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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
. 1982 Nov;79(21):6742–6746. doi: 10.1073/pnas.79.21.6742

Localization of immunoreactive dynorphin in neurons cultured from spinal cord and dorsal root ganglia.

P M Sweetnam, J H Neale, J L Barker, A Goldstein
PMCID: PMC347205  PMID: 6128736

Abstract

Antisera specific for dynorphin were used to study the cellular distribution of opioid peptides in spinal cord and dorsal root ganglion neurons in dissociated cell culture. Radioimmunoassay of 4-wk-old cultures yielded levels of dynorphin immunoreactivity similar to those in adult rodent spinal cord. Immunohistochemistry showed staining confined to the perinuclear region of neuronal cell bodies. In contrast, enkephalin immunoreactivity was found in extensive neurite fields as well as in neuronal perikarya. Opioid peptide immunoreactivity was observed in approximately equal to 5% of the spinal cord neurons either with dynorphin or enkephalin antiserum. No substantial increase in the number of reactive cells was observed when the two sera were applied simultaneously. These results suggest that the perinuclear region of opioid spinal cord neurons in culture contains peptide with an amino acid sequence similar to that of the midportion of dynorphin, whereas the neurites appear to contain smaller peptides related to NH2-terminal fragments of dynorphin. By using simple morphological criteria, spinal sensory neurons can be identified in these cell cultures and in cultures prepared from dorsal root ganglia without spinal cord. Approximately 1-2% of these ganglion cells showed intense immunostaining with an affinity-purified dynorphin antiserum. An additional few percent of the sensory neurons showed less intense opioid immunoreactivity. This result extends the observations of opioid peptides one step further along the pathway that processes sensory information.

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Selected References

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

  1. Barker J. L., Neale J. H., Smith T. G., Jr, Macdonald R. L. Opiate peptide modulation of amino acid responses suggests novel form of neuronal communication. Science. 1978 Mar 31;199(4336):1451–1453. doi: 10.1126/science.204016. [DOI] [PubMed] [Google Scholar]
  2. Barker J. L., Smith T. G., Jr, Neale J. H. Multiple membrane actions of enkephalin revealed using cultured spinal neurons. Brain Res. 1978 Oct 6;154(1):153–158. doi: 10.1016/0006-8993(78)91063-6. [DOI] [PubMed] [Google Scholar]
  3. Botticelli L. J., Cox B. M., Goldstein A. Immunoreactive dynorphin in mammalian spinal cord and dorsal root ganglia. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7783–7786. doi: 10.1073/pnas.78.12.7783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Fischli W., Goldstein A., Hunkapiller M. W., Hood L. E. Isolation and amino acid sequence analysis of a 4,000-dalton dynorphin from porcine pituitary. Proc Natl Acad Sci U S A. 1982 Sep;79(17):5435–5437. doi: 10.1073/pnas.79.17.5435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ghazarossian V. E., Chavkin C., Goldstein A. A specific radioimmunoassay for the novel opioid peptide dynorphin. Life Sci. 1980 Jul 7;27(1):75–86. doi: 10.1016/0024-3205(80)90021-1. [DOI] [PubMed] [Google Scholar]
  6. Goldstein A., Ghazarossian V. E. Immunoreactive dynorphin in pituitary and brain. Proc Natl Acad Sci U S A. 1980 Oct;77(10):6207–6210. doi: 10.1073/pnas.77.10.6207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hökfelt T., Ljungdahl A., Terenius L., Elde R., Nilsson G. Immunohistochemical analysis of peptide pathways possibly related to pain and analgesia: enkephalin and substance P. Proc Natl Acad Sci U S A. 1977 Jul;74(7):3081–3085. doi: 10.1073/pnas.74.7.3081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Jessell T. M., Iversen L. L. Opiate analgesics inhibit substance P release from rat trigeminal nucleus. Nature. 1977 Aug 11;268(5620):549–551. doi: 10.1038/268549a0. [DOI] [PubMed] [Google Scholar]
  9. Kakidani H., Furutani Y., Takahashi H., Noda M., Morimoto Y., Hirose T., Asai M., Inayama S., Nakanishi S., Numa S. Cloning and sequence analysis of cDNA for porcine beta-neo-endorphin/dynorphin precursor. Nature. 1982 Jul 15;298(5871):245–249. doi: 10.1038/298245a0. [DOI] [PubMed] [Google Scholar]
  10. Kangawa K., Matsuo H. alpha-Neo-endorphin : a "big" Leu-enkephalin with potent opiate activity from porcine hypothalami. Biochem Biophys Res Commun. 1979 Jan 15;86(1):153–160. doi: 10.1016/0006-291x(79)90394-2. [DOI] [PubMed] [Google Scholar]
  11. Lamotte C., Pert C. B., Snyder S. H. Opiate receptor binding in primate spinal cord: distribution and changes after dorsal root section. Brain Res. 1976 Aug 13;112(2):407–412. doi: 10.1016/0006-8993(76)90296-1. [DOI] [PubMed] [Google Scholar]
  12. Macdonald R. L., Nelson P. G. Specific-opiate-induced depression of transmitter release from dorsal root ganglion cells in culture. Science. 1978 Mar 31;199(4336):1449–1451. doi: 10.1126/science.204015. [DOI] [PubMed] [Google Scholar]
  13. Mudge A. W., Leeman S. E., Fischbach G. D. Enkephalin inhibits release of substance P from sensory neurons in culture and decreases action potential duration. Proc Natl Acad Sci U S A. 1979 Jan;76(1):526–530. doi: 10.1073/pnas.76.1.526. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Neale J. H., Barker J. L., Uhl G. R., Snyder S. H. Enkephalin-containing neurons visualized in spinal cord cell cultures. Science. 1978 Aug 4;201(4354):467–469. doi: 10.1126/science.351811. [DOI] [PubMed] [Google Scholar]
  15. Ransom B. R., Neale E., Henkart M., Bullock P. N., Nelson P. G. Mouse spinal cord in cell culture. I. Morphology and intrinsic neuronal electrophysiologic properties. J Neurophysiol. 1977 Sep;40(5):1132–1150. doi: 10.1152/jn.1977.40.5.1132. [DOI] [PubMed] [Google Scholar]
  16. Seizinger B. R., Höllt V., Herz A. Evidence for the occurrence of the opioid octapeptide dynorphin-(1-8) in the neurointermediate pituitary of rats. Biochem Biophys Res Commun. 1981 Sep 16;102(1):197–205. doi: 10.1016/0006-291x(81)91507-2. [DOI] [PubMed] [Google Scholar]
  17. Simantov R., Kuhar M. J., Uhl G. R., Snyder S. H. Opioid peptide enkephalin: immunohistochemical mapping in rat central nervous system. Proc Natl Acad Sci U S A. 1977 May;74(5):2167–2171. doi: 10.1073/pnas.74.5.2167. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Watson S. J., Akil H., Fischli W., Goldstein A., Zimmerman E., Nilaver G., van wimersma Griedanus T. B. Dynorphin and vasopressin: common localization in magnocellular neurons. Science. 1982 Apr 2;216(4541):85–87. doi: 10.1126/science.6121376. [DOI] [PubMed] [Google Scholar]
  19. Watson S. J., Akil H., Ghazarossian V. E., Goldstein A. Dynorphin immunocytochemical localization in brain and peripheral nervous system: preliminary studies. Proc Natl Acad Sci U S A. 1981 Feb;78(2):1260–1263. doi: 10.1073/pnas.78.2.1260. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Weber E., Evans C. J., Barchas J. D. Predominance of the amino-terminal octapeptide fragment of dynorphin in rat brain regions. Nature. 1982 Sep 2;299(5878):77–79. doi: 10.1038/299077a0. [DOI] [PubMed] [Google Scholar]
  21. Weber E., Roth K. A., Barchas J. D. Immunohistochemical distribution of alpha-neo-endorphin/dynorphin neuronal systems in rat brain: evidence for colocalization. Proc Natl Acad Sci U S A. 1982 May;79(9):3062–3066. doi: 10.1073/pnas.79.9.3062. [DOI] [PMC free article] [PubMed] [Google Scholar]

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