Anoctamin 1 (Ano1) is required for glucose-induced membrane potential oscillations and insulin secretion by murine ß-cells.

Anions such as Cl(-) and HCO3 (-) are well known to play an important role in glucose-stimulated insulin secretion (GSIS). In this study, we demonstrate that glucose-induced Cl(-) efflux from ß-cells is mediated by the Ca(2+)-activated Cl(-) channel anoctamin 1 (Ano1). Ano1 expression in rat ß-cells is demonstrated by reverse transcriptase-polymerase chain reaction, western blotting, and immunohistochemistry. Typical Ano1 currents are observed in whole-cell and inside-out patches in the presence of intracellular Ca(++): at 1 µM, the Cl(-) current is outwardly rectifying, and at 2 µM, it becomes almost linear. The relative permeabilities of monovalent anions are NO3 (-) (1.83 ± 0.10) > Br(-) (1.42 ± 0.07) > Cl(-) (1.0). A linear single-channel current-voltage relationship shows a conductance of 8.37 pS. These currents are nearly abolished by blocking Ano1 antibodies or by the inhibitors 2-(5-ethyl-4-hydroxy-6-methylpyrimidin-2-ylthio)-N-(4-(4-methoxyphenyl)thiazol-2-yl)acetamide (T-AO1) and tannic acid (TA). These inhibitors induce a strong decrease of 16.7-mM glucose-stimulated action potential rate (at least 87 % on dispersed cells) and a partial membrane repolarization with T-AO1. They abolish or strongly inhibit the GSIS increment at 8.3 mM and at 16.7 mM glucose. Blocking Ano1 antibodies also abolish the 16.7-mM GSIS increment. Combined treatment with bumetanide and acetazolamide in low Cl(-) and HCO3 (-) media provokes a 65 % reduction in action potential (AP) amplitude and a 15-mV AP peak repolarization. Although the mechanism triggering Ano1 opening remains to be established, the present data demonstrate that Ano1 is required to sustain glucose-stimulated membrane potential oscillations and insulin secretion.

Interaction between cAMP, volume­regulated anion channels and the Na+­HCO3­­cotransporter, NBCe1, in the regulation of nutrient­ and hypotonicity­induced insulin release from isolated rat pancreatic islets and tumoral insulin­producing BRIN­BD11 cells.

Soluble adenylyl cyclase (sAC) has been hypothesized to play a role in insulin secretion. The present study aimed to investigate the interaction between adenosine 3',5'­cyclic monophosphate (cAMP), volume­regulated anion channels (VRACs) and the electrogenic sodium bicarbonate (Na+­HCO3­) cotransporter, NBCe1, in the regulation of nutrient­ and hypotonicity­induced insulin release from rat pancreatic islets and tumoral insulin­producing BRIN­BD11 cells. In the islets, 5­nitro­2­(3­phenylpropylamino)benzoic acid (NPPB) and 5­chloro­2­hydroxy­3­(thiophene­2­carbonyl)indole­1­carboxamide (tenidap) reduced glucose­stimulated insulin release, however, only NPPB suppressed the enhancing action of cAMP analogs upon such a release. Insulin output from the BRIN­BD11 cells was stimulated by 2­ketoisocaproate (KIC) or extracellular hypoosmolarity. cAMP analogs and 3­isobutyl­1­methylxanthine increased the insulin output recorded in the isotonic medium to a greater relative extent than that in the hypotonic medium. The secretory response to KIC or hypotonicity was inhibited by NPPB or tenidap, which both also opposed the enhancing action of cAMP analogs. Inhibitors of mitogen­activated protein (MAP) kinase decreased insulin output in isotonic and hypotonic media. The inhibitor of sAC, 2­hydroxyestriol, caused only a modest inhibition of insulin release, whether in the isotonic or hypotonic medium, even when tested at a concentration of 100 µM. The omission of NaHCO3 markedly decreased the secretory response to KIC or extracellular hypotonicity. The omission of Na+ suppressed the secretory response to extracellular hypotonicity. The observations of the present study do not support the hypothesis of a major role for sAC in the regulation of insulin release.

Pendrin: the thyrocyte apical membrane iodide transporter?

In the thyroid, the transport of iodide from the extracellular space to the follicular lumen requires two steps: the transport in the cell at the basal side and in the lumen at the apical side. The first step is mediated by the Na(+)/I(-) symporter (NIS). In most reviews and textbooks, the second step is presented as mediated by pendrin. In this review, we analyze this assumption. There are several arguments supporting the concept that indeed pendrin plays an important role in thyroid physiology. However, biochemical, clinical and histological data on the thyroid of a patient with Pendred syndrome do not suggest an essential role in iodide transport, which is corroborated by the lack of a thyroid phenotype in pendrin knockout mice. Experiments in vivo and in vitro on polarized and unpolarized cells show that iodide is transported transport of iodide at the apex of the thyroid cell. Moreover, ectopic expression of pendrin in transfected non-thyroid cells is capable of mediating iodide efflux. It is concluded that pendrin may participate in the iodide efflux into thyroid lumen but not as the unique transporter. Moreover, another role of pendrin in mediating Cl(-)/HCO(3)(-) exchange and controlling luminal pH is suggested.

Expression and localization of cystic fibrosis transmembrane conductance regulator in the rat endocrine pancreas.

Impaired glucose tolerance and overt diabetes mellitus are becoming increasingly common complications of cystic fibrosis (CF), most probably merely as a result of increased life expectancy. In order to understand the pathophysiology of cystic fibrosis-related diabetes (CFRD), knowledge on the possible expression and cell distribution of the cystic fibrosis transmembrane conductance regulator (CFTR) protein within the endocrine pancreas is required. In this report, we establish the first evidence for expression of CFTR protein in rat pancreatic islets by using independent techniques. First reverse transcriptase-polymerase chain reaction (RT-PCR) amplification showed that CFTR mRNA is present in isolated islets of Langerhans. Furthermore, the analysis of flow cytometry-separated islet cells indicated that the level of CFTR transcripts is significantly higher in the non-beta than in beta-cell populations. The expression of CFTR protein in rat islet cells was also demonstrated by Western blotting and the level of expression was also found significantly higher in the non-beta than in beta-cell populations. Last, in situ immunocytochemistry studies with two monoclonal antibodies recognizing different CFTR epitopes indicated that CFTR expression occurs mainly in glucagon-secreting alpha-cells.

Evaluation of somatostatin inhibitory effect on pancreatic exocrine function using secretin-enhanced dynamic magnetic resonance cholangiopancreatography: a crossover, randomized, double blind, placebo-controlled study.

OBJECTIVES: Somatostatin inhibitory effect on the exocrine pancreas has been demonstrated by clinical and experimental studies performed with invasive investigative methods. The aim of this study was to quantify the inhibitory effect of low doses of somatostatin (62.5, 125, and 250 microg) on secretin-stimulated pancreatic exocrine secretions using magnetic resonance cholangiopancreatography (MRCP). METHODS: Ten healthy volunteers underwent 4 MRCP at a 1-week interval. At each MRCP, 1 of the 3 doses of Somatostatin or the placebo was given by the intravenous route for a period of 40 minutes. After 20 minutes from the beginning of drug infusion, secretin was injected (0.3 CU/kg). MRCP was performed before and every 30 to 45 seconds for 15 minutes after secretin administration. Pancreatic exocrine secretions were quantified by the measurements of pancreatic flow output and total excreted volume, derived from a linear regression between MRCP calculated volumes and time. RESULTS: For the 3 doses of somatostatin, pancreatic flow output was significantly reduced compared to placebo (P < 0.05). Total excreted volume was significantly reduced only for the doses of 62.5 and 250 microg. No statistical significant differences were observed among the 3 doses. CONCLUSIONS: Low doses of somatostatin inhibit pancreatic exocrine secretions as demonstrated noninvasively with MRCP.

Primary antibiotic resistance and effectiveness of Helicobacter pylori triple therapy in ulcero-inflammatory pathologies of the upper digestive tract.

OBJECTIVES: To determine firstly, the rates of primary antimicrobial resistance for Helicobacter pylori-associated upper-digestive lesions in relation to the success rate of triple therapy; and secondly, the performance of HpSA stool antigen detection test for control of eradication after treatment. METHODS: Prospective open study of 436 patients who underwent upper-digestive tract endoscopy with biopsies for histological examination and culture between January 1 and July 31, 2002 at a University hospital in Brussels, Belgium. The primary resistance to antibiotics of H. pylori isolates was determined by disc diffusion method. Seventy of 164 infected patients agreed to be included in the treatment study with standard triple therapy with amoxicillin + clarithromycin + omeprazole adjusted on the basis of antibiogram results. Control of eradication was tested by 14C-Urea breath test and H. pylori Stool Antigen test (HpSA test). RESULTS: Primary resistance to clarithromycin and metronidazole was observed in 3% and 31% of the isolates, respectively. No primary resistance to amoxicillin and tetracycline was observed. By intention to treat analysis, H. pylori was eradicated in 56 (80%) patients included in the therapeutic study. Three (4%) patients were lost to follow-up. The rate of eradication failure was 20% (14/70), included 11 cases documented by a positive control test (14C-Urea breath test). In comparison with 14C-Urea breath test, the H. pylori Stool Antigen test showed a sensitivity of 100%, a specificity of 91%, PPV of 69%, and NPV of 100%. CONCLUSION: Standard triple therapy achieved 80% bacterial eradication in this patient population with a low prevalence of H. pylori primary antibiotic resistance. Our data confirm that the H. pylori Stool Antigen test displays a diagnostic performance similar to the breath test for control of eradication.