We evaluated the effects of mediators that can be produced by smooth muscle and endothelial cells on growth responses in isolated arteries. Segments of carotid and renal arteries, denuded of endothelium, were isolated from adult rats and studied during tissue culture in the presence of indomethacin. Three days of culture in the presence of serum stimulated DNA synthesis in the media. During long-term culture new layers of cells developed at the borders of the arterial segments. Medial DNA synthesis depended less on serum than extramedial cell proliferation. During moderate stimulation, basic fibroblast growth factor and endothelin-1 enhanced and interleukin-1 and transforming growth factor-beta reduced medial DNA synthesis, whereas insulin-like growth factor-1, platelet-derived growth factor AA, platelet-derived growth factor BB, and angiotensin II were without effect. Of these factors, only endothelin-1 stimulated extramedial cell proliferation. In addition, serum-stimulated but not basic fibroblast growth factor-stimulated medial DNA synthesis was less marked in arteries that had not been denuded of endothelium than in ee-endothelialized arteries. Differences between preparations with and without endothelium persisted in the absence of L-arginine and in the presence of an inhibitor of nitric oxide synthase. These observations confirmed that DNA synthesis in the arterial media and extramedial cell proliferation are influenced by different factors. They further indicated that endothelial modulation of medial DNA synthesis does not seem to involved endothelium-derived prostaglandins, nitric oxide, or interleukin-1 and that it can be blunted by basic fibroblast growth factor.