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. 1994 Oct;14(5):507–521. doi: 10.1007/BF02088834

The application of antisense oligonucleotide technology to the brain: Some pitfalls

B J Chiasson 1, J N Armstrong 1,2, M L Hooper 1, P R Murphy 3, H A Robertson 1,✉
PMCID: PMC11566955  PMID: 7621510

Abstract

1. Amphetamine-induced c-fos andegr-1 expression in the striatum was used as a model in which to study the effects of antisense oligodeoxynucleotides (ODNs) directed at c-fos. Using direct infusions of ODNs into the striata of animals we have demonstrated that c-fos antisense ODNs retain most of their biological activity with 2- or 3-base substitutions. The c-fos antisense and mismatch ODNs attenuated Fos immunoreactivity but had little effect on Egr-1 immunoreactivity.

2. In another group of studies examining the role of c-fos in amygdala kindling, we have demonstrated that ODNs cause neurotoxic damage following repeated daily infusions into the amygdala. The damage observed was greatly diminished when the time interval between infusions was extended.

Key words: c-fos, egr-1, antisense oligodeoxynucleotides (ODNs), amygdala kindling, immediate-early genes, ODN-induced toxicity

References

  1. Altschul, S. F., Gish, W., Miller, W., Myers, E. W., and Lipman, D. J. (1990). Basic local alignment search tool.J. Mol. Biol.215403–410. [DOI] [PubMed] [Google Scholar]
  2. Campbell, J. M., Bacon, T. A., and Wickstrom, E. (1990). Oligodeoxynucleoside phosphorothioate stability in subcellular extracts, culture media, sera and cerebrospinal fluid.J. Biochem. Biophys. Meth.20259–267. [DOI] [PubMed] [Google Scholar]
  3. Chiang, M.-Y., Chan, H., Zounes, M. A., Freier, S. M., Lima, W. F., and Bennett, C. F. (1991). Antisense oligonucleotides inhibit intercellular adhesion molecule 1 expression by two distinct mechanisms.J. Biol. Chem.26618162–18171. [PubMed] [Google Scholar]
  4. Chiasson, B. J., and Robertson, H. A. (1994). Toxicity following in vivo infusion of antisense oligonucleotide: Role of the inter-infusion interval.Soc. Neurosci. Abstr. (in press).
  5. Chiasson, B. J., Hooper, M. L., and Robertson, H. A. (1992a). Amphetamine induced rotational behavior in non-lesioned rats: A role for c-fos expression in the striatum.Soc. Neurosci. Abstr.18562. [Google Scholar]
  6. Chiasson, B. J., Hooper, M. L., Murphy, P. R., and Robertson, H. A. (1992b). Antisense oligonucleotide eliminatesin vivo expression of c-fos in mammalian brain.Eur. J. Mol. Pharmacol.227451–453. [DOI] [PubMed] [Google Scholar]
  7. Chiasson, B. J., Dennison, Z., and Robertson, H. A. (1995). Amygdala kindling and immediate-early genesMol. Brain. Res. (in press). [DOI] [PubMed]
  8. Cooney, M., Czernuszewick, G., Postel, E. H., Plint, S. J., and Hogan, M. E. (1988). Site-specific oligonucleotide binding represses transcription of the human c-myc gene in vitro.Science241456–459. [DOI] [PubMed] [Google Scholar]
  9. Dragunow, M., and Robertson, H. A. (1987). Kindling stimulation induces c-fos protein(s) in granule cells of the rat dentate gyrus.Nature329441–442. [DOI] [PubMed] [Google Scholar]
  10. Dragunow, M., Robertson, H. A., and Robertson, G. S. (1988). Effects of kindled seizures on the induction of c-fos protein(s) in mammalian neurons.Exp. Neurol.102261–263.3141205 [Google Scholar]
  11. Dragunow, M., Lawlor, P., Chiasson, B. J., and Robertson, H. A. (1993). C-fos antisense generates apomorphine and amphetamine induced rotation.Neuroreport5305–306. [DOI] [PubMed] [Google Scholar]
  12. Gao, W.-Y., Hand, F.-S., Storm, C., Egan, W., and Cheng, Y.-C. (1991). Phosphorothioate oligonucleotides are inhibitors of human DNA polymerases and RNase H: Implications for antisense technology.Mol. Pharmacol.41223–229. [PubMed] [Google Scholar]
  13. Goddard, G. V., McIntyre, D., and Leech, C. (1969). A permanent change in brain function resulting from daily electrical stimulation.Exp. Neurol.25295–330. [DOI] [PubMed] [Google Scholar]
  14. Hooper, M. L., Chiasson, B. J., and Robertson, H. A. (1994). Infusion into the brain of an antisense oligonucleotide to the immediate-early gene c-fos suppresses production of Fos and produces a beharioral effect.Neurosci.63917–924. [DOI] [PubMed] [Google Scholar]
  15. Koelle, C. G., and Friedenwald, J. S. (1949). A histochemical method for localizing cholinesterase activity.Proc. Soc. Exp. Biol. Med.70617–622. [DOI] [PubMed] [Google Scholar]
  16. Kreig, A. M. (1993). Uptake and efficacy of phosphodiester and modified antisense oligonucleotides in primary cell cultures.Clin. Chem.39 710–712. [Google Scholar]
  17. Krieg, A. M., Tonkinson, J., Matson, S., Zhao, Q., Saxon, M., Zhang, L.-M., Bhanja, U., Yakubov, L., and Stein, C. A. (1993). Modification of antisense phosphodiester oligodeoxynucleotides by a 5′ cholesteryl moiety increases cellular association and improves efficacy.Proc. Natl. Acad. Sci. USA901048–1052. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Lewis, P. R. (1961). The effect of varying the conditions in the Koelle method.Biblthca. Anat. Vol. 2, Karger, Basel, pp. 11–20. [Google Scholar]
  19. Molino, A., and McIntyre, D. C. (1972). Another inexpensive headplug for the electrical recording and/or stimulation of rats.Physiol. Behav.9273–275. [DOI] [PubMed] [Google Scholar]
  20. Morgan, J. I., and Curran, T. (1991). Stimulus-transcription coupling in the nervous system: Involvement of the inducible proto-oncogenesfos andjun.Annu. Rev. Neurosci.14421–451. [DOI] [PubMed] [Google Scholar]
  21. Nadler, J. V., and Evenson, D. A. (1983). Use of excitatory amino acids to make axon-sparing lesions of hypothalamus.Methods Enzymol.103393–400. [DOI] [PubMed] [Google Scholar]
  22. Perez, J. R., Li, Y., Stein, C. A., Majumder, S., van Oorschot, A., and Narayanan, R. (1994). Sequence-independent induction of Sp 1 transcription factor activity by phosphorothioate oligonucleotides.Proc. Natl. Acad. Sci. USA915959–5961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Robertson, G. S., and Robertson, H. A. (1989). Evidence that L-dopa-induced rotational behavior is dependent on both striatal and nigral mechanisms.J. Neurosci.93326–3331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Robertson, H. A. (1992). Immediate-early genes, neuronal plasticity, and memory.Biochem. Cell Biol.70729–737. [DOI] [PubMed] [Google Scholar]
  25. Robertson, H. A., Peterson, M. R., Murphy, K., and Robertson, G. S. (1989). D1-dopamine receptor agonists selectively activate striatal c-fos independent of rotational behaviour.Brain Res.503346–349. [DOI] [PubMed] [Google Scholar]
  26. Standifer, K. M., Chien, G.-C., Wahlestedt, C., Brown, G. P., and Pasternak, G. W. (1994). Selective loss of delta opioid analgesia and binding by antisense oligodeoxynucleotides to a delta opioid receptor.Neuron12805–810. [DOI] [PubMed] [Google Scholar]
  27. Stein, C. A., and Cheng, Y.-C. (1993). Antisense oligonucleotides as therapeutic agents—Is the bullet really magical?Science261 1004–1012. [DOI] [PubMed] [Google Scholar]
  28. Stein, C. A., and Cohen, J. S. (1988). Oligonucleotides as inhibitors of gene expression: A review.Cancer Res.482659–2668. [PubMed] [Google Scholar]
  29. Stein, C. A., Subasinghe, C., Shinozuka, K., and Cohen, J. S. (1988). Physicochemical properties of phosphorothioate oligodeoxynucleotides.Nucl. Acids Res.163209–3221. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Stull, R. A., Taylor, L. A., and Szoka, J. C., Jr. (1992). Predicting antisense oligonucleotide inhibitory efficacy: A computational approach using histograms and thermodynamic indices.Nucl. Acids. Res.203501–3508. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Ungerstedt, U. (1971). Postsynaptic supersensitivity after 6-hydroxydopamine induced degeneration of the nigro-striatal dopamine system.Acta. Physiol Scand. (Suppl.)36769–93. [DOI] [PubMed] [Google Scholar]
  32. Wahlestedt, C. (1994). Antisense oligonucleotide strategies in neuropharmacology.Trends Pharmacol. Sci.1542–46. [DOI] [PubMed] [Google Scholar]
  33. Wahlestedt, C., Golanov, E., Yamamoto, S., Yee, F., Ericson, H., Yoo, H., Inturrisi, C. E., and Reis, D. J. (1993a). Antisense oligonucleotides to NMDA-R1 receptor channel protect cortical neurons from excitotoxicity and reduce focal ischaemic infarctions.Nature363260–263. [DOI] [PubMed] [Google Scholar]
  34. Wahlestedt, C., Pich, E. M., Koob, G. F., Yee, F., and Heilig, M. (1993b). Modulation of anxiety and neuropeptide Y-Y1 receptors by antisense oligodeoxynucleotides.Science259528–531. [DOI] [PubMed] [Google Scholar]
  35. Woolf, T. M., Jennings, G. B., Rebagliati, M., and Melton, D. A. (1990). The stability, toxicity and effectiveness of unmodified and phosphorothioate antisense oligodeoxynucleotides in Xenopus oocytes and embryos.Nucl. Acids Res.181763–1769. [DOI] [PMC free article] [PubMed] [Google Scholar]

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