The pharmacological study of the blood-brain barrier has often been hampered by the unavailability of a large number of pure and fully differentiated brain capillary endothelial cells. Here we describe a homogeneous culture of brain capillary endothelial cells isolated from bovine brain (BBECs), which retain at least some phenotypic characteristics of the functional blood-brain barrier: intracellular tight junctions and monoamine oxidase activity. These cells were subcultured in vitro, in the absence of any neuronal or glial influences, for greater than 100 doublings without any sign of senescence. The present study is focused on the expression of beta-adrenergic receptors on BBECs. By Northern blot hybridization, subtype-specific ligand binding, and cyclic AMP accumulation experiments, we demonstrate that beta 1- and beta 2-adrenergic receptors are coexpressed (in the respective proportions of 42 and 58%) on BBEC membranes and are functionally coupled to adenylate cyclase. This is the first report documenting a significant number of beta 1-adrenergic receptors on brain capillary endothelial cells. The results are discussed in light of the known noradrenergic innervation of brain capillaries.