Vicencio Jose Miguel, Ibarra Cristian, Estrada Manuel, Chiong Mario, Soto Dagoberto, Parra Valentina, Diaz-Araya Guillermo, Jaimovich Enrique, Lavandero Sergio
Centro FONDAP (Fondo de Invesigación Avanzada en Areas Prioritarias) Estudios Moleculares de la Celula, Universidad de Chile, Olivos 1007, Santiago 6640750, Chile.
Endocrinology. 2006 Mar;147(3):1386-95. doi: 10.1210/en.2005-1139. Epub 2005 Dec 8.
Androgens are associated with important effects on the heart, such as hypertrophy or apoptosis. These responses involve the intracellular androgen receptor. However, the mechanisms of how androgens activate several membrane signaling pathways are not fully elucidated. We have investigated the effect of testosterone on intracellular calcium in cultured rat cardiac myocytes. Using fluo3-AM and epifluorescence microscopy, we found that exposure to testosterone rapidly (1-7 min) led to an increase of intracellular Ca2+, an effect that persisted in the absence of external Ca2+. Immunocytochemical analysis showed that these effects occurred before translocation of the intracellular androgen receptor to the perinuclear zone. Pretreatment of the cells with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethylester and thapsigargin blocked this response, suggesting the involvement of internal Ca2+ stores. U-73122, an inhibitor of phospholipase C, and xestospongin C, an inhibitor of inositol 1,4,5-trisphosphate receptor, abolished the Ca2+ signal. The rise in intracellular Ca2+ was not inhibited by cyproterone, an antagonist of intracellular androgen receptor. Moreover, the cell impermeant testosterone-BSA complex also produced the Ca2+ signal, indicating its origin in the plasma membrane. This effect was observed in cultured neonatal and adult rat cardiac myocytes. Pertussis toxin and the adenoviral transduction of beta- adrenergic receptor kinase carboxy terminal peptide, a peptide inhibitor of betagamma-subunits of G protein, abolished the testosterone-induced Ca2+ release. In summary, this is the first study of rapid, nongenomic intracellular Ca2+ signaling of testosterone in cardiac myocytes. Using various inhibitors and testosterone-BSA complex, the mechanism for the rapid, testosterone-induced increase in intracellular Ca2+ is through activation of a plasma membrane receptor associated with a Pertussis toxin-sensitive G protein-phospholipase C/inositol 1,4,5-trisphosphate signaling pathway.
雄激素对心脏有重要影响，如导致心肌肥大或凋亡。这些反应涉及细胞内雄激素受体。然而，雄激素如何激活多种膜信号通路的机制尚未完全阐明。我们研究了睾酮对培养的大鼠心肌细胞内钙的影响。使用fluo3-AM和落射荧光显微镜，我们发现暴露于睾酮后迅速（1-7分钟）导致细胞内Ca2+增加，这种效应在无细胞外Ca2+时持续存在。免疫细胞化学分析表明，这些效应在细胞内雄激素受体转位至核周区之前就已出现。用1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸-乙酰甲酯和毒胡萝卜素预处理细胞可阻断此反应，提示细胞内钙库参与其中。磷脂酶C抑制剂U-73122和肌醇1,4,5-三磷酸受体抑制剂西司他汀C可消除Ca2+信号。细胞内雄激素受体拮抗剂环丙孕酮不抑制细胞内Ca2+升高。此外，细胞不能透过的睾酮-牛血清白蛋白复合物也产生Ca2+信号，表明其起源于质膜。在培养的新生和成年大鼠心肌细胞中均观察到这种效应。百日咳毒素和β-肾上腺素能受体激酶羧基末端肽（一种G蛋白βγ亚基的肽抑制剂）的腺病毒转导可消除睾酮诱导的Ca2+释放。总之，这是第一项关于睾酮在心肌细胞中快速、非基因组细胞内Ca2+信号传导的研究。使用各种抑制剂和睾酮-牛血清白蛋白复合物，睾酮诱导细胞内Ca2+快速增加的机制是通过激活与百日咳毒素敏感的G蛋白-磷脂酶C/肌醇1,4,5-三磷酸信号通路相关的质膜受体。
