Distinctive patterns of GABAA receptor subunit mRNAs in 13 cell lines

Abstract

We have investigated the GABAA receptor mRNA composition in 13 cell lines, using 13 subunit-specific oligo-primers (alpha 1-6, beta 1-3, gamma 1-3, and delta) and reverse transcriptase PCR amplification. Cell lines (B35, B65, B103, B104, RINm5F, Rat1, PC12, C6, C17, C27, beta TC3, NB41A3, AtT-20), derived from diverse tissue origins, were investigated in order to identify homogeneous cellular sources with distinctive GABAA receptor subunits. Fifteen GABAA receptor subunits have been cloned from mammalian tissue (those listed above plus the retinal subunits rho 1 and rho 2). This multiplicity of GABAA receptor subunits underlies the diverse pharmacology of the GABAA receptor. Attempts to understand the regulation and pharmacology of individual subunits and of the heterooligomeric receptor combinations have been impeded by a lack of pure populations of cells expressing GABAA receptor subunits. Permanent cell lines provide such a resource. Each GABAA receptor subunit mRNA, alpha 1-5, beta 1-3, gamma 1-3, and delta, was detected in at least one cell line. All cell lines examined contained detectable levels of at least one GABAA receptor subunit mRNA. Each cell line contained distinctive combinations of subunit mRNAs. None of the cell lines examined contained detectable amounts of alpha 6 mRNA. These cell lines, which transcribe GABAA receptor subunit mRNAs, provide useful cellular sources for transcriptional and pharmacological studies. Our data also suggest that endogenous GABAA receptor subunit mRNAs may be present in cells that are routinely used for transfection studies, and that this expression might confound interpretation of the studies. In the following companion article, we have looked for functional GABAA receptor Cl- ion channels in these cell lines, using the patch-clamp technique (Hales and Tyndale, 1994).

Effect of the α subunit subtype on the macroscopic kinetic properties of recombinant GABAA receptors

“…Neuronal receptors are likely to be subject to modulation by post-translational modifications such as phosphorylation (Brandon et al, 2002), interactions with cytoskeletal proteins (Chen and Olsen, 2007), and differences in assembly and membrane targeting (Fritschy et al, 1998;Brünig et al, 2002;Klausberger et al, 2002). However, virtually all neurons and neuronal cell lines that express GABARs produce multiple subunit subtypes (Wisden et al, 1992;Laurie et al, 1992a;Tyndale et al, 1994;Neelands et al, 1998) and neuronal populations which express only one or two of the α subtypes are extremely rare. Therefore, in order to control the subunit composition of the receptors and describe the characteristics of a homogeneous population of receptors, these studies must be done in cell lines that do not normally express functional GABARs.…”

Critical interval of somal calcium transient after neurite transection determines B104 cell survival

“…B104 cells display specific functional characteristics, including propagation of action potentials, expression of plasmalemmal ion channels and transporters, and secretion of neurotransmitter that qualify them as a neuronal model (Tan et al, 2003, Dib-Haji et al, 1996, Gu and Waxman, 1996, Luo et al, 1999, Luo et al, 1997, Tyndale et al, 1994, Bottenstein et al, 1979, Schubert et al, 1975.…”

Few cell lines with GABAA mRNAs have functional receptors

“…In the present study we used the whole-cell configuration of the patch-clamp technique to determine which of these cell lines express functional GABA, receptors. All of the cell lines contain detectable levels of at least one GABA, receptor subunit mRNA (Tyndale et al, 1994); however, only RINm5F and @TC3 cells exhibited GABA-evoked currents. GABA activated currents in all RINm5F cells, but currents were only barely detectable in 50% of BTC3 cells tested.…”

“…In the preceding companion article (Tyndale et al, 1994) we used the PCR to investigate the occurrence of 13 GABA, receptor subunit mRNAs in several cell lines, including those derived from brain (865,8104,and NB41A3), cerebellum (C17), glia (C6), pituitary (AtT-PO), adrenal medulla (PC12), and the endocrine pancreas (RINm5F and BTCB). In the present study we used the whole-cell configuration of the patch-clamp technique to determine which of these cell lines express functional GABA, receptors.…”