Locomotor activity measures are often used in behavioral neuroscience. There is, however, a large variability in the protocols assessing locomotor activity which may, more or less, strongly be influenced by exploration and reactivity to novelty in unfamiliar situations. Using Long-Evans male rats, we investigated how far changes, such as placing rats in a cage physically identical to the home cage supplied with fresh sawdust but kept in a familiar room, or placing the familiar home cage with the rat inside in another (unfamiliar) room, may influence the level of locomotor activity. We showed that both changes resulted in significantly increased locomotion in the first 2 h after placing the rats in the respective test situation, but there is no significant additive effect. These changes performed right before the start of the test do not alter diurnal or nocturnal locomotor activity once the first 2 h have elapsed. The results illustrate that rats kept in an environment with stable proximal features (cage, sawdust) can react by increased activity in response to more distal novelty (experimental room), and conversely, that rats in a familiar environment react to proximal changes in the home cage.