Effects of adrenergic agents on the expression of zebrafish (Danio rerio) vitellogenin Ao1

Teleost vitellogenins (VTGs) are large multidomain apolipoproteins, traditionally considered to be estrogen-responsive precursors of the major egg yolk proteins, expressed and synthesized mainly in hepatic tissue. The inducibility of VTGs has made them one of the most frequently used in vivo and in vitro biomarkers of exposure to estrogen-active substances. A significant level of zebrafish vtgAo1, a major estrogen responsive form, has been unexpectedly found in heart tissue in our present studies. Our studies on zebrafish cardiomyopathy, caused by adrenergic agonist treatment, suggest a similar protective function of the cardiac expressed vtgAo1. We hypothesize that its function is to unload surplus intracellular lipids in cardiomyocytes for "reverse triglyceride transportation" similar to that found in lipid transport proteins in mammals. Our results also demonstrated that zebrafish vtgAo1 mRNA expression in heart can be suppressed by both (x-adrenergic agonist, phenylephrine (PE) and beta-adrenergic agonist, isoproterenol (ISO). Furthermore, the strong stimulation of zebrafish vtgAo1 expression in plasma induced by the beta-adrenergic antagonist, MOXIsylyl, was detected by Enzyme-Linked ImmunoSorbent Assay (ELISA). Such stimulation cannot be suppressed by taMOXIfen, an antagonist to estrogen receptors. Thus, Our present data indicate that the production of teleost VTG in vivo can be regulated not only by estrogenic agents, but by adrenergic signals as well. (c) 2009 Elsevier Inc. All rights reserved.

Effects of adrenergic agonists on the extrahepatic expression of vitellogenin Ao1 in heart and brain of the Chinese rare minnow (Gobiocypris rarus)

Teleost vitellogenins (VTGs) are large multidomain apolipoproteins and traditionally considered as the estrogen responsive precursors of the major egg yolk proteins. We identified five clones encoding VTGs, about 16% of the random EST clones from our constructed cDNA library from Chinese rare minnow liver tissue treated with 17 beta-estradiol (E2). Full-length vtgAo1 has been obtained based on the sequence information of four partial cDNA inserts by RACE. The inducibility of the vtgAo1 expression in liver by E2 was confirmed by RT-PCR. The presence of vtgAo1 transcripts have been observed primarily in liver. However. a significant level of the vtgAo1 was found in an unexpected location, heart, particularly in atrial cells by RT-PCR and whole mount in situ hybridization analyses. The vtgAo1 mRNA expression in heart and liver tissue could be suppressed by both alpha-adrenergic agonist, phenylephrine (PE) and beta-adrenergic agonist, isoproterenol (ISO). The expression of VTG in the heart observed in the present studies suggested it may provide protection from surplus intracellular lipids in fish cardiomyocytes as triglyceride transport proteins do in mammals. The results also indicated that the production of teleost vtg in vivo can be regulated by riot only estrogenic agents, but adrenergic signals as well. (c) 2008 Elsevier B.V. All rights reserved.

Differential proteomic analysis of neonatal cardiomyocytes in response to beta-adrenergic receptor stimulation

beta-Adrenoceptors(beta-ARs) play a critical role in regulating cardiac functions under both physiological and pathological conditions. To further explore the mechanisms through which beta-ARs perform its actions, proteomic approaches were adopted to study the global protein patterns in cultured neonatal rat cardiomyocytes exposed to isoproterenol (ISO). A modified method, "Mirror Images in One Gel", was used to improve the reproducibility and resolution power of two-dimensional electrophoresis. A 2-DE map with a good reproducibility was obtained in which 1281 70 spots were detected and about 1191 +/- 54 spots were matched, with an average matching rate of 92.9%. Nine proteins with significant changes were identified by using peptide mass fingerprinting(PMF) data obtained via MALDI-MS.

Pharmacological and immunocytochemical investigation of the role of catecholamines on larval metamorphosis by beta-adrenergic-like receptor in the bivalve Meretrix meretrix

Catecholamines regulate several physiological processes in mollusks. Many pharmacological experiments have been conducted to determine the effects of adrenergic agonist and antagonist of catecholamine receptors on Meretrix meretrix metamorphosis. Results showed that adrenaline (AD) and noradrenaline (NA) had substantial effects (p < 0.05) on larval metamorphosis at concentrations ranging from 10 mu M to 100 mu M. 10 mu M beta-adrenergic receptor (AR) agonist isoproterenol showed the same inducement effect as that of NA and AD on metamorphosis, whereas the alpha-AR agonist phenylephrine had no significant effect at concentrations between 0.1 mu M and 100 mu M concentrations (p > 0.05). Furthermore, I mu M beta-AR antagonist propanolol, but not alpha-AR antagonist prazosin, depressed the larval metamorphosis induced by NA or AD. By immunocytochemistry, two cell bodies of beta-adrenergic-like receptor, C/A1, C/A2, were observed in the cerebral/apical ganglion of competent larvae. In addition, there were other immunoreactive dots near C/A1 and C/A2. The results of pharmacology and immunocytochemistry suggests that beta-adrenergic-like receptor located in the larval CNS, might play a considerable role in the larval metamorphosis of M meretrix by AD or NA. (c) 2006 Elsevier B.V. All rights reserved.

调控文蛤幼虫变态的肾上腺素能受体类型及幼虫发育中神经和肌肉的变化

大部分贝类幼虫在发育过中要经历从浮游生活到底栖生活的变化过程，同时形态结构也要经历巨大的变化，这个过程称为变态。 变态是文蛤幼虫发育过程中非常重要的一个阶段。 药理学和细胞免疫学证据表明β肾上腺素样受体在文蛤幼虫变态过程中有重要作用。药理学实验分别采用了几种儿茶酚胺类受体的激动剂和抑制剂来处理幼虫，检验它们在幼虫变态过程中的作用。结果表明，在10μM和100μM的浓度下，肾上腺素(AD)和去甲肾上腺素(NA)中能够显著提高幼虫的变态率(p<0.05)。10μM和100μM浓度的AD能够提高幼虫变态率30%左右。10μM和100μM浓度的NA能分别提高幼虫变态率35.3%和27.6%。10μM的β受体激动剂－isoproterenol也能够显著的提高幼虫的变态率30%(p<0.05)，但是α受体激动剂－phenylephrine在0.1μM到100μM的浓度范围内不能显著提高幼虫的变态率(p>0.05)。而且，1μM的β受体抑制剂－propanolol能显著的抑制AD或NA提高幼虫变态率的作用(p<0.05)；但是α受体抑制剂－prazosin对AD或NA提高幼虫变态率没有显著性影响(p>0.05)。 此外本文还利用整装免疫细胞化学的方法进一步研究了文蛤幼虫不同发育阶段，神经系统和β肾上腺素样受体的发育情况。 幼虫的神经系统在担轮幼虫时期(受精后18h)开始发育，这时还不能检测到β肾上腺素受体。面盘幼虫时期(受精后1d)具备了顶神经节、脑神经节和脏神经节组成的中枢神经系统，在口附近有一些外周神经。β肾上腺素受体在受精后24h首次出现在面盘幼虫的顶神经节和脑神经节，分别命名为AR(apical receptor)和CR1(cerebral receptor 1)。 在受精后5d顶神经节已经检测不到。脑神经节和脏神经节由腹部向背部迁移，口的背腹两侧都出现了一些神经元。并且脏神经节周围也出现了一些神经细胞。AR在受精后3d就检测不到了。同时在CR1的后部新出现了一些β肾上腺素受体，命名为CR2。此后CR2发育迅速，在受精后5d就和CR1差不多大小。并且在CR1和CR2之间还出现了很多小的阳性信号。 变态过程中中枢神经系统中顶神经节消失了，并且出现了足神经节。另外，外套膜上出现了更多的外周神经。除了在稚贝的脑神经节和脏神经节外，在足、外套膜和水管上都有β肾上腺素受体存在。成体的鳃、足、心脏、水管、唇掰和外套膜上也有β肾上腺素受体分布。 变态信号传递到靶器官后，文蛤幼虫在形态结构和生态习性上开始了快速巨大的变化。其中幼虫的肌肉系统是变化最大的系统，并且直接与文蛤运动、摄食等习性转变相关。本文采用免疫组化的方法对文蛤幼虫肌肉系统的发育进行了研究。结果表明，文蛤幼虫具有由幼虫收缩肌、面盘收缩肌和闭壳肌组成的十分复杂的肌肉系统。幼虫收缩肌和前闭壳肌最早出现于担轮幼虫阶段(受精后18h)。孵化后幼虫的肌肉系统迅速发育，受精后22h幼虫的肌肉系统基本发育完全，并一直维持到变态前。变态过程中幼虫收缩肌和面盘收缩肌逐渐萎缩消失。同时，足收缩肌、后闭壳肌和外套膜肌肉等稚贝的肌肉系统快速形成。变态后稚贝幼虫收缩肌和面盘收缩肌完全消失，其肌肉系统由前后闭壳肌、足收缩肌和外套膜肌肉组成。