Aim:The most prevalent central nervous system malignancy is glioblastoma multiforme. Despite current therapy strategies, the recurrence and mortality rates remain high. Ceramides are reportedly as the main mediators of antiproliferative cellular responses such as apoptosis and growth inhibition. We aimed to evaluate the cytotoxic and proapoptotic effects of C6 ceramide on the C6 rat glioma cell line.Material and Methods:The C6 rat glioma cell line was evaluated. Using a confocal microscope and the appropriate software, the cytotoxic effects of C6 ceramide were identified using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H tetrazolium bromide (MTT) colorimetric experiments. Transmission electron microscopy (TEM) was utilized to examine the ultrastructural changes following treatment with IC50 concentrations of C6 ceramideResults:Condensation and fragmentation of nuclei and DNA laddering was observed, indicating apoptotic cell death. C6 ceramide induced apoptosis and effectively caused cytotoxicity in the C6 glioblastoma cells. MTT assay demonstrated >90% cell death after short-term application of C6 ceramide, confirming its apoptosis-triggering effect. Apoptosis was also confirmed via confocal microscopy and TEM.Conclusion:Glioblastoma cells undergo apoptosis when exposed to C6 ceramide, which makes it a potential chemotherapeutic agent for the treatment of this aggressive brain cancer.