Effects of protein tyrosine kinase inhibitors on voltage‐operated calcium channel currents in vascular smooth muscle cells and pp60c‐src kinase activity

Publisher Website
Google Scholar
Abstract
Tyrosine kinases have been proposed as regulators of voltage‐operated calcium channels. The effects of a range of structurally different inhibitors of protein tyrosine kinases (PTK) were examined on voltage‐operated calcium channel currents (IBa) and pp60c‐src kinase (c‐src) activity in vitro. IBa was measured in single myocytes isolated from rabbit ear artery by conventional whole cell voltage‐clamp techniques. The activity of purified human c‐src was measured in vitro using a non‐radioactive assay. Bath application of tyrphostin‐23 and genistein (non‐selective PTK inhibitors), bistyrphostin (a receptor‐PTK‐selective inhibitor) and PP1 (a src family‐selective inhibitor) inhibited IBa in a concentration‐dependent manner over a range of test membrane potentials. Intracellular application of peptide‐A, a peptide inhibitor of c‐src also inhibited currents. Inhibitor potency series against IBa was PP1 > genistein > tyrphostin 23 > bistyrphostin. Tyrphostin‐23, genistein, PP1, and peptide‐A shifted the steady‐state inactivation curves in a hyperpolarized direction without altering their slope. The inhibitors had no significant effects on IBa activation calculated from current‐voltage relationships. The agents inhibited c‐src activity in a concentration‐dependent manner. The order of potency was PP1 > genistein > peptide‐A > tyrphostin‐23 > bistyrphostin. The IC50 for inhibition of c‐src activity was similar to the IC50 for inhibition of IBa in all cases. Western blot analysis with a specific antibody to c‐src showed the presence of this cytoplasmic tyrosine kinase in rabbit ear artery cells. A range of structurally dissimilar inhibitors of PTKs inhibit IBa and c‐src activity with similar potency. These data provide further evidence implicating endogenous c‐src in the modulation of L‐type calcium channels in vascular smooth muscle cells. British Journal of Pharmacology (2000) 129, 1347–1354; doi:10.1038/sj.bjp.0703186

This publication has 40 references indexed in Scilit: