Lactate has diverse roles in the brain at the molecular and behavioral levels under physiological and pathophysiological conditions. This study investigates whether lysine lactylation (Kla), a lactate-derived post-translational modification in macrophages, occurs in brain cells and if it does, whether Kla is induced by the stimuli that accompany changes in lactate levels. Here, we show that Kla in brain cells is regulated by neural excitation and social stress, with parallel changes in lactate levels. These stimuli increase Kla, which is associated with the expression of the neuronal activity marker c-Fos, as well as with decreased social behavior and increased anxiety-like behavior in the stress model. In addition, we identify 63 candidate lysine-lactylated proteins and find that stress preferentially increases histone H1 Kla. This study may open an avenue for the exploration of a role of neuronal activity-induced lactate mediated by protein lactylation in the brain.