Type IV collagen is a major basement membrane component that has been implicated in the regulation of angiogenesis. The purpose of this study was to evaluate the effect of type IV collagen on the angiogenic response of native endothelial cells in three-dimensional vascular organ culture. Rings of rat aorta were cultured under serum-free conditions in gels of type I collagen with or without type IV collagen. In the absence of type IV collagen, aortic rings generated neovessels, which proliferated until day 9 and gradually regressed during the second and third weeks of culture. Type IV collagen promoted neovessel elongation and survival in a dose-dependent manner. Microvascular length increased by 43, 57, and 119% over control values in cultures treated with 3, 30, and 300 μg/ml type IV collagen, respectively. When used at high concentrations (300 μg/ml) type IV collagen stabilized the neovascular outgrowths and prevented vascular regression. Type IV collagen also promoted the formation of neovessels, but significant stimulatory effects were observed only at an intermediate concentration (30 μg/ml) and were no longer significant at the high concentration (300 μg/ml). The observation that type IV collagen has dose-dependent effects on vascular elongation, proliferation, and stabilization, supports the concept that the developing basement membrane of neovessels acts as a solid-phase regulator of angiogenesis, whose function varies depending on the concentration of its molecular components.