Endogenous [3H]flunitrazepam binding in human embryonic kidney cell line 293

https://doi.org/10.1016/0922-4106(95)90172-8Get rights and content

Abstract

Specific endogenous [3H]flunitrazepam binding sites were identified and characterized in membranes from the human embryonic kidney (HEK) cell line 293. A large part of these binding sites exhibited an intermediate affinity for [3H]flunitrazepam and a μM affinity for diazepam, clonazepam, 1-(2-chlorophenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinolinecarboxamide (PK 11195) or 4′-chlorodiazepam (Ro 5-4864). These sites, thus, resembled neither γ-aminobutyric acidA (GABAA) receptor associated nor ‘peripheral’ benzodiazepine binding sites.

A small part of the binding sites labeled by [3H]flunitrazepam seemed to belong to ‘peripheral’ benzodiazepine binding sites exhibiting a nM affinity for PK 11195, and another small part of the binding sites seemed to exhibit a high affinity for flunitrazepam and PK 11195.

Although small amounts of mRNA for α1-, β3- and γ2-subunits of GABAA receptors could be identified in HEK 293 cells, neither the actual expression of GABAA receptors in these cells nor a coassembly of endogenous subunits with transfected GABAA receptor subunits could be demonstrated by binding studies.

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