Adrenaline (ADR) and noradrenaline (NA) can simultaneously activate inhibitory α2‐ and stimulatory β‐adrenoceptors (AR). However, ADR and NA differ significantly in that ADR is a potent β2‐AR agonist while NA is not. Only recently has the interaction resulting from the simultaneous activation of α2‐ and β2‐AR been examined at the cellular level to determine the mechanisms of α2‐AR regulation following concomitant activation of both α2‐ and β2‐ARs by chronic ADR. This study evaluates β2‐AR regulation of α2A‐AR signalling following chronic ADR (300 nM) and NA (1 and 30 μM) treatments of BE(2)‐C human neuroblastoma cells that natively express both β2‐ and α2A‐ARs. Chronic (24 h) treatment with ADR (300 nM) desensitized the response to the α2A‐AR agonist, brimonidine, in BE(2)‐C cells. Addition of the β‐AR antagonist, propranolol, blocked the ADR‐induced α2A‐AR desensitization. Unlike ADR, chronic NA (1 μM) treatment had no effect on the α2A‐AR response. However if NA was increased to 30 μM for 24 h, α2A‐AR desensitization was observed; this desensitization was partially reversed by propranolol. Chronic ADR (300 nM) treatment reduced α2A‐AR binding levels, contributing to the α2A‐AR desensitization. This decrease was prevented by addition of propranolol during ADR treatment. Chronic NA (30 μM), like ADR, treatment lowered specific binding, whereas 1 μM NA treatment was without effect. Chronic ADR treatment produced a significant increase in GRK3 levels and this was blocked by propranolol or GRK2/3 antisense DNA treatment. This antisense DNA, common to both GRK2 and GRK3, also blocked chronic ADR‐induced α2A‐AR desensitization and down‐regulation. Acute (1 h) ADR (300 nM) or NA treatment (1 μM) produced α2A‐AR desensitization. The desensitization produced by acute treatment was β‐AR independent, as it was not blocked by propranolol. We conclude that chronic treatment with modest levels of ADR produces α2A‐AR desensitization by mechanisms that involve up‐regulation of GRK3 and down‐regulation of α2A‐AR levels through interactions with the β2‐AR. British Journal of Pharmacology (2003) 138, 921–931. doi:
Serum prolactin concentrations and dopamine turnover in the striatum and median eminence were studied in male rats after the administration of estradiol benzoate. The alpha-methyltyrosine-induced reduction of dopamine concentrations in these brain regions was used to evaluate relative rates of turnover. Steady state dopamine concentrations in the median eminence and striatum were not altered by 1, 3 or 5 days of estradiol treatment. However, 3 or 5 days of estradiol administration enhanced dopamine turnover in the median eminence but not in the striatum. Estradiol treatment failed to alter dopamine turnover in the median eminence of hypophysectomized rats. Estradiol increased serum prolactin concentrations at all of the times examined. Although alpha-methyltyrosine also increased serum prolactin, this increase was further enhanced in estradiol-treated rats. The increased prolactin response to alpha-methyltyrosine and increased dopamine turnover in the median eminence of estradiol-treated rats suggests that tuberoinfundibular dopaminergic neurons may be part of a hormonal-neuronal negative feedback loop which functions to regulate prolactin secretion.
We recently reported that ␣ 2A -adrenoceptor (AR) desensitization and down-regulation occurs after 24-h treatment with epinephrine (EPI) (0.3 M) in BE(2)-C cells that express both ␣ 2 -and  2 -ARs. The same concentration of norepinephrine (NE) has no effect. The effect of EPI is prevented by  2 -AR blockade and is associated with an increase in G protein-coupled receptor kinase 3 (GRK3) expression. Because differences in agonist-induced down-regulation of the ␣ 2A -versus ␣ 2B -ARs have been reported, the present study examines the effects of simultaneous activation of ␣ 2B -and  2 -ARs on ␣ 2B -AR number and signaling. We studied NG108 cells that naturally express ␣ 2B -ARs, and BN17 cells, NG108 cells transfected to express the human  2 -AR. In NG108 cells, ␣ 2B -AR desensitization and down-regulation require treatment with 20 M EPI or NE; GRK expression was not changed. In BN17 cells expressing  2 -ARs, the threshold EPI concentration for ␣ 2B -AR desensitization and down-regulation was reduced to 0.3 M; 10 M NE was required for the same effect. Furthermore, 24-h EPI or NE treatments that produced desensitization also resulted in a selective 2-fold up-regulation of GRK3; GRK2 was unchanged. The -AR antagonist alprenolol (1 M) and GRK3 antisense (but not sense) DNA blocked 0.3 M EPI-and 10 M NE-induced desensitization and down-regulation of the ␣ 2B -AR as well as GRK3 up-regulation. In conclusion, simultaneous activation of ␣ 2B -and  2 -ARs results in a 67-fold decrease in the threshold concentration of EPI required for ␣ 2B -AR down-regulation. This lower threshold for down-regulation is associated with ␣ 2B -and  2 -AR dependent up-regulation of GRK3 expression.Over the last decade, considerable attention has been given to the short-and long-term regulation of ␣ 2 -adrenoceptor signaling. Primarily using transfected receptors in cell lines that do not endogenously express the ␣ 2 -adrenoceptor, Liggett and coworkers have conducted a series of experiments examining the short-term and long-term desensitization of the three ␣ 2 -adrenoceptor subtypes (␣ 2A/D,2B and 2C ) (Liggett, 1998). Through these studies, structural components of the ␣ 2 -adrenoceptor have been identified as sites for GRK phosphorylation that are important for the short-term desensitization of these receptors. For example, four serines in the third intracellular loop are reported to be critical for shortterm desensitization of the ␣ 2A -adrenoceptor (Eason et al., 1995). However, there are structural differences within the ␣ 2 -adrenoceptor family that may lead to heterogeneity in the regulation of the three ␣ 2 -subtypes (Eason and Liggett, 1992). For example, significantly higher agonist concentrations are reportedly required for desensitization of ␣ 2A compared with ␣ 2B -adrenoceptors (Heck and Bylund, 1997). Moreover, studies have suggested that phosphorylation sites critical to short-term ␣ 2 -adrenoceptor desensitization are not important for long-term desensitization and down-regulation (Jewell-Motz and Liggett, 19...
Exposure of rats to unpredictable, inescapable stress results in two distinct behaviors during subsequent escape testing. One behavior, suggestive of lack of stress resilience, is prolonged escape latency compared to non-stressed rats and is labeled learned helplessness (LH). The other behavior suggestive of stress resilience is normal escape latency and is labeled non-helpless (NH). This study examines the effects of unpredictable, inescapable tail-shock stress (TSS) on alpha2-adrenoceptor (α2-AR) and corticotropin-releasing factor 1 receptor (CRF1) regulation as well as protein levels of G protein-coupled receptor kinase 3 (GRK3), GRK2, tyrosine hydroxylase (TH) plus carbonylated protein levels in locus coeruleus (LC), amygdala (AMG), cortex (COR) and striatum (STR). In NH rats, α2-AR and CRF1 receptors were significantly down-regulated in LC after TSS. No changes in these receptor levels were observed in the LC of LH rats. GRK3, which phosphorylates receptors and thereby contributes to α2-AR and CRF1 receptor down-regulation, was reduced in the LC of LH but not NH rats. GRK2 levels were unchanged. In AMG, GRK3 but not GRK2 levels were reduced in LH but not NH rats, and receptor regulation was impaired in LH rats. In STR, no changes in GRK3 or GRK2 levels were observed. Finally, protein carbonylation, an index of oxidative stress, was increased in the LC and AMG of LH but not NH rats. We suggest that reduced stress resilience after TSS may be related to oxidative stress, depletion of GRK3 and impaired regulation of α2-AR and CRF1 receptor in LC.
Chronic coactivation of ␣ 2B -and  2 -adrenoceptors (AR) was recently reported to down-regulate the ␣ 2B -AR at a lower threshold epinephrine (EPI) concentration compared with the activation of ␣ 2B -AR alone. This is the result of a modest  2 -AR-dependent up-regulation of G protein-coupled receptor kinase 3 (GRK3). In the present study, we determined that increasing GRK2 or GRK3 levels, independent of  2 -AR activation, decreases the EC 50 concentration for agonist-induced down-regulation of the ␣ 2B -AR using NG108 cells with or without overexpression (2-to 10-fold) of GRK2 or GRK3. In parental NG108 cells, the EC 50 concentration of EPI required for downregulation of the ␣ 2B -AR is 30 M. A 2-to 3-fold overexpression of GRK3 in NG108 cells, however, reduces the EC 50 to 0.2 M (a 150-fold decrease), whereas a comparable overexpression of GRK2 reduces it to 1 M (a 30-fold decrease). However, when GRK3 or GRK2 in NG108 cells are overexpressed 8-to 10-fold, the EC 50 concentration (0.02 M EPI) for ␣ 2B -AR down-regulation is reduced 1000-fold. These data clearly suggest that a modest (2-to 3-fold) up-regulation of GRK3 is more effective at enhancing the sensitivity of ␣ 2B -AR to down-regulation after exposure to EPI than a modest up-regulation of GRK2, but that both GRK2 and GRK3 are equally effective at inducing ␣ 2B -AR down-regulation when up-regulated 8-to 10-fold. To our knowledge, this is the first report to systematically demonstrate that GRKs, particularly GRK3, play a pivotal role in modulating the agonist EC 50 concentration that down-regulates the ␣ 2B -AR and thus adds a new dimension to an already intricate signaling network.
G-protein coupled receptor kinase 3 (GRK3) mediates desensitization of alpha 2 -adrenergic (a 2 -AR) and CRF 1 receptors. CRF 1 receptors, a 2 -AR and GRK3, are localized to the primary source of noradrenergic inputs to higher brain centers critical in both the response to stress and the development of depression, namely, locus coeruleus (LC). This study utilizing CATH.a cells (derived from the LC), demonstrates for the first time, that the stress hormone, CRF selectively up-regulates GRK3 expression via an ERK1/2-mediated mechanism accompanied by the activation of Sp-1 and Ap-2 transcription factors. This observation has important implications for the regulation of stress signaling in the brain.
The nomenclature of the Adrenoceptors has been agreed by the NC-IUPHAR Subcommittee on Adrenoceptors [58], see also [180]. Adrenoceptors, α1α1-Adrenoceptors are activated by the endogenous agonists (-)-adrenaline and (-)-noradrenaline. phenylephrine, methoxamine and cirazoline are agonists and prazosin and cirazoline antagonists considered selective for α1- relative to α2-adrenoceptors. [3H]prazosin and [125I]HEAT (BE2254) are relatively selective radioligands. S(+)-niguldipine also has high affinity for L-type Ca2+ channels. Fluorescent derivatives of prazosin (Bodipy PLprazosin- QAPB) are used to examine cellular localisation of α1-adrenoceptors. Selective α1-adrenoceptor agonists are used as nasal decongestants; antagonists to treat hypertension (doxazosin, prazosin) and benign prostatic hyperplasia (alfuzosin, tamsulosin). The α1- and β2-adrenoceptor antagonist carvedilol is used to treat congestive heart failure, although the contribution of α1-adrenoceptor blockade to the therapeutic effect is unclear. Several anti-depressants and anti-psychotic drugs are α1-adrenoceptor antagonists contributing to side effects such as orthostatic hypotension and extrapyramidal effects.Adrenoceptors, α2 α2-Adrenoceptors are activated by (-)-adrenaline and with lower potency by (-)-noradrenaline. brimonidine and talipexole are agonists and rauwolscine and yohimbine antagonists selective for α2- relative to α1-adrenoceptors. [3H]rauwolscine, [3H]brimonidine and [3H]RX821002 are relatively selective radioligands. There is species variation in the pharmacology of the α2A-adrenoceptor. Multiple mutations of α2-adrenoceptors have been described, some associated with alterations in function. Presynaptic α2-adrenoceptors regulate many functions in the nervous system. The α2-adrenoceptor agonists clonidine, guanabenz and brimonidine affect central baroreflex control (hypotension and bradycardia), induce hypnotic effects and analgesia, and modulate seizure activity and platelet aggregation. clonidine is an anti-hypertensive and counteracts opioid withdrawal. dexmedetomidine (also xylazine) is used as a sedative and analgesic in human and veterinary medicine with sympatholytic and anxiolytic properties. The α2-adrenoceptor antagonist yohimbine has been used to treat erectile dysfunction and mirtazapine as an anti-depressant. The α2B subtype appears to be involved in neurotransmission in the spinal cord and α2C in regulating catecholamine release from adrenal chromaffin cells.Adrenoceptors, ββ-Adrenoceptors are activated by the endogenous agonists (-)-adrenaline and (-)-noradrenaline. Isoprenaline is selective for β-adrenoceptors relative to α1- and α2-adrenoceptors, while propranolol (pKi 8.2-9.2) and cyanopindolol (pKi 10.0-11.0) are relatively β1 and β2 adrenoceptor-selective antagonists. (-)-noradrenaline, xamoterol and (-)-Ro 363 show selectivity for β1- relative to β2-adrenoceptors. Pharmacological differences exist between human and mouse β3-adrenoceptors, and the 'rodent selective' agonists BRL 37344 and CL316243 have low efficacy at the human β3-adrenoceptor whereas CGP 12177 and L 755507 activate human β3-adrenoceptors [88]. β3-Adrenoceptors are resistant to blockade by propranolol, but can be blocked by high concentrations of bupranolol. SR59230A has reasonably high affinity at β3-adrenoceptors, but does not discriminate well between the three β- subtypes whereas L 755507 is more selective. [125I]-cyanopindolol, [125I]-hydroxy benzylpindolol and [3H]-alprenolol are high affinity radioligands that label β1- and β2- adrenoceptors and β3-adrenoceptors can be labelled with higher concentrations (nM) of [125I]-cyanopindolol together with β1- and β2-adrenoceptor antagonists. [3H]-L-748337 is a β3-selective radioligand [474]. Fluorescent ligands such as BODIPY-TMR-CGP12177 can be used to track β-adrenoceptors at the cellular level [8]. Somewhat selective β1-adrenoceptor agonists (denopamine, dobutamine) are used short term to treat cardiogenic shock but, chronically, reduce survival. β1-Adrenoceptor-preferring antagonists are used to treat hypertension (atenolol, betaxolol, bisoprolol, metoprolol and nebivolol), cardiac arrhythmias (atenolol, bisoprolol, esmolol) and cardiac failure (metoprolol, nebivolol). Cardiac failure is also treated with carvedilol that blocks β1- and β2-adrenoceptors, as well as α1-adrenoceptors. Short (salbutamol, terbutaline) and long (formoterol, salmeterol) acting β2-adrenoceptor-selective agonists are powerful bronchodilators used to treat respiratory disorders. Many first generation β-adrenoceptor antagonists (propranolol) block both β1- and β2-adrenoceptors and there are no β2-adrenoceptor-selective antagonists used therapeutically. The β3-adrenoceptor agonist mirabegron is used to control overactive bladder syndrome.
Cellular levels of G protein-coupled receptor kinase (GRK)3 determine the sensitivity of the ␣ 2A/B -adrenoceptor (␣ 2 -AR) to agonist-induced down-regulation. Using human neuroblastoma BE(2)-C cells, this study examines how cellular GRK3 levels are affected by several mechanisms reported to influence stability and degradation of other GRKs. We first examined the interaction between the 90-kDa heat shock protein (Hsp90) and GRK3; Hsp90 reportedly affects the maturation and stability of GRK2. In unstimulated cells, GRK3 coimmunoprecipitates with Hsp90, suggesting a physical interaction. Moreover, when GRK3 protein expression was increased by 24-h epinephrine (EPI) treatment, Hsp90 protein expression increased with a similar but slightly delayed time course. To investigate the influence of Hsp90 on GRK3 protein stability, we determined the effect of the Hsp90 inhibitor geldanamycin (GA) on cellular GRK3 levels. GA eliminated the interaction between Hsp90 with GRK3 and produced a rapid, proteasome-mediated, 70% decrease in GRK3 levels within 24 h. To investigate the influence of Hsp90 on up-regulation of GRK3 expression, we examined the effect of GA on EPI-induced up-regulation. GA reduced the absolute increase in GRK3; however, the percentage of increase in GRK3 by EPI was not significantly different in the absence versus presence of GA (141 Ϯ 41 versus 94 Ϯ 12%). Finally, we examined the influence of Ca 2ϩ -activated proteases on cellular GRK3. Treatment with the calcium ionophore ionomycin produced a rapid decrease in GRK3 levels that was inhibited by the calpain inhibitor calpeptin. In conclusion, several mechanisms influence the degradation of GRK3 and therefore have the potential to affect GPCR signaling by regulating GRK3 levels in neurons.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.