Regulation of GABAA receptor subunit mRNA expression by the pesticide dieldrin in embryonic brainstem cultures: A quantitative, competitive reverse transcription‐polymerase chain reaction study

Abstract

Cyclodiene organochlorine pesticides, such as dieldrin, inhibit γ-aminobutyric acid (GABA)ergic neurotransmission by blocking the Cl− channel of GABAA receptors. This action may make the developing nervous system especially vulnerable to these neurotoxins, which could interfere with the trophic actions of GABA on developing neurons and alter expression of GABAA receptors. We have used an in vitro model to determine whether exposure to dieldrin alters developmental expression of GABAA receptor subunit mRNA transcripts. Dissociated cell cultures were prepared from embryonic day 14 (E14) brainstem and cultured in serum-containing medium for 1 day in vitro (DIV), then treated for 2 DIV with 10 μM dieldrin in serum-free medium. This dose was based on preliminary experiments and previous studies (Nagata et al.: Brain Res 645:19–26, 1994; Pomes et al.: J Pharmacol Exp Ther 271:1616–1623, 1994). Absolute amounts of α1, β3, γ1, γ2S and γ2L mRNA transcripts were quantified in these cultures by quantitative, competitive reverse transcription–polymerase chain reaction (RT-PCR) using subunit-selective internal standards. The most abundant GABAA subunit transcript was β3, which was much more highly expressed than γ2S, γ1, γ2L, or α1 subunit mRNAs. Dieldrin differentially regulated expression of these transcripts. Levels of β3 subunit transcripts were significantly increased (by 300%) by dieldrin, whereas expression of γ2S and γ2L transcripts were decreased (by 50% and 40%, respectively). However, dieldrin did not alter the ratio of γ2S to γ2L transcripts, indicating that it did not affect alternative splicing of γ2 transcripts. Dieldrin appeared to increase expression of α1 subunit transcripts, but this effect was not statistically significant. Dieldrin did not significantly alter expression of γ1 subunit transcripts. These results support the hypothesis that in utero exposure to cyclodiene pesticides could pose a risk to the developing brain by virtue of their ability to alter gene expression of GABAA receptor subunits, which could produce GABAA receptors with altered functional properties. J. Neurosci. Res. 49:645–653, 1997. © 1997 Wiley-Liss, Inc.