Mammalian cells require a constant O₂ supply to maintain adequate energy production and sophisticated mechanisms have evolved to allow cells to sense hypoxia. In this study, we investigated the role of transient receptor potential (TRP) channels and the Na-Ca exchanger (NCX) in mediating hypoxia-induced [Ca²⁺]_i elevation in a rat pheochromocytoma (PC12) cell line, a model of O₂-sensing. In these cells, PCR and western blot confirmed the presence of TRPC1, 3 and 6 as well as NCX. Hypoxia consistently induced a reversible, Ca²⁺ elevation, which was sensitive to nifedipine (20mM). The block of TRPC by 2-APB (100 mM) and SKF 96365 (40 mM) significantly reduced hypoxia-mediated [Ca²⁺]_i elevation by 52 ± 7.5% and 56 ± 9.1%, respectively. The activation of TRPC in mediating the Ca²⁺ response to hypoxia was, in part, via the G-protein and PLC pathway as it was blocked by 27-30% by suramin (50mM) and U73122 (10mM). In addition to TRPC, NCX also contributed to the hypoxia response and blockage of NCX by KBR9473 (10mM) significantly attenuated the hypoxia induced [Ca²⁺]_i elevation by 41 ± 7.6%. Our results show that hypoxia induced Ca²⁺ can occur via TRPC and NCX channels in PC12 cells. We suggest that TRPC channels may play a major role in mediating the hypoxia response in these cells, perhaps by contributing to cell depolarization.