title: Stable guanosine 5'-triphosphate-analogues inhibit specific (+)-[3H]isradipine binding in rat hearts by a Ca(2+)-lowering, G protein-independent mechanism.
abstract: We investigated if and how stable guanosine 5'-triphosphate-analogues affect (+)-[3H]isradipine binding in rat hearts. Gpp(NH)p and GTP-gamma-S inhibit specific (+)-[3H]isradipine binding in membranes and cell-homogenates by reducing the binding density without changing the Kd of the k-1. Inhibition by Gpp(NH)p was less in crude tissue homogenates than in membranes apparently due to a soluble factor. Pretreatment of cardiomyocytes with cholera toxin or the presence of the protein kinase A inhibitor, PKI6-22, did not influence the effect of 10(-3) M Gpp(NH)p on binding. The inhibitory effect of 10(-3) M Gpp(NH)p was not significantly altered in membranes from in vivo pertussis toxin treated rats. The addition of 10(-3) M Ca2+ or Mg2+ abolished the inhibitions. Gpp(NH)p in the concentration that inhibits binding, reduced the free concentration of Ca2+. The Ca(2+)-lowering effect of 10(-3) M Gpp(NH)p produced 70%, 60% and 100% of the inhibition in membranes, sonicated and unsonicated cell homogenates. Thus, Gpp(NH)p inhibited specific (+)-[3H] isradipine binding mainly by lowering the free concentration of Ca2+ by chelation and not by activation of cholera toxin or pertussis toxin-sensitive G proteins or protein kinase A.
authors.affiliation: Clinical Chemistry Department, Ullevål University Hospital, Oslo, Norway.
authors.forename: M
authors.initials: M
authors.lastname: Stokke
authors.forename: E M
authors.initials: EM
authors.lastname: Hagelin
authors.forename: U
authors.initials: U
authors.lastname: Mende
authors.forename: O
authors.initials: O
authors.lastname: Brørs
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology*; D006321:Heart / Q000187:drug effects; D066298:In Vitro Techniques; D017275:Isradipine / Q000493:pharmacokinetics*; D008274:Magnesium / Q000494:pharmacology; D008297:Male; D008566:Membranes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D009206:Myocardium / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D011869:Radioligand Assay; D051381:Rats; D017208:Rats, Wistar; D013696:Temperature; D014665:Vasodilator Agents / Q000493:pharmacokinetics*
chemical_list: D015220:Calcium Channels; D014665:Vasodilator Agents; D006165:Guanylyl Imidodiphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D017868:Cyclic AMP-Dependent Protein Kinases; D019204:GTP-Binding Proteins; D008274:Magnesium; D002118:Calcium; D017275:Isradipine