Diazepam is not a direct allosteric modulator of α1‐adrenoceptors, but modulates receptor signaling by inhibiting phosphodiesterase‐4

Pharmacology Research & Perspectives

Paul R. Gooley¹  Spencer J. Williams²  Alaa Abdul‐Ridha³  Xiaoji He³  Daniel J. Scott³  Tasneem M. Vaid³  Alice R. Whitehead³  Ross A.D. Bathgate³  Lisa M. Williams³ 

[1] Department of Biochemistry and Molecular Biology University of Melbourne Parkville Vic Australia;School of Chemistry and Bio21 Molecular Science and Biotechnology Institute University of Melbourne Parkville Vic Australia;The Florey Institute of Neuroscience and Mental Health University of Melbourne Parkville Vic Australia;

关键词: α1 adrenergic receptors;    α1‐adrenoceptors;    allosteric modulator;    benzodiazepines;    GPR68;    phosphodiesterase;   

DOI  :  10.1002/prp2.455

来源: DOAJ

【 摘 要 】

Abstract α1A‐ and α1B‐adrenoceptors (ARs) are G protein‐coupled receptors (GPCRs) that are activated by adrenaline and noradrenaline to modulate smooth muscle contraction in the periphery, and neuronal outputs in the central nervous system (CNS). α1A‐ and α1B‐AR are clinically targeted with antagonists for hypertension and benign prostatic hyperplasia and are emerging CNS targets for treating neurodegenerative diseases. The benzodiazepines midazolam, diazepam, and lorazepam are proposed to be positive allosteric modulators (PAMs) of α1‐ARs. Here, using thermostabilized, purified, α1A‐ and α1B‐ARs, we sought to identify the benzodiazepine binding site and modulatory mechanism to inform the design of selective PAMs. However, using a combination of biophysical approaches no evidence was found for direct binding of several benzodiazepines to purified, stabilized α1A‐ and α1B‐ARs. Similarly, in cell‐based assays expressing unmodified α1A‐ and α1B‐ARs, benzodiazepine treatment had no effect on fluorescent ligand binding, agonist‐stimulated Ca2+ release, or G protein activation. In contrast, several benzodiazepines positively modulated phenylephrine stimulation of a cAMP response element pathway by α1A‐ and α1B‐ARs; however, this was shown to be caused by off‐target inhibition of phosphodiesterases, known targets of diazepam. This study highlights how purified, stabilized GPCRs are useful for validating allosteric ligand binding and that care needs to be taken before assigning new targets to benzodiazepines.

【 授权许可】

Unknown