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Characterization of High‐Affinity Dopamine D 2 Receptors and Modulation of Affinity States by Guanine Nucleotides in Cholate‐Solubilized Bovine Striatal Preparations
Author(s) -
Kazmi Syed M. I.,
Ramwani Jai,
Srivastava Lalit K.,
Rajakumar G.,
Ross Gregory M.,
Cullen Marjorie,
Mishra Ram K.
Publication year - 1986
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1986.tb00784.x
Subject(s) - solubilization , nucleotide , guanine , chemistry , receptor , dopamine , biochemistry , affinity chromatography , stereochemistry , enzyme , biology , neuroscience , gene
3,4‐Dihydroxyphenylethylamine (dopamine) D 2 receptors, solubilized from bovine striatal membranes using a cholic acid‐NaCl combination, exhibited the typical pharmacological characteristics of both agonist and antagonist binding. The rank order potency of the agonists and antagonists to displace [ 3 H]spiroperidol binding was the same as that observed with membrane‐bound receptors. Computer‐assisted analysis of the [ 3 H]spiroperidol/agonist competition curves revealed the retention of high‐ and low‐affinity states of the D 2 receptor in the solubilized preparations and the proportions of receptor subpopulations in the two affinity states were similar to those reported in membrane. Guanine nucleotide almost completely converted the high‐affinity sites to low‐affinity sites for the agonists. The binding of the high‐affinity agonist [ 3 H] N‐n ‐propylnor‐apomorphine ([ 3 H]NPA) was clearly demonstrated in the solubilized preparations for the first time. Addition of gua‐nylyl‐imidodiphosphate completely abolished the [ 3 H]NPA binding. When the solubilized receptors were subjected to diethylaminoethyl‐Sephacel chromatography, the dopaminergic binding sites eluted in two distinct peaks, showing six‐ to sevenfold purification of the receptors in the major peak. Binding studies performed on both peaks indicated that the receptor subpopulation present in the first peak may have a larger proportion of high‐affinity binding sites than the second peak. The solubilized preparation also showed high‐affinity binding of [ 35 S]guanosine‐5′‐(γ‐thio)triphos‐phate, a result suggesting the presence of guanine nucleotide binding sites, which may interact with the solubilized D 2 receptors. These data are consistent with the retention of the D 2 receptor‐guanine nucleotide regulatory protein complex in the solubilized preparations and should provide a suitable model system to study the receptor‐effector interactions.