Since a growing number of studies based on the real-time reverse transcriptase polymerase chain reaction (RT-PCR) continue to be published in order to highlight genes specifically involved in brain development, maturation, and function, the identification of reference genes suitable for this kind of experiments is now an urgent need in the neuroscience field. The aim of this work was to verify the suitability of some very common housekeeping genes (such as Gapdh, 18s, and B2m) and of some relatively new control genes (such as Pgk1, Tfrc, and Gusb) during mouse brain maturation. We tested the candidate reference genes in mouse whole brain, cerebellum, brain stem, hippocampus, medial septum, frontal neocortex, and olfactory bulb. Moreover, we reported the first complete study of Pgk1 expression throughout the development and the aging of mouse brain. Although no tested gene showed to be the optimal reference for all mouse brain regions, in general, the new housekeeping genes were highly stable in most of the analyzed regions. Above all, with few exceptions, Pgk1 showed to be a reliable control for the analyzed mouse brain regions during development, maturation, and aging.