Canonical transient receptor potential (TRPC) channels are Ca2+-permeable, non-selective cation channels that are widely expressed in mammalian cells.Various molecules have been found to regulate TRPC both in vitro and in vitro, but it is unclear how heterotrimeric G proteins transmit external stimuli to affect TRPC5 function.Here, we demonstrated that TRPC5 was regulated in a bidirectional manner by the Gαs regulatory pathway.We found that the whole-cell TRPC5 current was increased by treatment with the β-adrenergic receptorspecific agonist isoproterenol (246 ± 36%, n=6), the adenylatecyclase activator forskolin (273 ± 6%, n=5), and the membrane permeable cAMP analogue 8-Br-cAMP (251 ± 63%, n=7).In addition, we observed robust Ca2+transients induced by isoproterenol utilizing a Ca2+ imaging technique in HEK293 cells.After pretreatment with Gαs signaling cascade agonists or effectors, the membrane localization of TRPC5 was significantly increased by treatment with ISO (155 ± 17%, n=3), FSK (172 ± 39%, n=3) or 8-Br-cAMP (216 ± 59%, n=3).However, when GTPγS or GTPγS and IP3 were included in the pipette solution, the ISO-induced TRPC5 current was attenuated or inhibited, respectively.In conclusion, we suggest that the Gαs pathway regulates TRPC5 not only via the inhibition of PKA phosphorylation but also through potentiating actions that are dependent on the facilitation of intracellular Ca2+ dynamics and increased channel trafficking.