Psychedelic-assisted psychotherapy gained considerable interest as a novel treatment strategy for fear-related mental disorders. Promising clinical results have been obtained with 3,4-m methylenedioxyamphetamine (MDMA)-assisted psychotherapy in patients with post-traumatic stress disorder (PTSD) but the underlying mechanism remains poorly understood. The serotonin 2A (5-HT2A) receptor is a key target underlying the effects of psychedelics on emotional arousal but its role in fear processing remains controversial. Using the psychedelic 5-HT2A/5-HT2C agonist 2,5-dimethoxy-4-iodoamphetamine (DOI) and 5-HT2A receptor knockout (KO) mice we investigated the effect of 5-HT2A receptor activation on emotional processing. We show that DOI administration did not impair performance in a spontaneous alternation task but reduced anxiety-like behavior in the elevated plus maze task. Moreover, we found that DOI did not block memory recall but diminished fear expression in a passive avoidance task. Similarly, we found that DOI reduced fear expression in an auditory fear conditioning paradigm, while it did not affect fear extinction learning in wild type (WT) mice. The effect of DOI on fear expression was abolished in 5-HT2A receptor KO mice. Administration of DOI induced a significant increase of c-Fos expression in specific amygdalar nuclei. Moreover, local infusion of the 5-HT2A receptor antagonist M100907 into the amygdala reversed the effect of systemic administration of DOI on fear expression while local administration of DOI into the amygdala was sufficient to suppress fear expression. Our data demonstrate that activation of 5-HT2A receptors in the amygdala suppresses fear expression, but provide no evidence for an effect on fear extinction.