Data from: Beauty is more than skin deep: a non-invasive protocol for in vivo anatomical study using micro-CT

Microcomputed tomography (μCT) is a widely used tool in biomedical research, employed to investigate tissues and bone structures of small mammals in vivo. The application of in vivo μCT scanning in non-medical studies greatly lags behind the rapid advancements made in the biomedical field wherein the methodology has evolved to allow for longitudinal studies and eliminate the need to sacrifice the animal. Ecological and evolutionary studies often involve morphological measurements of a large sample of live animals; however, the potential of in vivo μCT imaging as a method for data acquisition has yet to be delineated.

Here, we describe a protocol for in vivo μCT imaging of the internal anatomy of reptiles and amphibians, commonly used study organisms in ecological and evolutionary research. We consider the skeletal and extraskeletal (i.e. osteoderms) bones of a lizard as a case study to elucidate the potential of in vivo μCT imaging. First, we explore the effects of various parameter settings on radiation dose, scan time and image quality. Secondly, we develop a protocol to immobilize and restrain study organisms during scanning without need for the administration of anaesthetics and compare the results of the in vivo protocol to images obtained post-mortem.

To immobilize animals, we replace the use of anaesthetics by cooling, thereby allowing the use of previously unsuitable rotating gantry μCT scanners that are readily available in scientific institutions. The resultant image quality of in vivo μCT scans is similar to that of post-mortem μCT scans, especially in the abdominal region. We discuss the effect of tube voltage, distance to X-ray source and metal filtration on radiation dose, and how these parameters could be altered to reduce the cumulative radiation dose while maintaining optimal image quality.

The proposed in vivo μCT protocol offers a new approach to acquire anatomical information for non-biomedical studies. We offer specific suggestions as to how the protocol can be employed to suit a variety of model organisms.

显微计算机断层扫描（Microcomputed tomography，μCT）是生物医学研究中广泛应用的工具，可用于在体研究小型哺乳动物的组织与骨骼结构。然而，非医学研究中的在体μCT扫描应用，远落后于生物医学领域的快速发展——后者的方法学已演进至可支持纵向研究，且无需牺牲实验动物。生态与进化研究通常需要对大量活体动物开展形态学测量，但作为数据获取手段的在体μCT成像，其应用潜力尚未得到充分阐明。

本研究针对生态与进化研究中常用的模式生物——爬行动物与两栖动物，提出一套用于其内部解剖结构的在体μCT成像方案。我们以蜥蜴的骨骼与骨骼外结构（即真皮骨）为案例，阐明在体μCT成像的应用潜力。首先，我们探究了不同参数设置对辐射剂量、扫描时长与图像质量的影响；其次，我们开发了一套扫描过程中固定实验动物的方案，无需施加麻醉，并将在体成像方案的结果与死后扫描获得的图像进行对比。

在动物固定环节，我们采用冷却替代麻醉，从而可使用科研机构中普遍配备的、此前不适用于该场景的旋转机架μCT扫描仪。在体μCT扫描所得的图像质量与死后扫描结果相近，尤其在腹部区域表现更为突出。我们还探讨了管电压、X射线源距离与金属滤过对辐射剂量的影响，以及如何在维持最优图像质量的前提下，调整这些参数以降低累积辐射剂量。

本研究提出的在体μCT成像方案，为非生物医学研究提供了一种获取解剖学信息的新途径。我们还针对该方案如何适配不同模式生物，提供了具体的应用建议。