Understanding the mechanism of how fear memory can be extinguished could provide potential therapeutic strategies for the treatment of posttraumatic stress disorders. Here we show that infusion of CB1 receptor antagonist into the infralimbic (IL) subregion of the medial prefrontal cortex (mPFC) retarded cue-alone-induced reduction of fear-potentiated startle. Conversely, cannabinoid agonist WIN55212-2 (WIN) facilitated the extinction. Unexpectedly, administration of WIN without cue-alone trials reduced startle potentiation in a dose-dependent manner. The effect of cannabinoid agonists was mimicked by endocannabinoid uptake or fatty acid amide hydrolase inhibitors. Rats were trained with 10 conditioned stimulus (CS+) (yellow light)-shock pairings. Extinction training with CS+ (yellow light)-alone but not CS- (blue light)-alone trials decreased fear-potentiated startle. Intra-IL infusion of WIN before CS--alone trials decreased startle potentiation, suggesting that the cannabinoid agonist decreased conditioned fear irrespective of whether the rats underwent CS+- or CS --alone trials. Cannabinoid agonists activated extracellular signal-regulated kinases (ERKs) in mPFC slices, and ERK inhibitor blocked the effect of cannabinoid agonists on fear-potentiated startle. These results suggest that CB1 receptors acting through the phosphorylation of ERK are involved not only in the extinction of conditioned fear but also in the adaptation to aversive situations in general.