title: G protein dependent alterations in [125I]iodocyanopindolol and +/- cyanopindolol binding at 5-HT1B binding sites in rat brain membranes.
abstract: Several manipulations that affect G protein/receptor coupling also alter the binding of [125I]iodocyanopindolol ([125I]ICYP) and [corrected] +/- cyanopindolol (+/- CYP) to rat brain 5-HT1B binding sites in radioligand binding assays. Inclusion of 5 mM MgSO4 in these assays results in a small but significant increase in the affinity of [125I]ICYP (from KD = 0.046 nM to KD = 0.037 nM). In contrast, 100 microM Gpp(NH)p, GTP, or GDP reduce [125I]ICYP affinity (KD = 0.056 nM with GTP) while ATP and GMP are less effective. +/- CYP affinity for 5-HT1B sites labeled by [3H]dihydroergotamine [( 3H]DE) also displays a small but significant reduction (from Ki = 1.4 nM to Ki = 3.5nM) by the inclusion of 100 microM GTP. Pre-treatment of the brain membranes with N-ethylmaleimide (NEM) in concentrations known to inactivate many G proteins reduces 5-HT1B specific binding of [125I]ICYP. The NEM induced reduction in [125I]ICYP binding can be reversed by reconstitution with purified exogenous G proteins (Go and Gi), demonstrating directly that high affinity binding of [125I]ICYP to 5-HT1B sites is dependent on G proteins. The effects of Mg2+ ion, guanine nucleotides, NEM and G protein reconstitution on [125I]ICYP and +/- CYP binding are all hallmarks of agonist binding to G protein linked receptors. The effect of GTP, however, is quantitatively much less for the binding of these pindolol derivatives than for the binding of 5-HT, a presumed full agonist at 5-HT1B sites.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, California 94305.
authors.forename: K
authors.initials: K
authors.lastname: Ariani
authors.forename: M W
authors.initials: MW
authors.lastname: Hamblin
authors.forename: G L
authors.initials: GL
authors.lastname: Tan
authors.forename: C A
authors.initials: CA
authors.lastname: Stratford
authors.forename: R D
authors.initials: RD
authors.lastname: Ciaranello
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D004087:Dihydroergotamine / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000502:physiology*; D006153:Guanosine Diphosphate / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D020120:Iodocyanopindolol; D008278:Magnesium Sulfate / Q000494:pharmacology; D010869:Pindolol / Q000031:analogs & derivatives* / Q000378:metabolism; D051381:Rats; D011985:Receptors, Serotonin / Q000378:metabolism*
chemical_list: D011985:Receptors, Serotonin; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D004087:Dihydroergotamine; C047975:cyanopindolol; D008278:Magnesium Sulfate; D020120:Iodocyanopindolol; D006160:Guanosine Triphosphate; D010869:Pindolol; D019204:GTP-Binding Proteins; D005033:Ethylmaleimide