Jones Vicky C, Rodríguez José J, Verkhratsky Alexei, Jones Owen T
Faculty of Life Sciences, The University of Manchester, 1.124 Stopford Building, Oxford Road, Manchester, M13 9PT, UK.
Pflugers Arch. 2009 Aug;458(4):809-18. doi: 10.1007/s00424-009-0663-1. Epub 2009 Mar 19.
The endoplasmic reticulum (ER) is the largest intracellular membranous organelle. Functions of the ER are many and diverse, which include various biosynthetic, transport and signalling roles, central to cellular physiology, such as the biosynthesis of membrane and secretory proteins and the regulation of intracellular calcium. Its continuous lumen also serves as a highway for the distribution of proteins and ions to different regions of the cell, independent of the cytosol. The ER is an excitable organelle, capable of generating a regenerative wave of calcium release, which can propagate along the endomembrane throughout the entire cell, serving as a system of intracelluar communication in polarised cells. Nowhere is this feature of ER function more crucial than in neurones. The extremely polarised nature of nerve cells presents a unique challenge for the global co-ordination of localised physiological events such as growth cone guidance and synaptic plasticity. Clearly, the physical continuity of the neuronal ER lumen is central to its functionality as a conduit for communication. To further probe the continuity of ER in neurones and glia, we developed LV-PA-pIN-KDEL, a photoactivatable analogue of our recently described vector LV-pIN-KDEL, a lentivirally delivered ER-targeting soluble GFP. We demonstrate the ability of this vector to transduce astrocytes and neurones in culture and in cortical explants. Furthermore, we exploit the photoactivatable attributes of the vector together with a focal laser photoactivation protocol to reveal the continuous nature of the ER lumen in these cell types, presenting the first direct evidence of an astrocytic ER luminal continuum and providing more data to support the existence of a single ER lumen in neurones.
内质网（ER）是细胞内最大的膜性细胞器。内质网的功能多样，包括各种生物合成、运输和信号传导作用，这些对于细胞生理至关重要，例如膜蛋白和分泌蛋白的生物合成以及细胞内钙的调节。其连续的管腔还作为蛋白质和离子向细胞不同区域分布的通道，独立于细胞质溶胶。内质网是一种可兴奋的细胞器，能够产生钙释放的再生波，该波可沿内膜在整个细胞中传播，在极化细胞中作为细胞内通讯系统。内质网功能的这一特性在神经元中最为关键。神经细胞的极端极化性质对局部生理事件（如生长锥导向和突触可塑性）的全局协调提出了独特挑战。显然，神经元内质网管腔的物理连续性对于其作为通讯管道的功能至关重要。为了进一步探究神经元和神经胶质细胞内质网的连续性，我们开发了LV-PA-pIN-KDEL，它是我们最近描述的载体LV-pIN-KDEL的光激活类似物，LV-pIN-KDEL是一种通过慢病毒递送的内质网靶向可溶性绿色荧光蛋白。我们证明了该载体在培养物和皮质外植体中转导星形胶质细胞和神经元的能力。此外，我们利用该载体的光激活特性以及聚焦激光光激活方案来揭示这些细胞类型中内质网管腔的连续性质，首次直接证明了星形胶质细胞内质网管腔的连续性，并提供了更多数据支持神经元中单一内质网管腔的存在。
