Effects of diphenylhydantoins on (Na⁺ + K⁺)-ATPase activity in rat and cat brain microsomes were studied. 5,5-diphenylhydantoin (DPH) in concentrations of 5-20 μg ml^-1 produces an apparent stimulation of the rat brain (Na⁺ + K⁺)-activated ATPase of 55-65% in media containing 50 mM Na⁺, 0.15 mM K⁺, 3 mM Mg⁺⁺, and 3 mM ATP. No effects are found on the Mg-ATPase. At constant K⁺ levels of 0.05 mmole/liter and 0.15 mmole/liter, increasing the Na⁺ concentration activates the enzyme similarly with and without DPH. However, Na⁺ concentrations greater than 5 mmoles/liter and 10 mmoles/liter, respectively, which are inhibitory in these low K⁺ media, produce less inhibition in the presence of DPH. In media containing 10 mM Na⁺, the K⁺ activation, on the other hand, is potentiated by DPH. In preparations from cat brain qualitatively similar results are obtained. No effect of DPH is seen on the inhibition produced by high K⁺ in low Na⁺ media. DPH produces an approximately constant apparent stimulation of 45% in the (Na⁺ + K⁺) increments when these ions are varied simultaneously at a fixed ratio of 150 Na⁺:1 K⁺ with cat brain extracts. 5-(p-hydroxyphenyl)-5-phenylhydantoin (HPPH) has the same potency as DPH in reducing the Na⁺ inhibition at high Na:K ratios. The hydantoins appear to act by decreasing the Na⁺ inhibition that occurs at high Na:K ratios.