The vitamin A derivative all-trans-retinoic acid (ATRA) is an important biologically active metabolite that regulates a variety of essential biological processes in particular via gene-regulatory mechanisms. In the retina, ATRA is a light-dependent byproduct of the phototransduction cascade. Here, ATRA is not only needed for proper retinal development, but it also acts as a neuromodulator on horizontal cells, second-order inhibitory neurons in the outer retina, which reveal morphological and physiological changes when the retina is treated with ATRA. There is evidence that gene-regulatory mechanisms may only be partially involved in these neuromodulatory processes and the underlying nontranscriptional mechanisms are still elusive. This is, among other things, due to the lack of appropriately labeled ATRA, which would allow the tracking of ATRA in cells or a given tissue. To overcome this obstacle, we designed, synthesized, and evaluated two conjugates of ATRA, one conjugated with biotin (biotin-ATRA) and one conjugated with diaminoterephthalate fluorophore (DAT-ATRA), as molecular tools for different fields of application. The biocompatibility of both compounds was demonstrated via cell viability assays in cultured N2a-cells. N2a-cells exposed to the compounds showed no significant changes in the viability rate. The functionality of synthesized ATRA-conjugates was verified using retinal tissue derived from adult carp. The binding of ATRA-conjugates to distinct retinal cells was assessed in primary cultures of carp retina. Hereby, horizontal and Müller cells have been identified as specific target cells of the new ATRA compounds. Electron microscopy further confirmed that the new substances are still able to induce synaptic plasticity at horizontal cell dendrites resulting in formation of spine synapses, as it is shown for native ATRA. Taken together, the novel ATRA-conjugates represent biocompatible and functional molecular tools, which may further provide the possibility to track ATRA in neuronal cells and study its modulatory effects in different cell systems.

中文翻译：

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两种新型全反式维甲酸缀合物的合成和评价：用于视网膜研究的生物相容性和功能性工具

维生素A衍生物全反式-视黄酸（ATRA）是一种重要的生物活性代谢物，尤其是通过基因调控机制来调控多种重要的生物过程。在视网膜中，ATRA是光转导级联的光依赖性副产物。在这里，ATRA不仅是适当的视网膜发育所必需的，而且还充当水平细胞的神经调节剂，是外部视网膜中的二阶抑制神经元，当用ATRA治疗视网膜时，它们揭示了形态和生理变化。有证据表明，基因调节机制可能仅部分参与这些神经调节过程，而潜在的非转录机制仍然难以捉摸。除其他外，这是由于缺少适当标记的ATRA，这将允许追踪细胞或给定组织中的ATRA。为了克服这一障碍，我们设计，合成和评估了两种ATRA偶联物，一种与生物素偶联（biotin-ATRA），另一种与二氨基对苯二甲酸二氨基酯荧光团（DAT-ATRA）偶联，作为针对不同应用领域的分子工具。两种化合物的生物相容性通过培养的N2a细胞中的细胞活力测定法得到证实。暴露于该化合物的N2a细胞的存活率没有显着变化。使用衍生自成年鲤鱼的视网膜组织验证了合成的ATRA-缀合物的功能。在鲤鱼视网膜的原代培养中评估了ATRA-缀合物与不同的视网膜细胞的结合。因此，水平细胞和Müller细胞已被鉴定为新ATRA化合物的特定靶细胞。电子显微镜进一步证实，新物质仍然能够在水平细胞树突处诱导突触可塑性，导致形成脊柱突触，如天然ATRA所示。综上所述，新型ATRA偶联物代表了生物相容性和功能性分子工具，这可能进一步提供跟踪神经元细胞中ATRA并研究其在不同细胞系统中的调节作用的可能性。