To study whether nipradilol, which is used as an ophthalmic solution for the treatment of glaucoma, has a cytoprotective effect, we investigated its effect on the apoptosis induced by serum withdrawal in PC12 cells. Nipradilol has alpha1- and beta-adrenoceptor-blocking and nitric oxide (NO)-donating properties. We also investigated the effects of timolol, prazosin and S-nitroso-N-acetylpenicillamine (SNAP) on PC12 cell death. Serum withdrawal from PC12 cells resulted in apoptosis, and the survival rate was decreased in a time-dependent manner. The addition of nipradilol to the medium showed a cytoprotective effect on PC12 cell death in a dose-dependent manner, but timolol and prazosin did not. We measured caspase-3 activity to clarify the mechanism of the inhibition of apoptosis in the presence or absence of dithiothreitol (DTT). The caspase-3 activity could be reactivated by DTT. In addition, to investigate the relationship of the cGMP-dependent pathway to the nipradilol-induced cytoprotective effect, we tested the effect of the protein kinase G inhibitor KT5823. KT5823 partially reversed the nipradilol-mediated cytoprotective effect. These results indicate that the cytoprotective effect of nipradilol in PC12 cell death was due to the caspase-3 inhibition mediated by NO-related S-nitrosylation and activation of protein kinase G.