DICE microCT and MRI data for Mandibular musculature constrains brain-endocast disparity in sarcopterygians

The scan of the adult <i>Neoceratodus</i> specimen was obtained using the 3T GE Signa Exite HDx human MRI scanner (Signa Excite 750; GE Healthcare; Milwaukee, WI), equipped with an 8-Channel Cardiac Coil, as part of the Digital Fish Library project Berquist et al. (2012). The pulse sequence parameters used were: flip angle of 35 degrees, 12.4 ms repetition time, 3.9 ms echo time, and three averages. Images were collected with slice thickness of 0.7 mm and resulting image resolution of 703 microns. Data were converted to DICOM format for image processing and visualisation. The Magnetic Resonance Imaging (MRI) scan of the adult <i>Neoceratodus</i> specimen was obtained from the Digital Fish Library catalogue, University of California San Diego (http://www.digitalfishlibrary.org). Image segmentation of the brain and braincase was performed using MIMICS v.17 (Materialise MedicalCo Belgium). <br> <i>Protopterus aethiopicus and Protopterus dolloi.</i> Specimens <br> were stained in IKI using the method of Metscher (2009) for three months. Micro-CT (uCT) scans were obtained using a Feinfocus 10-160kV transmission X-ray source and a Perkin Elmer XRD0822 1 MP flat panel X-ray camera. The peak energy used was 120kV, and 2 second exposures were corrected with offset and gain images. The resulting exposures were reconstructed as a tif stack using OctopusV9 resulting in a voxel size of 87.3 and 86.9 microns for <i>P. aethiopicus </i>and <i>P. dolloi</i> respectively. <br> <br> <i>Salamanders. </i>DiceCT contrast staining was completed using a Lugol's Iodine (I2KI) solution. 11.75% Lugol's Iodine was prepared following the protocol of Gignac &amp; Kley (2014) . <i>Ambystoma</i> was stained for 5 days prior to uCT scanning, whereas <i>Cynops</i> was scanned after only 72 hours. Both specimens were imaged using a Skyscan1173 (Bruker) desktop uCT scanner and were reconstructed as a stack of jpegs using NRecon 1.6.6. In both cases, the files were cropped in ImageJ (to reduce file size and sampled images from the stack at an interval of 2 in the Z-plane. The resultant voxel size for <i>Ambystoma</i> is 16.69 x 16.69 x 33.38 microns, and <i>Cynops</i> is 15.868 x 15.868 x 31.727 microns.<br>

本成年<i>Neoceratodus</i>（角齿鱼属）标本的扫描数据，采自搭载8通道心脏线圈的3T GE Signa Exite HDx人体磁共振扫描仪（Signa Excite 750；通用电气医疗集团；威斯康星州密尔沃基市），作为数字鱼类图书馆（Digital Fish Library）项目（Berquist等，2012）的一部分获取。所采用的脉冲序列参数如下：翻转角35°，重复时间12.4 ms，回波时间3.9 ms，平均采集次数3次。图像采集的切片厚度为0.7 mm，最终图像分辨率为703微米。数据被转换为DICOM格式以用于图像处理与可视化。
本成年<i>Neoceratodus</i>标本的磁共振成像（Magnetic Resonance Imaging，MRI）扫描数据源自加利福尼亚大学圣迭戈分校数字鱼类图书馆馆藏目录（http://www.digitalfishlibrary.org）。脑与颅腔的图像分割采用MIMICS v.17（比利时Materialise Medical公司）完成。

<i>Protopterus aethiopicus</i>（非洲肺鱼）与<i>Protopterus dolloi</i>（多氏肺鱼）标本：采用Metscher（2009）的方法，以IKI溶液染色三个月。显微CT（Micro-CT，μCT）扫描使用Feinfocus 10-160kV透射X射线源与珀金埃尔默（Perkin Elmer）XRD0822 1 MP平板X射线相机完成。管电压峰值设置为120kV，2秒曝光通过偏移与增益图像进行校正。最终的投影数据经OctopusV9软件重建为TIFF图像堆栈，非洲肺鱼与多氏肺鱼的体素尺寸分别为87.3微米与86.9微米。

<i>Salamanders</i>（蝾螈类）标本：采用卢戈氏碘液（Lugol's Iodine，I2KI）完成弥散碘对比增强计算机断层扫描（DiceCT）染色。按照Gignac与Kley（2014）的实验方案配制11.75%卢戈氏碘液。<i>Ambystoma</i>（钝口螈属）标本在显微CT扫描前染色5天，而<i>Cynops</i>（蝾属）标本仅染色72小时后即进行扫描。两类标本均使用Skyscan1173（布鲁克（Bruker）公司）台式显微CT扫描仪成像，并通过NRecon 1.6.6软件重建为JPEG图像堆栈。两类数据均在ImageJ中进行裁剪以减小文件体积，并在Z轴平面以2的间隔对图像堆栈进行采样。钝口螈属标本的最终体素尺寸为16.69 × 16.69 × 33.38微米，蝾属标本则为15.868 × 15.868 × 31.727微米。