AbstractAimIn our pilot study, we found an increase in tyrosine hydroxylase (Th) mRNA expression in the prefrontal cortex of 72‐h REM sleep‐deprived (SD) rats, a mania model. Additionally, the expression levels of miR‐325‐3p, miR‐326‐3p, and miR‐330‐5p, the predicted target miRNAs on TH, were significantly decreased. Based on these results, in this study, we investigated whether miRNA‐325‐3p, miR‐326‐3p, and miR‐330‐5p modulate TH and manic‐like behaviors in SD rats.MethodsManic‐like behaviors were assessed using the open field test (OFT) and elevated plus‐maze (EPM) test. The direct binding activity of miRNAs to the 3′‐untranslated region (3′‐UTR) of theThgene was measured in HEK‐293 cells using a luciferase reporter system. We also examined mRNA and protein expression of TH after intracerebroventricular (ICV) injection of miR‐330‐5p agomir to SD rats, along with manic‐like behaviors.ResultsWe observed an upregulation in mRNA and protein expression of TH and downregulation in miRNA‐325‐3p, miR‐326‐3p, and miR‐330‐5p expressions in the prefrontal cortex of SD rats, together with increased manic‐like behaviors. The luciferase reporter assay showed that miR‐330‐5p could repress TH expression through direct binding to its target site in the 3′‐UTR of Th, whereas miR‐326‐3p and miR‐330‐5p could not. In addition, ICV injection of miR‐330‐5p agomir alleviated the increase in TH expression in the prefrontal cortex of SD rats and manic‐like behaviors.ConclusionsTH expression regulation through miR‐330‐5p may be implicated in the pathophysiology of mania in SD rats.