Wu Xin, Muthuchamy Mariappan, Reddy Doodipala Samba
Department of Neuroscience and Experimental Therapeutics, Texas A&M Health Science Center College of Medicine Bryan, TX, USA.
Department of Medical Physiology, Texas A&M Health Science Center College of Medicine Bryan, TX, USA.
Front Aging Neurosci. 2016 May 4;8:88. doi: 10.3389/fnagi.2016.00088. eCollection 2016.
Physiological interactions between extracellular matrix (ECM) proteins and membrane integrin receptors play a crucial role in neuroplasticity in the hippocampus, a key region involved in epilepsy. The atomic force microscopy (AFM) is a cutting-edge technique to study structural and functional measurements at nanometer resolution between the AFM probe and cell surface under liquid. AFM has been incrementally employed in living cells including the nervous system. AFM is a unique technique that directly measures functional information at a nanoscale resolution. In addition to its ability to acquire detailed 3D imaging, the AFM probe permits quantitative measurements on the structure and function of the intracellular components such as cytoskeleton, adhesion force and binding probability between membrane receptors and ligands coated in the AFM probe, as well as the cell stiffness. Here we describe an optimized AFM protocol and its application for analysis of membrane plasticity and mechanical dynamics of individual hippocampus neurons in mice with chronic epilepsy. The unbinding force and binding probability between ECM, fibronectin-coated AFM probe and membrane integrin were strikingly lower in dentate gyrus granule cells in epilepsy. Cell elasticity, which represents changes in cytoskeletal reorganization, was significantly increased in epilepsy. The fibronectin-integrin binding probability was prevented by anti-α5β1 integrin. Thus, AFM is a unique nanotechnique that allows progressive functional changes in neuronal membrane plasticity and mechanotransduction in epilepsy and related brain disorders.
细胞外基质（ECM）蛋白与膜整合素受体之间的生理相互作用在海马体的神经可塑性中起着关键作用，海马体是癫痫涉及的关键区域。原子力显微镜（AFM）是一种前沿技术，用于在液体环境下研究AFM探针与细胞表面之间纳米级分辨率的结构和功能测量。AFM已逐渐应用于包括神经系统在内的活细胞研究。AFM是一种独特的技术，能够在纳米级分辨率下直接测量功能信息。除了能够获取详细的三维成像外，AFM探针还可以对细胞内成分的结构和功能进行定量测量，如细胞骨架、粘附力以及AFM探针上包被的膜受体与配体之间的结合概率，还有细胞硬度。在此，我们描述了一种优化的AFM方案及其在分析慢性癫痫小鼠单个海马神经元的膜可塑性和力学动态方面的应用。癫痫小鼠齿状回颗粒细胞中，ECM、纤连蛋白包被的AFM探针与膜整合素之间的解离力和结合概率显著降低。代表细胞骨架重组变化的细胞弹性在癫痫中显著增加。抗α5β1整合素可阻止纤连蛋白 - 整合素的结合概率。因此，AFM是一种独特的纳米技术，可用于研究癫痫及相关脑部疾病中神经元膜可塑性和机械转导的渐进性功能变化。
