γ-Aminobutyric acid enhancement of CL 218,872 affinity and evidence of benzodiazepine receptor heterogeneity

J. W. Regan, W. R. Roeske, J. B. Malick, S. H. Yamamura, H. I. Yamamura

Research output: Contribution to journalArticle

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Abstract

The inhibition of [3H]flunitrazepam binding by a novel anxiolytic agent [CL 218,872; 3-methyl-6-[3-trifluoromethyl)phenyl]-1,2,3-triazolo-[4,3-b]pyridazone] was studied in membranes prepared from bovine retina and rat cerebral cortex, cerebellum, and kidney. The order of potency for the inhibition of [3H]flunitrazepam binding by CL 218,872 in these tissues was cerebellum > retina (approx. equal) cerebral cortex >> kidney. The slope factors (Hill coefficients) for CL 218,872 inhibition of [3H]flunitrazepam were approximately 1.0 for kidney, 0.9 for cerebellum, and 0.7 for cerebral cortex and retina. In thoroughly washed membrane preparations from all of the central tissues, K(i) values were insignificantly decreased an average of 60% in the presence of 100 μM γ-aminobutyric acid (GABA) (p < 0.01). With kidney membranes there was no apparent affect of GABA on the K(i) of CL 218,872. (+)-Bicuculline (100 μM) could antagonize the effect of GABA on membranes from central tissues. Nonlinear least-squares regression analyses were used to reanalyze these data in terms of receptor models describing the interaction of a ligand with either one or two classes of independent binding sites. A two-site regression model resulted in a highly significant improvement in the fit of data obtained from retina, cerebral cortex, and cerebellum (p < 0.01), but not with data from kidney (p > 0.05). GABA was found to enhance the affinity of CL 218,872 for both of the sites without changing the proportion of sites. The results of these studies show that GABA enhances the afifnity of CL 218,872 for the central benzodiazepine receptor(s) and that the inhibition of [3H]flunitrazepam binding by CL 218,872 in bovine retina rat cerebellum, and cerebral cortex may be explained by interactions with two classes of independent binding sites.

Original languageEnglish (US)
Pages (from-to)477-483
Number of pages7
JournalMolecular Pharmacology
Volume20
Issue number3
StatePublished - 1981

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Aminobutyrates
GABA-A Receptors
Flunitrazepam
Cerebral Cortex
Cerebellum
Retina
gamma-Aminobutyric Acid
Kidney
Membranes
Anti-Anxiety Agents
CL 218872
Binding Sites

ASJC Scopus subject areas

  • Pharmacology

Cite this

γ-Aminobutyric acid enhancement of CL 218,872 affinity and evidence of benzodiazepine receptor heterogeneity. / Regan, J. W.; Roeske, W. R.; Malick, J. B.; Yamamura, S. H.; Yamamura, H. I.

In: Molecular Pharmacology, Vol. 20, No. 3, 1981, p. 477-483.

Research output: Contribution to journalArticle

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abstract = "The inhibition of [3H]flunitrazepam binding by a novel anxiolytic agent [CL 218,872; 3-methyl-6-[3-trifluoromethyl)phenyl]-1,2,3-triazolo-[4,3-b]pyridazone] was studied in membranes prepared from bovine retina and rat cerebral cortex, cerebellum, and kidney. The order of potency for the inhibition of [3H]flunitrazepam binding by CL 218,872 in these tissues was cerebellum > retina (approx. equal) cerebral cortex >> kidney. The slope factors (Hill coefficients) for CL 218,872 inhibition of [3H]flunitrazepam were approximately 1.0 for kidney, 0.9 for cerebellum, and 0.7 for cerebral cortex and retina. In thoroughly washed membrane preparations from all of the central tissues, K(i) values were insignificantly decreased an average of 60{\%} in the presence of 100 μM γ-aminobutyric acid (GABA) (p < 0.01). With kidney membranes there was no apparent affect of GABA on the K(i) of CL 218,872. (+)-Bicuculline (100 μM) could antagonize the effect of GABA on membranes from central tissues. Nonlinear least-squares regression analyses were used to reanalyze these data in terms of receptor models describing the interaction of a ligand with either one or two classes of independent binding sites. A two-site regression model resulted in a highly significant improvement in the fit of data obtained from retina, cerebral cortex, and cerebellum (p < 0.01), but not with data from kidney (p > 0.05). GABA was found to enhance the affinity of CL 218,872 for both of the sites without changing the proportion of sites. The results of these studies show that GABA enhances the afifnity of CL 218,872 for the central benzodiazepine receptor(s) and that the inhibition of [3H]flunitrazepam binding by CL 218,872 in bovine retina rat cerebellum, and cerebral cortex may be explained by interactions with two classes of independent binding sites.",
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