Type-III sodium-dependent phosphate transporters 1 and 2 (PiT-1 and PiT-2, respectively) are proteins encoded by SLC20A1 and SLC20A2, respectively. The ubiquitous distribution of PiT-1 and PiT-2 mRNAs in mammalian tissues is in agreement with the housekeeping maintenance of homeostasis of intracellular inorganic phosphate (Pi), which is absorbed from interstitial fluid for normal cellular functions. Recently, mutations of SLC20A2 have been found in patients with idiopathic basal ganglia calcification (IBGC), also known as Fahr's disease. However, the localization of PiT-2 in the brain has not been clarified yet. Therefore, the aim of this study is to clarify the distribution of PiT-2 expression in the mouse brain. Our biochemical and immunohistochemical analyses using a polyclonal antibody (Ab) and a monoclonal Ab showed that PiT-2 was ubiquitously expressed throughout the brain. In terms of the cellular type, PiT-2 was predominantly detected in neurons; it colocalized with β-tubulin III in the cerebral cortex and with calbindin D-28K in Purkinje cells. Additionally, PiT-2 immunopositivity was detected in the microtubule-associated protein 2-positive neuronal dendrites in the cerebral cortex. However, colocalization with PiT-2 immunopositivity was not observed in the synaptophysin-positive nerve terminals. PiT-2 was also expressed in astrocytes and vascular endothelial cells. Our results indicate that PiT-2 plays an important role in the maintenance of cellular Pi homeostasis in neurons, astrocytes, and endothelial cells. This finding is a milestone in the study of the function of PiT-2 in the normal mouse brain and particularly in the brains of patients with Fahr's disease.