Brain histamine H₃ receptors are predominantly presynaptic and serve an important autoregulatory function for the release of histamine and other neurotransmitters. They have been implicated in a variety of brain functions, including arousal, locomotor activity, thermoregulation, food intake, and memory. The recent cloning of the H₃ receptor in our laboratory has made it possible to create a transgenic line of mice devoid of H₃ receptors. This paper provides the first description of the H₃receptor-deficient mouse (H₃ ^−/−), including molecular and pharmacologic verification of the receptor deletion as well as phenotypic screens. The H₃ ^−/− mice showed a decrease in overall locomotion, wheel-running behavior, and body temperature during the dark phase but maintained normal circadian rhythmicity. H₃ ^−/− mice were insensitive to the wake-promoting effects of the H₃ receptor antagonist thioperamide. We also observed a slightly decreased stereotypic response to the dopamine releaser, methamphetamine, and an insensitivity to the amnesic effects of the cholinergic receptor antagonist, scopolamine. These data indicate that the H₃receptor-deficient mouse represents a valuable model for studying histaminergic regulation of a variety of behaviors and neurotransmitter systems, including dopamine and acetylcholine.