Slices of cat brain respiring in a Krebs–Ringer bicarbonate medium were found to incorporate radioactive inorganic phosphate (P32) into the phospholipid fraction. The addition of glucose or mannose increased the incorporation of P32into the phospholipids. Fructose caused a small increase, whereas galactose was without effect. Pyruvate and lactate increased the incorporation of P32into the phospholipids. Succinate, L (+)-glutamate, D (−)-glutamate, α keto-glutarate, citrate, and L-malate failed to support the incorporation.Anaerobic conditions and homogenization of the tissue prevented the in vitro incorporation of P32into the phospholipids of cat brain. A wide range of metabolic inhibitors (cyanide, azide, malononitrile, chloretone, nembutal, iodoacetate, and fluoride), in concentrations that inhibit the oxygen consumption of brain slices, inhibited the incorporation. The incorporation was also inhibited by 2,4-dinitrophenol in concentrations that do not decrease the oxygen consumption of brain slices.These findings indicated that the incorporation of P32into the phospholipids of slices of cat brain is a metabolic phenomenon and is dependent upon the maintenance of an adequate phosphorylating mechanism within the slice.