G-CSF promotes survival of immunopurified rat RGCs after neurotrophic factor deprivation. RGCs from 6- to 8-day old rat pups were immunopurified and cultured under full neurotrophic support for 24 hours before neurotrophins were withdrawn. (A) RT-PCR from primary RGCs demonstrates G-CSFR expression at the mRNA level. The resulting PCR product has the expected 233 bp size. (B, C) G-CSFR immunocytochemistry on primary RGCs demonstrates receptor expression at the protein level. (C) shows higher power magnification. (D) RGCs were cultured under full neurotrophic support for 24 h. Cells were then deprived of neurotrophins, and at the same time G-CSF (1, 10, 50 or 100 ng/ml) or vehicle was added. The number of surviving RGCs was determined by MTT staining after an additional 2 days in vitro. A bell-shaped dose-response curve is observed, demonstrating significantly enhanced survival of RGCs compared with the vehicle-treated control cultures at a concentration of 10 and 50 ng/ml. * p < 0.05 (compared with vehicle). Data are given as mean ± S.E.M.