We have recently documented that cocaine-induced MEF2C expression in rat brain is mediated by the activation of the salt-inducible kinase SIK1, which is itself regulated by phosphorylation mechanisms. We report here that acute or chronic treatment of rats with cocaine increased the expression of the catalytic subunit of protein phosphatase PP2A in the prefrontal cortex and striatum. Cocaine treatment also reduced the number of cortical neurons expressing SIK1 phosphorylated on Ser-577, but not on Thr-182. To further support the hypothesis that phospho-SIK1-S577 is a substrate of PP2A, we used Neuro-2A cell cultures overexpressing either the wild-type PP2A, in which the amount of phospho-SIK1-S577 was found to be decreased, or a mutant PP2A devoid of enzymatic activity, in which the level of phospho-SIK1-S577 was increased when compared to control cells. The data indicate that, by inducing PP2AC, cocaine regulates the nuclear location and activity of SIK1 and HDAC5, which ultimately govern the activity of CREB and MEF2C transcription factors. The results highlight PP2A as a novel target for regulating long-term effects of cocaine.