Properties of recombinant gamma-aminobutyric acid A receptor isoforms containing the alpha 5 subunit subtype.
The cDNAs encoding alpha 5 and gamma 2L subunit subtypes of the gamma-aminobutyric acid (GABA) type A receptor (GABAR) were transfected into L929 cells together with cDNAs encoding either the beta 1, beta 2, or beta 3 subunit subtype. Properties of expressed recombinant alpha 5 beta X gamma 2L (where X = 1,2, or 3) GABARs were studied with the use of whole-cell, patch-clamp techniques. In cells voltage-clamped at -70 mV with equlvalent bath and pipette chloride concentrations, the application of GABA produced a concentration-dependent inward chloride current with all three alpha 5 beta X gamma 2L isoforms. Minimal or no responses were recorded from cells transfected with only two subunit cDNAs, demonstrating that all three subunits were required for functional receptor assembly in these cells. The GABA concentration producing a half-maximal current was similar for beta 2 and beta 3 subtype-containing receptors (6 microM) but higher for beta 1 subtype-containing receptors (26 microM). alpha 5 beta 3 gamma 2L receptors were zinc and diazepam sensitive but zolpidem insensitive. In response to low GABA concentrations, beta 1 and beta 3 subtype-containing receptors showed outward rectification of the current-voltage relationship, whereas current-voltage responses of beta 2 subtype-containing receptors were relatively linear. Likewise, at high GABA concentrations, beta 1 and beta 3 subtype-containing receptors showed less desensitization at positive than at negative membrane potentials. Beta 2 subtype-containing receptors displayed faster desensitization at depolarized potentials. These voltage-dependent properties were characteristic of alpha 5 but not alpha 1 or alpha 6 subtype-containing receptors and were similar to responses recorded from hippocampal CA1 pyramidal neurons. Based on the pharmacological and biophysical similarities to hippocampal GABAR responses, the alpha 5 beta 3 gamma 2L isoform could represent a native GABAR subtype.
Burgard
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Department of Neurology, University of Michigan, Ann Arbor 48104-1687, USA.
Tietz
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Neelands
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Macdonald
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D000818:Animals; D001569:Benzodiazepines / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D003001:Cloning, Molecular; D018076:DNA, Complementary; D003975:Diazepam / Q000494:pharmacology; D006993:Hypnotics and Sedatives / Q000494:pharmacology; D066298:In Vitro Techniques; D007739:L Cells; D008024:Ligands; D008564:Membrane Potentials / Q000187:drug effects; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D017966:Pyramidal Cells / Q000187:drug effects / Q000502:physiology*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D011963:Receptors, GABA-A / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D014162:Transfection; D015032:Zinc / Q000494:pharmacology; D000077334:Zolpidem; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology*
D018076:DNA, Complementary; D006993:Hypnotics and Sedatives; D008024:Ligands; D011725:Pyridines; D011963:Receptors, GABA-A; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D001569:Benzodiazepines; D005680:gamma-Aminobutyric Acid; D000077334:Zolpidem; D015032:Zinc; D003975:Diazepam