title: meta- and para-isothiocyanato-t-butylbicycloorthobenzoate: irreversible ligands of the gamma-aminobutyric acid-regulated chloride ionophore.
abstract: The meta- and para-isothiocyanato derivatives of t-butylbicycloorthobenzoate (TBOB) were synthesized by catalytic reduction of the corresponding nitro compounds, followed by treatment with thiophosgene. p-NCS-TBOB (2) inhibited the binding of both [3H]TBOB and [35S]t-butylbicyclophosphorothionate (TBPS) with potencies (IC50 of 61 and 23 nM, respectively) similar to the parent compound. In contrast, the meta derivative (m-NCS-TBOB, 1) was more than 1 order of magnitude less potent (IC50 of 1588 and 149 nM, respectively). The IC50 values for both 1 and 2 were strongly dependent on the tissue concentration, in a manner characteristic of irreversible inhibitors. Moreover, preincubation of tissue with these compounds, followed by extensive washing, resulted in a concentration-dependent reduction in the number of [35S]TBPS binding sites and in the apparent affinity of this radioligand. Similar effects were not observed in tissues treated in identical fashion with either TBOB or picrotoxin. Preincubation with p-NCS-TBOB at concentrations that significantly inhibit [35S]TBPS or [3H]TBOB binding did not affect radioligand binding to either benzodiazepine or gamma-aminobutyric acid receptors. These findings suggest that m- and p-NCS-TBOB bind irreversibly to sites labeled by cage convulsants such as TBOB and TBPS, which are on or near GABA-gated chloride channels. p-NCS-TBOB should prove useful in determining the molecular characteristics of the benzodiazepine receptor-coupled GABA-gated chloride ionophore.
authors.affiliation: Laboratory of Neuroscience, National Institute of Diabetes, Digestive, and Kidney Diseases, Bethesda, MD 20892.
authors.forename: A H
authors.initials: AH
authors.lastname: Lewin
authors.forename: B R
authors.initials: BR
authors.lastname: de Costa
authors.forename: K C
authors.initials: KC
authors.lastname: Rice
authors.forename: P
authors.initials: P
authors.lastname: Skolnick
mesh_terms: D000818:Animals; D001643:Bridged Bicyclo Compounds / Q000378:metabolism / Q000494:pharmacology*; D019086:Bridged Bicyclo Compounds, Heterocyclic; D001952:Bridged-Ring Compounds / Q000494:pharmacology*; D018118:Chloride Channels; D002712:Chlorides / Q000378:metabolism*; D066298:In Vitro Techniques; D017879:Isothiocyanates; D008024:Ligands; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains; D013329:Structure-Activity Relationship; D013861:Thiocyanates / Q000494:pharmacology; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology*
chemical_list: D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D001952:Bridged-Ring Compounds; D018118:Chloride Channels; D002712:Chlorides; D017879:Isothiocyanates; D008024:Ligands; D008565:Membrane Proteins; D013861:Thiocyanates; C037152:isothiocyanic acid; D005680:gamma-Aminobutyric Acid; C037476:tert-butylbicyclophosphorothionate; C046308:tert-butylbicyclo-2-benzoate