1. Regulation of a swelling-activated chloride current in bovine endothelium by protein tyrosine phosphorylation and G proteins.
- Author
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Voets T, Manolopoulos V, Eggermont J, Ellory C, Droogmans G, and Nilius B
- Subjects
- Action Potentials drug effects, Action Potentials physiology, Animals, Benzylidene Compounds pharmacology, Cattle, Cell Line, Cell Size drug effects, Cell Size physiology, Chloride Channels drug effects, Drug Synergism, Endothelium, Vascular drug effects, Endothelium, Vascular physiology, Enzyme Inhibitors pharmacology, Genistein pharmacology, Guanosine 5'-O-(3-Thiotriphosphate) pharmacology, Hydroquinones pharmacology, Hypotonic Solutions pharmacology, Nitriles pharmacology, Phosphorylation, Protein Tyrosine Phosphatases antagonists & inhibitors, Protein-Tyrosine Kinases antagonists & inhibitors, Sensitivity and Specificity, Vanadates pharmacology, Chloride Channels physiology, Endothelium, Vascular cytology, GTP-Binding Proteins physiology, Protein Tyrosine Phosphatases metabolism, Protein-Tyrosine Kinases metabolism, Tyrphostins
- Abstract
1. The role of protein tyrosine phosphorylation and of G proteins in the activation of a swelling-activated Cl- current (ICl,swell) in calf pulmonary artery endothelial (CPAE) cells was studied using the whole-cell patch clamp technique. ICl,swell was activated by reducing the extracellular osmolality by either 12.5% (mild hypotonicity) or 25% (strong hypotonicity). 2. The protein tyrosine kinase (PTK) inhibitors tyrphostin B46, tyrphostin A25 and genistein inhibited ICl,swell with IC50 values of, respectively, 9.2 +/- 0.2, 61.4 +/- 1.7 and 62.9 +/- 1.3 microM. Tyrphostin A1, a tyrphostin analogue with little effect on PTK activity, and daidzein, an inactive genistein analogue, were without effect on ICl,swell. 3. The protein tyrosine phosphatase (PTP) inhibitors Na3VO4 (200 microM) and dephostatin (20 microM) potentiated ICl,swell activated by mild hypotonicity by 47 +/- 9 and 69 +/- 15%, respectively. 4. Intracellular perfusion with GTP gamma S (100 microM) transiently activated a Cl- current with an identical biophysical and pharmacological profile to ICl,swell. This current was inhibited by the tested PTK inhibitors and potentiated by the PTP inhibitors. Hypertonicity-induced cell shrinkage completely inhibited the GTP gamma S-activated Cl- current. 5. Intracellular perfusion with GDP beta S (1 mM) caused a time-dependent inhibition of ICl,swell, which was more pronounced when the current was activated by mild hypotonicity. 6. Our results demonstrate that the activity of endothelial swelling-activated Cl- channels is dependent on tyrosine phosphorylation and suggest that G proteins regulate the sensitivity to cell swelling.
- Published
- 1998
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