George W Woodruff School of Mechanical Engineering, Georgia Institute of Technology, Atlanta, United States.
Ocular and Stem Cell Translational Research Section, Ophthalmic Genetics and Visual Function Branch, National Eye Institute, National Institute of Health, Bethesda, United States.
Elife. 2024 Nov 18;13:RP97399. doi: 10.7554/eLife.97399.
Significant technical challenges exist when measuring synaptic connections between neurons in living brain tissue. The patch clamping technique, when used to probe for synaptic connections, is manually laborious and time-consuming. To improve its efficiency, we pursued another approach: instead of retracting all patch clamping electrodes after each recording attempt, we cleaned just one of them and reused it to obtain another recording while maintaining the others. With one new patch clamp recording attempt, many new connections can be probed. By placing one pipette in front of the others in this way, one can 'walk' across the mouse brain slice, termed 'patch-walking.' We performed 136 patch clamp attempts for two pipettes, achieving 71 successful whole cell recordings (52.2%). Of these, we probed 29 pairs (i.e. 58 bidirectional probed connections) averaging 91 μm intersomatic distance, finding three connections. Patch-walking yields 80-92% more probed connections, for experiments with 10-100 cells than the traditional synaptic connection searching method.
在活体脑组织中测量神经元之间的突触连接存在重大技术挑战。当用于探测突触连接时，膜片钳技术既繁琐又耗时。为了提高其效率，我们采用了另一种方法：不是在每次记录尝试后都缩回所有的膜片钳电极，而是只清洁其中一个，并在保持其他电极不变的情况下重新使用它来获得另一次记录。通过一次新的膜片钳记录尝试，可以探测到许多新的连接。通过以这种方式将一个吸管放在其他吸管前面，可以在小鼠脑切片上“行走”，称为“膜片钳行走”。我们对两个吸管进行了 136 次膜片钳尝试，成功获得了 71 个全细胞记录（52.2%）。其中，我们探测了 29 对（即 58 个双向探测连接），平均体细胞间距离为 91 μm，发现了 3 个连接。与传统的突触连接搜索方法相比，膜片钳行走在 10-100 个细胞的实验中可获得 80-92%更多的探测连接。
