During post-weaning development, a marked increase in peer–peer interactions is observed in mammals, including humans, which is signified by the abundance of social play behaviour. Social play is highly rewarding, and known to be modulated through monoaminergic neurotransmission. Recently, the habenula has received widespread attention because of its role in the regulation of monoaminergic neurotransmission as well as in a variety of emotional and cognitive functions. Therefore, in the present study, we investigated the involvement of the habenula in social play behaviour. Using the neuronal activity maker c-fos, we showed that the habenula was activated after 24 h of social isolation in adolescent rats, and that a subsequent social play interaction reduced c-fos activity in the medial part of the lateral habenula. This suggested that habenula activity modulated the aversive properties of social isolation, which was alleviated by the positive effects of social play. Furthermore, after functional inactivation of the habenula, using a mixture of the GABA receptor agonists baclofen and muscimol, social play behaviour was markedly reduced, whereby responsiveness to play solicitation was more sensitive to habenula inactivation than play solicitation itself. Together, our data indicate an important role for the habenula in the processing of positive (i.e. social play behaviour) and negative (i.e. social isolation) social information in adolescent rats. Altered habenula function might therefore be related to the social impairments in childhood and adolescent psychiatric disorders such as autism, attention deficit/hyperactivity disorder and early-onset schizophrenia.