We report on a series of experiments which examines the factors controlling lymphocyte adhesion to brain endothelium in vitro and the factors which control cell migration across the endothelium, using a new migration assay. Although lymphocyte adhesion preceded migration across the brain endothelium, the two processes are not identical. We noted that activated CD4+ T cells were particularly good at migrating across endothelia. CD8+ T cells and B cells did not migrate but adhered well to endothelia. Moreover, the endothelium maintained high levels of cell traffic without being disrupted and without exhausting the molecular systems which allowed migration. From the viewpoint of migration of dividing cells, the state of lymphocyte activation appeared to be the most important controlling factor--these cells migrated equally well across endothelium activated with cytokines or untreated endothelium. The kinetics of adhesion suggested that the LFA-1/ICAM-1 and VLA-4/VCAM combinations of adhesion molecules were important in controlling migration. With antibody blocking studies, the role of the LFA-1/ICAM-1 system was equivocal. While anti-LFA-1 blocked lymphocyte adhesion, anti-ICAM-1 did not, suggesting that the level of ICAM-1 was not critical.