Most brain functions engage a network of distributed regions. Full investigation of these functions thus requires assessment of whole brains; however, whole-brain functional imaging of behaving animals remains challenging. This protocol describes how to follow brain-wide activity in awake head-fixed mice using functional ultrasound imaging, a method that tracks cerebral blood volume dynamics. We describe how to set up a functional ultrasound imaging system with a provided acquisition software (miniScan), establish a chronic cranial window (timing surgery: ~3–4 h) and image brain-wide activity associated with a stimulus at high resolution (100 × 110 × 300 µm and 10 Hz per brain slice, which takes ~45 min per imaging session). We include codes that enable data to be registered to a reference atlas, production of 3D activity maps, extraction of the activity traces of ~250 brain regions and, finally, combination of data from multiple sessions (timing analysis averages ~2 h). This protocol enables neuroscientists to observe global brain processes in mice.

大多数大脑功能都涉及分布式区域网络。因此，对这些功能的全面调查需要对整个大脑进行评估；然而，行为动物的全脑功能成像仍然具有挑战性。该协议描述了如何使用功能超声成像跟踪清醒的头部固定小鼠的全脑活动，这是一种跟踪脑血容量动态的方法。我们描述了如何使用提供的采集软件 (miniScan) 建立功能性超声成像系统，建立慢性颅窗（定时手术：~3-4 小时）和图像与高分辨率刺激相关的全脑活动（100 × 110 × 300 µm 和每个脑切片 10 Hz，每个成像会话需要约 45 分钟）。我们包括使数据能够注册到参考图集的代码，生成 3D 活动地图，提取约 250 个大脑区域的活动轨迹，最后，组合来自多个会话的数据（时间分析平均约 2 小时）。该协议使神经科学家能够观察小鼠的全球大脑过程。