ArticleJournal of neuroscience methods2009
The optimization of TaqMan real-time RT-PCR assay for transcriptional profiling of GABA-A receptor subunit plasticity.
Article in Journal of neuroscience methods, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 25 papers.
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Who cites it
25 citing papers in PubMed, 57 citations in OpenAlex.
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- PR-independent neurosteroid regulation of α2-GABA-A receptors in the hippocampus subfields.Brain research · 2017Article
- Cysteine mutagenesis improves the production without abrogating antigenicity of a recombinant protein vaccine candidate for human chagas disease.Human vaccines & immunotherapeutics · 2017Article
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- Perimenstrual-like hormonal regulation of extrasynaptic δ-containing GABAA receptors mediating tonic inhibition and neurosteroid sensitivity.The Journal of neuroscience : the official journal of the Society for Neuroscience · 2014Article
- Neurosteroid interactions with synaptic and extrasynaptic GABA(A) receptors: regulation of subunit plasticity, phasic and tonic inhibition, and neuronal network excitability.Psychopharmacology · 2013Review
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2 authors.
Funding
Abstract
The GABA-A receptor plays a critical role in inhibitory neurotransmission in the brain. Quantitation of GABA-A receptor subunits in various brain regions is essential to understand their role in plasticity and brain disorders. However, conventional RNA assays are tedious and less sensitive for use in studies of subunit plasticity. Here we describe optimization of a sensitive assay of GABA-A receptor subunit gene expression by TaqMan real-time PCR. For each subunit gene, a set of primers and TaqMan fluorogenic probe were designed to specifically amplify the target template. The TaqMan methodology was optimized for quantification of mouse GABA-A receptor subunits (alpha(1-6), beta(1-3), gamma(2), and delta) and GAPDH. The TaqMan reaction detected very low levels of gene expression ( approximately 100 template copies of cDNA). A standard curve for GAPDH and one of the target genes, constructed using the cDNA, revealed slopes around -3.4 (r(2)=0.990), reflecting similar optimum PCR efficiencies. The methodology was utilized for quantification of the GABA-A receptor alpha(4)-subunit, which is known to upregulate following withdrawal from chronic progesterone or neurosteroids. Our results show that the alpha(4)-subunit expression increased threefold in the hippocampus following neurosteroid withdrawal in mice. The TaqMan PCR assay allows sensitive, high-throughput transcriptional profiling of complete GABA-A receptor subunit family, and thus provides specific tool for studies of GABA-A receptor subunit plasticity in neurological and psychiatric animal models.
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