Impact of wavefront distortion and scattering on 2-photon microscopy in mammalian brain tissue.
Impact of wavefront distortion and scattering on 2-photon microscopy in mammalian brain tissue.
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DOI：
10.1364/oe.19.022755
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发表时间：
2011-11-07
期刊：
影响因子：
3.8
通讯作者：
Silver RA
中科院分区：
文献类型：
--
作者：
Chaigneau E;Wright AJ;Poland SP;Girkin JM;Silver RA
双光子显微镜在神经科学中应用广泛，但对脑组织的光学特性了解甚少。我们通过活体皮质切片成像荧光珠，研究了脑组织对2P点扩散函数（PSF2P）的影响。通过结合激发光的平均自由程测量、自适应光学和基于矢量的建模（包括相位调制和散射），我们发现组织诱导的波前畸变是中等成像深度下PSF2P放大和畸变的主要决定因素。此外，它们产生的周围叶包含超过一半的2P激发。这些影响降低了精细结构的分辨率和对比度，并与散射一起限制了2P激发。我们的研究结果解开了散射和波前畸变对形成皮层PSF2P的贡献，从而为改进2P显微镜提供了基础。
Two-photon (2P) microscopy is widely used in neuroscience, but the optical properties of brain tissue are poorly understood. We have investigated the effect of brain tissue on the 2P point spread function (PSF2P) by imaging fluorescent beads through living cortical slices. By combining this with measurements of the mean free path of the excitation light, adaptive optics and vector-based modeling that includes phase modulation and scattering, we show that tissue-induced wavefront distortions are the main determinant of enlargement and distortion of the PSF2P at intermediate imaging depths. Furthermore, they generate surrounding lobes that contain more than half of the 2P excitation. These effects reduce the resolution of fine structures and contrast and they, together with scattering, limit 2P excitation. Our results disentangle the contributions of scattering and wavefront distortion in shaping the cortical PSF2P, thereby providing a basis for improved 2P microscopy.