[3H]t-butylbicycloorthobenzoate binding to recombinant alpha1beta2gamma2s GABA(A) receptor.
The interaction of several selected compounds with the binding of the cage convulsant t-[3H]butylbicycloorthobenzoate ([3H]TBOB) to membranes isolated from human embryonic kidney (HEK) 293 cells stably transfected with alpha1beta2gamma2s subtype of GABA(A) receptors was studied. Scatchard analysis of binding data revealed the existence of a single type of binding site for [3H]TBOB with a Kd of 47.06+/-4.06 nM and a Bmax value of 6.72+/-0.52 pmol/mg protein. GABA, thiopental, TBOB, picrotoxin and the neurosteroid dehydroepiandrosterone sulfate displaced concentration-dependently the binding of [3H]TBOB to this recombinant receptor. Dehydroepiandrosterone sulfate reversed the 5 microM GABA-induced inhibition of specific [3H]TBOB binding. It is concluded that membranes isolated from HEK 293 cells stably transfected with alpha1beta2gamma2s subunits exhibit specific high-affinity [3H]TBOB binding. The potency of drugs to inhibit [3H]TBOB binding mainly corresponded to that observed for the inhibition of the binding of cage convulsants to the native receptors or to transiently transfected HEK 293 cells.
Pericić
D
D
Ruder Bosković Institute, Laboratory for Molecular Neuropharmacology, Zagreb, Croatia. pericic@rudjer.irb.hr
Mirković
K
K
Jazvinsćak
M
M
Besnard
F
F
D001667:Binding, Competitive / Q000187:drug effects; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D019314:Dehydroepiandrosterone Sulfate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D018756:GABA Antagonists / Q000494:pharmacology; D018757:GABA Modulators / Q000494:pharmacology; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000648:ultrastructure; D010852:Picrotoxin / Q000494:pharmacology; D011869:Radioligand Assay; D011963:Receptors, GABA-A / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013874:Thiopental / Q000494:pharmacology; D014316:Tritium; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology
D019086:Bridged Bicyclo Compounds, Heterocyclic; D019314:Dehydroepiandrosterone Sulfate; D018756:GABA Antagonists; D018757:GABA Modulators; D010852:Picrotoxin; D011963:Receptors, GABA-A; D011994:Recombinant Proteins; D013874:Thiopental; D014316:Tritium; D005680:gamma-Aminobutyric Acid; C046308:tert-butylbicyclo-2-benzoate