GABArho 1/GABAAalpha 1 receptor chimeras to study receptor desensitization.
gamma-Aminobutyrate type C (GABA(C)) receptors are ligand-gated ion channels that are expressed preponderantly in the vertebrate retina and are characterized, among other things, by a very low rate of desensitization and resistance to the specific GABA(A) antagonist bicuculline. To examine which structural elements determine the nondesensitizing character of the human homomeric rho1 receptor, we used a combination of gene chimeras and electrophysiology of receptors expressed in Xenopus oocytes. Two chimeric genes were constructed, made up of portions of the rho1-subunit and of the alpha1-subunit of the GABA(A) receptor. When expressed in Xenopus oocytes, one chimeric gene (rho1/alpha1) formed functional homooligomeric receptors that were fully resistant to bicuculline and were blocked by the specific GABA(C) antagonist (1,2,5, 6-tetrahydropyridine-4-yl)methylphosphinic acid and by zinc. Moreover, these chimeric receptors had a fast-desensitizing component, even faster than that of heterooligomeric GABA(A) receptors, in striking contrast to the almost nil desensitization of wild-type rho1 (wt rho1) receptors. To see whether the fast-desensitizing characteristic of the chimera was determined by the amino acids forming the ion channels, we replaced the second transmembrane segment (TM2) of rho1 by that of the alpha1-subunit of GABA(A). Although the alpha1-subunit forms fast-desensitizing receptors when coexpressed with other GABA(A) subunits, the sole transfer of the alpha1TM2 segment to rho1 was not sufficient to form desensitizing receptors. All this suggests that the slow-desensitizing trait of rho1 receptors is determined by a combination of several interacting domains along the molecule.
Martínez-Torres
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A
Laboratory of Cellular and Molecular Neurobiology, Department of Neurobiology and Behavior, University of California, Irvine, CA 92697, USA.
Demuro
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Miledi
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R
D000818:Animals; D018387:Codon, Initiator; D005260:Female; D018756:GABA Antagonists / Q000494:pharmacology; D006801:Humans; D007473:Ion Channels / Q000187:drug effects; D008564:Membrane Potentials / Q000187:drug effects; D009865:Oocytes / Q000502:physiology; D010721:Phosphinic Acids / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D018079:Receptors, GABA / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis
C101879:(1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid; D018387:Codon, Initiator; D018756:GABA Antagonists; D007473:Ion Channels; D010721:Phosphinic Acids; D011725:Pyridines; D018079:Receptors, GABA; D011993:Recombinant Fusion Proteins