Effect of hypoxia on Na+-K+-Cl- cotransport in cultured brain capillary endothelial cells of the rat

The effect of hypoxia on Na+,K+-ATPase and Na+-K+-Cl- cotransport activity in cultured rat brain capillary endothelial cells (RBECs) was investigated by measuring Rb-86(+) uptake as a tracer for K+. RBECs expressed both Na+,K+-ATPase and Na+-K+-Cl- cotransport activity (4.6 and 5.5 nmol/mg of protein/min, respectively). Hypoxia (24 h) decreased cellular ATP content by 43.5% and reduced Na+,K+-ATPase activity by 38.9%, whereas it significantly increased Na+-K+-Cl- cotransport activity by 49.1% in RBECs. To clarify further the mechanism responsible for these observations, the effect of oligomycin-induced ATP depletion on these ion transport systems was examined. Exposure of RBECs to oligomycin led to a time-dependent decrease of cellular ATP content (by similar to 65%) along with a complete inhibition of Na+,K+-ATPase and a coordinated increase of Na+-K+-Cl- cotransport activity (up to 100% above control values). Oligomycin augmentation of Na+-K+-Cl- cotransport activity was not observed in the presence of 2-deoxy-D-glucose (a competitive inhibitor of glucose transport and glycolysis) or in the absence of glucose. These results strongly suggest that under hypoxic conditions when Na+,K+-ATPase activity is reduced, RBECs have the ability to increase K+ uptake through Na+-K+Cl- cotransport.