Expressional potency of mRNAs encoding receptors and voltage-activated channels in the postmortem rat brain.
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.
Ragsdale
D S
DS
Department of Psychobiology, University of California, Irvine 92717.
Miledi
R
R
D000818:Animals; D015152:Blotting, Northern; D001921:Brain / Q000378:metabolism*; D005260:Female; D005615:Freezing; D005998:Glycine / Q000494:pharmacology; D007473:Ion Channels / Q000235:genetics* / Q000502:physiology; D007608:Kainic Acid / Q000494:pharmacology; D008297:Male; D018377:Neurotransmitter Agents / Q000494:pharmacology*; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D011061:Poly A / Q000235:genetics* / Q000302:isolation & purification; D011180:Postmortem Changes; D012313:RNA / Q000235:genetics* / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000502:physiology; D012701:Serotonin / Q000494:pharmacology; D014982:Xenopus laevis; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D007473:Ion Channels; D018377:Neurotransmitter Agents; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D011061:Poly A; D012701:Serotonin; D005680:gamma-Aminobutyric Acid; D012313:RNA; D007608:Kainic Acid; D005998:Glycine