For patients experiencing inadequate analgesia and intolerable opioid-related side effects on one strong opioid analgesic, pain relief with acceptable tolerability is often achieved by rotation to a second strong opioid. These observations suggest subtle pharmacodynamic differences between opioids in vivo. This study in rats was designed to assess differences between opioids in their in vivo profiles. Male Sprague Dawley rats were given single i.c.v. bolus doses of morphine, morphine-6-glucuronide (M6G), fentanyl, oxycodone, buprenorphine, DPDPE ([D-penicillamine(2,5) ]-enkephalin) or U69,593. Antinociception, constipation and respiratory depression were assessed using the warm water tail-flick test, the castor oil-induced diarrhoea test and whole body plethysmography respectively. These opioid agonists produced dose-dependent antinociception, constipation and respiratory depression. For antinociception, morphine, fentanyl and oxycodone were full agonists, buprenorphine and M6G were partial agonists, whereas DPDPE and U69,593 had low potency. For constipation, M6G, fentanyl and buprenorphine were full agonists, oxycodone was a partial agonist, morphine produced a bell-shaped dose-response curve, whereas DPDPE and U69,593 were inactive. For respiratory depression, morphine, M6G, fentanyl and buprenorphine were full agonists, oxycodone was a partial agonist, whereas DPDPE and U69,593 were inactive. The respiratory depressant effects of fentanyl and oxycodone were of short duration, whereas morphine, M6G and buprenorphine evoked prolonged respiratory depression. For the seven opioids we assessed, no two had the same profile for evoking antinociception, constipation and respiratory depression, suggesting that these effects are differentially regulated. Our findings may explain the clinical success of 'opioid rotation'. This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2.