Astrocytes are morphologically complex and use local translation to regulate distal functions. To study the distribution of mRNA in astrocytes, we combined mRNA detection via in situ hybridization with immunostaining of the astrocyte-specific intermediate filament glial fibrillary acidic protein (GFAP). mRNAs at the level of GFAP-immunolabelled astrocyte somata, and large and fine processes were analysed using AstroDot, an ImageJ plug-in and the R package AstroStat. Taking the characterization of mRNAs encoding GFAP-α and GFAP-δ isoforms as a proof of concept, we showed that they mainly localized on GFAP processes. In the APPswe/PS1dE9 mouse model of Alzheimer's disease, the density and distribution of both α and δ forms of Gfap mRNA changed as a function of the region of the hippocampus and the astrocyte's proximity to amyloid plaques. To validate our method, we confirmed that the ubiquitous Rpl4 (large subunit ribosomal protein 4) mRNA was present in astrocyte processes as well as in microglia processes immunolabelled for ionized calcium binding adaptor molecule 1 (Iba1; also known as IAF1). In summary, this novel set of tools allows the characterization of mRNA distribution in astrocytes and microglia in physiological or pathological settings.

中文翻译：

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AstroDot - 一种研究星形胶质细胞中 mRNA 空间分布的新方法。

星形胶质细胞形态复杂，利用局部翻译来调节远端功能。为了研究星形胶质细胞中 mRNA 的分布，我们将原位杂交的 mRNA 检测与星形胶质细胞特异性中间丝胶质纤维酸性蛋白 (GFAP) 的免疫染色相结合。使用 AstroDot、ImageJ 插件和 R 软件包 AstroStat 分析 GFAP 免疫标记的星形胶质细胞胞体水平的 mRNA，以及大的和精细的过程。以编码 GFAP-α 和 GFAP-δ 亚型的 mRNA 的表征作为概念证明，我们发现它们主要定位于 GFAP 过程。在阿尔茨海默病的 APPswe/PS1dE9 小鼠模型中，Gfap mRNA 的 α 和 δ 形式的密度和分布随着海马区域和星形胶质细胞与淀粉样斑块的接近程度而变化。为了验证我们的方法，我们确认普遍存在的 Rpl4（大亚基核糖体蛋白 4）mRNA 存在于星形胶质细胞突起以及小胶质细胞突起中，并被离子钙结合接头分子 1（Iba1；也称为 IAF1）免疫标记。总之，这套新颖的工具可以表征生理或病理环境中星形胶质细胞和小胶质细胞中的 mRNA 分布。