Diazepam protects against rat hippocampal neuronal cell death induced by antisense oligodeoxynucleotide to GABA(A) receptor gamma2 subunit.
Antisense oligodeoxynucleotides (ODNs) are used for the selective inhibition of gene expression. Antisense ODNs are promising tools for the investigation of physiological implications of proteins in the central nervous system of rodents in vivo. We have previously demonstrated that a phosphorothioate antisense ODN to the GABA(A) receptor gamma2 subunit, but not sense or mismatch control ODNs, induces a decrease in ex vivo benzodiazepine receptor radioligand binding in rat hippocampus when infused into the hippocampus in vivo [Karle et al., Neurosci. Lett., 202 (1995) 97-100]. This effect is parallelled by a decrease in the number of GABA(A) receptors and an extensive loss of hippocampal neurones. There is increasing awareness of risks of toxic 'non-antisense' effects induced by ODNs, and in particular phosphorothioate ODNs. The present experiments were designed to investigate the specificity of effects induced by the gamma2 subunit antisense ODN. The temporal development of changes in [3H]flunitrazepam and [3H]quinuclidinyl benzilate binding as well as in tissue protein levels supports the notion that the antisense ODN primarily acts by blocking the expression of the targeted receptor subunit protein. Furthermore, it is shown that a threshold for the elicitation of neurodegenerative changes exists. Finally, it is demonstrated that diazepam treatment of rats protects against the development of neuronal cell death induced by the antisense ODN. Collectively, the results support the hypothesis that the neurodegeneration induced by the antisense ODN is a consequence of diminished GABAergic inhibitory tonus following a selective down-regulation of gamma2 subunit-containing GABA(A) receptor complexes.
Karle
J
J
Research Institute of Biological Psychiatry, St. Hans Hospital, Roskilde, Denmark. resshh@inet.uni-c.dk
Witt
M R
MR
Nielsen
M
M
D000818:Animals; D016923:Cell Death / Q000187:drug effects; D003975:Diazepam / Q000494:pharmacology*; D005445:Flunitrazepam / Q000494:pharmacology; D018757:GABA Modulators / Q000494:pharmacology*; D006624:Hippocampus / Q000166:cytology* / Q000187:drug effects; D008297:Male; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009410:Nerve Degeneration; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D011813:Quinuclidinyl Benzilate / Q000494:pharmacology; D011869:Radioligand Assay; D051381:Rats; D017208:Rats, Wistar; D011963:Receptors, GABA-A / Q000235:genetics*; D011976:Receptors, Muscarinic / Q000378:metabolism; D013997:Time Factors
D018757:GABA Modulators; D018727:Muscarinic Antagonists; D016376:Oligonucleotides, Antisense; D011963:Receptors, GABA-A; D011976:Receptors, Muscarinic; D005445:Flunitrazepam; D011813:Quinuclidinyl Benzilate; D003975:Diazepam