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

We studied the ionic currents activated by basic fibroblast growth factor ( bFGF) and insulin-like growth factor-I (IGF-I) in cultured bovine aortic en dothelial cells (BAE-1) by using patch-clamp and single-cell fluorimetric c alcium measurements. In whole-cell, voltage-clamp experiments at V-h = -50 mV, the addition of either bFGF (20 ng/ml) or IGF-I (50 ng/ml) induced an i nward current with similar amplitude, time course, and permeation propertie s. The response was dependent on receptor occupancy and showed a desensitis ation in the continued presence of the factors. Ionic substitutions in whol e-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 extrace llular medium, whereas no detectable release from internal stores was obser ved. 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, a nd tyrphostin 1. Moreover, addition of 200 muM Na3VO4, an inhibitor of prot ein tyrosine phosphatase (PTP) activity, evoked the responses to the factor s both in patch-clamp and in fluorimetric measurements. Cell-attached recor dings 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. (C) 2000 Wiley-Liss, Inc.