We have previously shown that ethanol-induced injury to the gastric mucosa triggers increased expression of the angiogenic factors, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) and angiogenesis. To further investigate ethanol-induced angiogenesis, we used an in vitro angiogenesis model which employs the ability of an endothelial-derived cell line (EA hy926) to form tubelike structures resembling capillaries when plated on the matrix material, Matrigel. We report that serum-starved EA hy926 cells, incubated for as little as 5 minutes with ethanol concentrations of 1.0-2.5%, formed tubelike structures reflecting in vitro angiogenesis. Control cells, not incubated with ethanol, did not form tubelike structures. Incubation for 5 minutes with 2.5% ethanol resulted in increased activities of PKC and MAP kinase (ERK2) by 1.6-fold (p < 0.05) and 2.3-fold (P < 0.001), respectively. Furthermore, inhibitors of the MAPK kinase, MEK (PD98059) and PKC (GF 109203X) prevented the induction of in vitro angiogenesis by ethanol.