RESUMEN
Drugs that act on α-adrenoceptors may contain morpholine and pyrimidinone heterocycles. The aim of this study was to synthesize a series of pyrimidinones (S6a-e and S8) and characterize their α-adrenoceptor activity. Cytotoxicity assays (MTT and LDH) were performed in A7r5 and HUVECs. Concentration-effect curves to phenylephrine (Phe) were performed in rat aortic rings in the presence of compounds S6a-e and S8 or vehicle. Nitric oxide (NO) production and NO stable metabolic products, nitrite and nitrate, expressed as total nitrogen oxides (NOx) were assessed in HUVECs by confocal microscopy with the DAF-2DA probe and by the Griess reaction, respectively. Molecular docking simulations were performed using the 6a compound and α2A-adrenoceptor. In the evaluated conditions, the percentage of viable cells and the release of LDH were similar between control cells and cells exposed to the tested pyrimidinones. S6d, S6e, S8, and the positive control prazosin (but not S6a, S6b, and S6c) decreased Phe-induced contractions in endothelium-denuded aortic rings. S6a, S6b, and S6c decreased Phe-induced contractions in endothelium-intact aortic rings. The effect of S6a was abolished by L-NAME. NO production and NOx levels were inhibited in the presence of the α2 receptor antagonist yohimbine and the NOS inhibitor L-NAME. The 6a docking simulation estimated that the mean binding free energy of the compound was lower than the estimated value for yohimbine. These data suggest that S6d, S6e, and S8 may be α1-adrenoceptor antagonists while S6a acts as an agonist of α2-adrenoceptors.
Asunto(s)
Células Endoteliales de la Vena Umbilical Humana , Simulación del Acoplamiento Molecular , Morfolinas , Pirimidinonas , Animales , Humanos , Ratas , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Pirimidinonas/farmacología , Pirimidinonas/química , Morfolinas/farmacología , Morfolinas/química , Óxido Nítrico/metabolismo , Masculino , Receptores Adrenérgicos alfa 2/metabolismo , Línea Celular , Aorta/efectos de los fármacos , Aorta/citología , Aorta/metabolismo , Ratas WistarRESUMEN
Epinephrine influences the function of pancreatic ß-cells, primarily through the α2A-adrenergic receptor (α2A-AR) on their plasma membrane. Previous studies indicate that epinephrine transiently suppresses insulin secretion, whereas prolonged exposure induces its compensatory secretion. Nonetheless, the impact of epinephrine-induced α2A-AR signaling on the survival and function of pancreatic ß-cells, particularly the impact of reprogramming after their removal from sustained epinephrine stimulation, remains elusive. In the present study, we applied MIN6, a murine insulinoma cell line, with 3 days of high concentration epinephrine incubation and 2 days of standard incubation, explored cell function and activity, and analyzed relevant regulatory pathways. The results showed that chronic epinephrine incubation led to the desensitization of α2A-AR and enhanced insulin secretion. An increased number of docked insulin granules and impaired Syntaxin-2 was found after chronic epinephrine exposure. Growth curve and cell cycle analyses showed the inhibition of cell proliferation. Transcriptome analysis showed the occurrence of endoplasmic reticulum stress (ER stress) and oxidative stress, such as the presence of BiP, CHOP, IRE1, ATF4, and XBP, affecting cellular endoplasmic reticulum function and survival, along with UCP2, OPA1, PINK, and PRKN, associated with mitochondrial dysfunction. Consequently, we conclude that chronic exposure to epinephrine induces α2A-AR desensitization and leads to ER and oxidative stress, impairing protein processing and mitochondrial function, leading to modified pancreatic ß-cell secretory function and cell fate.
Asunto(s)
Estrés del Retículo Endoplásmico , Epinefrina , Células Secretoras de Insulina , Insulina , Estrés Oxidativo , Animales , Epinefrina/farmacología , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Ratones , Estrés del Retículo Endoplásmico/efectos de los fármacos , Insulina/metabolismo , Secreción de Insulina/efectos de los fármacos , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/genética , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/efectos de los fármacosRESUMEN
Most α2-AR agonists derived from dexmedetomidine have few structural differences between them and have no selectivity for α2A/2B-AR or Gi/Gs, which can lead to side effects in drugs. To obtain novel and potent α2A-AR agonists, we performed virtual screening for human α2A-AR and α2B-AR to find α2A-AR agonists with higher selectivity. Compound P300-2342 and its three analogs significantly decreased the locomotor activity of mice (p < 0.05). Furthermore, P300-2342 and its three analogs inhibited the binding of [3H] Rauwolscine with IC50 values of 7.72 ± 0.76 and 12.23 ± 0.11 µM, respectively, to α2A-AR and α2B-AR. In α2A-AR-HEK293 cells, P300-2342 decreased forskolin-stimulated cAMP production without increasing cAMP production, which indicated that P300-2342 activated α2A-AR with coupling to the Gαi/o pathway but without Gαs coupling. P300-2342 exhibited no agonist but slight antagonist activities in α2B-AR. Similar results were obtained for the analogs of P300-2342. The docking results showed that P300-2342 formed π-hydrogen bonds with Y394, V114 in α2A-AR, and V93 in α2B-AR. Three analogs of P300-2342 formed several π-hydrogen bonds with V114, Y196, F390 in α2A-AR, and V93 in α2B-AR. We believe that these molecules can serve as leads for the further optimization of α2A-AR agonists with potentially few side effects.
Asunto(s)
Agonistas de Receptores Adrenérgicos alfa 2 , Simulación del Acoplamiento Molecular , Receptores Adrenérgicos alfa 2 , Humanos , Animales , Células HEK293 , Receptores Adrenérgicos alfa 2/metabolismo , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Ratones , Subunidades alfa de la Proteína de Unión al GTP Gi-Go/metabolismo , AMP Cíclico/metabolismo , Masculino , Unión ProteicaRESUMEN
The present study investigated the impact of central α2-adrenergic mechanisms on sympathetic action potential (AP) discharge, recruitment and latency strategies. We used the microneurographic technique to record muscle sympathetic nerve activity and a continuous wavelet transform to investigate postganglionic sympathetic AP firing during a baseline condition and an infusion of a α2-adrenergic receptor agonist, dexmedetomidine (10 min loading infusion of 0.225 µg kg-1; maintenance infusion of 0.1-0.5 µg kg h-1) in eight healthy individuals (28 ± 7 years, five females). Dexmedetomidine reduced mean pressure (92 ± 7 to 80 ± 8 mmHg, P < 0.001) but did not alter heart rate (61 ± 13 to 60 ± 14 bpm; P = 0.748). Dexmedetomidine reduced sympathetic AP discharge (126 ± 73 to 27 ± 24 AP 100 beats-1, P = 0.003) most strongly for medium-sized APs (normalized cluster 2: 21 ± 10 to 5 ± 5 AP 100 beats-1; P < 0.001). Dexmedetomidine progressively de-recruited sympathetic APs beginning with the largest AP clusters (12 ± 3 to 7 ± 2 clusters, P = 0.002). Despite de-recruiting large AP clusters with shorter latencies, dexmedetomidine reduced AP latency across remaining clusters (1.18 ± 0.12 to 1.13 ± 0.13 s, P = 0.002). A subset of six participants performed a Valsalva manoeuvre (20 s, 40 mmHg) during baseline and the dexmedetomidine infusion. Compared to baseline, AP discharge (Δ 361 ± 292 to Δ 113 ± 155 AP 100 beats-1, P = 0.011) and AP cluster recruitment elicited by the Valsalva manoeuvre were lower during dexmedetomidine (Δ 2 ± 1 to Δ 0 ± 2 AP clusters, P = 0.041). The reduction in sympathetic AP latency elicited by the Valsalva manoeuvre was not affected by dexmedetomidine (Δ -0.09 ± 0.07 to Δ -0.07 ± 0.14 s, P = 0.606). Dexmedetomidine reduced baroreflex gain, most strongly for medium-sized APs (normalized cluster 2: -6.0 ± 5 to -1.6 ± 2 % mmHg-1; P = 0.008). These data suggest that α2-adrenergic mechanisms within the central nervous system modulate sympathetic postganglionic neuronal discharge, recruitment and latency strategies in humans. KEY POINTS: Sympathetic postganglionic neuronal subpopulations innervating the human circulation exhibit complex patterns of discharge, recruitment and latency. However, the central neural mechanisms governing sympathetic postganglionic discharge remain unclear. This microneurographic study investigated the impact of a dexmedetomidine infusion (α2-adrenergic receptor agonist) on muscle sympathetic postganglionic action potential (AP) discharge, recruitment and latency patterns. Dexmedetomidine infusion inhibited the recruitment of large and fast conducting sympathetic APs and attenuated the discharge of medium sized sympathetic APs that fired during resting conditions and the Valsalva manoeuvre. Dexmedetomidine infusion elicited shorter sympathetic AP latencies during resting conditions but did not affect the reductions in latency that occurred during the Valsalva manoeuvre. These data suggest that α2-adrenergic mechanisms within the central nervous system modulate sympathetic postganglionic neuronal discharge, recruitment and latency strategies in humans.
Asunto(s)
Potenciales de Acción , Agonistas de Receptores Adrenérgicos alfa 2 , Dexmedetomidina , Sistema Nervioso Simpático , Humanos , Dexmedetomidina/farmacología , Femenino , Adulto , Masculino , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Sistema Nervioso Simpático/fisiología , Sistema Nervioso Simpático/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Adulto Joven , Frecuencia Cardíaca/efectos de los fármacos , Frecuencia Cardíaca/fisiología , Presión Sanguínea/fisiología , Presión Sanguínea/efectos de los fármacos , Músculo Esquelético/fisiología , Músculo Esquelético/inervación , Músculo Esquelético/efectos de los fármacos , Receptores Adrenérgicos alfa 2/fisiología , Receptores Adrenérgicos alfa 2/metabolismoRESUMEN
Enhancing the selectivity of alpha2-adrenoceptor (α2A-AR) agonists remains an unresolved issue. Herein, we reported the design of an α2A-AR agonist using the conformation constraint method, beginning with medetomidine. The structure-activity relationship indicated that the 8-substituent of chromane derivatives exerted the most pronounced effect on α2A-AR agonistic activity. Compounds A9 and B9 were identified as the most promising, exhibiting EC50 values of 0.78 and 0.23 nM, respectively. Their selectivity indexes surpassed dexmedetomidine (DMED) by 10-80 fold. In vivo studies demonstrated that both A9 and B9 dose-dependently increased the loss of righting reflex in mice, with ED50 values of 1.54 and 0.138 mg/kg, respectively. Binding mode calculations and mutation studies suggested the indispensability of the hydrogen bond between ASP1283.32 and α2A-AR agonist. In particular, A9 and B9 showed no dual reverse pharmacological effect, a characteristic exhibited by DMED in α2A-AR activation.
Asunto(s)
Agonistas de Receptores Adrenérgicos alfa 2 , Cromanos , Diseño de Fármacos , Receptores Adrenérgicos alfa 2 , Animales , Receptores Adrenérgicos alfa 2/metabolismo , Cromanos/farmacología , Cromanos/química , Cromanos/síntesis química , Relación Estructura-Actividad , Ratones , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Agonistas de Receptores Adrenérgicos alfa 2/química , Humanos , Conformación Molecular , Simulación del Acoplamiento Molecular , MasculinoRESUMEN
Yohimbine, a natural indole alkaloid and a nonselective adrenoceptor antagonist, possesses potential benefits in treating inflammatory disorders and sepsis. Nevertheless, its broader clinical use faces challenges due to its low receptor selectivity. A structure-activity relationship study of novel yohimbine analogues identified amino esters of yohimbic acid as potent and selective ADRA2A antagonists. Specifically, amino ester 4n, in comparison to yohimbine, showed a 6-fold higher ADRA1A/ADRA2A selectivity index (SI > 556 for 4n) and a 25-fold higher ADRA2B/ADRA2A selectivity index. Compound 4n also demonstrated high plasma and microsomal stability, moderate-to-low membrane permeability determining its limited ability to cross the blood-brain barrier, and negligible toxicity on nontumor normal human dermal fibroblasts. Compound 4n represents an important complementary pharmacological tool to study the involvement of adrenoceptor subtypes in pathophysiologic conditions such as inflammation and sepsis and a novel candidate for further preclinical development to treat ADRA2A-mediated pathologies.
Asunto(s)
Antagonistas de Receptores Adrenérgicos alfa 2 , Diseño de Fármacos , Receptores Adrenérgicos alfa 2 , Yohimbina , Humanos , Receptores Adrenérgicos alfa 2/metabolismo , Yohimbina/farmacología , Yohimbina/química , Relación Estructura-Actividad , Antagonistas de Receptores Adrenérgicos alfa 2/farmacología , Antagonistas de Receptores Adrenérgicos alfa 2/química , Antagonistas de Receptores Adrenérgicos alfa 2/síntesis química , AnimalesRESUMEN
Apoptosis is the crucial pathological mechanism following cerebral ischemic injury. Our previous studies demonstrated that clonidine, one agonist of alpha2-adrenergic receptor (α2-AR), could attenuate cerebral ischemic injury in a rat model of middle cerebral artery occlusion/reperfusion (MCAO/R). However, it's unclear whether clonidine exerts neuroprotective effects by regulating neuronal apoptosis. In this study, we elucidated whether clonidine can exert anti-apoptotic effects in cerebral ischemic injury, and further explored the possible mechanisms. Neurological deficit score was measured to evaluate the neurological function. TTC staining was used for the measurement of brain infarct size. Hematoxylin-Eosin (HE) staining was applied to examine the cell morphology. TUNEL and DAPI fluorescent staining methods were used to analyze the cell apoptosis in brain tissue. Fluorescence quantitative real-time PCR was performed to assess the gene expression of Caspase-3 and P53. Western blotting assay was applied to detect the protein expression of Caspase-3 and P53. The results showed that clonidine improved neurological function, reduced brain infarct size, alleviated neuronal damage, and reduced the ratio of cell apoptosis in the brain with MCAO/R injury. moreover, clonidine down-regulated the gene and protein expression of Caspase-3 and P53 which were over-expressed after MCAO/R injury. Whereas, yohimbine (one selective α2-AR antagonist) mitigated the anti-apoptosis effects of clonidine, accompanied by reversed gene and protein expression changes. The results indicated that clonidine attenuated cerebral MCAO/R injury via suppressing neuronal apoptosis, which may be mediated, at least in part, by activating α2-AR.
Asunto(s)
Agonistas de Receptores Adrenérgicos alfa 2 , Apoptosis , Clonidina , Neuronas , Fármacos Neuroprotectores , Ratas Sprague-Dawley , Daño por Reperfusión , Animales , Clonidina/farmacología , Clonidina/uso terapéutico , Apoptosis/efectos de los fármacos , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Daño por Reperfusión/prevención & control , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Agonistas de Receptores Adrenérgicos alfa 2/uso terapéutico , Masculino , Ratas , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/prevención & control , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Caspasa 3/metabolismo , Receptores Adrenérgicos alfa 2/metabolismo , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patologíaRESUMEN
The prepositus hypoglossi nucleus (PHN) and the interstitial nucleus of Cajal (INC) are involved in the control of horizontal and vertical gaze, respectively. A previous study showed that PHN neurons exhibit depolarized or hyperpolarized responses to noradrenaline (NA). However, the adrenoceptor types that participate in NA-induced responses and the effects of NA on INC neurons have not yet been investigated. Furthermore, the relationship between NA-induced responses and neuron types defined by neurotransmitter phenotypes has not been determined. In this study, we investigated NA-induced current responses in PHN and INC neurons and the relationships between these responses and neuron types using whole cell recordings in wild-type and transgenic rat brainstem slices. Local application of NA to the cell soma induced slow inward (SI) and slow outward (SO) currents that were mainly mediated by α1 and α2 adrenoceptors, respectively. These current responses were observed in both PHN and INC neurons, although the proportion of INC neurons that responded to NA was low. Analyses of the distributions of the current responses revealed that in the PHN, all fluorescently identified inhibitory neurons exhibited SI currents, whereas glutamatergic and cholinergic neurons exhibited both SI and SO currents. In the INC, glutamatergic and inhibitory neurons preferentially exhibited SI and SO currents, respectively. When the PHN and INC neurons were characterized by their firing pattern, we found that the proportions of the currents depended on their firing pattern. These results suggest that various modes of noradrenergic modulation in horizontal and vertical neural integrators are dependent on neuron type.NEW & NOTEWORTHY Noradrenergic modulation of oculomotor neural integrators involved in gaze control has not been elucidated. Here, we report that noradrenaline (NA)-induced slow inward (SI) and outward (SO) currents are mediated mainly by α1 and α2 adrenoceptors in neurons that participate in horizontal and vertical gaze control. The NA-induced current responses differed depending on the neurotransmitter phenotype and firing pattern. These results suggest various modes of noradrenergic modulation in horizontal and vertical integrator neurons.
Asunto(s)
Norepinefrina , Animales , Norepinefrina/farmacología , Ratas , Masculino , Ratas Transgénicas , Neuronas/fisiología , Neuronas/efectos de los fármacos , Receptores Adrenérgicos alfa 1/metabolismo , Receptores Adrenérgicos alfa 1/fisiología , Neuronas Adrenérgicas/fisiología , Neuronas Adrenérgicas/efectos de los fármacos , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/fisiología , Técnicas de Placa-Clamp , Tronco Encefálico/fisiología , Tronco Encefálico/citología , Tronco Encefálico/efectos de los fármacos , Neuronas Colinérgicas/fisiología , Neuronas Colinérgicas/efectos de los fármacosRESUMEN
Catecholamines binding to α- and ß-adrenergic receptors on immune cells have recently been shown to play an important role in regulating immune responses. Although α2-adrenergic receptors are known to modulate the immune response in different ways, the therapeutic exploration of their utility has been limited by the lack of agonists selective for the three α2-adrenergic subtypes. We report in this study the identification of the agonist AGN-762, which activates α2B- and α2C-adrenergic subtypes, but not the α2A subtype. We show that AGN-762 reduced clinical disease in an experimental autoimmune encephalitis model of autoimmune disease via direct or indirect effects on T regulatory cells. The activity of AGN-762 was abrogated by depletion of T regulatory cells, which express the α2B-adrenergic receptor. Furthermore, a drug-induced shift to an anti-inflammatory phenotype was demonstrated in immune cells in the spleen of drug-treated experimental autoimmune encephalitis mice. AGN-762 does not display sedative and cardiovascular side effects associated with α2A subtype agonists. Immune modulation by selective α2-adrenergic agonists represents a novel, to our knowledge, approach for treating autoimmune disease.
Asunto(s)
Agonistas de Receptores Adrenérgicos alfa 2 , Autoinmunidad , Receptores Adrenérgicos alfa 2 , Animales , Ratones , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/inmunología , Autoinmunidad/inmunología , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Agonistas de Receptores Adrenérgicos alfa 2/uso terapéutico , Ratones Endogámicos C57BL , Linfocitos T Reguladores/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Humanos , Femenino , Modelos Animales de EnfermedadRESUMEN
Inflammatory bowel disease (IBD) is characterized by dysbiosis of the gut microbiota and dysfunction of intestinal stem cells (ISCs). However, the direct interactions between IBD microbial factors and ISCs are undescribed. Here, we identify α2A-adrenergic receptor (ADRA2A) as a highly expressed GPCR in ISCs. Through PRESTO-Tango screening, we demonstrate that tyramine, primarily produced by Enterococcus via tyrosine decarboxylase (tyrDC), serves as a microbial ligand for ADRA2A. Using an engineered tyrDC-deficient Enterococcus faecalis strain and intestinal epithelial cell-specific Adra2a knockout mice, we show that Enterococcus-derived tyramine suppresses ISC proliferation, thereby impairing epithelial regeneration and exacerbating DSS-induced colitis through ADRA2A. Importantly, blocking the axis with an ADRA2A antagonist, yohimbine, disrupts tyramine-mediated suppression on ISCs and alleviates colitis. Our findings highlight a microbial ligand-GPCR pair in ISCs, revealing a causal link between microbial regulation of ISCs and colitis exacerbation and yielding a targeted therapeutic approach to restore ISC function in colitis.
Asunto(s)
Colitis , Ratones Noqueados , Receptores Adrenérgicos alfa 2 , Células Madre , Tiramina , Animales , Tiramina/metabolismo , Tiramina/farmacología , Colitis/microbiología , Colitis/inducido químicamente , Colitis/metabolismo , Ratones , Receptores Adrenérgicos alfa 2/metabolismo , Células Madre/metabolismo , Humanos , Ratones Endogámicos C57BL , Tirosina Descarboxilasa/metabolismo , Enterococcus faecalis/metabolismo , Microbioma Gastrointestinal , Mucosa Intestinal/microbiología , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patología , Yohimbina/farmacología , Modelos Animales de Enfermedad , Enterococcus/metabolismo , Intestinos/microbiología , Intestinos/patología , Proliferación Celular , Enfermedades Inflamatorias del Intestino/microbiología , Enfermedades Inflamatorias del Intestino/metabolismo , Sulfato de DextranRESUMEN
Sympathetic nervous system (SNS) hyperactivity is mediated by elevated catecholamine (CA) secretion from the adrenal medulla, as well as enhanced norepinephrine (NE) release from peripheral sympathetic nerve terminals. Adrenal CA production from chromaffin cells is tightly regulated by sympatho-inhibitory α2-adrenergic (auto)receptors (ARs), which inhibit both epinephrine (Epi) and NE secretion via coupling to Gi/o proteins. α2-AR function is, in turn, regulated by G protein-coupled receptor (GPCR)-kinases (GRKs), especially GRK2, which phosphorylate and desensitize them, i.e., uncouple them from G proteins. On the other hand, the short-chain free fatty acid (SCFA) receptor (FFAR)-3, also known as GPR41, promotes NE release from sympathetic neurons via the Gi/o-derived free Gßγ-activated phospholipase C (PLC)-ß/Ca2+ signaling pathway. However, whether it exerts a similar effect in adrenal chromaffin cells is not known at present. In the present study, we examined the interplay of the sympatho-inhibitory α2A-AR and the sympatho-stimulatory FFAR3 in the regulation of CA secretion from rat adrenal chromaffin (pheochromocytoma) PC12 cells. We show that FFAR3 promotes CA secretion, similarly to what GRK2-dependent α2A-AR desensitization does. In addition, FFAR3 activation enhances the effect of the physiologic stimulus (acetylcholine) on CA secretion. Importantly, GRK2 blockade to restore α2A-AR function or the ketone body beta-hydroxybutyrate (BHB or 3-hydroxybutyrate), via FFAR3 antagonism, partially suppress CA production, when applied individually. When combined, however, CA secretion from PC12 cells is profoundly suppressed. Finally, propionate-activated FFAR3 induces leptin and adiponectin secretion from PC12 cells, two important adipokines known to be involved in tissue inflammation, and this effect of FFAR3 is fully blocked by the ketone BHB. In conclusion, SCFAs can promote CA and adipokine secretion from adrenal chromaffin cells via FFAR3 activation, but the metabolite/ketone body BHB can effectively inhibit this action.
Asunto(s)
Catecolaminas , Receptores Adrenérgicos alfa 2 , Receptores Acoplados a Proteínas G , Animales , Células PC12 , Ratas , Receptores Acoplados a Proteínas G/metabolismo , Catecolaminas/metabolismo , Receptores Adrenérgicos alfa 2/metabolismo , Adipoquinas/metabolismo , Células Cromafines/metabolismo , Transducción de Señal , Norepinefrina/metabolismo , Norepinefrina/farmacologíaRESUMEN
BACKGROUND: Acute kidney injury (AKI) was reported to be one of the initiators of chronic kidney disease (CKD) development. Necroinflammation may contribute to the progression from AKI to CKD. Dexmedetomidine (Dex), a highly selective α2-adrenoreceptor (AR) agonist, has cytoprotective and "anti-" inflammation effects. This study was designed to investigate the anti-fibrotic properties of Dex in sepsis models. METHODS: C57BL/6 mice were randomly treated with an i.p. injection of lipopolysaccharides (LPS) (10â¯mg/kg) alone, LPS with Dex (25⯵g/kg), or LPS, Dex and Atipamezole (Atip, an α2-adrenoreceptor antagonist) (500⯵g/kg) (n=5/group). Human proximal tubular epithelial cells (HK2) were also cultured and then exposed to LPS (1⯵g/ml) alone, LPS and Dex (1⯵M), transforming growth factor-beta 1 (TGF-ß1) (5â¯ng/ml) alone, TGF-ß1 and Dex, with or without Atip (100⯵M) in culture media. Epithelial-mesenchymal transition (EMT), cell necrosis, necroptosis and pyroptosis, and c-Jun N-terminal kinase (JNK) phosphorylation were then determined. RESULTS: Dex treatment significantly alleviated LPS-induced AKI, myofibroblast activation, NLRP3 inflammasome activation, and necroptosis in mice. Atip counteracted its protective effects. Dex attenuated LPS or TGF-ß1 induced EMT and also prevented necrosis, necroptosis, and pyroptosis in response to LPS stimulation in the HK2 cells. The anti-EMT effects of Dex were associated with JNK phosphorylation. CONCLUSIONS: Dex reduced EMT following LPS stimulation whilst simultaneously inhibiting pyroptosis and necroptosis via α2-AR activation in the renal tubular cells. The "anti-fibrotic" and cytoprotective properties and its clinical use of Dex need to be further studied.
Asunto(s)
Agonistas de Receptores Adrenérgicos alfa 2 , Dexmedetomidina , Fibrosis , Ratones Endogámicos C57BL , Receptores Adrenérgicos alfa 2 , Animales , Humanos , Ratones , Lesión Renal Aguda/tratamiento farmacológico , Lesión Renal Aguda/patología , Lesión Renal Aguda/metabolismo , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/prevención & control , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Agonistas de Receptores Adrenérgicos alfa 2/uso terapéutico , Antagonistas de Receptores Adrenérgicos alfa 2/farmacología , Línea Celular , Dexmedetomidina/farmacología , Transición Epitelial-Mesenquimal/efectos de los fármacos , Inflamación/tratamiento farmacológico , Inflamación/patología , Inflamación/metabolismo , Riñón/patología , Riñón/efectos de los fármacos , Riñón/metabolismo , Lipopolisacáridos/farmacología , Necroptosis/efectos de los fármacos , Fenotipo , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Receptores Adrenérgicos alfa 2/metabolismoRESUMEN
Prenatal opioid exposure (POE) and postnatal adverse experiences are early life adversities (ELA) that often co-occur and increase problematic alcohol (EtOH) drinking during adolescence. We investigated the relationship between POE, postnatal adversity, and adolescent EtOH drinking in rats. We also sought to determine whether ELAs affect alpha-adrenoceptor density in the brain because the noradrenergic system is involved in problematic alcohol drinking and its treatment. We hypothesized that the combination of POE and postnatal adversity will increase alcohol drinking in rats compared to rats with exposure to either adversity alone or to control. We also predicted that POE and postnatal adversity would increase α1-adrenoceptor density and decrease α2-adrenoceptor density in brain to confer a stress-responsive phenotype. Pregnant rats received morphine (15 mg/kg/day) or saline via subcutaneous minipumps from gestational day 9 until birth. Limited bedding and nesting (LBN) procedures were introduced from postnatal day (PD) 3-11 to mimic early life adversity-scarcity. Offspring rats (PD 31-33) were given opportunities to drink EtOH (20 %, v/v) using intermittent-access, two-bottle choice (with water) procedures. Rats given access to EtOH were assigned into sub-groups that were injected with either yohimbine (1 mg/kg, ip) or vehicle (2 % DMSO, ip) 30 min prior to each EtOH access session to determine the effects of α2-adrenoceptor inhibition on alcohol drinking. We harvested cortices, brainstems, and hypothalami from EtOH-naïve littermates on either PD 30 or PD 70 and conducted radioligand receptor binding assays to quantify α1- and α2-adrenoceptor densities. Contrary to our hypothesis, only LBN alone increased EtOH intake in female adolescent rats compared to female rats with POE. Neither POE nor LBN affected α1- or α2-adrenoceptor densities in the cortex, brainstem, or hypothalamus of early- or late-aged adolescent rats. These results suggest a complex interaction between ELA type and sex on alcohol drinking.
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Consumo de Bebidas Alcohólicas , Etanol , Efectos Tardíos de la Exposición Prenatal , Animales , Femenino , Ratas , Embarazo , Consumo de Bebidas Alcohólicas/metabolismo , Efectos Tardíos de la Exposición Prenatal/metabolismo , Etanol/administración & dosificación , Etanol/farmacología , Masculino , Receptores Adrenérgicos alfa 2/metabolismo , Morfina/farmacología , Encéfalo/metabolismo , Encéfalo/efectos de los fármacos , Receptores Adrenérgicos alfa 1/metabolismo , Ratas Sprague-DawleyRESUMEN
Autism spectrum disorder (ASD) is a neurodevelopmental disorder that severely affects individuals' daily life and social development. Unfortunately, there are currently no effective treatments for ASD. Dexmedetomidine (DEX) is a selective agonist of α2 adrenergic receptor (α2AR) and is widely used as a first-line medication for sedation and hypnosis in clinical practice. In recent years, there have been reports suggesting its potential positive effects on improving emotional and cognitive functions. However, whether dexmedetomidine has therapeutic effects on the core symptoms of ASD, namely social deficits and repetitive behaviors, remains to be investigated. In the present study, we employed various behavioral tests to assess the phenotypes of animals, including the three-chamber, self-grooming, marble burying, open field, and elevated plus maze. Additionally, electrophysiological recordings, western blotting, qPCR were mainly used to investigate and validate the potential mechanisms underlying the role of dexmedetomidine. We found that intraperitoneal injection of dexmedetomidine in ASD model mice-BTBR T+ Itpr3tf/J (BTBR) mice could adaptively improve their social deficits. Further, we observed a significant reduction in c-Fos positive signals and interleukin-6 (IL-6) expression level in the prelimbic cortex (PrL) of the BTBR mice treated with dexmedetomidine. Enhancing or inhibiting the action of IL-6 directly affects the social behavior of BTBR mice. Mechanistically, we have found that NF-κB p65 is a key pathway regulating IL-6 expression in the PrL region. In addition, we have confirmed that the α2AR acts as a receptor switch mediating the beneficial effects of dexmedetomidine in improving social deficits. This study provides the first evidence of the beneficial effects of dexmedetomidine on core symptoms of ASD and offers a theoretical basis and potential therapeutic approach for the clinical treatment of ASD.
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Agonistas de Receptores Adrenérgicos alfa 2 , Trastorno del Espectro Autista , Dexmedetomidina , Modelos Animales de Enfermedad , Interleucina-6 , FN-kappa B , Receptores Adrenérgicos alfa 2 , Conducta Social , Animales , Dexmedetomidina/farmacología , Ratones , Trastorno del Espectro Autista/tratamiento farmacológico , Trastorno del Espectro Autista/metabolismo , Masculino , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , FN-kappa B/metabolismo , Interleucina-6/metabolismo , Transducción de Señal/efectos de los fármacos , Ratones Endogámicos C57BL , Conducta Animal/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Inflamación/metabolismo , Inflamación/tratamiento farmacológicoRESUMEN
The α2A adrenergic receptor (α2A-AR) serves as a critical molecular target for sedatives and analgesics. However, α2A-AR ligands with an imidazole ring also interact with an imidazoline receptor as well as other proteins and lead to undesirable effects, motivating us to develop more novel scaffold α2A-AR ligands. For this purpose, we employed an ensemble-based ligand discovery strategy, integrating long-term molecular dynamics (MD) simulations and virtual screening, to identify new potential α2A-AR agonists with novel scaffold. Our results showed that compounds SY-15 and SY-17 exhibited significant biological effects in the preliminary evaluation of protein kinase A (PKA) redistribution assays. They also reduced levels of intracellular cyclic adenosine monophosphate (cAMP) in a dose-dependent manner. Upon treatment of the cells with 100 µM concentrations of SY-15 and SY-17, there was a respective decrease in the intracellular cAMP levels by 63.43% and 53.83%. Subsequent computational analysis was conducted to elucidate the binding interactions of SY-15 and SY-17 with the α2A-AR. The binding free energies of SY-15 and SY-17 calculated by MD simulations were -45.93 and -71.97 kcal/mol. MD simulations also revealed that both compounds act as bitopic agonists, occupying the orthosteric site and a novel exosite of the receptor simultaneously. Our findings of integrative computational and experimental approaches could offer the potential to enhance ligand affinity and selectivity through dual-site occupancy and provide a novel direction for the rational design of sedatives and analgesics.
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Analgésicos , Receptores Adrenérgicos alfa 2 , Ligandos , Receptores Adrenérgicos alfa 2/metabolismo , Hipnóticos y SedantesRESUMEN
In the central nervous system, noradrenaline transmission controls the degree to which we are awake, alert, and attentive. Aberrant noradrenaline transmission is associated with pathological forms of hyper- and hypo-arousal that present in numerous neuropsychiatric disorders often associated with dysfunction in serotonin transmission. In vivo, noradrenaline regulates the release of serotonin because noradrenergic input drives the serotonin neurons to fire action potentials via activation of excitatory α1-adrenergic receptors (α1-AR). Despite the critical influence of noradrenaline on the activity of dorsal raphe serotonin neurons, the source of noradrenergic afferents has not been resolved and the presynaptic mechanisms that regulate noradrenaline-dependent synaptic transmission have not been described. Using an acute brain slice preparation from male and female mice and electrophysiological recordings from dorsal raphe serotonin neurons, we found that selective optogenetic activation of locus coeruleus terminals in the dorsal raphe was sufficient to produce an α1-AR-mediated excitatory postsynaptic current (α1-AR-EPSC). Activation of inhibitory α2-adrenergic receptors (α2-AR) with UK-14,304 eliminated the α1-AR-EPSC via presynaptic inhibition of noradrenaline release, likely via inhibition of voltage-gated calcium channels. In a subset of serotonin neurons, activation of postsynaptic α2-AR produced an outward current through activation of GIRK potassium conductance. Further, in vivo activation of α2-AR by systemic administration of clonidine reduced the expression of c-fos in the dorsal raphe serotonin neurons, indicating reduced neural activity. Thus, α2-AR are critical regulators of serotonin neuron excitability.
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Núcleo Dorsal del Rafe , Locus Coeruleus , Receptores Adrenérgicos alfa 2 , Neuronas Serotoninérgicas , Transmisión Sináptica , Animales , Núcleo Dorsal del Rafe/efectos de los fármacos , Núcleo Dorsal del Rafe/fisiología , Núcleo Dorsal del Rafe/metabolismo , Masculino , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/fisiología , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Locus Coeruleus/efectos de los fármacos , Locus Coeruleus/fisiología , Femenino , Neuronas Serotoninérgicas/efectos de los fármacos , Neuronas Serotoninérgicas/fisiología , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Ratones , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Optogenética , Agonistas de Receptores Adrenérgicos alfa 2/farmacología , Ratones Endogámicos C57BL , Norepinefrina/metabolismo , Ratones TransgénicosRESUMEN
One of the reasons of the development of pathologies causing death is hypoxia. The purposes of this study were (1) to study some physiological and biochemical mechanisms of α2-adrenoblockers, which ensure the tissue resistance increase to hypoxia; (2) to offer new drugs contributing to the increase of tissues' stability towards the hypoxic affection; and (3) to submit new medications to surpass by their anti-hypoxic activity of those already used in modern medicine and have some advantages. The reactivity of postsynaptic vascular α2-adrenoceptors was determined on the damaged spinal cord expressed by the blood pressure increase in response to intravenous administration of azepexole that selectively binds to α2-adrenoceptors. Determination of the systemic hemodynamic values and the vascular resistance to the blood flow was performed by the method with plastic microspheres of marked isotopes. pO2 in the blood and the oxygen-transporting function were determined in a sample of 0.1 ml of blood in 30, 90, and 180 min after the α2-adrenoblockers' injections. It has been found that one of the major hemodynamic effects of mesedin and beditin was an improvement in cardiac output, as well as a prolonged increase in coronary blood flow and vasodilation of the heart vessels. Some anti-hypoxic mechanisms of the studied α2-adrenoblockers are an improvement of blood oxygen-transporting function followed by tissue oxygenation and the increased level of corticosterone and resistance to hypoxia. Revealing the mechanisms of action of the postsynaptic α2-adrenoceptors suggests that mesedin and beditin are potentially effective therapeutic means for many hypoxic conditions.
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Antagonistas de Receptores Adrenérgicos alfa 2 , Hipoxia , Ratas Wistar , Animales , Masculino , Hipoxia/tratamiento farmacológico , Hipoxia/fisiopatología , Hipoxia/metabolismo , Antagonistas de Receptores Adrenérgicos alfa 2/farmacología , Ratas , Sistema Cardiovascular/efectos de los fármacos , Sistema Cardiovascular/metabolismo , Presión Sanguínea/efectos de los fármacos , Resistencia Vascular/efectos de los fármacos , Oxígeno/metabolismo , Hemodinámica/efectos de los fármacos , Receptores Adrenérgicos alfa 2/metabolismo , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Corticosterona/sangreRESUMEN
Introduction: Nicotine is a major component of cigarette smoke with various detrimental cardiovascular effects, including increased oxidative stress in the heart. Agonism of α2-adrenergic receptors (ARs), such as with dexmedetomidine, has been documented to exert cardioprotective effects against oxidative stress and related apoptosis and necroptosis. α2-ARs are membrane-residing G protein-coupled receptors (GPCRs) that primarily activate Gi/o proteins. They are also subjected to GPCR-kinase (GRK)-2-dependent desensitization, which entails phosphorylation of the agonist-activated receptor by GRK2 to induce its decoupling from G proteins, thus terminating α2AR-mediated G protein signaling. Objective: In the present study, we sought to examine the effects of nicotine on α2AR signaling and effects in H9c2 cardiomyocytes exposed to H2O2 to induce oxidative cellular damage. Methods and Results: As expected, treatment of H9c2 cardiomyocytes with H2O2 significantly decreased cell viability and increased oxidative stress, as assessed by reactive oxygen species (ROS)-associated fluorescence levels (DCF assay) and superoxide dismutase activity. Both H2O2 effects were partly rescued by α2AR activation with brimonidine in control cardiomyocytes but not in cells pretreated with nicotine for 24 hours, in which brimonidine was unable to reduce H2O2-induced cell death and oxidative stress. This was due to severe α2AR desensitization, manifested as very low Gi protein activation by brimonidine, and accompanied by GRK2 upregulation in nicotine-treated cardiomyocytes. Finally, pharmacological inhibition of adenylyl cyclase (AC) blocked H2O2-dependent oxidative damage in nicotine-pretreated H9c2 cardiomyocytes, indicating that α2AR activation protects against oxidative injury via its classic coupling to Gai-mediated AC inhibition. Discussion/Conclusions: Nicotine can negate the cardioprotective effects of α2AR agonists against oxidative injury, which may have important implications for patients treated with this class of drugs that are chronic tobacco smokers.
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Miocitos Cardíacos , Nicotina , Humanos , Nicotina/farmacología , Nicotina/metabolismo , Peróxido de Hidrógeno/farmacología , Peróxido de Hidrógeno/metabolismo , Receptores Adrenérgicos alfa 2/metabolismo , Estrés Oxidativo , Apoptosis , Tartrato de Brimonidina/metabolismo , Tartrato de Brimonidina/farmacologíaRESUMEN
The G protein-coupled α2-adrenoceptor subtype C (abbreviated α2C-AR) has been implicated in peripheral vascular conditions and diseases such as cold feet-hands, Raynaud's phenomenon, and scleroderma, contributing to morbidity and mortality. Microvascular α2C-adrenoceptors are expressed in specialized smooth muscle cells and mediate constriction under physiological conditions and the occlusion of blood supply involving vasospastic episodes and tissue damage under pathological conditions. A crucial step for receptor biological activity is the cell surface trafficking of intracellular receptors, triggered by cAMP-Epac-Rap1A GTPase signaling, which involves protein-protein association with the actin-binding protein filamin-2, mediated by critical amino acid residues in the last 14 amino acids of the receptor carboxyl (C)-terminus. This study assessed the role of the C-terminus in Rap1A GTPase coupled receptor trafficking by domain-swapping studies using recombinant tagged receptors in transient co-transfections and compared with wild-type receptors using immunofluorescence microscopy. We further tested the biological relevance of the α2C-AR C-terminus, when introduced as competitor peptides, to selectively inhibit intracellular α2C-AR surface translocation in transfected as well as in microvascular smooth muscle cells expressing endogenous receptors. These studies contribute to establishing proof of principle to target intracellular α2C-adrenoceptors to reduce biological activity, which in clinical conditions can be a target for therapy.
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Miocitos del Músculo Liso , Péptidos , Receptores Adrenérgicos alfa 2 , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/metabolismo , Péptidos/metabolismo , Péptidos/farmacología , Receptores Adrenérgicos/metabolismo , Receptores Adrenérgicos alfa 2/efectos de los fármacos , Receptores Adrenérgicos alfa 2/metabolismo , Transducción de Señal/fisiologíaRESUMEN
Lots of adrenergic receptors (ARs) are widely present across the auditory pathways and are positioned to affect auditory and vestibular functions. However, noradrenergic regulation in the cochlea has not been well characterized. In this study, a rat model of noise-induced hearing loss was developed to investigate the expression of α2A-adrenergic receptor (AR) after acoustic trauma, then, we investigated the expression of α2A-AR in the developing rat cochlea using immunofluorescence, qRT-PCR, and Western blotting. We found that the expression of α2A-AR significantly increased in rats exposed to noise compared with controls. Immunofluorescence analysis demonstrated that α2A-AR is localized on hair cells (HCs), spiral ganglion neurons (SGNs), and the stria vascularis (SV) in the postnatal developing cochlea from post-natal day (P) 0 to P28. Furthermore, we observed α2A-AR mRNA reached a maximum level at P14 and P28 when compared with P0, while no significant differences in α2A-AR protein levels at the various stages when compared with P0. This study provides direct evidence for the expression of α2A-AR in HCs, SGNs, and the SV of the cochlea, indicating that norepinephrine might play a vital role in hearing function within the cochlea through α2A-AR.