Murayama Masanori, Larkum Matthew E
Physiologisches Institut, Universität Bern, Bern, Switzerland.
Nat Protoc. 2009;4(10):1551-9. doi: 10.1038/nprot.2009.142. Epub 2009 Oct 1.
Dendritic recordings in freely moving animals present great challenges using the current approaches. Here we present in detail a microendoscopic technique (the 'periscope' method) for measuring intracellular calcium activity directly from the apical dendrites of L5 pyramidal neurons from the pia down to depths of approximately 700 mum in anesthetized and freely moving rats. This method gives high signal-to-noise dendritic fluorescence responses to sensory stimuli, and has been proven to be inexpensive, straightforward and reliable, allowing essentially unrestricted behavior. We describe refinements and practical optimizations of procedures aimed at achieving dendritic Ca(2+) imaging in freely moving animals. The periscope imaging technique presented here is also ideal for combining with other in vivo recording techniques. The protocol, from the beginning of anesthesia to starting dendritic imaging, can be completed in 5 h.
使用当前方法在自由活动的动物中进行树突记录面临巨大挑战。在此，我们详细介绍一种显微内窥镜技术（“潜望镜”方法），用于在麻醉和自由活动的大鼠中，直接从L5锥体神经元的顶端树突测量细胞内钙活性，从软膜向下至约700微米深处。该方法对感觉刺激能产生高信噪比的树突荧光反应，且已被证明成本低廉、操作简单且可靠，能让动物基本不受限制地活动。我们描述了旨在实现自由活动动物树突钙成像的程序改进和实际优化。这里介绍的潜望镜成像技术也非常适合与其他体内记录技术相结合。从麻醉开始到开始树突成像的整个流程可在5小时内完成。
