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Why Does Insensitivity to Opioid Narcotics Develop?

Maze, Mervyn MB, ChB, FRCP

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Opiate receptors hold a place of prominence for practitioners of anesthesia. Not only are these targets for several analgesic and anesthetic drugs that are used commonly but the primary structure of each of the opiate receptor subtypes was first reported in 1993 by an anesthesiologist, Dr. Kazuhiko Fukuda working in Professor Kenjiro Mori's department in Kyoto, Japan. [1] The isolation and cloning of this family of proteins proved particularly difficult because of its relatively low abundance. However, following the example of Fukuda's pivotal studies, molecular genetic reagents have been developed that have prompted a profusion of cell biology studies that has greatly increased our understanding of opioid action.
Of particular interest to investigators in Dr. Robert Peterfreund's laboratory at the Massachusetts General Hospital are factors that regulate sensitivity to opiate narcotics. Biochemical changes are induced in chronic pain states, which reduce the analgesic efficacy of opioids. In addition, patients develop tolerance to the analgesic properties of opioids after they are administered continuously. In both these settings, a similar cascade of neuroplastic changes are induced, resulting in activation of the NMDA receptor [2] and activation and translocation of protein kinase C. [3]
These investigators have directed their attention to the “downstream” effects of the activation of protein kinase C. [4] In a model cell system derived from humans (SH‐SY5Y cell line) that contains the micro‐opioid receptor and the entire signaling system responsible for neuroplasticity, the authors directly activated protein kinase C with the phorbol ester. Then they determined whether transcription of the micro‐opioid receptor was affected by this treatment by measuring the amount of messenger RNA (mRNA) using a technique called slot blot hybridization. This technique takes advantage of the fact that the mRNA is complementary to the template DNA with which it will “hybridize.” Therefore, the amount of mRNA specific for the micro‐opioid receptor can be determined by reacting the harvested mRNA with a32 P‐labeled cDNA probe (i.e., the template for the micro‐opioid receptor) and determining the amount of radioactivity that “sticks” by autoradiography followed by laser densitometry. Care was to taken to establish that the sup 32 P‐labeled cDNA probe directed against the micro‐opioid receptor did not react nonspecifically with all mRNA. In addition, they confirmed that the same amount of mRNA was used by normalizing their data with the “housekeeping gene” beta actin, whose content does not change under these conditions.
The authors report that the amount of micro‐opioid receptor mRNA present is significantly reduced when protein kinase C activity is stimulated with phorbol esters. Further, they demonstrated that the decrement in transcription of the micro‐opioid receptor did not depend on new protein synthesis, which effectively precludes a genomically mediated transcription factor in the pathogenesis of this event. Rather, the decrement in transcription is likely to be produced by posttranslational modification by phosphorylation of certain factors(s).
The importance of this finding is that the decrease in micro‐opioid receptor mRNA is expected to cause a concomitant decrease in micro‐opioid receptor protein expression or receptor downregulation. In turn, this may cause a decrease in sensitivity to opioids and thereby a loss in the analgesic properties of this class of compound.
Mervyn Maze, M.B., Ch.B., F.R.C.P.
Professor and Director of Research; Department of Anesthesia; Stanford University; Staff Physician, Anesthesiology Service, VAPAHCS; Palo Alto, California 943042
maze@leland.stanford.edu
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References

1. Fukuda K, Kato S, Mori K, Nishi M, Takeshima H: Primary structures and expression from cDNAs of rat opioid receptor delta- and mu-subtypes. FEBS Letters 1993; 327:311-4.

2. Zhang L, Rzigalinski BA, Ellis EF, Satin LS: Reduction of voltage-dependent Mg2+ blockade of NMDA current in mechanically injured neurons. Science 1996; 274:1921-3.

3. Lin Q, Peng YB, Willis WD: Possible role of protein kinase C in the sensitization of primate spinothalamic tract neurons. J Neurosci 1996; 16:3026-34.

4. Gies EK, Peters DM, Gelb CR, Knag KM, Peterfreund RA: Regulation of mu opioid receptor mRNA levels by activation of protein kinase C in human SH-SY5Y neuroblastoma cells. Anesthesiology 1997; 87:1127-38.

Keywords:
mRNA; micro‐opioid receptor; Phorbol esters; Protein kinase C; SH‐SY5Y neuroblastoma cells.

© 1997 American Society of Anesthesiologists, Inc.

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