This study aimed to investigate the effects of ketanserin on micturition mediated via the 5-HT2A receptor in the motoneuron nucleus of the Lumbosacral cord, as reflected in high frequency oscillations (HFOs) of intravesical pressure and the external urethral sphincter electromyogram (EUS-EMG) in anesthetized male rats. METHODS：: Male Sprague-Dawley rats were used. Cystometry and EUS-EMG were performed in all rats under urethane anesthesia to examine the variations after successive intrathecal (i.t.) administration of various doses of ketanserin into the lumbosacral cord. Immunofluorescence staining and Western blotting were made to observe the distribution of 5-HT2 A and -2C receptors in the lumbosacral cord motor neurons. Compared to the controls, ketanserin-treated rats showed a declined trend of dose-dependent manner in the HFOs, in accordance with the variation of EUS-EMG, while decreased micturition volume, voiding efficiency, and increased post-void residual volume was only observed at the dose of 0.1 mg/kg. The effects of ketanserin on the HFO and EUS-EMG activity were partially or completely reversed by the 5-HT2A/2C receptor agonist, DOI. Meanwhile, immunofluorescence staining and Western blot analysis showed that immunoreactivity of 5-HT2A receptor was higher than that of 5-HT2C, labeling in the lumbosacral cord motoneurons. The intrathecally administrated 5-HT2A receptor antagonist ketanserin can weaken the EUS bursting activity, decrease HFOs, and reduce voiding efficiency as dose dependently. The effects of ketanserin on micturition may be mainly mediated via the 5-HT2A receptors in the motoneuron nucleus of the lumbosacral cord.