Morphine, one of the strongest analgesics known, is produced by and commercially isolated from the opium poppy plant. Over the past thirty years, claims have been published on the detection of trace quantities of morphine in mammals. The question of whether this morphine is synthesized de novo or whether it is of dietary origin was not solved until the recent success of incorporation of 18O2 regiospecifically into morphine in cultured human cells. It has now been demonstrated that human cells and mammalian tissues are capable of synthesizing low concentrations (circa 10 nM) of morphine, a concentration that is potentially sufficient to activate the u receptor. Whether this powerful analgesic produced in situ is physiologically active in mammals in, for instance, pain relief or mood modulation, is an open question that need now be addressed. Results from the research proposed herein may impact how pain and emotional disorders are both viewed and treated. The biosynthesis of morphine in mammalians diverges from the plant pathway in the early steps, but is then identical in the latter steps, suggesting a convergent evolution of the genes and encoded enzymes. A clear and complete understanding of the biosynthetic pathway in mammals will make available new biological targets for the potential treatment of pain and conditions related to mood. Towards this goal, several specific aims will be addressed. Techniques will be established by mass spectrometry and radioimmunoassay that allow routine measurement of low quantities of morphine in brain tissue. Stable isotope-labeled biosynthetic precursors such as L-DOPA will be synthesized with a labeling pattern that will allow the recognition of intact incorporation into morphine or incorporation after metabolic scrambling of label. Injection of labeled L-DOPA into mice labels the brain and spinal cord, but not the heart, lung, liver or kidneys. The precise region(s) of the brain and spinal cord in which morphine accumulates will now be determined by mass spectrometric / radioimmunioassay analyses of brain and spinal cord explants. These results will be compared to the in situ hybridization results published in brainatlas.org for genes of known function that are candidates for involvement in mammalian morphine biosynthesis such as those encoding tyrosine hydroxylase, DOPA decarboxylase and catechol O-methyltransferases. Once a brain region is identified, the general gene expression pattern provided in brainatlas.org may yield insight into the identities of additional morphine biosynthetic genes. In addition, blood leucocytes contain morphine and adrenal glands are hypothesized to produce morphine;both this systems will be subjected to feeding of stable isotope labeled precursors followed by mass spectrometric analysis of the morphine isolated from these cells/tissues to establish whether these are sites of accumulation, transport or synthesis. The cumulative results from these experiments will lay the ground work for future experiments designed to address the question of function of endogenous morphine and its interaction with other endogenous (endorphins, enkephalins) and exogenous (synthetic analgesic) compounds. The Public Health Relevance: Our research group is the first to prove unequivocally that morphine is produced endogenously by the mammalian body (human cells and mouse brain). We wish now to study the site of morphine formation, the pathway by which it is produced and how this production is regulated. In the long term, we will examine the influence of this endogenous morphine on pain and mood in mammals, which may lead to a whole new approach to the understanding of pain abatement, and eventually could enable the development of new drugs and therapies to alleviate pain and to treat emotional disorders.
| Grobe, Nadja; Kutchan, Toni M; Zenk, Meinhart H (2012) Rat CYP2D2, not 2D1, is functionally conserved with human CYP2D6 in endogenous morphine formation. FEBS Lett 586:1749-53 |
| Frölich, Nadine; Dees, Christian; Paetz, Christian et al. (2011) Distinct pharmacological properties of morphine metabolites at G(i)-protein and ?-arrestin signaling pathways activated by the human ?-opioid receptor. Biochem Pharmacol 81:1248-54 |
| Grobe, Nadja; Ren, Xuan; Kutchan, Toni M et al. (2011) An (R)-specific N-methyltransferase involved in human morphine biosynthesis. Arch Biochem Biophys 506:42-7 |
| Han, Xu; Lamshoft, Marc; Grobe, Nadja et al. (2010) The biosynthesis of papaverine proceeds via (S)-reticuline. Phytochemistry 71:1305-12 |
| Grobe, Nadja; Lamshoft, Marc; Orth, Robert G et al. (2010) Urinary excretion of morphine and biosynthetic precursors in mice. Proc Natl Acad Sci U S A 107:8147-52 |
