Studies of glucose transporter activity and anti-glucose transporter (GLUT1) immunoblots were performed on different endothelial cell primary cultures (brain capillary, adrenal capillary and aortic) to determine their response to glucose deprivation. Cell cultures were exposed to glucose deprivation (0.5 mM) for 48 h periods and refed (11.0 mM) for 36 additional hours. Control cultures were kept in 11.0 mM glucose for the duration of these studies. Measurements of 2-[3H]deoxy-D-glucose uptake and membrane fraction purification were performed every 12 h during these timecourses. Baseline cytochalasin-B sensitive uptake of 2-deoxy-D-glucose was near three times larger in brain capillary endothelial cells than in adrenal or aortic endothelial cultures. In all three endothelial cell cultures, 2-deoxy-D-glucose uptake increased during glucose deprivation, and returned to control values upon refeeding. Aortic and adrenal cortical endothelia expressed the starvation induced increases 12 h sooner than brain capillary endothelia. Return to control values was also 12 h faster in these cultured endothelia. Immunoblot studies showed that in all three endothelial cell cultures the increases in transporter activity during glucose starvation correlate with increased membrane expression of GLUT1. Quantitative analysis of the anti-GLUT1 immunoblots indicated that induction of GLUT1 following glucose starvation was slower in brain capillary endothelia than in aortic or adrenal endothelia. The slower response by brain capillary endothelial cells may be related to the higher transport rate of glucose in these cells.