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Potent stimulation and inhibition of the CFTR Cl(-) current by phloxine B.

ABSTRACT: The effects of the fluoresceine derivative, phloxine B, on the Cl(-) current through the cystic fibrosis transmembrane conductance regulator (CFTR) were examined in Xenopus oocytes expressing human CFTR. In whole oocytes, the CFTR Cl(-) current (I(CFTR)) was activated by superfusion with isobutylmethylxanthine and forskolin. I(CFTR) was stable during activation and deactivated rapidly upon washout of the activation solution. Phloxine B slowed deactivation and, at high concentrations, inhibited I(CFTR) weakly. In excised inside-out macropatches, I(CFTR) was activated by the catalytic subunit of protein kinase A (cPKA) and MgATP. Phloxine B (0.01 - 3 microM), applied after activation, increased I(CFTR) within 30 s followed by a slow decrease which became dominant at high concentrations. Slowing of deactivation of the CFTR was observed at all concentrations. The effect of phloxine B after 30 s had a bell-shaped concentration-dependence with midpoints at 45 and 1600 nM for the stimulatory and the inhibitory limb, respectively; maximum stimulation was about 1.8 times. The slow inhibitory component, measured after 6 min, occurred with an IC(50) value of approximately 1 microM. In the absence of cPKA, phloxine B did not stimulate I(CFTR). In the presence of cPKA and MgATP, the effects of phloxine B were more prominent at low (0.02 mM) than at high ATP (2 mM). The data show that phloxine B modulates I(CFTR) by increasing channel activity and slowing channel deactivation; at high concentrations inhibition dominates. The effects may be mediated by direct interactions with CFTR from the inside of the cell.

SUBMITTER: Bachmann A

PROVIDER: S-EPMC1572357 | biostudies-literature | 2000 Oct

REPOSITORIES: biostudies-literature

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Potent stimulation and inhibition of the CFTR Cl(-) current by phloxine B.

Bachmann A A Russ U U Waldegger S S Quast U U

British journal of pharmacology 20001001 3

The effects of the fluoresceine derivative, phloxine B, on the Cl(-) current through the cystic fibrosis transmembrane conductance regulator (CFTR) were examined in Xenopus oocytes expressing human CFTR. In whole oocytes, the CFTR Cl(-) current (I(CFTR)) was activated by superfusion with isobutylmethylxanthine and forskolin. I(CFTR) was stable during activation and deactivated rapidly upon washout of the activation solution. Phloxine B slowed deactivation and, at high concentrations, inhibited I ...[more]

PMID: 11015292

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Project description:Aquaporins (AQPs) facilitate the transepithelial water flow involved in epithelial fluid secretion in numerous tissues; however, their function in the pancreas is less characterized. Acute pancreatitis (AP) is a serious disorder in which specific treatment is still not possible. Accumulating evidence indicate that decreased pancreatic ductal fluid secretion plays an essential role in AP; therefore, the aim of this study was to investigate the physiological and pathophysiological role of AQPs in the pancreas. Expression and localization of AQPs were investigated by real-time PCR and immunocytochemistry, whereas osmotic transmembrane water permeability was estimated by the dye dilution technique, in Capan-1 cells. The presence of AQP1 and CFTR in the mice and human pancreas were investigated by immunohistochemistry. Pancreatic ductal HCO3- and fluid secretion were studied on pancreatic ducts isolated from wild-type (WT) and AQP1 knock out (KO) mice using microfluorometry and videomicroscopy, respectively. In vivo pancreatic fluid secretion was estimated by magnetic resonance imaging. AP was induced by intraperitoneal injection of cerulein and disease severity was assessed by measuring biochemical and histological parameters. In the mice, the presence of AQP1 was detected throughout the whole plasma membrane of the ductal cells and its expression highly depends on the presence of CFTR Cl- channel. In contrast, the expression of AQP1 is mainly localized to the apical membrane of ductal cells in the human pancreas. Bile acid treatment dose- and time-dependently decreased mRNA and protein expression of AQP1 and reduced expression of this channel was also demonstrated in patients suffering from acute and chronic pancreatitis. HCO3- and fluid secretion significantly decreased in AQP1 KO versus WT mice and the absence of AQP1 also worsened the severity of pancreatitis. Our results suggest that AQP1 plays an essential role in pancreatic ductal fluid and HCO3- secretion and decreased expression of the channel alters fluid secretion which probably contribute to increased susceptibility of the pancreas to inflammation.

Dataset Information

Potent stimulation and inhibition of the CFTR Cl(-) current by phloxine B.

Publications

Potent stimulation and inhibition of the CFTR Cl(-) current by phloxine B.

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