Hanssen Katrine Sjaastad, Witter Menno P, Sandvig Axel, Sandvig Ioanna, Kobro-Flatmoen Asgeir
Kavli Institute for Systems Neuroscience, Centre for Neural Computation, Egil and Pauline Braathen and Fred Kavli Centre for Cortical Microcircuits, Norwegian University of Science and Technology (NTNU), Trondheim, Norway; Department of Neuromedicine and Movement Science, Faculty of Medicine and Health Sciences, NTNU, Trondheim, Norway.
Kavli Institute for Systems Neuroscience, Centre for Neural Computation, Egil and Pauline Braathen and Fred Kavli Centre for Cortical Microcircuits, Norwegian University of Science and Technology (NTNU), Trondheim, Norway; K.G. Jebsen Centre for Alzheimer's Disease, Faculty of Medicine and Health Sciences, NTNU, Trondheim, Norway.
J Neurosci Methods. 2023 Apr 15;390:109840. doi: 10.1016/j.jneumeth.2023.109840. Epub 2023 Mar 21.
Primary neuronal cultures enable cell-biological studies of Alzheimer's disease (AD), albeit typically non-neuron-specific. The first cortical neurons affected in AD reside in layer II of the lateralmost part of the entorhinal cortex, and they undergo early accumulation of intracellular amyloid-β, form subsequent tau pathology, and start degenerating pre-symptomatically. These vulnerable entorhinal neurons uniquely express the glycoprotein reelin and provide selective inputs to the hippocampal memory system. Gaining a more direct access to study these neurons is therefore highly relevant.
We demonstrate a methodological approach for dissection and long-term culturing of adult lateral entorhinal layer II-neurons from AD-model mice.
We maintain adult dissected lateralmost entorhinal layer II-neurons beyond two months in culture. We show that they express neuronal markers, and that they are electrophysiologically active by 15 days in vitro and continuing beyond 2 months.
Primary neurons are typically harvested from embryonic or early postnatal brains because such neurons are easier to culture compared to adult neurons. Methods to culture adult primary neurons have been reported, however, to our knowledge, culturing of adult entorhinal neuron-type specific primary neurons from AD-model animals have not been reported.
Our methodological approach offers a window to study initial pathological changes in the AD disease-cascade. This includes the study of proteinopathy, single-neuron changes, and network-level dysfunction.
原代神经元培养可用于阿尔茨海默病（AD）的细胞生物学研究，尽管通常并非神经元特异性的。AD中最早受影响的皮质神经元位于内嗅皮质最外侧部分的第II层，它们会经历细胞内淀粉样β蛋白的早期积累，形成后续的tau病理变化，并在症状出现前就开始退化。这些易损的内嗅神经元独特地表达糖蛋白Reelin，并为海马记忆系统提供选择性输入。因此，更直接地研究这些神经元具有高度相关性。
我们展示了一种从AD模型小鼠中解剖和长期培养成年外侧内嗅皮质第II层神经元的方法。
我们在培养中将成年解剖的最外侧内嗅皮质第II层神经元维持了两个多月。我们表明它们表达神经元标志物，并且在体外15天时具有电生理活性，并且这种活性持续超过2个月。
原代神经元通常从胚胎或出生后早期的大脑中获取，因为与成年神经元相比，此类神经元更容易培养。虽然已经报道了培养成年原代神经元的方法，但据我们所知，尚未报道从AD模型动物中培养成年内嗅神经元类型特异性原代神经元的方法。
我们的方法为研究AD疾病级联反应中的初始病理变化提供了一个窗口。这包括对蛋白病、单神经元变化和网络水平功能障碍的研究。
