G‐protein activation by putative antagonists at mutant Thr<sup>373</sup>Lys α<sub>2A</sub> adrenergic receptors

Wurch, Thierry; Colpaert, Françis C.; Pauwels, Petrus J.

doi:10.1038/sj.bjp.0702379

Cited by 36 publications

(43 citation statements)

References 31 publications

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“…Combining expression of human a 2A -receptors (particularly with a mutation at Thr 373 ) with rat G á0 protein gives rise to a constitutively active receptor that tonically suppresses production of cyclic AMP (87,113,116). Under these rather artificial circumstances, RX 821002, in common with yohimbine/rauwolscine, usually behaves as an inverse agonist at á 2A -adrenoceptors (87,113).…”

Section: Rx 821002 As An Inverse Agonist At á 2 -Adrenoceptorsmentioning

confidence: 99%

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

Clarke

Harris

2002

CNS Drug Reviews

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RX 821002 is the 2-methoxy congener of idazoxan. In binding and tissue studies it behaves as a selective antagonist of á 2 -adrenoceptors, with at least 5 times greater affinity for these receptors than any other binding site. It does not select between the different types of á 2 -receptor. Although this drug probably has no future as a therapeutic agent, it remains a good probe for physiological activity at á 2 -adrenoceptors in animal experiments. A particularly useful feature of this compound is its lack of binding at I 1 and I 2 imidazoline receptors. However, it has relatively high affinity for 5-HT 1A receptors (at which it acts as an antagonist) and a tendency to behave as an inverse agonist at á 2A -adrenoceptors in some cell culture systems. These potential drawbacks may be overcome by careful design of experiments, and the greater selectivity of RX 821002 renders it much superior to yohimbine or idazoxan as a tool for probing physiological actions at á 2 -receptors. It can be compared favorably with other selective antagonists such as atipamezole.In physiological studies, RX 821002 augments norepinephrine release in the frontal cortex and increases drinking behavior in rat. In rabbit, intrathecal administration of this drug enhances somatic and autonomic motor outflows, showing that tonic adrenergic descending inhibition of withdrawal reflexes and sympathetic pre-ganglionic neurons is strong in this species. The potentiation of reflexes may be considered a pro-nociceptive action. In the same model, RX 821002 antagonizes the inhibitory effects of the ì opioid fentanyl, indicating that exogenous opioids synergize with endogenously released norepinephrine in the spinal cord. Thus, the careful use of RX 821002 has revealed several aspects of the physiological activity of á 2 -adrenoceptors in rabbit spinal cord and rat brain. We recommend that RX 821002 and/or compounds with similar selectivity for á 2 -adrenoceptors (atipamezole, MK-912, RS-79948) should be used in preference to yohimbine or idazoxan in all future studies of this type.

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Section: Rx 821002 As An Inverse Agonist At á 2 -Adrenoceptorsmentioning

confidence: 99%

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

Clarke

Harris

2002

CNS Drug Reviews

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“…Earlier studies had reported little or no difference in antagonist binding for the CAM versus WT receptor (Ren et al, 1993;Wurch et al, 1999). Since yohimbine had previously been shown to be an inverse agonist for G i activation (Tian et al, 1994), we carefully determined its K d value for WT and CAM receptor-expressing membranes to see if the change in receptor conformation expected with the activating mutation would alter the [ 3 H]yohimbine binding.…”

Section: Radiochemicalsmentioning

confidence: 99%

“…The identification of ␣ 2A adrenergic inverse agonists and neutral antagonists that are more selective than yohimbine would be desirable. Furthermore, recent literature is contradictory as to whether the classical ␣ 2 -AR antagonists are inverse agonists or neutral antagonists (Tian et al, 1994;Wurch et al, 1999).…”

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confidence: 99%

Inverse Agonist Activity at the α_2A-Adrenergic Receptor

et al. 2001

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Constitutive activation of G protein-coupled receptors (GPCRs) is now well recognized and many classical GPCR antagonists have been found to be inverse agonists. For the ␣ 2A -adrenergic receptor (AR) we determine the relative inverse efficacies of a series of antagonists and utilize the extended ternary complex model to estimate the fraction of constitutively active mutant (CAM) receptors in the active state. Stable Chinese hamster ovary cell lines expressing the porcine ␣ 2A -AR in its wild-type (WT) and constitutively activated (CAM-T373K) form were isolated. Activation of both G i and G s was enhanced for CAM receptors. cAMP production was suppressed in cells with the CAM ␣ 2A -AR and this suppression was reversed by ␣ 2 -adrenergic antagonists with an order of inverse efficacy of rauwolscine Ͼ yohimbine Ͼ RX821002 Ͼ MK912, whereas phentolamine and idazoxan were essentially neutral antagonists. This striking difference in inverse efficacy between idazoxan and RX821002 may account for in vivo pharmacological differences between these two ␣ 2 -adrenergic antagonists. Agonist binding affinity to the non-G protein-coupled CAM receptor was 3-to 9-fold higher than to WT, whereas binding of the most efficacious inverse agonists, yohimbine and rauwolscine, was 1.7-and 2.1-fold weaker. Analysis of this difference by the extended ternary complex model indicates that approximately 50% of the CAM ␣ 2A -AR is in the active (R*) state although there is no detectable constitutive activity of the WT receptor in the absence of agonist.

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“…In the ␤-adrenergic (40) and 5HT2A (41) Mutations in a number of GPCRs have been described that confer agonism to antagonists. Mutations in TM3 of the AT1 angiotensin receptor (39,42), rhodopsin (43), and the ␤-adrenergic receptor (40), in TM4 of -and ␦-opioid receptors (44), in TM6 of the -opioid receptor (45), in IC3 of the ␣ 2A -adrenergic receptor (46), and in IC2 of the V2 vasopressin receptor (47) conferred agonistic activity to antagonist ligands. The present study is the first description of a mutation in an extracellular domain that confers agonism to an antagonist.…”

Section: Chicken Gnrh Receptor Ec2 Confers Agonism To Antagonistsmentioning

confidence: 99%

A Chicken Gonadotropin-releasing Hormone Receptor That Confers Agonist Activity to Mammalian Antagonists

Sun

Flanagan

Illing

et al. 2001

Journal of Biological Chemistry

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Mammalian receptors for gonadotropin-releasing hormone (GnRH) have over 85% sequence homology and similar ligand selectivity. Biological studies indicated that the chicken GnRH receptor has a distinct pharmacology, and certain antagonists of mammalian GnRH receptors function as agonists. To explore the structural determinants of this, we have cloned a chicken pituitary GnRH receptor and demonstrated that it has marked differences in primary amino acid sequence (59% homology) and in its interactions with GnRH analogs. The chicken GnRH receptor had high affinity for mammalian GnRH (K i 4.1 ؎ 1.2 nM) , similar to the human receptor (K i 4.8 ؎ 1.2 nM). But, in contrast to the human receptor, it also had high affinity for chicken GnRH or D-isopropyl-Lys 6 moieties, functioned as pure antagonists in the human receptor but were full or partial agonists in the chicken receptor. This suggests that the Lys side chain interacts with functional groups of the chicken GnRH receptor to stabilize it in the active conformation and that these groups are not available in the activated human GnRH receptor. Substitution of the human receptor extracellular loop two with the chicken extracellular loop two identified this domain as capable of conferring agonist activity to mammalian antagonists. Although functioning of antagonists as agonists has been shown to be species-dependent for several GPCRs, the dependence of this on an extracellular domain has not been described.

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G‐protein activation by putative antagonists at mutant Thr³⁷³Lys α_2A adrenergic receptors

Cited by 36 publications

References 31 publications

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

Inverse Agonist Activity at the α_2A-Adrenergic Receptor

A Chicken Gonadotropin-releasing Hormone Receptor That Confers Agonist Activity to Mammalian Antagonists

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G‐protein activation by putative antagonists at mutant Thr373Lys α2A adrenergic receptors

Cited by 36 publications

References 31 publications

RX 821002 as a Tool for Physiological Investigation of α2‐Adrenoceptors

RX 821002 as a Tool for Physiological Investigation of α2‐Adrenoceptors

Inverse Agonist Activity at the α2A-Adrenergic Receptor

A Chicken Gonadotropin-releasing Hormone Receptor That Confers Agonist Activity to Mammalian Antagonists

Contact Info

Product

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G‐protein activation by putative antagonists at mutant Thr³⁷³Lys α_2A adrenergic receptors

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

RX 821002 as a Tool for Physiological Investigation of α₂‐Adrenoceptors

Inverse Agonist Activity at the α_2A-Adrenergic Receptor