title: Pentylenetetrazole-induced inhibition of recombinant gamma-aminobutyric acid type A (GABA(A)) receptors: mechanism and site of action.
abstract: Pentylenetetrazole (PTZ) is a central nervous system convulsant that is thought, based on binding studies, to act at the picrotoxin (PTX) site of the gamma-aminobutyric acid type A (GABA(A)) receptor. In the present study, we have investigated the mechanism and site of action of PTZ in recombinant GABA(A) receptors. In rat alpha 1 beta 2 gamma 2 receptors, PTZ inhibited GABA-activated Cl(-) current in a concentration-dependent, voltage-independent manner, with an IC(50) of 0.62 +/- 0.13 mM. The mechanism of inhibition appeared competitive with respect to GABA in both rat and human alpha 1 beta 2 gamma 2 receptors. Varying subunit configuration (change or lack of alpha subunit isoform or lack of gamma 2 subunit) had modest effects on PTZ-induced inhibition, as evidenced by comparable IC(50) values (0.6-2.2 mM) in all receptor configurations tested. This contrasts with PTX and other PTX-site ligands, which have greater affinity in receptors lacking an alpha subunit. Using a one-site model for PTZ interaction with alpha 1 beta 2 gamma 2 receptors, the association rate (k(+1)) was found to be 1.14 x 10(3) M(-1) s(-1) and the dissociation rate (k(-1)) was 0.476 s(-1), producing a functional k(d) of 0.418 mM. PTZ could only gain access to its binding site extracellularly. Single-channel recordings demonstrated that PTZ decreased open probability by increasing the duration of closed states but had no effect on single-channel conductance or open state duration. alpha-Isopropyl-alpha-methyl-gamma-butyrolactone, a compound known to antagonize effects of PTX, also diminished the effects of PTZ. Taken together, our results indicate that pentylenetetrazole and picrotoxin interact with overlapping but distinct domains of the GABA(A) receptor.
authors.affiliation: Department of Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, Texas 76107, USA.
authors.forename: R Q
authors.initials: RQ
authors.lastname: Huang
authors.forename: C L
authors.initials: CL
authors.lastname: Bell-Horner
authors.forename: M I
authors.initials: MI
authors.lastname: Dibas
authors.forename: D F
authors.initials: DF
authors.lastname: Covey
authors.forename: J A
authors.initials: JA
authors.lastname: Drewe
authors.forename: G H
authors.initials: GH
authors.lastname: Dillon
mesh_terms: D000465:Algorithms; D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D003292:Convulsants / Q000494:pharmacology*; D004594:Electrophysiology; D018756:GABA Antagonists / Q000494:pharmacology; D058787:GABA-A Receptor Antagonists; D006801:Humans; D007700:Kinetics; D009865:Oocytes; D018408:Patch-Clamp Techniques; D010433:Pentylenetetrazole / Q000494:pharmacology*; D010852:Picrotoxin / Q000494:pharmacology; D014981:Xenopus
chemical_list: D003292:Convulsants; D018756:GABA Antagonists; D058787:GABA-A Receptor Antagonists; D010433:Pentylenetetrazole; D010852:Picrotoxin