4-Phenyl-N-methylpyridinium (MPP⁺), the xxidation product of the neurotoxic amine MPTP, is considerably more inhibitory to the oxidation of NAD⁺-linked substrates in intact mitochondria in State 3 than is 4-phenylpyridine. On adding uncouplers, the inhibition by MPP⁺ progressively diminishes, while the effect of 4-phenylpyridine remains. This is in accord with the fact that MPP⁺ is rapidly concentrated in the mitochondria by an energy-dependent process, while 4-phenylpyridine seems to enter passively with the concentration gradient. Collapse of the electrical gradient after addition of uncouplers thus leaves the inhibiton by 4-phenylpyridine unaffected but causes efflux of MPP⁺ from the mitochondria and a reversal of its inhibitory action. In isolated inner membranes the inhibition of NADH oxidation via the respiratory chain by 4-phenylpyridine is much greater than by MPP⁺. MPTP and 4-phenyl-N-methylpyridinone also inhibit more than MPP⁺, whereas N-methylpyridinium has relatively little effect. The block is not at the point of entry of electrons into the flavoprotein since the NADH-ferricyanide activity is not inhibited by MPP⁺ at V_max.