Colonies of naked mole-rats (Heterocephalus glaber, NMRs) are characterised by an extreme skew in lifetime reproductive success with only one female and one to three male consorts in a colony. The rest of the individuals in a colony are reproductively suppressed and much research has been focussed on elucidating that mechanism. The dopamine system and prolactin have recently been implicated in the suppression of reproduction of subordinate NMRs. To investigate the changes in prolactin during the removal of an aged reproductive female (queen) and succession of a new queen, blood samples were collected during different stages of queen removal: before queen removal, after separation, but in olfactory contact with the queen and after the total removal of the queen. Further, plasma cortisol and testosterone concentrations were determined. The colony appeared unstable prior to queen removal as indicated by high concentrations of cortisol and testosterone and lack of successful breeding. A new queen succeeded the old queen whilst she was still in olfactory contact. The time preceding queen succession was characterised by high levels of aggression, the death of a number of individuals, high cortisol and testosterone and low prolactin concentrations. Once the older queen was removed entirely and the new queen had given birth, prolactin concentrations increased and cortisol and testosterone concentrations decreased in subordinate NMRs. The results suggest that low prolactin levels are associated with low reproductive suppression during times of colony instability due to the removal or death of a queen.