Valproic acid (VPA) is a widely used anti-epileptic drug. If administered to pregnant women, it causes autism spectrum disorder (ASD) in their progeny, however, the molecular mechanism by which VPA causes ASD remains unknown. Since it is reported that VPA inhibits several classes of histone deacetylases (HDACs) that regulate gene expression, we hypothesized that the alteration in autism-related gene expression is involved in VPA-induced ASD pathogenesis. Utilizing neuroblastoma cell line Neuro2a, we found that VPA reduces mRNA level of Setd5 gene whose heterozygous loss-of-function mutations are identified in autism patients. To investigate whether HDACs are involved in the regulation of Setd5 expression, we then treated another HDAC inhibitor Entinostat which inhibits class I HDACs, revealing that Entinostat also reduces Setd5 mRNA. Class I HDACs is constituted by HDAC1, HDAC2, HDAC3 and HDAC8. To determine which HDAC regulates Setd5 expression, we then individually deleted these genes and identified HDAC3 that positively regulate Setd5 expression, which is contrary to the well-established function of HDAC3 that represses gene expression. Therefore, we speculated that HDAC3 indirectly controls Setd5 expression. Consistently, inhibition of de novo protein synthesis rescued VPA-induced downregulation of Setd5 mRNA. Taken together, our data suggest that VPA represses Setd5 expression that is mediated by HDAC3-regulated gene, which at least in part contributes to ASD pathogenesis.