Distribution and postnatal ontogeny of adenosine A2A receptors in rat brain: comparison with dopamine receptors

In adult rat brain, adenosine A_2A receptors and dopamine D₂ receptors are known to be located on the same cells where they interact in an antagonistic manner. In the present study we wanted to examine when this situation develops and compared the postnatal ontogeny of the binding of the adenosine A_2A receptor agonist [³H]CGS 21680, the binding of the dopamine D₁ receptor antagonist [³H]SCH 23390 and the dopamine D₂ receptor antagonist [³H]raclopride.

All three radioligands bound to the striatum at birth and this binding increased several-fold during the postnatal period. [³H]SCH 23390 binding developed first (mostly during the first week), followed by [³H]raclopride binding (first to third week) and [³H]CGS 21680 binding (only during second and third week). For all three radioligands the binding tended to decrease between 21 days and adulthood. This occurred earlier and was more pronounced in the globus pallidus than in the other examined structures. The increase in [³H]CGS 21680 binding from newborn to adult was mainly due to four-fold increase in the number of binding sites. The pharmacology of [³H]CGS 21680 binding to caudate–putamen was similar in newborn, one-week-old and adult animals, and was indicative of A_2A receptors. The binding was inhibited by guanylyl imidodiphosphate at all ages, indicating that A_2A receptors are G-protein-coupled already at birth. In contrast to the large increase in [³H]CGS 21680 binding, there was a decrease in the levels of A_2A messenger RNA during the postnatal period in the caudate–putamen. In cerebral cortex [³H]CGS 21680 bound to a different site than the A_2A receptor. From birth to adulthood cortical binding of [³H]CGS 21680 increased four-fold and that of the adenosine A₁ agonist [³H]cyclohexyladenosine 19-fold. During early postnatal development [³H]SCH 23390 binding was higher in deep than in superficial cortical layers, but this difference disappeared in adult animals. There was binding of both [³H]CGS 21680 and [³H]cyclohexyladenosine to the olfactory bulb, suggesting a role of the two adenosine receptors in processing of olfactory information. [³H]CGS 21680 binding was present in the external plexiform layer and glomerular layer, and increased during development, but the density of binding sites was about one tenth of that seen in caudate–putamen. [³H]cyclohexyladenosine showed a very different labelling pattern, resembling that observed with [³H]SCH 23390.

Postnatal changes in adenosine receptors may explain age-dependent differences in stimulatory caffeine effects and endogenous protection against seizures. Since A_2A receptors show a co-distribution with D₂ receptors throughout development, caffeine may partly exert such actions by regulating the activity of D₂ receptor-containing striatopallidal neurons     

Aging does not alter cytosolic calcium levels of cortical synaptosomes in Fischer 344 rats

Several lines of experimental evidence support an association between altered Ca²⁺ regulation and aging. It has been supposed that free cytosolic Ca²⁺ concentrations ([Ca²⁺]_i) may decrease or increase in aged animals. In this study, both resting and KCl-stimulated [Ca²⁺]_i were measured in purified cortical synaptosomes from young (3 mo.), middle-aged (12 mo.), and old (24 mo.) Fischer 344 rats. Two additional groups of rats were included, one middle-aged and one old which were trained on a treadmill for 6 months prior to experimentation. The [Ca²⁺]_i was determined using the fluorescent Ca²⁺ chelator fura-2. Net KCl-dependent changes (ΔK) in [Ca²⁺]_i were determined by the difference between stimulatory (100 μM Ca²⁺/60 mM KCl) and resting (100 μM Ca²⁺/5 mM KCl buffer) conditions among the 3 age groups. Significant increases in [Ca²⁺]_i were observed in each age group upon depolarization with 60 mM KCl. However, there were no significant age-dependent differences in either resting [Ca²⁺]_i or KCl-stimulated [Ca²⁺]_i.     

Changes in somatotropin and thyrotropin secretory patterns in aging rats

In order to clarify whether pituitary enlargement influences the secretory patterns of growth hormone (GH) and thyrotropin (TSH) in old rats, we studied the correlation between pituitary weight and plasma levels of GH and TSH in Sprague-Dawley rats of different age and sex. Young female (3–4 months; YF), old female (25 months; OF), and senescent female (33–35 months; SF) rats and young male (3–4 months; YM) and old male (24–26 months; OM) rats carrying chronic intraatrial cannulas were used. Sequential blood samples were removed through the cannulas while the animals remained conscious and undisturbed. Plasma TSH and GH as well as serum thyroxine (T₄) and triiodothyronine (T₃) were measured by radioimmunoassay. At two years of age, both males and females showed a consistent decline in GH pulse amplitude without change in trough levels. By 33–35 months of age, females showed a reversal in the previous pattern of change for GH secretion: pulse amplitude, trough levels, and mean plasma GH increased significantly with respect to the old females. The correlation between mean plasma GH and anterior pituitary (AP) weight was positive and significant (p<0.01) for females but nonsignificant for males. Old and senescent rats showed significantly lower serum T₄, but not T₃, than young animals while plasma TSH increased with age in both sexes. The present results show for the first time that senescent females hypersecrete GH and suggest that the age-related alteration of TSH secretion in rats may be due to the low levels of T₄ present in the aged animals. The correlation analysis shows that pituitary enlargement is in general associated with increased secretion of both GH and TSH in senescent female rats.     

Effects of tibolone on lipoprotein(a) and HDL subfractions

Objective: To determine the effects of tibolone, a synthetic steroid used to alleviate climacteric symptoms and prevent osteoporosis, on lipoprotein metabolism, with particular reference to lipoprotein(a) levels and HDL subfraction profiles.Design: Thirty nine postmenopausal women were treated with tibolone (Livial) 2.5 mg/day for 6 months and fasting serum lipoprotein levels were estimated at 0, 2, 4 and 6 months. Results: Lipoprotein(a) levels were reduced significantly over the 6 months from a median level of 245 (range <60–780) mg/I to 152 (range <60–530) mg/l, a reduction of 39% in the median level. A decrease was observed in approximately two thirds of the women. Reductions were noted in all 6 subjects whose pretreatment levels were high, although concentrations remained at a level associated with increased risk in all but one. There were significant decreases in triglycerides and VLDL cholesterol and no significant change in LDL cholesterol. There was a significant reduction of 18% in HDL cholesterol and a 26% reduction in the HDL₂:HDL₃ ratio. Conclusion: The reduction in lipoprotein(a) levels may have a beneficial effect on cardiovascular risk, which could go some way towards balancing the potentially adverse effect on the cardiovascular system caused by the reduction in HDL cholesterol.     

正常成人圆孔骨性通道相关参数的多层螺旋CT研究

目的 探讨MSCT测量圆孔骨性通道相关参数的数值范围。方法 回顾性分析2014年1月1日—2014年8月20日在南通市第一人民医院行头颅、鼻窦和眼眶MSCT检查的152例非外伤性患者的临床资料。其中男80例,年龄16～89岁;女72例,年龄17～79岁。所有患者行MSCT扫描,从冠状面、横断面、矢状面上显示152例304侧圆孔骨性通道,在MSCT图像上测量圆孔的深径D、圆孔中轴线与翼腭窝中轴线的夹角B、两侧圆孔中点连线与圆孔中轴线的夹角A₁、同侧圆孔中轴线与翼腭窝中轴线的夹角A₂、同侧圆孔外口外侧和内口内侧连线与翼腭窝中轴线的夹角A₃。对比相同性别不同侧别间及同一侧别不同性别间圆孔骨性通道的相关参数,计算测量数据的变异系数(CV) 和测量者个体间的CV,并采用组间相关系数(ICC)分析各测量参数的可信度。结果 MSCT能够清晰显示152例304侧圆孔的形态、大小及走行,图像质量良好。圆孔深径D值为(4.76±1.61) mm,角B、A₁、A₂、A₃分别为93.13°±8.68°、87.78°±8.64°、134.49°±8.63°、160.16°±8.22°。相同性别不同侧别间比较:角A₁右侧均大于左侧,差异均有统计学意义(P值均＜0.05);而圆孔深径D和角B、A₂、A₃右侧和左侧间差异均无统计学意义(P值均＞0.05)。同一侧别不同性别间比较:圆孔深径D和角A₁、A₂、A₃ 、B在男性、女性间差异均无统计学意义(P值均＞0.05)。D、 B、 A₁、A₂、A₃的CV范围分别为31.58%～37.14%、8.15%～9.78%、8.35%～9.94%、5.50%～6.74%、4.43%～5.59%;测量者间的CV分别为20.63%～21.54%、9.13%～9.16%、2.54%～3.77%、2.00%～2.59%、1.58%～2.69%。角B测量可信度为较好,圆孔深径D和角A₁、A₂、A₃的测量可信度为极佳。结论 在MSCT图像上对圆孔骨性通道进行测量,其结果稳定、可靠,可以为临床应用提供参考依据。     

Behavioral and neurochemical modifications induced by apomorphine treatment

By means of quantitative receptor autoradiography, the modifications of dopamine (DA) receptors in selected target regions of A₉ and A₁₀ DA neurons were studied after chronic apomorphine treatment, at a dosage able to induce behavioral changes that are claimed to be due to the activation of the target areas of A₉ neurons. An increase in [³H]spiperone binding sites (cold (+)-butaclamol) was observed in the dorsal and ventrolateral striatum, receiving fibers from A₉ neurons, while there were no changes in the ventromedial and ventrocentral striatum, in the nucleus accumbens and in the cerebral cortex receiving fibers from A₁₀ neurons. Our results suggest that the anatomical division of the DA target regions corresponds with a functional one.     

The Production of Anti-A1 Agglutinins in A-Like Rabbits Injected with Human Group A1 Secretor Saliva

A-like and non-A-like rabbits were injected with human group A₁ and A₂ secretor saliva. The sera were then examined for their agglutinative capacity against human group A₁, A₂, B and O erythrocytes before and after sequential adsorption with group O, B and A₂ erythrocytes. The A-like rabbits produced specific anti-A₁ agglutinins when injected with group A₁ saliva but not when injected with group A₂ saliva. The non-A-like rabbits produced anti-A and anti-A₁ agglutinins in response to the A₁ antigen but only anti-A agglutinins when group A₂ saliva was injected. These experiments support the interpretation that a qualitative difference exists between subgroups A₁ and A₂. The non-A-like rabbits also formed cross-reacting antibodies as shown by the increase in the agglutinins reactive with human group B red cells.     

Modulation of Ca channels by activation of adenosine A1 receptors in rat striatal glutamatergic nerve terminals

We determined that activation of adenosine A₁ receptors in striatal synaptosomes with 100 nM N⁶-cyclopentyladenosine (CPA) inhibited both the release of endogenous glutamate and the increase of intracellular free Ca²⁺ concentration ([Ca²⁺]_i), due to 4-aminopyridine (4-AP) stimulation, by 28 and 19%, respectively. Furthermore, CPA enhanced the inhibition of endogenous glutamate release due to ω-conotoxin GVIA (ω-Cgtx GVIA), ω-Cgtx MVIIC or ω-Cgtx GVIA plus ω-Cgtx MVIIC. Similar effects were observed in the [Ca²⁺]_i signal. The inhibitory effects of CPA and ω-Cgtx GVIA were additive, but the effects of CPA and ω-Cgtx MVIIC were only partially additive. These results suggest that P/Q-type Ca²⁺ channels and other type(s) of Ca²⁺ channel(s), coupled to glutamate release, are inhibited subsequently to activation of adenosine A₁ receptors.     

Effects of cholinergic depletion on experience-dependent plasticity in the cortex of the rat

Clinical and functional studies have strongly suggested that acetylcholine input from the nucleus basalis of Meynert is important for the cortex's adaptive response to experience. The purpose of this study was to investigate the effects of depletion of acetylcholine inputs from nucleus basalis of Meynert on experience-dependent plasticity in the cortex of young adult male rats. The posteromedial barrel subfield in the primary somatosensory cortex was studied. Experience-dependent plasticity was elicited using a whisker-pairing paradigm in which all whiskers except D₂ and D₃ were trimmed daily. Plasticity within barrel D₂ of the posteromedial barrel subfield was measured using the electrophysiological extracellular recording technique. An index of plasticity was determined in two ways: as an increase in the magnitude of evoked activity to stimulation of whisker D₂ and as a bias in the ratio of evoked activity for stimulation of paired whisker D₃ and cut whisker D₁ (D₃/D₁). Whiskers D₂, D₃ and D₁ were stimulated (deflected) by a Chubbuck electromechanical stimulator. Cholinergic neurons in the nucleus basalis of Meynert were selectively lesioned with an immunotoxin, 192 IgG-saporin, injected into the left lateral ventricle. Lesions of cholinergic neurons in the nucleus basalis of Meynert were verified using choline acetyltransferase immunocytochemistry and radioenzymatic assay. Experience-dependent plasticity was significantly reduced in cholinergic-depleted animals. The magnitude of evoked activity to stimulation of whisker D₂ increased by 16–100% in control animals compared with 0–20% in cholinergic-depleted animals. Similarly, compared to a 60–100% increase in the D₃/D₁ ratio of evoked activity for phosphate-buffered saline-injected control animals, cholinergic-depleted rats showed no significant increase in the D₃/D₁ ratio (0–15%) after undergoing the whisker-pairing paradigm. After whisker trimming, the D₃/D₁ response ratio in immunotoxin-treated animals was essentially the same as in control animals that had not been subjected to the whisker-pairing paradigm.

This study showed that no significant plasticity response was observed in the absence of cholinergic input from the nucleus basalis of Meynert. The mechanisms of the action of acetylcholine in cortical plasticity are still not known, but we hypothesize that this type of plasticity is activity dependent and is significantly enhanced in the presence of acetylcholine.     

脂氧素A4抑制内毒素诱导的人支气管上皮细胞环氧合酶2及前列腺素E2的表达

目的: 探讨脂氧素A₄对人支气管上皮细胞(HBECs)环氧合酶2(COX-2)表达的影响。方法: 应用不同浓度(0.1、1、10 mg/L)的内毒素(LPS)刺激HBECs 9 h,或者用1 mg/L LPS分别刺激HBECs不同时点(3 h、6 h、9 h)后,测定HBECs的COX-2 mRNA表达和细胞上清液前列腺素E₂(PGE₂)水平。应用不同浓度 (0、100、400 μmol/L) 的脂氧素A₄作用于经过LPS(1 mg/L)刺激培养9 h的HBECs,采用酶联免疫吸附法(ELISA)检测细胞上清液PGE₂的水平, 同时分别应用RT-PCR和Western blotting分别检测HBECs COX-2 mRNA及蛋白的表达。结果: LPS刺激培养条件下HBECs的COX-2 mRNA表达及其上清液PGE₂水平增加,并呈时间、剂量依赖性。脂氧素A₄能抑制LPS刺激培养HBECs COX-2蛋白和mRNA的表达及上清液PGE₂的水平,并呈剂量依赖性。结论: 脂氧素A₄能抑制LPS诱导的HBECs COX-2表达及上清液PGE₂的水平。     

Adenosine receptors in the nervous system: pathophysiological implications

Adenosine is a ubiquitous homeostatic substance released from most cells, including neurones and glia. Once in the extracellular space, adenosine modifies cell functioning by operating G-protein-coupled receptors (GPCR; A₁, A_2A, A_2B, A₃) that can inhibit (A₁) or enhance (A₂) neuronal communication. Interactions between adenosine receptors and other G-protein-coupled receptors, ionotropic receptors and receptors for neurotrophins also occur, and this might contribute to a fine-tuning of neuronal function. Manipulations of adenosine receptors influence sleep and arousal, cognition and memory, neuronal damage and degeneration, as well as neuronal maturation. These actions might have therapeutic implications for neurodegenerative diseases such as Parkinson’s disease, Alzheimer’s disease, as well as for other neurological situations such as epilepsy, idiopathic pain or even drug addition. Peripheral side effects associated with adenosine receptor agonists limit their usefulness in therapeutics; in contrast, adenosine receptor antagonists appear to have less side effects as it is the case of the well-known non-selective antagonists theophylline (present in tea) or caffeine (abundant in coffee and tea), and their emerging beneficial actions in Parkinson’s disease and Alzheimer’s disease are encouraging. A₁ receptor antagonism may also be useful to enhance cognition and facilitate arousal, as well as in the periphery when deficits of neurotransmitter release occur (e.g. myasthenic syndromes). Enhancement of extracellular adenosine levels through drugs that influence its metabolism might prove useful approaches in situations such as neuropathic pain, where enhanced activation of inhibitory adenosine A₁ receptors is beneficial. One might then consider adenosine as a fine-tuning modulator of neuronal activity, which via subtle effects causes harmonic actions on neuronal activity. Whenever this homeostasis is disrupted, pathology may be installed and selective receptor antagonism or agonism required.     

Pancreatic phospholipase A2 induces bacterial translocation in rats

The importance of pancreatic enzymes, particularly phospholipase A₂ (PLA₂), for bacterial translocation, which is considered to be one of the aggravating causes of acute pancreatitis, was investigated. Male rats were administered an intraperitoneal or intravenous injection of normal saline, PLA₂, or amylase. Four days later, the mesenteric lymph nodes (MLNs) and portal blood of the animals were cultured. None of the animals had a positive portal blood culture. The MLNs contained enteric bacteria in 78 % of the animals given 50 mg/kg of PLA₂ intraperitoneally. 5 mg/kg of PLA₂ intraperitoneally, 50 mg/kg of amylase intraperitoneally, or 50 mg/kg of PLA₂ intravenously showed positive cultures in 25 %, 20 %, and 11 %, respectively. None of the animals given intraperitoneal or intravenous normal saline had positive cultures of their MLNs. Intraperitoneal injection of 25mg/kg of nafamostat mesilate just before intraperitoneal PIA₂ (50 mg/kg) resulted in a reduction of positive MLNs from 70 % to 30 %. The cecal myeroperoxidase (MPO) activity of animals administered 50 mg/kg of PIA₂ intraperitoneally was significantly higher compared with animals administered saline intraperitoneally. These results indicate that intraperitoneal leakage of PLA₂ plays an important role in bacterial translocation during acute pancreatitis and that administration of a protease inhibitor may be effective against the bacterial translocation.     

Selective detection of adenosine A1 receptor-dependent G-protein activity in basal and stimulated conditions of rat brain [S]guanosine 5′-(γ-thio)triphosphate autoradiography

[³⁵S]Guanosine 5′-(γ-thio)triphosphate autoradiography is a novel technique to detect receptor-dependent activation of G-proteins in brain tissue sections. While an increasing number of reports using this approach are beginning to appear, little effort has been directed to the identification of factors responsible for the heterogeneously distributed [³⁵S]guanosine 5′-(γ-thio)triphosphate signal in basal conditions. The present study demonstrates that endogenously formed adenosine generates a widespread and prominent adenosine A₁ receptor-dependent signal in basal conditions using this technique. Treatment of rat brain tissue sections with the A₁-selective antagonist 8-cyclopentyl-1,3-dipropylxanthine dose-dependently ( ₅₀<10 nM) suppressed basal [³⁵S]guanosine 5′-(γ-thio)triphosphate binding in a region-specific manner, an effect fully mimicked by the adenosine-depleting enzyme adenosine deaminase, and less so by the A₁ antagonist cirsimarin and by caffeine. That adenosine was continuously formed during the incubation is supported by the constant requirements of adenosine deaminase in order to suppress basal radioligand binding and further by the fact that low micromolar concentrations of adenine nucleotides evoked only adenosine-mimicking and fully 8-cyclopentyl-1,3-dipropylxanthine-sensitive binding responses. In the presence of adenosine deaminase, all responses to adenine nucleotides were abolished, indicating that prior conversion to adenosine was required. Upon stimulation, this technique selectively detected A₁ receptor-activated G-proteins, as the non-selective agonists adenosine and 2-chloroadenosine and the A₁-selective agonist N⁶-p-sulfophenyladenosine all evoked only 8-cyclopentyl-1,3-dipropylxanthine-sensitive responses in identical gray matter areas, and also in several white matter areas such as the corpus callosum, anterior commissure, optic tract and cerebellar white matter. Dose–response studies revealed region-specific differences in the magnitude of A₁ receptor-stimulated G-protein activation, with the highest response (nine-fold over basal) detectable in the hippocampus. No response to the A_2A-selective agonist 2-[(2-aminoethylamino)carbonylethylphenylethylamino]-5′-N-ethylcarboxamidoadenosine or the A₃-selective agonist 2-chloro-N⁶-(3-iodobenzyl)-adenosine-5′-N-methyluronamide was detected in any region.

These data reveal that a significant amount of noise inherent to [³⁵S]guanosine 5′-(γ-thio)triphosphate autoradiography can be eliminated by removal of the adenosine signal, a step likely facilitating detection of responses to other receptors. Furthermore, the data establish [³⁵S]guanosine 5′-(γ-thio)triphosphate autoradiography as a novel and selective approach to directly assess A₁ receptor–G-protein coupling in anatomically defined regions of the central nervous system.     

Measurements of intracellular calcium in sensory neurons of adult and old rats

Cytosolic free calcium ([Ca²⁺]_i) was measured using fluorescent digital imaging microscopy in rat dorsal root ganglion neurons isolated from animals of two age groups (adult: seven months; and old: 30 months). Neurons were enzymatically isolated and maintained in primary culture for 14 days. Cultured neurons were loaded with the fluorescent dye, Fura-2. The spatial distribution of resting [Ca²⁺]_i was even in both adult and old rats, but the value of cytoplasmic free calcium in old neurons was significantly higher (207 ± 37 nmol/l vs96 ± 23 nmol/l) in comparison with adult ones. Depolarization with 50 mmol/l K⁺ produced a rapid increase in [Ca²⁺]_i in all neurons, but the values of depolarization-induced increase of [Ca²⁺]_i in old neurons were significantly lower (423 ± 54 nmol/l) compared with cells isolated from adult rats (1011 ± 91 nmol/l). The time of the complete restoration of [Ca²⁺]_i to the resting level was 10-times longer in old neurons. The caffeine-induced rise of intracellular calcium was somewhat higher in neurons from old animals, and its restoration to normal level was delayed.

The findings indicate a substantial alteration of the mechanisms of regulation of intracellular calcium homeostasis with neuronal ageing.     

Deleterious effect of null phenoloxidase mutation on the survival rate in Drosophila melanogaster

The effect of null activity of phenoloxidase on the survival rate was investigated in mutants of Drosophila melanogaster. Mox^GM95 and Dox-3^KD95, structural genes for prophenoloxidase A₁ and A₃, were found in natural populations in the former Soviet Union, and affected the phenoloxidase activity in active A₁ or A₃, respectively. After linking the visible markers located on the second chromosome together with the variants, cross experiments were performed to make homozygote, rdo Dox-3^KD95 pr c Mox^GM95 wt. No double mutant had emerged. In the mutant, c Mox^GM95 wt Pu², the viability was greatly reduced. These results suggested that phenoloxidase and tyrosine-3-hydroxylase act as indispensable proteins to maintain life in Drosophila.     

The Production of Anti-A1 Agglutinins in A-Like Rabbits Injected with Human Group A1 Erythrocytes

A-like and non-A-like rabbits were injected with human group A₁ erythrocytes. The rabbit sera were examined serologically for their hemagglutination patterns with human red cells of groups A₁ and A₂ before and after sequential adsorption with human group O, B and A₂ erythrocytes. The A-like rabbits produced specific anti-A₁ heteroagglutinins that resisted further adsorptions with group A₂ cells. The tissues of A-like rabbits obviously do not contain antigenic configurations equivalent to those on human group A₁ erythrocytes. Cross-reacting antibodies were also formed, by the non-A-like rabbits especially, as evidenced by the increased titer of agglutinins reactive with human group B red cells.     

Cardiac angiotensin II type I and type II receptors are increased in rats submitted to experimental hypothyroidism

This study assessed the behaviour of angiotensin II (Ang II) receptors in an experimental hypothyroidism model in male Wistar rats. Animals were subjected to thyroidectomy and resting for 14 days. The alteration of cardiac mass was evaluated by total heart weight (HW), right ventricle weight (RVW), left ventricle weight (LVW), ratio of HW, RVW and LVW to body weight (BW) and atrial natriuretic factor (ANF) expression. Cardiac and plasma Ang II levels and serum T₃ and T₄ were determined. The mRNA and protein levels of Ang II receptors were investigated by RT-PCR and Western blotting, respectively. Functional analyses were performed using binding assays. T₃ and T₄ levels and the haemodynamic parameters confirmed the hypothyroid state. HW/BW, RVW/BW and LVW/BW ratios and the ANF expression were lower than those of control animals. No change was observed in cardiac or plasma Ang II levels. Both AT1/AT2 mRNA and protein levels were increased in the heart of hypothyroid animals due to a significant increase of these receptors in the RV. Experiments performed in cardiomyocytes showed a direct effect promoted by low thyroid hormone levels upon AT1 and AT2 receptors, discarding possible influence of haemodynamic parameters. Functional assays showed that both receptors are able to bind Ang II. Herein, we have identified, for the first time, a close and direct relation of elevated Ang II receptor levels in hypothyroidism. Whether the increase in these receptors in hypothyroidism is an alternative mechanism to compensate the atrophic state of heart or whether it may represent a potential means to the progression of heart failure remains unknown.     

Stimulation of adenosine A2 receptors induces catalepsy.

The central administration of the adenosine A₂ agonist CGS 21680 induced catalepsy in the rat. This effect was counteracted by the previous systemic administration of the adenosine antagonist theophylline or the D₂ agonist BHT-920. These results are in agreement with the view that adenosine A₂ receptors regulate central dopamine D₂ transmission and underline the potential antipsychotic activity of A₂ agonists.     

Adenosine receptors linked to adenylate cyclase activity in human neuroblastoma cells: Modulation during cell differentiation

In IMR32 neuroblastoma cells, the two adenosine receptor agonists N⁶-R-phenylisopropyl-adenosine and5'-N-ethylcarboxamidoadenosine dose-dependently stimulated membrane adenylate cyclase activity with potencies consistent with the presence of adenosine receptors of the A₂-subtype. The S enantiomer of N⁶-R-phenylisopropyladenosine induced a significantly lower stimulation of adenylate cyclase, accordingly to its lower ability to activate adenosine receptors. These effects were selectively counteracted by the adenosine receptor antagonist theophylline and, conversely, were not affected by the A₁-adenosine receptor selective blocker 8-cyclopentyl-1,3-dipropylxanthine. No adenosine receptors belonging to the A₁-subtype seem, therefore, to be present in this cell line, as also shown by the lack of inhibitory activity of_{N⁶-R-phenylisopropyladenosine} on both basal and forskolin-stimulated adenylate cyclase activity. Activation of A₂-receptors did not modify intracellular basal calcium levels, did not influence calcium influx through voltage-dependent calcium channels and did not modify calcium influx and redistribution induced by muscarinic receptor activation. Prolonged exposure of cells to either N⁶-R-phenylisopropyladenosine or5'-N-ethylcarboxamidoadenosine was associated with a small but significant degree of morphological differentiation, comparable to that induced by dibutyryl cAMP, and therefore presumably related to the prolonged increase of intracellular cAMP levels elicited by the two adenosine agonists. After cellular differentiation induced with either dibutyryl cAMP or 5-bromode-oxyuridine, a selective desensitization of A₂-receptor stimulated adenylate cyclase activity was found.

IMR32 cells are a good model for studying the characteristics of the A₂-adenosine receptors in a human neuronal tissue and its regulation during neuronal differentiation or pharmacological treatments.