BRL-37344对心力衰竭大鼠β肾上腺素受体表达水平的影响(英文)

目的　探讨 β3 肾上腺素受体 (β3 AR)及其激动剂BRL 37344 (BRL)在心力衰竭中的作用。方法大鼠分为对照组、异丙肾上腺素 (Iso)组和Iso +BRL组。Iso组和Iso +BRL组大鼠间隔 2 4h ,scIso 340mg·kg- 12次制作心力衰竭模型。 8周后 ,Iso +BRL组大鼠从尾静脉给予BRL 0 .4nmol·kg- 1·min- 1,10min ,每周 2次 ,给药 2或 6周。分别于给Iso后 10或 14周测定死亡率、血流动力学、左室重 /体重、心肌组织 β1 ,β2 和 β3 ARmRNA水平。结果　 14周时对照组、Iso组和Iso +BRL组分别死亡 0 / 2 0 ,3/ 18和 5 / 2 2 (P >0 .0 5 )。Iso使左室收缩末压、±dp/dtmax明显降低 ,左室等容舒张时间常数、左室舒张末压明显增高 ,左室重 /体重明显增加 ;与Iso组比 ,Iso +BRL组的心脏舒缩功能进一步下降 ,左室重 /体重进一步增加。Iso组 β1 ARmRNA水平降低 ,β3 ARmRNA水平升高 ,与Iso组比较 ,Iso +BRL组 β1 ARmRNA水平更低 ,β3 ARmRNA水平更高。β2 ARmRNA 3组均有下降趋势 ,但无统计学差异。结论衰竭心脏β1 AR水平下降及 β3 AR水平增高可导致心功能降低。β3 ARmRNA水平在衰竭心脏比非衰竭心脏明显增高 ,应用 β3 AR激动剂可明显加重心力衰竭。     

Lateral Paracapsular GABAergic Synapses in the Basolateral Amygdala Contribute to the Anxiolytic Effects of ��3 Adrenoceptor Activation

Norepinephrine (NE) is known to play an integral role in the neurobiological response to stress. Exposure to stressful stimuli increases NE levels in brain regions that regulate stress and anxiety, like the basolateral amygdala (BLA). NE is thought to increase excitability in these areas through α- and β-adrenoceptors (ARs), leading to increased anxiety. Surprisingly, recent studies have shown that systemic β3-AR agonist administration decreases anxiety-like behaviors, suggesting that β3-ARs may inhibit excitability in anxiety-related brain regions. Therefore, in this study we integrated electrophysiological and behavioral approaches to test the hypothesis that the anxiolytic effects of β3-AR agonists may be mediated by an increase in BLA GABAergic inhibition. We examined the effect of a selective β3-AR agonist, BRL37344 (BRL), on GABAergic synapses arising from local circuit interneurons and inhibitory synapses originating from a recently described population of cells called lateral paracapsular (LPCS) interneurons. Surprisingly, BRL selectively enhanced LPCS-evoked inhibitory postsynaptic currents (eIPSCs) with no effect on local GABAergic inhibition. BRL also had no effect on glutamatergic synaptic excitation within the BLA. BRL potentiation of LPCS eIPSCs was blocked by the selective β3-AR antagonist, SR59230A, or by intracellular dialysis of Rp-CAMPS (cAMP-dependent protein kinase inhibitor), and this enhancement was not associated with any changes in spontaneous IPSCs or LPCS paired-pulse ratio. BRL also increased the amplitude of unitary LPCS IPSCs (uIPSCs) with no effect on uIPSC failure rate. Finally, bilateral BLA microinjection of BRL reduced anxiety-like behaviors in an open-field assay and the elevated plus-maze. Collectively, these data suggest that β3-AR activation selectively enhances LPCS, but not local, BLA GABAergic synapses, and that increases in LPCS-mediated inhibition may contribute to the anxiolytic profile of β3-AR agonists.     

Functional characterization of beta-adrenoceptors mediating relaxation in sheep gallbladder

The purpose of this study was to characterize beta-adrenoceptor subtype(s) mediating relaxation in smooth muscle strips of the sheep gallbladder. Experiments were performed on isolated smooth muscle strips suspended in tissue baths containing Krebs' solution. Isoprenaline (10(-8) M-10(-5) M) and salbutamol (10(-7) M-10(-4) M) produced concentration-dependent relaxation of carbachol (10(-7) M-3 x 10(-7) M) contracted smooth muscles of the sheep gall bladder. Isoprenaline-induced relaxation was significantly antagonized by propranolol with -logKB values of 7.81 +/- 0.11 (n = 7) and 7.73 +/- 0.12 (n = 6) in the fundic and ductal strips respectively. Atenolol (10(-5) M), a selective beta 1-adrenoceptor antagonist, also significantly antagonized isoprenaline-induced relaxation with -logKB values of 5.82 +/- 0.11 and 6.09 +/- 0.09 in the fundic and ductal strips respectively. However, ICI 118551, a selective beta 2-adrenoceptor antagonist, at concentrations up to 10(-6) M had little or no effect on isoprenaline-induced relaxation in either of these preparations. BRL 37344A, a selective beta 3-adrenoceptor agonist produced concentration-dependent relaxation of carbachol-precontracted fundic and ductal strips. BRL 37344 was approximately 9-fold more potent in the ductal than fundic strips. In both preparations, BRL 37344-induced relaxation was not significantly (p > 0.05) antagonized by propranolol (3 x 10(-7) M). This would confirm that the response was mediated via beta 3-adrenoceptors. It was concluded that beta 1- and beta 3-adrenoceptors coexist in the sheep gallbladder and mediate smooth muscle relaxation.     

Therapeutic potential of α2 adrenoceptor antagonism for antipsychotic-induced extrapyramidal motor disorders

We examined the effects of JP-1302 (a selective α_2C antagonist), BRL-44408 (a selective α_2A antagonist) and yohimbine (a non-selective α₂ antagonist) on haloperidol-induced bradykinesia and catalepsy in mice to elucidate the role of α₂ adrenoceptor subtypes in modifying extrapyramidal motor disorders. JP-1302 (0.1–1 mg/kg, s.c.) dose-dependently ameliorated haloperidol-induced bradykinesia in the pole-test and reversed the catalepsy time increased by haloperidol. Antibradykinetic and anticataleptic actions of JP-1302 were statistically significant at 0.3 and 1 mg/kg, and these doses did not alter the ambulatory distance, rearing or center–perimeter residence time in the open-field test. BRL-44408 (1–10 mg/kg, s.c.) and yohimbine (0.3–3 mg/kg, i.p.) also ameliorated haloperidol-induced bradykinesia and catalepsy. However, both agents significantly decreased ambulatory distance and rearing in the open-field test, possibly reflecting their anxiogenic actions associated with α_2A antagonism. The present study shows for the first time that blockade of α_2C receptors can alleviate antipsychotic-induced extrapyramidal motor disorders without affecting gross behaviors.     

Effects of salbutamol and BRL 37344 on diastolic arterial blood pressure, plasma glucose and plasma lactate in rabbits

Summary— The aim of this study was to investigate in rabbits the diastolic arterial blood pressure, plasma glucose and plasma lactate responses to salbutamol (a selective beta-2 adrenoceptor agonist) and BRL 37344 (a selective beta-3 adrenoceptor agonist) in comparison with CGP 12177 (a potent beta-1 and beta-2 adrenoceptor antagonist which also acts as a partial beta-3 agonist), isoprenaline (a non-selective beta-1, beta-2 and beta-3 adrenoceptor agonist) and adrenaline (a non-selective beta and alpha adrenoceptor agonist). All drugs were iv infused at the same dose: 0.3 μg/kg/min (30 min). In sodium pentobarbitone (40 mg/kg)-anasthetized animals none of these compounds altered diastolic arterial blood pressure. BRL 37344 (0.1, 0.3, 1 μg/kg/min) did not modify this parameter either. In conscious 24-h fasted rabbits, only adrenaline was able to increase plasma glucose levels. By contrast, under the same experimental conditions, salbutamol, isoprenaline and adrenaline, but not BRL 37344 or CGP 12177, induced a significant increase in plasma lactate levels. Finally, the salbutamol-mediated plasma lactate response was inhibited in the presence of clonidine (2 μg/kg/min, an alpha-2 adrenoceptor agonist), a drug considered to have opposite effects (stimulatory and inhibitory) on the adenylate cyclase system. In conclusion, these data suggest that only beta-2 adrenoceptor stimulation is able to increase plasma lactate levels, a response which is inhibited by alpha-2 adrenoceptor stimulation.     

Inhibition of Frizzled-2 by small interfering RNA protects rat hepatic BRL-3A cells against cytotoxicity and apoptosis induced by Hypoxia/Reoxygenation

Frizzled-2 plays an important role in maintaining normal hepatic cell functionality. This study aimed to investigate the role of inhibition of Frizzled-2 in protecting rat liver BRL-3A cells from Hypoxia/Reoxygenation (H/R). In vitro H/R hepatic cell model was established by culturing BRL-3A cells under H/R condition. Frizzled-2 siRNA was transfected into BRL-3A cells to inhibit Frizzled-2 signaling. Wnt5a and Frizzled-2 were significantly increased in BRL-3A cells upon H/R treatment. H/R treatment induced cell cytotoxicity, the early apoptosis rate and the intracellular Ca²⁺ level in BRL-3A cells while silencing frizzled-2 gene decreased the H/R induced cell cytotoxicity, apoptosis and intracellular Ca²⁺ level. In vivo mice study further showed the up-regulation of Frizzled-2/Wnt 5 pathway and cleaved Caspase-3 expression in liver tissues under ischemia and reperfusion injury (IRI). In summary, inhibition of Frizzled-2 by its siRNA may protects BRL-3A cells by attenuating the H/R induced cell cytotoxicity and apoptosis.     

Atypical pharmacologies at beta-adrenoceptors

Beta-Adrenoceptors are one of the most widely studied groups of G-protein-coupled receptors but continue to provide surprises and insights that have general relevance to pharmacology. Atypical pharmacologies have been described for ligands formerly (and still) described as antagonists acting at beta(1)-, beta(2)- and beta(3)-adrenoceptors that involve ligand-directed signalling and possibly allosteric interactions at the receptors. In the article by Ngala et al., in this issue of the Br J Pharmacol, another example of atypical interactions with beta-adrenoceptors is described, this time for agonists. Some of the responses to BRL37344 and clenbuterol can be explained in terms of actions at beta(2)-adrenoceptors, whereas others such as the increased glucose uptake and palmitate oxidation observed with pM concentrations of BRL37344 may involve interactions with other (possibly allosteric) sites. Atypical pharmacologies of ligands acting at beta-adrenoceptors have already indicated new ways in which ligands can interact with G-protein-coupled receptors and these mechanisms are likely to have important consequences for future drug development.     

缝隙连接增强肿瘤坏死因子-α的肝细胞毒性

【目的】 观察缝隙连接(GJ)功能改变对肿瘤坏死因子-α(TNF-α)肝细胞毒性的影响?【方法】 使用3种不同的方法:(1)GJ抑制剂预处理;(2)高低密度接种细胞;(3)小分子RNA转染抑制缝隙连接蛋白32(Cx32)表达等3种不同的方法抑制GJ的功能,观察不同浓度TNF-α作用不同时间后对肝细胞毒性的影响?【结果】 TNF-α的肝细胞毒性呈浓度和时间依赖性;抑制剂预处理后,TNF-α的肝细胞毒性明显降低(P < 0.01);低密度接种细胞(细胞间无GJ形成)时,TNF-α的肝细胞毒性较高密度接种细胞(细胞间有GJ形成)明显降低(P < 0.01);抑制Cx32的表达后,TNF-α的肝细胞毒性明显降低(P < 0.01)? 【结论】 抑制由Cx32组成的GJ的功能后,TNF-α的肝细胞毒性明显降低,Cx32的表达对于TNF-α的肝细胞毒性作用具有重要的作用?     

β3—肾上腺素能受体兴奋剂BRL37344对非肥胖和肥胖小白鼠游离足底肌葡萄糖摄取糖原合成的作用

目的：研究β3－肾上腺素能受体兴奋剂BRL 37344对非肥胖和肥胖小白鼠游离足底肌葡萄糖摄取,糖原合成的作用,方法：纯种非肥胖（＋／＋）和肥胖（ob/ob）小白鼠,年龄范围17－22周;体重,非肥胖小白鼠75－100g/只,肥胖小白鼠150－180g/只。肥胖和非肥胖小白鼠各36只,用生理和生化以及放射标记的方法和技术,测定β2－肾上腺素能受体兴奋剂BRL37344刺激葡萄糖摄取及糖原合成的作用。结果：BRL37344刺激葡萄糖摄取和糖原合成在非肥胖和肥胖小白鼠所需的BRL37344浓度不同,在非肥胖小白鼠,BRL37344的浓度在10^-10M引起最大作用的葡萄糖摄取和10^-11M引起最大作用的糖原合成,对肥胖小白鼠BRL37344浓度在10^-7M,引起最大作用的葡萄糖摄取,以及BRL37344浓度为10^-6M引起最大作用的糖原合成,结论：研究提示一个不典型的β－肾上腺素能受体（称为β3－肾上腺素能受体）存在于小白鼠的游离足底肌,肥胖小白鼠β3－肾上腺素能受体的数目,可能较非肥胖小白鼠者少以及可能存在胰岛素抵抗,β3－肾上腺素能受 兴奋剂BRL37344在一定的浓度促进骨骼肌最大作用的葡萄糖摄取和糖原合成,使骨骼肌利用葡萄糖增加,因此,BRL37344作为开发治疗糖尿病的新药物,则具有实际意义。     

Metabolic responses to BRL37344 and clenbuterol in soleus muscle and C2C12 cells via different atypical pharmacologies and beta2-adrenoceptor mechanisms

Background and purpose:

Picomolar concentrations of the β₃-adrenoceptor agonist BRL37344 stimulate 2-deoxyglucose uptake in soleus muscle via undefined receptors. Higher concentrations alter uptake, apparently via β₂-adrenoceptors. Effects of BRL37344 and β₂-adrenoceptor agonists are compared.

Experimental approach:

Mouse soleus muscles were incubated with 2-deoxy[1-¹⁴C]-glucose, [1-¹⁴C]-palmitate or [2-¹⁴C]-pyruvate, and BRL37344, β₂-adrenoceptor agonists and selective β-adrenoceptor antagonists. Formation of 2-deoxy[1-¹⁴C]-glucose-6-phosphate or ¹⁴CO₂ was measured. 2-Deoxy[1-¹⁴C]-glucose uptake and β-adrenoceptor mRNA were measured in C2C12 cells.

Key results:

10 pM BRL37344, 10 pM clenbuterol and 100 pM salbutamol stimulated 2-deoxyglucose uptake in soleus muscle by 33–54%. The effect of BRL37344 was prevented by 1 μM atenolol but not by 300 nM CGP20712A or IC3118551, or 1 μM SR59230A; that of clenbuterol was prevented by ICI118551 but not atenolol. 10 nM BRL37344 st4mulated 2-deoxyglucose uptake, whereas 100 nM clenbuterol and salbutamol inhibited uptake. These effects were blocked by ICI118551. Similar results were obtained in C2C12 cells, in which only β₂-adrenoceptor mRNA could be detected by RT-PCR. 10 nM BRL37344 and 10 pM clenbuterol stimulated muscle palmitate oxidation. In the presence of palmitate, BRL37344 no longer stimulated 2-deoxyglucose uptake and the effect of clenbuterol was not significant.

Conclusions and implications:

Stimulation of glucose uptake by 10 pM BRL37344 and clenbuterol involves different atypical pharmacologies. Nanomolar concentrations of BRL37344 and clenbuterol, probably acting via β₂-adrenoceptors, have opposite effects on glucose uptake. The agonists preferentially stimulate fat rather than carbohydrate oxidation, but stimulation of endogenous fat oxidation cannot explain why 100 nM clenbuterol inhibited 2-deoxyglucose uptake.