Calcium influx induced by activation of tyrosine kinase receptors in cultured bovine aortic endothelial cells

• Published in: Journal of cellular physiology 2000-12, Vol.185 (3), p.454-463
• Main Authors: Munaron, Luca, Fiorio Pla, Alessandra
• Format: Article
• Language: English
• Subjects: Animals; Aorta - physiology; Calcium - physiology; Cattle; Cells, Cultured; Endothelium, Vascular - physiology; Fibroblast Growth Factor 2 - pharmacology; Insulin-Like Growth Factor I - pharmacology; Ion Transport - physiology; Receptor Protein-Tyrosine Kinases - physiology; Signal Transduction - drug effects; Signal Transduction - physiology
• ISSN: 0021-9541; 1097-4652
• DOI: 10.1002/1097-4652(200012)185:3<454::AID-JCP17>3.0.CO;2-A

We studied the ionic currents activated by basic fibroblast growth factor (bFGF) and insulin‐like growth factor‐I (IGF‐I) in cultured bovine aortic endothelial cells (BAE‐1) by using patch‐clamp and single‐cell fluorimetric calcium measurements. In whole‐cell, voltage‐clamp experiments at Vh = −50 mV, the addition of either bFGF (20 ng/ml) or IGF‐I (50 ng/ml) induced an inward current with similar amplitude, time course, and permeation properties. The response was dependent on receptor occupancy and showed a desensitisation in the continued presence of the factors. Ionic substitutions in whole‐cell experiments indicated that the current barely discriminated among Na+, Ca+, and K+ ions. Accordingly, stimulation with bFGF or IGF‐I induced a dose‐dependent [Ca2+]i elevation completely due to entry from the extracellular medium, whereas no detectable release from internal stores was observed. Calcium influx was dependent on protein tyrosine kinase (PTK) activity; it was significantly inhibited by treatment with genistein or tyrphostin 47, two PTK inhibitors, and not affected by inactive analogues, daidzein, and tyrphostin 1. Moreover, addition of 200 μM Na3VO4, an inhibitor of protein tyrosine phosphatase (PTP) activity, evoked the responses to the factors both in patch‐clamp and in fluorimetric measurements. Cell‐attached recordings using 100 mM CaCl2 in the pipette showed that bFGF and IGF‐I activate calcium‐permeable channels with similar properties. These results provide evidence for a calcium influx induced by two factors that bind to tyrosine kinase receptors (RTK) in endothelial cells. J. Cell. Physiol. 185:454–463, 2000. © 2000 Wiley‐Liss, Inc.