This study was designed to investigate the effects of pheromonal cues and specific behaviors within the male copulatory sequence on c-fos expression in the medial nucleus of the amygdala (Me), the bed nucleus of the stria terminalis (BNST), and the medial preoptic area (MPOA) of the Syrian hamster brain. Sexually experienced male hamsters were placed into clean testing arenas and were either: 1) left alone as handled controls; 2) exposed to female hamster vaginal secretion (FHVS) on cotton swabs; or mated to various end points of copulation with a sexually receptive female: 3) five intromissions, 4) one ejaculation, 5) five ejaculations, or 6) long intromissions, A seventh group of sexually naive control males 7) was left alone in the arena. The brains of these males were compared to those of the sexually experienced controls to determine whether exposure to cues associated with prior sexual experience could alter c-fos expression. In males exposed only to FHVS, Fos immunoreactivity (Fos-ir) increased within the posterodorsal Me, the anterodorsal part of the posteromedial BNST, and the magnocellular medial preoptic nucleus (MPNmag). Following one ejaculation, Fos-ir increased within the caudal posterodorsal Me, the dorsolateral MPOA, and the paraventricular nucleus of the hypothalamus. After multiple ejaculations, additional labeling was observed within the posteroventral part of the posteromedial BNST, the medial preoptic nucleus (MPN), the central tegmental field, and in cell clusters of the caudal posterodorsal Me and rostral posteromedial BNST. Fos-ir also increased within the posterodorsal Me, MPN, and MPNmag in sexually experienced control males exposed to the empty test chamber compared to sexually naive males exposed to an identical chamber. These results demonstrate that the mating-induced pattern of neuronal activation in sexually experienced males is dependent upon multiple factors, including prior sexual experience in the testing environment, investigation of FHVS, and the number of ejaculations achieved.