Center for Neurodegeneration and Experimental Therapeutics, Department of Neurology, University of Alabama at Birmingham, USA.
Center for Neurodegeneration and Experimental Therapeutics, Department of Neurology, University of Alabama at Birmingham, USA.
Neuroscience. 2022 Aug 21;498:1-18. doi: 10.1016/j.neuroscience.2022.06.029. Epub 2022 Jun 23.
Dendritic spines are small protrusions on dendrites that serve as the postsynaptic site of the majority of excitatory synapses. These structures are important for normal synaptic transmission, and alterations in their density and morphology have been documented in various disease states. Over 130 years ago, Ramón y Cajal used Golgi-stained tissue sections to study dendritic morphology. Despite the array of technological advances, including iontophoretic microinjection of Lucifer yellow (LY) fluorescent dye, Golgi staining continues to be one of the most popular approaches to visualize dendritic spines. Here, we compared dendritic spine density and morphology among pyramidal neurons in layers 2/3 of the mouse medial prefrontal cortex (mPFC) and pyramidal neurons in hippocampal CA1 using three-dimensional digital reconstructions of (1) brightfield microscopy z-stacks of Golgi-impregnated dendrites and (2) confocal microscopy z-stacks of LY-filled dendrites. Analysis of spine density revealed that the LY microinjection approach enabled detection of approximately three times as many spines as the Golgi staining approach in both brain regions. Spine volume measurements were larger using Golgi staining compared to LY microinjection in both mPFC and CA1. Spine length was mostly comparable between techniques in both regions. In the mPFC, head diameter was similar for Golgi staining and LY microinjection. However, in CA1, head diameter was approximately 50% smaller on LY-filled dendrites compared to Golgi staining. These results indicate that Golgi staining and LY microinjection yield different spine density and morphology measurements, with Golgi staining failing to detect dendritic spines and overestimating spine size.
树突棘是树突上的小突起，作为大多数兴奋性突触的突触后位点。这些结构对于正常的突触传递很重要，并且已经在各种疾病状态下记录了它们的密度和形态的改变。130 多年前，Ramón y Cajal 使用高尔基染色组织切片来研究树突形态。尽管有一系列技术进步，包括离子导入 Lucifer yellow（LY）荧光染料的微注射，但高尔基染色仍然是可视化树突棘的最流行方法之一。在这里，我们比较了小鼠内侧前额叶皮层（mPFC）第 2/3 层的锥体神经元和海马 CA1 中的锥体神经元的树突棘密度和形态，使用（1）高尔基浸渍树突的明场显微镜 z 堆叠和（2）LY 填充树突的共聚焦显微镜 z 堆叠进行三维数字重建。对棘密度的分析表明，在这两个脑区，LY 微注射方法能够检测到的棘数大约是高尔基染色方法的三倍。与 LY 微注射相比，高尔基染色法测量的棘体积更大。在 mPFC 和 CA1 中，两种技术的棘长度大多相当。在 mPFC 中，高尔基染色和 LY 微注射的头部直径相似。然而，在 CA1 中，与高尔基染色相比，LY 填充树突上的头部直径小了约 50%。这些结果表明，高尔基染色和 LY 微注射产生不同的棘密度和形态测量值，高尔基染色不能检测到树突棘，并高估了棘的大小。
