Abstract: Recent studies have shown that keratinocytes can sense temperature via thermo-transient receptor potential (TRP) channels. It is not known whether other thermosensitive ion channels such as TREK-1, TREK-2 and TRAAK (TREKs/TRAAK) that are members of the two-pore domain K+ (K2P) channel family are expressed in human keratinocytes. Here, we identified the expression of TREKs/TRAAK in human keratinocytes-derived cell line HaCaT cells using RT-PCR, immunocytochemistry, Western blot analysis and patch-clamp technique. RT-PCR showed that all six K2P channels tested (TASK-1, TASK-3, TREK-1, TREK-2, TRAAK and TASK-2) were expressed in HaCaT cells, as well as in skin and dorsal root ganglion (DRG) of rat. The expression of TREKs/TRAAK mRNA identified by RT-PCR was further studied at the protein level. Using anti-TREK-1, -TREK-2 and -TRAAK, bands of ∼46, ∼60 and ∼43 kDa, respectively, were observed at plasma membrane of HaCaT cells. Immunostaining also showed that TREK-1, TREK-2 and TRAAK were expressed in all area of cells including plasma membrane. Whole-cell K+ currents recorded from HaCaT cells were activated by arachidonic acid and heat. These results suggest that TREKs/TRAAK channels could act as thermosensors in human keratinocytes.