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.

Quantification of the salivary volume flow rate in the parotid duct using the time-spatial labeling inversion pulse (Time-SLIP) technique at MRI: A feasibility study.

BACKGROUND: We developed a method to quantify the volume flow rate (VFR) using the time-spatial labeling inversion pulse (Time-SLIP) technique to evaluate salivary function. PURPOSE/HYPOTHESIS: To investigate the accuracy of quantification of the salivary VFR using the Time-SLIP technique in phantoms and to examine the feasibility of its use in human subjects. STUDY TYPE: This was a prospective phantom and volunteer study. POPULATION/SUBJECTS/PHANTOM/SPECIMEN/ANIMAL MODEL: A phantom and 23 normal volunteers who fasted at least 2 hours study was performed. FIELD STRENGTH/SEQUENCE: Flow images of the phantom and the parotid duct of 23 volunteers were acquired on a 3T-MRI scanner using the Time-SLIP technique. ASSESSMENT: Hypothesizing that flow aggregates in the conducting duct, we measured the VFR on flow images. In the phantom study, the actual VFR (slow, medium, fast flow) was controlled by an automatic pump system and the measured VFR was compared with the actual VFR on flow images. In the human study we injected citric acid into the mouth of healthy volunteers to stimulate saliva secretion and recorded the VFR. STATISTICAL TESTS: As this study was a feasibility study, statistical tests were not performed. RESULTS: In the phantom study, the VFR at slow, medium, and fast flow was 5.7 ± 0.4 (SD), 8.4 ± 0.3, and 12.2 ± 1.1 mm3 /sec, respectively. The error between the measured and actual VFR values was 2.8-3.7%. Salivary flow in the parotid duct was visualized in 22 of the 23 volunteers. The mean VFR was 8760 mm3 /10 min. DATA CONCLUSION: When salivary flow was stimulated with citric acid in normal volunteers, the salivary VFR could be obtained using the Time-SLIP technique. LEVEL OF EVIDENCE: 2 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2018;47:928-935.

Dental microwear textural analysis as an analytical tool to depict individual traits and reconstruct the diet of a primate.

OBJECTIVES: Dental microwear is a promising tool to reconstruct animals' diet because it reflects the interplay between the enamel surface and the food items recently consumed. This study examines the sources of inter-individual variations in dietary habits in a free-ranging population of mandrills (Mandrillus sphinx) using a combination of feeding monitoring and in vivo dental microwear textural analysis (DMTA). METHODS: We investigated the impact of seasonality and individual traits on four DMTA parameters. In parallel, we further studied the influence of the physical properties of the food items consumed on these four parameters, using three proxies (mechanical properties, estimates of phytolith and external grit contents). RESULTS: We found that seasonality, age, and sex all impact DMTA parameters but those results differ depending on the facet analyzed (crushing vs. shearing facets). Three DMTA parameters (anisotropy, complexity, and heterogeneity of complexity) appear sensitive to seasonal variations and anisotropy also differs between the sexes while textural fill volume tends to vary with age. Moreover, the physical properties of the food items consumed vary seasonally and also differ depending on individual sex and age. CONCLUSION: Considering the interplay between the tested variables and both dental microwear and diet, we reaffirm that food physical properties play a major role in microwear variations. These results suggest that DMTA parameters may provide valuable hints for paleoecological reconstruction using fragmentary fossil dental remains.

Intravoxel incoherent motion imaging measurement of perfusion changes in the parotid gland provoked by gustatory stimulation: A pilot study.

PURPOSE: To demonstrate the feasibility of intravoxel incoherent motion imaging (IVIM) for quantification of perfusion changes in the parotid gland after gustatory stimulation. MATERIALS AND METHODS: Eight healthy volunteers underwent diffusion-weighted magnetic resonance imaging (MRI) of the neck at 3T with 11 dynamic acquisitions (9 b-values between 0 and 980 s/mm2 , 2:40 min each). After 5:20 minutes, a lemon-mint-drop was administered orally. Perfusion fraction (Fp ), pseudodiffusion (D*), tissue diffusion (Dt ) coefficients, and optimal b-value threshold were measured using a multistep variable b-value threshold fitting approach. Dynamic changes in the coefficients between three exemplary timepoints (baseline, after stimulation, after dissolution) were compared using a Mann-Whitney U-test with Bonferroni correction (P < 0.016 significance level). RESULTS: Mean values (95% confidence interval [CI]) for IVIM parameters at baseline were Fp : 0.11 (0.08-0.15), D*: 56.48 mm2 /s (39.71-98.27), Dt : 1.01 mm2 /s (0.84-1.06), b-value threshold: 30 s/mm2 (21.25-105). After stimulation: Fp 0.16 (0.15-0.24; P < 0.01), D* 93.83 mm2 /s (77.98-129.53, P = 0.25), Dt 0.93 mm2 /s (0.87-1.08, P = 0.94), b-value threshold 20 s/mm2 (13.75-26.25 s/mm2 , P = 0.10), reflecting the increase in tissue perfusion. After dissolution of the drop: Fp : 0.13 (0.11-0.18, P = 0.38), D*: 101.61 mm2 /s (90.68-144.55, P = 0.07), Dt : 0.91 mm2 /s (0.85-1.05, P = 0.64), b-value threshold: 15 s/mm2 (11.25-40, P = 0.38). CONCLUSION: The IVIM method allows for simultaneous quantification of changes in perfusion and diffusion effects after gustatory stimulation of the parotid gland. LEVEL OF EVIDENCE: 2 J. Magn. Reson. Imaging 2017;45:570-578.

Outpatient versus observation/inpatient parotidectomy: patient factors and perioperative complications.

The objective of this manuscript is to review a single institution's experience with superficial or total parotidectomy in outpatient and observation/inpatient groups. All patients who underwent superficial or total parotidectomy between 2009 and 2015 were identified. Patients were excluded if they had undergone concurrent surgery such as neck dissection, had prior radiation treatment or surgery at the operative site. Main outcomes were perioperative complications in both groups. 215 consecutive patients were included in the study, 116 (54%) patients in the inpatient group and 99 (46%) in the outpatient group. Aside from a higher observed rate of cardiac disease in the outpatient group (24.2 vs. 11.2%, p = 0.014) and larger mean body mass index (BMI) in the inpatient group (32.448 vs. 30.034, p = 0.017), there were no significant differences for age, sex or smoking status. Average operative time differed between groups with 2 h 42 min for inpatients and 2 h 18 min for outpatients (p < 0.001). There were 26 complications in the inpatient group (22.4%, including two hematomas) and 8 in the outpatient group (8.1%). The rate of seroma/sialocele formation was significantly higher in the inpatient group at 15.5% (n = 18) compared with the outpatient group at 3% (n = 3, p = 0.001). Our study shows that parotidectomy, superficial or total, was performed safely as an outpatient procedure without significant increase in complications when compared to patients observed for at least one night after surgery.

Digestive and Nondigestive Functions of Rodents' Salivary Glands.

Major salivary glands play a role not only in digestion, but also in regulation of other functions in rodents. In this review, we analyzed and summarized the data about the rodents' parotid, submandibular and sublingual salivary glands functions, which is not limited to the production of saliva and action of its hydrolytic enzymes on food in the oral cavity. In recent decades significantly expanded understanding of major salivary glands nondigestive functions. They are involved in excretion of metabolic products, maintaining fluid and electrolyte balance. Special attention has been paid to the characteristics of specific (parotin, sialorphin, etc.) and nonspecific (epidermal growth factor, nerve growth factor, kallikrein, etc.) active substances of the major salivary glands and their involvement in wound healing, mineral metabolism, regulation of hematopoiesis and immunity system. Summarized and analyzed major salivary glands endocrine function in the organs and systems. Available literature data suggest: the structure of the major salivary glands, as well as the synthesis and secretion of a number of biologically active substances are controlled by sex hormones. In turn, these biologically active factors of the salivary glands, as epidermal growth factor, and parotin, sialorphin, whose expression is regulated by androgens, have an impact on the morphological and functional state of the gonads. Thus, major salivary glands operate a wide range of functions and involved in the regulation of sexual behavior of reproductive function and maintaining homeostasis in the body.

Activation of muscarinic receptors in rat parotid acinar cells induces AQP5 trafficking to nuclei and apical plasma membrane.

BACKGROUND: The subcellular distribution of aquaporin-5 (AQP5) in rat parotid acinar cells in response to muscarinic acetylcholine receptor (mAChR) activation remains unclear. METHODS: Immunoconfocal and immunoelectron microscopy were used to visualize the distribution of AQP5 in parotid acinar cells. Western blotting was used to analyze AQP5 levels in membranes. To clarify the characteristics of membrane domains associated with AQP5, detergent solubility and sucrose-density flotation experiments were performed. RESULTS: Under control conditions, AQP5 was diffusely distributed on the apical plasma membrane (APM) and apical plasmalemmal region and throughout the cytoplasm. Upon mAChR activation, AQP5 was predominantly located in the nucleus, APM and lateral plasma membrane (LPM). Subsequently, localization of AQP5 in the nucleus, APM and LPM was decreased. Prolonged atropine treatment inhibited mAChR agonist-induced translocation of AQP5 to the nucleus, APM and LPM. AQP5 levels were enhanced in isolated nuclei and nuclear membranes prepared from parotid tissues incubated with mAChR agonist. mAChR agonist induced AQP5 levels in both soluble and insoluble nuclear fractions solubilized with Triton X-100 or Lubrol WX. Small amounts of AQP5 in nuclei were detected using low-density sucrose gradient. When AQP5 was present in the nuclear membrane, nuclear size decreased. CONCLUSION: The activation of mAChR induced AQP5 translocation to the nucleus, APM and LPM, and AQP5 may trigger water transport across the nuclear membrane and plasma membrane in rat parotid acinar cells. GENERAL SIGNIFICANCE: AQP5 translocates to the nuclear membrane and may trigger the movement of water, inducing shrinkage of the nucleus and the start of nuclear functions.

Evaluation of Parotid Glands With Real-time Ultrasound Elastography in Children.

OBJECTIVES: The aim of this study was to determine the strain index for parotid glands in children by using ultrasound elastography. METHODS: In this prospective study, apparently healthy children were referred from the ear-nose-throat clinic to the radiology clinic for elastographic examinations. Conventional sonographic and elastographic examinations of the parotid glands were performed. A linear 5-12-MHz transducer was used to obtain the images. RESULTS: A total of 54 children were enrolled in this prospective study. The normal mean strain index value ± SD for the parotid glands was 1.24 ± 0.67 (range, 0.29-1.39) regardless of sex. The mean age of girls was 7.42 ± 2.94 years (range, 3-14 years), and the age of boys was 8.50 ± 3.46 years (range, 4-16 years). The strain index values for the parotid glands in boys was 1.25 ± 0.76, and in girls it was 1.22 ± 0.55. There was no statistically significant difference in the strain index values between girls and boys (P= .986). There was no correlation between the strain index and age (r = 0.026) or body mass index (r = 0.066). CONCLUSIONS: This study determined the mean strain index values for apparently healthy children. Such information can serve as a baseline from which pathologic parotid diseases can be diagnosed with ultrasound elastography in combination with other sonographic criteria.

The aflatoxin-detoxifizyme specific expression in mouse parotid gland.

The aflatoxin-detoxifizyme (ADTZ) gene derived from Armillariella tabescens was cloned into parotid gland-specific expression vector (pPSPBGPneo) to construct the parotid gland-specific vector expressing ADTZ (pPSPBGPneo-ADTZ). Transgenic mice were generated by microinjection and identified by using PCR and Southern blotting analysis. PCR and Southern blotting analysis showed that total six transgenic mice carried the ADTZ gene were generated. RT-PCR analysis indicated that the expression of ADTZ mRNA could be detected only in parotid glands of the transgenic mice. The ADTZ activity in the saliva was found to be 3.72 ± 1.64 U/mL. After feeding a diet containing aflatoxin B1 (AFB1) for 14 days, the effect of ADTZ on serum biochemical indexes and AFB1 residues in serum and liver of mice were evaluated. The results showed that total protein and globulin contents in the test treatment (transgenic mice) produced ADTZ were significantly higher than that of the positive control, while alanine aminotransferase and aspartate aminotransferase activity in serum of the test treatment (transgenic mice) were remarkably lower compared to that of the positive control (P < 0.05). Moreover, AFB1 residues in serum and liver of the test treatment (transgenic mice) were significantly lower compared with that of the positive control (P < 0.05). These results in the study confirmed that ADTZ produced in transgenic mice could reduce, even eliminate the negative effects of AFB1 on mice.

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.

Short communication: urea transporter protein UT-B in the bovine parotid gland.

Ruminant nutrition relies upon the symbiotic relationship that exists with microbial populations in the rumen. Urea transported across the ruminal epithelia and secreted by the salivary glands is a key source of nitrogen for microbial growth in the rumen. As ruminal urea transport can be mediated by specific UT-B urea transporters, this study investigated whether UT-B urea transporters were also present in the bovine salivary gland. Western blotting experiments detected only small amounts of UT-B protein in whole-cell lysate from the bovine parotid gland. In contrast, strong 32 to 34 and 40 kDa UT-B proteins were detected in parotid plasma membrane-enriched protein, showing the importance of using enriched samples. These signals were also detected in rumen and correspond to bovine UT-B1 and UT-B2 urea transporters, respectively. Further immunolocalization studies identified that these proteins were located in the ductal system of the parotid gland. This study, therefore, confirmed the presence of UT-B urea transporter protein in the bovine parotid salivary gland.

Distinct contributions by ionotropic purinoceptor subtypes to ATP-evoked calcium signals in mouse parotid acinar cells.

There is emerging consensus that P2X4 and P2X7 ionotropic purinoceptors (P2X4R and P2X7R) are critical players in regulating [Ca²âº]i dynamics and fluid secretion in the salivary gland. In contrast, details regarding their compartmentalization and selective activation, contributions to the spatiotemporal properties of intracellular signals and roles in regulating protein exocytosis and ion channel activity have remained largely undefined. To address these concerns, we profiled mouse parotid acinar cells using live-cell imaging to follow the spatial and temporal features of ATP-evoked Ca²âº dynamics and exocytotic activity. Selective activation of P2X7Rs revealed an apical-to-basal [Ca²âº]i signal that initiated at the sub-luminal border and propagated with a wave speed estimated at 17.3 ± 4.3 µm s⁻¹ (n =6). The evoked Ca²âº spike consisted of Ca²âº influx and Ca²âº-induced Ca²âº release from intracellular Ca²âº channels. In contrast, selective activation of P2X4Rs induced a Ca²âº signal that initiated basally and propagated toward the lumen with a wave speed of 4.3 ± 0.2 µm s⁻¹ (n =8) that was largely independent of intracellular Ca²âº channel blockade. Consistent with these observations, P2X7R expression was enriched in the sub-luminal regions of acinar cells while P2X4R appeared localized to basal areas. In addition, we showed that P2X4R and P2X7R activation evokes exocytosis in parotid acinar cells. Our studies also demonstrate that the P2X4R-mediated [Ca²âº]i rise and subsequent protein exocytosis was enhanced by ivermectin (IVR). Thus, in addition to furthering our understanding of salivary gland physiology, this study identifies P2X4R as a potential target for treatment of salivary hypofunction diseases.

Effects of extracorporeal shock-wave lithotripsy directed at the parotid gland on oxidative stress parameters and some trace element levels in facial nerve of rats.

INTRODUCTION: This study was designed to assess the effect of extracorporeal shock-wave lithotripsy (ESWL) exposure of the parotid gland on oxidative stress and some trace element levels in the facial nerves of rats. METHODS: Twelve male Wistar albino rats were divided into two groups, each consisting of 6 animals. The rats in the first group served as controls. The left parotid glands of animals in the second group were treated with 1000 18-kV shock waves while anesthetized with ketamine. The animals in both groups were euthanized 72 h after the ESWL treatment, and the right facial nerve was harvested for determination of oxidant/antioxidant status and trace element levels. RESULTS: Lipid peroxidation product malondialdehyde (MDA) and antioxidant glutathione (GSH) levels increased, and the activities of antioxidant enzymes, such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px), decreased in the facial nerves of ESWL-treated rats. The levels of iron, lead, manganese, and cobalt increased, and magnesium, cadmium, and copper levels decreased. CONCLUSIONS: ESWL treatment of the parotid gland may increase lipid peroxidation and decrease antioxidant enzyme activity in adjacent tissues such as the facial nerve. It also causes a decrease or increase in many mineral levels of the facial nerve, which is an undesirable condition for normal physiological function.

A Stimulator of the Salivary Excretion Based on Physical Vibration of the Parotid Glands.

Oral dryness causes significant health problems both functional (difficulty speaking, chewing and swallowing) and structural in teeth (increased number of infections) and oral mucosa. The main objective of this study is to show an alternative treatment to help stimulate the salivary secretion thus improving the quality of life of the patient. In this study, a salivary stimulation equipment using vibrotactile stimuli is shown. The system has been placed bilaterally in the parotid glands and assessed the efficacy of the salivary secretion by sialometry before and after the stimulation. The new proposal is capable of stimulating salivary secretion, in a significative way after 7 minutes of use, at least in the cases analyzed, and fulfills low-cost, easy-to-use, and safe technical restrictions. In this setting, this paper suggests the performance of a deep clinical trial to measure the exact efficacy of the prototype and the times and frequencies needed to state the optimal treatment depending in each case.

The antifungal activity of human parotid secretion is species-specific.

Candida albicans is the major fungal colonizer of the oral cavity and causes oral candidiasis in immunocompromised patient populations. While antifungal proteins in saliva have been identified and the virulence factors of C. albicans have been well studied, little is known about the role saliva plays in the preferential colonization of the oral cavity by C. albicans. We report that the fungistatic activity of human parotid secretion toward six C. albicans strains is considerably lower than towards nine non-C. albicans fungal species (average IC50 values >1000 mg/l and <70 mg/l, respectively). The species-specific activity of parotid secretion suggests that saliva may play a determining role in oral fungal colonization patterns.

Long non-coding RNA and mRNA profile analysis in the parotid gland of mouse with type 2 diabetes.

AIMS: Salivary gland dysfunction is a common complication of diabetes mellitus (DM). Long non-coding RNA (lncRNA) is evidenced to involve in the functional regulation of salivary gland, however, its role in DM-impaired gland is unknown. Therefore, this study aimed to investigate the expression profiles and functional networks of lncRNA in the parotid glands (PGs) of DM mice. MAIN METHODS: Microarray was used to detect lncRNA and messenger RNA (mRNA) expression profiles in the PGs from db/db and db/m mice. Eleven differently expressed (DE) lncRNAs validated by qRT-PCR were selected for coding-non-coding gene co-expression (CNC) and competing endogenous RNA (ceRNA) network analysis, as well as the following Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Pearson's coefficient correlation analysis was used to analyze the correlations between DE lncRNAs expression and DM pathology. KEY FINDINGS: By using a 2-fold change and P < 0.05 as the cutoff criteria, 1650 DE lncRNAs (758 upregulated and 892 downregulated) and 1073 mRNAs (563 upregulated and 510 downregulated) were identified in the PGs of db/db mice compared to db/m mice. GO and KEGG analysis of DE mRNA suggested that activated inflammation response and downregulated ion transport might count for the dysfunction of diabetic PG. CNC and ceRNA networks analysis of 11 DE lncRNAs showed that the inflammation process and its related signaling pathways including advanced glycation end product (AGE)-receptor for AGE (RAGE) signaling pathway in diabetic complications, cytokine-cytokine receptor interaction, chemokine signaling pathway, apoptosis, and cell adhesion molecules were significantly enriched. The alterations of lncRNAs were closely correlated with higher blood glucose and serum insulin levels in mice. SIGNIFICANCE: We identified multiple lncRNAs/mRNAs and several signaling pathways that may involve in the pathogenesis of diabetic salivary injury, providing new insight into potential target of diabetic hyposalivation.

Variation in size and shape of toxin glands among cane toads from native-range and invasive populations.

If optimal investment in anti-predator defences depends on predation risk, invading new regions (and thus, encountering different predators) may favour shifts in that investment. Cane toads offer an ideal system to test this prediction: expensive anti-predator toxins are stored mainly in parotoid glands whose dimensions are easy to measure, and toad invasions have changed the suites of predators they encounter. Although plasticity may influence parotoid morphology, comparisons between parents and progeny revealed that gland dimensions were highly heritable. That heritability supports the plausibility of an evolved basis to variation in gland dimensions. Measurements of 3779 adult toads show that females have larger glands than males, invasive populations have larger glands than in the native-range, and that parotoid sexual size dimorphism varies strongly among invaded areas. Geographic variation in parotoid morphology may be driven by predation risk to both adult toads and offspring (provisioned with toxins by their mother), with toxins allocated to eggs exacerbating the risk of cannibalism but reducing the risk of interspecific predation. Investment into chemical defences has evolved rapidly during the cane toad's international diaspora, consistent with the hypothesis that organisms flexibly adjust resource allocation to anti-predator tactics in response to novel challenges.

Permeant anions contribute to voltage dependence of ClC-2 chloride channel by interacting with the protopore gate.

It has been shown that the voltage (V(m)) dependence of ClC Cl(-) channels is conferred by interaction of the protopore gate with H(+) ions. However, in this paper we present evidence which indicates that permeant Cl(-) ions contribute to V(m)-dependent gating of the broadly distributed ClC-2 Cl() channel. The apparent open probability (P(A)) of ClC-2 was enhanced either by changing the [Cl(-)](i) from 10 to 200 mM or by keeping the [Cl(-)](i) low (10 mM) and then raising [Cl(-)](o) from 10 to 140 mM. Additionally, these changes in [Cl(-)] slowed down channel closing at positive V(m) suggesting that high [Cl(-)] increased pore occupancy thus hindering closing of the protopore gate. The identity of the permeant anion was also important since the P(A)(V(m)) curves were nearly identical with Cl(-) or Br(-) but shifted to negative voltages in the presence of SCN() ions. In addition, gating, closing rate and reversal potential displayed anomalous mole fraction behaviour in a SCN(-)/Cl() mixture in agreement with the idea that pore occupancy by different permeant anions modifies the V(m) dependence ClC-2 gating. Based on the ec1-ClC anion pathway, we hypothesized that opening of the protopore gate is facilitated when Cl(-) ions dwell in the central binding site. In contrast, when Cl(-) ions dwell in the external binding site they prevent the gate from closing. Finally, this Cl(-)-dependent gating in ClC-2 channels is of physiological relevance since an increase in [Cl(-)](o) enhances channel opening when the [Cl(-)](i) is in the physiological range.

Botulinum toxin type A for the treatment of sialorrhoea in amyotrophic lateral sclerosis: a clinical and neurophysiological study.

Botulinum toxin type A (BoNT/A) has been proposed as an alternative treatment for sialorrhoea in patients with amyotrophic lateral sclerosis (ALS). In an open-label prospective study, BoNT/A was injected into the parotid glands bilaterally using anatomic landmarks in 26 ALS patients with bulbar symptoms. Two weeks after injection the severity of sialorrhoea and the related disability were evaluated subjectively and objectively. A group of healthy subjects acted as controls for saliva production. Patients also underwent electrophysiological tests to evaluate possible toxin effects in the nearby non-injected muscles by comparing the amplitude of compound motor action potentials (cMAPs) elicited by electrical stimulation and recorded from the orbicularis oculi and masseter muscles. After BoNT/A injections, of the 26 patients treated, 23 reported that the severity of sialorrhoea improved and the disabling symptoms diminished. Cotton roll weight also decreased after BoNT/A injection, suggesting a reduction in saliva production. Two patients complained of dry mouth. BoNT/A injection left the cMAP amplitude unchanged, suggesting that botulinum toxin does not significantly affect the non-injected facial and masticatory muscles. In conclusion, intraparotid anatomically-guided BoNT/A injection is an effective, easy, and safe treatment for sialorrhoea in patients with bulbar symptoms related to ALS.