['Despite success in treating many forms of cancer, pain associated with malignancy remains a serious clinical issue with a poorly understood etiology. This study determined if certain sarcoma cell lines produced a soluble factor that activates the TRPV1 ion channel expressed on nociceptive sensory neurons, thereby activating a major pain transduction system.', 'Trigeminal ganglia were harvested from rats and cultured. A rhabdomyosarcoma (CRL1598) and osteosarcoma (CRL 1543) cell line were grown to 75% confluency. Conditioned media (CM) was collected after 24\xa0h of exposure and subjected to reverse phase chromatography. Neuronal activation in the presence of CM was measured using iCGRP RIA and calcium imaging after treatment with vehicle or I-RTX, a potent TRPV1 antagonist. Data were analyzed by ANOVA/Bonferroni or t test.', 'The rhabdomyosarcoma CM produced a 4-fold increase in iCGRP release compared with control media (P\xa0<\xa00.001). The osteosarcoma cell line CM produced a 7-fold increase in iCGRP release compared with control media (P\xa0<\xa00.001). This evoked iCGRP release was via TRPV1 activation since the effect was blocked by the antagonist I-RTX. The application of rhabdomyosarcoma CM produced about a 4-fold increase in [Ca(2+)]I levels (P\xa0<\xa00.001), and this effect was blocked by pretreatment with the TRPV1 antagonist, I-RTX.'] Do in vitro sarcoma cells release a lipophilic substance that activates the pain transduction system via TRPV1?