell death and changes in actin,EGF and V-ATPase immunoexpression in rat submandibular glands

Submandibular gland (SMG) is responsive to androgens via androgen receptor (AR). We verified whether cimetidine induces androgenic dysfunction in SMG, and evaluated the structural integrity, cell death and immunoexpression of actin, EGF and V-ATPase in androgen-deficient SMG. Male rats received cimetidine (CMTG) and control animals (CG) received saline. Granular convoluted tubules (GCTs) diameter and number of acinar cell nuclei were evaluated. TUNEL and immunofluorescence reactions for detection of AR, testosterone, actin, EGF and V-ATPase were quantitatively analysed. In CG, testosterone immunolabelling was detected in acinar and ductal cells cytoplasm. AR-immunolabelled nuclei were observed in acinar cells whereas ductal cells showed AR-immunostained cytoplasm, indicating a non-genomic AR action. In CMTG, the weak testosterone and AR immunoexpression confirmed cimetidine-induced androgenic failure. A high cell death index was correlated with decreased number of acinar cells, GCTs diameter and EGF immunoexpression under androgenic dysfunction. Actin immunofluorescence decreased in the SMG cells, but an increased and diffuse cytoplasmic V-ATPase immunolabelling was observed in striated ducts, suggesting a disruption in the actin-dependent V-ATPase recycling due to androgenic failure. Our findings reinforce the androgenic role in the maintenance of SMG histophysiology, and point to a potential clinical use of cimetidine against androgen-dependent glandular tumour cells.     

Secretory and Motor Effects in the Submaxillary Gland of the Rat on Intraarterial Administration of Some Polypeptides and Autonomic Drugs

Bradykinin, oxytocin, physalaemin and some autonomic drugs were injected into the common carotid artery. Physalaemin evoked secretion and a pressure rise in the submaxillary duct. A duct pressure rise could be elicited by bradykinin which did not evoke secretion. Autonomic blocking agents did not diminish secretion evoked by physalaemin and did not change pressure responses elicited by bradykinin or physalaemin. Neither secretion, nor duct pressure changes could be recorded after administration of oxytocin. In agreement with previous experiments secretion evoked by autonomic drugs was found to be mediated via cholinergic, α- and β-adrenergic receptors, while motor effects were due to activation of cholinergic and a-adrenergic receptors.     

Recovery of rat submandibular salivary gland function following removal of obstruction: a sialometrical and sialochemical study

Functional recovery of the rat submandibular gland following ligation of the main excretory duct was examined. Rat submandibular glands were ligated for 1, 4 and 8 weeks using a micro-clip with a plastic tube. Micro-clips were removed and glands were allowed to recover for periods of 8, 16 and 24 weeks. Submandibular glands were stimulated with autonomimetic drugs (methacholine and isoprenaline) and salivas were collected from atrophic or de-ligated and contralateral control glands. Glands recovered almost full size (92% of control gland) following 24 weeks of de-ligation. Saliva volume secreted by ligated/de-ligated (RSM) and control (LSM) glands were similar with different doses of agonists. Protein output expressed per gram of tissue wet weight was similar from both ligated/de-ligated and control glands with all doses of agonist. Sodium and chloride levels were higher from de-ligated glands than contralateral control glands. Protein electrophoresis showed similar profiles of salivary proteins in all samples with some minor differences. Acinar cells in de-ligated glands showed a normal morphology, as indicated by light microscopy, whilst granular ductal cells were fewer and contained fewer secretory granules. Sodium potassium ATPase staining of striated ducts in de-ligated glands was similar to that of control glands. It can be concluded that rat submandibular glands can regenerate following severe atrophy and secrete normal amounts of saliva containing broadly a full profile of secretory proteins. In contrast to acinar cells, ductal cells appear not to recover full function.     

Morphological evidence that pentagastrin regulates secretion in the human parotid gland

Salivary secretion is principally regulated by autonomic nerves. However, recent evidence from in vivo animal experiments suggests that gastrointestinal peptide hormones can also influence saliva production. The aim of the present study was to define the secretagogue activity of the gastrin-analogue pentagastrin in human salivary glands. For this purpose, parotid tissues were exposed to pentagastrin in vitro. Morphological techniques were used to evaluate modifications to serous acinar cells associated with secretion. Using a variant of the osmium maceration method, high resolution scanning electron microscopy allowed assessment of the morphology of the cytoplasmic aspect of the plasmalemma to demonstrate secretory activity. To quantify responses to pentagastrin, we recorded morphometric data on microvilli, microbuds, and protrusions. Dose-dependent morphological changes were observed, whereas protein concentration increased in the incubate. The use of selective receptor antagonists showed pentagastrin to act principally via cholecystokinin-A receptors. The morphological responses observed following exposure to pentagastrin differed from those elicited following exposure to the pan-muscarinic agonist carbachol. This study provides the first demonstration of a direct secretory action of gastrointestinal peptides on salivary glands in humans.     

神经激肽A在大鼠空肠发育中的定性定量分析

目的：探讨神经激肽A(NKA)在大鼠空肠发育中的表达及变化。方法：应用免疫组织化学PAP法和图像分析系统研究大鼠胚胎13d至成年空肠中NKA的表达情况。结果：(1)在空肠,首先于胚胎15d的肌间丛处出现NKA-IR的表达,随发育相继出现的纵肌、环肌、绒毛、小肠腺周、粘膜下层、深肌丛,生后30d已具备成年组的分布特征;(2)NKA-IR细胞具有典型的肠道内分泌细胞的形态特征和分布特征。结论：(1)NKA在空肠的发生发育主要在生前1周～生后4周内;(2)NKA在空肠的发生发育有2个关键时期,即生前1周～生后1周和生后1月末;(3)NKA-IR细胞可能是肠道内分泌NKA的内分泌细胞。