Adrenoceptor mRNA Levels Research Articles

Adrenergic modulation of migration, CD11b and CD18 expression, ROS and interleukin-8 production by human polymorphonuclear leukocytes.

Adrenergic modulation of immunity has been extensively characterized, however, few information exist regarding polymorphonuclear leukocytes (PMN), despite their key role in immunity and inflammation. We investigated the effect of adrenergic agents on human PMN migration, CD11b and CD18 expression, reactive oxygen species (ROS) and interleukin (IL)-8 production, and on adrenoceptor (AR) expression. Migration was measured by the Boyden chamber assay, CD11b/CD18 expression was assessed by flow cytometry, intracellular ROS were detected by spectrofluorimetry, and IL-8 was quantitated by standard ELISA assay. AR mRNA levels were measured by real-time PCR and PMN morphology was studied by scanning electron microscopy. Adrenaline(A), noradrenaline and the β-AR agonist isoprenaline reduced N-formyl-Met-Leu-Phe (fMLP)-induced migration, CD11b/CD18 expression, and ROS production, without affecting IL-8. The effect of A on CD11b was antagonized by yohimbine and propranolol, and increased by prazosin. The effect on ROS production was completely abolished by propranolol. PMN expressed α(1A)-, α(1B)-, α(1D)-, α(2A)-, α(2C)-, β(1)-, β(2)-, and β(3)-AR mRNA. A prevented fMLP-induced morphological changes of PMN. Adrenergic agents reduced PMN responses mainly through β-AR, although α-AR may contribute at least to CD11b expression. AR-operated pathways in PMN should be investigated in disease conditions and in the response to therapeutic agents.

Regulation of Catecholamine-Synthesising Enzymes and ß-Adrenoceptors Gene Expression in Ventricles of Stressed Rats

Stress exposure activates the sympathoneural system, resulting in catecholamine release. Chronic stress is associated with development of numerous disorders, including cardiovascular diseases. Here we investigated the expression of mRNAs for catecholamine biosynthetic enzymes tyrosine-hydroxylase, dopamine-beta-hydroxylase and phenylethanolamine N-methyl-transferase, and for beta(1)- and beta(2)-adrenoceptors in the right and left ventricles of rats exposed to chronic unpredictable mild stress. The tyrosine-hydroxylase and dopamine-beta-hydroxylase mRNA levels were not affected by stress, whereas the phenylethanolamine N-methyltransferase mRNA levels significantly increased in both right and left ventricles. No changes in beta(1)-adrenoceptor mRNA levels in either right or left ventricles were observed. At the same time, stress produced a significant increase of beta(2)-adrenoceptor mRNA levels in left ventricles. These results suggest that elevated expression of phenylethanolamine N-methyltransferase in both ventricules and beta(2)-adrenoceptor genes in left ventricles could provide a molecular mechanism that leads to altered physiological response, which is important for the organism coping with stress.

Relation between expression of α1-adrenoceptor mRNAs in bladder mucosa and urodynamic findings in men with lower urinary tract symptoms

To investigate the possible relationship between the bladder mucosal expression of α(1)-adrenoceptor (AR) mRNAs, storage symptoms and urodynamic findings in patients with lower urinary symptoms (LUTS) and benign prostatic obstruction (BPO). Mucosa was obtained from the posterior bladder wall from 20 patients with a diagnosis of LUTS and BPO undergoing prostatectomy. α(1)-AR subtype mRNA was quantified using competitive real-time reverse transcription-polymerase chain reaction. A preoperative video urodynamic study measured volumes for first desire to void (FDV) and strong desire to void (SDV). Patients were divided into group 1, with volumes for FDV ≤ 200 ml or SDV ≤ 300 ml, and group 2, with volumes FDV ≥ 201 ml and SDV ≥ 301 ml. Preoperative and postoperative international prostate symptom score (IPSS) and Overactive Bladder Symptom Score (OABSS) were evaluated. The group 1 FDV (141.0 ± 13.5 ml) was significantly lower than that of group 2 (299.0 ± 48.1 ml). Similarly, the SDV for group 1 (247.0 ± 21.9 ml) was also significantly lower than for group 2 (444.0 ± 32.5 ml). There were no significant differences between groups 1 and 2 with respect to mucosal α(1a)- and α(1b)-AR mRNA levels. However, group 1 patients had significantly more α(1d)-mRNA than those in group 2. No relationship between the expression of α(1)-AR mRNAs and preoperative or postoperative OABSS or IPSS was detected. There was a relationship between the expression of α(1d)-AR mRNA in the bladder mucosa and storage-phase urodynamics in LUTS/BPO patients, suggesting a role of α(1D)-ARs in bladder sensation.

Gene Expression of Adrenoceptors in the Hearts of Cold‐Acclimated Rats Exposed to a Novel Stressor

Changes in the heart rate and force of contraction are regulated by catecholamines via adrenoceptors (AR). In this work, we measured gene expression of AR in left heart atria and ventricles in rats exposed to cold stress and in cold-acclimated rats exposed to a novel stressor (immobilization). We found a significant increase in beta(3)-AR in left ventricle of rats exposed to acute (1 day) and long-term (28 days) cold, but no changes in beta(1)- and beta(2)-AR mRNA levels. However, single immobilization significantly decreased beta(2)-AR mRNA levels both in left atria and ventricles compared to acute cold stress. Application of a novel stressor (immobilization) to previously cold-acclimated animals did not show decrease of beta(2)-AR mRNA levels as seen in intact animals. Moreover, beta(1)- and beta(2)-AR did not show any significant changes. Surprisingly, the most prominent changes in the heart were detected for alpha(1B)-AR gene expression. We found decreased levels of alpha(1B)-AR mRNA in the heart of rats exposed to cold and immobilization. We also found that exposure of cold-acclimated rats to immobilization is responsible for additional decrease of alpha(1B)-AR mRNA levels in heart. It seems that while beta-AR undergoes adaptation, alpha(1B)-AR is probably prepared to modulate heart functions. Proposed mechanism of beta-AR adaptation needs to be elucidated. Thus, we have shown that gene expression of different AR subtypes in the heart is regulated differently by various stressors. A protective role of beta(2)-, beta(3)-AR, and alpha(1B)-AR in the process of heart adaptation to chronic stress exposure is proposed.

Heart Adrenoceptor Gene Expression and Binding Sites in the Human Failing Heart

Adrenergic regulation of the heart function is well documented by many studies. Catecholamines act through alpha(1)-, beta(1)-, beta(2)-, and beta(3)-adrenoceptors (ARs) in the heart. There are many findings about the changes of beta(1)- and beta(2)-AR in heart failure (HF). On the other hand, the role of other AR subtypes is not clear yet. We focused on determining how HF could affect gene expression and specific ligand binding to alpha(1A)-, alpha(1B)-, alpha(1D)-, beta(1)-, beta(2)-, and beta(3)-AR. Hearts from 11 patients with HF subjected to transplantation were investigated. As a control, corresponding parts from hearts not suitable for transplantation were used. We have found significantly higher mRNA levels of alpha(1A)-, alpha(1B)-,beta(1)-, and beta(2)-AR in the left ventricle of failing hearts compared to the levels in controls. beta(3)-AR mRNA levels in the left ventricle of failing hearts were not changed. No changes in mRNA levels of all receptors studied in other cardiac areas were found. On the other hand, binding studies showed a substantial decrease in left ventricles of failing hearts in all alpha(1)-AR subtypes and in beta(1)- and beta(2)-AR. However, the binding to beta(3)-AR was not changed. Our results suggest that alpha(1)-AR changes might be part of a compensatory mechanism, by which the heart suffering from the HF tries to secure its function, and it could be hypothesized that ineffective beta(3)-AR regulation might be involved in development of HF. According to our knowledge, this is the first report about the beta(3)-AR binding in HF.

β1-Adrenoceptor Expression in Rat Anterior Pituitary Gonadotrophs and in Mouse αT3-1 and LβT2 Gonadotrophic Cell Lines

The rat anterior pituitary expresses beta(2)-adrenoceptors (ARs) on somatotrophs, lactotrophs, and corticotrophs. The present study investigates whether beta(1)-ARs exist in the anterior pituitary, in which cell type(s) they are found, and whether they are regulated by glucocorticoids. As determined by quantitative RT-PCR and Western immunoblotting, the rat anterior pituitary expressed beta(1)-AR mRNA and protein. Unlike the beta(2)-AR, expression decreased to very low levels after 5-d aggregate cell culture but was strongly up-regulated in a dose- and time-dependent manner by dexamethasone (DEX). Glucocorticoids attenuated isoproterenol-induced down-regulation of beta(1)-AR mRNA levels. As examined by immunofluorescence confocal microscopy, beta(1)-AR immunoreactivity was detected in a subpopulation of gonadotrophs, but not in somatotrophs, lactotrophs, corticotrophs, thyrotrophs, or folliculo-stellate cells. beta(1)-AR-immunoreactivity cells were often surrounded by cup-shaped lactotrophs. Consistent with these findings, beta(1)-AR mRNA was considerably more abundant in the gonadotrophic alphaT3-1 and LbetaT2 cell lines than in the GHFT, GH3, and TtT/GF cell lines. DEX did not affect expression level in the cell lines. DEX also failed to up-regulate beta(1)-AR mRNA levels in aggregates from a subpopulation enriched in large gonadotrophs obtained by gradient sedimentation. In contrast, excessive DEX-dependent up-regulation of beta(1)-AR mRNA was found in a subpopulation enriched in small nonhormonal cells. The present data indicate that beta(1)-AR is expressed in a subpopulation of gonadotrophs with a topographical relationship to lactotrophs. However, the glucocorticoid-induced up-regulation does not seem to occur directly in the gonadotrophs but within (an)other unidentified cell type(s), or is transduced by that cell type on gonadotrophs.

Elevated beta2-adrenoceptor protein concentration in adipose tissue from obese subjects is closely related to the body mass index and waist/hip ratio.

The aim of the present study was to quantify beta(2)-adrenoceptor protein content in adipose tissue during fasting, and to study the relationships between beta(2)-adrenoceptor protein and mRNA levels and changes in metabolites related to lipolysis. Groups of male subjects with a body mass index of <25 kg/m(2) or >30 kg/m(2) fasted for 60 h. Abdominal subcutaneous fat biopsies were analysed for receptor mRNA levels by reverse transcription-PCR-HPLC. The beta(2)-adrenoceptor protein concentration was measured by Western blotting using fluorescence laser scanning for detection. The beta(2)-adrenoceptor protein concentration per cell (on a DNA basis) was higher in obese subjects ( P <0.03). There were highly significant relationships between beta(2)-adrenoceptor protein concentration and both body mass index and waist/hip ratio ( P <0.001 for both). Furthermore, there was an inverse relationship between the receptor protein concentration and the serum beta-hydroxybutyrate level during fasting ( P <0.005). beta(2)-Adrenoceptor protein levels decreased in both groups during fasting, to a similar degree. Basal beta(2)-adrenoceptor mRNA levels were similar in the two groups, but there was a smaller increase in the obese group during fasting ( P <0.03). The increased beta(2)-adrenoceptor protein level in obese subjects is likely to be related to the greater plasma membrane area of their adipocytes. The decrease during fasting may be due to increased binding of noradrenaline and subsequent internalization and degradation of the receptor. Elevated levels of less responsive beta(2)-adrenoceptor protein in obese subjects may contribute to the development of obesity.

Effects of Single and Repeated Restraint Stresses on the Expression of β1-Adrenoceptor mRNA in the Rat Hypothalamus and Midbrain

We examined the effects of single and repeated restraint stresses on the expression of β₁-adrenoceptor mRNA in the rat midbrain and hypothalamus using reverse transcriptase-polymerase chain reaction (RT-PCR). After the rats had been restrained for 4 h (single stress), or for 4 h per day during 2 or 3 consecutive days, the hypothalamus and midbrain were removed immediately and β₁-adrenoceptor mRNA levels in these regions were determined by RT-PCR. Single stress significantly decreased the mRNA level in the hypothalamus, but the mRNA level was near control levels after 2 and 3 days of stress. In the midbrain, single stress had no effect on the mRNA level, but 2 days of stress significantly increased it.

Beta(1)-Adrenoceptors compensate for beta(3)-adrenoceptors in ileum from beta(3)-adrenoceptor knock-out mice.

1. This study examines beta(1)-, beta(2)- and beta(3)-adrenoceptor (AR)-mediated responses, mRNA levels and radioligand binding in ileum from beta(3)-AR knock-out (-/-) (KO) and wild type (+/+) (FVB) mice. 2. In KO and FVB mice, SR59230A (100 nM) (beta(3)-AR antagonist) antagonized responses to (-)-isoprenaline in both KO and FVB mice. (-)-Isoprenaline mediated relaxation of ileum was antagonized weakly by ICI118551 (100 nM) (beta(2)-AR antagonist). Responses to (-)-isoprenaline were more strongly antagonized by CGP20712A (100 nM) (beta(1)-AR antagonist), propranolol (1 microM) (beta(1)-/beta(2)-AR antagonist), carvedilol (100 nM) (non-specific beta-AR antagonist), and CGP12177A (100 nM) (beta(1)-/beta(2)-AR antagonist) in ileum from KO than in FVB mice. 3. Responses to CL316243 (beta(3)-AR agonist) in ileum from FVB mice were antagonized by SR59230A (100 nM) but not by propranolol (1 microM) or carvedilol (100 nM). CL316243 was ineffective in relaxing ileum from KO mice. 4. CGP12177A had no agonist actions in ileum from either KO or FVB mice. 5. beta(1)-AR mRNA levels were increased 3 fold in ileum from KO compared to FVB mice. This was associated with an increased maximum number of beta(1)-/beta(2)-AR binding sites (B(max)). beta(2)-AR mRNA levels were unaffected while no beta(3)-AR mRNA was detected in KO mice. 6. In mouse ileum, beta(3)-ARs and to a lesser extent beta(1)-ARs are the predominant adrenoceptor subtypes mediating relaxation in ileum from FVB mice. In KO mice beta(1)-ARs functionally compensate for the lack of beta(3)-ARs, and this is associated with increased beta(1)-AR mRNA and levels of binding.

Beta(3)-adrenoceptor regulation and relaxation responses in mouse ileum.

1. This study examines the relationship between beta(3a)- and beta(3b)-adrenoceptor (AR) mRNA levels, beta(3)-AR binding and changes in ileum responses in mice treated with the beta(3)-AR agonist (R, R)-5-[2[[2-(3-chlorophenyl)-2-hydroxyethyl]-amino]-propyl]1, 3-benzodioxole-2,2-dicarboxylate (CL316243), or the beta(3)-AR antagonist 3-(2-ethylphenoxy)-1-[(1S)-1,2,3, 4-tetrahydronapth-1-ylamino]-2S-2-propanol oxalate (SR59230A), or dexamethasone or forskolin. 2. Levels of beta(3a)- and beta(3b)-AR mRNA and the maximum number of binding sites (B(max)) in ileum were unaffected following CL316243 treatment, although responses to CL316243 were reduced by 50% following 4 and 24 h treatment, indicating another desensitization mechanism not involving changes in receptor expression or number. beta(3a)-AR mRNA levels were reduced in both brown (BAT) and white adipose tissue (WAT) but beta(3b)-AR mRNA levels were significantly reduced only in WAT. Levels of beta(3a)- and beta(3b)-mRNA returned towards normal with continued treatment. 3. SR59230A treatment markedly increased beta(3)-AR mRN levels in ileum and BAT but not in WAT. The increase in beta(3)-AR mRNA levels in ileum was associated with increased B(max) levels in binding analysis and increased responses to CL316243, suggesting these as the cause of sensitization. 4. Treatment with forskolin (4 h) or dexamethasone (4 h) significantly reduced beta(3a)-AR mRNA levels in BAT and WAT but did not alter levels in ileum. Responses to CL316243 in ileum were unaffected by either treatment. 5. In summary, the beta(3)-AR is differently regulated in adipose tissue and ileum: Treatment with SR59230A increased beta(3)-AR number, mRNA and responsiveness in ileum, whereas treatment with CL316243 reduced responses without affecting beta(3)-AR number or mRNA levels.

Effects of restraint stress on α 1 adrenoceptor mRNA expression in the hypothalamus and midbrain of the rat

We examined the effects of restraint stress on α 1 adrenoceptor mRNA expression in the rat brain using reverse transcriptase–polymerase chain reaction (RT-PCR). After rats had been restrained for 10, 30, 60, 120 or 240 min, the hypothalamus and midbrain were removed immediately and α 1 adrenoceptor mRNA levels in these regions were determined by RT-PCR. Blood samples were also collected for simultaneous measurement of serum adrenocorticotropic hormone (ACTH) and corticosterone. Restraint stress resulted in a variety of changes in the hypothalamus and midbrain. In the hypothalamus, 30 and 60 min of stress resulted in a significant fall in the level of α 1 adrenoceptor mRNA relative to the control. This was associated with a rise in serum ACTH and corticosterone. In the midbrain, significant elevation of α 1 adrenoceptor mRNA was noted after 60, 120 and 240 min of restraint stress. Our findings indicated that the influence of restraint stress on α 1 adrenoceptor mRNA level in the hypothalamus is different to that of the midbrain region in rats.