Significance: To expand our understanding of the roles of astrocytes in neural circuits, there is a need to develop optical tools tailored specifically to capture their complex spatiotemporal Ca2 + dynamics. This interest is not limited to 2D, but to multiple depths. Aim: The focus of our work was to design and evaluate the optical performance of an enhanced version of a two-photon (2P) microscope with the addition of a deformable mirror (DM)-based axial scanning system for live mammalian brain imaging. Approach: We used a DM to manipulate the beam wavefront by applying different defocus terms to cause a controlled axial shift of the image plane. The optical design and performance were evaluated by an analysis of the optical model, followed by an experimental characterization of the implemented instrument. Results: Key questions related to this instrument were addressed, including impact of the DM curvature change on vignetting, field of view size, image plane flatness, wavefront error, and point spread function. The instrument was used for imaging several neurobiological samples at different depths, including fixed brain slices and in vivo mouse cerebral cortex. Conclusions: Our implemented instrument was capable of recording z-stacks of 53 μm in depth with a fine step size, parameters that make it useful for astrocyte biology research. Future work includes adaptive optics and intensity normalization.

中文翻译：

---

基于变形镜的轴向扫描，用于双光子哺乳动物脑成像

意义：为了扩大我们对星形胶质细胞在神经回路中作用的理解，需要开发专门为捕获其复杂的时空Ca2 +动力学而设计的光学工具。这种兴趣不仅限于2D，还包括多个深度。目的：我们的工作重点是设计和评估增强型双光子（2P）显微镜的光学性能，并添加基于可变形镜（DM）的轴向扫描系统进行哺乳动物活体脑成像。方法：我们使用DM通过应用不同的散焦项来控制像面的轴向偏移，从而控制光束的波前。光学设计和性能通过对光学模型的分析进行评估，然后对所实现的仪器进行实验表征。结果：解决了与该仪器有关的关键问题，包括DM曲率变化对渐晕的影响，视场大小，像平面平坦度，波前误差和点扩散函数。该仪器用于对不同深度的几种神经生物学样本进行成像，包括固定的脑切片和体内小鼠大脑皮层。结论：我们实现的仪器能够记录深度为53μm的z堆栈，步长很小，参数可用于星形胶质细胞生物学研究。未来的工作包括自适应光学和强度归一化。包括固定的脑片和体内小鼠的大脑皮层。结论：我们实现的仪器能够记录深度为53μm的z堆栈，步长很小，参数可用于星形胶质细胞生物学研究。未来的工作包括自适应光学和强度归一化。包括固定的脑片和体内小鼠的大脑皮层。结论：我们实现的仪器能够记录深度为53μm的z堆栈，步长很小，参数可用于星形胶质细胞生物学研究。未来的工作包括自适应光学和强度归一化。