Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1992 Dec 1;89(23):11234–11238. doi: 10.1073/pnas.89.23.11234

Regulation of an opioid-binding protein in NG108-15 cells parallels regulation of delta-opioid receptors.

C M Lane 1, R Elde 1, H H Loh 1, N M Lee 1
PMCID: PMC50524  PMID: 1333602

Abstract

An opioid-binding protein has recently been purified from bovine brain and cloned, and its cDNA sequence has been obtained. Indirect evidence suggests that this protein has a role in opioid-receptor function. However, because direct testing of its function by expression of its cDNA has not yet been possible and because its structure bears no resemblance to G protein-coupled receptors, the role of this protein in opioid-receptor activity is still in question. An antibody raised to a portion of the predicted amino acid sequence of opioid-binding cell-adhesion molecule (OBCAM) specifically labeled the surface of NG108-15 cells, as visualized by immunofluorescence with confocal microscopy. Furthermore, chronic treatment of these cells with opioid agonist, which down-regulates opioid receptors, reduced OBCAM immunoreactivity (ir). Down-regulation of both opioid receptors and OBCAM-ir was greatest after chronic treatment of NG108-15 cells with delta-opioid agonists, as well as with nonselective agonists such as etorphine, whereas other agonists including [D-Ala2-N-MePhe4-Gly-ol]enkephalin, morphine, levorphanol, dynorphin A-(1-13), and U-50,488H were less effective or ineffective. Chronic treatment of NG108-15 cells with muscarinic agonists had no effect on OBCAM-ir. Furthermore, NG108-15 cells transfected with an antisense construct to OBCAM have a reduced density of opioid-binding sites as well as reduced OBCAM-ir. Taken together, these results strongly suggest that OBCAM has a role in opioid-receptor function in NG108-15 cells.

Full text

PDF

Images in this article

11236Image
on p.11236
11237Image
on p.11237

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ann D. K., Hasegawa J., Ko J. L., Chen S. T., Lee N. M., Loh H. H. Specific reduction of delta-opioid receptor binding in transfected NG108-15 cells. J Biol Chem. 1992 Apr 15;267(11):7921–7926. [PubMed] [Google Scholar]
  2. Brelje T. C., Scharp D. W., Sorenson R. L. Three-dimensional imaging of intact isolated islets of Langerhans with confocal microscopy. Diabetes. 1989 Jun;38(6):808–814. doi: 10.2337/diab.38.6.808. [DOI] [PubMed] [Google Scholar]
  3. Chang K. J., Cuatrecasas P. Multiple opiate receptors. Enkephalins and morphine bind to receptors of different specificity. J Biol Chem. 1979 Apr 25;254(8):2610–2618. [PubMed] [Google Scholar]
  4. Cho T. M., Hasegawa J., Ge B. L., Loh H. H. Purification to apparent homogeneity of a mu-type opioid receptor from rat brain. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4138–4142. doi: 10.1073/pnas.83.12.4138. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dohlman H. G., Caron M. G., Lefkowitz R. J. A family of receptors coupled to guanine nucleotide regulatory proteins. Biochemistry. 1987 May 19;26(10):2657–2664. doi: 10.1021/bi00384a001. [DOI] [PubMed] [Google Scholar]
  6. Gioannini T. L., Howard A. D., Hiller J. M., Simon E. J. Purification of an active opioid-binding protein from bovine striatum. J Biol Chem. 1985 Dec 5;260(28):15117–15121. [PubMed] [Google Scholar]
  7. Hassan A. H., Almeida O. F., Gramsch C., Herz A. Immunocytochemical demonstration of opioid receptors in selected rat brain areas and neuroblastoma x glioma hybrid (NG108-15) cells using a monoclonal anti-idiotypic antibody. Neuroscience. 1989;32(1):269–278. doi: 10.1016/0306-4522(89)90126-7. [DOI] [PubMed] [Google Scholar]
  8. Hassell J., Hand A. R. Tissue fixation with diimidoesters as an alternative to aldehydes. I. Comparison of cross-linking and ultrastructure obtained with dimethylsuberimidate and glutaraldehyde. J Histochem Cytochem. 1974 Apr;22(4):223–229. doi: 10.1177/22.4.223. [DOI] [PubMed] [Google Scholar]
  9. Hazum E., Chang K. J., Cuatrecasas P. Opiate (Enkephalin) receptors of neuroblastoma cells: occurrence in clusters on the cell surface. Science. 1979 Nov 30;206(4422):1077–1079. doi: 10.1126/science.227058. [DOI] [PubMed] [Google Scholar]
  10. Johnson G. D., Davidson R. S., McNamee K. C., Russell G., Goodwin D., Holborow E. J. Fading of immunofluorescence during microscopy: a study of the phenomenon and its remedy. J Immunol Methods. 1982 Dec 17;55(2):231–242. doi: 10.1016/0022-1759(82)90035-7. [DOI] [PubMed] [Google Scholar]
  11. Klein W. L., Nathanson N., Nirenberg M. Muscarinic acetylcholine receptor regulation by accelerated rate of receptor loss. Biochem Biophys Res Commun. 1979 Sep 27;90(2):506–512. doi: 10.1016/0006-291x(79)91264-6. [DOI] [PubMed] [Google Scholar]
  12. Law P. Y., Hom D. S., Loh H. H. Loss of opiate receptor activity in neuroblastoma X glioma NG108-15 hybrid cells after chronic opiate treatment. A multiple-step process. Mol Pharmacol. 1982 Jul;22(1):1–4. [PubMed] [Google Scholar]
  13. Law P. Y., Hom D. S., Loh H. H. Opiate receptor down-regulation and desensitization in neuroblastoma X glioma NG108-15 hybrid cells are two separate cellular adaptation processes. Mol Pharmacol. 1983 Nov;24(3):413–424. [PubMed] [Google Scholar]
  14. Law P. Y., Hom D. S., Loh H. H. Opiate regulation of adenosine 3':5'-cyclic monophosphate level in neuroblastoma X glioma NG108-15 hybrid cells. Relationship between receptor occupancy and effect. Mol Pharmacol. 1983 Jan;23(1):26–35. [PubMed] [Google Scholar]
  15. Loh H. H., Smith A. P. Molecular characterization of opioid receptors. Annu Rev Pharmacol Toxicol. 1990;30:123–147. doi: 10.1146/annurev.pa.30.040190.001011. [DOI] [PubMed] [Google Scholar]
  16. Ornatowska M., Glasel J. A. Two- and three-dimensional distributions of opioid receptors on NG108-15 cells visualized with the aid of fluorescence confocal microscopy and anti-idiotypic antibodies. J Chem Neuroanat. 1992 Mar-Apr;5(2):95–106. doi: 10.1016/0891-0618(92)90037-q. [DOI] [PubMed] [Google Scholar]
  17. Roy S., Zhu Y. X., Lee N. M., Loh H. H. Different molecular weight forms of opioid receptors revealed by polyclonal antibodies. Biochem Biophys Res Commun. 1988 Jan 15;150(1):237–244. doi: 10.1016/0006-291x(88)90511-6. [DOI] [PubMed] [Google Scholar]
  18. Roy S., Zhu Y. X., Loh H. H., Lee N. M. A monoclonal antibody that inhibits opioid binding to rat brain membranes. Biochem Biophys Res Commun. 1988 Jul 29;154(2):688–693. doi: 10.1016/0006-291x(88)90194-5. [DOI] [PubMed] [Google Scholar]
  19. Schofield P. R., McFarland K. C., Hayflick J. S., Wilcox J. N., Cho T. M., Roy S., Lee N. M., Loh H. H., Seeburg P. H. Molecular characterization of a new immunoglobulin superfamily protein with potential roles in opioid binding and cell contact. EMBO J. 1989 Feb;8(2):489–495. doi: 10.1002/j.1460-2075.1989.tb03402.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Simonds W. F., Burke T. R., Jr, Rice K. C., Jacobson A. E., Klee W. A. Purification of the opiate receptor of NG108-15 neuroblastoma-glioma hybrid cells. Proc Natl Acad Sci U S A. 1985 Aug;82(15):4974–4978. doi: 10.1073/pnas.82.15.4974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. White J. G., Amos W. B., Fordham M. An evaluation of confocal versus conventional imaging of biological structures by fluorescence light microscopy. J Cell Biol. 1987 Jul;105(1):41–48. doi: 10.1083/jcb.105.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Xie G. X., Miyajima A., Goldstein A. Expression cloning of cDNA encoding a seven-helix receptor from human placenta with affinity for opioid ligands. Proc Natl Acad Sci U S A. 1992 May 1;89(9):4124–4128. doi: 10.1073/pnas.89.9.4124. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES