Optical recording of action potentials with second-harmonic generation microscopy

被引:144
作者
Dombeck, DA
Blanchard-Desce, M
Webb, WW
机构
[1] Cornell Univ, Sch Appl & Engn Phys, Ithaca, NY 14853 USA
[2] Univ Rennes 1, Inst Chim, CNRS, Unite Mixte Rech 6510, F-35042 Rennes, France
关键词
imaging; action potential; membrane potential; second-harmonic generation; microscopy; nonlinear; optical; voltage-sensitive dye;
D O I
10.1523/JNEUROSCI.4840-03.2004
中图分类号
Q189 [神经科学];
学科分类号
071006 ;
摘要
Nonlinear microscopy has proven to be essential for neuroscience investigations of thick tissue preparations. However, the optical recording of fast (similar to1 msec) cellular electrical activity has never until now been successfully combined with this imaging modality. Through the use of second-harmonic generation microscopy of primary Aplysia neurons in culture labeled with 4-[4-(dihexylamino)phenyl][ethynyl]-1-(4-sulfobutyl)pyridinium (inner salt), we optically recorded action potentials with 0.833 msec temporal and 0.6 mum spatial resolution on soma and neurite membranes. Second-harmonic generation response as a function of change in membrane potential was found to be linear with a signal change of similar to6%/100 mV. The signal-to-noise ratio was similar to1 for single-trace action potential recordings but was readily increased to similar to6-7 with temporal averaging of similar to50 scans. Photodamage was determined to be negligible by observing action potential characteristics, cellular resting potential, and gross cellular morphology during and after laser illumination. High-resolution (micrometer scale) optical recording of membrane potential activity by previous techniques has been limited to imaging depths an order of magnitude less than nonlinear methods. Because second-harmonic generation is capable of imaging up to similar to400 mum deep into intact tissue with submicron resolution and little out-of-focus photodamage or bleaching, its ability to record fast electrical activity should prove valuable to future electrophysiology studies.
引用
收藏
页码:999 / 1003
页数:5
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