Autism spectrum disorder (ASD) is a serious neurodevelopmental disorder characterized by impairments in social interaction, language, and communication and induction of stereotypic behavior. In rodents, prenatal administration of valproic acid (often on 12.5 gestational days) is used for the induction of an ASD-like model. In the present study, we aimed to assess the potential therapeutic effects of crocin (a major component of Saffron, a neuroprotective and anti-inflammatory agent) on behavioral dysfunctions with respect to the level of brain-derived neurotrophic factor (BDNF) and glycogen synthase kinase-3 beta (GSK-3beta) in the medial prefrontal cortex. Valproic acid was intraperitoneally injected at the dose of 600 mg/kg on 12.5 gestational days. BDNF and GSK-3beta expression levels were also measured using real-time PCR. Locomotion, anxiety-like behavior, grooming, and sniffing were also measured in the open-field test. The results showed that prenatal valproic acid administration induced hyperactivity, anxiety-like behavior, increased grooming and sniffing (stereotyped behavior), decreased BDNF levels, and increased GSK-3beta levels in the medial prefrontal cortex. However, crocin dose-dependently restored the effects of prenatal valproic acid administration on behavioral functions and gene expressions. In conclusion, we suggested that BDNF and GSK-3beta expression changes in the medial prefrontal cortex may underlie the pathophysiology of ASD. The therapeutic effects of crocin may be also related to counteracting BDNF and GSK-3beta expression changes induced by prenatal valproic acid.