Inhibition by inositoltetrakisphosphates of calcium- and volume-activated Cl^– currents in macrovascular endothelial cells

Abstract

 We have used the whole-cell patch-clamp technique to study the effects of inositol 1,4,5,6-tetrakisphosphate [Ins(1,4,5,6)P ₄], inositol 3,4,5,6-tetrakisphosphate [Ins(3,4,5,6)P ₄] and inositol 1,3,4,5,6-pentacisphosphate [Ins(1,3,4,5,6)P ₅] on volume-activated Cl^– currents (I _Cl,vol) in cultured endothelial cells from bovine pulmonary artery (CPAE cells). Ins(1,4,5,6)P ₄ and Ins(3,4,5,6)P ₄ were applied intracellularly via the patch pipette at concentrations between 10 and 100 μM. Both tetrakisphosphates inhibited the Cl^– current I _Cl,Ca, which was activated by intracellular loading of the cells with 500 nM Ca²⁺ [for inhibition by Ins(1,4,5,6)P ₄: 58% at 10 μM, 75% at 100 μM; for Ins(3,4,5,6)P ₄: 44% at 10 μM, 65% at 100 μM]. Inhibition of I _Cl,Ca occurred without significant changes in its kinetic properties. The amplitude of I _Cl,vol activated by a 13.5 or 27% hypotonic solution at +100 mV was strongly reduced in cells loaded with either tetrakisphosphate, i.e. a 73% reduction for Ins(3,4,5,6)P ₄ and 89% for Ins(1,4,5,6)P ₄ at 100 μM. Both tetrakisphosphates also inhibited a current probably identical to I _Cl,vol which was activated by dialysing the cell with 100 μM guanosine 5′-O-(3-thiotriphosphate) (GTP[γ-S]). Ins(1,3,4,5,6)P ₅ at a concentration of 30 μM did not significantly reduce I _{Cl, vol}. The effects of Ins(3,4,5,6)P ₄ may represent an inhibitory pathway for the I _Cl,Ca and I _Cl,vol in macrovascular endothelium after sustained receptor-mediated activation of phospholipase C.