title: Peripheral-type mitochondrial binding sites for benzodiazepines in GH3 pituitary cells.
abstract: Benzodiazepines (BZs) interact with two classes of high affinity binding sites, equilibrium dissociation constants in the nanomolar range, a neuronal or central-type and a non-neuronal or peripheral-type. The peripheral-type binding site has been shown to be present on the outer mitochondrial membrane and appears to be involved in regulation of cholesterol transport in steroid hormone-producing endocrine cells. In rat pituitary GH3 cells, BZs bind to receptors for thyrotropin-releasing hormone (TRH) and via interaction at a different site block Ca2+ influx through voltage-sensitive channels. These, however, are low affinity interactions occurring at micromolar BZ concentrations. Here, using [3H]Ro 5-4864, we report that GH3 cells also have high affinity peripheral-type BZ binding sites. Apparent equilibrium dissociation constants of 7.8 +/- 1.7 nM and 9.3 +/- 4.5 nM for [3H]Ro 5-4864 were measured with intact cells and isolated mitochondria, respectively. As predicted from studies of these sites in other cells, the order of potencies of BZs to displace [3H]Ro 5-4864 was Ro 5-4864 greater than diazepam (DZP) much greater than clonazepam (CIZP); chlordiazepoxide (CDE) did not affect binding. Nifedipine, a dihydropyridine antagonist of Ca2+ channels that has been shown to displace BZs from peripheral-type sites in other cells, was shown to be a competitive inhibitor of [3H]Ro 5-4864 binding with a half-effective concentration in the micromolar range. Ro 5-4864, however, had no effect on Ca2+ influx or efflux in mitochondria isolated from GH3 cells. Hence, GH3 cells exhibit mitochondrial, peripheral-type BZ binding sites but the role of these putative receptors in these neuroendocrine cells, which do not produce steroid hormones, is unclear.
authors.affiliation: Department of Medicine, Cornell University Medical College, New York, New York 10021.
authors.forename: M C
authors.initials: MC
authors.lastname: Gershengorn
authors.forename: V S
authors.initials: VS
authors.lastname: Purrello
authors.forename: E
authors.initials: E
authors.lastname: Geras-Raaka
mesh_terms: D000818:Animals; D001569:Benzodiazepines / Q000378:metabolism*; D001570:Benzodiazepinones / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002707:Chlordiazepoxide / Q000378:metabolism / Q000494:pharmacology; D002998:Clonazepam / Q000378:metabolism / Q000494:pharmacology; D003975:Diazepam / Q000378:metabolism / Q000494:pharmacology; D008928:Mitochondria / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D051381:Rats
chemical_list: D001570:Benzodiazepinones; D001569:Benzodiazepines; C028513:4'-chlorodiazepam; D002998:Clonazepam; D002707:Chlordiazepoxide; D009543:Nifedipine; D003975:Diazepam; D002118:Calcium