AIM: To investigate possible intracellular signal molecules involved in diphenylhydantoin (DPH)-mediated apoptosis of cerebellar granule neurons (CGN) and explore possible molecular mechanisms of neurotoxicity of DPH. METHODS: Fluorescein diacetate (FDA) stain, hochest 33258 stain, and agar gel electrophoresis were used to test morphological and biological characters of primary CGN and cortical neurons (CN) in the presence or absence of 100 μmol/L DPH; Western blot and RT-PCR were employed to further investigate apoptotic/survival signal moleculars involved in the neuronal apoptotic signal transdution. RESULTS: DPH 100 μmol/L induced a typical apoptosis of CGN but had no toxicity on CN. Cerebellar granule neural apoptosis induced by 100 μmol/L DPH was significantly inhibited by pre-treatment with SB203580 (10 μmol/L) or CEP-11004 (1 μmol/L) for 1 h. DPH markedly upregulated the levels of phospho-c-Jun (active c-Jun), total c-Jun protein and c-jun mRNA in CGN. The levels of phosphoc-Jun dramatically elevated by DPH at 8 h were significantly inhibited by SB203580 (10 μmol/L) or CEP-11004 (1 μmol/L). Moreover, the activities of p44/42 (ERK1/ERK2), other members of MAP kinases and generally believed to be important survival effetors in CGN, were markedly suppressed. However, the activities of both JNK and p38 were little affected in the process of apoptosis of CGN induced by 100 μmol/L DPH. CONCLUSION: The selective toxicity of DPH on CGN is likely due to its ability to induce apoptosis of CGN, it is a process involved activation of c-Jun and suppression of the activity of p44/42.