Date

September 2010

Document Type

Thesis

Degree Name

Ph.D.

Institution

Oregon Health & Science University

Abstract

Morphine has a potency and efficacy for producing analgesia and a pharmacokinetic profile that make it favorable for use in pain treatment. Continued use of morphine and other opioids results in some degree of tolerance, such that the dose needs to be escalated to achieve the same effect. This can become problematic in the treatment of chronic pain, as elevated doses of opioids can produce unwanted side effects such as respiratory depression and dependence. Methadone, a synthetic agonist, is being increasingly used in the treatment of pain. In cancer patients, less dose escalation is required for those who are receiving sustained release methadone versus morphine. Thus the development of tolerance may differ depending on the agonist being used and understanding the adaptations that produce tolerance will be useful for pain treatment. The goal of this work is to understand the cellular mechanisms involved in desensitization and resensitization of mu opioid receptors in the locus coeruleus (LC) and how these processes are regulated by acute and long-term administration of various opioids. To do this electrophysiological recording was used to measure mu opioid receptor (MOR) stimulated hyperpolarizations and 2-photon imaging of epitope-tagged receptors was used to monitor MOR trafficking in neurons in live brain slices. Desensitization and internalization induced by various agonists was examined in opiate naïve LC neurons. [Met[superscript 5]]-enkephalin, methadone and etorphine resulted in both desensitization and internalization while oxycodone caused neither. Morphine and oxymorphone were unique in that they caused desensitization without internalization of MOR, demonstrating that these processes are separately engaged. Chronic treatment with morphine resulted in reduced recovery from ME-induced desensitization that was associated with less recycling of internalized receptors back to the plasma membrane. Reduced recovery from desensitization did not occur in slices from mice lacking functional G protein receptor kinase (GRK) or ßarrestin2 (ß-arr2). Thus, chronic morphine treatment altered GRK2 and ß-arr2 function resulting in inhibited recovery from desensitization. Changes in the GRK2/ß-arr2 pathway were specifically engaged by morphine and reduced recovery from desensitization and receptor recycling were not observed after treatment with methadone. The frequency of presynaptic excitatory inputs onto LC neurons was enhanced following chronic morphine treatment and this was dependent on PKA activity. However, no post-synaptic changes in efficacy or potency of ME were observed in mouse LC neurons after chronic morphine treatment. Thus, reports of increased excitability in LC neurons during withdrawal are likely due to a presynaptic mechanism that involves an upregulation in the cAMP/PKA pathway. The results of this work highlight the complexity of signaling mechanisms involved in mu-opioid receptor regulation. Acute application of opioid agonists can specifically engage regulatory mechanisms to various degrees, and differences in acute regulation may contribute to the morphine specific adaptations. Interestingly, morphine, which does not induce internalization of opioid receptors, alters trafficking induced by other agonists in such a way that MOR cannot recover from desensitization, and thus long-term signaling is dampened.

Identifier

doi:10.6083/M4736NWT

Division

Neuroscience Graduate Program

School

School of Medicine

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