Until recently, the visualization of cerebral microvessels was hampered by the fact that only short segments of vessels could be evaluated in brain sections by histochemistry. These limitations have been overcome by light sheet microscopy, which allows the 3D analysis of microvasculature in cleared brains. A major limitation of light sheet microscopy is that antibodies do not sufficiently penetrate cleared brains. We herein describe a technique of reverse clearing and rehydration, which after microvascular network analysis allows brain sectioning and immunohistochemistry employing a broad set of antibodies. Performing light sheet microscopy on brains of mice exposed to intraluminal middle cerebral artery occlusion (MCAO), we show that in the early phase of microvascular remodeling branching point density was markedly reduced, more strongly than microvascular length. Brain infarcts in light sheet microscopy were sharply demarcated by their autofluorescence signal, closely corresponding to brain infarcts revealed by Nissl staining. Neuronal survival, leukocyte infiltration, and astrocytic reactivity could be evaluated by immunohistochemistry in rehydrated brains, as shown in direct comparisons with non-cleared brains. Immunohistochemistry revealed microthrombi in ischemic microvessels that were likely responsible for the marked branching point loss. The balance between microvascular thrombosis and remodeling warrants further studies at later time-points after stroke.

直到最近，脑微血管的可视化还受到以下事实的阻碍：通过组织化学只能在大脑切片中评估血管的短段。通过光片显微镜已经克服了这些局限性，该技术可以对大脑清洁后的微血管进行3D分析。光学片显微镜的主要局限性在于抗体不能充分穿透清澈的大脑。我们在本文中描述了反向清除和补液的技术，该技术在微血管网络分析后允许采用多种抗体进行脑切片和免疫组化。对暴露于腔内大脑中动脉闭塞（MCAO）的小鼠的大脑进行光片显微镜检查，我们发现在微血管重构的早期，分支点密度显着降低，比微血管长度更强。薄板显微术中的脑梗死通过其自身荧光信号被清晰地标出，这与尼氏染色所揭示的脑梗死非常相似。神经元的存活，白细胞浸润和星形胶质细胞反应性可以通过免疫组织化学法在补水的大脑中进行评估，如与未清理过的大脑的直接比较所示。免疫组织化学显示缺血性微血管中的微血栓可能是明显的分支点丢失的原因。微血管血栓形成与重塑之间的平衡值得在卒中后的较晚时间点进行进一步研究。可以通过免疫组织化学方法对复水大脑中的白细胞浸润和星形胶质细胞反应性进行评估，如与未清除大脑的直接比较所示。免疫组织化学显示缺血性微血管中的微血栓可能是明显的分支点丢失的原因。微血管血栓形成与重塑之间的平衡值得在卒中后的较晚时间点进行进一步研究。可以通过免疫组织化学方法对复水大脑中的白细胞浸润和星形胶质细胞反应性进行评估，如与未清除大脑的直接比较所示。免疫组织化学显示缺血性微血管中的微血栓可能是明显的分支点丢失的原因。微血管血栓形成与重塑之间的平衡值得在卒中后的较晚时间点进行进一步研究。