GABA-site antagonism and pentobarbital actions do not depend on the alpha-subunit type in the recombinant rat GABA receptor.
OBJECTIVE
The roles of alpha-subunits on the gamma-aminobutyric acid (GABA)-site antagonism and pentobarbital actions were examined in rat recombinant GABA(A) receptors in Xenopus oocytes.


METHODS
Experiments were performed with binary and ternary GABA(A) receptors containing alpha1-, alpha4- or alpha5-subunit by the two-electrode voltage-clamp technique.


RESULTS
The potency of GABA was significantly higher in the alpha1beta2, alpha4beta2 and alpha5beta2 receptors compared with the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors. However, the alpha5beta2 receptor possessed significantly lower GABA efficacy compared with the alpha5beta2gamma2L receptor. While the gamma2-subunit was essential to the potency of GABA, its influence on the apparent GABA-site antagonism was less profound. The antagonist affinity constants (K(B)) of bicuculline inhibition and slopes of Schild plots were similar between all types of ternary and binary receptors except alpha5beta2 receptor which was not tested. The pK(B)s and IC(50)s of the GABA-site antagonism were not significantly different between the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors. Bicuculline blocked pentobarbital-activated currents in a reversible and non-competitive manner with the alpha1beta2gamma2L, alpha4beta2gamma2L, and alpha5beta2gamma2L receptors, indicating an allosteric inhibition of the GABA-site. No significant difference of bicuculline potencies in inhibiting GABA- and pentobarbital-activated currents was found between the alpha1beta2gamma2L, alpha4beta2gamma2L and alpha5beta2gamma2L receptors.


CONCLUSIONS
The GABA-site antagonism does not depend on the subtype of alpha-subunits. Similarly, pentobarbital activates ternary receptors composed of different alpha-subunits in a bicuculline-sensitive manner. The potencies of bicuculline to inhibit pentobarbital-activated currents are identical with receptors containing alpha1, alpha4 or alpha5-subunit. The alpha1beta2 and alpha4beta2 receptors possess higher GABA potencies compared with the alpha1beta2gamma2L and alpha4beta2gamma2L receptors.
Rahman
M
M
Department of Clinical Science, Umeå Neurosteroid Research Center, Obstetrics and Gynecology, Umeå University, Umeå, Sweden.
Zhu
D
D
Lindblad
C
C
Johansson
I-M
IM
Holmberg
E
E
Isaksson
M
M
Taube
M
M
Bäckström
T
T
Wang
M-D
MD
D000818:Animals; D001640:Bicuculline / Q000031:analogs & derivatives / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005260:Female; D018756:GABA Antagonists / Q000494:pharmacology*; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010424:Pentobarbital / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D051381:Rats; D011963:Receptors, GABA-A / Q000187:drug effects* / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D014981:Xenopus
D018756:GABA Antagonists; D011963:Receptors, GABA-A; D011994:Recombinant Proteins; C017069:bicuculline methiodide; D010424:Pentobarbital; D001640:Bicuculline