3 alpha-hydroxy-5 alpha-pregnan-20-one modulation of solubilized GABA/benzodiazepine receptor complex

• Published in: The Journal of steroid biochemistry and molecular biology Vol. 45; no. 4; pp. 309 - 314
• Main Authors: Giusti, L, Belfiore, M S, Martini, C, Lucacchini, A
• Format: Journal Article
• Language: English
• Published: England 01.04.1993
• Subjects: Animals; Binding Sites; Bridged Bicyclo Compounds - metabolism; Bridged Bicyclo Compounds, Heterocyclic; Cerebral Cortex - drug effects; Cerebral Cortex - metabolism; Flunitrazepam - metabolism; Pregnanolone - pharmacology; Radioligand Assay; Rats; Receptors, GABA-A - chemistry; Receptors, GABA-A - drug effects; Receptors, GABA-A - metabolism; Solubility
• ISSN: 0960-0760
• DOI: 10.1016/0960-0760(93)90347-Y

The allosteric modulation of the progesterone metabolite 3 alpha- hydroxy-5 alpha-pregnan-20-one (DHP) on [3H]Flunitrazepam and [35S]t-butylbicyclophosphorothionate (TBPS) binding was investigated on a soluble receptor preparation. Better results in the solubilization occurred by the use of the zwitterionic detergent CHAPS with the inclusion of the phospholipid asolectin: this treatment was found suitable to study the steroidal modulation on [3H]Flunitrazepam and [35S]TBPS binding. We found that DHP was able to enhance [3H]Flunitrazepam binding in the presence of Cl- ions, while [35S]TBPS binding was inhibited by DHP. Scatchard analysis of specific [35S]TBPS and [3H]Flunitrazepam binding yielded in a single straight line both in the controls and in the presence of the hormone; DHP increased the apparent affinity of [3H]Flunitrazepam binding without altering the apparent Bmax value. In the case of [35S]TBPS, DHP decreased the apparent Bmax value whereas the Kd value remained nearly the same.