Extracellular site for econazole-mediated block of Ca2+ release-activated Ca2+ current (I-crac) in T lymphocytes

1 Standard whole cell patch clamp recording techniques were used to study the pharmacological characteristics and site of econazole-mediated inhibition of calcium release-activated calcium current (I-crac) in the human leukaemic T cell line, Jurkat. 2 Extracellularly applied econazole blocked I-carc in a concentration-dependent manner (IC(50)approximate to 14 mu M). Block developed over a relatively slow timecourse of 30-60 s (10 mu M), and only partially reversed over minutes. 3 Econazole dialysed from the pipette into the cytosol at concentrations ranging from 0.1 to 30 mu M did not reduce I-carc, or quantitatively affect I-crac block by extracellularly applied econazole. 4 A less lipophilic quaternary iodide derivative of econazole was synthesized to retard absorption through the cell membrane. When applied extracellularly, this compound blocked I-crac in a concentration-dependent manner with onset kinetics comparable to econazole. 5 Results with intracellularly dialysed econazole and the quaternary econazole derivative provide convergent evidence that econazole blocks I-crac via an extracellular interaction. 6 The inability of intracellularly applied econazole to inhibit I-crac argues against the notion that econazole inhibits capacitative Ca2+ entry pathways secondary to its known inhibitory effects on cytochrome P-450.