Howard Hughes Medical Institute, Janelia Research Campus, Ashburn, Virginia, USA.
Nat Protoc. 2014 Dec;9(12):2784-95. doi: 10.1038/nprot.2014.190. Epub 2014 Nov 6.
Intracellular recording allows precise measurement and manipulation of individual neurons, but it requires stable mechanical contact between the electrode and the cell membrane, and thus it has remained challenging to perform in behaving animals. Whole-cell recordings in freely moving animals can be obtained by rigidly fixing ('anchoring') the pipette electrode to the head; however, previous anchoring procedures were slow and often caused substantial pipette movement, resulting in loss of the recording or of recording quality. We describe a UV-transparent collar and UV-cured adhesive technique that rapidly (within 15 s) anchors pipettes in place with virtually no movement, thus substantially improving the reliability, yield and quality of freely moving whole-cell recordings. Recordings are first obtained from anesthetized or awake head-fixed rats. UV light cures the thin adhesive layers linking pipette to collar to head. Then, the animals are rapidly and smoothly released for recording during unrestrained behavior. The anesthetized-patched version can be completed in ∼4-7 h (excluding histology) and the awake-patched version requires ∼1-4 h per day for ∼2 weeks. These advances should greatly facilitate studies of neuronal integration and plasticity in identified cells during natural behaviors.
细胞内记录允许对单个神经元进行精确的测量和操作，但它需要电极和细胞膜之间的稳定机械接触，因此在行为动物中进行仍然具有挑战性。通过刚性固定（“锚定”）将玻璃微电极固定在头部，可以在自由移动的动物中进行全细胞记录；然而，以前的固定程序速度较慢，并且经常导致大量的玻璃微电极移动，导致记录丢失或记录质量下降。我们描述了一种透明的 UV 管套和 UV 固化胶技术，该技术可以快速（在 15 秒内）将玻璃微电极固定在原位，几乎没有任何移动，从而大大提高了自由移动全细胞记录的可靠性、产量和质量。首先从麻醉或清醒的头部固定大鼠中获得记录。UV 光使连接玻璃微电极和管套的薄胶层固化到头部。然后，动物可以在不受约束的行为期间快速平稳地释放进行记录。麻醉贴片版本大约需要 4-7 小时（不包括组织学）完成，清醒贴片版本每天需要大约 1-4 小时，持续大约 2 周。这些进展应该极大地促进在自然行为过程中对已鉴定细胞的神经元整合和可塑性的研究。
