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Volume 16, Number 17, Issue of September 1, 1996 pp. 5415-5424
Copyright ©1996 Society for Neuroscience

Stoichiometry of a Recombinant GABA_A Receptor

Received May 1, 1996; revised June 14, 1996; accepted June 18, 1996.

Yongchang Chang, Ruoping Wang, Sonal Barot, and David S. Weiss

Neurobiology Research Center, Department of Physiology and Biophysics, University of Alabama at Birmingham, Birmingham, Alabama 35294

GABA is the main inhibitory neurotransmitter in the mammalian brain. The postsynaptic GABA_A receptor/pore complex is presumed to be a pentamer typically composed of a combination of , , and  subunits, although the stoichiometry remains controversial. We probed the stoichiometry of the GABA_A receptor by site-directed mutagenesis of a conserved leucine (to serine) in the putative second membrane-spanning domain of the rat 1(L263S), 2(L259S), and 2(L274S) subunit isoforms. Coexpression of wild-type and mutant subunits of each class (e.g.,  and L263S), along with their wild-type counterparts (e.g.,  and ), in Xenopus laevis oocytes resulted in mixed populations of receptors with distinct GABA sensitivities. This is consistent with the interpretation that the leucine mutation increased the GABA sensitivity in proportion to the number of incorporated mutant subunits. The apparent number of incorporated subunits for each class (, , and ) could then be determined from the number of components comprising the compound GABA dose-response relationships. Using this approach, we conclude that the recombinant 122 GABA_A receptor is a pentamer composed of two  subunits, two  subunits, and one  subunit.

Key words: GABA; stoichiometry; receptor; ion channel; mutagenesis

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