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<p>Miniaturized two-photon microscopes (m2PMs) enable high-resolution, deep-brain imaging in freely behaving animals. However, the trade-offs among weight, form factor, and optical performance in conventional refractive objectives present major obstacles to achieving large field-of-view (FOV), high-resolution imaging in weight-sensitive or multiregion applications. Metalenses offer ultralight, high-NA, aberration-corrected alternatives, but face persistent challenges, including restricted FOV, off-axis aberrations, and dual-band optimization for two-photon imaging. Here, we present Meta-m2PM 2.0, a 1.06&#xa0;g headpiece featuring a compound metalens architecture with a two-stage imaging design that decouples scanning and focusing. We designed and fabricated a large-angle (± 12°) metalens-based scan lens and a high-NA (0.48) dual-layer metalens-based objective, both derived from our established polarization-insensitive Si₃N₄ meta-atom library co-optimized for focusing and transmission at 920&#xa0;nm excitation and 500–560&#xa0;nm fluorescence collection. Meta-m2PM 2.0 achieves a 350 × 330 µm<sup>2</sup> FOV (&gt; 50 × that of Meta-m2PM 1.0), with 1.17&#xa0;µm lateral and 19.85&#xa0;µm axial resolution at the center, while maintaining &gt; 80% lateral resolution uniformity across the FOV, and extending imaging depth to 210&#xa0;µm (~ 3 × deeper). We demonstrate dendritic imaging in head-fixed awake mice and, to our knowledge, the first metalens-based two-photon calcium imaging in freely behaving mice with a high signal-to-noise ratio, paving the way for ultralight multiregion neuroimaging and compact nonlinear microendoscopy.</p>

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Compound metalens-based miniature two-photon microscope for large-FOV imaging in freely behaving animals

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年12月17日

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  • Zeyu Hao,
  • Yao Zhang,
  • Yufei Zhu,
  • Bianhua Sun,
  • Hongchen Jiao,
  • Yanhui Hu,
  • Zhili Lin,
  • Lishuang Feng,
  • Aimin Wang,
  • Shumin Xiao,
  • Runlong Wu

摘要

Miniaturized two-photon microscopes (m2PMs) enable high-resolution, deep-brain imaging in freely behaving animals. However, the trade-offs among weight, form factor, and optical performance in conventional refractive objectives present major obstacles to achieving large field-of-view (FOV), high-resolution imaging in weight-sensitive or multiregion applications. Metalenses offer ultralight, high-NA, aberration-corrected alternatives, but face persistent challenges, including restricted FOV, off-axis aberrations, and dual-band optimization for two-photon imaging. Here, we present Meta-m2PM 2.0, a 1.06 g headpiece featuring a compound metalens architecture with a two-stage imaging design that decouples scanning and focusing. We designed and fabricated a large-angle (± 12°) metalens-based scan lens and a high-NA (0.48) dual-layer metalens-based objective, both derived from our established polarization-insensitive Si₃N₄ meta-atom library co-optimized for focusing and transmission at 920 nm excitation and 500–560 nm fluorescence collection. Meta-m2PM 2.0 achieves a 350 × 330 µm2 FOV (> 50 × that of Meta-m2PM 1.0), with 1.17 µm lateral and 19.85 µm axial resolution at the center, while maintaining > 80% lateral resolution uniformity across the FOV, and extending imaging depth to 210 µm (~ 3 × deeper). We demonstrate dendritic imaging in head-fixed awake mice and, to our knowledge, the first metalens-based two-photon calcium imaging in freely behaving mice with a high signal-to-noise ratio, paving the way for ultralight multiregion neuroimaging and compact nonlinear microendoscopy.