Most associative learning tests in rodents use negative stimuli, such as electric shocks. We investigated if young rats can learn to associate the presence of an odour with the experience of being tickled (i.e. using an experimenter’s hand to mimic rough-and-tumble play), shown to elicit 50 kHz ultrasonic vocalisations (USVs), which are indicative of positive affect. Male, pair-housed Wistar rats (N = 24) were all exposed to two neutral odours (A and B) presented in a perforated container on alternate days in a test arena. Following 60s of exposure, the rats were either tickled on days when odour A (n = 8) or odour B (n = 8) was present, or never tickled (n = 8). When tickled, rats produced significantly more 50 kHz USVs compared to the days when not being tickled, and compared to control rats. The level of anticipatory 50 kHz USVs in the 60s prior to tickling did not differ significantly between the tickled and control rats. As a retrieval test following the odour conditioning, rats were exposed successively in the same arena to three odours: an unknown neutral odour, extract of fox faeces, and either odours A or B. Compared to controls, 50 kHz USVs of tickled rats increased when exposed to the odour they had previously experienced when tickled, indicating that these rats had learned to associate the odour with the positive experience of being tickled. In a test with free access for 5 min to both arms of a T-maze, each containing one of the odours, rats tickled with odour A spent more time in the arm with this odour. This work is the first to test in a fully balanced design whether rats can learn to associate an odour with tickling, and indicates that positive odour conditioning has potential to be used as an alternative to negative conditioning tests.

Most associative learning tests in rodents use negative stimuli, such as an electric shock. We investigated if young rats can learn to associate the presence of an odour with the experience of being tickled (i.e. using an experimenter’s hand to mimic rough-and-tumble play), shown to elicit 50 kHz ultrasonic vocalisations (USVs), which are indicative of positive affect. Male, pair-housed Wistar rats (N=24) were all exposed to two neutral odours (A and B) presented in a perforated container on alternate days in a test arena. Following 60s of exposure, the rats were either tickled on days when odour A (n=8) or odour B (n=8) was present, or never tickled (n=8). When tickled, rats produced significantly more 50 kHz USVs compared to the days when not being tickled, and compared to control rats. The level of anticipatory 50 kHz USVs in the 60s prior to tickling did not differ significantly between the tickled and control rats. Following the odour conditioning, rats were exposed successively in the same arena to three odours: an unknown neutral odour, extract of fox faeces, and either odours A or B. Compared to controls, 50 kHz USVs of tickled rats increased when exposed to the odour they had previously experienced when tickled, indicating that these rats had learned to associate the odour with the positive experience of being tickled. In a test with free access for 5 min to both arms of a T-maze, each containing one of the odours, rats tickled with odour A spent more time in the arm with this odour. This work is the first to test in a fully balanced design whether odours can be conditioned to tickling, and indicates that positive odour conditioning has potential to be used as an alternative to negative conditioning tests.