Abstract This study aimed to identify behaviours that could be used to assess post-operative pain and analgesic efficacy in male rabbits. In consideration of the ‘Three Rs’, behavioural data were collected on seven male New Zealand White rabbits in an ethically approved experiment requiring abdominal implantation of a telemetric device for purposes other than behavioural assessment. Prior to surgery, rabbits were anaesthetised using an isoflurane/oxygen mix and given Carprofen (2 mg kg −1 ) as a peri-operative analgesic. Rabbits were housed individually in standard laboratory cages throughout. Data were collected at three time periods: 24-21 h prior to surgery (T1) and, post-surgery, 0-3 h (T2) and 3-6 h (T3). Behavioural changes were identified using Observer XT, significance of which was assessed using a Friedman's test for several related samples. The frequency or duration of numerous pre-operative behaviours was significantly reduced in T2 and T3, as compared to T1. Conversely, novel or rare behaviours had either first occurrence or significant increase in T2 into T3 as compared to T1, these include ‘full-body-flexing’, ‘tight-huddling’, ‘hind-leg-shuffling’. We conclude that reduced expression of common pre-operative behaviours and the appearance of certain novel post-operative behaviours may be indicative of pain in rabbits. Behaviours identified as increased in T2 as compared to T1 but not consistently elevated into T3 were considered separately due to the potentially confounding effect of anaesthesia recovery. These included lateral lying, ‘drawing-back’, ‘staggering’ and ‘closed eyes’. We postulate that for effective application of best-practice post-operative care, informed behavioural observation can provide routes by which carers may identify requirements for additional post-operative analgesia. Additionally, improvement of the peri-operative pain management regimen may be required to ameliorate the immediate effects of abdominal surgery. Comparisons with other studies into post-operative pain expression in rabbits suggest behavioural indicators of pain may differ, depending on housing and surgical procedure.