Kelly Emily A, Majewska Ania K
Neurobiology and Anatomy, University of Rochester, USA.
J Vis Exp. 2010 Oct 25(44):2060. doi: 10.3791/2060.
In vivo imaging using two-photon laser scanning microscopy (2PLSM) allows the study of living cells and neuronal processes in the intact brain. The technique presented here allows the imaging of the same area of the brain at several time points (chronic imaging) with microscopic resolution allowing the tracking of dendritic spines which are the small structures that represent the majority of postsynaptic excitatory sites in the CNS. The ability to clearly resolve fine cortical structures over several time points has many advantages, specifically in the study of brain plasticity in which morphological changes at synapses and circuit remodeling may help explain underlying mechanisms. In this video and supplementary material, we show a protocol for chronic in vivo imaging of the intact brain using a thinned-skull preparation. The thinned-skull preparation is a minimally invasive approach, which avoids potential damage to the dura and/or cortex, thus reducing the onset of an inflammatory response. When this protocol is performed correctly, it is possible to clearly monitor changes in dendritic spine characteristics in the intact brain over a prolonged period of time.
使用双光子激光扫描显微镜（2PLSM）进行的体内成像能够研究完整大脑中的活细胞和神经突。此处介绍的技术能够在多个时间点对大脑的同一区域进行成像（长期成像），其微观分辨率足以追踪树突棘，这些小结构代表了中枢神经系统中大多数突触后兴奋性位点。在多个时间点清晰分辨精细皮质结构的能力具有诸多优势，尤其在研究脑可塑性方面，突触处的形态变化和回路重塑可能有助于解释潜在机制。在本视频及补充材料中，我们展示了一种使用薄颅骨制备方法对完整大脑进行长期体内成像的方案。薄颅骨制备是一种微创方法，可避免对硬脑膜和/或皮质造成潜在损伤，从而减少炎症反应的发生。如果正确执行该方案，就能够在较长时间内清晰监测完整大脑中树突棘特征的变化。
