Background: Learning and memory is a complex process. Some reports have shown that protein kinases (PKs) and phosphatases (PPs) are important mediators in this process. And it is also well known that protein serine/threonine phosphatase 1 (PP1) and DNA methylation are critically involved in learning and memory. Methods: In the current study, the mice and cultured cells (NG108-15) were treated with vehicle or 5-Aza-20-deoxycytidine (5-aza-cdR), a DNA methyltransferase (DNMT) inhibitor. The ability of learning and memory of mice was detected by Morris Water Maze, while the mRNA and protein expression levels of DNMTs and PP1 gamma in mice hippocampus were measured by real-time PCR and western-blot. To further clarify whether the 5-aza-cdR effects on learning and memory depend on cell proliferation/apoptosis or not, the effects of 5-aza-cdR on the cell proliferation, apoptosis, and PP1 gamma transcriptional activity were analyzed by using the xCelligence system, flow cytometer and Luciferase reporter assay, respectively. Results: The ability of learning and memory was increased while the expressions of DNMTs and PP1 gamma were decreased in the hippocampus of mice which were injected with 5-aza-cdR. In vitro experiments showed 10 mM 5-aza-cdR inhibited cell proliferation, decreased PP1 gamma transcription without inducing apoptosis. Conclusion: Our data demonstrate that the 5-aza-cdR restrains the expression of PP1 gamma which is related to learning and memory in the mice. (C) 2016 Elsevier Masson SAS. All rights reserved.