Complex Effects of Antisense Oligonucleotides Complementary to the Mouse g₂ Subunit of the GABA_A Receptor

R. Dayne Mayfield*

T.K. Booker*

Sherry S. Leonard* ** ***

Margaret J. Velardo* **

Nancy R. Zahniser* **

*Department of Pharmacology and **Neuroscience Program, University of Colorado Health Sciences Center, 4200 East Ninth Avenue, Denver, CO 80262

***Department of Veterans Affairs Medical Center, Denver, CO 80220

Send correspondence to:
Dr. R. Dayne Mayfield
Department of Pharmacology (C236)
University of Colorado Health Sciences Center
4200 East Ninth Avenue
Denver, Colorado 80262 U.S.A.
E-mail: Dayne Mayfield

Keywords: Antisense oligonucleotide, Phosphorothioate oligonucleotide, Long Sleep (LS) mouse, g-Aminobutyric acid (GABA)_A receptor, benzodiazepine receptor, Radioligand binding, Osmotic Pump

ABSTRACT
(Neuroscience-Net, Volume 2, Article #10012; March 7, 1997)

Benzodiazepines allosterically modulate the inhibitory effects mediated by the GABA_A receptor, and the g₂ subunit of the receptor is necessary for this action. Since no antagonists selective for the g₂ subunit are available, our goal was to determine whether expression of this subunit could be decreased in vivo using antisense phosphorothioate oligonucleotides (S-ODNs) to prevent its expression. S-ODNs were administered into the lateral ventricle of Long Sleep mice for 3.5 days via miniosmotic pump. Saturation binding parameters were determined using [³H]-flunitrazepam ([³H]-FNZ) and washed membrane preparations. In initial experiments, Bmax values were decreased significantly in cortex, striatum, and cerebellum by antisense S-ODN infusion compared to phosphate buffered saline controls. However, these effects were not consistent in subsequent experiments and Bmax values in cortex were also decreased by random-base S-ODN treatment. Approximately 30% of the animals infused with S-ODNs with 100% S-ODN linkages displayed overt signs of toxicity. Modified S-ODNs produced no apparent toxicity, but had no significant effect on [³H]FNZ binding parameters. These results demonstrate that antisense S-ODN treatment can decrease [³H]FNZ binding in vivo, but also suggest that toxic effects of the S-ODN derivatives may interfere with the interpretation of the data.