title: Characterization of [3H]forskolin binding sites in the iris-ciliary body of the albino rabbit.
abstract: [3H]Forskolin binding sites were identified using membranes prepared from the iris-ciliary body of adult, albino rabbits. Scatchard analysis of saturation binding experiments demonstrated that [3H]forskolin bound to a single population of high affinity sites. The Kd and Bmax values were 8.7 +/- 0.9 nM and 119.0 +/- 30.9 fmol/mg prot. using membranes prepared from frozen tissue and 17.0 +/- 6.2 nM and 184.4 +/- 47.2 fmol/mg prot. using fresh tissue. The binding of [3H]forskolin was magnesium-dependent. The Bmax was enhanced by sodium fluoride and Gpp(NH)p, a nonhydrolyzable guanine nucleotide analog. Forskolin was the most potent inhibitor of [3H]forskolin binding; two commercially-available analogs were weaker inhibitors. In an adenylate cyclase assay, there was the same rank order of potency to enhance enzyme activity. Based upon binding affinities, magnesium-dependence, sensitivity to sodium fluoride and Gpp(NH)p, rank order of potencies of analogs and correlation of binding with adenylate cyclase activity, these studies suggest that the [3H]forskolin binding site in the iris-ciliary body is similar to the binding site in other tissues.
authors.affiliation: Department of Microbial Pharmacometrics, Merck Sharp and Dohme Research Laboratories, West Point, PA 19486.
authors.forename: M E
authors.initials: ME
authors.lastname: Goldman
authors.forename: P
authors.initials: P
authors.lastname: Mallorga
authors.forename: D J
authors.initials: DJ
authors.lastname: Pettibone
authors.forename: M F
authors.initials: MF
authors.lastname: Sugrue
mesh_terms: D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D002924:Ciliary Body / Q000378:metabolism*; D005576:Colforsin / Q000378:metabolism* / Q000494:pharmacology; D005260:Female; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D007498:Iris / Q000378:metabolism*; D008274:Magnesium / Q000494:pharmacology; D011817:Rabbits; D012969:Sodium Fluoride / Q000494:pharmacology; D014316:Tritium
chemical_list: D014316:Tritium; D005576:Colforsin; D006165:Guanylyl Imidodiphosphate; D012969:Sodium Fluoride; D000262:Adenylyl Cyclases; D008274:Magnesium