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Evidence of primary cilia in the developing rat heart

机译：发育中的大鼠心脏中原发性纤毛的证据

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A transient increase in cytosolic Ca2+ (the “Ca2+ transient”) determines the degree and duration of myocyte force development in the heart. However, we have previously observed that, under the same experimental conditions, the Ca2+ transients from isolated cardiac myocytes are reduced in amplitude in comparison to those from multicellular cardiac preparations. We therefore questioned whether the enzymatic cell isolation procedure might remove structures that modulate intracellular Ca2+ in some way. Primary cilia are found in a diverse range of cell types, and have an abundance of Ca2+-permeable membrane channels that result in Ca2+ influx when activated. Although primary cilia are reportedly ubiquitous, their presence and function in the heart remain controversial. If present, we hypothesized they might provide an additional Ca2+ entry pathway in multicellular cardiac tissue that was lost during cell isolation. The aim of our study was to look for evidence of primary cilia in isolated myocytes and ventricular tissue from rat hearts. Immunohistochemical techniques were used to identify primary cilia-specific proteins in isolated myocytes from adult rat hearts, and in tissue sections from embryonic, neonatal, young, and adult rat hearts. Either mouse anti-acetylated α-tubulin or rabbit polyclonal ARL13B antibodies were used, counterstained with Hoechst dye. Selected sections were also labelled with markers for other cell types found in the heart and for myocyte F-actin. No evidence of primary cilia was found in either tissue sections or isolated myocytes from adult rat ventricles. However, primary cilia were present in tissue sections from embryonic, neonatal (P2) and young (P21 and P28) rat hearts. The lack of primary cilia in adult rat hearts rules out their contribution to myocyte Ca2+ homoeostasis by providing a Ca2+ entry pathway. However, evidence of primary cilia in tissue from embryonic and very young rat hearts suggests they have a role during development.
机译：胞质Ca2 +的瞬时增加（“ Ca2 +瞬时”）决定了心脏中心肌细胞力发展的程度和持续时间。但是，我们之前已经观察到，在相同的实验条件下，与多细胞心脏制剂相比，离体心肌细胞的Ca2 +瞬变幅度减小。因此，我们质疑酶促细胞分离程序是否可能以某种方式去除调节细胞内Ca2 +的结构。原发纤毛存在于多种细胞类型中，并具有丰富的Ca2 +渗透膜通道，在激活时会导致Ca2 +流入。尽管原发纤毛普遍存在，但它们在心脏中的存在和功能仍存在争议。如果存在，我们假设它们可能会在多细胞心脏组织中提供额外的Ca2 +进入途径，而该途径在细胞分离过程中会丢失。我们研究的目的是寻找大鼠心脏分离的心肌细胞和心室组织中原发纤毛的证据。免疫组织化学技术用于鉴定成年大鼠心脏分离的心肌细胞以及胚胎，新生儿，幼年和成年大鼠心脏组织切片中的主要纤毛特异性蛋白。使用小鼠抗乙酰化α-微管蛋白抗体或兔多克隆ARL13B抗体，均用Hoechst染料复染。选定的切片还用标记物标记了心脏中发现的其他细胞类型和肌细胞F-肌动蛋白。在成年大鼠心室的组织切片或分离的心肌细胞中均未发现原发纤毛的证据。但是，原发纤毛存在于胚胎，新生儿（P2）和幼年（P21和P28）大鼠心脏的组织切片中。成年大鼠心脏中缺乏初级纤毛，通过提供Ca2 +进入途径，排除了它们对肌细胞Ca2 +稳态的贡献。但是，来自胚胎和非常年轻的大鼠心脏的组织中原发性纤毛的证据表明它们在发育过程中发挥了作用。

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