Astroglial cell-induced expression of na-k-cl cotransporter in brain microvessel endothelial cells. Sun, Dandan, Christian Lytle, and Martha E. O'donnell. Department of Human Physiology, School of Medicine, University of California, Davis, CA 95616, Division of Biomedical Sciences, University of California, Riverside, CA 92521
APStracts 2:0277C, 1995.
Endothelial cells of the blood-brain barrier (BBB) are specifically characterized and the maintenance of the BBB endothelial phenotype depends on astrocyte-endothelial interactions. We showed previously that cultured cerebral microvessel endothelial cells (CMEC) exhibit robust Na-K-Cl cotransport activity. In the present study, we evaluated the expression of Na-K-Cl cotransport protein in CMEC by quantitative Western blot analysis and found that a protein of approximately 170 kDa was recognized by a monoclonal antibody against the cotransporter. Exposure of CMEC to astroglial cells or their conditioned media increased the expression of the CMEC cotransport protein by about 55%. Using a monoclonal antibody against the [alpha] subunit of chicken Na/K ATPase, we found that these treatments also increased expression of Na/K ATPase protein by a similar amount. By comparing bumetanide-sensitive K influx and 3H-bumetanide binding of apical versus basolateral surfaces of CMEC, We found both cotransporter activity and 3H-bumetanide binding to be approximately 90% apical and 10% basolateral. Co-culture of the CMEC with astroglial cells increased cotransport activity and 3H-bumetanide binding at both surfaces, with the asymmetric distribution maintained. These results indicate that the cotransporter is regulated by astroglial cells and an apically distributed CMEC cotransporter may function in tandem with the basolateral Na/K ATPase to mediate vectorial transport of Na and Cl across the BBB. 

Received 9 June 1995; accepted in final form 10 July 1995.
APS Manuscript Number C323-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 30 July 1995.