Transient receptor potential vanilloid subtype 1 (TRPV1) is a polymodel sensory receptor and can be activated by moderate temperature (≥ 43 °C). Though extensive researches on the heat-activation mechanism revealed some key elements that participate in the heat-sensation pathway, the detailed thermal-gating mechanism of TRPV1 is still unclear. We investigate the heat-activation process of TRPV1 channel using the molecular dynamics simulation method at different temperatures. It is found that the favored state of the supposed upper gate of TRPV1 cannot form constriction to ion permeation. Oscillation of S5 helix originated from thermal fluctuation and forming/breaking of two key hydrogen bonds can transmit to S6 helix through the hydrophobic contact between S5 and S6 helix. We propose that this is the pathway from heat sensor of TRPV1 to the opening of the lower gate. The heat-activation mechanism of TRPV1 presented in this work can help further functional study of TRPV1 channel.