To study the cellular mechanism of vascular responsiveness by vasoactive hormones, such as catecholamines and angiotensin (A), the vascular smooth muscle cells (VSMC) of two clonal cell line (A7r5 and A 10) from rat embryo, and of adult rat aorta were established in culture. Binding studies using 125I-labeled-hydroxyphenylethylaminoethyltetralone as an α-adrenergic ligand and 125I-labeled-iodocyanopindolol as a β-adrenergic ligand, revealed that cultured VSMCs contain both α- and β-adrenergic receptors ; the binding was specific, rapid, reversible, and saturable. α-Adrenergic receptors appear to be a single class of high-affinity binding sites with an apparent dissociation constant (Kd) of ∼ 2× 10-10 M and a maximal binding capacity (Bmax) of ∼300, 000-400, 000 sites/cell, and exclusively of α1-subtype that is responsible for smooth muscle contraction. On the other hand, β-adrenergic receptors show almost comparable characteristics with the apparent Kd of ∼ -0.7-1.1 10-10 M and Bmax of 50, 000-130, 000 sites/cell, and consist predominantly of 2-subtype that mediates smooth muscle relaxation. Furthermore, -adrenergic receptors are coupled to adenylate cyclase system, of which activation by -agonists induces intracellular cyclic AMP formation. In contrast, the binding of 125I-labeled-AII was demonstrated only in A7r5. The binding declined rapidly during incubation possibly due to faster degradation of AII by proteolytic enzyme (s). AII receptors appear to be a single class of high-affinity binding sites with the apparent Kd of 0.9 10-10 M and Bmax of 11, 000 sites/cell, of which affinity is higher than any of vascular AII receptors previously reported. Therefore, cultured VSMCs isolated from neurogenic and systemic influence should provide a valuable in vitro system to investigate the characteristics of specific receptors for vasoactive hormones, the cellular mechanisms involved in activation and regulation of the receptors and subsequent biochemical responses.