Temporary retinal ischemia in the rat leads to a proliferation of endothelial cells and glial cells. We tested the hypothesis that this proliferation is caused by a release of soluble mitogens from the ischemic retina. Conditioned media were prepared from normal rat retina and from retina that had undergone 2 h ischemia and 48 h reperfusion, at which time it showed intense mitotic activity. The conditioned media were placed in cultures of fibroblasts, bovine adrenal capillary endothelial cells, and rat brain astrocytes. Cell proliferation in vitro was stimulated by the retinal extracts in all cell cultures. However, the cell proliferation in cultures with conditioned media from normal retina was similar to that in cultures with conditioned media from ischemic and proliferating retina. Although these data are consistent with the presence of soluble growth factors in the retina, they also indicate that release of these growth factors into the surrounding milieu after transient retinal ischemia is not altered to a degree that would explain the dramatic increase in mitosis.