Na(+)-K(+)-Cl- cotransport system in brain capillary endothelial cells: response to endothelin and hypoxia.

Abstract

Effect of endothelin-1 and chemically induced hypoxia on Na(+)-K(+)-Cl- cotransport activity in cultured rat brain capillary endothelial cells was examined by using 86Rb+ as a tracer for K+; bumetanide-sensitive K+ uptake was defined as Na(+)-K(+)-Cl- cotransport activity. Endothelin-1, phorbol 12-myristate 13-acetate (PMA), or thapsigargin increased Na(+)-K(+)-Cl- cotransport activity. A protein kinase C inhibitor, bisindolylmaleimide, inhibited PMA- and endothelin-1- (but not thapsigargin-) induced Na(+)-K(+)-Cl- cotransport activity, indicating the presence of both protein kinase C-dependent regulatory mechanisms and protein kinase C-independent mechanisms which involve intracellular Ca2+. Oligomycin, sodium azide, or antimycin A increased Na(+)-K(+)-Cl- cotransport activity by 80-200%. Oligomycin-induced Na(+)-K(+)-Cl- cotransport activity was reduced by an intracellular Ca2+ chelator (BAPTA/AM) but not affected by bisindolylmaleimide, suggesting the involvement of intracellular Ca2+, and not protein kinase C, in hypoxia-induced Na(+)-K(+)-Cl- cotransport activity.