ObjectiveTo examine the effect of ketanserin and naloxone on fentanyl-induced motor activity in isoflurane-anaesthetized pigs. Study designRandomized blinded prospective two-group study. AnimalsA group of 12 crossbred pigs weighing 22-31 kg. MethodsFentanyl was administered to isoflurane anaesthetized pigs at 7.5 μg kg-1 hour-1 for 40 minutes intravenously, followed by an intravenous injection of naloxone 0.1 mg kg-1 or ketanserin 1 mg kg-1. Electromyography (EMG) and accelerometry were used to record motor unit activity and tremors, respectively. To test the effect of drug administration on motor activity, data from a 5-minute period at baseline, immediately before and after antagonist injection was compared in a mixed model. p < 0.05. ResultsResults are reported with the median difference, 95% confidence intervals and corresponding p-values in brackets. Fentanyl significantly increased EMG activity [30.51 (1.84–81.02) μV, p = 0.004] and induced tremors [0.09 (0.02–0.18) m s-2, p < 0.001] in 10 of 12 pigs. EMG [32.22 (6.29–136.80) μV, p = 0.001] and tremor [0.10 (0.03–0.15) m s-2, p = 0.007] activity was significantly reduced by ketanserin. No significant effect was found for naloxone on EMG [26.76 (-13.28–91.17) μV, p = 0.4], or tremors [0.08 (-0.01–0.19) m s-2, p = 0.08]. Conclusions and clinical relevanceFentanyl can induce motor activity in anaesthetized pigs, with a suggested link to the serotonergic system. This study shows that ketanserin can antagonise this activity, which supports the role of serotonin. This knowledge contributes to the general understanding of the motor effects of fentanyl and especially the problem of tremors in anaesthetized pigs.