Monocrotaline (MCT) is a pyrrolizidine alkaloid (PA) of large occurrence in plants of Crotalaria genus (leguminosae) and causes intoxication in humans and in animals. It has been demonstrated that MCT is metabolised to dehydromonocrotaline (DHMCT) by hepatic cytochrome P450, and it may alkylate macromolecules, such as proteins and DNA, leading to formation of adducts, which can initiate a chronic or acute intoxication. However, more susceptible animals to its intoxication, such as horses, may present nervous clinical signs. In this study, we evaluated if MCT, extracted from Crotalaria retusa, can induce genetic damage in a human glioblastoma cell line (GL-15). Confluent cell cultures, grown in DMEM supplemented medium, were treated for 72 h with 1 μM, 10 μM or 100 μM MCT. Cells treated with the MCT vehicle (0.5% DMSO) were used as the negative control, and cells directly exposed to UV light for 1h at 1m from the source was used as the positive control. The results showed that unlike what was found for UV exposure, 1 μM MCT failed to induce a significant increase on cell DNA damage evidenced by the Comet test. However, exposure of GL-15 cells to 10-100 μM MCT caused significant increases in cell damage index. The data obtained provide support to the view that the alkaloid MCT acts directly on glial cells and induces genotoxicity.