Methadone, a potent opioid analgesic, has long been considered a mu-opioid, based upon the similarities between its actions and those of morphine. This classification is supported by the sensitivity of methadone analgesia to the highly mu-opioid receptor-selective antagonist beta-funaltrexamine. Yet, CXBK mice respond normally to methadone despite their insensitivity to systemic morphine, distinguishing between the receptor mechanisms of the two drugs. Beta-funaltrexamine antagonizes methadone analgesia in CXBK mice, implying that the opioid is still acting through a mu-opioid receptor. These results reveal distinct analgesic mechanisms for morphine and methadone and provide further support for multiple subtypes of mu-opioid receptors.