Pain - facilitating medullary neurons contribute to opioid - induced respiratory depression 1 2 Abbreviated title : Separating analgesia from respiratory depression 3 4 5

40 Respiratory depression is a therapy-limiting side-effect of opioid analgesics, yet our 41 understanding of the brain circuits mediating this potentially lethal outcome remains incomplete. 42 Here we studied the contribution of the rostral ventromedial medulla (RVM), a region long 43 implicated in pain modulation and homeostatic regulation, to opioid-induced respiratory 44 depression. Microinjection of the μ-opioid agonist DAMGO in the RVM of lightly anesthetized 45 rats produced both analgesia and respiratory depression, showing that neurons in this region can 46 modulate breathing. Blocking opioid action in the RVM by microinjecting the opioid antagonist 47 naltrexone reversed the analgesic and respiratory effects of systemically administered morphine, 48 showing that this region plays a role in both the analgesic and respiratory depressant properties 49 of systemically administered morphine. The distribution of neurons directly inhibited by RVM 50 opioid microinjection was determined using a fluorescent opioid peptide, dermorphin-Alexa594, 51 and found to be concentrated in and around the RVM. The non-opioid analgesic improgan, like 52 DAMGO, produced antinociception but, unlike DAMGO, stimulated breathing when 53 microinjected into the RVM. Concurrent recording of RVM neurons during improgan 54 microinjection showed that this agent activated RVM ON-cells, OFF-cells and NEUTRAL-cells. 55 Since opioids are known to activate OFF-cells but suppress ON-cell firing, the differential 56 respiratory response to these two analgesic drugs is best explained by their opposing effects on 57 the activity of RVM ON-cells. These findings show that pain relief can be separated 58 pharmacologically from respiratory depression, and identify RVM OFF-cells as important 59 central targets for continued development of potent analgesics with fewer side-effects. 60 61