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

The scan of the adult Neoceratodus 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 Neoceratodus 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). Protopterus aethiopicus and Protopterus dolloi. Specimens 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 P. aethiopicus and P. dolloi respectively. Salamanders. DiceCT contrast staining was completed using a Lugol's Iodine (I2KI) solution. 11.75% Lugol's Iodine was prepared following the protocol of Gignac & Kley (2014) . Ambystoma was stained for 5 days prior to uCT scanning, whereas Cynops 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 Ambystoma is 16.69 x 16.69 x 33.38 microns, and Cynops is 15.868 x 15.868 x 31.727 microns.

成年角齿鱼（Neoceratodus）标本的扫描数据采集自3T GE Signa Exite HDx型人体磁共振成像（Magnetic Resonance Imaging）扫描仪（型号为Signa Excite 750；通用电气医疗（GE Healthcare）；威斯康星州密尔沃基），该设备配备8通道心脏线圈，相关扫描属于《数字鱼类图书馆（Digital Fish Library）》项目（Berquist等，2012）的一部分。所用脉冲序列参数如下：翻转角35°，重复时间12.4ms，回波时间3.9ms，平均采集次数为3次。图像采集层厚为0.7mm，最终图像分辨率达703微米。所有数据均转换为DICOM格式，以用于图像处理与可视化。该成年角齿鱼标本的磁共振成像扫描数据源自加利福尼亚大学圣地亚哥分校数字鱼类图书馆馆藏（http://www.digitalfishlibrary.org）。
使用MIMICS v.17（比利时Materialise Medical公司）完成脑部与脑颅的图像分割工作。
非洲肺鱼（Protopterus aethiopicus）与细鳞肺鱼（Protopterus dolloi）的标本采用Metscher（2009）提出的方法，通过碘-碘化钾（IKI）染色剂染色三个月。显微计算机断层扫描（micro-CT，μCT）使用Feinfocus 10-160kV透射X射线源以及珀金埃尔默（Perkin Elmer）XRD0822型1MP平板X射线相机完成。扫描峰值能量设置为120kV，2秒曝光数据通过偏移与增益图像进行校正。使用OctopusV9软件将所得曝光数据重建为TIFF图像栈，非洲肺鱼与细鳞肺鱼的体素尺寸分别为87.3微米与86.9微米。
蝾螈类标本采用卢戈氏碘液（I2KI）进行双能CT对比染色（DiceCT）。按照Gignac与Kley（2014）的实验方案配制11.75%浓度的卢戈氏碘液。钝口螈（Ambystoma）在显微CT扫描前染色5天，而真螈（Cynops）仅染色72小时后即开展扫描。两类标本均使用Skyscan1173（布鲁克（Bruker））台式显微CT扫描仪成像，并通过NRecon 1.6.6软件重建为JPEG图像栈。所有文件均在ImageJ中进行裁剪以减小文件体积，并在Z轴平面以2的间隔对图像栈进行采样。钝口螈的最终体素尺寸为16.69×16.69×33.38微米，真螈为15.868×15.868×31.727微米。