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. 1991 Mar 1;88(5):1854–1858. doi: 10.1073/pnas.88.5.1854

Expressional potency of mRNAs encoding receptors and voltage-activated channels in the postmortem rat brain.

D S Ragsdale 1, R Miledi 1
PMCID: PMC51124  PMID: 1705710

Abstract

The stability and integrity of mRNAs encoding neurotransmitter receptors and voltage-activated channels in the postmortem rat brain was investigated by isolating poly(A)+ mRNA, injecting it into Xenopus oocytes, and then examining the expression of functional neurotransmitter receptors and voltage-activated channels in the oocyte membrane by electrophysiological recording. This approach was also used to assess the stability of mRNAs in brains that were incubated in oxygenated mammalian Ringer's solution for various lengths of time and from brains that were freshly frozen and then thawed at room temperature. Oocytes injected with mRNA from up to 21-hr postmortem brains gave large agonist- and voltage-activated responses, indicating that mRNAs encoding neurotransmitter receptors and voltage-activated channels are relatively stable in postmortem brain tissue. In contrast, oocytes injected with mRNA from brains incubated in Ringer's solution exhibited smaller responses, and oocytes injected with mRNA from tissue that was frozen and then thawed displayed very small or undetectable responses. Northern blot analysis using a nucleic acid probe for rat brain Na(+)-channel mRNA indicated that the size of the Na+ currents in injected oocytes reflected the levels of mRNA for Na+ channels in the different mRNA preparations. Thus, the expressional potency of mRNAs encoding neurotransmitter receptors and voltage-activated channels is quite stable in postmortem brains in situ, but it is reduced if the brains are kept in oxygenated saline, and freezing and thawing of tissue results in rapid degeneration of mRNA.

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Selected References

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

  1. Akagi H., Miledi R. Heterogeneity of glycine receptors and their messenger RNAs in rat brain and spinal cord. Science. 1988 Oct 14;242(4876):270–273. doi: 10.1126/science.2845580. [DOI] [PubMed] [Google Scholar]
  2. Akagi H., Patton D. E., Miledi R. Discrimination of heterogenous mRNAs encoding strychnine-sensitive glycine receptors in Xenopus oocytes by antisense oligonucleotides. Proc Natl Acad Sci U S A. 1989 Oct;86(20):8103–8107. doi: 10.1073/pnas.86.20.8103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Alberghina M., Giuffrida Stella A. M. Age-related changes of ribonuclease activities in various regions of the rat central nervous system. J Neurochem. 1988 Jul;51(1):21–24. doi: 10.1111/j.1471-4159.1988.tb04829.x. [DOI] [PubMed] [Google Scholar]
  4. Barish M. E. A transient calcium-dependent chloride current in the immature Xenopus oocyte. J Physiol. 1983 Sep;342:309–325. doi: 10.1113/jphysiol.1983.sp014852. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Carpenter M. K., Parker I., Miledi R. Expression of GABA and glycine receptors by messenger RNAs from the developing rat cerebral cortex. Proc R Soc Lond B Biol Sci. 1988 Jul 22;234(1275):159–170. doi: 10.1098/rspb.1988.0042. [DOI] [PubMed] [Google Scholar]
  6. Franzoni L., García Argiz C. A. Rat brain ribonucleases. Acta Physiol Lat Am. 1978;28(4-5):185–192. [PubMed] [Google Scholar]
  7. Gilbert J. M., Brown B. A., Strocchi P., Bird E. D., Marotta C. A. The preparation of biologically active messenger RNA from human postmortem brain tissue. J Neurochem. 1981 Mar;36(3):976–984. doi: 10.1111/j.1471-4159.1981.tb01689.x. [DOI] [PubMed] [Google Scholar]
  8. Gundersen C. B., Miledi R., Parker I. Glutamate and kainate receptors induced by rat brain messenger RNA in Xenopus oocytes. Proc R Soc Lond B Biol Sci. 1984 Apr 24;221(1223):127–143. doi: 10.1098/rspb.1984.0027. [DOI] [PubMed] [Google Scholar]
  9. Gundersen C. B., Miledi R., Parker I. Messenger RNA from human brain induces drug- and voltage-operated channels in Xenopus oocytes. 1984 Mar 29-Apr 4Nature. 308(5958):421–424. doi: 10.1038/308421a0. [DOI] [PubMed] [Google Scholar]
  10. Gundersen C. B., Miledi R., Parker I. Serotonin receptors induced by exogenous messenger RNA in Xenopus oocytes. Proc R Soc Lond B Biol Sci. 1983 Aug 22;219(1214):103–109. doi: 10.1098/rspb.1983.0062. [DOI] [PubMed] [Google Scholar]
  11. Gundersen C. B., Miledi R., Parker I. Voltage-operated channels induced by foreign messenger RNA in Xenopus oocytes. Proc R Soc Lond B Biol Sci. 1983 Nov 22;220(1218):131–140. doi: 10.1098/rspb.1983.0092. [DOI] [PubMed] [Google Scholar]
  12. Houamed K. M., Bilbe G., Smart T. G., Constanti A., Brown D. A., Barnard E. A., Richards B. M. Expression of functional GABA, glycine and glutamate receptors in Xenopus oocytes injected with rat brain mRNA. 1984 Jul 26-Aug 1Nature. 310(5975):318–321. doi: 10.1038/310318a0. [DOI] [PubMed] [Google Scholar]
  13. Johnson S. A., Morgan D. G., Finch C. E. Extensive postmortem stability of RNA from rat and human brain. J Neurosci Res. 1986;16(1):267–280. doi: 10.1002/jnr.490160123. [DOI] [PubMed] [Google Scholar]
  14. Julius D., MacDermott A. B., Axel R., Jessell T. M. Molecular characterization of a functional cDNA encoding the serotonin 1c receptor. Science. 1988 Jul 29;241(4865):558–564. doi: 10.1126/science.3399891. [DOI] [PubMed] [Google Scholar]
  15. Kusano K., Miledi R., Stinnakre J. Cholinergic and catecholaminergic receptors in the Xenopus oocyte membrane. J Physiol. 1982 Jul;328:143–170. doi: 10.1113/jphysiol.1982.sp014257. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. McGann L. E., Yang H. Y., Walterson M. Manifestations of cell damage after freezing and thawing. Cryobiology. 1988 Jun;25(3):178–185. doi: 10.1016/0011-2240(88)90024-7. [DOI] [PubMed] [Google Scholar]
  17. Miledi R. A calcium-dependent transient outward current in Xenopus laevis oocytes. Proc R Soc Lond B Biol Sci. 1982 Jul 22;215(1201):491–497. doi: 10.1098/rspb.1982.0056. [DOI] [PubMed] [Google Scholar]
  18. Miledi R., Parker I. Chloride current induced by injection of calcium into Xenopus oocytes. J Physiol. 1984 Dec;357:173–183. doi: 10.1113/jphysiol.1984.sp015495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Morrison M. R., Griffin W. S. The isolation and in vitro translation of undegraded messenger RNAs from human postmortem brain. Anal Biochem. 1981 May 15;113(2):318–324. doi: 10.1016/0003-2697(81)90083-x. [DOI] [PubMed] [Google Scholar]
  20. Noda M., Ikeda T., Kayano T., Suzuki H., Takeshima H., Kurasaki M., Takahashi H., Numa S. Existence of distinct sodium channel messenger RNAs in rat brain. Nature. 1986 Mar 13;320(6058):188–192. doi: 10.1038/320188a0. [DOI] [PubMed] [Google Scholar]
  21. Parker I., Sumikawa K., Miledi R. Neurotensin and substance P receptors expressed in Xenopus oocytes by messenger RNA from rat brain. Proc R Soc Lond B Biol Sci. 1986 Nov 22;229(1255):151–159. doi: 10.1098/rspb.1986.0079. [DOI] [PubMed] [Google Scholar]
  22. Parker I., Sumikawa K., Miledi R. Neurotensin and substance P receptors expressed in Xenopus oocytes by messenger RNA from rat brain. Proc R Soc Lond B Biol Sci. 1986 Nov 22;229(1255):151–159. doi: 10.1098/rspb.1986.0079. [DOI] [PubMed] [Google Scholar]
  23. Reichert G. H., Issinger O. G. In vitro study of the biological activity of RNAs after incubation of hog liver, heart and brain tissue at room temperature. Biochimie. 1985 Jun;67(6):657–661. doi: 10.1016/s0300-9084(85)80208-x. [DOI] [PubMed] [Google Scholar]
  24. Rowe A. W., Lenny L. L. Cryopreservation of granulocytes for transfusion: studies on human granulocyte isolation, the effect of glycerol on lysosomes, kinetics of glycerol uptake and cryopreservation with dimethyl sulfoxide and glycerol. Cryobiology. 1980 Jun;17(3):198–212. doi: 10.1016/0011-2240(80)90027-9. [DOI] [PubMed] [Google Scholar]
  25. Sajdel-Sulkowska E. M., Marotta C. A. Alzheimer's disease brain: alterations in RNA levels and in a ribonuclease-inhibitor complex. Science. 1984 Aug 31;225(4665):947–949. doi: 10.1126/science.6206567. [DOI] [PubMed] [Google Scholar]
  26. Sajdel-Sulkowska E., Coughlin J. F., Marotta C. A. In vitro synthesis of polypeptides of moderately large size by poly(A)-containing messenger RNA from postmortem human brain and mouse brain. J Neurochem. 1983 Mar;40(3):670–680. doi: 10.1111/j.1471-4159.1983.tb08032.x. [DOI] [PubMed] [Google Scholar]
  27. Schulz-Harder B., Graf von Keyserlingk D. Comparison of brain ribonucleases of rabbit, guinea pig, rat, mouse and gerbil. Histochemistry. 1988;88(3-6):587–594. doi: 10.1007/BF00570329. [DOI] [PubMed] [Google Scholar]
  28. Sigel E. Properties of single sodium channels translated by Xenopus oocytes after injection with messenger ribonucleic acid. J Physiol. 1987 May;386:73–90. doi: 10.1113/jphysiol.1987.sp016523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sumikawa K., Parker I., Miledi R. Messenger RNA from rat brain induces noradrenaline and dopamine receptors in Xenopus oocytes. Proc R Soc Lond B Biol Sci. 1984 Dec 22;223(1231):255–260. doi: 10.1098/rspb.1984.0093. [DOI] [PubMed] [Google Scholar]
  30. Sumikawa K., Parker I., Miledi R. Partial purification and functional expression of brain mRNAs coding for neurotransmitter receptors and voltage-operated channels. Proc Natl Acad Sci U S A. 1984 Dec;81(24):7994–7998. doi: 10.1073/pnas.81.24.7994. [DOI] [PMC free article] [PubMed] [Google Scholar]

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